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Significant Cell Death (significant + cell_death)
Selected AbstractsMaternal hypoxia increases the activity of MMPs and decreases the expression of TIMPs in the brain of neonatal ratsDEVELOPMENTAL NEUROBIOLOGY, Issue 3 2010Wenni Tong Abstract A recent study has shown that increased activity of matrix metalloproteinases-2 and metalloproteinases-9 (MMP-2 and MMP-9) has detrimental effect on the brain after neonatal hypoxia. The present study determined the effect of maternal hypoxia on neuronal survivability and the activity of MMP-2 and MMP-9, as well as the expression of tissue inhibitors of metalloproteinase 1 and 2 (TIMP-1 and TIMP-2) in the brain of neonatal rats. Pregnant rats were exposed to 10.5% oxygen for 6 days from the gestation day 15 to day 21. Pups were sacrificed at day 0, 4, 7, 14, and 21 after birth. Body weight and brain weight of the pups were measured at each time point. The activity of MMP-2 and MMP-9 and the protein abundance of TIMP-1 and TIMP-2 were determined by zymography and Western blotting, respectively. The tissue distribution of MMPs was examined by immunofluorescence staining. The neuronal death was detected by Nissl staining. Maternal hypoxia caused significant decreases in body and brain size, increased activity of MMP-2 at day 0, and increased MMP-9 at day 0 and 4. The increased activity of the MMPs was accompanied by an overall tendency towards a reduced expression of TIMPs at all ages with the significance observed for TIMPs at day 0, 4, and 7. Immunofluorescence analysis showed an increased expression of MMP-2, MMP-9 in the hippocampus at day 0 and 4. Nissl staining revealed significant cell death in the hippocampus at day 0, 4, and 7. Functional tests showed worse neurobehavioral outcomes in the hypoxic animals. © 2009 Wiley Periodicals, Inc. Develop Neurobiol 2010 [source] Prostaglandin E2 production and viability of cells cultured in contact with freshly mixed endodontic materialsINTERNATIONAL ENDODONTIC JOURNAL, Issue 5 2006K. K. Melegari Abstract Aim, To determine whether commonly used endodontic sealers could either induce or increase the release of prostaglandin E2 (PGE2) when in contact with cell types found in the periapical tissues. Methodology, Freshly mixed samples of Roth 801 sealer, Sealapex® and ProRoot® mineral trioxide aggregate (MTA) were placed in contact with cultured macrophages and fibroblasts for 24 h. The supernatant from the cultures was assayed for PGE2 using enzyme-linked immunosorbent assay. Cell viability counts were made. As a positive control, similar cultures were also exposed to lipopolysaccharide and the supernatant analysed for PGE2. Data were compared by anova. Results, The three materials examined in these experiments did not stimulate increased PGE2 release from either of the cell lines. In control cultures, lipolysaccharide increased PGE2 release from macrophages but not from fibroblasts. Viability counts revealed that, whilst Roth 801 sealer caused some cell death in both fibroblasts and macrophages, Sealapex® led to cell death only in the macrophage cultures. ProRoot® MTA did not lead to statistically significant cell death in either culture. Conclusions, Under 24-h culture conditions, the three freshly mixed test materials did not increase directly either production or release of PGE2 from either macrophages or gingival fibroblasts. Roth 801 decreased cell viability counts for both fibroblasts and macrophages. Sealapex® decreases macrophage viability. ProRoot® MTA did not affect viability in either cell line. [source] The small heat shock protein Hsp27 protects cortical neurons against the toxic effects of ,-amyloid peptideJOURNAL OF NEUROSCIENCE RESEARCH, Issue 14 2009Michael King Abstract Neurofibrillary tangles and amyloid plaques are considered to be hallmarks of Alzheimer's disease (AD), and the toxic effects of amyloid-, peptide (A,) lead to activation of stress-related signaling and neuronal loss. The small heat shock protein Hsp27 is reported to be increased in AD brains and to accumulate in plaques, but whether this represents a potentially protective response to stress or is part of the disease process is not known. We hypothesized that increased expression of Hsp27 in neurons can promote neuronal survival and stabilize the cytoskeleton in the face of A, exposure. By using neonatal rat cortical neurons, we investigated the potential role of Hsp27 in neuronal cultures in the presence or absence of A,. We initially tested whether a heat stress (HS) would be sufficient to induce endogenous Hsp27 expression. HS not only did not result in