Paracetamol disposition in normal subjects and in patients treated with antiepileptic drugs. 1 The serum concentration profile of paracetamol has been determined after administration of single 1000 mg intravenous and oral doses in six normal subjects and six epileptic patients on chronic antiepileptic drug therapy. The urinary excretion of free and conjugated paracetamol has also been determined. 2 Following intravenous administration, serum paracetamol concentration declined with first-order kinetics. Both elimination rate and total body clearance were higher in the epileptic patients, although in neither case was the difference statistically significant. 3 The oral bioavailability (mean +/- s.e. mean) was significantly lower in the epileptic patients than in the normal subjects (0.77 +/- 0.03 and 0.89 +/- 0.02 respectively, P less than 0.01), whereas the urinary excretion total (free+conjugated) paracetamol was almost identical in the two groups. 4 It is suggested that the lower bioavailability of paracetamol in the epileptic patients results from enhancement of first-pass metabolism, secondary to enzyme induction.


Serum, tear and salivary concentrations of methotrexate in man. 1. Methotrexate handling has been studied in four patients with psoriasis and eleven patients with neoplastic disease. 2. Methotrexate levels in serum, tears and saliva were measured by radioimmunoassay while protein binding studies were performed by continuous ultrafiltration. 3. There was a close correlation between methotrexate concentrations in tears and serum (P less than 0.001, r = 0.714). Parotid salivary and serum methotrexate levels were similarly correlated (P less than 0.001, r = 0.557) but not mixed salivary and serum levels (P greater than 0.1, r = 0.232). 4. The mean protein binding was 95.11% +/- 2.26 (s.d.) while the ratio of methotrexate levels in tears to the free serum methotrexate level was 1:1.04. The corresponding parotid salivary level: free serum level ratio was 1:18.11. 5. No relationship could be determined between the methotrexate levels in tears and conjunctivitis observed in some of the patients under study.


Pupil responses to intravenous heroin (diamorphine) in dependent and non-dependent humans. 1. Intravenous heroin was administered to volunteers, in doses of 2.5 and 5 mg to non-dependent subjects and does of 1/6, 1/3 and 1/2 of their prescribed daily does of opiates to dependent subjects, and pupillary responses measured before and three times during the 2 h after injection. 2. Tolerance to the miotic effects of heroin in the dependent subjects was demonstrated--larger doses of heroin were needed to produce the same pupil response in dependent subjects than in non-dependent subjects and the duration of action was shorter in the former group. 3. The effect of concurrent oral methadone medication on pupil response to heroin was demonstrated. Subjects prescribed both methadone and heroin showed smaller control pupil diameters and a reduced dose effect to heroin than did subjects prescribed heroin alone.


Amniocentesis with and without ultrasound guidance. The increasing use of amniocentesis requires that the technique be made as safe as possible. The results of 284 amniocenteses done under ultrasound control are presented and compared with the results of 140 amniocenteses performed without ultrasound control. The use of ultrasound guidance increased the success rate from 80 to 99.6 per cent and decreased the incidence of feto-maternal haemorrhage from 8.5 to 2.8 per cent. The incidence of blood-stained amniotic fluid was reduced from 43.5 to 17.6 per cent. The clinical significance of these findings is discussed and the results interpreted as indicating that amniocentesis done with ultrasound guidance is a safer and less traumatic procedure.


The incidence of different types of fetal movements during pregnancy. In high risk pregnancies with severe fetal distress, a reduction of fetal movements may take place before fetal death occurs. This decrease is accompanied by a weakening of the fetal movements. One hundred and twenty pregnant women (310 recordings) between 20 and 41 weeks gestation recorded fetal movements and classified them into weak, strong, and rolling. The movements noted by the woman were correlated to those recorded by a fetal movements recorder. The rate of weak movements gradually decreased until the 36 to 37th week, while the strong and rolling movements increased. From the 36 to 37th week till term weak movements increased again with a decline in strong and rolling movements. Before fetal death, or in severe fetal distress with decreased total movements, the relative rate of weak movements increased. Reduction in daily total movements without a change in the distribution of the type of movement may not indicate fetal distress.


The recurrence rate and significance of low oestriol excretion in successive pregnancies. A total of 440 women who had low oestriol excretion in a previous pregnancy was investigated by urinary oestriol assays in one or more subsequent pregnancies. The incidence of low oestriol excretion in the subsequent pregnancy was 29.1 per cent, or more than double that in the total obstetric population (13.4 per cent; p less than 0.001). Patients with persistently low oestriol excretion had a 40.8 per cent recurrence rate in subsequent pregnancies. When oestriol excretion was low in successive pregnancies it retained a significant association with increased incidences of stillbirths, neonatal deaths and fetal growth retardation. It was concluded that low oestriol excretion in a previous pregnancy is a definite indication to test fetoplacental function in subsequent pregnancies even when the clinical findings are normal.


A survey of 59 triplet pregnancies.

A survey of 59 triplet pregnancies. The outcome of 59 triplet pregnancies from four hospitals was analyzed. The combined stillbirth and neonatal mortality rate was 232/1000. Preterm labour and birth order were the most important identifiable factors in relation to neonatal death. Birth weight within a triplet set and malpresentation were not related to neonatal death. Elective cervical suture appeared to be of no value in prolonging gestation.


Melatonin in the maternal and umbilical circulations during human parturition. The concentration of melatonin in maternal peripheral plasma was measured during late pregnancy, term and pre-term labour. There was a small increase in the mean concentration of melatonin during labour which was significant in term labour. Umbilical arterial and venous plasma, whether obtained at term, after spontaneous vaginal delivery or at Caesarean section, contained significantly greater concentrations of melatonin than maternal plasma. A significant arterio-venous difference was demonstrated for both groups of umbilical samples with raised venous levels after spontaneous vaginal delivery but higher arterial levels at Caesarean section.


Prostaglandin E2 tablets used intravaginally for the induction of labour. Prostaglandin E2 tablets were administered intravaginally 505 patients for the induction of labour, without amniotomy, using doses ranging from 1 to 5 mg as a single insertion. The advantages and limitations of this method of induction of labour are discussed.


Normal patterns of fetal activity in the third trimester. The incidence and frequency of fetal respiratory movements (FRM) and fetal trunk movements (FTM) were measured in 21 normal pregnancies between 28 and 39 weeks gestation. Fetal activity was assessed with a real-time ultrasound scanner and all observed movements were punched directly onto computer tape. Recordings were made for one hour in three over a 24 hour period, providing a total of 168 hours for analysis. The overall mean percentage incidence of FRM was 31 per cent at a mean breathing rate of 43 breaths/minute. The mean percentage incidence of FRM during the day (0800-1759 hours) was 37 per cent. The overall mean percentage incidence of FTM was 18 per cent with a mean of 29 moves per 30 minute observation period. The mean percentage incidence of FTM during the day (0800-1759 hours) was 16 per cent. There was a well-defined circadian variation in fetal activity with FRM peaking between 1900 and 2200 hours, and FTM between 2200 and 0100 hours. There was considerable variation in the amount of FRM and FTM recorded from hour to hour. Total fetal activity rarely fell below 10 per cent in any one recording period and this measurement may be a more useful clinical test of fetal well-being than FRM or FTM alone.


Studies on the maternal immune response to placental antigens: absence of a blocking factor from the blood of abortion-prone women. The leucocyte migration-inhibition assay was employed to assess the reactivity of cells from postpartum women to placental antigen preparations. Leucocytes from normal primigravidae responded to their own placental antigens and displayed a limited degree of cross-reactivity with allogeneic preparations. Autologous plasma suppressed this antigen recognition and the factor responsible was shown to be an IgG antibody. Leucocytes from five women with spontaneous abortions also responded to autologous placental preparations but no immunosuppressive factor could be detected in their blood. It is suggested that target antigens for maternal lymphocytes are present on the placenta, a structure critical to the continuance of pregnancy, and that the presence of the blocking antibody is necessary for fetal well-being.


The effect of danazol on menorrhagia, coagulation mechanisms, haematological indices and body weight. Eighteen patients with objective evidence of menorrhagia (more than 80 ml menstrual blood loss) were treated with danazol for twelve weeks. Danazol significantly reduced the menstrual blood loss from 231 +/- 39 ml (mean +/- SEM) to 135 +/- 33 ml in the first treatment month and the mean loss thereafter was only 21 ml and 3 ml for the second and third months respectively. A rapid increase in haemoglobin level and a reduction in the number of days of bleeding were also observed on danazol treatment. No important effect on the coagulation profile was observed during the period of study. Three months after stopping danazol, menstrual blood loss (103 +/- 27 ml) was still significantly less than the pre-treatment loss.


Plasma ferritin levels as an index of iron deficiency in women using intrauterine devices. Forty-seven women were studied before and for one year after insertion of an intrauterine contraceptive device (IUCD). The menstrual blood loss increased in 44 subjects (94 per cent), and this was related to the surface area of the IUCD. Plasma ferritin levels in the iron deficient range (below 16 microgram/l) were present in 9 subjects (19 per cent) before insertion of the IUCD, whereas by the end of one year, 21 subjects (45 per cent) had reached these levels. The incidence of depressed ferritin levels was highest in subjects with a monthly blood loss exceeding 80 ml. The haemoglobin, mean corpuscular haemoglobin and mean corpuscular volume measurements declined progressively, but were less sensitive predictors of iron deficiency. These findings indicate the need to monitor iron status and administer oral iron replacement therapy in subjects with an IUCD.


Plasma levels of oestrone and oestradiol-17beta in postmenopausal women. The plasma oestrone and oestradiol levels of 34 postmenopausal women were studied and related to various of their other characteristics. The plasma oestrone and oestradiol-17beta levels (Mean +/- SE) were, respectively, 32.09 +/- 4.6 pg/ml and 13.9 +/- 3.1 pg/ml. There was a highly significant negative linear association between the years elapsed since the menopause and oestrone levels (P less than 0.001) and oestradiol levels (P less than 0.02). Both plasma oestrone and oestradiol levels were directly related to weight, but the relation was only significant for oestradiol. A significant positive correlation was found between vaginal bleeding and excess weight. The degree of oestrogenicity of the vaginal smear was directly related to the levels of oestrone (P less than 0.08) and oestradiol (P less than 0.005) and to the degree of endometrial hyperplasia (P less than 0.02).


