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Stem Cells International
Volume 2017, Article ID 4758930, 12 pages
Human Suprapatellar Fat Pad-Derived Mesenchymal Stem Cells Induce Chondrogenesis and Cartilage Repair in a Model of Severe Osteoarthritis
1Unidad de Traumatología, Hospital Casa de la Salud, Valencia, Spain
2Neuronal and Tissue Regeneration Lab, Centro de Investigación Príncipe Felipe, Valencia, Spain
3Molecular Pathology and Translational Research in Oncology, Unidad Mixta Universidad Católica de Valencia y Centro de Investigación Príncipe Felipe, Valencia, Spain
4Unidad de Artroscopia y Unidad de Traumatología del Hospital Quiron, Barcelona, Spain
5Fundación García Cugat, Barcelona, Spain
6Universidad Católica de Valencia, Valencia, Spain
7FactorStem Ltd., Valencia, Spain
Correspondence should be addressed to Victoria Moreno-Manzano; se.fpic@moneromv
Received 5 December 2016; Accepted 23 April 2017; Published 9 July 2017
Academic Editor: Gerald A. Colvin
Copyright © 2017 Ignacio Muñoz-Criado et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Cartilage degeneration is associated with degenerative bone and joint processes in severe osteoarthritis (OA). Spontaneous cartilage regeneration is extremely limited. Often the treatment consists of a partial or complete joint implant. Adipose-derived stem cell (ASC) transplantation has been shown to restore degenerated cartilage; however, regenerative differences of ASC would depend on the source of adipose tissue. The infra- and suprapatellar fat pads surrounding the knee offer a potential autologous source of ASC for patients after complete joint substitution. When infrapatellar- and suprapatellar-derived stromal vascular fractions (SVF) were compared, a significantly higher CD105 (+) population was found in the suprapatellar fat. In addition, the suprapatellar SVF exhibited increased numbers of colony formation units and a higher population doubling in culture compared to the infrapatellar fraction. Both the suprapatellar- and infrapatellar-derived ASC were differentiated in vitro into mature adipocytes, osteocytes, and chondrocytes. However, the suprapatellar-derived ASC showed higher osteogenic and chondrogenic efficiency. Suprapatellar-derived ASC transplantation in a severe OA mouse model significantly diminished the OA-associated knee inflammation and cartilage degenerative grade, significantly increasing the production of glycosaminoglycan and inducing endogenous chondrogenesis in comparison with the control group. Overall, suprapatellar-derived ASC offer a potential autologous regenerative treatment for patients with multiple degenerative OA.
1. Introduction
Osteoarthritis (OA) is a degenerative joint disease with no efficient treatment due to limited endogenous regenerative capacity. It affects the knees of nearly a quarter of the population aged 60 and older [1, 2]. Currently, patients with severe OA are inexorably relegated to prosthetic joint substitution. OA is a disorder involving movable joints characterized by cell stress and extracellular matrix degradation initiated by micro- and macroinjury that activates maladaptive repair responses including proinflammatory pathways of innate immunity. The disease is first manifested as a molecular derangement (abnormal joint tissue metabolism) followed by anatomic and/or physiologic derangements (characterized by cartilage degradation, bone remodelling, osteophyte formation, joint inflammation, and loss of normal joint function) culminating in disease [3, 4].
Since the OA pathological processes are well described involving among others, the loss of functional chondrocytes, several cell-based therapeutic approaches have already been successfully developed including bone-marrow stimulation [5], implantation of osteochondral autograft [6] or allografts (ACI) [7], and transplantation of expanded autologous chondrocytes [8], or amplified mesenchymal stem cells (MSCs) [9], which help to restore articular cartilage. The intra-articular administration of MSC directly in the synovial fluid has been the predominant cell-based approach, with already demonstrated clinical effectiveness, with probed regenerative and immunosuppressant activities [9, 10]. To date, it continues to be an important avenue of research and clinical development due to its extraordinary therapeutic aptitude. However, the direct differentiation of multipotent MSC into cells of the chondrogenic lineage has led to a variety of experimental strategies to investigate whether tissue-specific MSCs are preferential for the regeneration and maintenance of articular cartilage [9]. Additional efforts on engineered cartilage implants are also done by using MSC synergistically activated with biomolecules to potentially improve chondral and osteochondral lesion repair, converting those in specialized trophic producers to initiate endogenous regenerative activities in the OA joint [11–13]. The long-term durability, increased tissue integration, and specific activity depend first on better transplantation survival rates and better adaptation to the hostile environment which still comprises a challenge. To date, there is not a precise description of which donor sample would be more efficient in the generation of MSC for the treatment and repair of joints in a degenerative process. Subcutaneous fat tissue is the most accessible source; however, after prosthetic implementation for joint substitution, the supra- and infrapatellar fat pads are commonly resected, constituting a suitable autologous adipose-derived MSC source (ASC). The infrapatellar or Hoffa's fat pad is an intracapsular but extrasynovial structure, situated in the knee under the patella, between the patellar tendon, femoral condyle, and tibia plateau [14]. The suprapatellar or quadricep fat pad is externally interposed between the joint capsule and the synovium, lined to the joint cavity showing a triangular shape and extended through the patellar base [15]. Previous studies have already described the potential regenerative capability of ASC derived from the infrapatellar pad in a model of OA [16, 17], showing in fact a higher percentage of immunophenotypical positive stromal cells in comparison with those obtained from subcutaneous fat [18]. However, no previous data has been reported regarding the suprapatellar tissue regenerative capacity. In addition, recent reports also focus on the role of the Hoffa's-derived cells, such as inflammatory cells releasing or inducing the release of inflammatory mediators such as IL-6, IL-8, TNFα, and PGE2 when derived from OA patients, suggesting that the infrapatellar fat pad is an active joint tissue in the initiation and progression of knee OA [19–22].
There are stem cell niches with no spontaneous capacity to mobilize to the injury to repair the severe OA defects. The amplification of these cells from these specialized sources could render an optimal source of autologous or even allogenic applications to promote cartilage regeneration. Indeed, the patellar fat pad-derived ASC would offer a potential autologous regenerative treatment for patients with multiple degenerative OA.
2. Material and Methods
2.1. Adipose Tissue Processing, SVF Isolation, and ASC Culture
Twenty-four patients between 50 and 80 years old indicated for complete joint substitution were included in the study. All patients were asked to sign an informed consent for the use of surplus fat tissue in the prosthesis surgery as well as a donation of peripheral blood (~20 ml) for autologous serum isolation. The samples, adipose tissue from supra- or infrapatellar areas, were anonymized and individually housed and collected inside the surgery room in sterile containers with sterile saline. This experimental procedure has been evaluated and accepted by the Regional Ethics Committee for Clinical Research with Medicines and Health Products following the Code of Practice 2014/01. As exclusion criteria, no samples were collected from patients with a history of cancer and infectious diseases active at the time of the surgery (viral or bacterial).
The adipose tissue, from supra- or infrapatellar areas, was transferred from the surgery room to the laboratory in a hermetic container at 4°C in sterile solution. The samples were washed multiple times in PBS plus antibiotics to clean the tissue and remove residual blood. The samples were distributed within 10 g of adipose tissue per 100 mm petri dish, with a solution containing PBS, 100 units/ml penicillin and 100 μg/ml streptomycin (Gibco 15,140), collagenase type IA (0.07%, Sigma C9891 CA, USA), and dispase I (0.2 mM Sigma). The tissue was cut into small pieces using sterile surgical scissors in a laminar flow hood and digested in a closed cell flask overnight in a shaker at 37°C, 20% O2, 5% CO2. The following day, the digested adipose tissue was collected and washed multiple times with PBS plus antibiotic by serial centrifugation. The cell pellet constitutes the stromal vascular fraction (SVF) and was suspended in 1 ml of PBS for cell counting via a Neubauer® chamber. For FACS analysis, 105 cells were utilized for each pair of antibodies. The remaining cells were employed for cell amplification and posterior analysis, distributed in two groups, growth in 10% human serum containing medium or 10% fetal bovine serum (in DMEM medium containing 2 mM L-glutamine, 30% L-glucose, 100 units/ml penicillin, and 100 μg/ml streptomycin), plated in petri dishes, and incubated overnight. The following day, the medium was removed and replaced with fresh medium and attached cells were allowed to grow until nearly confluent and then subjected to cell proliferative analysis, clonicity assay, morphological assessments, and FACS analysis and cell differentiation assays.
