A volatile factor inducing transmissible lysis in Gaeumannomyces graminis (Sacc.) Arx and Olivier var. tritici Walker. Filtered water extract of Gabalong soil with a recent history of take-all in wheat caused lytic plaques to form in agar cultures of a virulent strain of Gaeumannomyces graminis var. tritici. The plaques resembled those produced by Bdellovibrio on plate seeded with bacteria. However, there was no evidence of the presence of bacteria, viruses, or mycoplasmas. The lytic factor was transmissible in culture filtrates to fresh subcultures of the fungus. Exposure of young healthy colonies to sublethal doses of ultraviolet light also induced transmissible lysis. The lytic factor was heat-stable, passed through a 25-nm filter, and was not affected by nuclease (enzymes) or severe irradiation with UV light. It also induced bysis in several other strains of G. graminis. Lysis was always preceded by a growth-stimulatory effect on the fungus. The lytic factor was active as a volatile chemical which induced transmissible lysis and continued to be formed, apparently as a self-perpetuating agent, in lysing cultures of the fungus.


Some effects of petroleum on estuarine and marine microorganisms. Degradation of mixed hydrocarbon substrate in a system comprising water from an environment relatively free of oil and a sediment inoculum from an oil-contaminated site was significantly greater than when sediment from the non-oil-contaminated environment served as inoculum. Mixed hydrocarbon substrate, however, was observed to have a limiting effect on the growth of autochthonous bacteria from the non-oil-contaminated estuarine source. Growth and cell yield were similarly reduced when marine sediment bacteria were cultured in seawater supplemented with mixed hydrocarbon substrate. The addition of a South Louisiana crude oil or a No. 2 fuel oil to water and sediment collected from a marsh area of Chesapeake Bay showed no limiting effects on growth of the total heterotrophic microbial flora when examined over a 28-day period. However, results of these studies indicate that the effects of petroleum on microoganisms should be examined carefully under conditions closely approximating those in situ.


Measurement of bacterial and fungal contributions to respiration of selected agricultural and forest soils. A technique using selective inhibitors was used to estimate the relative contributions of bacterial and fungal populations to the respiration of six soils and one litter sample. The ratios of bacterial to fungal respiration in the four agricultural soils, given in percentage of the total microbial activity, ranged from 10/90 to 35/65, with the average ratio being about 30/70. In the forest soils, the ratios were 20/80 and 30/70, and in a beech litter sample, the ratio was 40/60. The fungi clearly dominated in all samples. The ratios were not found to be pH related. The difficulties which had previously limited the use of selective inhibitors for in situ soil ecological investigations, such as insufficient inhibitor specificity, inhibitor inactivation or degradation, and errors of measurement caused by elimination of competitor populations, were either resolved or methodologically avoided in the experiments. Inhibitor selectivity was demonstrated using both mixed and pure cultures of microorganisms from each soil. Through the use of experiments with short incubation periods (6-8 h), problems with population shifts and inhibitor degradation were eliminated.


Variation in the activity levels of selected enzymes of Erwinia amylovora 595 in response to changes in dissolved oxygen tension and growth rate of D-glucose-limited chemostat cultures. Chemostat cultures of Erwinia amylovora 595, grown in mineral salts-nicotinic acid medium at 30 degrees C, and limited by D-glucose concentrations in the presence of dissolved oxygen tensions (D.O.T.) greater than about 6mm Hg, became limited by oxygen availability below about 4 mm Hg. This latter limitation was accompanied by a marked increase in acid production as the D.O.T. was depressed. The transition between D-glucose- and oxygen-limitation was also characterized by a maximum in succinate oxidase activity, and a minimum in the in situ respiration. D-Glyceraldehyde-3-phosphate dehydrogenase and D-fructose-1, 6-diphosphate aldolase showed small reductions in specific activity in the region 4-6 mm Hg D.O.T., but further reduction to 2 mm Hg resulted in a marked increase in the specific activity of aldolase. Malate dehydrogenase followed the converse trend, and attained very low activity levels when the D.O.T. decreased beyond the lower limits of detection. The in situ respiration was maximal at 2 mm Hg D.O.T., while potential respiration values were minimal at 2 mm Hg, and maximal at about 8 mm Hg D.O.T. The insitu respiration rate was proportional to dilution rate (D), in presence of excess oxygen, up to 0.18 h-1, after which a marked diminution occurred and continued until the wash-out rate was attained. Succinate oxidase activity decreased with increase in dilution rate, but remained constant above D equals 0.18 h-1. Malate dehydrogenase showed a persistent decline with increase in dilution rate, while D-glyceraldehyde-3-phosphate activity increased somehwat at higher dilution rates. The data are interpreted in terms of two transition points, at 6 and 2 mm Hg D.O.T., and of a change from respiratory to fermentative metabolism at low D.O.T., and at high dilution rates.


A nutritional and taxonomic survey of Arthrobacter soil isolates. One hundred and sixty Arthrobacter soil isolates, four groups of 40 from each of four soil types, and 17 Arthrobacter named strains were characterized on the basis of 203 different nutritional tests performed on each culture. One hundred and twenty-one compounds were examined as sole sources of carbon, 44 compounds as sole sources of nitrogen, 22 carbohydrates for the production of acid, and 16 tests were done for tolerance to various dyes. The four groups of soil arthrobacters differed markedly in acid production from carbohydrates, and in utilization of aliphatic amino acids and aromatic hydrocarbons as sole sources of carbon. The named strains did not exhibit as much nutritional diversity as any of the four groups of soil isolates. The results of the nutritional tests were subjected to a computerized taxonomic analysis. Eighty-four of the isolates were contained in nine separate clusters with 87 to 94% similarity; 39 isolates were contained in five mixed but recognizable clusters with 84 to 86% similarity; 60 isolates were randomly mixed with no recognizable clusters and similarity of 72 to 83%. All clusters were composed of nearly equal numbers of isolates from each of the four soils; the named strains were not found associated with any of the clusters of soil isolates. The results indicated that the diversity of the genus Arthrobacter is not represented by the named species already described, and the possibility of using a series of standardized tests to categorize soil arthrobacters into readily identifiable nutritional groups is discussed.


A cluster analysis of some bacteria in the water column of Green Lake, Washington. Mass inoculation and computer clustering techniques were used in an abbreviated procedure to group similar bacteria isolated from four depths in the water column of Green Lake, Washington. Four groups of bacteria were differentiable and were catagorized as orange-yellow Flavobacterium-Cytophaga, yellow Flavobacterium-Cytophaga, Vibrio-Aeromonas-, and Pseudomonas-like. Some of the groups were most prevalent in certain sample depths in the water column.


Reversal of fungitoxicity of 8-quinolinols and their copper (II) bischelates. I. Amino acids and related compounds. The effect of amino acids and related compounds on the toxicity of 8-quinolinols and their copper (II) bischelates to Aspergillus oryzae (ATCC 1011) Was studied. None of the compounds tested except the thiol-containing compounds, cysteine, cysteamine, glutathione, and N-acetylcysteine reversed the inhibitory action of 8-quinolinol but not that of 5-iodo-8-quinolinol or bis (8-quinolinolato) copper (II). It appears that the mechanism(s) of fungitoxicity of 8-quinolinol is different from that of 5-iodo-8-quinolinol or bis(8-quinolinolato) copper (II.).


Interpreting the rate of survival in carcinoma. Carcinomas originate in one organ, spread away from that site, cannot be accurately staged, and require total ablation for cure. Cure is achieved when all of the tumour is located within the tissue ablated by the method used. The rate of survival indicates the proportion of patients with such localized disease. A comparison of survival rates is not a comparison of therapeutic accomplishments; it is a comparison of the cumulative localization of the tumours in different series. The superior method is that which ablates the primary tumour and the largest amount of surrounding expendable tissue at the earliest time. Selecting the method according to this principle rather than by a comparison of survival rates provides the highest prospect for cure. That is the purpose of treatment.


Fractures of the medial humeral epicondyle in children. A review of 50 children with fractures of the medial humeral epicondyle facilitated a simple classification of this injury and its management. The "apparent isolated" fracture of the medial humeral epicondyle is uncommon; it is more frequently associated with elbow dislocation, with or without spontaneous reduction, at the time of injury. Analysis of the results of treatment of medial humeral epicondylar fractures in this group of children showed that, in the case of those with an "apparent isolated" injury, treatment with a sling only gave good results in 90% whereas poorer results followed open treatment or fractures associated with dislocation. Closed treatment is therefore recommended. Open reduction is indicated only with (a) ulnar neuritis at the time of injury or (b) intra-articular fragment.


Effects and prevention of frostbite in wound healing. Observations on wounds sustained at subfreezing temperatures in husky dogs and in man suggested that exposed wet tissues readily become frozen. Frostbite of wounds caused tissue necrosis, wound sepsis and delayed healing, but immediate wound suture protected against this sequence. To explore the effect of wound closure on healing frostbitten tissue, healing of paired dorsal wounds was studied in 20 rats. In each animal one wound was sutured and the other was left open. In 10 animals both wounds were frozen with Dry Ice. All sutured wounds healed primarily. Most of the control open wounds had healed at 15 days, but in the frozen wounds healing was delayed and infection ensued. It is suggested that at temperatures much below freezing the immediate treatment of a wound should include prevention of frostbite by wound closure.


Osteoid osteoma and osteoblastoma: reclassification of 43 cases using Schajowicz's classification. Forty-three cases of benign osteoblastic tumour of bone from the University of British Columbia bone tumour registry have been reviewed and reclassified according to the classification of Schajowicz and Lemos. Their classification has the advantage that the relationship of the two lesions is recognized and overlap, both radiologically and microscopically, is allowed for. The terminology used in the new classification, however, is cumbersome and confusing and appears to be of no great advantage. We therefore recommend instead the terminology modified from that of Dias and Frost, which incorporates the observations put forth by Schajowicz and Lemos: (a) cortical osteoblastoma, (b) medullary osteoblastoma, (c) periosteal osteoblastoma and (d) multifocal osteoblastoma.


Vascular compartment syndromes.

Vascular compartment syndromes. In a 5-year period (1968 to 1973) 33 cases of vascular compartment syndrome were seen. Seven case reports illustrate various etiopathogenetic factors in the development of vascular compartment syndrome. A review of the 33 cases has permitted discussion of the pathophysiology and clinical features of these syndromes, together with introduction of a clinical classification on which treatment is based. Mild cases may be treated by application of ice, elevation, and observation; for severe cases fasciotomy is the treatment of choice. Two techniques of fasciotomy are available: multiple skin incisions with fasciotomy between the incisions for single-compartment sydnromes and extensive skin incisions over the length of the fasciotomy for multicompartment syndromes or severe single-compartment syndromes.


