Apoptosis Of Colonic Epithelial Cells Research Articles

Modified apple polysaccharide prevents against tumorigenesis in a mouse model of colitis-associated colon cancer: role of galectin-3 and apoptosis in cancer prevention

Colorectal cancer (CRC) is one of the most common and preventable cancers. Regular consumption of apples is conducive to reduction in CRC risk. To evaluate effects of modified apple polysaccharide (MAP) on tumorigenesis in a mouse model of colitis-associated colon cancer. One hundred male ICR mice were administered with 1, 2-dimethyl-hydrazine (DMH) and dextran sodium sulfate (DSS). Forty mice were given no further treatment, the rest were fed basal diet blended with three different doses of MAP; 2.5, 5, and 10% (20 mice in each group). MAP significantly protected ICR mice against DMH/DSS-induced tumorigenesis. The incidence of tumor development was 90% (18/20) in the mice treated with DMH/DSS, but that was reduced to 25% (5/20), 15% (3/20), and 5% (1/20), respectively, in the mice treated with basal diets plus 2.5, 5, and 10% of MAP. Study of apoptosis of colonic epithelial cells revealed that MAP moderately increased apoptosis, suggesting that the anti-tumor potency of MAP was probably attributed to its ability to induce apoptosis. Western blot analysis demonstrated that carbohydrate-binding protein galectin-3 changed in both the nucleus and the cytoplasm during the process from colitis to colon cancer in the model. And MAP could inhibit the binding of galectin-3 to its ligand: this is, at least in part, the possible mechanism of MAP by enhancing apoptosis and preventing tumorigenesis. These data suggest that MAP has a potential role in clinical prevention and treatment for colon cancer.

Moxibustion down-regulates colonic epithelial cell apoptosis and repairs tight junctions in rats with Crohn’s disease

To investigate the effects of moxibustion on down-regulation of the colonic epithelial cell apoptosis and repair of the tight junctions in rats with Crohn's disease (CD). Sixty male Sprague-Dawley rats were randomly divided into a normal control (NC) group, a model control (MC) group, an herbs-partitioned moxibustion (HPM) group, a mild-warm moxibustion (MWM) group and a salicylazosulphapyridine (SASP) group, with 12 rats in each group. The CD model rats were treated with trinitrobenzene sulphonic acid to induce intestinal inflammation. The rats in the HPM and MWM groups were treated at the Tianshu (ST25) and Qihai (CV6) acupoints once daily for 14 d, and the SASP group was fed SASP twice daily for 14 d. No additional treatment was given to the MC and NC groups. The microstructure of the colonic epithelium was observed under a transmission electron microscope, the transepithelial resistance was measured using a short-circuit current, colonic epithelial cell apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling assay, and the expression of occludin, claudin-1 and zonula occludens-l (ZO-1) in the colonic epithelial junction was determined by Western blotting and immunofluorescence staining. Compared with the MC group, the microstructure of the colonic epithelial barrier was significantly improved in rats treated with HPM, MWM or SASP, meanwhile, the current flow was reduced significantly, with values of 168.20 ± 6.14 vs 99.70 ± 3.13, 99.10 ± 4.28 and 120.30 ± 3.65 mA, respectively (P = 0.001). However, the HPM and MWM groups had higher current flow rates than the SASP group (99.70 ± 3.13, 99.10 ± 4.28 vs 120.30 ± 3.65 mA, P = 0.001). The number of the apoptotic colonic epithelial cells in HPM, MWM and SASP groups was largely reduced (61.5 ± 16.91 vs 15.5 ± 8.89, 14.8 ± 6.27 and 24.7 ± 9.68, respectively (P = 0.001); and the expression of occludin, claudin-1 and ZO-1 in the MWM and HPM groups was significantly enhanced (0.48 ± 0.10, 0.64 ± 0.09 vs 0.18 ± 0.05 for occludin, 0.12 ± 0.02, 0.17 ± 0.03 vs 0.05 ± 0.01 for claudin-1, and 0.08 ± 0.01, 0.11 ± 0.01 vs 0.02 ± 0.01 for ZO-1). And in SASP group, the expression of occludin and ZO-1 was also significantly increased (0.27 ± 0.04 vs 0.18 ± 0.05 for occludin and 0.05 ± 0.01 vs 0.02 ± 0.01 for ZO-1), but there was no significant difference for claudin-1. The HPM and MWM groups had higher expression of occludin, claudin-1 and ZO-1 than the SASP group. HPM and MWM treatment can down-regulate apoptosis of colonic epithelial cells, repair tight junctions and enhance colonic epithelial barrier function in rats with CD.