Oestrogen-gonadotrophin feedback abnormalities in hyperprolactinaemic amenorrhoea demonstrated by the response to clomiphene administration. Clomiphene was administered to 16 patients with elevated serum prolactin levels in doses of 100, 200 and 300 mg/day for five days in succeeding months and total urinary oestrogens estimated on days 0, 5, 8, 12 and 15 following commencement of treatment. The responses were compared with six patients who were amenorrhoeic but had normal serum prolactin levels and absent positive feedback to oestrogen. The increased outputs of oestrogens were similar in the two groups. In the hyperprolactinaemic group 5 out of 16 subjects showed evidence of ovulation whilst the remainder showed a secondary failure of response. Six subjects who failed to ovulate were treated with clomiphene and human chorionic gonadotrophin (HCG) and ovulations were induced in 31 out of 34 treatment cycles but no pregnancies were achieved. The responses to clomiphene therapy in the hyperprolactinaemic subjects were compared to the assessment of positive feedback mechanisms by means of oestrogen provocation and oestrogen amplification tests and good correlation was obtained. Only those with evidence of positive feedback to these tests were likely to ovulate on clomiphene.


Management of acute primary angle-closure glaucoma: a long-term follow-up of the results of peripheral iridectomy used as an initial procedure. For the past 12 years peripheral iridectomy has been used as the initial surgical procedure after an acute attack of angle-closure glaucoma. The operation has been performed 2 to 5 days after the pressure has been controlled medically and has been followed routinely by prophylactic peripheral iridectomy in the fellow eye. Peripheral iridectomy for primary acute angle-closure glaucoma controlled the condition without any need for further medical or surgical therapy in 72% of cases. Careful and frequent follow-up within the first 4 postoperative months is essential to prevent further field loss, which can occur very rapidly if the pressure is not adequately controlled. If the pressure remains uncontrolled, further surgery should not be delayed. It was impossible to decide which patients would eventually need trabeculectomy when they first attended in the acute attack. Peripheral iridectomy as a prophylactic procedure was entirely effective in preventing an acute angle-closure attack in the fellow eye.


Management of chronic or intermittent primary angle-closure glaucoma: a long-term follow-up of the results of peripheral iridectomy used as an initial procedure. Patients presenting with either intermittent closed-angle glaucoma which gave to subacute congestive attacks or with chronic angle-closure glaucoma were followed up over 12 years. Peripheral iridectomy was performed as a primary procedure on these patients during this period. It was found to be a highly effective procedure in those patients without field loss at the time of presentation, but because of the figures presented here we would recommend that any patient presenting with angle closure and disc and field changes should have a trabeculectomy performed as a primary procedure. We found no way of predicting which patients would require further  surgery from the history, initial intraocular pressure, or the gonioscopic findings. No patient in this series developed malignant glaucoma after trabeculectomy although it occurred in 2 eyes after peripheral iridectomy.


Acute aphakic pupil block glaucoma: an alternative surgical approach. In a series of 7 cases of acute aphakic pupil block glaucoma a surgical approach designed to break adhesions between the iris and the anterior hyaloid face was uniformly successful. Reasons for the success of the method are discussed and comparisons made with other surgical approaches.


Trabeculotomy in late onset congenital glaucoma. All eyes of children above the age of 1 year referred because of glaucoma were treated with trabeculotomy as the primary procedure. The surgery was modified by the excision of a rectangle of tissue in the deep scleral bed under the scleral flap instead of the classical radial incision. This serves to exteriorise Schlemm's canal while thinning the scleral tissue, without, however, entering the anterior chamber. A total of 7 eyes in 5 children between the ages of 1 and 9 years were operated upon. In 6 of the 7 eyes intraocular pressure has remained under 20 mmHg without treatment after follow-up of between 18 months and 4 years. In the seventh eye intraocular pressure was controlled for 2 years but has since required epinephrine drops twice daily for the maintenance of pressure below 20 mmHg. In 4 of the 7 eyes flat, diffuse conjunctival filtering blebs are present, but no complications have been encountered. We consider that trabeculotomy may be recommended for children with late-onset congenital glaucoma.


Rieger's syndrome with pericentric inversion of chromosome 6. Pericentric inversion of chromosome 6 (6p+q-) was found in a girl with Rieger's syndrome and in her father. The only ocular signs in the father were prominent iris mounds and Schwalbe's line. The association of chromosomal anomalies with Rieger's syndrome indicates the need for a chromosome banding test in familial or sporadic patients with the syndrome and in patients with mild anomalies of the anterior chamber angle.


The trabecular wall of Schlemm's canal: a study of the effects of pilocarpine by scanning electron microscopy. The scanning electron microscope was used to study the endothelium lining the trabecular wall of Schelemm's canal in 10 human eyes enucleated in treatment of choroidal melanomas. The eyes of 5 patients had been treated before enucleated with pilocarpine drops, and the remaining 5 were untreated and served as controls. In the pilocarpine-treated tissue there was an increased prominence of cellular bulges in the endothelial monolayer, and the endothelial pores were both larger and more frequent than in the untreated eyes. By the use of Poiseuille's formula it was calculated that the resistance offered by the pore system to the drainage of aqueous in the pilocarpine-treated group was approximately one-ninth of that in the control series.


The relationship between the lifetime and yield of the 735 nm fluorescence of chloroplasts at low temperatures. The lifetime and relative yield of the 735 nm fluorescence of chloroplasts, over a range of low temperatures (-60 to -196 degrees C) where the yield of fluorescence changes markedly, were found to be directly proportional. It is concluded that the species of chlorophyll responsible for the 735 nm fluorescence, C-705, is present over the entire temperature range but is less fluorescent at the higher temperatures because of greater energy transfer to P-700. It is also concluded from attempts to measure the rise-time of the 735 nm fluorescence at -196 degrees C that the rise-time is less than 50 ps.


Dichlorophenylurea-resistant oxygen evolution in Chlorella after cerulenin treatment. Fluorescence spectra at 77 K, oxygen evolution at 30 degrees C and delayed fluorescence at 25 degrees C were measured in Chlorella pyrenoidosa cultures with and without cerulenin and subsequent 3-(3,4-dichlorophenyl)-1,1-dimethyl-urea (DCMU) treatment, respectively. In pure algal cultures the oxygen evolution was inhibited by DCMU and the long-time component of fluorescence was highly influenced by DCMU, as expected. In contrast, both oxygen evolution and delayed fluorescence became DCMU-resistant in cerulenin-treated cultures. The DCMU-resistance is correlated with a change in the fatty acid distribution of the thylakoid membrane, which also leads to changes in the prompt fluorescence. Cerulenin appears to be a promising new tool of diagnostics for the hitherto unsatisfactorily understood processes of oxygen evolution in photosynthesizing organisms.


Fractionation of membranes from Acholeplasma laidlawii A on the basis of their surface properties by partition in two-polymer aqueous phase systems. Acholeplasma laidlawii A consists of pleomorphic cell clusters surrounded by a single membrane. When lysed, a cell gives rise to several membrane fragments which cannot be separated from each other by isopycnic sucrose gradient centrifugation. A heterogeneous lateral organization of the cell membranes was detected by countercurrent distribution of membrane fragments in a two-polymer aqueous phase system. It revealed that the membranes consist of at least two subpopulations with respect to surface properties. Changes in the fatty acid and cholesterol content of the membranes revealed that the resolution of different subpopulations was predominantly due to a critical ratio of monoglucosyldiglyceride to diglucosyldiglyceride. The heterogeneity of the membrane probably depends on lipid-lipid and lipid-protein steric interactions. Charged lipids, an apolar monoglucolipid and the ratio between lipids and proteins also affect membrane partition. The differences in the subpopulations were further reflected by different specific activities of NADH dehydrogenase, NADH oxidase and ATPase. These activities varied independently. Minor quantitative differences in the protein patterns of different subpopulations were apparent. The origin and the preservation of the membrane subpopulations are discussed in terms of lipid-lipid and lipid-protein interactions, their age and energy metabolism.


Two-dimensional electrophoresis of surface glycoproteins of normal BHK cells and ricin resistant mutants. The surface glycoproteins of baby hamster kidney (BHK) cells were iodinated by lactoperoxidase and submitted to a two-dimensional electrophoresis procedure involving isoelectric focusing in the first dimension and SDS gel electrophoresis in the second dimension. After autoradiography a complex but reproducible pattern was obtained. The technique was then applied to the study of three ricin-resistant mutant clones with reduced rates of cell-cell and/or cell-substratum adhesion. Abnormal patterns were observed in all three mutant clones indicating different mechanisms of ricin resistance and identifying glycoproteins which may be involved in cellular interactions.


Role of unstirred layer in intestinal absorption of phenylalanine in vivo. The appearance rate of L- and D-phenylalanine in the venous blood of rat jejunal loops in vivo is increased up to 60% if the intraluminal solution is mixed more efficiently by the simultaneous perfusion of air. The effect decreases as the luminal concentration is increased to 100 mmol/l. Thus, the apparent Michaelis constants are by 50% lower in the case of the reduced unstirred layer (26 to 17 for L- and 9 to 6 mmol/l for D-phenylalanine). The enhancement of the absorption and the reduction of the Michaelis constants can be attributed to the reduction of the effective unstirred layer thickness by about 400--500 micrometer.


Effect of membrane cholesterol enrichment or depletion on the partition behavior of human erythrocytes in dextran-poly(ethylene glycol) aqueous phases. It has previously been shown that by appropriate manipulation of polymer concentrations and ionic composition and concentration one can select whether charge-associated or lipid-related membrane surface properties are reflected by cell partition in dextran-poly(ethylene glycol) aqueous two-phase systems (Walter, H. (1977) in Methods of Cell Separation ((Catsimpoolas, N., ed.), Vol. 1, pp. 307-354, Plenum Press, New York). In the current experiments we have studied that partition behavior of human erythrocytes and found that not only lipid-related but also charge-associated membrane properties are altered as a consequence of cholesterol-enrichment or -depletion. Results further indicate that, just as cell partition in charged phase systems reflects membrane charge-associated properties not readily measured by means other than partition (Brooks, D.E., Seaman, G.V.F. and Walter, H. (1971) Nat. New Biol. 234, 61--62; Walter, H., Tung, R., Jackson, L.J. and Seaman, G.V.F. (1972) Biochem. Biophys. Res. Commun. 48, 565--571), cell partition in uncharged phases reflects membrane lipid-related properties also not readily measured by other means.


Lipid reorganization in biological membranes: a study by Fourier transform infrared difference spectroscopy. The first application of infrared difference spectroscopy to the study of a natural biological membrane is described. Perdeuterated palmitic acid was incorporated biosynthetically into the lipids of the plasma membrane of Acholeplasma laidlawii and the temperature-induced structural rearrangement of the endogenous lipids monitored via their C--2H vibrational modes. Changes in infrared parameters were studied between 0 and 50 degrees C and contrasted with those occurring in the model membrane system of 1,2-diperdeuteropalmitoyl-sn-glycero-3-phosphocholine. The phase transition of the biomembrane occurs over a 20 degrees C range with the temperature of the maximum rate of change of absorbance coinciding with that of the sharp phase transition of the model membrane.