neuronal Hsp27 up-regulation but made the cells more vulnerable to A, exposure. We then used cDNA transfection to overexpress EGFP-Hsp27 (or the empty vector) in cultures and then assessed neuronal survival and growth. Transfected neurons appeared healthy and had robust neuritic outgrowth. A, treatment induced significant cell death by 48,72 hr in nontransfected and empty-vector-expressing cultures. In contrast, cultures expressing Hsp27 did not display significant apoptosis. Our results show that Hsp27-expressing neurons were selectively protected against the deleterious effects of A, treatment; neuronal degeneration was prevented, and A,-induced alterations in mitochondrial size were attenuated. We also demonstrate that Hsp27 expression can enhance neurite growth in cortical neurons compared with control vector-transfected cells. Overall, our study provides new evidence that Hsp27 can provide a protective influence in primary cortical neurons in the face of toxic concentrations of amyloid. © 2009 Wiley-Liss, Inc. [source] EDTA enhances high-throughput two-dimensional bioprinting by inhibiting salt scaling and cell aggregation at the nozzle surfaceJOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE, Issue 4 2009Cheryl A. Parzel Abstract Tissue-engineering strategies may be employed in the development of in vitro breast tissue models for use in testing regimens of drug therapies and vaccines. The physical and chemical interactions that occur among cells and extracellular matrix components can also be elucidated with these models to gain an understanding of the progression of transformed epithelial cells into tumours and the ultimate metastases of tumour cells. The modified inkjet printer may be a useful tool for creating three-dimensional (3D) in vitro models, because it offers an inexpensive and high-throughput solution to microfabrication, and because the printer can be easily manipulated to produce varying tissue attributes. We hypothesized, however, that when ink is replaced with a biologically based fluid (i.e. a ,bio-ink'), specifically a serum-free cell culture medium, printer nozzle failure can result from salt scale build-up as fluid evaporates on the printhead surface. In this study, ethylene diamine tetra-acetic acid (EDTA) was used as a culture medium additive to prevent salt scaling and cell aggregation during the bioprinting process. The results showed that EDTA, at a concentration typically found in commercially available trypsin solutions (0.53 mM), prevented nozzle failure when a serum-free culture medium was printed from a nozzle at 1000 drops/s. Furthermore, increasing concentrations of EDTA appeared to mildly decrease aggregation of 4T07 cells. Cell viability studies were performed to demonstrate that addition of EDTA did not result in significant cell death. In conclusion, it is recommended that EDTA be incorporated into bio-ink solutions containing salts that could lead to nozzle failure. Copyright © 2009 John Wiley & Sons, Ltd. [source] Effect of intracellular lipid droplets on cytosolic Ca2+ and cell death during ischaemia,reperfusion injury in cardiomyocytesTHE JOURNAL OF PHYSIOLOGY, Issue 6 2009Ignasi Barba Lipid droplets (LD) consist of accumulations of triacylglycerols and have been proposed to be markers of ischaemic but viable tissue. Previous studies have described the presence of LD in myocardium surviving an acute coronary occlusion. We investigated whether LD may be protective against cell death secondary to ischaemia,reperfusion injury. The addition of oleate,bovine serum albumin complex to freshly isolated adult rat cardiomyocytes or to HL-1 cells resulted in the accumulation of intracellular LD detectable by fluorescence microscopy, flow cytometry and 1H-nuclear magnetic resonance spectroscopy. Simulated ischaemia,reperfusion of HL-1 cells (respiratory inhibition at pH 6.4 followed by 30 min of reperfusion) resulted in significant cell death (29.7 ± 2.6% of total lactate dehydrogenase release). However, cell death was significantly attenuated in cells containing LD (40% reduction in LDH release compared with control cells, P= 0.02). The magnitude of LD accumulation was inversely correlated (r2= 0.68, P= 0.0003) with cell death. The protection associated with intracellular LD was not a direct effect of the fatty acids used to induce their formation, because oleate added 30 min before ischaemia, during ischaemia or during reperfusion did not form LD and did not protect against cell death. Increasing the concentration of free oleate during reperfusion progressively decreased the protection afforded by LD. HL-1 cells labelled with