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131.
Jinping Luo Lynda K. McGinnis William H. Kinsey 《Molecular reproduction and development》2009,76(9):819-831
The objective of the present study was to determine whether Fyn kinase participated in signaling events during sperm–egg interactions, sperm incorporation, and meiosis II. The functional requirement of Fyn kinase activity in these events was tested through the use of the protein kinase inhibitor SKI‐606 (Bosutinib) and by analysis of Fyn‐null oocytes. Suppression of Fyn kinase signaling prior to fertilization caused disruption of the functional polarity of the oocyte with the result that sperm were able to fuse with the oocyte in the immediate vicinity of the meiotic spindle, a region that normally does not allow sperm fusion. The loss of functional polarity was accompanied by disruption of the microvilli and cortical granule‐free zone that normally overlie the meiotic spindle. Changes in the distribution of cortical granules and filamentous actin provided further evidence of disorganization of the oocyte cortex. Rho B, a molecular marker for oocyte polarity, was unaffected by suppression of Fyn activity; however, the polarized association of Par‐3 with the cortex overlying the meiotic spindle was completely disrupted. The defects in oocyte polarity in Fyn‐null oocytes correlated with a failure of the MII chromosomes to maintain a position close to the oocyte cortex which seemed to underlie the above defects in oocyte polarity. This was associated with a delay in completion of meiosis II. Pronuclei, however, eventually formed and subsequent mitotic cleavages and blastocyst formation occurred normally. Mol. Reprod. Dev. 76: 819–831, 2009. © 2009 Wiley‐Liss, Inc. 相似文献
132.
Studelska DR Mandik-Nayak L Zhou X Pan J Weiser P McDowell LM Lu H Liapis H Allen PM Shih FF Zhang L 《The Journal of biological chemistry》2009,284(4):2354-2362
In the K/BxN mouse model of rheumatoid arthritis, autoantibodies specific for glucose-6-phosphate isomerase (GPI) can transfer joint-specific inflammation to most strains of normal mice. Binding of GPI and autoantibody to the joint surface is a prerequisite for joint-specific inflammation. However, how GPI localizes to the joint remains unclear. We show that glycosaminoglycans (GAGs) are the high affinity (83 nm) joint receptors for GPI. The binding affinity and structural differences between mouse paw/ankle GAGs and elbows/knee GAGs correlated with the distal to proximal disease severity in these joints. We found that cartilage surface GPI binding was greatly reduced by either chondroitinase ABC or beta-glucuronidase treatment. We also identified several inhibitors that inhibit both GPI/GAG interaction and GPI enzymatic activities, which suggests that the GPI GAG-binding domain overlaps with the active site of GPI enzyme. Our studies raise the possibility that GAGs are the receptors for other autoantigens involved in joint-specific inflammatory responses. 相似文献
133.
Shigeru Amano Keisuke Sekine Lynda F. Bonewald Yoshihiro Ohmori 《Journal of cellular physiology》2009,221(1):40-53
Osteoclasts are bone‐resorbing multinucleated cells differentiated from monocyte/macrophage lineage precursors. A novel osteoclast precursor cell line, 4B12 was established from Mac‐1+c‐Fms+RANK+ cells from calvaria of 14‐day‐old mouse embryos using immunofluorescence and cell‐sorting methods. Like M‐CSF‐dependent bone marrow macrophages (M‐BMMs), M‐CSF is required for 4B12 cells to differentiate into TRAP‐positive multinucleated cells [TRAP(+) MNCs] in the presence of RANKL. Bone‐resorbing osteoclasts differentiated from 4B12 cells on dentine slices possess both a clear zone and ruffled borders and express osteoclast‐specific genes. Bone‐resorbing activity, but not TRAP, was enhanced in the presence of IL‐1α. The number of TRAP(+) MNCs and the number of pits formed from 4B12 cells on dentine slices was fourfold higher than that from M‐BMMs. 4B12 cells were identified as macrophages with Mac‐1 and F4/80, yet lost these markers upon differentiation into osteoclasts as determined by confocal laser scanning microscopy. The 4B12 cells do not have the potential to differentiate into dendritic cells indicating commitment to the osteoclast lineage. 4B12 cells are readily transfectable with siRNA transfection before and after differentiation. These data show that 4B12 cells faithfully replicate the properties of primary cells and are a useful and powerful model for analyzing the molecular and cellular regulatory mechanisms of osteoclastogenesis and osteoclast function. J. Cell. Physiol. 221: 40–53, 2009. © 2009 Wiley‐Liss, Inc 相似文献
134.
135.