Inflammation-induced tumorigenesis in the colon is regulated by caspase-1 and NLRC4

Chronic inflammation is a known risk factor for tumorigenesis, yet the precise mechanism of this association is currently unknown. The inflammasome, a multiprotein complex formed by NOD-like receptor (NLR) family members, has recently been shown to orchestrate multiple innate and adaptive immune responses, yet its potential role in inflammation-induced cancer has been little studied. Using the azoxymethane and dextran sodium sulfate colitis-associated colorectal cancer model, we show that caspase-1-deficient (Casp1(-/-)) mice have enhanced tumor formation. Surprisingly, the role of caspase-1 in tumorigenesis was not through regulation of colonic inflammation, but rather through regulation of colonic epithelial cell proliferation and apoptosis. Consequently, caspase-1-deficient mice demonstrate increased colonic epithelial cell proliferation in early stages of injury-induced tumor formation and reduced apoptosis in advanced tumors. We suggest a model in which the NLRC4 inflammasome is central to colonic inflammation-induced tumor formation through regulation of epithelial cell response to injury.

The Mechanisms of Sai-rei-to, a Herbal Drug, in the Treatment of Experimental Colitis

Purpose: Current medical management of IBD consists of aminosalicylats (ASAs), steroids, and immunosuppressant therapies including azathioprine, and recently TNF-α antibody therapy has also been used. But treatment with these drugs reduces immunity which may be a trigger for adverse events including infection. Sai-rei-to (SRT), a type of kampo medicine (Japanese herbal medicine), is a compound drug made from the crude extracts of various medicinal plants. These plants and extracts have been reported to have an antiinflammatory action. Recently in Japan, the administration of SRT was shown to reduce the amount of steroid administration required for IBD patients. But the mechanisms of the effect of this drug for IBD are unknown. In this study, we demonstrated the effect of SRT on colonic epithelial cell proliferation and apoptosis in rats with TNBS-induced experimental colitis. Methods: SRT, prednisolone (PSL) and 5-ASA (ASA) were administered in various combinations to TNBS rats for 4 weeks. After the rats were sacrificed, any inflammatory change of the colonic mucosa was studied using Matt's classification. The cell proliferation of colonic mucosa was studied the BrdU uptake, and apoptosis cells were stained by Tunel method. Results: The treatment of SRT, PSL and ASA alone significantly reduced colonic inflammation (TNBS control:39.75±1.26, vs. SRT:34.33±1.09, PSL:31.11±1.89, ASA:33.57±1.26) and there was no significant difference on the inflammation between these drugs. PSL administration significantly decreased the number of BrdU-labeled cells (TNBS control:4.66±1.1, vs. PSL:3.44±2.1), but SRT and ASA did not influence the number of BrdU-labeled cells (SRT:4.63±1.1, ASA:4.64±3.2). Both ASA and SRT administration increased the number of total cells in colonic pits (TNBS control:42.61±21.8, ASA:57.11±13.7, SRT:74.34±19.9). In the SRT+ASA group, administration of SRT further increased the number of total cells which had already been increased by ASA treatment (SRT+ASA:72.64±22.7). SRT and ASA treatment alone increased the number of cells above the proliferative zone (TNBS control:24.8±17.3, vs. SRT:56.63±26.3, ASA:39.19±17.8). ASA, PSL and SRT alone significantly decreased the number of apoptosis cells (TNBS control:9.53±2.17, vs. ASA:1.52±0.29, PSL:1.62±0.6, SRT:0.4±0.18). In the SRT+ASA group (ASA:1.52±0.29, vs. SRT+ASA:0.36±0.05) administration of SRT further decreased apoptosis cells which had been decreased by ASA. Conclusion: The treatment of SRT alone decreased colonic inflammation similar to PSL and ASA. SRT reduced both the colonic cells above the PZ zone and the apoptosis cells. From these data, the effect of SRT treatment for IBD is not an inducer of proliferation, and reduces and protects the desquamated colonic epithelial cells.

