Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We recently demonstrated that Giardia lamblia rearranges cytoskeletal proteins and reduces transepithelial electrical resistance. The effect of G. lamblia on enterocyte apoptosis is unknown, and a possible link between microbially induced enterocyte apoptosis and altered epithelial permeability has yet to be established. The aim of this study was to assess whether G. lamblia induces enterocyte apoptosis in duodenal epithelial monolayers and whether this effect increases epithelial permeability. Monolayers of nontransformed human duodenal epithelial cells were incubated with sonicated or live G. lamblia trophozoites (NF, S2, WB, or PB strains) for 8, 24, and 48 h. Cell cultures were assessed for apoptosis by Hoechst fluorescence staining, enzyme-linked immunosorbent assay for apoptotic nucleosomes, and electron microscopy. In separate experiments, monolayers were pretreated with or without 120 microM caspase-3 inhibitor (Z-DEVD-FMK) for 1 h and were assessed for production of apoptotic nucleosomes, tight junctional integrity (with fluorescent ZO-1 staining followed by confocal laser microscopy), and transepithelial permeability for fluorescein isothiocyanate-dextran. G. lamblia strains NF and S2, but not strains WB or PB, induced enterocyte apoptosis within the monolayers, and this effect was inhibited by Z-DEVD-FMK pretreatment. Using the G. lamblia NF isolate, additional experiments investigated the possible link between enterocyte apoptosis and altered epithelial permeability. G. lamblia NF disrupted tight junctional ZO-1 and increased epithelial permeability, but these effects were also prevented by pretreatment with the caspase-3 inhibitor. These findings indicate that strain-dependent induction of enterocyte apoptosis may contribute to the pathogenesis of giardiasis. This effect is responsible for a loss of epithelial barrier function by disrupting tight junctional ZO-1 and increasing permeability in a caspase-3-dependent manner.
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PMID:Strain-dependent induction of enterocyte apoptosis by Giardia lamblia disrupts epithelial barrier function in a caspase-3-dependent manner. 1206 9

Cell polarity regulates diverse biological events such as localization of embryonic determinants and establishment of tissue and organ architecture. Epithelial cell polarity is regulated by the polarity complex Par6/Par3/atypical protein kinase C (aPKC). We previously found that the nucleotide exchange factor ECT2 associates with this polarity complex and regulates aPKC activity, but the role of ECT2 in cell polarity is still unclear. Here we show that expression of a dominant negative (ECT2-N2) or constitutively active (ECT2-DeltaN5) form of ECT2 inhibits normal cyst formation of MDCK cells in 3-dimensional collagen gels. Central lumens were not observed in cysts formed by cells expressing either ECT2-DeltaN5 or ECT2-N2. Apical localization of ZO-1 and basolateral localization of beta-catenin were no longer observed in these cells. Interestingly, cells expressing ECT2-N2 did form normal cysts when cultured in the basement membrane matrix Matrigel instead of collagen gels. Addition of a major Matrigel component, laminin, partially rescued the normal cyst formation inhibited by ECT2-N2 in 3-dimensional collagen gels. Thus, signaling through laminin might override the defects of signaling through collagen and ECT2. Whereas ECT2-N2 inhibited the lumen formation of MDCK cysts, caspase-3, which is reportedly involved in lumen formation through apoptosis, was activated at various locations of cells in the cysts. It is likely that perturbation of ECT2 signaling inhibits the establishment of epithelial cell polarity leading to the inhibition of selected elimination of cells at the center of cysts. Thus, ECT2 appears to play a critical role in epithelial cell polarity.
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PMID:Nucleotide exchange factor ECT2 regulates epithelial cell polarity. 1649 35