Characterization of the membrane fraction isolated by the fluorescein mercuric acetate technique of Barland and Schroeder. Using scanning electron microscopy we have demonstrated that tha membrane fraction isolated by the fluorescein mercuric acetate technique of Barland and Schroeder (Barland, P. and Schroeder, E.A. (1975) J. Cell Biol. 45, 662-668) represents a topologically distinct membrane which circumscribes the cell nucleus. Our data suggest that not all the cells within a non-synchronized cell population release a membrane fraction after treatment according to the technique of Barland and Schroeder, but rather that the efficiency of membrane release achieved using this preparative technique is dependent on the morphology of individual cells. Our work has also demonstrated that the peptide composition of the membrane fraction isolated by the technique of Barland and Schroeder differs from the peptide composition of the plasma membrane-enriched fraction isolated by the technique of Brunette and Till (Brunette, D.M. and Till, J.E. (1971) J. Membrane Biol. 5, 215-224). This difference in peptide composition is particularly noticeable among the higher molecular weight proteins, glycoproteins and iodineateable membrane components. The data which we have accumulated suggest that the compositional differences noted between the two membrane isolates do not result from differential extraction of membrane components during the ZnCl2-fluorescein mercuric acetate treatments required in the isolation technique originally described by Barland and Schroeder. However, our data do clearly demonstrate that the membrane isolation technique of Barland and Schroeder cannot be used to study the general composition of the plasma membrane.


Can regular solution theory be applied to lipid bilayer membranes? Direct measurement of the partition coefficient of n-hexane into phosphatidylcholine and phosphatidylcholine-cholesterol bilayers showed that (a) isotropic liquids are not good models for lipid bilayers and (b), Regular Solution Theory cannot, in general, be applied to lipid bilayer membranes at temperatures above their phase transition. Theoretical and experimental evidence given.


Phase properties of senescing plant membranes: role of the neutral lipids. Wide-angle X-ray diffraction studies have indicated that rough and smooth microsomal membranes from bean cotyledons acquire increasing proportions of gel phase lipid at physiological temperature as the tissue senesces. In addition, for both types of membrane the lipid phase transition temperature, defined as the highest temperature at which gel phase lipid can be detected, progressively rises with advancing senescence. Liposomes prepared from total lipid extracts of the membranes show a similar increase in transition temperature with age, indicating that separation of the polar lipids into distinct gel and liquid-crystalline domains is not attributable to peculiar protein-lipid interactions. Liposomes prepared from purified phospholipid fractions of the membranes show little change in transition temperature with age, indicating that the altered phase properties of the lipid do not reflect an increase in fatty acid saturation. However, the formation of gel phase lipid that occurs naturally during senescence can be stimulated by preparing liposomes from a mixture of the phospholipid fraction from young membrane and the neutral lipid fraction from old membrane. By adding the separated components of the neutral lipid fraction to purified phospholipid it was found that sterol esters and several unidentified lipids are able to raise the transition temperature of the polar lipids. Sterols have no effect on the phospholipid transition temperature. The data have been interpreted as indicating that several neutral lipids, which presumably increase in abundance with advancing senescence, induce a lateral phase separation of the polar lipids resulting in distinct gel and liquid-crystalline domains of lipid in the senescent membranes.


The membrane valve: a consequence of asymmetrical inhibition of membrane carriers. I. Equilibrating transport systems. Facilitated membrane transport systems act as valves, or rectifiers, when the substrate affinities on the two sides of the membrane differ substantially, i.e. when the system is strongly asymmetric. The asymmetry may be intrinsic or imposed by a reversible competitive inhibitor acting on only one side of the membrane. Under non-equilibrium conditions such systems allow net movements of substrate to proceed faster, sometimes much faster, in one direction than the other, though the final equilibrium is unaffected. Obligatory exchange systems may also function as valves when inhibited unsymmetrically, permitting exchange to occur more rapidly with one distribution of substrates than with the reversed distribution. Here, unequal flux rates do not depend on unequal concentrations of the substrate on either side of the membrane, but may also occur with equal concentrations, provided the affinities of the two substrates differ. The kinetic theory leading to these conclusions is given here, and it is shown how individual parameters of a carrier system affect the efficiency, or tightness, of the valve. In addition, simple kinetic tests for the operation of a valve are outlined. Examples are cited of transport systems having inhibitor-binding sites on only one surface of the cell membrane, which could function normally as valves. Systems implicated are glucose transport in various cells, the ADP-ATP exchanger of mitochondria, the anion transporter of erythrocytes, and the Na+-K+ pump.


Cell adhesion and spreading factor: chemical modification studies. The purified fetal calf serum factor that promotes cell adhesion and spreading of baby hamster kidney cells on tissue culture substrata has been subjected to a variety of chemical modifications and then tested for activity. These studies have shown that modification of the carbohydrate portions of the factor by glycosidic enzymes or by periodate oxidation did not alter its ability to promote cell spreading. On the other hand, modification of some protein portions of the factor by proteolytic enzymes or by specific modification of -COOH groups, tyrosine residues, or tryptophan residues resulted in a marked inhibition of factor activity. Modification of protein -SH groups, -NH2 groups, or methionine residues did not affect factor activity. Control experiments indicate that the various modifications were directed at the activity of the factor and not its adsorption onto the substrata.


Direct hydroxylation in the biosynthesis of hydroxy fatty acid in lipid a of Pseudomonas ovalis. It was found that Pseudomonas ovalis IAM 1177 had an abundance of hydroxy fatty acids such as 3-hydroxy-decanoic acid, 3-hydroxy-dodecanoic acid and 2-hydroxy-dodecanoic acid in the lipophilic part of the lipopolysaccharide fraction, which comprise 80% of total fatty acids. By using 18O2, it was shown that one oxygen atom from molecular oxygen was incorporated into 2-hydroxy-dodecanoic acid, but not into 3-hydroxy-decanoic acid. The incorporated oxygen atom was specifically located at the hydroxyl group of 2-hydroxy-dodecanoic acid. The biosynthetic pathways of these hydroxy fatty acids are discussed.


The effect of ethanol on glycerolipid biosynthesis by primary monolayer cultures of adult rat hepatocytes. This study evaluates the effects of ethanol exposure on glycerolipid production and release by hepatocyte monolayers. Glycerolipid formation from [1,3-14C]glycerol was increased in monolayers exposed to ethanol (1--50 mM) for 6 h. Monolayers exposed to 1 mM ethanol for 24 h also exhibited a rise in glycerolipid formation from either [1,3-14C]glycerol or [1-14C]palmitate; however, higher ethanol concentrations produced a dose dependent decrease in glycerolipid formation. Glycerolipids released into the medium by monolayers declined after all periods of ethanol exposure. The effects of ethanol on the enzymatic reactions involved in glycerolipid biosynthesis were determined in homogenates prepared from monolayers exposed to ethanol. Two enzymes, phosphatidate phosphohydrolase and glycerol kinase, exhibited ethanol-induced alterations in enzyme activity; however, only glycerol kinase activity correlated well with monolayer glycerolipid formation. These ethanol-induced alterations in enzyme activities and glycerolipid biosynthesis were reduced by simultaneously exposing monolayers to pyrazole or cycloheximide.


Alterations in lipid metabolism of developing muscle cells in culture. Embryonic chick muscle cells in the infused (myoblast) and fused (myotube) states differentially incorporate [3H]palmitic acid into their cellular lipids. Myoblasts incorporated [3H]palmitic acid 6--7-fold greater than myotubes in the triglyceride fraction. The endogenous triglyceride levels, however, were similar for the two developmental stages indicating increased triglyceride turnover in the myoblast compared to that of the myotube. Myotubes showed an increased incorporation of [3H]palmitic acid into sphingomyelin compared to myoblasts. Myoblasts and myotubes had a similar phospholipid composition. The data indicate that fusion is characterized by a decrease in triglyceride turnover possibly reflecting a shift from oxidative to glycolytic metabolism to coincide with the fusion process.


Influence of reduced cholesterol synthesis on the activity of cerebroside sulfotransferase in cultured glioblastoma cells treated with estradiol. Cultured glioblastoma cells were inoculated with estradiol in concentrations of 0.5--10 microliter/ml medium in order to check the effect of this hormone on the activity of cerebroside sulfotransferase, an enzyme whose activity is strongly related to myelination. Thereby we could show that the cerebroside-sulfotransferase activity increases to a value of 200% of normal. Concomitant to this effect, the cholesterol content of the membrane bearing cerebroside sulfotransferase activity decreases to 60% of normal. The effect is fully reversible: after 48 h, cholesterol synthesis as well as cerebroside sulfotransferase activity reach normal values again. We suggest that cerebroside sulfotransferase activity is modulated by the changing cholesterol/phospholipid ratio in the cells during the inoculation period.


Alteration of the lipase activities of muscle, adipose tissue and liver by rapeseed oil feeding of rats. Feeding rapeseed oil, rich in erucic acid, for 4 days results in a significant increase of the lipoprotein lipase activities of heart and adipose tissue. The lipase activity of liver, which in earlier studies has been shown to be releasable by heparin perfusion, also increases by the dietary regimen. The increased lipoprotein lipase activity of heart may contribute to lipid accumulation in this organ. The higher intracellular lipid store probably results in higher (hormone-sensitive) tissue lipase activity. The increase of lipoprotein lipase of adipose tissue, however, is not accompanied by an increase of hormone-stimulated tissue lipase activity in fat cells. This activity may even become lower, and might contribute to the decrease of the lipid store in heart after an initial rapid phase of fat accumulation during erucic acid feeding.


Cholesterol biosynthesis in a variety of cultured cells. Lack of correlation between synthesis and activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase caused by dexamethasone. In a variety of cell lines grown in the presence of 10(-6) M dexamethasone, the incorporation of acetate into cellular cholesterol was partially suppressed. However, the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (mevalonate:NADP+ oxidoreductase (CoA-acylating), EC 1.1.1.34) was either significantly elevated or remained unchanged providing evidence that under certain conditions 3-hydroxy-3-methylglutaryl coenzyme A reductase activity is not a valid measure of cholesterogenesis.