2.2. FACS Analysis
The SVF was assayed for cell surface protein expression by flow cytometry (FC500, Beckman Cultek, USA). 105 cells, diluted into 100 μl of PBS were incubated with 1 : 50 dilution for every pair of antibodies: CD90-PE and CD29-APC, CD44-PE-Cy, CD117-APC, CD105-PE, CD45-FITC, and CD34-PE (BD Pharmigen, USA). As a negative control, cell suspension without antibody was employed following the same procedure. Cells were incubated in the dark for 45 min at room temperature and then washed three times with PBS and suspended in 0.3 ml of cold PBS for FACS analysis. The mean ± SD of every identified population (in percentage) of all tested samples was determined.
2.3. Colony Formation Assay
The amplification and expansion of the ASC population from the SVF involves first the colony forming units after attachment onto a substrate and then a subsequent amplification in the presence of appropriate growth factors [23]. Fifty cells at passage three of every sample, supra- and infrapatellar, of four different patients were seeded in 6-well plates in the presence of 10% human serum or 10% fetal bovine serum containing medium for 10 days. The medium was replaced every third day. Subsequently, the cells were fixed with cold methanol for 5 minutes and washed with PBS before incubating in Giemsa solution (Sigma, USA) for 30 minutes. The excess stain was removed by subsequent washes with tap water. The cells were allowed to air dry and then visualized under the microscope. The mean ± SD of the total number of colonies at each condition of all tested samples was determined.
2.4. ASC Proliferative Analysis
At passage three of every sample, supra- and infrapatellar, 104 cells were seeded in 24-well plates in the presence of 10% human serum or 10% fetal bovine serum containing medium. Cells were seeded for up to ten days of analysis, by quantifying the number of cells in a Neubauer chamber every day in every well. The mean ± SD at every time point (every consecutive day) was represented.
2.5. Transmission Electron Microscopy
Cells were seeded at 2000 cells/cm2 in Lab-Tek chamber slides of 2 wells (Nalge Nunc International, Naperville, IL) and were fixed in 3% glutaraldehyde for 1 hour at 37°C. Cells were postfixed in 2% OsO4 for 1 hour at room temperature and stained in 1% uranyl acetate in the dark for 2 h at 4°C. Finally, cells were rinsed in distilled water, dehydrated in ethanol, and infiltrated overnight in Durcupan resin (Fluka, Sigma-Aldrich, St. Louis, USA). Following polymerization, embedded cultures were detached from the chamber slide and glued to araldite blocks. Serial semithin sections (1.5 μm) were cut with an Ultracut UC-6 (Leica, Heidelberg, Germany) and mounted onto slides and stained with 1% toluidine blue. Selected semithin sections were glued with Super Glue-3, Loctite (Henkel, Düsseldorf, Germany), to araldite blocks and detached from the glass slide by repeated freezing (in liquid nitrogen) and thawing. Ultrathin sections (0.06–0.08 μm) were prepared with the Ultracut and stained with lead citrate. Finally, photomicrographs were obtained under a transmission electron microscope FEI Tecnai G2 Spirit (FEI Europe, Eindhoven, Netherlands) using a digital camera Morada (Olympus Soft Image Solutions GmbH, Münster, Germany).
2.6. ASC Directed-Differentiation
ASC at or after passage 4 were subjected to directed differentiation [24], to induce adipogenesis, osteogenesis, and chondrogenesis, for each sample, supra- and infrapatellar from three different patients. All directed-differentiation media were obtained from Lonza (Lonza Co., Basel, Switzerland). Adipogenesis: ASC were seeded at a cell density of 10,000 cells/cm2 and when ASC were >90% confluent, the growth medium was substituted for differentiation medium containing insulin, dexamethasone, IBMX (3-isobutyl-methyl-xantine), and indomethacin (adipose-derived stem cell basal medium; Lonza Co.). The cells were then incubated in standard cell culture conditions for 12 days. The adipogenic differentiation was evaluated by Oil Red staining of the lipid vacuoles in formalin-fixed cultures; Oil Red O stock solution (5 g/l of isopropanol) was diluted 6/10 in water and incubated for 1 hour at RT. Serial washes with water to remove the excess staining were applied. After bright-field picture acquisition, total Oil Red O stain was extracted with 100% isopropanol for 5 min with gentle rocking and the absorbance at 492 nm was quantified. The mean ± SD of the absolute numbers at each condition of all tested samples was determined. Osteogenesis: ASC were seeded at a cell density of 10,000 cells/cm2 in collagen I (Sigma; 10 mM) coated plates in medium containing 0.1 μM dexamethasone, 50 μM Asc2P, and 10 mM μ-glycerophosphate (osteogenic basal medium; Lonza Co.) with 10% human serum. ASC cultures were maintained in this medium for 4 weeks (with medium changes every 3 days). For detection of extracellular calcium deposits, Alizarin red staining was used in formalin-fixed cultures; Alizarin red solution (0.2 g/l water) was incubated for 2-3 minutes, until the reaction was observed microscopically. Excess dye was removed by several washing steps using water. The orange-red calcium precipitates were quantified from at least four different pictures of each sample by using ImageJ software. Chondrogenesis: The ASC culture was performed from cell "Micromass" starting with a high concentration of cells in a minimal volume (1 × 105 cells/100 μl) in the presence of TGF-β 1 and 3 (10 ng/ml), Asc 2P (50 μM), and insulin (6.25 μg/ml) (Chondro BulletKit; Lonza Co.) for four weeks with medium changes every 3 days. Alcian blue (0.1 g/l in water, pH 1.0) was used to detect the presence of enrichment of sulphated proteoglycans in the extracellular matrix. Before staining, the micromass cultures were fixed in formalin, embedded in paraffin, and sectioned into 10 μm. Blue staining was quantified from 4 different pictures of each sample with ImageJ software.
2.7. OA Mouse Model and Cell Transplantation
Twelve-week old adult male C57BL6 mice were experimentally induced to have severe OA in both knees by a single injection of 13 U collagenase II in 6 μl of 5 mM CaCl2 [25]. Five days later, when the OA by severe cartilage damage was induced, all animals were distributed into 4 groups ( per group). The control group was injected with 6 μl of growth cell culture medium; 105 cells in 6 μl of SVF or hASC (from suprapatellar fat pad) were injected in the SVF or hASC groups, respectively. Six μl of thawed pooled human platelet rich plasma (PRP) was injected in the PRP group. All intra-articular injections were performed by using a 30G Hamilton syringe connected to a Remote Infuse/withdraw pump 11 elite nanomite programmable syringe pump (Harvard Apparatus) at a 2 μl/min infusion rate. The PRP was isolated from blood collected from all patients in citrate tubes and was prepared following the standardized method described in Anitua et al. [26] and pooled to minimize individual modifications. All animals were sacrificed one month after OA induction, and knee joints were collected. The experimental protocol was approved by the Animal Care Committee of the Centro de Investigación Principe Felipe (Valencia, Spain) in accordance with the National Guide to the Care and Use of Experimental Animals (Real Decreto 53/2013). The leg was isolated and osteomized in the middle of the femoral shaft and in the middle of the tibia and fibula to get the whole joint. Most of the adherent connective tissue, including muscle, ligaments, and tendons, surrounding the knee were removed preventing any damage to the cartilage. One knee was used for histological analysis and the other knee for the quantification of glycosaminoglycans (GAGs).