Esophageal reconstruction: an experimental approach to the control of reflux after esophageal resection. A study of the properties of replacement gastric tubes and colonic segments, and their use in the prevention of reflux after esophageal resection indicates that, in order to prevent reflux, these tubes must be maintained in a subdiaphragmatic position. Gastric tubes have a higher intrinsic pressure barrier than colonic tubes--a 2.5- to 6-cm segment prevents reflux and the tube maintains a pressure barrier 10 cm H2O higher than stomach presure, whereas colonic segments require 12 cm of subdiaphragmatic length to control reflux and maintain a pressure barrier only 2 cm H2O above gastric pressure. Removal of the intrinsic pressure barrier by myotomy allows free reflux in tubes that previously had prevented reflux.


Methymethacrylate cement: its curing temperature and effect on articular cartilage. The increasing use of methylmethacrylate in orthopedic surgery makes it desirable to determine whether the curing temperature of methylmethacrylate is high enough to cause bone necrosis and whether methylmethacrylate damages articular cartilage. Studies in dogs showed that methylmethacrylate plut temperatures ranged from 95 to 107degrees C and that temperatures at the cement-bone interface ranged from 50 to 95degrees. Curing temperatures therefore are high enough to cause necrosis of bone. Other studies revealed no harmful action of methylmethacrylate on articular cartilage. Fractures stabilized with methylmethacrylate can therefore be expected to heal provided the normal criteria of fracture management are fulfilled.


Pacemaker infections: a 12-year review.

Pacemaker infections: a 12-year review. In a 12-year period, 546 pacemaker-related operations were complicated by 23 infections (infection rate, 4.2%). Most of these infections were treated successfully, but two cases of infection were fatal. The most frequently encountered organism was Staphylococcus epidermidis. In 14 cases, infection occurred despite the preoperative administration of cloxacillin to which the organism was sensitive in 13 of the cases. The skin flora constitute the most likely source of contamination. The ideal treatment consists of removal of all foreign material and replacement with a new pacing system by an alternative route.


Gastrocolic fistula complicating benign gastric ulcer: case report and review of literature. A review of the literature and report of a case of benign gastrocolic fistula indicate that epigastric pain is the most frequent early symptom. Later, the predominant symptoms are diarrhea, weight loss and feculent vomiting. Patients suffering from rheumatoid arthritis and taking steroids appear to be particularly at risk of developing a gastrocolic fistula. Barium enema is the most reliable method of demonstrating the fistula. Preferred management is a one-stage gastrocolic resection and primary anastomosis.


[Emphysematous cholecystitis: presentation of two clinical cases]. Emphysematous cholecystitis is relatively rare: in 1971 the cumulative total of cases reported in the world literature was 115. Two cases seen in the period 1967 to 1972 can be added. The features of these two cases, and those described in the literature support the view that emphysematous cholecystitis results from a primary obliterative endarteritis and a secondary proliferation of microorganisms.


Inuit myopia: an environmentally induced "epidemic"? Among Inuit less than 30 years old the prevalence of myopia is far in excess of that of their elders. This is especially true for females. There seems to be little, if any, genetic contribution to this "epidemic" of myopia in the young. The age and sex distribution indicates the likelihood of an environmental factor, probably cultural, being responsible for the current pattern. Other data implicate school attendance as a possible etiologic factor.


Electrocardiographic changes during exercise in asymptomatic men: 3-year follow-up. Electrocardiographic (ECG) changes during maximal bicycle exercise and risk factors for coronary heart disease (CHD) were studied in 510 male civic employees who were followed for 3 years. Clinical CHD developed in 15 (24.6 percent) of the 61 men with an ischemic exercise ECG on the initial examination and in 11 (2.4 percent) of the 449 subjects with a normal initial exercise ECG. A normal maximal exercise ECG is no guarantee that severe CHD does not exist and that a subject will not soon sustain major myocardial damage; and an ischemic exercise ECG does not necessarily indicate underlying CHD. In the former group angina was the most frequent clinical CHD episode; in the latter group, infarction. Among those with an abnormal initial exercise ECG, CHD was most likely to develop in association with a poor exercise capacity. Subjects with subsequent clinical CHD and those with abnormal ECGs after 3 years tended to have a higher frequency of risk factors; subjects whose abnormal ECGs reverted to normal after 3 years tended to have a lower frequency of risk factors.


Massive transplacental hemorrhage: clinical manifestations in the newborn. Thirteen newborn infants had transplacental hemorrhage in excess of 30 ml. Fetal blood in the maternal circulation was demonstrated in all cases by the acid elution technique. Anemia was noted in five babies either at birth or within the first 24 hours of life. One baby was stillborn, the death possibly being related to fetal hemorrhage. The other seven babies were clinically normal in spite of massive transplacental hemorrhage. The hemoglobin values and reticulocyte counts were normal at birth and the first 5 days of life. The data on this group of babies suggest that the clinical manifestations of transplacental hemorrhage are related not only to the size of the hemorrhage but also to the time at which the hemorrhage occurs.


Blood glucose in intoxicated chronic alcoholics. Chronic alcoholics may present with hyperglycemia or hypoglycemia. Because alcohol induces glycogenolysis, chronic alcoholics usually have higher blood glucose values than do nonalcoholic subjects. In a prospective study of blood glucose concentration in 201 chronic alcoholics, blood alcohol concentration, sex, weight, type of beverage consumed and time since last eating were not generally associated with lower blood glucose values. The infrequency of hypoglycemia in ambulatory chronic alcoholics may reflect the relatively ready availability of hostels, detoxification centres and hospitals in large cities. It is, however, important to be aware of the possible occurrence of hypoglycemia in chronic alcoholics, particularly when community facilities for the chronic alcoholic are not available.


Abdominal and endometrial actinomycosis associated with an intrauterine device. Actinomycotic endometrial infection associated with an intrauterine device (IUD) complicated chronic abdominal inflammatory disease in a 28-year-old woman. Colonies of organisms with morphologic resemblance to and staining reactions of Actinomyces israelii were observed in tissue adherent to the IUD and in inflamed omental and pericolic tissues. However, the organism could not be cultured. Because intact tissues are resistant to actinomyces it is likely that the IUD created an environment favouring the establishment and growth of the organism.


Self-conversion of drug-resistant paroxysmal atrial tachycardia. A 56-year-old woman had paroxysmal atrial tachycardia (PAT) that was recurrent, drug-resistant and required frequent electroversion. The mode of onset and termination of initiated episodes of PAT suggested an AV junctional reentrant mechanism. The problem was successfully managed by a permanent electrode implanted transvenously in the coronary sinus and connected to a subcutaneous radio frequency receiver. For the past 24 months the patient has successfully terminated all episodes of PAT (at least 60) by activating an external radio frequency transmitter that transmits impulses to the implanted receiver and electrode catheter. Conversion to sinus rhythm requires 5 seconds or less of pacing.


A combined modality approach to the treatment of Hodgkin's disease. Preliminary results of a prospectively randomized clinical trial. Eighty-seven previously untreated patients with pathologic Stage IA, II (A or B), or IIIA Hodgkin's disease were randomized over a 48-month period to receive either megavoltage extended field radiotherapy alone or megavoltage radiotherapy limited to involved lymph node sites (including at least an upper mantle field) followed by combination chemotherapy with nitrogen mustard, vincristine, prednisone, and procarbazine (MOPP). Four patients (4.6%) failed to achieve remission with initial radiotherapy. Seventy-two evaluable patients have currently completed therapy. Ten of 41 patients achieving remission with radiation alone have relapsed, compared to only 1 of 31 receiving radiation plus chemotherapy. Seven patients have died, 3 of whom failed to achieve remission with initial radiotherapy. The other 4 had Stage IIIA disease treated with radiation alone. Severe myelosuppression occurred infrequently during chemotherapy, and neither serious infections nor second neoplasms have observed. Although these preliminary results are encouraging, longer followup is required to determine the ultimate effects of combined modality therapy on survival and long-term complications.


Hepatic artery ligation and postoperative chemotherapy for hepatic metastases: clinical and pathophysiological results. Nineteen patients underwent hepatic artery ligation for metastatic cancer of the liver. Metastases were from colon cancer in 13 and from other primary cancers in 6. Postoperative chemotherapy was given to 15 by the systemic route, and to 1 by infusion into the ligated hepatic artery. Sixteen patients survived the procedure. In these patients, the mean survival of those with metastases from colon cancer was 27+ (5-72) weeks, and from other cancer 50+ (8-23) weeks. Among the patients evaluated, 57% had symptomatic improvement, 43% showed improvement in the liver scan, and 33% had a decrease in liver size. Our experience suggests that hepatic artery ligation can offer temporary but significant palliation in carefully selected patients.


Biological markers in breast carcinoma. I. Incidence of abnormalities of CEA, HCG, three polyamines, and three minor nucleosides. Patients with breast carcinoma were screened for abnormal concentrations of CEA, HCG, putrescine, spermidine, spermine, pseudouridine, N2, N2-dimethylguanosine, and 1-methylinosine. Abnormal polyamine levels occurred in less than 15% of the patients. Among the nucleosides, N2, N2-dimethylguanosine was the most frequently abnormal, occurring in 57% of the patients with metastatic disease. CEA levels were abnormal in 30% of postoperative N+ patients and 74% of patients with metastatic disease, while HCG elevations were found in 45% and 50%, respectively. All the patients with one or more marker abnormalities could be detected by measuring only CEA, N2, N2-dimethylguanosine, and HCG. Among these three tests, a singular marker abnormality occurred in 35.8% of the patients, and all three tests were abnormal in 21.8% of the patients. The performance of these three tests in each patient revealed one or more abnormalities 97% of the patients with metastatic disease, and 67% of the postoperative N+ patients.


An effective low-dose intermittent cyclophosphamide, methotrexate, and 5-fluorouracil treatment regimen for metastatic breast cancer. A low-dose, three-drug regimen, C.M.F. (cyclophosphamide 50 mg, p.o., days 1-14; methotrexate, 25 mg, and 5-fluorouracil, 500 mg, i.v., days 1 and 8; cycled every 28 days) was used in 46 consecutive chemtherapy-eligible women (41 previously hormonally treated) with recurrent breast cancer. Thirteen percent of the patients had complete regressions (C.R.); 33% had partial regressions (P.R.); 26% stabilized; and 28% progressed. In evaluating response by sites of metastases, lymph nodes (30%), lung nodules (22%), and subcutaneous deposits (2/3) had the highest incidence of C.R.; 46-71% of patients with lymph node, lung, subcutaneous, liver, breast, or peritoneal disease showed C.R. or P.R. Skin and pleural disease responded in 30% of patients whereas no patients had radiographic healing of bony metastases. The toxicity was minimal: 7% gastrointestinal, 26% marrow-suppressive, and 7% infectious. This low-dose C.M.F. regimen resulted in regression resulted in regression rates similar to higher dose C.M.F. protocols, which use approximately twice these drug dosages with commensurate toxicity.