The effects of fatty acids and antioxidants in the culture medium on membrane composition and properties of the microsomal enzymes aryl hydrocarbon hydroxylase and cytochrome c reductase of cultured liver cells. Rat liver epithelial cells grown in medium without serum show characteristic changes in fatty acid composition and decreased levels of activity of the microsomal enzyme function aryl hydrocarbon hydroxylase. Manipulation of cell fatty acid composition was performed using albumin-fatty acid complexes. Both GLC analysis of fatty acid methyl esters and Arrhenius plots of enzyme activity indicate changes in membrane lipid composition without major changes in aryl hydrocarbon hydroxylase enzyme activity or inducibility by benzanthracene in the presence of butylated hydroxytoluene. When essential fatty acids were added without antioxidant, increased aryl hydrocarbon hydroxylase activity was found. These changes in microsomal aryl hydrocarbon hydroxylase activity were independent of changes in cytochrome c reductase. It is concluded that expression and induction of aryl hydrocarbon hydroxylase is not dependent upon cell fatty acid composition but may be enhanced under conditions of limited lipid peroxidation.


Purification and characterization of an agglutinin in the red alga Agardhiella tenera. The red alga, Agardhiella tenera was found to contain a glycoprotein which agglutinates mouse leukemia cells, L5178Y but not L1210. It also agglutinates guinea pig and rabbit erythrocytes, and has weak activity against human A, B and O, mouse, horse and sheep erythrocytes and hamster and mouse lymphocytes. The agglutination was not inhibited by simple sugars. The major active component was purified and determined to be a beta-structure protein containing 2.7% glucose as sugar moiety. The molecular weight was estimated to be 12,000 by gel filtration and 13,000 by polyacrylamide gel electrophoresis, in the presence of sodium dodecylsulfate. Its isoelectric point was 6.1, and it contained high amounts of glycine, serine and threonine, but no half cystine or histidine. It had no subunit structure, and the C- and N-terminal amino acids were threonine and arginine, respectively.


Interaction of C1-inhibitor with the C1r and C1s subcomponents in human C1. 1. Insoluble IgG-ovalbumin aggregates were used to bind and activate C1 from human serum. The bound C1 provided a useful reagent for studying the interaction of C1 subcomponents with C1-inhibitor. 2. C1-inhibitor bound to both subcomponents (C1r and C1s in C1 and formed stable complexes of respective apparent molecular weights 197,000 and 185,000, as determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The binding reaction proceeded more readily with C1s than with C1r and was correlated with the inhibition of C1s esterase activity. 3. At physiological ionic strength, binding of C1-inhibitor to subcomponents C1r and C1s caused release of these subcomponents from the C1-immune aggregates complex, indicating that C1-inhibitor binding decreased the inter-subcomponent binding forces in C1. At low ionic strength, however, this release did not occur.


Hemoprotein quantitation in isolated hepatocytes. Methods for quantitation of catalase, cytochromes P-450 and b5 and mitochondrial cytochromes a + a3, b561 + b566, and c + c1 in isolated hepatocytes were developed in analogy to methods established for subcellular systems and were used to measure changes in specific hemoprotein concentrations due to pretreatment and to change in incubation conditions. Pretreatment of rats with phenobarbital or 3-methylcholanthrene resulted in increased concentrations of cytochromes P-450 and b5 on a cellular basis, but had no effect on the other hemoproteins. Chronic ethanol pretreatment resulted in increased cytochrome P-450 and decreased cytochromes a + a3 concentrations. Hemoprotein concentrations in hepatocytes decreased following 4-10-h incubations in rotating round-bottom flasks. Rates of decrease were dependent upon both incubation conditions and prior in vivo treatments with phenobarbital or 3-methylcholanthrene.


Mechanisms of spectral shifts in lobster carotenoproteins. The resonance Raman spectra of ovoverdin and the crustacyanins. Resonance Raman data have been used to elucidate the mechanisms of the absorption spectral shifts occurring for astaxanthin upon binding to the carotenoproteins, ovoverdin and alpha-,beta- and gamma-crustacyanins, from the lobster Homarus americanus. Although distinguishable on the basis of small differences in their resonance Raman spectra the binding sites of the crustacyanins, giving rise to lambdamax at 605 +/- 25 nm, are essentially the same. The large red shift in lambdamax for the crustacyanins compared to free astaxanthin (lambdamax 480 nm), is accounted for by a charge-polarisation mechanism in which charged groups and possibly hydrogen bonds in the binding site set up pi electron polarisation in the ligand. Several alternate mechanisms can be eliminated. Ovoverdin is found to consist of three polypeptide chains of molecular weight 105,000, 95,000 and 78,000 which are not linked by disulfide bridges. The visible absorption peaks of ovoverdin at 460 and 640 nm are shown to arise from two astaxanthin molecules each bound at a different site. The spectral characteristics of the 460 nm site suggest a rigid hydrophobic environment for astaxanthin, in which no charge-ligand interactions occur. The mechanism of the spectral shift in the 640 nm site is the same as in the crustacyanins, i.e. a charge-polarisation effect. Resonance Raman spectra of ovoverdin and the crustacyanins could be obtained in situ; they were identical to the spectra of the purified proteins showing that the carotenoid sites were unperturbed by protein isolation.


Nitrogenase XI: Mössbauer studies on the cofactor centers of the MoFe protein from Azotobacter vinelandii OP. We have studied the MoFe protein from Azotobacter vinelandii OP with Mössbauer spectroscopy in applied magnetic fields up to 50 kG. The results are as follows. (1) The Mössbauer spectra of the S = 3/2 centers, which reside on the cofactor of nitrogenase, have been decomposed into six subcomponents. This suggests that each center contains 5-7, most probably 6, Fe atoms, thus confirming our earlier conclusions which were based on the quantitation of EPR data and on the assumption that the MoFe protein contains (30 +/- 2) Fe atoms. (2) Analysis of the high-field data shows that three subsites are characterized by a positive magnetic hyperfine coupling constant, A0, while A0 is negative for the other three sites. This observation demonstrates that the S = 3/2 centers are spin-coupled structures. (3) The zero-field splitting parameter D = +(6 +/- 1.5) cm-1 obtained from the Mössbauer data is in good agreement with our earlier EPR results, D approximately +5.5 cm-1. (4) The resolution of the Mössbauer spectra of the MoFe protein can be dramatically increased by employing Fourier transform deconvolution techniques. This allows a clear demonstration of spectral component S.


Local interactions as a structure determinant for protein molecules: II. Van der Waals interactions between sidechains are indicated to be important in determining the native state of the proteins of known structure by the following observations: 1. the average radial distribution of polarity increases continuously from the center of the molecule to its periphery. 2. nonpolar sidechains tend to occur in clusters. 3. the frequencies of long-range nearest-neighbor pairs are markedly non-random; each type of sidechain seeks nearest-neighbors of similar polarity. To investigate how these interactions affect the overall structure of the protein molecule, three simplified models are treated: a sheath-core model composed of independent residues, a modification accounting approximately for the connected nature of the chain, and a model consisting of three concentric spherical phases.


Local interactions as a structure determinant for protein molecules: III. Investigation of the known protein structures has led to the generalization that the native folding permits each sidechain to select those nearest-neighbors which maximize stabilization from van der Waals interactions. With regard to secondary structure: 1. Helical and beta regions exhibit characteristic patterns of short-range contacts (residue numbers k and k + t with [t] less than or equal to 4) due to the geometries of these secondary structures. However, these are not strictly obligatory, and preferred short-range contacts which would result in unfavorable van der Waals interactions are replaced by favorable long-range contacts. 2. The generalization mentioned at the outset holds for individual proteins, both for short-range and long-range contacts, and without regard for the type or amount of secondary structure present. 3. These observations imply that van der Waals interactions arising from short-range contacts partially determine secondary structure, and this is demonstrated by tests based upon assignment of regions of secondary structure in the known proteins. The principle of optimizing van der Waals stabilization from long-range contacts is applied to predict the structure of the complex formed by the S-peptide and S-protein of ribonuclease-S. The formation of favorable pairs is found to be more important than the total number of intermolecular contacts, and 40 to 50% of this stabilization is contributed by two residues of the S-peptide, Phe-8 and Met-13.


Enzymic synthesis of opioid peptides.

Enzymic synthesis of opioid peptides. Protected opioid pentapeptides were synthesized by the papain-catalyzed condensation of small synthetic fragments. These peptides, free as well as protected, were identical with those prepared by the dicyclohexylcarbodiimide-1-hydroxybenzotriazole method.


Molecular weights and Stokes radii of soluble elastins. Soluble elastin was isolated from lathyritic chick aorta using neutral salt solutions in the presence of beta-amino propionitrile. The effect of a carboxy-methylation step in conjunction with proteolytic inhibitors was investigated. Hydrodynamic (Stokes) radii of soluble elastins were measured by gel filtration and the molecular size and weight distribution in purified fractions are reported.


The limiting collagen microfibril. The minimum structure demonstrating native axial periodicity. Collagen fibers were grown from solutions of acid-soluble or neutral salt-soluble collagen in 0.5 M acetic acid by rapid dialysis. The collagen was obtained under conditions where protease inhibitors were present at every stage of extraction and purification. Under the conditions used, length-wise but not lateral filament growth proceeded rapidly and gel-like networks were formed, Water readily exuded from the networks. The networks were stretched to fibrous form during drying. Small-angle X-ray diffraction showed the stretched fibrils to be highly ordered, showing up to 20 orders of the 670 A meridional periodicity. Intermediate- and wide-angle photographs show equatorial reflections at a spacing corresponding to approximately 12.5 A which is related to the intermolecular distance but none related to a microfibrillar packing at the 35-40 A level. Electron microscopy of the gel networks before stretching shows the presence of thin filaments with diameters predominantly in the 35-40 A range. No cross-striated fibrils are seen in electron micrographs of either stretched fibers or unstretched fibers. Thus, intermolecular packing in accord with the 670 A axial periodicity can take place within approximately 40 A diameter thin filaments. These correspond to the structures previously postulated to be collagen 'microfibrils'.


Cell surface charge correlation of partition with electrophoresis. Critical mixtures of aqueous solutions of polymers separate into two or more immiscible phases. Particulate materials distribute in such phase systems generally between one bulk phase and the interface between bulk phases. The distribution is described by a simple partition law, and is quantitatively determined by, inter alia, the nature of the particle surface, particularly net electrical charge. The partition behaviour of various cells, native or modified by treatment with trypsin, neuraminidase or maleic anhydride, correlate strongly with electrophoretic mobility. Partition behaviour and electrophoretic mobility are both dependent upon cell surface charge. Thus, in appropriate conditions, changes in surface charge may be registered as changes in partition.