2.8. GAG Analysis
Immediately after tissue dissections, GAGs were individually quantified for tibia and femur portions from one treated knee of all animals in all groups. Weighted samples were digested in 2.5% papain solution by overnight incubation at 65°C. The supernatant was incubated with 10% Alcian blue working solution containing 0.25% of Triton X-100 in 18 mM H2SO4 and then incubated with dissociation solution (4 M guanidine HCl, 0.375 Triton X-100 in 0.027 M H2SO4) for 30 min. The precipitated GAGs were suspended in 8 M guanidine HCl and quantified at 570 nm. A chondroitin sulfate standard curve was prepared in parallel.
2.9. Safranin-O Staining and Histological OA Scoring
Bones fixed in 4% PFA were incubated for 7 days in 0.1 M ethylene diamine tetra-acetic acid (EDTA) for decalcification before paraffin embedding and frontally sectioning the entire knee into 5 μm sections. Deparaffinized and hydrated slices were first stained for 5 min with hematoxylin (Sigma) and, after several washes, quickly destained in acid EtOH, stained with fast green (0.001%) solution for 5 minutes, quickly rinsed with 1% acetic acid solution, and then stained with 0.1% safranin-O solution for 5 minutes. Dehydrated samples were mounted in Eukit. Bright-field pictures of all series (every 4th section) were acquired for OA score quantification following the recommendations established by Glasson et al. [27]. The entire joint was analysed at all four quadrants and through multiple step sections through the joint following the 0–6 subjective scoring system, where 0 is normal; 0.5 means loss of safranin-O without structural changes; 1, small fibrillations without loss of cartilage; 2, vertical clefts down to the layer immediately below the superficial layer and some loss of surface lamina; 3, vertical clefts/erosion to the calcified cartilage extending to <25% of the articular surface; 4, vertical clefts/erosion to the calcified cartilage extending to 25–50% of the articular surface; 5, vertical clefts/erosion to the calcified cartilage extending to 50–75% of the articular surface; and 6, vertical clefts/erosion to the calcified cartilage extending to >75% of the articular surface.
2.10. Immunoassay
Both fixed cells on a cover slip from osteogenic-directed differentiation or the chondrogenic-directed differentiated as well as fixed bone (after demineralization by EDTA immersion) paraffin-embedding sections were subjected to protein expression analysis by specific immunostaining. The paraffin-embedded sections were first deparaffinized. Both were fixed with 4% paraformaldehyde at room temperature for 10 min and washed with PBS, permeabilized with a PBS solution containing 0.1% Triton X-100, and blocked with 5% goat serum in PBS for 1 h. The following primary antibodies were diluted in blocking solution and incubated overnight at 4°C. Polyclonal rabbit α-Sox9 (Millipore (1 : 200) or monoclonal mouse α-Connexin 43 (Invitrogen; (1 : 200). After being rinsed three times with PBS, the cells were incubated with Alexa647 dye conjugated goat anti-mouse IgG (1 : 400; Invitrogen, CA, USA) secondary antibody and Phalloidin-Alexa 488 (Invitrogen, CA, USA) for 1 h at room temperature. Tissues were incubated with Oregon Green-488 dye conjugated goat anti-rabbit IgG (1 : 400; Invitrogen, CA, USA) secondary antibody. All samples were counterstained by incubation with 4,6-diamidino-2-phenylindole hydrochloride (DAPI) from Molecular Probes (Invitrogen, USA) for 3 min at room temperature followed by washing steps. Samples were mounted using Fluor Save Reagent (Calbiochem, USA). Signals were visualized by confocal microscopy (Leica, Germany); at least 6 different fields per condition and assay were analysed.
2.11. PET Analysis
For PET image acquisition (Albira Si PET subsystem; Oncovision, Spain), an additional group of animals was included. After OA induction, the right knee was injected with medium (control) and the left knee was injected with 105 hASC in 6 μl each, in order to minimize individual differences of the NaF-18 probe signal distribution and acquisition. The probe was i.v. injected though the tail with a range of activities between 177 and 199 uCi with uptake times that varied between 83 and 110 min. In all cases, the whole body PET scan time was 10 minutes. The images were analysed by using Amide® software. The mean ± SD of the ROIs at each condition of all animals was determined.
2.12. Statistical Analysis
Statistical comparisons were assessed by Student's t-test. All values were derived from a two-tailed statistical test using the SPSS 11.5 software. A value <0.05 was considered statistically significant.
3. Results
3.1. Suprapatellar Fat Pad SVF Is Enriched in CD105 Positive Cells
The supra- and infrapatellar fat pads of twenty-four patients with severe OA were resected during the surgical intervention for prosthetic implantation. After tissue processing, total cell counting rendered no significant differences in the amount of nucleated cells between the supra- or infrapatellar samples (7.8 × 105 ± 2.8 × 105 infrapatellar cells versus 8.8 × 105 ± 4.4 × 105 suprapatellar cells/g of adipose tissue; Supplementary Figure available online at https://doi.org/10.1155/2017/4758930). The immunophenotypic analysis of the SVF showed in both cases a positive population for a panel of typical human mesenchymal cell markers including CD90, CD29, CD44, CD117, and CD105 (Figure 1) [28]. Interestingly, the suprapatellar-derived SVF contained a significant enrichment of CD105(+) cells in comparison with the infrapatellar-derived cell suspension (Figure 1). No significant differences were detected in any other assayed marker, including the hematopoietic precursor cell marker CD34 or the mature leucocyte cell marker CD45.
3.2. The Suprapatellar ASC Show a Faster Proliferative Profile
ASC cultures derived from both supra- and infrapatellar tissues were allowed to reach cell confluency and were kept for three passages before cell proliferative analysis in the presence of human serum (HS) or fetal bovine serum (FBS). For colony-forming unit (CFU) quantification, a clonogenic assay was performed (Figure 2(a)). When ASC were grown in the presence of FBS, no differences were found. However, when ASC were grown in the presence of HS, there was a significant increase in the number of CFU and indeed the suprapatellar-derived ASC showed a higher CFU in comparison with the infrapatellar fraction (Figure 2(a)). The colonies formed from the suprapatellar SVF were not only more abundant, but they were always bigger (Figure 2(a), right panel) and exhibited a faster proliferative profile. In fact, when the ASC were cultured with either FBS or HS, the proliferative rates, in terms of the number of cells per growth area, within a cell growth curve daily analysis, were significantly higher from those cells amplified from the suprapatellar fat tissue compared with infrapatellar. The differential growth rate was significantly different eight days after culture in the presence of HS and ten days after culture in FBS (Figure 2(b)). The ultrastructural analysis of the suprapatellar cell cultures grown in FBS or HS showed higher inclusion body accumulation (indicated by white arrows) and higher distribution of intermediate filaments (indicated by black arrows) in the presence of HS-containing medium (Figure 2(c)).
3.3. In Vitro Osteogenesis and Chondrogenesis Are Favored in Suprapatellar ASC
To further assess the stem cell characteristics from both adipose tissue sources, the supra- and infrapatellar fat pads, we compared the differentiation multipotency between both samples (Figure 3). Both underwent induced differentiation into the adipogenic, osteogenic, and chondrogenic lineages (Figure 3). However, the extent of differentiation was not identical. Suprapatellar-derived ASC showed higher osteogenic and chondrogenic potential in comparison with the infrapatellar-derived and infrapatellar-differentiated cells as indicated by quantification of Alizarin red and Alcian blue staining, respectively (Figure 3, right panels). The higher chondrogenic potential of the suprapatellar-derived cells was also supported by the increased expression of Sox9 (green), a chondrocyte precursor marker [29]. This differential multipotency yields the suprapatellar fat pad a promising cell source for cartilage tissue repair.
3.4. Suprapatellar-Derived ASC Significantly Improves Cartilage Regeneration in a Mouse Model of Severe OA
To assess whether the increased cell proliferative and cell differentiation properties shown in vitro of the suprapatellar-derived MSCs would be functionally efficient in cartilage regeneration, in vivo cell transplantation assay in a severe OA mouse model was performed.