Adriamycin plus vincristine compared to and combined with cyclophosphamide, methotrexate, and 5-fluorouracil for advanced breast cancer. The preliminary results of a controlled study with two independent combinations in metastatic breast cancer are reported. The first combination (Therapy A: 41 patients) consisted of Adriamycin (ADM) and vincristine (VCR), while the second combination (Therapy B: 41 patients) included cyclophosphamide (CTX), methotrexate (MTX), and 5-fluorouracil (FU), designated "CMF." Both treatments were administered as intermittent cycles in patients previously untreated with chemotherapy. After eight cycles, responsive patients on Therapy A were crossed over to Therapy B to avoid the risk of cardiomyopathy. In both groups, crossover was carried out at the time of progression or relapse. In the group given ADM plus VCR, the dominant site of disease was in soft tissues in 56%, in viscera in 22%, and in bones in 22%. These findings were present in 51%, 24%, and 24%, respectively in the group started on CMF. The comparison of response after primary treatment in patients receiving a minimum of two cycles failed to show a significant difference between Therapy A and Therapy B (overall response 58% vs. 65%). When the response rate was calculated only in patients who had completed the first eight cycles of therapy, these findings were 87% and 93%, respectively. Three patients receiving treatment A (8%) AND 4 patients given Treatment B (10%) achieved complete remission. The highest incidence of response was observed for soft tissue lesions (70% vs. 76%). However, complete or partial bone recalcification was seen in 33% and 24%, respectively. The duration of response was found to be longer in patients who crossed over to CMF after eight cycles of ADM plus VCR, as compared to those started and continued on CMF. No cross-resistance was observed after crossover for progressive disease. Both incidence and degree of side effects were found acceptable, and no drug-related death was seen. Virtually all patients were treated on an outpatient basis.


Systemic chemotherapy of advanced head and neck malignancies. Several Phase II chemotherapy protocols were evaluated in patients with advanced malignancies; 158 were evaluable head and neck cases. The protocols were as follows: five-drug combination (COMFP), four-drug (COMF), (CCNU, Adriamycin, DTIC, and cytosine arabinoside. Insufficient numbers and data were received to adequately evaluate Yoshi 864, 5 Azacytidine, porfiromycin, BCNU, and Azaserine. Significant responses to therapy were noted in the four and five-drug combinations in which 30-44% of the patients had 50% or greater regression, with an average duration of 2.2 months. Adriamycin and CCNU demonstrated lesser antitumor effects, while DTIC and cytosine arabinoside did not demonstrate significant antitumor activity in the head and neck areas. Usual toxicity consisted largely of nausea and vomiting, leukopenia, and thrombocytopenia. Alopecia was not pronouced in Adriamycin-treated patients. It appears that combination chemotherapy had a higher response rate compared to single agents used in the different cooperative protocols.


A phase II study of methyl CCNU in the treatment of solid tumors and lymphomas: a Southwest Oncology Group study. In March of 1972, the Southwest Oncology Group initiated a Phase II study, No. 7200, utilizing methyl-CCNU in the treatment of patients with solid tumors and lymphomas. Initially, they received 200 mg/m2 orally as a single dose every 6 weeks. The dose was subsequently reduced in poor-risk patients to 150 mg/m2. There were 69 responses noted in 675 evaluable patients (10%). The highest response rates were noted in patients with Hodgkin's disease (13/31, 35%), malignant gliomas of the brain (8/29, 28%), anaplastic carcinomas of the lung (5/20, 25%), and squamous cell carcinomas of the head and neck (5/29, 17%). Squamous cell tumors appeared to be more responsive than adenocarcinomas (15% vs. 5%, respectively). Hematologic toxicity was cumulative, and was influenced by dose and prior treatment. There appeared to be no cross-resistance in patients previously treated with alkylating agents. Methyl-CCNU is an active antineoplastic agent. Further studies are indicated in order to determine relative effectiveness.


The efficacy of combining radiation therapy with a surgical procedure in patients with cervical metastasis from squamous cancer of the oropharynx and hypopharynx. To determine whether combining radiation therapy and a surgical procedure was superior to either modality alone, the authors reviewed the records of 345 patients who had primary cancers of the base of the tongue, tonsillar fossa, or pyriform sinus associated with clinically positive nodes at the time of their first observation. Combining the two modalities of therapy produced a local and regional control of the disease in twice as many patients as did one modality alone. The absolute 5-year survival did not reflect this same trend, however, since as local and regional control increased, more patients lived longer but died before five years from distant metastases. Immunochemotherapy is suggested as a possible adjunct therapy to prevent distant  metastases.


Colonic neoplasms complicating ureterosigmoidostomy. A benign reactive polyp developed at each ureteral implantation in the sigmoid colon following bilateral ureterosigmoidostomy for exstrophy of the bladder. The first polyp was discovered 23 years after the operation. The second polyp occurred 4 years later at the other ureteral orifice. Each polyp contained benign cystic glandular epithelium within the submucosa and the stalk. These submucosal cysts resembled colitis cystica profunda, except that they were confined to the pedunculated polyps. A complete review of the literature of neoplasms complicating ureterosigmoidostomy is presented. It is concluded that a benign or a malignant colonic tumor may develop at the site of the ureterocolic anastomosis many years later. Therefore, patients who undergo this operation should be followed for the rest of their lives.


Risk of second tumors in survivors of childhood cancer. A new malignancy was diagnosed in 19 of 414 long-term survivors of cancer in childhood. All but two lesions were attributable to prior radiotherapy; eight were successfully treated. Excluding 4 patients ascertained in connection with the second malignancy, there were 15 in the series who developed a new cancer, in contrast to 0.7 cases expected (p less than 0.001). The 20-year (5-24 years after initial diagnosis) cumulative probability of a second cancer was 12% (S.E. 4%), and the radiation-related cancer rate was 1.8 cases in exposed tissues per million person-years per rad. Host susceptibility may have had an etiologic role, but an oncogenic effect of chemotherapy was not demonstrable. Another 13 study patients developed benign tumors. These findings emphasize the importance of long-term surveillance of children with cancer.


Association of asbestos and bronchogenic carcinoma in a population with low asbestos exposure. Quantitative counts of ferruginous bodies were performed on digests of lungs from 100 control and 30 lung cancer patients. It was found that the lung cancer group had significantly higher levels, although only 1 patient was known to be occupationally exposed to asbestos. It is suggested that even extremely low levels of asbestos exposure may have a carcinogenic effect.


Incidence of cancer in patients with leprosy. Individuals with congenital immunodeficiences and patients who are immunosuppressed for maintenance of organ allografts experience a marked increase in the occurrence of malignancy. Patients with lepromatous leprosy also have depressed cellular immunity, but or study of 195 autopsied subjects with leprosy did not reveal an increase in the occurrence of cancer. Thirty-three of the 195 subjects with leprosy or 16.9% died of cancer, which is comparable to an age-matched group of individuals.


Insulin goes metric: a time for reveiw.

Insulin goes metric: a time for reveiw. For most diabetics, a concentration of 100 units of insulin per cc (U-100 insulin) is the simplest and safest concentration. It is compatible with the metric system and reduces the volume of the injection. The introduction of U-100 gives the nurse an opportunity, not only to review her own knowledge of insulin use in diabetes, but also to assess the diabetic individual's understanding of it.


Project Alternative: the road away from isolation. The author describes why therapeutic social groups for long-term psychiatric patients sometimes don't work, and what makes them successful when they do. Project Alternative resulted in reduced rehospitalization, and staff were excited with the new life-style their clients adopted.


The effect of aging and interval between primary and secondary treatment in two-stage carcinogenesis on mouse skin. Two-stage carcinogenesis experiments on mouse skin (female ICR/Ha Swiss mice) were done by initiating mice at three age levels (6, 44, and 56 weeks) and promoting after a 2-week interval. In another series, mice were initiated at age 6 weeks, and three time intervals (2, 36, and 56 weeks) were used between initiation and promotion. The initiating agent was 7, 12-dimethylbenz(a)anthracene and the promoting agent was phorbol myristate acetate in all experiments. The results showed a general decrease in tumor production with increasing age at the time of promotion. However, the initiating effect persisted even when the interval between initiation and promotion was 56 weeks.


DNA synthesis in tumor-bearing rats.

DNA synthesis in tumor-bearing rats. Thmidine (TdR) incorporation into DNA increased in the livers and spleens of rats bearing Yoshida sarcoma (solid type) or AH130 (solid type). TdR kinase and DNA polymerase activities increased in the serum, liver, and spleen of these rats, while thymidine monophosphate kinase activity increased appreciably only in the liver and spleen. On diethylaminoethyl cellulose column chromatography, 2 peaks of TdR dinase activity were separated from the serum and tumor tissues of rats bearing Yoshida sarcoma (solid type) while only 1 peak was obtained from the liver. TdR kinase activity in the serum decreased abruptly 7 hr after removal of the Yoshida sarcoma, while that in the liver decreased more slowly.


The influence of thyroid stimulation on the incidence of 3-methylcholanthrene-induced tumors. The incidence of tumors induced by a single s.c. injection of 2 mg of 3-methylcholanthrene was observed in euthyroid and thyroidectomized rats and in rats fed thyroid powder. Thyroid feeding increased the metabolic rate by a factor of 1.6 and reduced the incidence of tumors at the site of injection from 92% in euthyroid rats to 36% in hyperthyroid rats.


Subcellular fate of protein antibiotic neocarzinostatin in culture of a lymphoid cell line from Burkitt's lymphoma. 14C-LABELED PROTEIN ANTIBIOTIC NEOCARZINOSTATIN (NCS) was prepared efficiently by chemical modification. With the use of lymphoma-derived cell line P3HR-1, the subcellular behavior of this antitumor antibiotic was studied by the uptake a.  nd autoradiography of isolated nuclei of radioactive NCS. The antibiotic was taken up by the cells, reaching the maximum value at 1.5 hr and decreasing in value at 4.0 hr to the level at 0.5 hr. The silver grains in the autoradiograms were also found in the isolated nuclei. The grain count in the nuclei showed a tendency similar to the uptake of NCS by the whole cells, i.e., a gradual increase at 0.5 hr, reaching the maximum value at 1.5 hr, and then decreasing after 4.0 hr to the level at 0.5 hr. These facts indicated that NCS reached not only to cytosol but also into the nucleus, and/or at least to the nuclear membrane of the lymphoid cell. The number of NCS molecules incorporated into the cells at 1.5 hr was calculated to be about 1 x 10-6/cells at a concentration of 3 mug NCS per ml of medium, which can be extrapolated to 1 x 10-4 molecules per cell at the minimum inhibitory concentration. The number of molecules should be even less within the nucleus. In cell-free systems, the interaction of DNA and NCS, which is an inhibitor of DNA synthesis, was investigated with use of a Sephadex G-100 column, with negative results. In the cell culture system, NCS molecules were degraded into smaller polypeptides of certain sizes by proteolysis either by serum component(s) or by cells themselves. An inactive isomer, pre-NCS, which is an antagonist of NCS and a partially denatured homologous molecule, behaved similarly to NCS in all of these experiments. Because the chemically modified NCS used in this study retained biological activity essentially similar to that of parental NCS, the results obtained here could be interpreted as similar to those of parental NCS in vitro.