Spermatogenesis, a tightly regulated developmental process of male germ cells in testis, is associated with temporal and spatial expression of gap junction proteins, such as the connexin family members. Perturbation of their expressions may lead to spermatogenic arrest as manifested by disruption of cell-cell interaction. To explore the role(s) of connexins during spermatogenesis, we utilized the small peptide antagonistic approach to specifically deplete connexin 31, connexin 33, and pan-connexin. Three connexin peptides corresponding to the extracellular binding domain of connexin 31 and connexin 33 and to the extracellular conserved domain of connexins were designed and synthesized commercially. Peptides (at single dosage of 0.5, 1, or 2 mg per animal) were injected into rat testes and testes were collected on day 0, 1, 3, 5, 10, 15, and 30 after microinjection. In situ TUNEL assay demonstrated the induction of apoptosis in the testes after pan-connexin peptide treatment in a dose-dependent manner from day 3 and onward. Unlike the pan-connexin peptide, connexin 31 and connexin 33 peptides appeared to have little effect on inducing apoptosis and germ cell loss. CD45 staining also detected the occasional presence of infiltrating lymphocytes in the seminiferous tubules. Accompanied with the apoptotic events, two apoptotic markers, NF-kappaB and caspase 3, demonstrated a general up-regulation in their expressions. In adjacent testis sections, eliminations of connexin 31, 32, and 43 were observed. However, an induction of connexin 33 expression was detected. This suggests the versatility and functional diversity of connexins in the testis. The expression of ZO-1, the only known adaptor of connexins in the testis, was reduced and remained in a low level in the seminiferous epithelium. As such, the alterations of connexins in seminiferous epithelium may induce apoptotic signaling in the testis via the caspase 3 and the NF-kappaB pathway. This demonstrates the significant role of testicular connexins to maintain the survival of germ cells by regulating inter-cellular communications among germ cells and adjacent supporting cells during spermatogenesis. In addition, the inter-relationship between connexins and other junction proteins and associated signaling protein were investigated. After pan-connexin peptide treatment, a dys-localization of N-cadherin, an adherens junction protein, and diminution of occludin, a tight junction protein, level were detected. In addition, inductions of junction regulatory protein, cathepsin L, was observed during the course of peptide-mediated germ cell loss in the testes. In summary, pan-connexin peptide treatment triggered apoptosis and germ cell loss in the testes. This event influenced the localization and expression of different junction proteins and junction-associated protein in the testes.
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PMID:Blockage of testicular connexins induced apoptosis in rat seminiferous epithelium. 1669 59

The mechanisms responsible for microbially induced epithelial apoptosis and increased intestinal permeability remain unclear. This study assessed whether purified bacterial lipopolysaccharide (LPS) increases epithelial apoptosis and permeability and whether these changes are dependent on caspase-3 activation. In nontumorigenic epithelial monolayers, Escherichia coli O26:B6 LPS increased apoptosis, as shown by nuclear breakdown, caspase-3 activation, and PARP cleavage, and induced disruption of tight junctional ZO-1. Apical, but not basolateral, exposure to LPS increased epithelial permeability. Addition of a caspase-3 inhibitor abolished the effects of LPS. The findings describe a novel mechanism whereby apical LPS may disrupt epithelial tight junctional ZO-1 and barrier function in a caspase-3-dependent fashion.
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PMID:The role of caspase-3 in lipopolysaccharide-mediated disruption of intestinal epithelial tight junctions. 1721 70

This study was performed to examine the maintenance of blood vessels in vitro in cortical organotypic slice cultures of mice with special emphasis on basic fibroblast growth factor (FGF-2), which is known to promote angiogenesis and to preserve the integrity of the blood-brain barrier. Slices of neonatal day 3 or 4 mouse brain were maintained for 3, 7, or 10 d in vitro (DIV) under standard culture conditions or in the presence of FGF-2. Immunohistochemistry for factor VIII-related antigen or laminin revealed a relative low number of blood vessels under standard conditions. In contrast, moderate FGF-2 concentrations increased the number of vessels: with 0.5 ng/ml FGF-2 it was 1.4-fold higher after DIV 3 or 1.5-fold after DIV 7 compared with controls; with 5 ng/ml it was almost doubled in both cases. With an excess of 50 ng/ml, FGF-2 vessels were reduced after DIV 3 or similar to controls after DIV 7. FGF receptor 1 was preferentially found on endothelial cells; its immunolabeling was reduced in the presence of the ligand. Cell death detected by an ethidium bromide analog or the apoptosis marker caspase-3 was barely detectable during the 10 d culture period. Immunolabeling of the tight junction proteins ZO-1 (zonula occludens protein 1), occludin, claudin-5, and claudin-3 revealed evidence for structural integrity of the blood-brain barrier in the presence of moderate FGF-2 concentrations. In conclusion, FGF-2 maintains blood vessels in vitro and preserves the composition of the tight junction. Hence, we propose FGF-2-treated organotypic cortical slices as a new tool for mechanistic studies of the blood-brain barrier.
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PMID:Basic fibroblast growth factor modulates density of blood vessels and preserves tight junctions in organotypic cortical cultures of mice: a new in vitro model of the blood-brain barrier. 1737 86