For the therapeutic efficacy testing of the suprapatellar-derived human ASC (hASC), severe OA was induced by single intra-articular injection of collagenase II (13 U) in both knees of 12-week old adult male C57BL6 mice. Five days after injection, when total cartilage destruction occurs (previously evaluated by histological analysis; data not shown), 105 hASC or the SVF or growth medium (control) was injected into the intra-articular space. It has been shown that PRP infiltration, increasingly implemented in regular clinical practice, can reduce pain and improve joint function with a notable improvement on the quality of life of patients [30, 31]. Therefore, PRP was also injected in an additional group of animals (PRP) in order to compare the regenerative effectiveness on cell transplantation in comparison with growth factor infiltration (Figure 4).
Once a week after treatments, the volume of the knees was measured using an automatic caliper. As shown in Figure 4(a), a significant increase in knee size occurs in all three groups after OA induction during the whole experiment. However, all treated groups, hASC, SVF, and PRP, showed a significant reduction of this increased volume one month after OA induction compared to the control group (Figure 4(a)). This result indicates a significant anti-inflammatory effect of the treatments. However, when we examined the structural regenerative effect by quantifying GAG (Figure 4(b)) and the OA damage score (Figure 4(c)), we observed a unique consistent structural regeneration of the loss of cartilage when the animals were treated with hASC in comparison with the control group (Figures 4(b) and 4(c)). Thus, PRP or SVF acts as an effective anti-inflammatory agent, but the regenerative effects of the stem cells do not seem to be bolstered by PRP or SVF. A representative safranin-O staining image for each treated group showed a complete absence of articular cartilage in the control animal and safranin-O-positive areas compatible with regenerating cartilage more prominent in the hASC-treated animal (Figure 4(d)).
To explore the contribution of the ectopic ASC to the regeneration of the articular cartilage, we evaluated the induced chondrogenic activity by the detection of Sox9 (a chondrocyte progenitor marker) positive cells in the control and supra-hASC-treated joints (Figure 4(e)). As shown in the representative images, an increase in Sox9 positive cells was detected in the hASC group (Figure 4(e)). However, few of the Sox9 positive cells costained with the specific immunodetection for human cells (antinuclei; data not shown) indicating an endogenous induction of the cartilage repair by the hASC transplantation.
3.5. Positron Emission Tomography (PET) Analysis Shows In Vivo Cartilage Regeneration Process after ASC Trasplantation
18F-NaF PET/CT is a high-performance diagnostic tool utilized by clinical practitioners and produces high-quality images within a shorter period from injection time in bone and joint disorders including osteoarthritis [32, 33]. PET analysis was employed to evaluate cartilage regeneration in vivo in a group of animals with induced OA in both knees. In order to minimize the individual differences of the NaF-18 probe signal distribution and acquisition (injected one month after OA induction), both conditions were included in the same animal; the left knee was transplanted with hASC and the right knee served as a control (Figure 5). After quantification of the ROIs from the acquired images at equivalent coronal and sagittal planes, a significantly lower detection of NaF-18 on the joints transplanted with hASC (Figure 5, right graph) compared to the control joints was observed. The bone at the hASC transplanted joints was less exposed, and the signal was masked by the regenerated cartilage.
4. Discussion
ASC-based therapies have already been tested in clinical trials, at the safety and efficacy phases, with remarkable restorative activity in the treatment of OA from subcutaneous abdominal fat [34, 35]. However, tissue-specific-derived stem cells would guarantee the improvement of ASC repair capabilities for instance, in terms of improved chondrogenic efficiency for cartilage repair [9]. In the knee, there are two major sources of ASC, the supra- and infrapatellar fat pads [14, 15]. Both fat pads are commonly resected during knee joint arthroplasty in patients with acute OA and are a suitable source of ASC for the autologous treatment of additional affected joints. Previous studies have already described the potential regenerative capability of ASC derived from the infrapatellar pad, recently shown to have demonstrated increased chondrogenic potential compared with those from subcutaneous fat in vitro [36] and in vivo in a model of OA [16, 17, 37]. However, no data have been reported yet with the suprapatellar-derived cells. Here, we show for the first time the regenerative capacity of the suprapatellar-derived ASC population in a severe OA mice model. The infrapatellar pad-derived cells have been considered an active joint tissue in the initiation and progression of knee OA, activating the inflammatory cells by producing inflammatory mediators influencing the cartilage and synovium metabolism [38]. When we compare in vitro the proliferative and differentiated properties among both derived ASCs, we demonstrate improved proliferative, chondrogenic, and osteogenic rates from the suprapatellar-derived ASC, showing a promising and improved source for efficient cell therapeutic resolution of joint degeneration [33].
The CD105 or endoglin receptor has been related to the more proliferative, migrating, and invading MSC size populations in the injury area during endogenous regeneration [39]. Moreover, transplantation of a CD105-negative stem cell population showed significant chondroid degeneration features compared to the CD105-positive transplanted group [40], showing an influence on chondrogenic cell fate induction. In fact, when we compare the directed chondrogenic capability between the two samples, we found a better yield from the suprapatellar-derived ASC that shows an enrichment of CD105-positive population versus those from the infrapatellar fat.
The ASC amplification in vitro with human serum was always better than with fetal bovine serum. Previous reports from subcutaneous adipose tissue have already shown detrimental proliferative activity of ASC in the presence of FBS [41]. Differential gene expression analysis of ASC cultured with HS or FBS showed a significant overexpression of a number of genes related with regulatory roles in cell cycle progression [42].
For translational and functional proof of concept, an in vivo model of severe OA [25] for intra-articular administration of the hASC was explored comparing the efficiency of cartilage regeneration in the SVF- and PRP-treated animals. Interestingly, all treated groups showed significantly reduced knee joint diameter (swelling).
PRP contains a multitude of growth factors including TGF-β, PDGF, IGF, bFGF, VEGF, and EGF [43] with anabolic chondral-promoting and chondral-protective properties [44, 45]. Abundant preclinical and clinical findings support that PRP is a promising coadjutant treatment for cartilage repair and relieving symptoms based on its anabolic effect on the resident cells and due to its potential to inhibit inflammation and alleviate OA symptoms with a clinically acceptable safety profile [46]. PRP treatment has shown additional benefits in the regeneration of other related tissues such as bone defects, indicating PRP plays a crucial role by influencing the local tissue microenvironment and thereby enhancing progenitor cell recruitment and proper matrix deposition [47]. Although only the ASC transplantation showed significant tissue regeneration, the PRP combination could create a synergistic effect by bridging and interfacing between the MSCs and the endogenous tissue, an effect not produced by an individual treatment alone. Chemically induced OA in the mice led to aggressive joint degeneration; however, even in such a severe condition, ASC transplantation reduced and rescued functional anatomy in the intra-articular space, reducing the OA damage scores and increasing the cartilage-like tissue. Continued expression of Sox9 is required to maintain hyaline-like cartilage, avoiding chondrocyte hypertrophy for efficient regeneration [29]. Only the amplified hASC were able to induce efficient regeneration one month after treatment showing a significant increase in SOX9 expression in the growing cartilage. One month after cell transplantation, very few human cells were found in the transplanted joints (data not shown), which is in accordance with previous reports [48, 49]. Rather than assuming cell replacement by ASC transplantation, the most contemporary hypothesis is that their actions are mostly associated with paracrine activity by secreting modulating factors. Accordingly, ASC secrete PGE2 which induces dendritic cells to upregulate the anti-inflammatory cytokine IL10 while reducing the proinflammatory TNFα and IL12 contributing to its known immune-modulatory effect and related promotion of tissue repair [50, 51]. However, this hypothesis needs to be addressed with further research to demonstrate its relevance for the treatment of OA.
5. Conclusion
Our results show that the human suprapatellar fat pad offers a proper ASC source for cartilage regeneration by promoting efficient endogenous chondrogenesis in a mouse model of severe OA.
Conflicts of Interest
The authors declare that they have no conflicts of interest.