2-chloroethanol formation as evidence for a 2-chloroethyl alkylating intermediate during chemical degradation of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea and 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea. Chemical degradation of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea or 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea in buffer under physiological conditions resulted in the formation of a significant quantity of 2-chlorethanol (18 to 25% of the initial nitrosourea concentration). Other degradation products observed included acetaldehyde (5 to 10%), vinyl chloride (1 to 2%), ethylene (1 to 2%), and cyclohexylamine (32%), but not 1,3-dicyclohexylurea. The 2-chlorethyl moiety of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea was trapped with halide ions, CI-, BR-, and I-, to form the corresponding dihaloethanes which were identified by gas chromatography-mass spectrometry techniques. High-pressure liquid chromatographic procedures were developed for the separation and quantiation of the nitrosoureas and many of their degradation products. It is postulated that a new mode of 1(2-chloreoethyl)-3-cyclohexyl-1-nitrosourea and 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea degradation can occur that is not the loss of the chloro group as chloride ion, but the loss of the N-3 hydrogen as a proton. Then the corresponding isocyanate and 2-chloroethyidiazene hydroxide are formed, with the latter intermidiate becoming an alkylating species, possibly in part as a 2-chloroethyl carbonium ion.


Effect of antimyeloma cell antiserum on immunological enhancement. Growth of Sarvoma 180 in AKR mice was enhanced by immunization with frozen-thawed homogenates of this tumor. Treatment of host mice with rabbit antimyeloma cell antiserum, either furing immunization or shortly after tumor implantation, resulted in a decreased incidence of tumor rejection or an increased rate of tumor growth. A slow-growing subline from a spontaneous, DBA/2Ha-DD mammary tumor is rejected after initial growth in DBA/2J mice. The incidence of rejection was reduced by treatment with the antiserum studied.


Chromatographic analyses of 3-methylcholanthrene metabolism in adult and fetal mice and the occurrence of conjugating enzymes in the fetus. Adult pregnant mice were given i.v. injections of (3H)3-methylcholanthrene (20 muCi in 1.1 mug/mouse) or (14C)3-methylcholanthrene (1.0 muCi in 48 mug/mouse). Ethanol extracts of their tissues were chromatographed on Sephadex LH-20. Three groups of 3-methylcholanthrene metabolites were obtained: one group as yet unidentified, one containing the hydrocarbon and hydroxylated derivatives, and a third consisting of conjugated metabolites from the treated adult mice and their fetuses. The conjugated metabolites in tissue and in bile were separated into two fractions; one was acted on by beta-glucuronidase and to a lesser extent by arylsulfatase, and the other was resistant to these enzymes but completely susceptible to acid hydrolysis. The hydrolysis resulted in altered chromatographic behavior characteristic of the hydroxy compounds, which also appear in tissue. The enzyme-resistant conjugates were predominant in brain, muscle, and lung, and the enzyme-labile conjugates were predominant in the kidney, liver, and bile of adult mice. These conjugated metabolites were also demonstrated in fetal mice; some appeared in the fetus as early as the thirteenth day of gestation, the most immature fetus so far examined. The resistant group was predominant in the early developmental stages of the fetus and the susceptible group was increased in the excretory organs such as the kidney, liver, and contents of the intestinal tract as the fetuses approached term. transplacental transfer of conjugated metabolites from the mother to the fetus did not take place, although the parent 3-methylcholanthrene and its nonconjugated metabolites were transferred. We therefore assume that drug-metabolizing enzymes, including hydroxylases and conjugases, are active in the fetal mouse tissues as well as in the adult.


Effect of core lipopolysaccharides from Salmonella minnesota R mutants on the survival times of mice bearing Ehrlich tumor. Injection of a homologous series of bacterial core lipopolysaccharides obtained from Salmonella minnesota R mutants to Ehrlich solid tumor-beating mice results in an increase of survival times of treated animals. Lower chain length favors greater antitumor activity. Oligosaccharides and polysaccharides derived from lipopolysaccharides were found to be ineffective in increasing survial time of tumore-bearing mice. Smaller-sugar-chain-length core lipopolysaccharides were found to be better adjuvant than were those with longer sugar chains. Implication of adjuvant action of lipopolysaccharides in the elicitation of antitumor activity is suggested.


Investigation of the possible involvement of ketonic derivatives of polycyclic hydrocarbons in hydrocarbon carcinogenesis. 5,6-Dihydro-5-oxo-7,12-dimethylbenz(a)anthracene and 5,6-dihydro-6-oxo-7,12-dimethylbenz(a)anthracene are described. These compounds, which are isomeric with 7,12-dimethylbenz(a)anthracene-5,6-oxide (the K-region epoxide), were inactive in tests for tumor-initiating activity in mouse skin and tumor production in the s.c. tissue of the mouse.


Carbodiimide enhancement of complement-dependent antibody-mediated tumor cell lysis in vitro and antitumor activity in vivo. The water-soluble carbodiimide salt 1-ethyl-3-(3'-dimethylaminopropyl)carbodiimide. HCl (EDCI-HCl) has been shown to increase the complement-dependent lysis of cultured mouse neuroblastoma C1300 cells by two types of antibody: (1) natural antibodies in the sera of normal (nonimmunized) rabbits, and (b) serum antibodies from snygeneic tumor-bearing A/HeJ mice. In the latter case, both the level of serum antibodies and the extent of carbodiimide enhancement of immune lysis were demonstrated in vitro to be substantially greater with sera from mice bearing 21-day-old tumors relative to 4-day-old tumors. The carbodiimide EDCI-HCl has also been found to increase the complement-dependent lysis of cultured TA3 carcinoma cells by serum antibodies from isogeneic LAF1/J mice bearing ascites tumors in advanced stages of growth. Finally, it has been shown that EDCI-HCl exerts an antitumor activity in vivo that is significantly greater against 21-day-old than against 4-day-old neuroblastoma c1300 tumors. The increase in EDCI-HCl activity with tumor age is contrary to the response that would be expected if this drug were serving as an antimetabolite. This is evidenced by data showing that the antimetabolite 6-thioguanine is most effective against young rapidly growing neuroblastoma C1300 tumors. The correlation between carbodiimide antitumor activity and host production of cytotoxic antibodies suggests that EDCI-HCl may operate in vivo by an immunological mechanism comparalbe to that demonstrated in vitro.


Isolation and characterization of nuclear RNA polymerase II from chicken myeloblastosis cells. Nuclear RNA polymerases of chicken myeloblastosis cells were solubilized and fractionated by diethylaminoethyl-Sephades A25 column chromatography. Both alpha-amanitin-insenstitive (polymerase I) and- sensitive (polymerase II) species were isolated. Polymerase activity, contained two peaks of enzyme (IIa and IIb), which were further purified by glycerol gradient centrifugation. The partially purified enzymes were characterized by their requirement of four nucleoside triphosphates and metal ions and by their sensitivity to several inhibitors. The enzymes were compared with RNA polymearases derived from normal chickent bone marrow cells,and the total extractable myeloblastosis than in bone marrow cells. Polymearse II from both cell types was shown to be sensitive to cytosine arabinoside triphosphate inhibiton.


Effects of double and multiple doses of hydroxyurea on mouse duodenum and mammary tumors. Previous data on the effects of a single dose of hydroxyurea on C3H mouse duodenum and mammary tumors from a fast growing line (S102F) were used to predict times that may be optimal (i.e., minimize killing of the duodenal S-phase cells while enhancing the killing of tumor S-phase cells) for the administration of subsequent doses of hydroxyurea. These predicted protocols were tested by giving tumor-bearing mice injections of 2 doses at 24 hr intervals. A preliminary in vivo tumor treatment experiment was also done wherein multiple doses (up to 10) were given either at 12, 20, or 24 hr intervals with the mouse survival, body weights, and tumor volumes being recorded daily. The data show that partial cell synchronization was achieved in both tissues and the initial knetics of the surviving cells was essentially the same after a single dose, 2 doses, or 4 doses of hydoxyurea. Also, the different intervals between the 2 doses did not affect the timing of the initital peaks of DNA synthesis in partially synchronized cells; however, the height of the peaks was affected The results demonstrate that kinetic data can be useful for predicting optimal intervals for 2-dose regimes and probably multiple-dose regimes involving a single cell-cycle phase-speeific drug when applied to a mouse tumor model. However, the recovery phenomena in the respective tissues are extremely complicated and more animal tumor data need to be collected before  one can make adquate use of cell-synchronizing agents and perturbed cellular kinetic data for routine clinical chemotherapy or combined modality therapy.


In vitro combination chemotherapy demonstrating potentiation of vincristine cytotoxicity by prednisolone. Human lymphoid cells grown in long-term tissue culture have been used to study the cytotoxic effects of the combination of vincristine and prednisolone. The potentiating effects of this drug combination in vitro cannot be consistently shown if the drugs are addeded without attention to the growth rate of the cultured population. If the cultured cells have achieved maximum density and have remained at this density for more than 24hr, they are readily killed by vincristine alone and no further kill is achieved by adding prednisolone. Rapidly growing cell cultures, however, are relatively ristant to vincristine. The addition of prednisolone to such cultures restores their sensitivity to vincriestine, but the combination is no more effective than is vincristine alone in stable cell populations. These findings indicate that the effects of vincristine on the mitotic spindle, which produce metaphase arrest, do not account entirely for its ability to destroy cells. A second mechanism of action of vincristine at low concentrations is proposed.


An experimental model for evaluation of factors in tumor escape from immunological attack. A model system is presented for studying the factors involved in tumor immunity. The initial observations with this system concern the importance of dose and route of administration of tumor cells on tumor growth. The data show that myeloma tumor cells, when inoculated i.v.in relatively large numbers, are eradicated by the immune response of an allogeneic host; tumor cells administered i.v. in smaller number escape from immune attack even though the host has the potential to mount an immune response. BALB/c mouse myeloma cells (MOPC-21) were transplanted s.c., i.p., or i.v. into H-2-compatible allogeneic DBA/ 2 mice. There was a marked difference in the response of the host to tumor given s.c. or i.p. as compared  to tumor given i.v. Thus s.c. or i.p. inoculation resulted in lethal tumor growth when 5 x 10-3 or more tumor cells were given. In contrast, the outcome of i.v. inoculation depended on tumor cell dose. Although small cell doses ( 5x 10-4 down to 10-2) resulted in lethal tumor gosulted in lethal tumor growth with only 10% survival, large cell doses (10-5 to 5 x 10-7) resulted in tumor rejection and 70% survival. DBA/2 mice possess the immunological ability to react agaist the tumor when large doses of tumor cells (10-7) are given i.v. or i.p., since spleen cells obtained from such mice were found to be able to suppress the growth of MOPC-21 when a mixture of spleen cells and tumor cells was inoculated. On the basis of these initial observations, our model appears to relate especially to the idea that, in autochithonous tumor development or in metastasis of tumor, a small number of antigenic tumor cells, perhaps even a single cell, usually grows into a frank tumor in spite of the immunological competence od the host to respond to the tumor cells.


