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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labelling (TUNEL) technique has been extensively used for the detection and quantification of apoptosis in histological tissue sections. However, the interpretation and specificity of this assay have been controversial. With accumulating knowledge of the molecular mechanisms of cell death and the discovery of the caspases as key mediators of apoptosis, more direct and earlier measurements of apoptosis in tissue sections have emerged. This study, using antibodies that specifically recognize activated caspase-3 and caspase-cleaved cytokeratin (CK) 18, evaluated whether immunohistochemical stains would improve the detection and quantification of apoptosis in tissue sections, compared with the TUNEL assay. Tumour xenografts of the prostate cancer cell line PC-3 were used as an example, since these tissues contain large numbers of cells undergoing apoptosis. Apoptotic cells were quantified and apoptotic indices were calculated by computer-assisted image analysis following identification of apoptotic cells by morphological analysis, the TUNEL assay, activated caspase-3 and cleaved CK18 immunohistochemistry. The results indicated that activated caspase-3 immunohistochemistry was an easy, sensitive, and reliable method for detecting and quantifying apoptosis in this model. An excellent correlation (R = 0.89) between the apoptotic indices obtained using activated caspase-3 and cleaved CK18 immunostaining was observed. A good correlation (R = 0.75) between the apoptotic indices obtained using activated caspase-3 immunostaining and the TUNEL assay was also found. Activated caspase-3 immunohistochemistry is therefore recommended for the detection and quantification of apoptosis in tissue sections.
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PMID:Comparison of immunohistochemistry for activated caspase-3 and cleaved cytokeratin 18 with the TUNEL method for quantification of apoptosis in histological sections of PC-3 subcutaneous xenografts. 1253 35

It has become clear that apoptosis is an essential part of normal folliculogenesis and that granulosa cells in particular demonstrate intense cellular activity as well as programmed cell death. Although the entire mechanism of apoptosis appears to be conserved through many species, it now becomes clear that different cells may use different pathways within this system. We examined human granulosa cells after ovulation induction during an in vitro fertilization procedure to study apoptosis in this particular situation. We demonstrated a loss of cytokeratin staining as granulosa cells differentiate. We also detected that granulosa cells with apoptotic morphology did not stain for terminal deoxy-UTP nick end labeling and we showed the absence of immunoreactivity for caspase-cleaved cytokeratin, caspase-cleaved poly-ADP-ribose polymerase and caspase-cleaved caspase-3 in apoptotic granulosa cells. These data provide strong arguments for a caspase-independent cell death mechanism in human granulosa-lutein cells.
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PMID:Human granulosa cells after ovulation induction show caspase-independent cell death. 1293 44

We have investigated the expression of P2X5, P2X7, P2Y1, and P2Y2 receptor subtypes in 8- to 11-wk-old human fetal epidermis in relation to markers of proliferation (proliferating cell nuclear antigen (PCNA) and Ki-67), keratinocyte differentiation (cytokeratin K10 and involucrin), and markers of apoptosis (TdT-mediated dUTP nick end labeling (TUNEL) and anti-caspase-3). Immunohistochemistry showed that each of the four receptors was expressed in spatially distinct zones of the developing epidermis: P2Y1 receptors were found in the basal layer, P2X5 receptors were predominantly in the basal and intermediate layers, and both P2Y2 and P2X7 receptors were in the periderm. Colocalization experiments suggested different functional roles for these receptors. P2Y1 receptors were found in fetal keratinocytes positive for PCNA and Ki-67, suggesting a role in proliferation. P2X5 receptors double labeled with differentiated fetal keratinocytes that were positive for cytokeratin K10, suggesting a role in differentiation. P2X7 receptors colocalized with anti-caspase-3 antibody and were also expressed in periderm cells positive for TUNEL, suggesting a role in periderm cell apoptosis. P2Y2 receptors were found only in periderm cells and may have a role in chloride and fluid secretion into the amniotic fluid.
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PMID:Purinergic receptors are part of a signaling system for keratinocyte proliferation, differentiation, and apoptosis in human fetal epidermis. 1470 18

Caspases are responsible for a cascade of events controlling the disassembly of apoptotic cells. We now demonstrate that caspase-9 is activated at an early stage of apoptosis in epithelial cells and all its detectable, catalytically active large subunits (both the p35 and p37) are concentrated on cytokeratin fibrils. Immunolabeling of distinctive neoepitopes, exposed by cleavage of procaspase-9 at either Asp315 or Asp330, was co-localized on these fibrils with active caspase-3, caspase-cleaved cytokeratin-18, death-effector-domain containing DNA-binding protein and ubiquitin. Cytokeratin filaments may thus provide a scaffold whereby active subunits of caspase-9 can activate caspase-3 which, in turn, can activate more caspase-9 so forming an amplification loop to facilitate cleavage of cytokeratin-18, disruption of the cytoskeleton and the ensuing formation of cytoplasmic inclusions. These inclusions, formed from the collapse of fibrils, together with their associated components, also contain ubiquitinated proteins, vimentin, heat-shock protein 72, and tumor necrosis factor receptor type-1-associated death domain protein. Many of their constituents, including active caspases, remain sequestered within these inclusions, even after detergent treatment and isolation. Thus, such inclusions do not merely accumulate disrupted cytokeratins but also sequestrate potentially noxious proteins that could injure healthy neighboring cells.
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PMID:Intermediate filaments control the intracellular distribution of caspases during apoptosis. 1474 46