Authors' Contributions
Ignacio Muñoz-Criado, Victoria Moreno-Manzano, Ramón Cugat, and Montserrat García contributed to the conception, design, collection, and/or assembly of data, data analysis and interpretation, manuscript writing, and final approval of manuscript. Victoria Moreno-Manzano, Ramón Cugat, and Montserrat García provided financial support and approved the final manuscript. Jose Meseguer-Ripolles, Ana Alastrue-Agudo, Jerónimo Forteza-Vila, Richard J Griffeth, and Maravillas Mellado-López contributed to the collection and/or assembly of data and data analysis and interpretation. Ignacio Muñoz-Criado and Jose Meseguer-Ripolles equally contributed to this work.
The authors specifically acknowledge the Fundación García Cugat for its valuable financial and intellectual support. This work was also supported by Ministerio de Economía y Competitividad (grants CONSOLIDER-INGENIO 2010 CSD2008-00005 and MAT2015-66666-C3-2-R, MINECO/FEDER, UE) and the Instituto de Salud Carlos III (PI13/00319). The authors also thank Oncovision for performing the PET analysis and for directly contributing to acquisition and analysis.
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TransLink has received $1.2 million in federal funding to install and operate overhead charging stations to get four electric buses on the road next year.
TransLink CEO Kevin Desmond says the four "fast charge" electric buses will operation on route 100 with overhead charging stations installed at Marpole Bus Loop and 22nd Avenue SkyTrain Station. Desmond says charging time is expected to take four to five minutes and adds that passengers can expect a "quieter ride" compared to a diesel-powered bus. The performance of the electric buses will be evaluated and data shared with B.C. Hydro.
Mayor Gregor Roberston says "we are thrilled here in the region to be leading the charge in getting electric buses on the road. "Here in Vancouver we have the largest electric fleet of any city in Canada right now in terms of our city vehicles and we're looking at an even more aggressive shift happening in these years ahead."
Robertson says the project has received $7 million from the federal gas tax fund. In total the project cost $10 million with the rest of the funding coming from the City of Vancouver, Natural Resources Canada and B.C. Hydro.
Josipa Petrunic, executive director of the Canadian Urban Transit Research & Innovation Consortium (CUTRIC) says the system "allows people to benefit from zero emissions transit and transportation while seamlessly being able to get on and off buses while they actually charge in operation. That is a world first."
Petrunic says buses can be charged at 450 or 600 kilowatts, which can bring charging times down to "a couple of seconds or less than a minute" if the bus is running behind schedule. During non-peak periods the bus can be charged at 350 or 100 kilowatts to reduce electricity costs and save the battery.
Canada's leading bus manufacturers New Flyer Industries in Manitoba and Nova Bus in Quebec will be developing two electric buses each. ABB and Siemens are developing the chargers.
Petrunic says the $40 million trial project includes 18 electric buses and seven overheard chargers in total in three cities across Canada. Vancouver is the first to launch ahead of Brampton and York Region in Ontario.

subset: commoncrawl


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Barchester Towers (Barnes & Noble Classics Series)
- New introductions commissioned from today's top writers and scholars
- Biographies of the authors
- Chronologies of contemporary historical, biographical, and cultural events
- Footnotes and endnotes
- Selective discussions of imitations, parodies, poems, books, plays, paintings, operas, statuary, and films inspired by the work
- Comments by other famous authors
- Study questions to challenge the reader's viewpoints and expectations
- Bibliographies for further reading
- Indices & Glossaries, when appropriate
The second and most popular of Trollope's six Barsetshire novels, Barchester Towers chronicles the struggles for power and position in an imaginary county in Victorian England. Passions start seething when an "outsider," Dr. Proudie, is appointed bishop of Barchester. Soon, his ambitious, domineering wife and the smarmy, scheming curate, Mr. Slope, are hatching plots and counter-plots as they try to control the choice of a new warden for Hiram's Hospital and a new husband for Eleanor, a lovely young widow and the daughter of the former warden, Mr. Harding.
The novel combines the realism of later fiction (including Trollope's own) with such Victorian devices as Dickensian character names and a comically interruptive narrator. The narrator's sharply satiric comments enhance the story's richness, while his playful, reassuring, and mocking asides subvert the reader's expectations, giving the book an unexpectedly post-modernist flavor. Ultimately, we see that Trollope's characters' petty jealousies, selfishness, and meanness are not metaphors for larger issues, they are the issuesthe same human failings that, in other contexts, can lead to serious social strife and civil unrest.
Edward Mendelson is Professor of English and Comparative Literature at Columbia University. He is W. H. Auden's literary executor and has written widely on nineteenth- and twentieth-century novels.
Product Details
|Barnes & Noble
|Publication date:
|Barnes & Noble Classics Series
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|Product dimensions:
|7.96(w) x 5.32(h) x 1.48(d)
About the Author
Anthony Trollope (1815-1882) was one of the most successful, prolific, and respected English novelists of the Victorian era. Some of his best-known books collectively comprise the Chronicles of Barsetshire series, which revolves around the imaginary county of Barsetshire and includes the books The Warden, Barchester Towers, Doctor Thorne, and others. Trollope wrote nearly 50 novels in all, in addition to short stories, essays, and plays.
Read an Excerpt
From Edward Mendelson's Introduction to Barchester Towers
Barchester Towers pretends to be nothing more than a comic novel about fierce but harmless battles over power, status, and marriage that divide the secure world of an English cathedral town, a place where nothing serious ever happens. Anthony Trollope narrates his story in a familiar and realistic style, as if he were telling a story after dinner, a style that prompted Nathaniel Hawthorne to praise Trollope's work as "solid, substantial, written on the strength of beef and through the inspiration of ale, and just as real as if some giant had hewn a great lump out of the earth and put it under a glass case." Barchester Towers is one of the greatest comic novels ever written, but it is also much more than that. In its portrait of a small community altered by vast changes in politics, religion, and culture, it is also an ambitious study of the ways in which large forces affect individual lives. And in its account of the anarchic power of sex in the repressed and proper world of an Anglican cathedral town, it rises to the level of myth: It is a story in which the instinctive force of Eros disrupts the civilized order and defeats those who want power rather than love.
Trollope can always be relied on for a readable story with a varied cast of plausible characters in a plausible social setting, and for many decades he was valued for the comfort and security that his books seemed to offer. One widely cited example: In England, sales of his novels markedly increased during World War II, especially in times of massive bombing raids over London. Trollope also had a darker edge than he wanted to admit to himself, especially in such later novels as The Way We Live Now, a cynical account of personal and public corruption, and He Knew He Was Right, a story of obsessive and destructive marital jealousy. Even his lighter novels, such as Barchester Towers, are complicated by Trollope's mixed feelings about relations between men and women, about historical and technological change, and about the nature of novel writing itself. Trollope often expresses his feelings about culture, society, and the sexes with table-thumping intensity, but his most intense feelings were always mixed feelings, even if he was not consciously aware of how mixed they were.
The love stories in his novels illustrate his way of combining overt simplicity and covert complexity. His love plots almost invariably involve a diffident young man who is too tongue-tied to express his love and a shyly feminine young woman who manages to misunderstand the young man's intentions until almost the last minute, when she accepts in the most modest way imaginable, and then withdraws to exult in private. Trollope repeatedly declares his preference for the reticent, unassertive women suitable to this kind of plot, and equally often declares his distaste for overbearing women such as Mrs. Proudie in Barchester Towers and for the independent-minded young feminists who appear in his later novels. Yet Trollope tends to give his unassertive young heroines at least one moment of assertiveness that he clearly regards as triumphant. Eleanor Bold, the retiring, beautiful young widow in the love plot of Barchester Towers, seems to be the exact opposite of the meddling, finger-pointing, domineering Mrs. Proudie; butt by the end of the book they have both acted against the same enemy (although neither knows about the other's action), and Eleanor has resorted to a small but momentous act of physical violence when defending herself against sexual aggression. Trollope devotes four extravagantly comic paragraphs to his own response to this violent act, first imagining the disgusted reactions of imaginary readers, then defending Eleanor against those reactions, then affirming the perfect suitability of her action to the provocation she received, finally pretending to conclude that she ought not to have done it—but only because she herself feels ashamed of it. Trollope admires enterprising, independent-minded women more than he wants to admit to himself, and his mixed feelings can be traced to his memories of his mother, the formidable Mrs. Frances Trollope, who took over the family finances from Trollope's father when his law practice failed and then transformed herself into a well-known and prolific author of novels and travel writings who wrote forty-one books—not as many as her son's sixty-seven, but an impressive number for any writer.