Glucose analogs as potential diagnostic tracers for brain tumors. No tumor-specific tracer has yet been found for the detection of brain tumors by external scintiscanning. Glucose is a substrate in high demand by almost all tumors, and therefore an investigation was undertaken into the potential value of glucose and its analogs as tracers for brain tumors. The compounds studied were D-)1-3H)glucose, D-(1-14C)glucose, (3H)3-O-methyl-D-glucose and L-(1-14C)glucose. The tracers were injected i.v. into C57BL/6J mice carrying a transplantable s.c. ependymoblastoma. At specific time intervlas after injection, mice were sacrificed and the radioactivity of 6 tissues including tumor and brain were assayed by means of an automated combustion technique and liquid scintillation counting. Tumor uptake, expressed as percentage mean body concentration, was 60% for 2 of the tracers, and 92 and 143%, respectively, for 2 others. Brain uptake was over 130% mean body concentration, with 3 of the 4 tracers studies. With L-glucose, brain uptake was only 15.4% mean body concentration, and a maximum tumor-to-brain ratio of 9.5 was achieved. The very high tumor uptake achieved with two of these carbohydrated demonstrates that a carbohydrate analog  may be found that shows high tumor specificity and uptake, and that may be useful for external scintiscanning.


Survival and differentiation of canine mammary tissues in the hamster cheek pouch. The survival and differentiation of canine normal, atypical nodular (preneoplastic, and neoplastic mammary tissues were examined after transplantation to the cheek pouches of immunosuppressed female golden hamsters. Without immunosuppression, none of the mammary transplants, survived 40 days of transplantation. With either prednisolone or antilymphocyte serum immunosuppression, 78% of the mammary transplants that were recovered from the cheek pouch survived at least 35 to 45 days.


Interaction of chemotherapeutic agents with methotrexate and 5-fluorouracil and its effect on de novo DNA synthesis. The transport of methotrexate is known to be affected by corticosteroids and vincristine in L1210 leukemia cells. The deoxyuridine suppression test was used to measure the metabolic consequences of using these drugs with the antimetabolites, methotrexate and 5-fluorouracil, in both L1210 leukemia cells and normal human marrow cells. The deoxyuridine suppression test can be utilized as a sensitive measure of methotrexate and 5-fluorouracil biological activity in producing defective de novo DNA synthesis. The deoxyuridine suppression test was found to detect changes in biological activity equal to 20 ng (0.044 nmole) of methotrexate and 200 ng (1.94 nmoles) of 5-fluorouracil. Hydrocortisone and prednisone, but not dexamethasone or prednisolone, decreased the methotrexate effect to one-half in both L1210 and human cells as measured by the deoxyuridine suppression test. 5-Fluorouracil biological activity was not affected by any steroid studied. Vincristine produced variable results, but on the average it decreased the methotrexate effect in human marrow. Vincristine consistently decreased the methotrexate effect in L1210 systems. Cephalosporin, 75 mug/ml (0.214 mumole), had no effect. In parallel studies, hydrocortisone decreased the uptake of methotrexate, but no folic acid, in human and L1210 cells. The deoxyuridine suppression test warants further investigation as a method of screening drugs for interaction with antagonists of de novo DNA synthesis. This tudy extends earlier evidence of drug interaction with methotrexate in a murine system to human cells and demonstrates that there is a metabolic consequence, reduced potency of methotrexate, as a result of reduced transport produced by certain corticosteroids.


Tumor induction in rats by feeding heptamethyleneimine and nitrite in water. Groups of 15 males and 15 females Sprague-Dawley rats were given 20 ml of drinking water solution containing either 0.2% heptamethyleneimine hydrochloride or this salt together with 0.2% sodium nitrite, 5 days a week for 28 weeks. Another group of 17 male and 30 female rats was given 0.2% sodium nitrite solution for 104 weeks. Most of the animals given heptamethyleneimine hydrochloride or sodium nitrite alone survived 2 years or more after the beginning  of the treatment, and no tumors attributable to the treatment were seen at death; tumors appearing were those of endocrine origin found commonly in untreated controls. In the group receiving the combined treatment, most females were dead at 50 weeks and most males were dead at 80 weeks, 27 of 30 having tumors not seen in either control group. A total of 16 had squamous carcinomas in the lung; 25 had tumors of the oropharynx, tongue, esophagus, and forestomach; and there were a few animals with tumors in the nasal cavity and trachea. The expericment showed that squamous tumors of the lung could be induced by ingestion an amine and sodium nitrite.


The isolation and characterization of polycyclic hydrocarbon-binding proteins from mouse liver and skin cytosols. The major protein to which metabolites of methylcholanthrene are covalently bound has been purified from C3H mouse liver cytosol. Its properties are identical to the mouse skin h-protein, which may be primary arget of carcinogenic hydrocarbon metabolites during transformation to caner. It has a molecular wight of 44,000, consists of 2 subunits o- M.W. 20,000, has an isoelectric point (pI) of 8.05 to 8.6, and a sedimentation coefficient of 3.6 S. These physical properties are rather similar to those of ligandin, a hepatic protein that binds carcinogen metabolites, steroid anionic metabolites, bilirubin, and exogenous organic anions, but not to those of the rat liver azo dye carcinogen binding 'slow h-2-5S' protein. The h-protein and ligandin consistently give different pl values. Two minor basic proteins (molecular weights around 44,000 each), to whcih methylcholanthrene metabolites are convalently bound, have been separated from the h-protein by carboxymethyl-cellulose chromatography. Prelininary results indicate that these 2 minor proteins are related to ligandin. A protein to which methylcholanthrene is noncovalently bound was also identified in the acidic fraction of the mouse liver and skin sytosols and has been partially purified and characterized. It has a molecular weight of 60,000, a pl of 5.0, and a sedimentation coefficient of 4.5S.


N-substitution of carbon 8 in guanosine and deoxyguanosine by the carcinogen N-benzoyloxy-N-methyl-4-aminoazobenzene in vitro. The major reaction products of the carcinogenic electrophile N-benzoyloxy-N-methyl-4-aminoazobenzene with guanosine or deoxyguanosine were characterized as N-(guanosin-8-yl)- and N-(DEOXYGUANOSIN-8-YL)-N-methyl-4-aminoazobenzene from the following chemical, radiochemical, and spectroscopic studies: (a) the presence of equimolar amounts of both N-methyl-4-aminoazobenzene (MAB) and guanosine or deoxyguanosine residues was shown by the 3H:14C ratios of the products from the reaction of (prime ring-3H)-N-benzoyloxy-N-methyl-4-aminoazobenzene with (8-14C)guanosine or (8-14C)deoxyguanosine and by the molecular weights of the trimethylsilyl derivatives of both products: (b) substitution of the dye residue on its amino nitrogen was indicated by the retention in the products of the 3H:14C ration of (CH2-3H + 14CH3)-N-BENZOYLOXY-N-methyl-4-aminoazobenzene and by the release of MAB on treatment of the nucleoside-dye derivatives with strong alkali in air; (c) substitution of the guanine residues in positon 7 or 8 was demonstrated by loss of 3H from (8-3H)guanosine or (8-3H)deoxyguanosine in the formation of the nucleoside-dye derivatives; (d) the stability of the products to mild alkali (as contrasted to the lability of 7-alkylguanosines) provided strong evidence that the substitution was in position 8 of the guanine residue; (e) direct evidence of 8-substitution came from the acid hydrolysis of guanosinyl- and deoxyguanosinyl-N-methyl-4-aminoazo benzene to N-(guan-8-yl)-N-methyl-4-aminoazobenze in up to 50% yield; (f) comparisons of the proton or 13C nuclear magnetic resonance spectra or both of N-(guan-8-yl)- N-methyl-4-aminoazobenzene, MAB, N-(guanosin-8-yl)-2-acetylaminofluorene, 2-acetylaminofluorene, guanosine, and 7 -methylguanosine with the spectra of the guanosine-MAB product further confirmed that substitution had occurred at position 8 of the guanosine residue. The new compound N-(guan-8-yl)-N-methyl-4-aminoazobenzene was synthesized. Attempts to devise an unambiguous synthesis of N-(GUANOSIN-8-YL)-N-methyl-4-aminoazobenzene were not successful.


Structures of hepatic nucleic acid-bound dyes in rats given the carcinogen N-methyl-4-aminoazobenzene. Rats were given the hepatocarcinogenic dye N-methyl-4-aminoazobenzene labeled in the prime ring with tritium. The hepatic ribosomal RNA and DNA from these rats were hydrolyzed to uoe;d imce;psode-dye products. The major nucleoside-dye derivatives cochromatographed with synthetic N-(guanosin-8-yl)-and N-(deoxyguanosin-8-yl)-N-(methyl-4-aminoazobenzene, respectively, on cellulose and silica gel thin layers in several solvent systems. Additional evidence for the characterization of the hepatic RNA-nucleoside derivative was obtained through its degradation by alkali in air to N-methyl-4-aminoazobenzene and -4-aminoazobenzene, as previously described for N-(guanosin-8-yl)-N-methyl-4-aminoazobenzene. The same nucleoside derivatives were also derived from ribosomal RNA and DNA reacted in vitro with the carcinogenic electrophilic derivative N-benzoyloxy-N-methyl-4-aminoazobenzene, esterification of the N-hydroxy derivative, and reaction of the resultant electrophilic esters with DNA and RNA.


Enhancing effect of hydrocortisone on hematogenous metastasis of Ehrlich ascites tumor in mice. The effect of hydrocortisone on blood-borne tumor metastasis was studied in an i.v. inoculation experiment with Ehrlich hypotetraploid clone 1, Ehrlich hypotetraploid stock, and Ehrlich hyperdiploid stock tumors. The administration of hydrocortisone before tumor inoculation resulted in increased tumor take, reduced mean survival time of mice, and concentration of tumor metastasis in a specific organ (i.e., lung metastasis for Ehrlich hypotetraploid clone 1 tumor, and liver metastasis for Ehrlich hypotetraploid stock and Ehrlich hyperdiploid stock tumors). Enhancement of tumor metastasis, as induced by hydrocortisone pretreatment, was not reproduced by the administration of 6-mercaptopurine, testosterone, or estradiol. The progress of tumor death in hydrocortisone-conditioned mice was not affected by either heparin or dextran sulfate. This indicated that the effect of hydrocortisone on tumor metastasis was independent of the effect of these agents on immune reaction or blood coagulation. In the tracer experiment with 125-I-labeled tumor cells, hydrocortisone pretreatment significantly increased over the control the intrapulmonary retention of Ehrlich hypotetraploid clone 1 tumor cells from 1 through 72 hr after tumor inoculation, the time lag required for the establishment of metastatic foci in the lung. The arrest of Ehrlich hypotetraploid stock and Ehrlich hyperdiploid stock tumors in the liver was also temporarily increased by hydrocortisone pretreatment. No correlation was found between tumor cell size and differential distribution of metastatic tumors with 3 Ehrlich tumors. An attempt was made to use this blood-borne metastasis system for chemotherapeutic study. Administration of cyclophosphamide gave rise to a significant prolongation of survival time and often to complete prevention of tumor metastasis in hydrocortisone-conditioned mice.