The ability of keratinocyte growth factor 1 to modulate apoptosis in the absence of proliferation was studied in vitro. A HaCaT scrape wound model was developed in which dense monolayers prior to wounding were cultured to quiescence in defined media with hydroxyurea at concentrations that blocked proliferation without loss of cell viability. Scrape wounding was then found to induce apoptosis, originating at the wound edge, but subsequently radiating away over a 24 h period to encompass areas not originally damaged. Keratinocyte growth factor 1 inhibited this radial progression of apoptosis in a concentration-dependent manner up to 20 ng per mL with induced migration present at the wound edge. The extent of this rescue was modulated by the concentration of Ca2+ prior to wounding. In control wound cultures apoptotic bodies were found in cells adjacent to the wound interface but were greatly reduced in keratinocyte-growth-factor-1-treated groups. Keratinocyte growth factor 1 receptor expression was significantly induced within two to three cell widths of the scraped wound edge, at levels far exceeding those found at the leading edge of a nonwounded epithelial sheet. Tumor necrosis factor alpha (1-5 ng per mL) or Escherichia coli lipopolysaccharide (10-50 ng per mL) exacerbated scrape-induced early apoptosis (1-4 h), but was largely ameliorated by coculture with keratinocyte growth factor 1. Keratinocyte growth factor 1 protection was associated with a reduction in both caspase-3 activation and cytokeratin-19 loss. Protected wound edges were also associated with the maintenance of e-cadherin expression and induction of beta1 integrin and actin stress fiber organization. These results suggest that keratinocyte growth factor 1 may play a role in limiting mechanically induced apoptotic processes at the epithelial wound edge in a manner that is distinct from its proliferative function.
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PMID:Keratinocyte growth factor 1 inhibits wound edge epithelial cell apoptosis in vitro. 1496 12

Diffuse malignant mesothelioma of the peritoneum is a rare diagnosis. Despite many histopathologic similarities between peritoneal and pleural tumors, clinical and prognostic features may be quite different. There is a paucity of data evaluating molecular features of peritoneal mesotheliomas. Therefore, we compared the results of a battery of immunohistochemical markers, some with therapeutic implications, in patients with primary peritoneal or pleural mesotheliomas. We examined 24 peritoneal and nine pleural malignant mesotheliomas with a battery of immunohistochemical markers (cytokeratin AE1/3, calretinin, c-kit/CD117, desmin, epidermal growth factor receptor (EGFR), estrogen receptors (ER), progesterone receptors (PR), MIB-1, and cleaved caspase-3) in an attempt to distinguish any differences in this tumor arising in these two distinct locations. The results indicate that the only marker to show a significant difference in its staining pattern between these two sites was EGFR (P=0.0004). In all, 92% (22/24) of peritoneal tumors demonstrated 3+ or 4+ immunoreactivity with EGFR, opposed to only 33% (3/9) pleural tumors. There was no significant difference in immunoreactivity between the pleural and peritoneal tumors with c-kit, ER, PR, cleaved caspase 3, calretinin, and desmin. There was a trend toward increased cytokeratin (P=0.07) and MIB-1 (P=0.08) expression in the peritoneal group. There was no significant difference in age, sex, or histologic subtype between the two locations. In conclusion, despite similarities between peritoneal and pleural mesothelioma, there are differences between this neoplasm arising in these two sites. The EGFR expression is more pronounced in peritoneal tumors compared to pleural tumors. The increased expression of EGFR in the peritoneal lesions may be of clinical significance with the recent emergence of epidermal growth factor receptor-targeted therapies.
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PMID:Diffuse malignant mesothelioma of the peritoneum and pleura, analysis of markers. 1497 33

When malignant cells undergo apoptosis, they exhibit many distinct patterns of behavior, with blebbing being one of the most spectacular and mysterious features. Despite huge advancements in our understanding of cell death, the mechanisms of apoptosis associated blebbing have not been elucidated. In order to verify the putative involvement of actin and tubulin in this process, Hep2 cells were treated with a combination of etoposide (10 microg/ml) and colchicine (0.2 microg/ml) for 24 h. Blebbing was analyzed using immunofluorescence staining of actin and tubulin, and the course of apoptosis was followed by time-lapse videomicroscopy, immunofluorescence detection of caspase-3 and cytokeratin fragment 18. The results indicate that microfilaments (actin) and not microtubules (tubulin) are involved in blebbing of Hep2 cells. Furthermore, despite the different mechanisms by which both chemicals act, their combined effects are not additive, but rather eliminate each other.
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PMID:Apoptosis in Hep2 cells treated with etoposide and colchicine. 1522 2