Judith K. Nzoughet John T. G. Hamilton Catherine H. Botting Alastair Douglas Lynda Devine John Nelson Christopher T. Elliott 《Molecular & cellular proteomics : MCP》2009,8(8):1811-1822
Azaspiracids are a class of recently discovered algae-derived shellfish toxins. Their distribution globally is on the increase with mussels being most widely implicated in azaspiracid-related food poisoning events. Evidence that these toxins were bound to proteins in contaminated mussels has been shown recently. In the present study characterization of these proteins in blue mussels, Mytilus edulis, was achieved using a range of advanced proteomics tools. Four proteins present only in the hepatopancreas of toxin-contaminated mussels sharing identity or homology with cathepsin D, superoxide dismutase, glutathione S-transferase Pi, and a bacterial flagellar protein have been characterized. Several of the proteins are known to be involved in self-defense mechanisms against xenobiotics or up-regulated in the presence of carcinogenic agents. These findings would suggest that azaspiracids should now be considered and evaluated as potential tumorigenic compounds. The presence of a bacterial protein only in contaminated mussels was an unexpected finding and requires further investigation. The proteins identified in this study should assist with development of urgently required processes for the rapid depuration of azaspiracid-contaminated shellfish. Moreover they may serve as early warning indicators of shellfish exposed to this family of toxins.Azaspiracids (AZAs)1 are a group of recently discovered algae-derived toxins following a shellfish poisoning event in 1995 in The Netherlands from consumption of Irish mussels (Mytilus edulis) (1). Initially the dinoflagellate Protoperidinium crassipes was proposed to be the organism producing AZAs (2); however, recent research has identified a new dinoflagellate, provisionally designated strain 3D9, as the source (3). Since the first AZA poisoning event in 1995 AZA incidents have been widely reported throughout Europe (4–6) and more recently in Morocco and eastern Canada (7, 8). AZA distribution thus appears to be on the increase and has become a public health concern and poses severe problems for the aquaculture industry. A regulatory limit of 160 μg of AZA/kg of shellfish in flesh has been proposed (9, 10) by the European Commission based on current information relating to the risks of consumption of contaminated shellfish.The most widely implicated species in AZA-associated food poisoning is mussels (7, 11). The blue mussel, M. edulis, has been widely used as a sentinel species for monitoring coastal environments and environmental pollution (12–14). Thus the recent appearance of AZAs could be considered as an indication of environmental changes that we do not as yet understand. A number of biochemical markers are known to be a good guide of the level of environmental stress to which living organisms have been subjected. It is also recognized that mussels produce proteins that can act as biomarkers to environmental contamination. Proliferating cell nuclear antigen and multixenobiotic resistance polyglycoprotein were revealed as biomarkers for genotoxic stress derived from benzo[a]pyrene in Baltic Sea blue mussels (15). Cu,Zn-superoxide dismutase (SOD), GSTs, and catalase are also well established biomarkers for the assessment of environmental stress in mussels following organic pollution and heavy metal exposure (16–21).Proteomics has proven to be a powerful technique for characterizing proteins expressed in specific tissues for many factors ranging from species differences to exposure to stress. For instance, López et al. (22) used proteomics to expand their understanding of the molecular differentiation between the mussels M. edulis and Mytilus galloprovincialis, whereas Apraiz et al. (23) identified the proteomic signatures in mussels exposed to marine pollutants.In the current study a range of advanced proteomics tools was used to further study the different protein profiles we recently observed between AZA-contaminated and non-contaminated mussels (24). Their identification and characterization may provide information toward identifying the mode of action of the toxins, which is currently unknown, and provide an indication as to why the AZA phenomenon has arisen so recently. If as recently suggested (24) prolonged AZA retention in shellfish is due to their association with proteins, then suitable processes could be developed to speed up the unusually low rates of depuration, which can take up to 8 months (25). A further important rationale for the work would be the identification of biomarkers that may serve as early warning indicators of AZA contamination in shellfish. 相似文献
136.
Martínez-Pardo ME Hernández L Reyes-Frías Mde L Luna-Zaragoza D 《Cell and tissue banking》2010,11(4):393-395
This short communication describes how some Mexican tissue banks have established their own system for coding and traceability
of tissues. 相似文献
137.
Kornberg MD Sen N Hara MR Juluri KR Nguyen JV Snowman AM Law L Hester LD Snyder SH 《Nature cell biology》2010,12(11):1094-1100
S-nitrosylation of proteins by nitric oxide is a major mode of signalling in cells. S-nitrosylation can mediate the regulation of a range of proteins, including prominent nuclear proteins, such as HDAC2 (ref. 2) and PARP1 (ref. 3). The high reactivity of the nitric oxide group with protein thiols, but the selective nature of nitrosylation within the cell, implies the existence of targeting mechanisms. Specificity of nitric oxide signalling is often achieved by the binding of nitric oxide synthase (NOS) to target proteins, either directly or through scaffolding proteins such as PSD-95 (ref. 5) and CAPON. As the three principal isoforms of NOS--neuronal NOS (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS)--are primarily non-nuclear, the mechanisms by which nuclear proteins are selectively nitrosylated have been elusive. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is physiologically nitrosylated at its Cys 150 residue. Nitrosylated GAPDH (SNO-GAPDH) binds to Siah1, which possesses a nuclear localization signal, and is transported to the nucleus. Here, we show that SNO-GAPDH physiologically transnitrosylates nuclear proteins, including the deacetylating enzyme sirtuin-1 (SIRT1), histone deacetylase-2 (HDAC2) and DNA-activated protein kinase (DNA-PK). Our findings reveal a novel mechanism for targeted nitrosylation of nuclear proteins and suggest that protein-protein transfer of nitric oxide groups may be a general mechanism in cellular signal transduction. 相似文献
138.