Aminopeptidases and arylamidases in normal and cancer tissues in humans. Multiforms of aminopeptidases and arylamidases in normal human liver, stomach, lung, ileum, colon, rectum, and kidney, and cancer tissue from human liver, stomach, and lung were separated by triethylaminoethyl cellulose column chromatography. The aminopeptidases and arylamidases were solubilized from human tissues by treatment with bromelain, and their column chromatograms on triethylaminoethyl-cellulose gave different patterns of multiforms of enzymes in these tissues. The fractions of enzymes separated specificities toward L-leucyl-beta-naphthylamide, L-leucinamide, L-methioninamide, L-phenylalaninamide, and L-alaninamide. The activity of aminopeptidase toward L-leucinamide and of arylamidase toward L-leucyl-beta-naphthylamide was higher in human stomach cancer tissue and lower in hepatic cancer tissue than in normal stomach and liver, respectively. In lung cancer tissue, the activity of aminopeptidase toward L-leucinamide was abnormally low, while the activity of arylamidase toward L-leucyl-beta-napthylamide was similar to that in normal lung. The substrate specificities or patterns of the multiforms of these enzymes in cancer tissue from human liver, stomach, and lung were shown to differ from those of normal liver, stomach, and lung, respectively, by triethylaminoethyl cellulose column chromatography.


Neoplastic transformation of guinea pig fetal cells in culture induced by chemical carcinogens. Twenty-four cell strains derived from freshly isolated diploid strain 2 guinea pig fetal cells exposed in utero or directly in culture to a carcinogenic or noncarcinogenic chemical were evaluated during 4 to 24 months of continuous cultivation. Morphological alterations in carcinogen, i.e., benzo(a)pyrene-, 7,12-dimethylbenz(a)anthracene-, 3-methylcholanthrene-, N-methyl-N'-nitro-N-nitrosoguanidine-, diethylnitrosamine-, aflatoxin B1-, or N-acetoxyacetylaminofluorene-treated cultures were seen shortly after treatment; transformation, a loss of cell orientation, did not occur for 4 or more months, was not seen in controls and did not uniformly appear simultaneously with the capacity of transformed cells to grow as tumors in irradiated syngeneic newborn guinea pigs. Changes in plating efficiency, saturation density, doubling time, and chromosome alterations of transformed cells also did not correlate with progressive tumor growth. Formation of colonies in 0.35% agar appeared subsequent to or concomitantly with morphological transformation, and in every case they developed concurrently with the potential for neoplastic growth. Controls originally treated with a noncarcinogen (acetone, polycyclic hydocarbon, or aromatic amide) did not form agar colonies or tumors after inoculation of 108 cells. Studies with guinea pig cells indicate that persistent alterations in some growth parameters occur prior to expression of tumor growth. The ability of cells to form colonies in agar and to produce tumors may require 4 to 18 months of culturing. Of the cell properties examined, colony formation in agar was the best indication of the neoplastic state.


Colony inhibition mediated by nonimmune leukocytes in vitro and skin reactivity in vivo as indices of tumorigenicity of guinea pig cultures transformed by chemical carcinogens. Two short-term quantitative assays that correlate with the tumorigenicity of strain 2 guinea pig fetal cells transformed in culture by chemical carcinogens are described; one measures inhibition of colony growth mediated by nonimmune leukocytes and the other measures skin reactivity in unimmunized syngeneic guinea pigs. Mineral oil-induced peritoneal exudate (PE) cells were obtained from healthy unimmunized syngeneic guinea pigs. The PE cells were cultured for 24 hr and the nonadherent cells with culture medium were incubated with tumorigenic or nontumorigenic target cells in ratios of 1000/1 to 10/1. After 7 to 9 days of incubation in the presence of peritoneal exudate cell culture (PEC), fewer target cell colonies were observed in cultures with tumorigenic than with nontumorigenic cells. The inhibition of tumorigenic cells was dependent on PEC concentration; at 1000/1 PEC/traget cell ratio, a reduction of as much as 80% in the number of colonies and a 100% decrease in colony size relative to controls were noted. Inhibitory activity was present primarily in the supernatant fluid of the culture medium of the PE cells. Phytohemagglutinin stimulation of the PE cells increased PEC and peritoneal exudate cell culture medium supernatant (PES) colony-inhibitory activity as much as twofold. The differential colony inhibition activity of PES with or without phytohemagglutinin was stable during storage of PES for 5 months at -35 degrees. In the 2nd assay, 2 or 5 X 106 tumorigenic or nontumorigenic cells were inoculated intradermally into 12- to 16-seek-old male unimmunized syngeneic guinea pigs; skin reactivity, in terms of the degree and persistence of induration during the next 4 days, was greater to tumorigenic than to nontumorigenic cells. In both assays, tumor-producing cells, morphologically transformed in culture, or tumor-derived cells, were affected more than early-passage-untreated fetal cells, morphologically nontransformed long-term-cultured cells previously exposed to noncarcinogenic chemicals, or chemical carcinogen-transformed but non-tumor-producing cells. The two assays, particularly the nonimmune leukocyte-mediated colony inhibition with its greater degree of discrimination, provide rapid estimations of the tumorigenic potential of cells transformed in in vitro model systems of chemical carcinogenesis.


Hexamethylmelamine-induced regression of human lung tumors growing in immune deprived mice. Hexamethylmelamine is known to be effective in humans in the treatment of certain malignant tumors, especially bronchial carcinoma. It is, however, quite inactive against a number of animal tumors, making difficult a study of its mechanism of action in experimental systems. In a reexamination of the effects of hexamethylmelamine, two tumors were found to be very sensitive, namely, a mouse plasma cell tumor (PC6) and a human bronchial carcinoma (P246) growing in immune deprived mice. Both tumors undergo a significant and almost complete regression, even when well established, and hence may serve as model systems for the study of the mechanism of action of hexamethylmelamine.


An improved Mantel-Bryan procedure for "safety" testing of carcinogens. A published method by Mantel and Bryan for calculating "safe" doses of carcinogens is updated by incorporating several improvements. These improvements include more effective procedures for taking into account any spontaneous tumor rate and for combining data at several dose levels. An added feature is that it permits the combining of data from several experiments by postulating that it is only the spontaneous rate that differs between experiments. The improved method is illustrated with data from five hypothetical experiments, using a risk level of 10-8, a conservative slope of one probit or normal deviate per tenfold dose increase, and a nominal assurance level of 99%. The hypothetical experiments were geared to bring out particular pointsas, for example, the applicability of the model in the absence of control data. A large variety of issues involved in the determination of "safe" doses are discussed, including questions of experiment design and extrapolitan between species. A statistical appendix is provided, laying the framework for the calculating procedure and detailing complications therein. The "safe" dose approach helps resolve certain dilemmas in questions relating to food additives. A "no-detectable-level" prescription for chemical residues may be dangerous to the public where detection techniques are insufficiently sensitive, but it can become far too restrictive as exquisitely sensitive detection techniquesare developed. Only levels in excess of the "safe" dose would require detection. Calculated values for the "safe" dose could be updated and increased as more clear evidence of safety becomes available.


The carcinogenic and electrophilic activities of N-benzoyloxy derivatives of N-methyl-4-aminoazobenzene and related dyes. The N-benzoyloxy derivatives of N-methyl-4-aminoazobenzene, its 4'-methyl and 4'-ethyl derivatives, and N-ethyl-4-aminoazobenzene were synthesized for comparison of their carcinogenic activities and their reactivities with nucleophilic reagents. Each of the 4 esters had a similar degree of nonenzymatic reactivity with methionine and guanosine at neutral pH. Each of the dyes induced sarcomas at the site of s.c. injection in rats, but N-benzoyloxy-N-methyl-4-aminoazobenzene was considerably more carcinogenic than were any of the other dyes. N-benzoyloxy-N-methyl-4-aminoazobenzene was also more stable in neutral lipid, and this stability may have contributed to its greater carcinogenicactivity. Neither the electrophilic reactivities nor the s.c. carcinogenicities of these dyes paralleled the hepatocarinogenic activities of the parent dyes.


Detection of nerve growth factor binding sites on neuroblastoma cells by rosette formation. A cloned suspension culture of mouse C1300 neuroblastoma cells bound, at 2degrees, sheep erythrocytes passively coated with nerve growth factor, with the formation of rosettes. When grown in tissue culture dishes to which they could attach, neuroblastoma cells rapidly transformed, within 48 hr emitting cytoplasmic processes some of which were several mm long. Most of the attached neuroblastoma cells formed rosettes. In contrast, normal mouse kidney cells or various murine tumor cell lines used as cell controls exhibited a poor capacity for binding nerve growth factor. Rosette formation was a specific reaction that could be prevented by pretreating cells with proteolytic enzymes, free nerve growth factor, or specific antibodies against neuroblastoma cell extracts.


Phagocytosis of nerve growth factor-coated erythrocytes in neuroblastoma rosette-forming cells. At 2 degrees, murine C1300 neuroblastoma cells bound NGF-coated sheep erythrocytes and formed rosettes. When the temperature was raised to 37 degrees, the neuroblastoma cells underwent a rapid transformation characterized by microtubule formation, which occurred under the membrane surface close to the points of contact with the attached red cells. Cytoplasmic processes filled wit- microtubules were then emitted by the cell body and surrounded the red cells. Within 20 to 30 min, the attached erythrocytes were phagocytized. Interiorization of membrane-bound erythrocytes-antibody-complement complexes by neuroblasto-a cells could be similarly induced at 37 degrees. In both cases, the extent of phagocytosis was decreased when microtubule formation was blocked with colchicine or vinblastine. Complete inhibition was obtained only by pretreatment of cells with cytochalasin B, a strong inhibitor of microfilament contraction. The role played by the microtubules and the microfilaments in promoting the phagocytosis of the attached erythrocytes is discussed.


The activity of regional nodes in the evolution of immune responses to allogeneic and isogeneic tumors. The evolution of immune responses of C57BL/6 mice to allogeneic tumor Sarcoma 180 and of A/J mice to isogeneic tumor Sarcoma 1 was investigated by colony and cell inhibition (CI) assays. The immune response of lymphocytes from regional popliteal nodes, distant nodes, and spleens was examined at varying times after s.c. implantation of known numbers of in vitro-grown tumor cells in the hind feet. In the allogeneic system, only regional node lymphocytes produced CI activity, amximum response appearing at Day 14 and gradually diminishing therafter with tumor regression. Serum-blocking activity was not observed until Day 21 and increased to significant levels by Day 39 when no lymphocyte CI activity was detectable. In the isogeneic system, CI activity was tumor-dose dependent. Responses to low-dose inocula were confined to regional nodes, whereas with high-dose inocula, initial responses were provided by regional nodes, but by Day 21 the spleen had become the primary source of CI activity. Examination of blocking activity in this system was not possible due to nonspecific serum cytotoxicity. Lymph nodes other than regional showed no CI response at any time in either tumor system. These studies demonstrate the importance of regional nodes in the development of immune responses to both allogeneic and osogeneic tumors.