ErbB4 promotes cyclooxygenase-2 expression and cell survival in colon epithelial cells

The ErbB4 receptor tyrosine kinase is expressed at high levels in human and mouse colitis, and inhibits colon epithelial cell apoptosis in the presence of proinflammatory cytokines. In this study, we investigated the molecular mechanisms responsible for ErbB4-induced cell survival. In cultured mouse colon epithelial cells, ErbB4 overexpression resulted in increased levels of cyclooxygenase-2 (COX-2) mRNA and protein; in contrast, ErbB4 knockdown with siRNA blocked COX-2 accumulation in response to tumor necrosis factor. Although ErbB4 is expressed as up to four isoforms in epithelial tissues, its ability to promote COX-2 expression was isoform independent. ErbB4-stimulated COX-2 induction was associated with an increase in mRNA half-life and was blocked by inhibition of Src, phosphatidylinositol (PI) 3-kinase, or epidermal growth factor receptor (EGFR). Furthermore, ErbB4 expression promoted EGFR phosphorylation in the presence of heregulin, implicating ErbB4-EGFR heterodimerization in these responses. As to the cellular responses to ErbB4 activation, increased survival of ErbB4-expressing cells in the presence of proinflammatory cytokines was sensitive to the COX-2 inhibitor celecoxib. Furthermore, ErbB4-overexpressing cells acquired the ability to form colonies in soft agar, indicative of cellular transformation, also in a celecoxib-sensitive manner. Together our data indicate that ErbB4 is a key regulator of COX-2 expression and cellular survival in colon epithelial cells, acting in concert with EGFR through a Src- and PI 3-kinase-dependent mechanism. These results suggest that chronic overexpression of ErbB4 in the context of inflammation could contribute to colitis-associated tumorigenesis by inhibiting colonocyte apoptosis.

S1694 Protease-Activated Receptor-2 Reduces TNFα-Induced Apoptosis in Colonic Epithelial Cells

overexpressed Plexin B1 compared to normal pancreas tissues. Endogenous Plexin B1 and Met protein were detected in eight pancreatic cancer cell lines, with a variety of expression levels. Sema 4D stimulation activated Met protein, and significantly induced cell migration. Discussion: Here we show that Sema 4D involves in the regulation of cell motility through its receptor Plexin B1. And the results of immunostaining suggest that Sema 4D could play a role in the interaction between tumor cells and tumor microenvironment in pancreatic cancer tissues. As many attempts are now ongoing to target the tumor stroma, these findings provide a new insight into molecular targeted therapies for pancreatic cancer.

Anti-mutagenic lichen extract has double-edged effect on azoxymethane-induced colorectal oncogenesis in C57BL/6J mice