Dexras1, a brain-enriched member of the Ras subfamily of GTPases, as a novel physiologic nitric oxide (NO) effector, anchor neuronal nitric oxide synthase (nNOS) that increased after spinal cord injury (SCI), to specific targets to enhance NO signaling, and is strongly and rapidly induced during treatment with dexamethasone. It is unknown how the central nervous system (CNS) trauma affects the expression of Dexras1. Here we used spinal cord transection (SCT) model to detect expression of Dexras1 at mRNA and protein level in spinal cord homogenates by real-time PCR and Western blot analysis. The results showed that Dexras1 mRNA upregulated at 3 day, 5 day, and 7 day significantly (P < 0.05) that was consistent with the protein level except at 7 day. Immunofluorescence revealed that both neurons and glial cells showed Dexras1 immunoreactivivty (IR) around SCT site, but the proportion is different. Importantly, injury-induced expression of Dexras1 was co-labeled by caspase-3 (apoptotic marker) and Tau-1 (marker for pathological oligodendrocyte). Furthermore, colocalization of Dexras1, carboxy-terminal PSD95/DLG/ZO-1 (PDZ) ligand of nNOS (CAPON) and nNOS was observed in neurons and glial cells, supporting the existence of ternary complexes in this model. Thus, the results that the transient high expression of Dexras1 which localized in apoptotic neurons and pathological oligodendrocytes might provide new insight into the secondary response after SCT.
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PMID:Spatiotemporal expression of Dexras1 after spinal cord transection in rats. 1821 71

Subarachnoid hemorrhage (SAH) causes secondary brain injury due to vasospasm and inflammation. Here, we studied a rat model of mild-to-moderate SAH intended to minimize ischemia/hypoxia to examine the role of sulfonylurea receptor 1 (SUR1) in the inflammatory response induced by SAH. mRNA for Abcc8, which encodes SUR1, and SUR1 protein were abundantly upregulated in cortex adjacent to SAH, where tumor-necrosis factor-alpha (TNFalpha) and nuclear factor (NF)kappaB signaling were prominent. In vitro experiments confirmed that Abcc8 transcription is stimulated by TNFalpha. To investigate the functional consequences of SUR1 expression after SAH, we studied the effect of the potent, selective SUR1 inhibitor, glibenclamide. We examined barrier permeability (immunoglobulin G, IgG extravasation), and its correlate, the localization of the tight junction protein, zona occludens 1 (ZO-1). SAH caused a large increase in barrier permeability and disrupted the normal junctional localization of ZO-1, with glibenclamide significantly reducing both effects. In addition, SAH caused large increases in markers of inflammation, including TNFalpha and NFkappaB, and markers of cell injury or cell death, including IgG endocytosis and caspase-3 activation, with glibenclamide significantly reducing these effects. We conclude that block of SUR1 by glibenclamide may ameliorate several pathologic effects associated with inflammation that lead to cortical dysfunction after SAH.
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PMID:Glibenclamide reduces inflammation, vasogenic edema, and caspase-3 activation after subarachnoid hemorrhage. 1885 40