Prolongation of chemotherapeutically induced remission of a syneneic murine leukemia by L-2,3,5,6-tetrahydro-6-phenylimidazo[2.1-BETA]thiazole hydrochloride. L-2,3,5,6-Tetrahydro-6-phenylimidazo[2,1-beta]thiazole hydrochloride (LMS), when used in concert with 1,3-bis(2-chloroethyl)-1-nitrosourea, resulted in a significantly higher percentage of long-term leukemic-free survivors. The additive effect provided by LMS treatment was evident during the immunosuppressed period induced by 1,3-bis(2-chloroethyl)-1-nitrosourea treatment and when tumor load was minimal. Treatment with LMS alone did not appear to possess any significant antitumor effect. The beneficial effect of LMS treatment may be attributable to the immunostimulatory activity reported for this drug. LMS appears to possess characteristics that make it an excellent candidate for use as an immunostimulant in cancer combined modality treatment.


Antitumor activity of macromomycin B (NSC 170105) against murine leukemias, melanoma, and lung carcinoma. Mice bearing either of the two rapidly growing mouse leukemias, L1210 or P388, or the slow-growing B16 melanoma responded to i.p. injections of Macromomycin B (NSC 170105) with significant increases in life-span. The maximal increases in life-span obtained in these experiments were 37% for L1210, 68% for P388, and 120% for B16. In addition, there were 7 of 30 cures for varying doses of Macromomycin in the B16 melanoma. Activity of over 50% increase in life-span in B16 was obtained with a daily i.p. injection on Days 1 to 9 of 16 to 40 mg/kg. Animals that had received s.c. implanted Lewis lung tumors responded to either single or repeated injections (8 to 16 mg/kg) given at the site of tumor implant by a marked reduction in growth of the primary tumor, increased life-span, and some cures. The same doses were without effect when administered i.p. The reported activity of Macromomycin against L1210, P388 leukemias, B16 melanoma, and Lewis lung carcinoma make it a good candidate for development for clinical trial against human solid tumors. A new method of evaluating activity against solid tumors, "responder analysis," is also presented.


The effects of an antimetastatic agent, (plus or minus)-1,2-bis(3,5-dioxopiperazin-1-yl)propane (ICRF 159), on platelet behavior. Some agents that inhibit platelet aggregation, e.g., aspirin and dipyridamole, ahve been reported to prevent metastasis formation. To see whether inhibition of platelet aggregation could account for the antimetastatic action of (plus or minus)-1,2- bis(3,5-dioxopiperazin-1-yl)propane, this compound was investigated in vivo for any effects on platelet behavior and thrombogenesis. (plus or minus)-1,2-Bis(3,5-dioxopiperazin-1-yl)propane inhibited the formation of platelet thrombi in blood vessels on the surface of the rat brain and in the hamster cheek pouch. 1,2-Bis(dioxo-4-methylpiperazin-1-yl)ethane, a closely related analog of (plus or minus)-1,2-bis(3,5-dioxopiperazin-1-yl)propane, but without antimetastatic action, inhibited thrombus formation in vivo as effectively as (plus or minus)-1,2-bis(3,5-dioxopiperazin-1-yl)propane. It seems unlikely therefore that the antimetastatic action of (plus or minus)-1,2-bis(3,5-dioxopiperazine-l-yl)propane derives from its effects on thrombogenesis. Neither platelet numbers nor the ionized plasma calcium concentrations were changed after (plus or minus)-1,2-bis(3,5-dioxopiperazin-l-yl)propane administration.


Induction of urinary bladder tumors in rats by administration nitrosomethyldodecylamine. Nitrosomethyl-n-dodecylamine, a product of the reaction of dimethyl-n-dodecylamine with nitrous acid, was given to Sprague-Dawley rats by gavage in olive oil solution. Twice-weekly treatments with 12 mg of the nitrosamine for 50 weeks gave rise to 100% incidence of transitional cell carcinomas of the urinary bladder. In contrast with the tests of other N-nitroso compounds, this compound seemed to be more effective in males than in females; females died later with the tumor although the dose per unit body weight was higher in females than in males.


The pharmacokinetics of 4-acetyl tritium vinblastine in two patients. Vinblastine, labeled with tritium in the 4-acetyl group, was given to two patients with malignant disease, and the pharmacokinetic behavior of the drug was determined. Clearance of radioactivity from the blood was biphasic, with t1/2 values for a first rapid phase of 4.25 and 4.78 min, and for a slower phase of 185 and 195 min. The volume of the central compartment was calculated as 29.7 and 39.4 liters, while the total fictive volume of distribution was 86.4 and 111.4 liters. Binding to blood components occurred in the order: plasma greater than platelets greater than red blood cells greater than white blood cells. Excretion of radiolabel occurred via the stool and the urine so that, after 72 hr, 25 and 41% of the total dose had appeared in the former and 19 a nd 23% had appeared in the latter. Appreciable amounts of unchanged drug appeared in the urine, while very little appeared in the stool, suggesting hepatic metabolism, consistent with prior animal studies.


Some properties of a DNA-unwinding protein unique to lymphocytes from chronic lymphocytic leukemia. Lymphocytes from a common human leukemia, chronic lymphocytic leukemia, chronic lymphocytic leukemia, have a greatly enhanced capability of DNA repair and a concomitantly prolonged survival in vitro after damage to DNA. From these lymphocytes, we isolated and purified a DNA-binding protein with a molecular weight of 24,000. It binds tightly to both ultraviolet light (UV)-irradiated and single-stranded DNA. At 35 degrees it enhances the helix-coil transition of poly[d(A-T)] AND the UV-irradiated calf thymus DNA but is inefficient in ordinary native DNA. This protein also facilitates the rate of UV-endonuclease incision of UV DNA but does not induce any nicks by itself. This finding suggests that the protein may be involved in DNA repair by enhancing such activity, and also offers an explanation for our observation of increased DNA repair in chronic lymphocytic leukemia cells. When human metaphase chromosomes are exposed to the protein, it induces marked lengthening of chromatids suggesting that this protein may also act on complex chromosomes. By quantitative immunochemical determinations, such protein could not be found in lymphocyte extracts of three normal individuals.


Dehydroretronecine-induced rhabdomyosarcomas in rats. Two groups of rat were given s.c. injections of either monocrotaline or its major detectable metabolite, dehydroretronecine, biweekly for 1 year. Tissues obtained from partial hepatectomies performed at 4 months on a portion of these animals showed that both compounds caused a decided inhibition of mitotic division in regenerating liver. Rhabdomyosarcomas developed at the site of dehydroretronecine injection in 51.6% of the rats and in 3.3% of the monocrotaline-treated rats. Metastatic lesions were recorded in 8.3% of these animals. In addition to the above, 10% of the monocrotaline-treated rats developed other tumors that included myelogenous leukemias, hepatocellular carcinomas, and pulmonary adenomas. These data indicate that either monocrotaline or its metabolite dehydroretronecine are capable of causing neoplastic transformations in the tissues of experimental animals.


Photoamidation of unsaturated carbohydrates. Synthesis of 1-(3-C-carbamoyl-3-deoxy-beta-D-glucopyranosyl)-thymine and -cytosine. The acetone-initiated photochemical addition of formamide to 1,2,4,6-tetra-O-acetyl-3-deoxy-alpha-D-erythro-hex-2-enopyranose (1) afforded 1,2,4,6-tetra-O-acetyl-3-C-carbamoyl-3-deoxy-alpha-D-glucopyransoe (2), 1,2,4,6-tetra-O-acetyl-3-C-carbamoyl-3-deoxy-alpha-D-allopyranose (3), 1,2,4,6-tetra-O-acetyl-3-C-carbamoyl-3-deoxy-alpha-D-altropyranose (4), and 1,2,4,6-tetra-O-acetyl-3-deoxy-3-C-(1-hydroxy-1-methylethyl)-alpha-D-mannopyranose (5), in 46, 13, 1, and 7% yields, respectively. Condensation of 2,4,6-tri-O-acetyl-3-C-carbamoyl-3-deoxy-alpha-D-glucopyranosyl bromide (produced from 2 with hydrogen bromide) with 2,4-bis(trimethylsilyl)thymine afforded 1-(2,4,6-tri-O-acetyl-3-C-carbamoyl-3-deoxy-beta-D-glucopyranosyl)thymine (6) in 40% yield. Deacetylation of 6 with sodium methoxide in methanol yielded the pure unprotected nucleoside 7. Similar condensation of the bromide from 2 with 2,4-bis(trimethylsilyl)-N4-acetylcytosine afforded the protected cytosine nucleoside 8 in 55% yield, which was converted into 1-(3-C-carbamoyl-3-deoxy-beta-D-glucopyranosyl)cytosine (9).


DNA-dependent RNA polymerase levels during the response of human peripheral lymphocytes to phytohemagglutinin. The cellular levels of the various RNA polymerases have been monitored in resting human peripheral lymphocytes and in lymphocytes stimulated by phytohemagglutin. Activity was measured in the presence of exogenous templates following solubilization and chromatographic resolution of the different RNA polymerases. Resting lymphocytes contain Class I, II, and III RNA polymerases, although the respective levels of activity are very low compared to the levels in metabolically active cell types. During the PHA-induced transformation of resting lymphocytes, the Class I, II, and III enzyme levels rise dramatically. During four days exposure to PHA, the levels of RNA polymerases I and III (which synthesize, respectively, rRNA and the transfer and 5S RNAs) increase 17 fold, while the level of RNA polymerase II (which synthezies heterogeneous nuclear RNA) increase 8 fold. The possible relationship between enzyme levels and the regulation of gene expression is discussed.


Further evidence of transcriptional and translational control of histone messenger RNA during the HeLa S3 cycle. Using a translational assay and new analytical procedures we have found that: -Histone mRNA can be detected both associated with polyribosomes and in the postribosomal supernatant of S phase HeLa S3 cells.-Inhibition of DNA replication by cytosine arabinoside treatment causes histone mRNA to completely disappear from polyribosomes, and little histone mRNA can be detected in the postribosomal supernatant of inhibited cells. These data indicate that histone mRNA does not accumulate in the cytoplasm after the inhibition of DNA replication. -Histone mRNA species cannot be detected in the postribosomal supernatant of G1 cells synchronized by selective detachment. This observation, together with the previous finding that histone mRNA is not present on G1 polyribosomes, is consistent with the idea of a transcriptional block in histone mRNA production and transport to the cytoplasm during G1.