Customer Reviews
Average Review
Most Helpful Customer Reviews
As a 52 year old accountant I'm enrolled in a one month class at my alma-mater entitled "Study in the Victorian Novel". We are required to read this book as one of four in a thirty day period. I was overwhelmed but this book went by so fast. The characters are so engaging, the story is interesting and the author is so amusing. I've already ordered other books by Trollope. I'm currently reading "Adam Bede" by George Eliot and it's not nearly as riveting. Don't be intimidated by this book - dive in!
This is a great classic! Anthony Trollop has the amazing insignts into human nature and brings out characters which are so close to life. One can always relate to situations and personalities in real life. None of the characters in his story are absolutely good or bad, good ones are not perfect, bad ones have some endearing trait as well. The plot is also well structured and entertaining.
Great stuff... wavered a bit at one point, this is a pretty long Trollope, but fun of course.... I'll eventually read all the Barsetshire books!
I enjoyed this second installment of the Barchester Chronicles much more than the first, The Warden, even though many of the same characters (Mr. Harding and his daughters) continue the story. Trollope has a unique way of getting inside the minutiae of Victorian society and behavior, and he does so with both affection and humor. I'll definitely continue reading the series.
Who will marry Eleanor Bold? That is one of several plot lines in Barchester Towers which is a novel set in the English countryside in the 1860s. Other plots ask Who will be the warden of Hiram Hospital? Which, if any, of Mr. Slope's devious plans will work out? Will the domineering Mrs. Proudie get her comeuppance? Will the archdeacon be scandalized by the conduct of his sister-in-law? Trollope's characters are memorable and very well drawn. They ring true in all eras. You will find you in fact know people like these. In developing his characters Trollope rises almost to the exquisite insights of Jane Austen; however, Trollope has more plots and more intricate plots than does Austen. His novels have a greater page-turning quality about them. Barchester Towers is the second in a series of six novels that make up the Barchester Series. The first is The Warden which is short and not necessary to a full appreciation of Barchester Towers. If you like Barchester Towers, and it is hard to imagine anyone who wouldn't, you will also enjoy the remaining novels in the series--Dr. Thorne, Framley Parsonage, The Small House at Allington, and The Last Chronicle of Barset. The last four I have read as e-books on my Nook which I find to be a good reading experience; however, I have one word of caution. When buying an e-book don't waste your time on the free ones from Google. I have tried about five of these and only found one to be satisfactory. The others were either not complete when downloaded or had serious defects; e.g., in one the letter "e" was systematically replaced with an "o". "The" became "Tho" and "She" became "Sho" which essentially made it unreadable. Even the $0.99 versions are not quite satisfactory. The two I tried had double or triple spacing between paragraphs so that not much text appeared on the page. Spring for the $2.99 editions for a decent reading experience.
Too much talking and an overabundance of footnotes.
Don't bother


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Part of Departure Lounge Festival 2019
Friday 19 July, 1:30pm - 2:20pm
Venue: Bistro
Work In Progress/ Midlands Work
How do I be a woman?
How will I know if I've fallen in love?
What shape should my eyebrows be?
Can your virginity grow back?
Can putting a tampon in kill you?!
Combining spoken word, stand up and storytelling set to a 90's and 00's pop and hip hop soundtrack, Georgie takes a break from haphazardly stumbling through adulthood to revisit first times, second chances, push up bras and waxing disasters in order to figure out the things she thought she would have known by now.
"Charming and touching, laced with warmth and hilarity at every turn" (LondonTheatre1)
"A warm, witty and engaging monologue that genuinely connects with the audience" (The Reviews Hub)
Festival Passes
Earlybird Festival Pass - Book by Friday 31 May
See all Departure Lounge events across all three days for just £60! (£90 after Friday 31 May)
DT Friends Festival Pass
£60 (if you are a member of our Friends scheme)
Friday Day Pass
See all events on Friday, the second day of Departure Lounge, for just £35.


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Michelle Obama Hilariously Breaks Down Why Melania Trump's Inauguration Gift Exchange Appeared So Awkward
The former FLOTUS also explained to Ellen Degeneres how Barack stepped in and saved the day.
Martavis Bryant needs to grab control of his life.
On Monday, the NFL suspended the Pittsburgh Steelers' star wide receiver for at least one year due to the 24-year-old violating the league's substance-abuse policy, as reported by ESPN.
With the suspension, he will lose out on the $600,000 he was set to earn over the 2016 NFL season.
"He is at a crossroads of his professional life, and he needs to understand significant changes need to occur in his personal life if he wants to regain his career as a Pittsburgh Steeler," Steelers GM Kevin Colbert told ESPN about the embattled Bryant. "We are hopeful that Martavis will take the necessary steps to develop the discipline in his personal life to become a successful player and a good teammate."
Perhaps acknowledging that he needs help, Bryant has decided to not appeal the suspension, according to ESPN.
"This is the biggest cry for help I've ever seen, and that hurts. It hurts us to see," one of his agents, Brian Fettner, told ESPN. "He's 24 years old, and he's got to get right, whatever it is. If you talk to anybody's family that has depression, they will be talking about these same things — the [despondence], the withdrawal, the head-in-the-sand despair — just trying to cope."
A source additionally told the Worldwide Leader in Sports that the suspension is related to the league's drug testing.
This comes a year after the NFL suspended Cleveland Browns receiver Josh Gordon for one year for also violating the league's substance-abuse policy.
BET Sports News — Get the latest news and information about African-Americans in sports, including weekly recaps, celebrity news and photos of your favorite Black athletes.
(Photo: Matthew Emmons/USA TODAY Sports/Reuters)
The former FLOTUS also explained to Ellen Degeneres how Barack stepped in and saved the day.
"When my teacher asks me a question thinking I'm not paying attention, but I get it right."


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Agra to Bhusawal Trains
Both these cities are located at a distance of 906kms. Traveling from one place to the other is easy because of the efficient rail connectivity. There are many trains that run between the two cities. On a daily basis, 7 trains run between Agra and Bhusawal. On a weekly basis, the count shoots up to 14.
First train that leaves Agra and takes you to Bhusawal is 12628 Karnataka Express. It departs from Agra at 00:05 AM and reaches Bhusawal at 01:24 AM. The last train that leaves Agra for Bhusawal is 12162 Lashkar Express. It leaves at 23:50 PM and reaches the destination at 00:45 AM.
22110 H Nizamuddin Lokmanyatilak Terminus Ac Express is the fastest train that will take you from Agra to Bhusawal. It takes 11h 36m and leaves Agra at 17:59. The maximum duration of 21h 0m is taken by Gondwana Express 12406. It departs from Agra at 18:50.
Out of all the trains that operate between Agra and Bhusawal,2 originate from Agra while the rest 19 pass from Agra.
If you are looking for a way to plan your travel between these two cities, then Cleartrip will have all your answers. With its easy-to-use mobile app and website, you may book your rail tickets in a jiffy. Cleartrip offers all the information you need and ensures that your experience is quick as well as seamless.
Agra Bhusawal Train Schedule
Trains Originating from Mumbai
Trains Passing from Agra
Frequently Asked Questions
- Which trains run from Agra to Bhusawal? The following trains start from Agra - Agcltt Special, Lashkar Express. In addition to this, trains like H Nizamuddin Lokmanyatilak Terminus Ac Express, H Nizamuddin Lokmanyatilak Terminus Ac Superfast, Karnataka Express pass through Agra to reach Bhusawal.
- How many weekly trains are there from Agra to Bhusawal? There are 66 weekly trains from Agra to Bhusawal. Following are the train names and their respective train number - Agcltt Special (1016), Lashkar Express (12162), Mngla Lksdp Express (12618).