Evidence for the utilization of extracellular [gamma-32P]ATP for the phosphorylation of intracellular proteins in the squid giant axon. Proteins in the squid giant axon were labeled with 32P by in vitro incubation of isolated axoplasm with radioactive [gamma-32P]adenosine triphosphate (ATP) and separated by polyacrylamide sodium dodecyl sulfate gel electrophoresis. The two major phosphorylated regions on the gel had molecular weights of 400,000 and 200,000. These two peaks appear to be neurofilament proteins of squid axoplasm. The same set of proteins was phosphorylated in the axoplasm regardless of whether the [gamma-32P]ATP was applied in situ intracellularly or extracellarly. These results suggest that ATP in the extracellular space is, by some ATP-translocation mechanism, utilized in the process of intracellular phosphorylation. Measurements of the apparent influx of ATP across the squid axon membrane yielded results consistent with the view that ATP in the extracellular fluid could be transported into the axoplasm.


Isolation and new biological properties of Arion empiricorum lectin. The lectin present in the mucus of the snail Arion empiricorum was isolated by ion exchange chromatography. Purity was demonstrated by immunelectrophoretic analysis, immunization studies, and polyacrylamide gel electrophoresis. With the latter we found a molecular weight of 43,000. Hemagglutination inhibition studies revealed that carbohydrates play a minor role in the agglutination reaction of A. empiricorum lectin. Stronger inhibition could be achieved with human serum and the serum of several animal species. These findings were clarified by the demonstration that some serum proteins were precipitated by A. empiricorum lectin. Besides its agglutinating and precipitating properties the purified A. empiricorum lectin possesses proteinase-inhibiting properties, as demonstrated by the inhibition of casein-digestion by trypsin and plasmin.


I-cell disease: intracellular desialylation of lysosomal enzymes using an influenza virus vector. It has been proposed that I-cell disease results from a primary deficiency of acid neuraminidase activity. Infection by influenza virus of fibroblasts from a patient with I-cell disease resulted in the production of abundant intracellular alpha2-3 neuraminidase activity. Despite electrophoretic evidence of desialylation of intracellular and fibroblast-secreted arylsulfatase (EC 3.1.6.1) and beta-hexosaminidase (EC 3.2.1.30) from the infected cells, there was no consequent alteration of the abnormal distribution of beta-hexosaminidase activity between the intracellular spaces characteristic of I-cell disease. This suggests that deficiency of  alpha2,3 neuraminidase activity is not the primary biochemical defect in I-cell disease.


Tissue specific regulation of prostaglandin production: stimulation of 6-ketoprostaglandin F1alpha biosynthesis by rat kidney cytosol. Boiled cytosol of various rat tissues each affected prostaglandin biosynthesis by bovine seminal vesicle microsomes in a specific way. Kidney cytosol enhanced 6-ketoprostaglandin F1alpha production in a dose-dependent manner. This stimulatory effect was lost after dialysis. Liver, spleen and carrageenin granuloma cytosol inhibited 6-ketoprostaglandin F1alpha production but enhanced prostaglandin E2 production.


The spherule wall of Physarum polycephalum: chemical analysis and electron microscopy. 1. The composition of purified wall fragments from the spherule wall of Physarum polycephalum has been studied. 2. Spherule wall fragments consist of two major components. More than one-half of the wall material is glycoproteins. The remaining wall material after fractionation has a high carbohydrate to protein ratio and is suggested to be composed of proteoglycan(s). 3. The predominant monosaccharide constituent of the proteoglycan is galactosamine. The protein component is unique. It is composed almost entirely of phenylalanine (87--92 mol%). 4. Approx. 50% of these proteoglycans are resistant to alkali extraction and in negative stained preparation for the electron microscope consist of short spindle-shaped fibrils which measure 100 nm in length and 8 nm in diameter. These fibrils show a weak but distinct X-ray diffraction pattern.


Effects of prostaglandin E1 on the metabolism in rat parathyroid gland in vitro. We investigated some effects of prostaglandin E1 on the metabolism of rat parathyroid glands using a culture system containing basal Eagle's medium supplemented with 5--10% heat-inactivated rat serum. Rat parathyroid glands incorporate [3H]fucose and 14C-labeled amino acids into cellular glycoproteins and secrete some of these into the culture medium. Gel filtration chromatography separates these glycoproteins into three classes, the smallest of which (peak 3) is secreted with immunoreactive parathyroid hormone. In cultures of 48 h, prostaglandin E1 (1 microgram/ml) specifically inhibits the secretion of peak 3 and of parathyroid hormone but has no effect on the incorporation of [3H]fucose, 14C-labeled amino acids, or [3H]uridine into parathyroid glands. Cytochalasin B inhibits the secretion of parathyroid hormone and the incorporation of isotopic fucose and amino acids. Cortisol stimulates incorporation of [3H]fucose and the secretion of parathyroid hormone even in the presence of inhibitory doses of prostaglandin E1. It is concluded that, in organ culture, prostaglandin E1 inhibits the secretion of parathyroid hormone and of a specific glycoprotein the function of which may be related to the secretion of the hormone.


Levels of glutathione, glutathione reductase and glutathione S-transferase activities in rat lung and liver. Levels of glutathione, glutathione reductase and glutathione S-transferase activities in rat lung and liver have been investigated. After perfusing the lung to remove contaminating blood, this organ was found to have an apparent concentration of glutathione (2mM) which is approx. 20% of that found in the liver. Both organs contain very low levels of glutathione disulfide. Neither phenobarbital nor methylcholanthrene had a significant effect on the levels of reduced glutathione in lung and liver. In addition, the activities of some glutathione-metabolizing enzymes--glutathione reductase and glutathione S-transferase activity assayed with four different substrates--were observed to be 5-to 60-fold lower in lung tissue than in the liver.


Cell partition: a study of parameters affecting the partition phenomenon. Critical mixtures of aqueous solutions of polymers can be separated into two or more immiscible phases. Particulate materials are distributed in such phase systems generally between one bulk phase and the interface between bulk phases. The distribution is described by a simple partition law and is quantitatively determined by temperature, interfacial tensions, the electromotic potential difference between phases and the nature of the particle surface. The effects on transfacial potential differences of varying polymer, NaCl and sodium phosphate concentrations in dextran/polyethylene glycol systems were studied: increase of polyethylene glycol concentration increased the potential; addition of up to 40 mM NaCl progressively reduced the potential to zero: very small (less than 10 mM) concentrations of sodium phosphate buffer increased the potential, but further increase caused a fall in potential, which was less marked than that caused by equivalent concentrations of NaCl. The partition properties of a variety of cells, native or modified by treatment with trypsin, neuraminidase or maleic anhydride, were studied. In systems containing greater than 40 mM NaCl no difference in partition patterns for modifications of each cell type was observed. In systems containing no NaCl the partition pattern was highly dependent on the nature of the modification. From the behaviour of such models, polymer-electrolyte phase systems suitable for study of cell surface modification involving change, or no change, in net surface have been identified.


Induction of a microsomal butyrylesterase in rat liver by phenobarbital treatment. A carboxylesterase (carboxylic-ester hydrolase, EC 3.1.1.1) was induced in the liver of rats after repeated administration of phenobarbital. This enzyme migrated most rapidly towards the anode among Triton X-100-solubilized liver esterases by electrophoresis on the cellulose acetate membrane and it was tentatively designated as L-I. L-I increased in the microsomal fraction and was mainly concentrated in the fraction of smooth endoplasmic reticulum that proliferated after phenobarbital treatment. L-I- was separated from other liver esterases and purified about 800-fold. The purified L-I was homogeneous as judged by polyacrylamide gel electrophoresis and exhibited a molecular weight of about 55 000 as determined by gel filtration. L-I split the 1-naphthyl ester of butyric acid faster than esters of acetic, propionic or valeric acids; it therefore seemed to be a butyrylesterase.


Synthesis and removal of phenylalanine ammonia-lyase activity in illuminated discs of potato tuber parenchyme. (1) The synthesis and removal of phenylalanine ammonia-lyase (EC 4.3.1.5) in illuminated discs of potato (Solanum tuberosum cv King Edward) tuber tissue has been investigated by density labelling with deuterium (2H) from deuterium oxide (2H2O) followed by centrifugation to equilibrium in a CsC1 density gradient. (2) Temporal changes in enzyme level have been described in terms of the equation (dE/dt) = ks-kdE where (dE/dt) is the rate of change of enzyme level per unit of tissue (E) with respect to time (t), ks is the rate constant for synthesis of the enzyme and kd is the rate constant for the removal of active enzyme. (3) The optimal concentration of 2H2O was determined by analysis of the relationship between 2H2O concentration, development of enzyme activity and the magnitude of the increase in buoyant density of phenylalanine ammonia-lyase. A concentration of 2H2O of about 40% (v/v) was found to be optimal, allowing achievement of maximal or near maximal increases in the buoyant density of the enzyme without inhibition of the development of enzyme activity, thereby circumventing the major drawback of 2H2O as a source of density label. (4) The overlapping distribution profiles of enzyme activity after density gradient centrifugation were resolved by an iterative method of best fit which allows estimation of the proportions of pre-existing, unlabelled enzyme and newly synthesised, labelled enzyme at the end of the labelling period. This technique has been developed to obtain the rate constants for enzyme synthesis and for removal of active enzyme throughout the period of rapid change in enzyme level. (5) It is demonstrated that the initial rapid increase in phenylalanine ammonia-lyase activity in illuminated discs reflects an increase in the rate constant for enzyme synthesis in the absence of activation of pre-existing enzyme and in the absence of removal of active enzyme. The abrupt transition to a phase of decline in enzyme activity is caused by (a) a reduction in the rate constant for enzyme synthesis and (b) a dramatic increase in the rate constant for removal of active enzyme. The subsequent stabilisation of the enzyme is caused by decay of both rate constants to relatively low levels. (6) The results are consistent with hypothesis that rapid modulation of enzyme levels during tissue differentiation is achieved by simultaneous changesin the rate constants for both enzyme synthesis and for removal of active enzyme.


Partial purificationand characterizationof a factor which stimulates an increase in ornithine decarboxylase activity in the liver from tumor cell-free ascites fluid. A factor responsible for stimulating an increase in ornithine decarboxylase activity in the liver of mice was found in tumor cell-free ascites fluid of mice 3 days after inoculation of tumor cells. The factor was purified about 70-fold in 25% yield from tumor cell-free ascites fluid. As little as 1 microgram of protein of purified fraction, injected intraperitoneally into normal mice, significantly increased the activity of ornithine decarboxylase in the liver. The most active preparation of the factor formed two major protein bands on analytical polyacrylamide gel electrophoresis and both these bands stained with periodic acid-Schiff's reagent. The factor was a heat-labile, alkaline-stable, acidic protein with a molecular weight of more than 300 000. It was inactivated by treatment with 10 mM dithiothreitol, 5 M urea, pronase or mixed glycosidase, but was stable on treatment with DNAase, RNAase or neuraminidase.