In vivo repair of rat liver DNA damaged by 3-hydroxyxanthine. Using alkaline and neutral sucrose gradients, it was observed that administration of the hepatocarcinogen, 3-hydroxyxanthine, to rats, resulted in the slower sedimentation of liver DNA in alkaline and neutral sucrose gradients. Slower sedimentation of liver DNA in alkaline sucrose gradients was apparent within 4 h following the administration of the carcinogen at a dose of 50 mug/g body weight. The fragmentation of liver DNA was progressive with an increase in the dose. Such fragmentation caused by a dose of 100 mug/g was largely repaired by 24 h following the administration of the carcinogen. Fragmentation and repair of liver DNA with a dose of 100 mug or more/g body weight was also seen using neutral sucrose gradients. In neutral sucrose gradients significant repair of the fragmented DNA was seen by 24 h after carcinogen. Liver DNA from rats given 3-hydroxyxanthine (100 mug/g for 4 h) or calf thymus DNA incubated in vitro with this carcinogen did not show any change in their melting profile.


Localization of heart vectors produced by epicardial burns and ectopic stimuli; validation of a dipole ranging method. Location of the equivalent cardiac dipole has been estimated but not fully verified in several laboratories. To test the accuracy of such a procedure, injury vectors were produced in 14 isolated, perfused rabbit hearts by epicardial searing. Strongly dipolar excitation fronts were produced in 6 additional hearts by left ventricular pacing. Twenty computer-processed signals, derived from surface electrodes on a spherical electrolyte-filled tank containing the test preparation, were optimally fitted with a locatable cardiac dipole that accounted for over 99% of the root-mean-square surface potential. For the 14 burns (mean radius 5.0 mm), the S-T injury dipole was located 3.4 plus or minus 0.7 (SD) mm from the burn center. For the 6 paced hearts, the dipole early in the ectopic beat was located 3.7 mm (range 2.6 to 4.6 mm) from the stimulating electrode. Phase inhomogeneities within the chamber appeared to have a small but predictable effect on dipole site determination. The study demonstrates that equivalent dipole location can be determined with acceptable accuracy from potential measurements of the external cardiac field.


Time course for reversal of electrophysiological and ultrastructural abnormalities in subendocardial Purkinje fibers surviving extensive myocardial infarction in dogs. The electrophysiological properties of subendocardial Purkinje fibers surviving in myocardial infarcts were studied with intracellular microelectrodes in isolated superfused preparations and correlated with subsequent light and electron microscopic studies. Transmembrane action potentials could always be recorded from one or two cell layers of subendocardial Purkinje fibers in infarcted regions 3 days to 7 weeks after coronary artery occlusion; ventricular muscle action potentials were rarely found. Microscopic studies also demonstrated several layers of intact subendocardial Purkinje fibers; the subjacent ventricular muscle cells were irreversibly injured and replaced by scar. At all time intervals, surviving Purkinje fibers had significantly reduced maximum diastolic potentials, action potential amplitudes, and depolarization velocities as well as prolonged action potential durations. These parameters normalized between 24 hours and 7 weeks after coronary artery occlusion. Surviving Purkinje fibers with electrophysiological abnormalities at 24 hours and 3 days contained vast lipid deposits. Lipid was less prevalent at 10 days when action potential characteristics had improved. By 7 weeks, action potentials were nearly normal and lipid was absent. Subendocardial Purkinje fibers surviving in infarcts are subject to conditions which cause electrophysiological and ultrastructural abnormalities. Persistent abnormalities in the electrophysiological properties of these surviving Purkinje fibers may cause persistent altered electrophysiological properties of the infarcted heart.


A model of psychosocial hypertension showing reversibility and progression of cardiovascular complications. The sequence of pathophysiological changes that can result from the stimulating effects of a sustained disturbance of the social environment was studied in ten colonies of socially deprived mice. Sixteen formerly isolated males were placed with 16 normal females in population cages consisting of  seven intercommunicating boxes. Six of these socially disturbed 32-member colonies were terminated after periods of interaction ranging from 2 days to 9 months. The remaining four were terminated a month or more after the males had been returned to individual isolation. Indirect blood pressure measurements, body and heart weights, and sections of hearts and aortas were studied in the males. Following the shorter exposures, blood pressure reverted to normal in a few days. Exposure of 6 months or more were associated with unchanged body weights and sustained increases in heart weight and blood pressure readings. In addition, there was a significant development of aortic arteriosclerosis and myocardial fibrosis. These changes persisted despite prolonged return to isolation.


Hemodynamic effects of acute experimental aortic coarctation in the dog. Pressure and flow pulses were recorded immediately proximal and distal to a temporary stenosis of the thoracic aorta of the dog. The severity of the stenosis was varied, and the magnitude of the pulse changes was correlated with the degree of stenosis expressed as the area ratio, As/All, where As is the cross-sectional area of the stenotic section and A-s that of the normal vessel. The harmonic amplitudes of the pressure and flow waves during the stenotic period were obtained by Fourier analysis and normalized to their values when the stenosis was released. It was found that as A-s/All yields 0 the proximal pressure amplitude increased and the distal pressure and the proximal and distal flow amplitudes decreased. These changes were such that the proximal fluid impedance calculated from the corresponding pressure and flow amplitude ratios increased as A-s/A-ll yields 0; the distal impedance remained relatively unaffected. These findings can be interpreted in terms of reflection of the pulse waves at the stenosis. The normalized proximal pressure amplitudes approximate to 1 + R and the distal pressure and both the proximal and the distal flow vary according to 1 minus R, where R is the reflection coefficient. The variation of R with A-s/A-ll can be determined from the proximal fluid impedance changes; we found that R increased relatively slowly for moderate stenoses, varying from 0 at A-s/A-ll 1.0 (no stenosis) to  similar to 0.2 at A-s/A-ll equals 0.2. At more severe stenoses, it increased more rapidly approaching 1.0 at A-s/A-ll equals 0. The generation of turbulence was most marked at A-s/A-ll yields 0.2.


Vascular endothelium-leukocyte interaction; sticking shear force in venules. To determine the shear force acting on a white blood cell sticking to the endothelium of a blood vessel, the flow field about a single white blood cell in a venule was determined by hign-speed motion picture photomicrography. The force acting on the white blood cell was then calculated according to the principles of fluid mechanics. In this paper, the calculation was made using an experimentally determined dimensionless shear force coefficient obtained from a kinematically and dynamically similar model. The large physical model of the hemodynamic system could be easily instrumented, and the shear force acting on the model cell and the flow field around it were measured. The data were then used to calculate a shear force coefficient. On the basis of dynamic similarity, this shear force coefficient was applied to the white blood cell in the venule. The shear force coefficient was strongly influenced by the hematocrit, so in vivo hematocrits were measured from electron micrographs. It was found that in the venules of the rabbit omentum a white blood cell sticking to the endothelial wall was subjected to a shear force in the range of 4 times 10--5 dynes to 234 times 10--5 dynes; the exact value depended on the size and motion of the white blood cell, the size of the blood vessel, the velocity of the blood flow, and the local hematocrit, which varied between 20% and 40% in venules of about 40 mum in diameter. The contact area between the white blood cell and the endothelial cell was estimated, and the shear stress was found to range between 50 dynes/cm-2 and 1060 dynes/cm-2. The normal stress of interaction between the white blood cell and the endothelium had a maximum value that was of the same order of magnitude as the shear stress. The accumulated relative error of the experimental procedure was about 49%. The instantaneous shear force was a random function of time because of random fluctuations of the hematocrit.


Comparison of 85krypton and 133xenon cerebral blood flow measurements before, during, and following focal, incomplete ischemia in the squirrel monkey. A comparison of regional cerebral blood flow measurements made with beta- and gamma-emitting isotopes revealed good correspondence in areas of normal perfusion and reactive hyperemia but poor correspondence in areas of focal ischemia. After middle cerebral artery occlusion at normocapnia, there was a 65% reduction in regional cerebral blood flow from 1.40 plus or minus 0.27 ml/g min--1 to 0.49 plus or minus 0.10 ml/g min--1 in monkeys studied with 85Kr but only a 27% reduction in regional cerebral blood flow from 0.84 plus or minus 0.09 ml/g min--1 to 0.61 plus or minus 0.08 ml/g min--1 in monkeys studied with 133Xe. The lack of correlation within areas of focal, incomplete ischemia was attributed to an impairment of isotope delivery to the area of ischemia coupled with the inherent lack of spatial resolution of determinations made with 133Xe. This finding may partly explain the numerous discrepancies in experimental and clinical studies of the effects of alterations in the arterial partial pressure of CO2 on regional cerebral blood flow in areas of ischemia; it may also explain the failure of such studies to reflect the true severity of focal ischemia.


Enhanced renal prostaglandin production in the dog. I. Effects on renal function. The changes in renal function produced by endogenous synthesis of prostaglandins by the kidney were evaluated by infusing sodium arachidonate, the prescursor of the prostaglandins, into one renal artery of the dog. These changes were compared with those produced by similar infusions on performed prostaglandin (PG) E2 and F2alpha.PGE2given at 0.01-0.3 mug/kg min--1 produced dose-related increases in urine flow, sodium and potassium excretion, free water clearance, and renal blood flow. The glomerular filtration rage increased only at the lowest dose and the calculated filtration fraction fell. Arachidonic acid at 1.0-30.0 mug/kg min--1 similarly produced dose-related increases in electrolyte excretion, but the increase in renal blood flow was much less than that produced by PGE2 and there were no changes in glomerular filtration rate, filtration fraction, or free water clearances. PGF2alpha had essentially no effects at infusion rates of 0.03-1.0 mug/kg min--1. All renal effects of arachidonic acid were inhibited by simultaneous infusions of an inhibitor of prostaglandin synthetase, 5, 8, 11,14-eicosatetraynoic acid (20:4). None of the effects produced by PGE2 were inhibited by 20:4. These results indicate that enhanced endogenous renal prostaglandin synthesis, which can be produced by arachidonate infusion, results in significant alterations of renal function. This finding strengthens the hypothesis that renal prostaglandins formed in vivo have physiological importance as regulators of renal function.


Enhanced renal prostaglandin production in the dog. II. Effects on intrarenal hemodynamics. The effects of enhanced endogenous production of prostaglandins by the kidney on the distribution of blood flow in the renal cortex were assessed by infusing sodium arachidonate, the precursor of the renal prostaglandins, into one renal artery of the dog. The changes produced with arachidonate (3 times 10--6 g/kg min-1 and 10--5 g/kg min--1) were compared with those produced by infusions of prostaglandin (PG) E2 (10--7 g/kg min--1) and PGF2alpha (3 times 10--7 g/kg min--1) into  one renal artery. Distribution of renal blood flow was measured by the radioactive microsphere technique. Sodium arachidonate caused an increase in blood flow to the inner cortical zones with no change in flow to the nephrons in the outermost quarter of the cortex. PGE2 increased flow to all cortical zones, and PGF2alpha produced no change in flow. Since arterial blood pressure did not change, changes in vascular resistance were reciprocal to changes in flow. Thus, vascular resistance fell in the inner cortical regions but not in the outer regions, with arachidonate infusions and in all regions of the cortex with PGE2 infusions; no changes were seen with PGF2alpha infusions. These data indicate that prostaglandin formed endogenously in the kidney affects the vascular resistance of only the inner cortical nephrons; the data thus support the hypothesis that renal prostaglandins are one mediator of renal autoregulation of blood flow.