This study compared the effects of three anti-mutagenic lichen extracts on colorectal oncogenesis in azoxymethane (AOM)-treated mice and determined whether the extracts also regulated the homeostatic response to genotoxic damage. C57BL/6J mice (n = 12 per group) were treated with the lichen extracts Antimutagen-He (AMH): AMH-C, AMH-D, or AMH-E dimethyl sulfoxide (DMSO, control) for 2 weeks. At the end of the treatment, mice were given a single AOM injection to induce DNA damage and killed 6 h later for measuring apoptosis and proliferation. Apoptotic and proliferation indexes in mice treated with AMH-C, AMH-D, and AMH-E were 0.61%, 1.41%, and 0.77%; and 30.62%, 21.93%, and 27.27%, respectively, which were significantly lower than those of control mice (5.88% and 38.69%) (p < 0.05). To examine the effects of lichen extracts on colorectal cancer, separate groups of mice (n = 25 per group) treated with AMH-C, AMH-D, AMH-E, or DMSO were given 4-weekly AOM injections to induce oncogenesis. Mice were killed 24 weeks after the last AOM injection for assessing colon tumor formation. Colonic tumor incidences were 47.3%, 13%, and 20%; the tumor volumes were 18.47, 2.75, and 10.78 mm3, respectively, in mice treated with AMH-C (p < 0.05), AMH-D (p < 0.05), and AMH-E (p > 0.05), compared to 24% and 13.28 mm3 in mice of control correspondingly. No lichen extract showed evident toxic effects on mice. No usnic acid was found in these lichen extracts. The regulation of acute apoptosis and cell proliferation in colonic epithelial cells and the anti-mutagenesis do not seem directly related to the cancer protective effect.

The Role of SAHA for Preventing IL-13-Induced Colon Epithelial Cell Dysfunction

IL-13 has been implicated as an effector cytokine in the pathophysiology of ulcerative colitis (UC) through increasing colon epithelial cell apoptosis and expression of the pore-forming tight junction protein claudin-2. Binding of IL-13 to its receptor leads to activation of STAT6. The STAT6 phosphorylation status in patients with UC, however, is unknown, as is the effect of STAT6 inhibition on colonic epithelium exposed to IL-13. Suberoylanilide hydroxamic acid (SAHA) is a FDA-approved histone deacetylase inhibitor that inhibits constitutive STAT6 activation in lymphoma cell lines. Therefore, the aim of this study was to determine if STAT6 phosphorylation is increased in patients with UC, and if inhibition of STAT6 signaling with SAHA attenuates IL-13-induced apoptosis and claudin-2 expression. To assess IL-13 signaling, immunohistochemical staining for phosphorylated (p) STAT6 was performed on colonic tissue from pediatric subjects with UC (early UC, n= 10) and Crohn's disease (CD, n= 10) at diagnosis, colectomy tissue from adults with UC (late UC, n= 5) and controls (n= 10). A pathologist blinded to the diagnosis scored histological sections on a 0-4 scale for pSTAT6. In cell culture, human colon HT-29 cells were pre-treated with SAHA (1 or 5 μM) or vehicle control prior to treatment with IL-13 (10 ng/ml). Western blot analysis was performed on cellular lysates for pSTAT6 and claudin-2 with results quantified by densitometry. Apoptosis was determined by flow cytometry of Annexin V-PE-stained cells and by immunodetection of cleaved caspase-3. Epithelial pSTAT6 staining was increased in UC with median score of 0 in control subjects, 2 in early UC (vs. control P= 0.004), 4 in late UC (P= 0.014), and 0 in CD. In HT-29 cell culture, SAHA inhibited IL-13-induced pSTAT6 in a dose-dependent fashion with a 57% reduction of pSTAT6 induction at 5 μM (P < 0.01). IL-13 induced a 2-fold increase in claudin-2 expression (p < 0.05), and SAHA reduced IL-13-induced claudin-2 expression by 70% to baseline levels (p<0.05). By Annexin V staining, IL-13 increased apoptosis 2.7-fold (3.7 ±1.6% vs. 10.0±1.4% Annexin V+, P<0.01). SAHA alone did not increase apoptosis and reduced IL-13-induced apoptosis in a dose-dependent manner with complete suppression to baseline at 5 μM (IL-13 + SAHA 1 μM 7.5±0.8%, + 5 μM 4.8±0.9, Ptrend< 0.001). Similarly, by immunodetection of cleaved caspase-3, SAHA reduced IL-13 induced apoptosis by 57% at 1 μM and 94% at 5 μM (P<0.001 for each compared to IL-13 alone). Increased STAT6 phosphorylation in the colonic epithelium of pediatric subjects at presentation with UC supports an effector role of IL-13 early in the disease. Treatment of colonic epithelial cells with SAHA, a drug that inhibits IL-13-induced STAT6 activation, reduces IL-13-induced apoptosis and claudin-2 expression, suggesting that STAT6 may be a novel therapeutic target for UC.