- When does the first train leave from Agra? The first train from Agra to Bhusawal is at 00:05 AM, which is Karnataka Express (12628) followed by Amritsar Junction Lokmanyatilak Terminus Express (11058) which leaves at 00:35 AM.
- When does the last train leave from Agra? The last train from Agra to Bhusawal is at 23:50 PM, which is Lashkar Express (12162).
- What are the different stations in Agra & it's respective code? Different station in Agra are Agra Cantt (AGC), Agra Fort (AF), Idgah Agra Junction (IDH), Raja Ki Mandi (RKM).
- What are the different stations in Bhusawal & it's respective code? Bhusaval Junction with station code BSL is the only station present in Bhusawal.
Other Routes From Agra
- Agra to New Delhi trains
- Agra to Mathura trains
- Agra to Jhansi Railway Settlement trains
- Agra to Morar Cantt trains
- Agra to Bhopal trains
- Agra to Itarsi trains
- Agra to Faridabad trains
- Agra to Delhi trains
- Agra to Morena trains
- Agra to Kanpur trains
- Agra to Dhaulpur trains
- Agra to Tundla trains
- Agra to Dabra trains
- Agra to Bina Etawa trains
- Agra to Datia trains
- Agra to Nagpur trains
- Agra to Hoshangabad trains
- Agra to Ambala trains
- Agra to Mughalsarai trains
- Agra to Khandwa trains
- Agra to Jaipur trains


text:
Kunstmuseum von Estland
56 Einheimischen empfohlen ·
Über diesen Ort
Kumu is the headquarters of the Art Museum of Estonia, as well as the largest and most impressive exhibition venue in Estonia.
Unterstützt von Foursquare
Tipps von Einheimischen
Go if you're into art and architecture. You can also visit Kadriorg park and president's castle on the way to Kumu.
Kumu, which opened in 2006, is the new and grand headquarters of the Art Museum of Estonia, which attracts everyone that is interested in Estonian art and culture. Both Estonian art classics as well as the works of artists that cultivate modern trends are on display at Estonia's largest museum. In…
Modern architecture and large art museum next to president palace and large park Kadriorg.
By far the nation's largest and most cutting-edge art museum, Kumu displays Estonian-created works from the 18th-21st centuries. A must-see for culture creatures, Kumu, the main building of Art Museum of Estonia, serves both as Estonia's national gallery and as a centre for contemporary art.
1 Valge
Kesklinn, Harju maakond 10127
Telefonnummer+372 602 6000
ÖffnungszeitenGeschlossen · Öffnet wieder: Tuesday 10:00 AM
Yes (incl. American Express & MasterCard)
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text:
NEW YORK (Reuters Health) - Obese men are more than three times as likely to have low sperm counts compared with their normal-weight peers, a study out this month in the journal Fertility and Sterility shows.
The heaviest men were also at triple the risk of having a low count of progressively motile sperm — sperm that swim forward in a straight line, Dr. Ahmad O. Hammoud of the University of Utah in Salt Lake City and colleagues found.
"There is a strong relationship between overweight and obesity and altered sperm parameters," Hammoud and his team write. Future studies should look at this relationship over time, they say, as well as how weight loss might affect sperm characteristics.
There has been some evidence that obesity may impair fertility in men, the researchers note, while increased body fat can contribute to lower testosterone levels and higher estrogen levels. To examine how body mass index (BMI) affects sperm quality, they evaluated 390 men who sought infertility treatment with their partners over a 2-year period.
Twenty-four percent of the men were normal weight, 43 percent were overweight, and 33 percent were obese. Overall, 10.5 percent had low sperm counts.
The prevalence of low sperm counts rose as BMI increased; obese men were 3.3 times more likely to have low sperm counts than normal-weight men. The risk of having a low count of progressively mobile sperm also rose with BMI; obese men were 3.4 times more likely than normal-weight men to have a low progressively mobile sperm count. Obese men were also 1.6 times more likely than overweight or normal-weight men to have a high percentage of abnormally shaped sperm.
While there was a trend toward increasing likelihood of erectile dysfunction with increasing BMI, the relationship wasn't statistically significant. However, the researchers note, other studies have found that obesity is associated with a greater risk of impotence.
The fact that the study was done among men seeking infertility treatment makes it likely that the study group had worse sperm quality than the population at large, Hammoud and colleagues point out. Nevertheless, they add, the correlation of poor sperm quality and increased BMI in the general population is probably similar.
SOURCE: Fertility and Sterility, December 2008.


text:
View My Grades (StudentVUE)
Antietam Teacher Weblinks
FLL Robotics
VEX IQ Robotics
Mustang Coders Club
Greetings Antietam Families,
All pre-paid Yearbooks were sent home with students today, June 5, 2017. If your student pre-purchased a Yearbook and did not receive one, please send a note in with your student June 6, 2017. If you have not purchased a Yearbook and would like to do so, please send in $20.00 with your student as Yearbooks will be sold on a first come first serve basis in the main lobby from 8:10 to 8:20 AM June 6, through June 15, 2017.
Thank you for your time and attention. Have a wonderful day!
Antietam Staff
School Board Representatives for this School
Warning! This web page may contain links to one of more websites outside the PWCS network, which are not PWCS venues and may not reflect the views or opinions of PWCS. PWCS does not control the content of such websites and does not sponsor or endorse any messages, products, or services contained on such websites.
Website by SchoolMessenger Presence. © 2019 West Corporation. All rights reserved.


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La Musa de las Flores Class | Michelle Chang Photography
I was very excited when Oliva from La Olivia invited me to attend this floral workshop taught by the world famous florist Gabriella from La Musa De Las Flores. As a wedding photographer, I work closely with florists, but a lot of times, I only got the see the end side of the amazing floral design, not behind the scenes.
At this workshop, I got to learn what it takes to create a stunning centerpiece. One thing that stood out so much to me is Gabriella's centerpiece looked stunning in all angles. She said the first step in creating a centerpiece is to create the "shape." Take the time to look at each branch see the way they naturally fall and base on that form the shape you desire. After your centerpiece is formed, you would then find flower pieces as "fillers" to support the bigger branches. She emphasized on "connection and levels," which are the key points to create a centerpiece where each flower pieces looks "happy," flow naturally and freely.
Lucy from The Flower Hunter is an award-winning landscape and floral designer. She created a hanging installation with lots of greens. It's incredible to see her working her magic on the piece. She also used fresh fruits such as the blueberries, pear, and strawberries to decorate the table settings. It's the little details that make a gorgeous table feels complete.
A little bonus from the day is when cake designer Soo Jung Noh from Rice and Bloom Flower Cake Studio brought in her famous bean paste flower cakes. I've always seen beautiful pictures of the bean paste cakes but never got to try it. Of course, after I took countless photos of the cakes, we got a little taste of them. Because the flowers were made by bean paste, the amount of sugar is reduced significantly compared to the typical sugar flowers. The cake itself is made with rice flour, which had a nice chewy texture like mochi. Needless to say, the bean paste cakes has quickly become one of my favorites, and I secretly hope all my brides will have bean paste cakes on their wedding day.
Special thanks to a team of amazing vendors who helped to make the class wonderful : @sweet_kvitka@dafioridesign @pompelmoooo | Dinnerware: @theark_ Take and Chairs: @foundrentals Studio sponsor: Max Grill Design
Some of the works from the students that attended the class:


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Weekly Camp Schedule
Our Global Kids Camp Director creates specially-tailored day programs with local partners. Our team is eager to make each week super fun and in line with your children's interests.
Week 8 Schedule
Daily program focuses on learning new skills with fun hands-on activities, such as designing a robot, creating art, developing a website and learning to launch a business
Weekly "Adventure Day" tailored to campers interests: swimming, basketball, tennis, scavenger hunts, dancing classes, and more!
'A la carte' - pick and choose which weeks to enroll your child based on your family's summer travel plans.


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If your workflow is set up so that receiving a new document means taking action, Zapier can keep you efficient with automation. This one will trigger whenever Docparser extracts new data from an uploaded PDF file, and parse in detail via CandidateZip when they're added to Docparser and create new contacts in Nimble using the parsed information. That way, you can easily keep up with all of your candidates' resume information and follow-up accordingly.
