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Extrinsic Pathway (extrinsic + pathway)
Selected AbstractsInvolvement of mitochondrial signaling pathways in the mechanism of Fas-mediated apoptosis after spinal cord injuryEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2009Wen Ru Yu Abstract Activation of the Fas receptor has been recently linked to apoptotic cell death after spinal cord injury (SCI). Although it is generally considered that Fas activation mediates apoptosis predominantly through the extrinsic pathway, we hypothesized that intrinsic mitochondrial signaling could be involved in the underlying mechanism of Fas-induced apoptosis after SCI. In the present study, we utilized the FejotaTM clip compression model of SCI at T5,6 in C57BL/6 Fas-deficient (lpr) and wild-type mice. Complementary studies were conducted using an in vitro model of trauma or a Fas-activating antibody to induce apoptosis in primary neuronal,glial mixed spinal cord cultures. After in vivo SCI, lpr mice, in comparison with wild-type mice, exhibited reduced numbers of terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells at the lesion, reduced expression of truncation of Bid (tBid), apoptosis-inducing factor, activated caspase-9 and activated caspase-3, and increased expression of the antiapoptotic proteins Bcl-2 and Bcl-xL. After in vitro neurotrauma or the induction of Fas signaling by the Jo2 activating antibody, lpr spinal cord cultures showed an increased proportion of cells retaining mitochondrial membrane integrity and a reduction of tBid expression, caspase-9 and caspase-3 activation, and TUNEL-positive cells as compared to wild-type spinal cord cultures. The neutralization of Fas ligand (FasL) protected against traumatically induced or Fas-mediated caspase-3 activation and the loss of mitochondrial membrane potential and tBid expression in wild-type spinal cord cultures. However, in lpr spinal cord cultures, FasL neutralization had no protective effects. In summary, these data provide direct evidence for the induction of intrinsic mitochondrial signaling pathways following Fas activation after SCI. [source] Gene expression demonstrates increased resilience toward harmful inflammatory stimuli in the proliferating epidermis of human skin woundsEXPERIMENTAL DERMATOLOGY, Issue 8 2010K. Markus Roupé Please cite this paper as: Gene expression demonstrates increased resilience toward harmful inflammatory stimuli in the proliferating epidermis of human skin wounds. Experimental Dermatology 2010; 19: e329,e332. Abstract:, We examined the epidermal gene expression during the proliferative phase of wound healing. Matrix metalloproteases were the group of proteases most prominently up-regulated in skin wounds, whereas serine protease inhibitors were the most strongly up-regulated protease inhibitors. Furthermore, we found down-regulation of genes involved in the extrinsic pathway of apoptosis. This together with the up-regulation of inhibitors of leukocyte serine proteases likely represents a protective step to ensure survival of keratinocytes in the inflammatory wound environment. The down-regulation of proapoptotic genes in the extrinsic pathway of apoptosis was not accompanied by a down-regulation of receptors indicating that the keratinocytes in skin wounds did not become less responsive to external stimuli. Examining the transcription factor binding sites in the promoters of the most differentially expressed genes between normal skin and skin wounds a significant overrepresentation of binding sites were found for STAT-5, SRY and members of the FOXO-family of transcription factors. [source] The initiator caspase, caspase-10,, and the BH-3-only molecule, Bid, demonstrate evolutionary conservation in Xenopus of their pro-apoptotic activities in the extrinsic and intrinsic pathwaysGENES TO CELLS, Issue 7 2006Katsuya Kominami Two major apoptotic signaling pathways have been defined in mammals, the extrinsic pathway, initiated by ligation of death receptors, and the intrinsic pathway, triggered by cytochrome c release from mitochondria. Here, we identified and characterized the Xenopus homologs of caspase-10 (xCaspase-10,), a novel initiator caspase, and Bid (xBid), a BH3-only molecule of the Bcl-2 family involved in both the extrinsic and intrinsic pathways. Exogenous expression of these molecules induced apoptosis of mammalian cells. By biochemical and cytological analyses, we clarified that xCaspase-10, and xBid exhibit structural and functional similarities to their mammalian orthologues. We also detected xCaspase-10, and xBid transcripts during embryogenesis by whole-mount in situ hybridization and RT-PCR analysis. Microinjection of mRNA encoding a protease-defect xCaspase-10, mutant into embryos resulted in irregular development. Enforced expression of active xBid induced cell death in developing embryos. Using transgenic frogs established to allow monitoring of caspase activation in vivo, we confirmed that this form of cell death is caspase-dependent apoptosis. Thus, we demonstrated that the machinery governing the extrinsic and intrinsic apoptotic pathways are already established in Xenopus embryos. Additionally, we propose that the functions of the initiator caspase and BH3-only molecule are evolutionarily conserved in vertebrates, functioning during embryonic development. [source] Involvement of the contact phase and intrinsic pathway in herpes simplex virus-initiated plasma coagulationJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2010E. S. GERSHOM Summary.,Background:,A hemostatic response to vascular