Primary human colonic epithelial cells are transiently protected from anoikis by a Src-dependent mechanism

Complete loss of cell anchorage triggers apoptosis in primary human colonic epithelial cells (CEC), a phenomenon known as anoikis. Besides the induction of pro-apoptotic events, activation of survival pathways was observed in detached intestinal epithelial cell lines, providing a transient apoptosis protection. However, nothing is known about molecular mechanisms protecting primary CEC from anoikis. In this study intact CEC crypts were isolated and kept in suspension, a condition which leads to the loss of cell–cell anchorage and induces anoikis. To reconstitute cell–cell contacts, cells were centrifuged to form cell aggregates. Induction of apoptosis was assessed by caspase-3 activity assay; activation of survival pathways was analyzed by Western blot. Immediately after loss of cell anchorage a rapid activation of survival proteins was observed before active caspase-3 could be detected. Src hyperactivation significantly contributed to transient protection from anoikis in CEC because its inhibition reversed the protecting effect of re-establishment of cell contacts. Basal levels of active Src in CEC from patients with inflammatory bowel disease were markedly reduced compared to control patients. These results demonstrate that loss of cell anchorage activates survival pathways in primary human CEC providing transient anoikis protection. Src is an important mediator of this mechanism and therefore constitutes a key regulatory molecule coordinating survival signals mediated by cell adhesion in primary human CEC.

T1749 Radiation-Induced Colonic Epithelial Cell Apoptosis and Crypt Killing Is Promoted By Intestinal Microbes and MyD88

T1749 Radiation-Induced Colonic Epithelial Cell Apoptosis and Crypt Killing Is Promoted By Intestinal Microbes and MyD88

Calpain-dependent degradation of transcription factors and X-linked inhibitor of apoptosis (XIAP) in colon epithelial cells during cell death induced by Entamoeba histolytica (133.4)

Abstract Entamoeba histolytica is a tissue-invasive protozoan parasite causing dysentery in human beings. During the infection in the colonic tissues, amoebic trophozoites have a capability to kill host cells via apoptosis or necrosis, which triggers IL-8-mediated acute inflammatory responses. However, signaling pathways involved in host cell death by E. histolytica is not fully defined. In this study, we examined whether calpain is closely involved in cleavage of transcription factors and XIAP, which are responsible for cell survival, during cell death of colon epithelial cells induced by live trophozoites of E. histolytica. Incubation with amoebic trophozoites induced PS externalization in CaCo2 cells in a time and dose-dependent manner. In addition, E. histolytica activated m-calpain and caspase-3 in CaCo2 cells. Moreover, incubation with amoeba resulted in marked degradation of STAT proteins (STAT1, 2, 3, 5, and 6), NF-¿B (p65 or p50), and XIAP in CaCo2 cells. However, I¿B, an inhibitor of NF-¿B, was not cleaved in CoCo2 cells following adherence of E. histolytica. Entamoeba-induced cleavage of STAT proteins, NF-kB, and XIAP was inhibited by pretreatment of cells with a cell permeable calpain inhibitor, calpeptin. Furthermore, calpeptin, but not z-VAD-fmk, reduced Entamoeba-induced DNA fragmentation in CoCo2 cells. These results suggest that calpain-mediated degradation of anti-apoptotic proteins, such as transcription factors and XIAP, is required for acceleration of cell death in colon epithelial cells induced by Entamoeba histolytica.