Extracellular matrix metabolism by chondrocytes. 5. The proteoglycans and glycosaminoglycans synthesized by chondrocytes in high density cultures. Proteoglycans were extracted from the extracellular matrix of cultures of embryonic chick chondrocytes grown at high density and were purified by CsC1 density gradient centrifugation. The chemical, physical and hyaluronate binding properties of the proteoglycans were similar to those observed in proteoglycans from other hyaline cartilages. Proteoglycans in the media were also purified and on analysis showed three populations of proteoglycans to be present. One population had the physical characteristics of a typical proteoglycan subunit and bound hyaluronate, the other two populations were unable to complex with hyaluronate but one had the physical characteristics of the proteoglycan subunit and the other was of smaller molecular weight. The small molecular weight appears to be a product of the enzymatic degradation of the larger molecular weight species.


Possibility of mitochondrial-cytosolic cooperation in gluconeogenesis from serine via hydroxypyruvate. Intracellular localization of D-glycerate dehydrogenase (D-glycerate: NAD+ oxidoreductase, EC 1.1.1.29), one of the enzymes of the pathway for gluconeogenesis from serine via hydroxypyruvate, was studied by differential centrifugation. Almost all enzyme activity was found in cytosol. Since the major activities of two other enzymes, serine: pyruvate aminotransferase (EC 2.6.1.51) and glycerate kinase (ATP: D-glycerate 2-phosphotransferase, EC 2.7.1.31), of the pathway via hydroxypyruvate are localized in mitochondrial inner membrane and/or matrix, the possible localization of D-glycerate dehydrogenase in mitochondria was examined. Detailed analysis of mitochondrial fraction prepared by differential centrifugation indicated that rat liver mitochondria do not contain any D-glycerate dehydrogenase activity. Based on these results, a cooperative connection between mitochondria and cytosol in gluconeogenesis from serine via hydroxypyruvate is proposed. Possible mechanisms for transport of intermediates of the pathway via hydroxypyruvate across the mitochondrial membranes are also discussed.


Preparation and characterization of antibody to galactosyl(alpha 1 leads to 4)galactosyl(beta 1 leads to 4)glucosylceramide. Antiserum against galactosyl(alpha 1 leads to 4)galactosyl(beta 1 leads to 4)glucosylceramide (globotriaosylceramide, Gb3) was raised in rabbits by the administration of four weekly intramuscular injections of 1.5 mg of the purified glycoplipid along with bovine serum albumin and Freund's complete adjuvant. AntiGb3 activity was quantitated initially by immunoprescipitation employing Gb3 mixed with 100-fold excess of lecithin and cholesterol (1 : 1 or 1 : 2, by wt.) as antigen. Subsequently, complement fixation tests done with antigen preparations containing Gb3/lecithin/cholesterol (1 :6 :20, by wt.) showed antiGb3 titres of up to 1 : 8192. Fractionation of the antiserum by BioGel A5m chromatography indicated the antibody was an IgM immunoglobulin. The partially purified antibody stimulated complement-dependent release of glucose from glucose-containing liposomes prepared with shingomyelin/cholesterol/dicetylphosphate/Gb3 (molar ratio, 100 : 75 :11 :1). The antibody crossreacted with the trisaccharide, Gal(alpha1 leads to 4)Gal(beta1 leads to 4)Glc, but not with galactosylerceramide, lactosylceramide, GM1 ganglioside, globotetraosylceramide, digalactosyldiglyceride or a number of low molecular weight saccharides.


The molecular size of glycans liberated by hydrazinolysis from Semliki Forest virus proteins. The glycans of well characterized, [6-3H]galactose-labelled glycopeptides, GC-4 from bovine IgG1 as well as GP-V-2 and GP-V-5 from alpha1-acid glycoprotein, were liberated by hydrazinolysis. Molecular weights close to the expected values were observed by gel filtration. Desialated glycans of Semliki Forest virus proteins were likewise liberated by hydrazinolysis and subjected to gel filtration. Metabolically labelled [1-3H]galactose-oligosaccharides of the mixed viral proteins revealed an apparent molecular weight of 1800. The bi-antennary glycan liberated from the reference glycopiptide GC-4 was of 1750 daltons. A mixture of [2-3H]mannose-labelled E1- and E2-proteins of the virus contained L-type glycans of 1800 daltons (formerly called A-type), and M-type glycans of 1200 daltons (formerly called B-type). A fraction of the E3-glycans isolated by affinity chromatography on Concanavlin A-Sepharose showed an average molecular weight of 2150, a value intermediate between the three- and four-antennary glycans liberated from the reference glycopeptides GP-V-5 and GP-V-2. The rest of the E3-glycans were of 1850 daltons, a value close to the bi-antennary GC-4 glycan. We suggest that the comparatively large size of the E3-glycans and the exposed position of E3-proteins on the viral surface may be interrelated.


Starvation-induced changes in secretin-like immunoreactivity of human plasma. Levels of secretin-like immunoreactivity in the plasma of 50 starved subjects were measured by radioimmunoassay and rose from 18 +/- 3 (S.E.) pg/ml after 12 h, to 103 +/- 12 pg/ml (P less than 0.005) after 36 h. The assay antibodies were found to be specific for a region of secretin located towards the C-terminal residue. Lactoperoxidase was used to label the secretin with 125I and ionexchange chromatography on SP-Sephadex C-25 was used to purify the labelled product. The plasma immunoreactivity was purified by immunoaffinity chromatography on antibody-Sepharose conjugates and characterised by gel-filtration on Sephadex G-50 calibrated with molecular weight markers. After a 12-h fast, 10-20% of the immunoreactivity had a molecular weight of about 12 000, possibly due to precursors of secretin. Most of the remainder was smaller than secretin with molecular weight of less than 3000. This material comprised over 90% of the plasma immunoreactivity after a 36-h fast and may be due to degradation products.


A comparison of the glucocorticoid receptor in cytosol from rat liver and hippocampus. The [3H]corticosterone- and [3H]dexamethasone-binding proteins in cytosol from liver and hippocampus of the rat were compared by isoelectric focusing analysis in slabs of polyacrylamide gel. A single peak of radioactivity with a pI of 6.1--6.2 was obtained during analysis of cytosol from both liver and hippocampus using either corticosterone or dexamethasone as radiolabelled ligand, provided the tissue was carefully perfused with buffer prior to preparation of cytosol. Rat serum or insufficiently perfused tissue contained a corticosterone-binding component with pI of 5.2--5.5 representing corticosteroid-binding globulin. Limited trypsin digestion resulted in fragmentation of the dexamethasone- and corticosterone-binding protein in cytosol from liver and hippocampus. Incubation of radiolabelled cytosol with 0.5 microgram of trypsin/A280--310nm of cytosol gave a sharp radioactive peak with a pI of 5.9--6.1 when analyzed by isoelectric focusing; when 5.0 microgram of trypsin/A280--310nm of cytosol was used, a double peak with pI values of 5.9--6.1 and 6.3--6.5, respectively, was seen. The same trypsin-induced peaks were seen with both [3H]dexamethasone and [3H]corticosterone as ligands. The substrate specificity and the sensitivity of this glucocorticoid binder for limited trypsin digestion is in good agreement with what was previously found for the glucocorticoid receptor in rat liver cytosol. It is concluded that cytosol from liver and hippocampus contains an identical or very similar receptor for glucocorticoid hormones.


Partial characterization of [3H]methyltrienolone binding in rat prostate cytosol. 3H-labelled R 1881 (methyltrienolone) was bound with high affinity (Kd, 2 . 10(-9) M) and low capacity (11--15 fmol/mg protein)in prostate cytosol from rats castrated 18 h earlier. The binding was inhibited by 5alpha-dihydrotestosterone, testosterone, cyproterone acetate and LEO 1727, but not by estradiol-17beta, R 5020 or corticosterone. The 3H-labelled R 1881-receptor complex was excluded from a Sephadex G=200 column, had a sedimentation coefficient of 3.5 S, was eluted from a DEAE-cellulose column at 0.2 M KCl concentration and precipitated between 20--40% of saturation of ammonium sulfate. 20% of specifically bound 3H-labelled R 1881 was retained by DNA-cellulose at low ionic strength. The fraction which was not bound to DNA-cellulose did not bind either, after rechromatography, not even following heating at 20 degrees C for 20 min or following fractionation with ammonium sulphate. In summary, the results indicate that the 3H-labelled R 1881-receptor complex in rat prostate cytosol has characteristics similar to those of the 5alpha-[3H]dihydrotestosterone-receptor complex.


A neural network model for the development of direction selectivity in the visual cortex. A neural network model is proposed to explain the development of direction selectivity of cortical cells. The model is constructed under the following three hypotheses that are very plausible from recent neurophysiological findings. (1) Direction selectivity is developed by modifiable inhibitory synapses. (2) It results not from the direct convergence of many excitatory inputs from LGN cells but from cortical neural networks. (3) Direction-selective mechanism is independent of orientation-selective mechanism.--The model was simulated on a computer for a few kinds of inhibitory connections and initial conditions. The results were consistent with neurophysiological facts not only for normal cats but for cats reared in an abnormal visual environment.


Mathematical analysis of optimal multichannel filtering for nerve signals. This paper extends recent analyses (Roberts and Hartline, 1975; Oğuztöreli and Stein, 1977) of optimal linear filters for separating neural signals from more than one electrode site. Roberts and Hartline's result, using a matched filter criterion, represents one of a class of optimal filters with different, but symmetrical, output waveforms derived by Oğuztöreli and Stein (1977). Another narrow bandwidth filter of this class will give the optimal results according to an energy criterion, but may be less useful in practical situations.


Analysis of recall and recognition in a certain class of adaptive networks. A general model of adaptive networks which perform recall is analyzed in view of qualitative psychological findings. The assumptions underlying the basic model are few and general in the sense that no specifications of structure or mechanisms of adaptation are imposed. The analysis of the model is towards the addition of various features drawn from the global input-output relations expected from the networks. The memorizing process of recognition is found to be intrinsic to the model, and four of the most prominent relations between the performance of recall and recognition are shown to either exist in model or to be realizable by means of few additional plausible features.


Additional comments on the use of Wiener filtering for the estimation of evoked potentials. The problem of average potentials filtration in the case of periodic stimulation is discussed in the paper. Some considerations proving the correctness of Wiener filtering processes suggested by Doyle are presented here. The basic assumption of the proof is an approximation of the noise power density by means of a step function.