Sheila A. Dugan Susan A. Everson‐Rose Kelly Karavolos Elizabeth F. Avery Deidre E. Wesley Lynda H. Powell 《Obesity (Silver Spring, Md.)》2010,18(6):1260-1265
The purpose of our study was to determine whether self‐reported physical activity (PA), including recreational, household, and exercise activities, is associated with intra‐abdominal fat (IAF) in community‐dwelling white and black midlife women. We performed a cross‐sectional study of 369 women from the Chicago site of the Study of Women's Health Across the Nation (SWAN) ancillary study, the SWAN Fat Patterning Study. PA level was the independent variable, and IAF, assessed by computerized tomography (CT) scan, was the dependent variable. Measures were obtained at SWAN Fat Patterning Baseline visit between August 2002 and December 2005. Linear regression models explored the association between PA and IAF. The first model included IAF as the outcome and total score PA as the main predictor, adjusting for total percent fat mass, age, and ethnicity. The second model included education, parity, sex hormone–binding globulin (SHBG) level, and depressive symptoms, measured by Center for Epidemiological Studies‐Depression (CES‐D) scale. Each 1‐point higher total PA score was associated with a 4.0 cm2 lower amount of IAF (P = 0.004), independent of total percent fat mass, age, ethnicity, SHBG level, educational level, CES‐D, and parity. Associations did not differ between white and black women. This study demonstrates a significant negative association between PA and IAF independent of multiple covariates in midlife women. Our findings suggest that motivating white and black women to increase PA during midlife may lessen IAF, which may have a positive impact on subsequent development of diabetes and cardiovascular disease. 相似文献
139.
Talbot HK Rock MT Johnson C Tussey L Kavita U Shanker A Shaw AR Taylor DN 《PloS one》2010,5(12):e14442
Background
Currently controversy exists about the immunogenicity of seasonal trivalent influenza vaccine in certain populations, especially the elderly. STF2.4×M2e (VAX102) is a recombinant fusion protein that links four copies of the ectodomain of influenza virus matrix protein 2 (M2e) antigen to Salmonella typhimurium flagellin, a TLR5 ligand. The objectives of this study were to assess the feasibility of giving VAX102 and TIV in combination in an effort to achieve greater immunogenicity and to provide cross-protection.Methodology/Principal Findings
Eighty healthy subjects, 18-49 years old, were enrolled in May and June 2009 in a double-blind, randomized, controlled trial at two clinical sites. Subjects were randomized to receive either TIV + VAX102 or TIV + placebo. Both arms tolerated the vaccines. Pain at the injection site was more severe with TIV + VAX102. Two weeks after immunization the HAI responses to the H1 and H3 antigens of TIV were higher in those that received TIV + VAX102 than in TIV + placebo (309 vs 200 and 269 vs 185, respectively), although statistically non-significant. There was no difference in the HAI of the B antigen. In the TIV + VAX102 arm, the geometric mean M2e antibody concentration was 0.5 µg/ml and 73% seroconverted.Conclusions/Significance
The combination of TIV + VAX102 has the potential to increase the immune response to the influenza A components of TIV and to provide M2e immunity which may protect against influenza A strains not contained in seasonal TIV.Trial Registration
ClinicalTrials.gov NCT00921973相似文献140.
The water-soluble tetrazolium salt (WST-1) assay is frequently used to assess cell proliferation. However, our study showed that in normal and cancerous keratinocytes, this assay is more responsive to changes in oxygenation than to rates of cell growth. Stimulation of keratinocyte proliferation by low Ca2+ and suppression of proliferation by nocodazole resulted in modest changes in WST-1 readings, whereas gradually reducing the level of oxygen in the cellular environment from ambient (21%) to near anoxic (0.1%) revealed a very strong negative correlation between cell oxygenation and WST-1 reagent reduction. In contrast, the very similar MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] cell proliferation assay, which uses a different tetrazolium salt, showed no sensitivity to the level of oxygen. Unlike MTT, WST-1 reagent is reduced extracellularly through trans-plasma membrane transport (tPMET), thereby suggesting that tPMET is oxygen dependent. We propose that the WST-1 assay can be developed into a sensitive quantitative method to evaluate cell oxygenation in vitro and used to study the role of hypoxia and tPMET in homeostasis and disease (e.g., cancer). At the same time, WST-1 assay should be used cautiously to assess cell viability or proliferation because readings can be affected by certain extrinsic (low atmospheric oxygen or high density culture) or intrinsic (defects in oxygen-sensing pathways) factors. 相似文献