injury is initiated by the extrinsic pathway of coagulation and amplified by the intrinsic pathway. We previously reported that purified herpes simplex virus type-1 (HSV1) has constitutive extrinsic pathway tissue factor (TF) and anionic phospholipid on its surface derived from the host cell, and can consequently bypass strict cellular control of coagulation. Objective:,The current work addresses the hypothesis that HSV1-induced plasma coagulation also involves intrinsic pathway, factor VIII (FVIII), and upstream contact activation pathway, factor XII (FXII). Results:,HSV1-initiated clotting was accelerated when purified FVIII was added to FVIII-deficient plasma and in normal plasma attenuated by an inhibitory anti-FVIII antibody (Ab). High HSV1 concentrations predictably reduced the effect of FVIII due to the availability of excess viral TF. To further define TF-independent clotting mechanisms initiated by HSV1, the extrinsic pathway was disabled using factor VII-deficient plasma. The intrinsic pathway is triggered by activation of FXII associated with surface-bound kallikrein, which subsequently activates factor XI. Here we found that an inhibitor of activated FXII, corn trypsin inhibitor, and anti-FXII, anti-kallikrein and anti-FXI Abs inhibited HSV1-initiated clotting. HSV1-enhanced activation of purified FXII was confirmed by Western blot, but required prekallikrein. Conclusion:,The current work shows that HSV1 can trigger and amplify coagulation through the contact phase and intrinsic pathway, and suggests an additional mechanism that may contribute to vascular pathology. [source] Expression of human tissue factor under the control of the mouse tissue factor promoter mediates normal hemostasis in knock-in miceJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 2 2008L. A. SNYDER Summary.,Background:,Tissue factor (TF) is expressed widely at the subluminal surface of blood vessels and serves as the primary cellular initiator of the extrinsic pathway of blood coagulation. Lack of TF in mice resulted in lethality in utero, but human TF (huTF) expressed at low levels from a human minigene rescued null mice from prenatal death. Although these low-TF expressing transgenic mice developed to term, they had a significantly shorter life span and exhibited hemorrhage and fibrosis in the heart. Methods:,Human TF knock-in (TFKI) mice were generated by replacing the first two exons of the mouse (murine) TF (muTF) gene with the huTF complete coding sequence, thus placing it under the control of the endogenous muTF promoter. Results:,Expression of huTF in the TFKI mice was similar to muTF in wild-type (wt) mice. The TFKI mice showed no microscopic evidence of spontaneous hemorrhage in the heart, nor cardiac fibrosis at up to 18 months of age. Immunohistochemistry showed that huTF was expressed in cells surrounding blood vessels in TFKI mice. Coagulation activity of brain homogenates from TFKI mice was comparable with that from wt brain. Cardiac hemorrhage similar to that of the low-TF transgenic mice occurred in the TFKI mice when huTF was blocked by a neutralizing anti-huTF monoclonal antibody. Conclusion:,We generated a transgenic mouse line that expresses huTF under the control of the endogenous muTF promoter at physiological levels. Our results suggest that huTF can fully reconstitute the murine coagulation system and mediate normal hemostasis. [source] Adverse effects associated with persistent stimulation of Leydig cells with hCG in vitroMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 11 2009Archana Aggarwal The detrimental effects of persistent stimulation with hCG were investigated in rat Leydig cells in vitro. Significant rise in lipid peroxidation and reactive oxygen species (ROS) with concomitant attenuation in the activities of antioxidant enzymes: superoxide dismutase, catalase, and glutathione- S -transferase was observed. Transcripts for catalase and superoxide dismutase were also depleted. Subsequent to each hCG challenge, the total antioxidant capacity in the target cells also declined significantly (P,<,0.05). There was an increase in cell apoptosis (23%), which was associated with a rise in caspase-3 activity, PARP cleavage, and Fas, FasL, caspase-8 expression. While Bax and Caspase-9 expression remained unchanged, Bcl-2 demonstrated a marked decline. Taken together, the above data indicate that persistent hCG stimulation of Leydig cells induced adverse effects leading to oxidative stress and apoptosis which was channeled primarily through the extrinsic pathway. Mol. Reprod. Dev. 76: 1076,1083, 2009. © 2009 Wiley-Liss, Inc. [source] Relevance of caspase activity during apoptosis in pubertal rat spermatogenesisMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 5 2008Veronica A. Codelia Abstract Caspases are a family of cysteine-proteases, activated upon several different stimuli, which execute apoptosis in many cell death models. Previous work of our group has shown rats have the highest rate of apoptosis during the first wave of spermatogenesis (between 20 and 25 days after birth), as evaluated by TUNEL and caspase activity. However, the hierarchical order of caspase activation and the relevance of each caspase during germ cell apoptosis are not clear. Thus, the goal of this work is to take a pharmacological approach to dissect the apoptosis pathway of caspase activation. Results showed that intratesticular injection of a caspase-8 inhibitor (z-IETD-fmk), or a pan-caspase inhibitor (z-VAD- fmk), significantly decreased the cleavage of p115 and PARP, two endogenous substrates of caspases, in 