Human psychophysics: functional interpretation for contrast sensitivity versus spatial frequency curve. The hypothesis that neural processing in the human visual pathways compensates for both optical degradation as well as noise contamination at the photoreceptor level is introduced and shown to be consistent with the high frequency portion of the contrast sensitivity function for threshold detection of sinusoidal gratings in addition to the suprathreshold phenomenon of matching sinusoidal gratings of different spatial frequencies. This offers a unifying interpretation for why, at threshold conditions, the high spatial frequency portion of the image is blurred as severely by the nervous system as it is by the optics (e.g. Campbell and Green, 1965) while in extreme suprathreshold conditions the nervous system effectively deblurs the image (e.g. Georgeson and Sullivan, 1975; Kulikowski, 1976). These conclusions do not necessitate a highly specific form of visual processing such as Fourier channeling.


[Hematopoietic organs in mice following a single administration of hydrocortisone]. Kinetics of the number and direction of CFU differentiating in mice after a single infusion of hydrocortisone in a dose of 5 mg per mouse was studied using the method of an exogenous cloning of the stem cells in the spleen and the bone marrow of the irradiated recipients. Against the background of the long-term involution of the thymicolymphoid apparatus a change of the number and direction of the CFU differentiating occurred. Along with this, CFU concentration in the spleen and bone marrow of mice remained constant. Under the hormone action CFU differentiating capacity in the spleen and bone marrow in the direction of erythro- and myelopoiesis sharply changes; CFU of the bone marrow behaves as CFU of the spleen, and CFU of the spleen behaves as CFU of the bone marrow of normal mice. It is suggested that the described effects of the CFU of the spleen and bone marrow of mice given hydrocortisone were due to the redistribution of T-lymphocytes and not to direct cytotoxic effect of hydrocortisone on the CFU population.


[Course of ether anesthesia and the post-anesthesia period against a background of prior vibration]. The experiment on albino rats showed that three-hour vibration (46.6 Hz, 1mm amplitude) decreases the functional activity of the central nervous system as well as that of the respiratory and circulatory apparatus. Ether anesthesia against this background aggravates the animals condition and they die during the post-anesthesia period (60% of cases). Survival of the animals under conditions of extreme actions (vibration, anesthesia) is estimated by the cerebral cortex resistance level.


[Effect of corazole in intensifying changes in the electrical activity of the brain in a constant magnetic field]. Corazol effects on the background and evoked biopotentials of the cerebral cortex, hypothalamus, and the cerebellar cortex were studied in rats exposed to a constant magnetic field (500, 1000, 4000 Oe). After the corazol pretreatement with subconvulsive doses there were sharp changes in the background rhythms; evoked potentials of the amplitude rose markedly, and there was also an increase of additional phases in its structure. Intensification of the constant magnetic field effect against the background of analeptic corazol action pointed to the importance of the central nervous system activation level in the response to the magnetic field action.


[Mechanism of action of eserine on DNA synthesis in rat liver]. Eserine (1-2 mg/kg) decreased incorporation of the radioactive precursors into total DNA of the rat liver by 40-50%. Only high eserine cincentrations (10-5 M) effected directly the DNA synthesis in the hepatocyte nuclei. It was shown in vitro that the DNA synthesis in the nuclei of hepatocytes, isolated from the liver of rats after eserine administration (1 mg/kg), decreased significantly. A conclusion is drawn that the eserine action on the incorporation of radioactive precursors into DNA of the rat liver in vivo is localized at the level of the DNA synthesis in the nuclei. The study of the acetylcholine action on the DNA synthesis in the isolated nuclei of hepatocytes suggests that the inhibitory effect of eserine on the DNA synthesis is mediated through acetylcholine.


[Changes in hemodynamic and endocrine-metabolic parameters in the early post-resuscitation period]. Changes in the system hemodynamics, renal and hind limb blood flow, as well as some endocrine-metabolic parameters were studied in 24 dogs which survived 17-minute circulatory arrest. As revealed, relative compensation and normalization of several functions and metabolic parameters in the course of 1 hour after reanimation were followed by a new wave of disturbances developing unsimultaneously, irregularly and gradually in the whole organism and some peripheral tissues. Thus, disturbances of the peripheral circulation and of the central hemodynamics served as one of the basic pathological symptoms of postreanimation disease.


[Detection of antigen-antibody reactions using optical beat spectroscopy]. The light beating spectroscopy technique was used for recording the results of serological reactions without visual phenomena. The investigated systems consisted of the antigens of influenza virus or Mycoplasma pneumoniae and the corresponding sera. A principal possibility of detecting the antigen-antibody complexes in serological reactions without visual phenomena by the light beating spectroscopy technique was demonstrated.


[Cytotoxic effect of BCG-activated macrophages on tumor target cells in vitro]. The macrophage-activating effect of fluid and lyophilized BCG vaccines and killed BCG was studied. Activation of murine peritoneal macrophages was determined by their cytotoxic action in vitro on the syngeneic tumour 51Cr-labeled target cells. Murine macrophages obtained 2 to 3 weeks after a single injection of the fluid BCG vaccine exerted a cytotoxic action on the tumour cells. A single injection of the lyophilized BCG vaccine or killed BCG caused no macrophage activation. Two injections of fluid or lyophilized BCG vaccines caused macrophage activation earlier--3 to 5 days after the second injection. Intraperitoneal BCG injection induced more macrophage activation than subcutaneous.


[Transplacental blastomogenic effect of p-hydroxyphenyl lactic acid]. Transplacental administration of tyrosine metabolite p-hydroxyphenyl lactic acid having blastomogenic activity induced development of malignant and benign neoplasms (leukemias and lymphosarcomas, adenomas and pulmonary carcinoma, hepatomas, papillomas, and cancer of the urinary bladder and other tumours) in progeny of CC57BR and C57BL mouse strains, in 88 and 78% of cases, respectively. The number of neoplasms was significantly higher, than in the control, they appeared earlier and were of more malignant character. The role of endogenous blastomogenic substances in the development of leukemias in the newborn and yound children is discussed.


[Autoimmune nature of spermatogenesis disorders in varicocele models in rats]. Partial decrease of venous drainage from the left testis in male rats induced development of destructive changes in the seminiferous epithelium at the site of operation and in the contralateral organ. Spermatogenesis disturbances in the rat testes (focal desquamation of seminiferous epithelium, disorganization and degeneration of germ cells, devastation of seminiferous tubules) resembles the lesion in men with varicocele. These data permit to consider the results of present experiment as varicocele modelling. Disturbance of permeability and of the fine blood-testis barrier structure in both testes was observed in experimental rats; the picture of morphological changes was similar to that of autoimmune orchitis; lymphocytes sensitized to spermatozoa antigens were revealed in the lymphoid organs of experimental rats. The results obtained permit to suggest the involvement of immunological mechanisms in the development of pathological changes in the testes affected with varicocele.


[Adrenergic innervation of the capillaries]. Adrenergic nerve fibers of the capillaries devoid of the smooth muscle coat were revealed by Falk's method in the mesentery and the kidney fibrous capsule of dogs after 6 daily noradrenaline infusions and with cardiogenic shock developing against this background.


[Determination of the alveolar dead space]. The author suggests a method of finding the alveolar dead space (AIDS), i.e. the part of the functional dead space (FDS) which presents the greatest difficulties for determination. A1DS determination is based on the difference in the CO2 concentration in the venous blood flowing to the lungs for oxygenation under conditions of respiration at rest, and of CO2 concentration in the alveolar portions of  expriation following 10-sex breath withholding at the level of the usual inspiration. The mean A1DS volume (VDA1) in % to the alveolar part of the tidal volume for 20 healthy persons in sitting position and in 12 of them in lying position was found: VDA1 (in sitting position)= = 8.0 +/- 1.5% (P = 0.95), sigma = 3%; VDA1 (in lying position) = = 5.0 +/- 0.7% (P = 0.95), sigma = 2.6%. It is suggested that FDS determined by this method be used to judge the efficacy of using the inspired air in the lung ventilation, to determine AIDS in the diagnosis of lung embolism, to judge the pulmonary circulation shunt by the difference between the value found according to Bohr's formula for FDS and the FDS value established according to the suggested formula.


Changes in the contractile activity of the rabbit myocardium during burn shock. As revealed on the papillary muscles of the rabbit heart in vitro, contractile function of the myocardium proved to be disturbed in 50% of cases an hour after the infliction of burn to these animals. This could serve as one of the causes of a sharp cardiac output derangement found in these experiments.


[Pathogenetic factors in experimental photogenic epilepsy]. Male albino rats were used in chronic experiments. The photogenic epilepsy syndrome was induced by local tetanus toxin injection into the lateral geniculate blody (LGB) which caused the formation of a pathologically enhanced excitation. The primary symptoms of photogenic epilepsy resulted from the pathologically enhanced specific sensory excitation in the LGB, and correlated with a significant increase of the evoked potential amplitude in the corresponding visual cortex field. The following progress of the neuropathological syndrome was connected with generalized disturbances of the rhythmic electrical activity of the brain, characterizing the state of increased epileptic sensibility. The experimental results demonstrate an important role of specific and nonspecific factors in the pathogenesis of experimental photogenic epilepsy.


Normal iron absorption and decreased red cell iron uptake in the aged. Absorption of iron was studied with a double-isotope technique that allowed differentiation between "mucosal uptake," "mucosal transfer," and ultimate "retention" of iron. A physiologic dose of ferrous sulfate was administered to 25 healthy young adults, 40 active aged persons, and 20 patients with uncomplicated iron deficiency. Radioactivity was measured with a whole-body scanner. Iron absorption values were not decreased in aged subjects compared to young adults. Mucosal uptake, mucosal transfer, and retention of iron were equally increased in both young and old patients with iron deficiency. In 12 young adults and 33 aged persons red cell iron uptake was studied in addition to iron absorption. Young adults utilized 91% of the retained, orally administered iron and the aged only 66%. An increase in ineffective erythropoiesis in old age is suggested.


Quantitative anisocytosis as a discriminant between iron deficiency and thalassemia minor. The coefficient of variation (CV) of red cell size, as measured by electronic red cell sizing (erythrography), was less than 14.0% in 20 normal subjects. In 22 of 25 patients with beta-thalassemia minor and microcytosis (mean corpuscular volume [MCV] less than 70 fl), CV was less than 14.0%; in the other 3, CV was 14.0%--14.9%. In 53 patients with iron deficiency anemia and MCV less than 70 fl, CV always was greater than 14.0%. In 7 patients with alpha-thalassemia minor and MCV less than 70 fl, CV was less than 14.0% in all 7. Among patients with microcytosis, erythrography appears to be an excellent technique for rapidly distinguishing between iron deficiency and alpha or beta thalassemia minor.