Reduced colonic apoptosis in mice overexpressing bovine growth hormone occurs through changes in several kinase pathways

Objective Growth hormone (GH) has antiapoptotic effects in several cell lines, including human colonic adenocarcinoma cells. In addition, it has been reported that patients with acromegaly have reduced apoptosis in colonic mucosa. The aim of the study was to investigate colonic apoptosis and underlying molecular mechanisms in transgenic mice overexpressing bovine GH (Acro) aged 3 months (young) or 9 months (elder). Design and methods Apoptosis in colonic epithelial cells was evaluated by TUNEL and Annexin V; expression of pro- and anti-apoptotic proteins was assessed by Western blot. GH action was blocked treating Acro with a selective GH receptor antagonist. Results Young and elder Acro had lower colonic apoptosis [driven by GH through p38, p44/42 and PI3 kinase pathways], than littermate controls; changes were abolished by treating Acro with a selective GH receptor antagonist. The effects of GH were consistent with an anti-apoptotic phenotype (reduced cytosolic cytochrome-c, Bad and Bax and increased Bcl-2, and Bcl-XL level) leading to lower activation of caspase-9 and caspase-3. Changes in apoptotic proteins reversed after treatment with a GH receptor antagonist, suggesting a direct effect of GH. In addition, antiapoptotic phenotype of Acro had a protective role against doxorubicin-induced apoptosis. Conclusions Our results suggest that GH leads to increased and reduced levels of anti- and pro-apoptotic proteins, respectively, lowering apoptosis in either young or elder transgenic animals through activation of several kinase pathways.

TNFR1 Promotes Tumor Necrosis Factor-mediated Mouse Colon Epithelial Cell Survival through RAF Activation of NF-κB

Tumor necrosis factor (TNF) is a therapeutic target in the treatment of inflammatory bowel disease; however, the exact role of TNF signaling in the colon epithelium remains unclear. We demonstrate that TNF activation of TNF receptor (R)1 stimulates both pro- and anti-apoptotic signaling pathways in the colon epithelium; however, TNFR1 protects against colon epithelial cell apoptosis following TNF exposure. To investigate anti-apoptotic signaling pathways downstream of TNFR1, we generated an intestinal epithelium-specific Raf knock-out mouse and identified Raf kinase as a key regulator of colon epithelial cell survival in response to TNF. Surprisingly, Raf promotes NF-kappaB p65 phosphorylation, independent of MEK signaling, to support cell survival. Taken together, these data demonstrate a novel pathway in which Raf promotes colon epithelial cell survival through NF-kappaB downstream of TNFR1 activation. Thus, further understanding of colon epithelial cell-specific TNFR signaling may result in the identification of new targets for inflammatory bowel disease treatment and define novel mediators of colitis-associated cancer.

The role of GSH efflux in staurosporine-induced apoptosis in colonic epithelial cells

Staurosporine (STP) was shown to induce cell apoptosis through formation of reactive oxygen species, but a role for cellular redox has not been defined. In this study, we report that STP (2 μM) caused apoptosis (24 ± 3% at 24 h) of human colon adenocarcinoma epithelial cell line HT29 that was preceded by significant glutathione (GSH) and glutathione disulfide (GSSG) efflux (6 h), but independent of changes in cellular glutathione/glutathione disulfide (GSH/GSSG) redox status. The blockade of GSH efflux by γ-glutamyl glutamate (γ-GG) or ophthalmic acid was associated with apoptosis attenuation; however, γ-GG administration after peak GSH efflux (8 h) did not confer cytoprotection. Moreover, lowering cellular GSH through inhibition of its synthesis prevented extracellular GSH accumulation and cell apoptosis, thus validating a link between cellular GSH export and the trigger of cell apoptosis. Inhibition of γ-glutamyl transferase (GGT1, EC 2.3.2.2)-catalyzed extracellular GSH degradation with acivicin significantly blocked GSH efflux, suggesting that GSH breakdown is a driving force for GSH export. Interestingly, acivicin treatment enhanced extracellular GSSG accumulation, consistent with GSH oxidation. STP-induced HT29 cell apoptosis was associated with caspase-3 activation independent of caspase-8 or caspase-9 activity; accordingly, inhibitors of the latter caspases were without effect on STP-induced apoptosis. STP similarly induced GSH efflux and apoptosis in a non-malignant human NCM460 colonic cell line in association with caspase-3 activation. Collectively, our results demonstrate that STP induction of apoptosis in malignant and non-malignant colonic cells is temporally linked to the export of cellular GSH and the activation of caspase-3 without caspase-8 or -9 involvement.

