Butyric acid-induced T-cell apoptosis is mediated by caspase-8 and -9 activation in a Fas-independent manner.
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Our previous study demonstrated that butyric acid, an extracellular metabolite of periodontopathic bacteria, induced apoptosis in murine thymocytes, splenic T cells, and human Jurkat cells. In this study, we examined whether CD95 ligand-receptor interaction is involved in butyric acid-induced T-cell apoptosis. Flow cytometry analysis indicated that expression of Fas in Jurkat and T cells from peripheral blood mononuclear cells was not affected by butyric acid treatment. Furthermore, the expression of Fas and FasL protein in Western blotting was not affected by butyric acid treatment. Coincubation with blocking anti-Fas antibodies prevented Fas-induced apoptosis but not butyric acid-induced apoptosis. Anti-FasL antibodies also did not prevent butyric acid-induced apoptosis at any dose examined. Although cytotoxic anti-Fas antibody affected butyric acid-induced apoptosis, a synergistic effect was not seen. Time-dependent activation of caspase-8 and -9 was recognized in butyric acid- as well as Fas-mediated apoptosis. IETD-CHO and LEHD-CHO, specific inhibitors of caspase-8 and -9, respectively, completely blocked Fas-mediated apoptosis and partially prevented butyric acid-induced apoptosis. These results suggest that the Fas-FasL interaction is not involved in butyric acid-induced apoptosis and that caspase-8 and -9-dependent apoptosis plays an important role in butyric acid-induced apoptosis, as well as Fas-induced apoptosis. (+info)
Desensitization of ileal vagal receptors by short-chain fatty acids in pigs.
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Coloileal reflux episodes trigger specialized ileal motor activities and inhibit gastric motility in pigs. The initiation of these events requires the detection by the distal ileum of the invading colonic contents that differ from the ileal chyme primarily in short-chain fatty acid (SCFA) concentrations. In addition to the already described humoral pathway, this detection might also involve ileal vagal afferents. Sensitivity to SCFA of 12 ileal vagal units was investigated in anesthetized pigs with single-unit recording at the left cervical vagus. SCFA mixtures (0.35, 0.7, and 1.4 mol/l) containing acetic, propionic, and butyric acids in proportions identical to that in the porcine cecocolon were compared with isotonic and hypertonic saline. All units behaved as slowly adapting mechanoreceptors (half-adaptation time = 35.4 +/- 15.89 s), and their sensitivity to local mechanical probing was suppressed by local anesthesia; 7 units significantly decreased their spontaneous firing with 0.7 and 1.4 but not 0.35 mol/l SCFA infusion compared with hypertonic or isotonic saline. Similarly, the response induced by distension in the same seven units was reduced (5 neurons) or abolished (2 neurons) after infusion of 0.7 (22.8 +/- 2.39 impulses/s) and 1.4 (30.3 +/- 2.12 impulses/s) mol/l SCFA solutions compared with isotonic saline (38.6 +/- 4.09 impulses/s). These differences in discharge were not the result of changes in ileal compliance, which remained constant after SCFA. In conclusion, SCFA, at concentrations near those found during coloileal reflux episodes, reduced or abolished mechanical sensitivity of ileal vagal afferents. (+info)
On-bipolar cells and depolarising third-order neurons as the origin of the ERG-b-wave in the RCS rat.
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In the retinas of Royal College of Surgeons (RCS) rats light induces an increase in distal extracellular potassium irrespective of the age, between days 19-24 and days 29-35 postpartum, but by days 29-35 the ERG b-wave has become reduced. The synaptic blocker 2-amino-4-phosphonobutyric acid (APB) causes the abolition of both the b-wave and the potassium increase at any age. MgCl2 greatly reduces the b-wave at all ages and abolishes the potassium increase in older rats, but in younger rats the potassium increase is enlarged. Since this increase occurs in the absence of the b-wave it is unlikely that the on-bipolar cells are the only sources of the b-wave. Because the NMDA receptor blocker ketamine reduces the b-wave, third order neurons, which possess NMDA receptors, could contribute to the b-wave. (+info)
Hydrogen peroxide-induced changes in intracellular pH of guard cells precede stomatal closure.
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Epidermal bioassay demonstrated that benzylamine, a membrane-permeable weak base, can mimick hydrogen peroxide (H2O2) to induce stomatal closure, and butyric acid, a membrane-permeable weak acid, can partly abolish the H2O2-induced stomatal closure. Confocal pH mapping with the probe 5-(and-6)-carboxy seminaphthorhodafluor-1-acetoxymethylester (SNARF-1-AM) revealed that H2O2 leads to rapid changes in cytoplasmic and vacuolar pH in guard cells of Vicia faba L, i. e. alkalinization of cytoplasmic areas occur red in parallel with a decrease of the vacuolar pH, and that butyric acid pretreatment can abolish alkalinization of cytoplasmic areas and acidification of vacuolar areas of guard cells challenged with H2O2. These results imply that the alkalinization of cytoplasm via efflux of cytosol protons into the vacuole in guard cells challenged with H2O2 is important at an early stage in the signal cascade leading to stomatal closure. (+info)
Synergistic growth inhibition of prostate cancer cells by 1 alpha,25 Dihydroxyvitamin D(3) and its 19-nor-hexafluoride analogs in combination with either sodium butyrate or trichostatin A.