We explored the role of Peyer's patch (PP) dendritic cell (DC) populations in the induction of immune responses to reovirus strain type 1 Lang (T1L). Immunofluorescence staining revealed the presence of T1L structural (sigma1) and nonstructural (sigmaNS) proteins in PPs of T1L-infected mice. Cells in the follicle-associated epithelium contained both sigma1 and sigmaNS, indicating productive viral replication. In contrast, sigma1, but not sigmaNS, was detected in the subepithelial dome (SED) in association with CD11c(+)/CD8alpha(-)/CD11b(lo) DCs, suggesting antigen uptake by these DCs in the absence of infection. Consistent with this possibility, PP DCs purified from infected mice contained sigma1, but not sigmaNS, and PP DCs from uninfected mice could not be productively infected in vitro. Furthermore, sigma1 protein in the SED was associated with fragmented DNA by terminal deoxy-UTP nick-end labeling staining, activated caspase-3, and the epithelial cell protein cytokeratin, suggesting that DCs capture T1L antigen from infected apoptotic epithelial cells. Finally, PP DCs from infected mice activated T1L-primed CD4(+) T cells in vitro. These studies show that CD8alpha(-)/CD11b(lo) DCs in the PP SED process T1L antigen from infected apoptotic epithelial cells for presentation to CD4(+) T cells, and therefore demonstrate the cross-presentation of virally infected cells by DCs in vivo during a natural viral infection.
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PMID:Peyer's patch dendritic cells process viral antigen from apoptotic epithelial cells in the intestine of reovirus-infected mice. 1526 30

Several papers report that the colon is one of the tissues regulated by estrogen receptor (ER)beta. To better understand the physiological role of ERbeta in colonic tissue, we have compared morphology, proliferation, and differentiation of colonic epithelium in ERbeta-/- mice and WT littermates. BrdUrd labeling revealed that the number of proliferating cells was higher in ERbeta-/- mice and that the migration of labeled cells toward the luminal surface was faster in ERbeta-/- mice than in WT littermates. Additionally, in the absence of ERbeta, there was a decrease in apoptosis, which was measured by immunohistochemical staining of cleaved caspase-3. The state of differentiation of the colonic epithelial cells was studied by using epithelial markers. In ERbeta-/- mice, there was a significant decrease in the expression of the differentiation marker cytokeratin (CK)20 and in the cellular adhesion molecules alpha-catenin (an adherens junction protein) and plectin (a hemidesmosomal protein). These changes were also evident by electron microscopy as abnormalities in tight junctions and in the number and shape of desmosomes in ERbeta-/- mice. These findings suggest a role for ERbeta in the organization and architectural maintenance of the colon. Furthermore, our results indicate that the rapidly proliferating cells of the colonic epithelium in ERbeta-/- mice are lost by increased shedding and not by increased apoptosis. In this way, hyperproliferative cells that lack ERbeta do not form hyperplastic lesions and do not accumulate in the superficial epithelium.
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PMID:Role of estrogen receptor beta in colonic epithelium. 1647 31

Endothelial cells play a crucial role in the pathogenesis of many diseases and are highly sensitive to low gravity conditions. Using a three-dimensional random positioning machine (clinostat) we investigated effects of simulated weightlessness on the human EA.hy926 cell line (4, 12, 24, 48 and 72 h) and addressed the impact of exposure to VEGF (10 ng/ml). Simulated microgravity resulted in an increase in extracellular matrix proteins (ECMP) and altered cytoskeletal components such as microtubules (alpha-tubulin) and intermediate filaments (cytokeratin). Within the initial 4 h, both simulated microgravity and VEGF, alone, enhanced the expression of ECMP (collagen type I, fibronectin, osteopontin, laminin) and flk-1 protein. Synergistic effects between microgravity and VEGF were not seen. After 12 h, microgravity further enhanced all proteins mentioned above. Moreover, clinorotated endothelial cells showed morphological and biochemical signs of apoptosis after 4 h, which were further increased after 72 h. VEGF significantly attenuated apoptosis as demonstrated by DAPI staining, TUNEL flow cytometry and electron microscopy. Caspase-3, Bax, Fas, and 85-kDa apoptosis-related cleavage fragments were clearly reduced by VEGF. After 72 h, most surviving endothelial cells had assembled to three-dimensional tubular structures. Simulated weightlessness induced apoptosis and increased the amount of ECMP. VEGF develops a cell-protective influence on endothelial cells exposed to simulated microgravity.
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PMID:Induction of three-dimensional assembly and increase in apoptosis of human endothelial cells by simulated microgravity: impact of vascular endothelial growth factor. 1652 71


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