Differentiation and transformation of untransformed and ts-Src-transformed canine kidney MDCK cells in 2D and 3D environment were investigated using microarray technique, RT-PCR, confocal microscopy and functional assays. Activated Src induced epithelial-mesenchymal transition (EMT) in 2D environment followed by translocation of junctional proteins to the cytoplasm, without significant changes in protein expression. In 3D environment untransformed MDCK cells formed cell cysts with apical domain facing a lumen, E-cadherin delineating the lateral membranes, ZO-1 at tight junctions and caspase-3 in apoptotic cells captured within the lumen. This was accompanied by reduced expression of an apoptosis inhibitor, survivin and vesicle transport effectors, rab 7 and 8, whereas rab 5 expression increased. In 3D environment activated Src induced changes in expression of over 100 genes as revealed by microarray analysis, mostly involved in cell signaling, division and energy metabolism. Only response in cytoskeletal components was decreased expression of actin and Arp2/3 by v-Src, whereas two p120catenin binding proteins Kaiso and Nanos increased their expression. Concomitantly, apoptosis was inhibited by v-Src resulting in formation of a sphere with epitheloid cells facing extracellular matrix and undifferentiated cells captured within the cluster. This was accompanied by increased expression of apoptosis inhibitor survivin, as revealed by western blotting. Mitochondrial membrane potential in untransformed MDCK cells was lower than in ts-Src-MDCK cells in early days of cluster formation correlating with the induction of apoptosis. Hence, v-Src activation in 3D environment did not induce EMT, but brought about inhibition of apoptosis and increased proliferation where increased expression of survivin and inhibition of the mitochondrial permeability have a role.
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PMID:Different responses in transformation of MDCK cells in 2D and 3D culture by v-Src as revealed by microarray techniques, RT-PCR and functional assays. 2021 54

Although enteric glial cells (EGCs) have been demonstrated to play a key role in maintaining intestinal epithelial barrier integrity, it is not known how EGCs regulate this integrity. We therefore hypothesized that glial-derived neurotrophic factor (GDNF) produced by EGCs might be involved in this regulation. Here we investigated the role of GDNF in regulating epithelial barrier function in vivo. Recombinant adenoviral vectors encoding GDNF (Ad-GDNF) were administered intracolonically in experimental colitis induced by dextran sulphate sodium (DSS). The disease activity index (DAI) and histological score were measured. Epithelial permeability was assayed using Evans blue dye. The anti-apoptotic potency of GDNF in vivo was evaluated. The expression of tumour necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and myeloperoxidase (MPO) activity were measured by ELISA assay and/or RT-PCR. The expression of ZO-1, Akt, caspase-3, and NF-kappaB p65 was analysed by western blot assay. Our results showed that GDNF resulted in a significant reduction in enhanced permeability, inhibited MPO activity, IL-1beta and TNF-alpha expression, and increased ZO-1 and Akt expression. Moreover, GDNF strongly prevented apoptosis in vivo and significantly ameliorated experimental colitis. Our findings indicate that GDNF participates directly in restoring epithelial barrier function in vivo via reduction of increased epithelial permeability and inhibition of mucosal inflammatory response, and is efficacious in DSS-induced colitis. These findings support the notion that EGCs are able to regulate intestinal epithelial barrier integrity indirectly via their release of GDNF in vivo. GDNF is namely an important mediator of the cross-talk between EGCs and mucosal epithelial cells. GDNF may be a useful therapeutic approach to the treatment of inflammatory bowel disease.
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PMID:Glial-derived neurotrophic factor regulates intestinal epithelial barrier function and inflammation and is therapeutic for murine colitis. 2063 86

The Fusarium derived mycotoxin deoxynivalenol (DON) is frequently found in cereals used for human and animal nutrition. We studied effects of DON in non-transformed, non-carcinoma, polarized epithelial cells of porcine small intestinal origin (IPEC-1 and IPEC-J2) in a low (200 ng/mL) and a high (2000 ng/mL) concentration. Application of high DON concentrations showed significant toxic effects as indicated by a reduction in cell number, in cellular reduction capacity measured by MTT assay, reduced uptake of neutral red (NR) and a decrease in cell proliferation. High dose toxicity was accompanied by disintegration of tight junction protein ZO-1 and increase of cell cycle phase G2/M. Activation of caspase 3 was found as an early event in the high DON concentration with an initial maximum after 6-8 h. In contrast, application of 200 ng/mL DON exhibited a response pattern distinct from the high dose DON toxicity. The cell cycle, ZO-1 expression and distribution as well as caspase 3 activation were not changed. BrdU incorporation was significantly increased after 72 h incubation with 200 ng/mL DON and NR uptake was only transiently reduced after 24 h. Low dose effects of DON on intestinal epithelial cells were triggered by mechanisms different from those responsible for the high dose toxicity.
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PMID:Mycotoxin deoxynivalenol (DON) mediates biphasic cellular response in intestinal porcine epithelial cell lines IPEC-1 and IPEC-J2. 2093 67


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