Regulation of intestinal barrier function by signal transducer and activator of transcription 5b

Background:Colon epithelial cell (CEC) apoptosis and nuclear factor-κB (NF-κB) activation may compromise barrier function, and it has been reported that signal transducer and activator of transcription 5b (STAT5b)-deficient mice exhibit...

The interaction of butyrate with TNF-α during differentiation and apoptosis of colon epithelial cells: Role of NF-κB activation

We demonstrated that TNF-α suppressed differentiation and potentiated cell death induced by butyrate (NaBt) in both adenocarcinoma HT-29 and fetal FHC human colon cells in vitro. Since TNF-α is a typical activator of NF-κB pathway, we studied the role of NF-κB activation in cell differentiation and death during the TNF-α and NaBt co-treatment. TNF-α induced rapid NF-κB activation in both HT-29 and FHC cell lines and this effect was differently modulated by NaBt in these two cell lines. In HT-29 cells, NaBt potentiated NF-κB activity induced by TNF-α after 4 h treatment. However, this initial potentiation of NF-κB activity was not observed in FHC cells. During additional time of TNF-α and NaBt co-treatment, NaBt decreased the TNF-α-mediated NF-κB activity in both cell types. We also detected a different response of HT-29 and FHC cells after the pre-treatment with the NF-κB inhibitor parthenolide. Our results indicated that NaBt-mediated differentiation and apoptosis of colon epithelial cells can be modulated by TNF-α. Furthermore, we found significant differences in the mechanism of the NaBt and TNF-α co-treatment effects between cells of non-cancer and cancer origin, suggesting that the NF-κB pathway may be more effectively involved in these processes in cancer cells.

Recombinant human granulocyte colony-stimulating factor reduces colonic epithelial cell apoptosis and ameliorates murine dextran sulfate sodium-induced colitis

Objective. Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a potentially effective therapy for Crohn's disease. The purpose of this study was to test the rhG-CSF in murine dextran sulfate sodium-induced colitis (DSS colitis). Material and methods. Murine colitis was induced by feeding with water containing 3% DSS for 9 days. Six to 7-week-old female BALB/c mice were given rhG-CSF (100 µg/kg) or phosphate-buffered saline (PBS) subcutaneously once a day from day 0 to day 8, and the mice were sacrificed at days 3, 5, 7 and 9. Tissue specimens from the transverse colon, descending colon and rectum were obtained and stained with hematoxylin and eosin. Inflammation was scored for severity, extent, epithelial damage and crypt loss. TUNEL staining was performed to assess epithelial cell apoptosis. Results. Treatment with rhG-CSF significantly attenuated body-weight loss, stool score and shortening of the colon length in comparison with treatment with PBS (p<0.01,<0.05,<0.01, respectively). Histological scores for inflammation, epithelial cell damage and crypt loss of the rectum were less severe at day 9 in the rhG-CSF group than in the PBS group (p<0.01, 0.05, 0.01, respectively). The number of TUNEL-positive cells in the rectum was smaller in the rhG-CSF group than in the PBS group (p<0.001). Conclusions. Treatment with rhG-CSF ameliorates murine DSS colitis by suppressing mucosal inflammation and epithelial damage in the rectum. The prevention of epithelial cell apoptosis seems to precede the anti-inflammatory action of rhG-CSF.