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Customer Support & Sales+919996186555
|» Products
|» Applied Mechanics Lab Equipments
|» Link Polygon Apparatus
|Advanced Technocracy Inc. is Manufacturer, Exporter & Supplier of Link Polygon Apparatus
|The apparatus is provided with five linkages and connectors, having hooks to which pans are hung.These are fixed between two pulleys around which rope is run to connect to two extension type spring balances. The unit is very useful for the study of arches, abutments and piers. The wooden rectangular beam is fitted with scale.Without weights.
ADVANCED TECHNOCRACY INC. All Rights Reserved.(Terms of Use)
Developed and Managed by Infocom Network Ltd.


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Xerocomus punctilifer (W. F. Chiu) F. L. Tai
|Sichuan: Xiangcheng Xian. Pass between Xiangcheng and Reda over Ma'an Shan. Open barren area at pass with scattered Quercus scrub and scattered Larix and Picea. 29°0'7"N, 99°44'34"E; 4200-4270 m.
On ground under Quercus at 4100 m. Pileus 3-5 cm in diam., convex to applanate, dry, grayish brown to brown, tomentose. Hymenophore yellow, unchanging or slowly bluing; tubes yellow, unchanging or slowly bluing. Stipe 5-9 x 1-1.5 cm, dirty white, upper part often yellowish, with longitudinally striations; context yellowish in apical region, whitish in lower part, bluing. Taste mild.
|Z. L. Yang
|20 July 1998


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Improvement science is like the opposite of paternalism.
Instead of people in power assuming that they know best for someone else—whether the mainstream for marginalized groups, teachers for students or bosses for workers—the essential idea of improvement science is to ask. Ask students, teachers, the community what might work so that they can propose a solution, with the power to collaborate and execute a solution held firmly in their hands.
College of Education (COE) Associate Professors Deborah Peterson and Susan Carlile and their students Ryan Carpenter and Cassandra Thonstad are part of a movement to apply this concept to education.
This quartet of academics are responsible for four chapters in "The Educational Leader's Guide to Improvement Science: Data, Design and Cases for Reflection," which came out in March. They co-wrote Chapter 8, called "Preparing School Leaders to Effectively Lead School Improvement Efforts: Improvement Science."
"The basic concept is in school systems, rather than having the school board or superintendent or building principals tell the students how to improve, have the teachers or the families or the students identify the problem," Peterson says.
They also wrote Chapter 10 together, called, "Redesign of the Licensure Field-Based Experience Using Improvement Science." Carlile explained that the essential truth behind every chapter is the same, that the people who are on the ground should be the ones guiding the changes in their field because they know it best. Then, once people realize how much they know, it helps further improve the environment around them.
"You create an environment where people collaborate or move ahead with what they think is important, and from there, they gain confidence," she said.
Peterson created Chapter 14 with Carpenter, called "Using Improvement Science in Professional Learning Communities: From Theory to Practice." Carpenter said he has seen improvement science create real results in professional learning communities (PLCs).
"Using improvement science tools in the PLCs in the district I serve has proven significant growth in student achievement," he said. "Our graduation rate has risen 10% in the last two years, we are seeing double digit gains in K-5 achievement, and our staff morale is high with a 93% job satisfaction rating among certified teachers."
In addition, Thonstad authored Chapter 13, "Growth and Grading: Overcoming 'Grades Don't Matter' in Middle School." Peterson said it is a "brilliant chapter" and that Thonstad is an "amazing leader" who leverages improvement science processes to help middle schoolers improve their academic performance.
Carlile and Peterson are not the only ones in the COE who are interested in improvement science, but they gained a special affinity for it after the dean in 2016 invited them to take a course on it led by nationally recognized PSU scholar Professor Sherril Gelmon. Before long, Peterson and Carlile were actively joining improvement science conferences.
Last year, the duo presented at two conferences on employing improvement science to get students' voices head. The conferences were put on by the University Council of Educational Administration, a consortium of institutions advancing the work of educational leaders, and Carnegie Foundation for the Advancement of Teaching, which espouses improvement science and networked communities as ways to resolve inequities in education.
Carlile and Peterson came to know and gain respect from people in the field, such as Robert Crow. Crow, an editor, was compiling a multi-author book on improvement science. He invited Peterson and Carlile to submit their work, and they earned a place in the book through a blind peer review process. Peterson said that Crow called the book a "Portland State book" because of the depth of involvement from the four PSU educators.
Peterson said that she, Carlile and their students are committed to improvement science because it matters.
"It's about empowerment, and as Susan said, it's an improvement process that empowers the most marginalized, underserved and oppressed in our communities so that one's race, gender or socioeconomic status doesn't decide their future," Peterson said.
The Educational Leader's Guide to Improvement Science is available online on the Stylus Publishing website and on Amazon.com.
Carlile and Peterson may have another book coming out soon, and Carlile said that their professional collaboration has helped them both be more successful and more fulfilled.
"Deborah is kind," she said. "She's always thinking about other people before herself, and she thinks of me before herself."
Peterson said that Carlile took work off of her plate that might otherwise have held her back.
"You, Susan, gave me feedback that was kind and critical to help me improve directly and in a supportive way, and you always find the best in me and help me to be the best that I could possibly be," Peterson said.
Carlile replied, "That's why (it works) and it keeps working."
Peterson agreed, "And it keeps working."
Photo: From left to right: College of Education (COE) Associate Professors Susan Carlile and Deborah Peterson are among the authors on a new work, "The Educational Leader's Guide to Improvement Science: Data, Design and Cases for Reflection."
To share stories about the College of Education, email Jillian Daley at email@example.com.


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|Re: screen-4.6.2 build fails
|Wed, 15 Nov 2017 21:30:32 +0900
2017-11-14 1:48 GMT+09:00 Arkadiusz Drabczyk <address@hidden>: > It's not possible to build screen on BSD without patches, that's > normal. If you have root on your BSD system you can download all > patches from official BSD repository. I'm not sure how it's done on Thank you ever so much, Arkadiusz. I was able to successfully build screen-4.6.2 by applying patches from pkgsrc/misc/screen/patches. However, I can't get the configure script to find libncursesw. I have tried setting various environment variables before running configure, but either gcc compile test fails, or still -lcurses is in the LIBS in Makefile. Do you have suggestions for using ncursesw rather than plain curses? TIA Henry
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What critical conditions are covered by Trauma Insurance?
The main trauma events that NobleOak Trauma Insurance covers are:
- Cancer
- Coronary Artery Angioplasty
- Coronary Artery By-pass Surgery
- Heart Attack
- Other Serious Coronary Artery Disease
- Stroke
Other Trauma Events covered include:
- Accidental HIV infection
- Alzheimer's Disease/Irreversible Organic Disorder
- Aplastic Anaemia
- Bacterial Meningitis
- Blindness
- Cardiomyopathy
- Chronic Liver Disease
- Chronic Lung Disease
- Coma
- Dementia
- Diplegia
- Heart Valve Replacement
- Hemiplegia
- Kidney Failure
- Leukaemia
- Loss of Hearing
- Loss of Independence
- Loss of Limbs and/or Sight
- Loss of Speech
- Major Brain Injury
- Major Burns
- Major Organ Transplant
- Motor Neurone Disease
- Multiple Sclerosis
- Muscular Dystrophy
- Paraplegia
- Parkinson's Disease
- Pulmonary Arterial Hypertension (Primary)
- Quadriplegia
- Surgery to Aorta
- Terminal Illness
- Viral Encephalitis
Please refer to the Product Disclosure Statement for the medical definitions of these conditions under our Trauma Cover. The medical definitions that NobleOak uses may differ from some common perceptions of these conditions.
From 1 July 2017 NobleOak abides by the Life Insurance industry approved definitions for at least the three main trauma conditions of cancer, heart-attack and stroke. The approved definitions can be found on the website of the Financial Services Council www.fsc.org.au