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Prostate cancer is a major cause of male cancer death. In vitro and in vivo data support a role for 1 alpha,25 Dihydroxyvitamin D(3) (1 alpha,25(OH)(2)D(3)) in regulating the growth and differentiation of the normal prostate gland yet prostate cancer cells appear significantly less sensitive to this action. Vitamin D(3) receptor (VDR) content or mutational status do not correlate clearly with the antiproliferative effects of 1 alpha,25(OH)(2)D(3) and therefore it is unclear why prostate cancer cell lines are significantly less sensitive to this action. We hypothesized that the antiproliferative responses of prostate cancer cells to 1 alpha,25(OH)(2)D(3) are suppressed by a process involving histone deacetylation. Sodium butyrate (NaB) and trichostatin A (TSA) are inhibitors of histone deacetylase (HDAC) activity. Low doses of NaB or TSA (300 microM and 15 nM respectively), which alone were relatively inactive, synergized with 1 alpha,25(OH)(2)D(3) in liquid and semi-solid agar to inhibit the growth of LNCaP, PC-3 and DU-145 prostate cancer cells. Still greater synergy was observed between vitamin D(3) hexafluoride analogs and either NaB or TSA. The mechanism appeared to involve neither the cyclin-dependent kinase inhibitor, p21((waf1/cip1)) nor cell cycle arrest, but rather induction of apoptosis. These data suggest that cells dysregulate the normal pro-apoptotic signals of 1 alpha,25(OH)(2)D(3) during prostate cancer development by a mechanism involving histone deacetylation. Combination therapy with potent vitamin D(3) analogs and clinically approved HDAC inhibitors may overcome this lesion and improve the treatment of both androgen-dependent and independent prostate cancer. (+info)
Downregulation of MLL-CBP fusion gene expression is associated with differentiation of SN-1 cells with t(11;16)(q23;p13).
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The translocation t(11;16)(q23;p13) has only been documented in patients with acute leukemia or myelodysplasia secondary to therapy with drugs targeting DNA topoisomerase II. We have established a myeloid cell line (SN-1) with the MLL-CBP fusion gene from an acute leukemia patient with t(11;16)(q23;p13). Although SN-1 cells were not induced to differentiate by all-trans retinoic acid (ATRA) and 1alpha,25-dihydroxyvitamin D(3) (VD3), retinoid X receptor (RXR) agonists, such as 9-cis retinoic acid and Ro48-2250, effectively induced differentiation of the cells. Downregulation of the expression of the MLL-CBP fusion gene occurred during the differentiation of SN-1 cells. When SN-1 cells were treated with MLL-CBP antisense oligonucleotide, the cells were induced to differentiate by ATRA or VD3, suggesting that the MLL-CBP fusion gene dominant-negatively suppresses ATRA- or VD3-induced differentiation. Moreover, suboptimal concentrations of sodium butyrate, a histone deacetylase inhibitor, had a cooperative effect with ATRA or VD3 in inducing the differentiation of SN-1 cells. The downregulation of the expression of MLL-CBP mRNA was accompanied by the induction of differentiation. These findings suggest that RXR agonists or a clinically applicable combination of ATRA and butyrate derivatives might be useful for differentiation therapy in leukemia patients with the MLL-CBP fusion gene. (+info)
Calmodulin-like skin protein: a new marker of keratinocyte differentiation.
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The expression of the calmodulin-like skin protein, a recently discovered new skin-specific calcium binding protein, was studied in cultured keratinocytes, reconstructed human epidermis, and normal human skin. Using a calmodulin-like skin protein specific polyclonal antibody and Western blot analysis we could show that in cultured keratinocytes calmodulin-like skin protein expression is strongly induced after stimulating cell differentiation by increasing the medium calcium concentration. Known modulators of epidermal differentiation such as sodium butyrate and the synthetic retinoid CD 367 strongly affected calmodulin-like skin protein expression. A more than 10-fold increase was observed in the presence of sodium butyrate, whereas CD 367 abolished almost completely calmodulin-like skin protein expression already at nanomolar concentrations. Calmodulin, another calcium binding protein that is expressed throughout the living layers of the epidermis, is not affected by these modulators. In normal human skin, calmodulin-like skin protein expression is restricted to the stratum granulosum and the lower layers of the stratum corneum. From these results we conclude that calmodulin-like skin protein is a new marker of late keratinocyte differentiation with a role distinct from calmodulin. (+info)
Reversal of methylation-mediated repression with short-chain fatty acids: evidence for an additional mechanism to histone deacetylation.
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We have constructed a stable cell line, human embryonal kidney 293M+, containing a lacZ reporter gene controlled by an in vitro methylated hormone-responsive enhancer. Methylation of the enhancer-promoter abolishes lacZ expression controlled by ponasterone A (an analogue of ecdysone). Ponasterone A-induced expression is restored by the short-chain fatty acids valeric > butyric > propionic > acetic acid, but not by the histone deacetylase inhibitors trichostatin A and suberoylanilide hydroxamic acid (SAHA). lacZ expression is restored to levels approaching that from an unmethylated counterpart. Incubation with short-chain fatty acids alone does not promote demethylation of the lacZ promoter, however, some demethylation (30%) is observed when transcription is triggered by addition of ponasterone A. Similar levels of hyperacetylated histones H3 and H4 were observed in cells treated with short-chain fatty acids, trichostatin A or SAHA. In vivo DNase I footprinting indicates a more open chromatin structure at the promoter region for butyric acid-treated cells. A synergistic effect in reversing the methylation-mediated repression of the lacZ gene is obtained by combined treatments with the normally ineffective compounds trichostatin A and the short-chain fatty acid caproic acid. Our results suggest the existence of an alternative silencing mechanism to histone deacetylation in executing methylation-directed gene silencing. (+info)