Mitochondrial GSH transport and intestinal cell injury: A commentary on “Contribution of mitochondrial GSH transport to matrix GSH status and colonic epithelial cell apoptosis”

Mitochondrial GSH transport and intestinal cell injury: A commentary on “Contribution of mitochondrial GSH transport to matrix GSH status and colonic epithelial cell apoptosis”

Contribution of mitochondrial GSH transport to matrix GSH status and colonic epithelial cell apoptosis

Previously, we showed that cellular glutathione/glutathione disulfide (GSH/GSSG) play an important role in apoptotic signaling, and early studies linked mitochondrial GSH (mtGSH) loss to enhanced cytotoxicity. The current study focuses on the contribution of mitochondrial GSH transport and mitochondrial GSH/GSSG status to apoptosis initiation in a nontransformed colonic epithelial cell line, NCM460, using menadione (MQ), a quinone with redox cycling bioreactivity, as a model of oxidative challenge. Our results implicate the semiquinone radical in MQ-mediated apoptosis, which was associated with marked oxidation of the mitochondrial soluble GSH and protein-bound thiol pools, mitochondria–to–cytosol translocation of cytochrome c, and activation of caspase-9. MQ-induced apoptosis was potentiated by inhibition of mtGSH uptake in accordance with exacerbated mitochondrial GSSG (mtGSSG) and protein–SSG and compromised mitochondrial respiratory activity. Moreover, cell apoptosis was prevented by N-acetyl- l-cysteine (NAC) pretreatment, which restored cellular redox homeostasis. Importantly, mtGSH transport inhibition effectively blocked NAC-mediated protection in accordance with its failure to attenuate mtGSSG. These results support the importance of mitochondrial GSH transport and the mtGSH status in oxidative cell killing.

Unusual Reasons for Incomplete Colonoscopy

Purpose: A 50 year old woman had h/o chronic constipation for 15 years. She could not have any bowel movement without the use of laxatives. She has been using laxatives daily for 15 years but she assured us that she only used a”safe, natural, herbal” laxative. Therewas no h/o hematemesis, melena or hematochezia. There were no constitutional symptoms. There were no other GI complaints, there was no F/H of colon cancer. The review of systems was unremarkablePhysical Examination was normal. CBC & Chemistry panel was normal. A screening colonoscopy was per formed after usual PEG preparation. The rectum and the sigmoid colon were very dark due to severe melanosis coli. As the scope entered the descending colon, the view became pitch dark; the endoscopist felt he was in a jungle at night without a flash light. Manually increasing the brightness to the maximum on the light source did not help. The procedure was abandoned. Biopsies were obtained in the sigmoid colon. Later on an elective BE ruled out any masses or polyps in the rest of the colon. The histologic examination of the biopsy showed extensive infiltration of the cells in the lamina propria & submucosa with pigment. On further questioning, the patient informed us that she has been using Nature's Secret—Super Cleanse laxative which she said had natural herbs. An evaluation of the composition of Super- Cleanse showed it contained Cascara sagrada bark which is rich in anthraquinones & anthracene which are known to cause melanosis coli. Methods: This is an unusual case in that SEVERE melanosis coli prevented completion of screening colonoscopy. The exact composition of so called”narural, herbal, safe” laxatives should be ascertained: besides cascara sagrada other herbs such as Aloe vera and Senna (Swiss Kriss) may contain chemicals which can cause melanosis coli & other serious side effects. The pigment in melanosis coli is not melanin so it should be called pseudo -melanosis coli. Melanosis coli involving pericolonic lymph nodes associated with the herbal laxative Swiss Kriss has been reported in a patient with colonic adenocarcinoma (Arch Path lab Med128:565–567, 2004). Rarely melanosis may involve other parts of the GI tract such as the ileum, esophagus & stomach. It may also involve the Vagina & Prostate. Anthraquinone-induced apoptosis of colonic epithelial cells may be important in pathogenesis.(Am J Path, 131:465–476, 1988).