fluo-4, a Ca2+ -sensitive fluorochrome, fluorescence within LD areas increased more throughout simulated ischaemia and reperfusion than in the cytosolic LD-free areas of the same cells. As a consequence, cells with LD showed less cytosolic Ca2+ overload than control cells. These results suggest that LD exert a protective effect during ischaemia,reperfusion by sequestering free fatty acids and Ca2+. [source] Hypoglycemia induced changes in cell death and cell proliferation in the organogenesis stage embryonic mouse heartBIRTH DEFECTS RESEARCH, Issue 3 2004Gautam S. Ghatnekar Abstract BACKGROUND Hypoglycemia is a side effect of diabetes therapy and causes abnormal heart development. Embryonic heart cells are largely resistant to teratogen-induced apoptosis. METHODS Hypoglycemia was tested for effects on cell death and cell proliferation in embryonic heart cells by exposing mouse embryos on embryonic day (E) 9.5 (plug = E0.5) to hypoglycemia (30,50 mg/dl glucose) in vivo or in vitro for 24 hr. Long-term effects of in vivo exposure on conceptus viability were evaluated at E18.5. Cell death was evaluated on E10.5 by: 1) two TUNEL assays in sectioned embryos to demonstrate DNA fragmentation; 2) confocal microscopy in whole embryos stained with Lysotracker; 3) flow cytometry in dispersed heart cells stained for TUNEL and myosin heavy chain (MHC) to quantify and characterize cell type susceptibility; and 4) immunohistochemistry (IHC) and Western analysis in sectioned embryos to evaluate potential involvement of caspase-3 active subunit and p53. Effects on cell proliferation were evaluated by IHC and Western analysis of proliferating cell nuclear antigen (PCNA). RESULTS In vivo hypoglycemic exposure on E9.5 reduced viability in conceptuses examined on E18.5. Hearts examined on E10.5 demonstrated increased TUNEL and Lysotracker staining. In hearts of embryos exposed to hypoglycemia, flow cytometry demonstrated increased TUNEL-positive cells and cells dual-labeled for TUNEL and MHC. Protein expression of caspase-3 active subunit and p53 was increased and PCNA was markedly reduced in hearts of embryos exposed to hypoglycemia. CONCLUSIONS Hypoglycemia reduces embryonic viability, induces significant cell death, and reduces cell proliferation in the E9.5 mouse heart, and these processes may involve active caspase-3 and p53. Birth Defects Research (Part A), 2004. © 2004 Wiley-Liss, Inc. [source] NV-128, a novel isoflavone derivative, induces caspase-independent cell death through the Akt/mammalian target of rapamycin pathwayCANCER, Issue 14 2009Ayesha B. Alvero MD Abstract BACKGROUND: Resistance to apoptosis is 1 of the key events that confer chemoresistance and is mediated by the overexpression of antiapoptotic proteins, which inhibit caspase activation. The objective of this study was to evaluate whether the activation of an alternative, caspase-independent cell death pathway could promote death in chemoresistant ovarian cancer cells. The authors report the characterization of NV-128 as an inducer of cell death through a caspase-independent pathway. METHODS: Primary cultures of epithelial ovarian cancer (EOC) cells were treated with increasing concentration of NV-128, and the concentration that caused 50% growth inhibition (GI50) was determined using a proprietary assay. Apoptotic proteins were characterized by Western blot analyses, assays that measured caspase activity, immunohistochemistry, and flow cytometry. Protein-protein interactions were determined using immunoprecipitation. In vivo activity was measured in a xenograft mice model. RESULTS: NV-128 was able to induce significant cell death in both paclitaxel-resistant and carboplatin-resistant EOC cells with a GI50 between 1 ,g/mL and 5 ,g/mL. Cell death was characterized by chromatin condensation but was caspase-independent. The activated pathway involved the down-regulation of phosphorylated AKT, phosphorylated mammalian target of rapamycin (mTOR), and phosphorylated ribosomal p70 S6 kinase, and the mitochondrial translocation of beclin-1 followed by nuclear translocation of endonuclease G. CONCLUSIONS: The authors characterized a novel compound, NV-128, which inhibits mTOR and promotes caspase-independent cell death. The current results indicated that inhibition of mTOR may represent a relevant pathway for the induction of cell death in cells resistant to the classic caspase-dependent apoptosis. These findings demonstrate the possibility of using therapeutic drugs, such as NV-128, which may have beneficial effects in patients with chemoresistant ovarian cancer. Cancer 2009. © 2009 American Cancer Society. [source] | |||||||||||||