Inhibition of granulocyte erythrophagocytosis by HLA antisera. The ability of HLA antisera to inhibit granulocyte erythrophagocytosis (EP) of opsonized red blood cells (RBC) was evaluated. Human granulocytes (PMN) were separated from heparinized whole blood by the Ficoll-Isopaque technique and suspended in McCoy's medium. EP occurred when the PMN were incubated with opsonized RBC. For six HLA antibody specificities evaluated, prior incubation of PMN with HLA antisera resulted in  significant inhibition of EP without cytolysis when the PMN donor was positive for the specific HLA antigen, but not when the donor was negative for the antigen. The inhibition was time- and dose-dependent. Prior absorption of HLA antisera with HLA-specific platelets reduced or abolished the inhibition. An example of anti-NB1 also inhibited EP in 4 individuals. These data suggest that HLA antibody may adversely affect granulocyte phagocytic function. Inhibition of EP might be useful in evaluating compatibility prior to granulocyte transfusion.


Reduction of salivary tissue factor (thromboplastin) activity by warfarin therapy. The coagulant of normal human saliva has been identified as tissue factor (thromboplastin, TF) by virtue of its ability to cause rapid coagulation in plasmas deficient in first-stage coagulation factors and to activate factor x in the presence of factor VII and by virtue of the fact that its activity is expressed only in the presence of factor VII and is inhibited by an antibody to TF. The TF is related to cells and cell fragments in saliva. Salivary TF activity has been found to be significantly reduced in patients taking warfarin. The decline in TF activity during induction of warfarin anticoagulation occurs during the warfarin-induced decline in vitamin-K-dependent clotting factor activity, as judged by the prothrombin time. The decrease in TF activity is not related to a reduction in salivary cell count or total protein content or to a direct effect of warfarin on the assay. It is hypothesized that the mechanism by which warfarin inhibits TF activity may be related to the mechanism by which it inhibits expression of the activity of the vitamin-K-dependent clotting factors. Inhibition of the TF activity may be involved in the antithrombotic effect of warfarin.


Treatment of acute nonlymphocytic leukemia: use of anthracycline-cytosine arabinoside induction therapy and comparison of two maintenance regimens. Patients with acute nonlymphocytic leukemia were given remission induction therapy consisting of cytosine arabinoside and an anthracycline. Those patients who experienced complete remission received two courses of consolidation therapy and were randomized to receive maintenance therapy consisting of either daily chemotherapy with reinforcements every 3 mo or reinforcement therapy only every 6 wk. The overall complete remission rate was 66%, with 80% complete remission for previously untreated patients less than 60 yr of age who did not have a prior history of malignancy. Remission durations were the same for patients treated with both maintenance regimens. The major determinant for successful remission induction therapy was patient age, with older patients frequently succumbing to intercurrent infection. Documented leukemic cell resistance to the therapy employed was only rarely encountered. Once remission was achieved, age was no longer a determinant of patient survival, since duration of remission was independent of age. Remission durations were directly related to leukemic cell retention of cytosine arabinoside triphosphate. Hence therapy for acute nonlymphocytic leukemia can be divided into two separate areas: remission induction and remission maintenance.


Erythrocyte cellular and membrane deformability in hereditary spherocytosis. In order to determine whether the relative rigidity of the hereditary spherocytosis (HS) red cell is due to membrane rididity or merely to an altered surface/volume ratio, we investigated the deformability of resealed red cell membranes from patients with HS. Whereas the osmotic fragility of intact red cells of HS patients showed the expected increase, the osmotic fragility of resealed HS membranes was normal, thus indicating that their surface/volume ratio was normal. Measurements with an ektacytometer showed that deformability of intact HS cells was markedly diminished, whereas deformability of resealed HS membranes was normal. These findings indicate that the HS red cell membrane is not intrinsically abnormally rigid, as has been suggested, but that the lack of deformability of the erythrocyte is primarily a function of the altered surface/volume ratio.


Oxidative metabolic responses of rabbit pulmonary alveolar macrophages. During phagocytosis of opsonized lipopolysaccharide-coated paraffin oil droplets, rabbit alveolar macrophages reduced nitroblue tetrazolium, which effect was in part inhibitable with the use of superoxide dismutase. Exposure of cytochalasin-B-treated rabbit alveolar macrophages to opsonized zymosan led to the generation of superoxide, as quantitated by ferricytochrome C reduction. It was found that nitroblue tetrazolium in the presence of ferricytochrome C could in turn serve as scavenger of superoxide during stimulation of cytochalasin-B-treated rabbit alveolar macrophages. Following challenge with either opsonized zymosan or the membrane perturbant digitonin, rabbit alveolar macrophages released hydrogen peroxide into the extracellular medium. Employment of the surface membrane stimulant phorbol myristrate acetate led to activation of the hexose monophosphate shunt, which activity could be further enhanced in the presence of superoxide dismutase or attenuated in the presence of catalase. These studies demonstrate that rabbit alveolar macrophages release superoxide and hydrogen peroxide during surface membrane perturbation. In turn, hydrogen peroxide generation can stimulate the hexose monophosphate shunt.


Protective effect of prophylactic penicillin on splenectomized mice exposed to an aerosolized suspension of type III Streptococcus pneumoniae. Prophylactic penicillin has been recommended for use in asplenic patients and postsplenectomy patients. A laboratory model using aerosolized pneumococci has been devised to test the effectiveness of prophylactic penicillin in a manner analogous to human experience. There is increased mortality, over time, in asplenic mice exposed to aerosolized type III Streptococcus pneumoniae. One hundred twenty-one male Swiss mice (mean weight 26 g) were divided into four groups: splenectomized, sham-operated, splenectomized + penicillin, and sham-operated + penicillin. After 2 wk the four groups were exposed for 30 min to an aerosolized atmosphere of 2.4 x 10(9) colony-forming units of type III S. pneumoniae using a Tri-R model A-42 airborne infection apparatus. Penicillin was given at a daily intramuscular dosage of 40,000 units procaine penicillin G beginning 2 days prior to exposure and continuing through the third day after exposure. The splenectomized and sham-operated mice given penicillin showed significantly lower mortality (p less than 0.001) than mice not given penicillin.


Splenectomy in myeloid metaplasia.

Splenectomy in myeloid metaplasia. Between 1960 and 1977, 50 patients with agnogenic myeloid metaplasia were splenectomized. Twenty-five of 26 patients with painful splenomegaly, 4 of 9 patients with refractory hemolytic anemia, 4 of 10 patients with refractory thrombocytopenia, and 4 of 4 patients with portal hypertension showed significant benefit from the procedure. There were five immediate postoperative deaths. Four of these deaths occurred early in our series of splectomies for myeloid metaplasia before 1970. Only one death has occurred in the last 21 patients operated on. Survival following splenectomy averaged 25.5 mo.


Hemoglobin biosynthesis in individual bursts in culture: studies of human umbilical cord blood. We cultured human umbilical cord blood erythropoietic precursors in methyl cellulose clonal assay and analyzed the synthetic rates of Hb A and Hb F in individual erythropoietic bursts. Hemoglobin was labeled with 14C-amino acids in culture, separated by slab gel isoelectric focusing techniques, and quantitated by fluorographic methods. Almost all bursts exhibited both Hb A and Hb F in varying ratios. Frequencies of the individual bursts differing in percentage Hb F biosynthesis had normal distributions. Natal erythropoietic precursors appeared to be randomly committed to Hb F synthesis.


Sodium in blood vessels. A brief review. Extracellular Na can be accurately partitioned into free and bound components; the latter component is directly related to the hydration and stiffness of the vascular wall. Cell Na can be similarly partitioned: [Na+]i is about 10mM and the transmembrane gradient about 15:1 in a physiological medium. The transmembrane gradient remains constant over a wide range of change in [Na+]0. Although active Na transport is largely responsible for developing the transmembrane ion distribution pattern, the membrane potential depends mainly on movements of K+ and C1-. No certain direct role can be assigned to Na+ in short-term vasoconstriction, but an indirect role through the regulation of catecholamine release and uptake is now established. There is evidence that the entry of Na+ into cells plays a fundamental role in the protein-synthesizing activity of the vascular smooth muscle cell and is thereby involved in the longer term determination of peripheral vascular resistance.


Analytical study of ionized or ionizable groups of platelet membrane. The knowledge of the molecular surface groups of the human blood platelets present a particular interest especially in some thrombotic processes or thrombocytopathies. The aim of this study was to approach quantitatively ionized or ionizable groups of the platelet membrane using liquid phase electrophoresis or physicochemical and analytical methods with specific reagents. A mean repartition of the principal groups (carboxyl, SH, amino and phosphate) on the membrane is obtained by cell electrophoresis.


Congenital factor VII deficiency. A report of four new cases. Four new cases with congenital homozygous factor VII deficiency are described. Factor VII levels were reduced to less than 1%, 3%, 8% and 10%, respectively. The incidence and severity of bleeding symptoms were well correlated with the measured factor VII activity. In the severe case of factor VII deficiency (less than 1%) a home treatment program was started because of severe recurrent hemarthroses. This entailed transfusions of 20 U/kg body weight prothrombin complex or factor VII concentrate in case of acute bleeding approximately every three weeks. These transfusions have been carried out successfully without any problems. In contradiction, two brothers with hypoproconvertinemia (factor VII 8% and 10% respectively) reached an age of more than 70 years. Despite replacement therapy postoperative bleeding followed one appendectomy, whereas no postoperative bleeding followed patients requiring Achilles tendon lengthening and an above knee amputation and only slight bleeding followed a tonsillectomy. Based on our experience we suggest that in patients with factor VII deficiency of less than 10%, when undergoing surgery, should be maintained a minimal factor VII activity of 10--15% during the first three postoperative days.


Capacity of the mucosal transfer system and absorption of iron after oral administration in rats. 1. The dose dependence of the iron absorption shows a saturation characteristic if iron (0.25--5 mumoles Fe/kg body weight) is administered in tied-off intestinal segments of normal and iron-deficient rats. 2. If the iron doses (2.1--570 mumoles Fe/kg body weight) are administered by stomach tube only in normal rats a similar dose dependence curve has been obtained as after administration in tied-off intestinal segments. In iron-deficient rats the shape of the dose dependence curve is changed and shows no clear-cut saturation characteristic. 3. The differences of these dose dependence curves are discussed with respect to the differences of the methodological conditions. A saturation type kinetic for absorption cannot be expected under all experimental conditions despite of the existence of a transfer system with limited capacity for the transport of iron.