22-day-old rats. Additionally, these inhibitors promoted a significant reduction in the number of apoptotic germ cells. On the other hand, intratesticular injection of two different inhibitors of the intrinsic pathway (z-LEHD-fmk and minocycline) did not have any effect upon caspase substrates cleavage (p115 and PARP) or the number of apoptotic germ cells. Therefore, we conclude that the extrinsic pathway of apoptosis plays an important role in physiological germ cell apoptosis during the first round of spermatogenesis in the rat. Mol. Reprod. Dev. 75: 881,889, 2008. © 2007 Wiley-Liss, Inc. [source] The roles of apoptotic pathways in the low recovery rate after cryopreservation of dissociated human embryonic stem cellsBIOTECHNOLOGY PROGRESS, Issue 3 2010Xia Xu Abstract Human embryonic stem (hES) cells have enormous potential for clinical applications. However, one major challenge is to achieve high cell recovery rate after cryopreservation. Understanding how the conventional cryopreservation protocol fails to protect the cells is a prerequisite for developing efficient and successful cryopreservation methods for hES cell lines and banks. We investigated how the stimuli from cryopreservation result in apoptosis, which causes the low cell recovery rate after cryopreservation. The level of reactive oxygen species (ROS) is significantly increased, F-actin content and distribution is altered, and caspase-8 and caspase-9 are activated after cryopreservation. p53 is also activated and translocated into nucleus. During cryopreservation apoptosis is induced by activation of both caspase-8 through the extrinsic pathway and caspase-9 through the intrinsic pathway. However, exactly how the extrinsic pathway is activated is still unclear and deserves further investigation. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [source] Defining the caspase-containing apoptotic machinery contributing to cornification in human epidermal equivalentsEXPERIMENTAL DERMATOLOGY, Issue 1 2006Vijaya Chaturvedi Abstract:, Whether terminal differentiation/stratum corneum formation of keratinocytes (KCs) represents a form of programmed cell death, utilizing mediators of classical apoptosis, is unclear. Apoptosis, an evolutionarily conserved death process, is comprised of extrinsic and intrinsic pathways, which converge using caspase 3. To define upstream and downstream caspases involved in terminal differentiation, we utilized human epidermal equivalents (EEs). Using submerged cultures comprised of human KCs, EEs were sequentially analyzed before and after being raised to an air/liquid (A/L) interface at 3,24 h intervals. At each time point, EEs were analyzed morphologically and for specific enzyme activity to distinguish different initiator (caspases 1, 2, 8, 9) and effector caspases (3, 6, 7). Terminal differentiation began at 6,8 h, as defined by stratum corneum with loricirin expression and completed at 18,24 h producing an epidermis resembling normal skin. Enzyme activity for caspases 1, 2, 3, 6, 7, 8, and 9 (but not 4, 5) was enhanced (>two-fold nmol/mg/h) at 3,6 h compared with submerged cultures. Processing of caspase 14 occurred at 18 h, and cleaved caspase 14 was increased at 24 h. Activated caspase 3-positive and terminal deoxynucleotidyl transferase-mediated nick end labeling-positive KCs were identified in EEs at 3,6 h corresponding to initiation sites of terminal differentiation. Addition of caspase inhibitors reduced levels of involucrin and loricrin in EEs raised to an A/L interface. We conclude caspases function as important death effectors strategically positioned at intersection of intrinsic and extrinsic pathways in KCs undergoing stratum corneum formation. [source] Tissue factor and tissue factor pathway inhibitorANAESTHESIA, Issue 5 2004G. C. Price Summary The classical ,cascade/waterfall' hypothesis formulated to explain in vitro coagulation organised the amplification processes into the intrinsic and extrinsic pathways. Recent molecular biology and clinical data indicate that tissue factor/factor-VII interaction is the primary cellular initiator of coagulation in vivo. The process of blood coagulation is divided into an initiation phase followed by a propagation phase. The discovery of tissue factor pathway inhibitor further supports the revised theory of coagulation. Tissue factor is also a signalling receptor. Recent evidence has shown that blood-borne tissue factor has an important procoagulant function in sepsis, atherosclerosis and cancer, and other functions beyond haemostasis such as immune function and metastases. [source] Deregulation of cell-death pathways as the cornerstone of skin diseasesCLINICAL & EXPERIMENTAL DERMATOLOGY, Issue 6 2010N. Zutterman Summary Deregulation of cell-death pathways plays a key role in the pathogenesis of various skin diseases. The different types of cell death are mainly defined by morphological criteria, and include apoptosis, autophagic cell death, and necrosis. The process of apoptosis is well characterized at the molecular level and involves the activation of two main pathways, the intrinsic and extrinsic pathways, converging into the execution of apoptosis by intracellular cysteine proteases, called caspases. The relevance and implication of these apoptotic pathways in the pathophysiology of skin diseases, such as toxic epidermal necrolysis, graft-versus-host disease and skin cancer, has been extensively studied. The role of autophagic cell death in progression of skin tumours and response to cytotoxic drugs is only beginning to be elucidated. [source] | |||||||||||||