Meei-Yn Lin

Transcriptionally Mediated Inhibition of Telomerase of Fungal Immunomodulatory Protein From Ganoderma tsugae in A549 Human Lung Adenocarcinoma Cell Line

Telomerase expression is the hallmark of tumor cells, and activation of this ribonucleoprotein complex may be a rate‐limiting or critical step in cellular immortalization and oncogenesis. Fungal immunomodulatory protein, FIP‐gts, has been isolated from Ganoderma tsugae. In the present study, we expressed and purified the recombinant fungal immunomodulatory protein reFIP‐gts in E. coli. We found that reFIP‐gts significantly and selectively inhibits the growth of A549 cancer cells while not affecting the growth of normal MRC‐5 fibroblasts. The reFIP‐gts suppression of telomerase activity is concentration‐dependent, due to the downregulation of the telomerase catalytic subunit (hTERT). It also happens at the mRNA level. These results were confirmed by transient transfections of A549 cells with pGL3‐Basic plasmid constructs containing the functional hTERT promoter and its E‐box‐deleted sequences cloned upstream of a luciferase reporter gene. With electrophoretic mobility shift assays and Western blotting, we demonstrated that in response to reFIP‐gts, binding of c‐myc transcriptional factor to the E‐box sequence on the hTERT promoter is inhibited. These results show that reFIP‐gts suppresses telomerase activity and inhibits transcriptional regulation of hTERT via a c‐myc‐responsive element‐dependent mechanism. Our findings provide new insight into both the anticancer function of reFIP‐gts and the regulation of hTERT/telomerase expression, which may be valuable in the development of a promising chemopreventive agent.

FIP-fve stimulates interferon-gamma production via modulation of calcium release and PKC-α activation.

Fungal immunomodulatory protein, FIP-fve, has been isolated from Flammulina velutipes, and its immunomodulatory effects are believed to be associated with the enhanced activation of IFN-gamma-releasing Th1 cells. However, the mechanisms of FIP-fve-mediated signal transduction in the regulation of interferon-gamma (IFN-gamma) gene expression in human peripheral blood mononuclear cells (PBMCs) are still poorly understood. Using fluo-3 AM, we found that FIP-fve induces a rapid elevation in calcium concentration. ELISA, RT-PCR and Western blot assays demonstrated significant increases in the production and mRNA expression of IFN-gamma and protein kinase C-alpha (PKC-alpha) activation in activated PBMCs, which were abolished by EGTA, nifedipine and GO6976. In conclusion, Ca2+ release and PKC-alpha activation are required for IFN-gamma production induced by FIP-fve in PBMCs.

Lactobacillus plantarum MYL26 induces endotoxin tolerance phenotype in Caco-2 cells

BackgroundCrohns disease and ulcerative colitis are the major types of chronic inflammatory bowel disease occurring in the colon and small intestine. A growing body of research has proposed that probiotics are able to attenuate the inflammatory symptoms of these diseases in vitro and in vivo. However, the mechanism of probiotic actions remains unclear.ResultsOur results suggested Lactobacillus plantarum MYL26 inhibited inflammation in Caco-2 cells through regulation of gene expressions of TOLLIP, SOCS1, SOCS3, and IκBα, rather than SHIP-1 and IRAK-3.ConclusionsWe proposed that live/ heat-killed Lactobacillus plantarum MYL26 and bacterial cell wall extract treatments impaired TLR4-NFκb signal transduction through Tollip, SOCS-1 and SOCS-3 activation, thus inducing LPS tolerance. Our findings suggest that either heat-killed probiotics or probiotic cell wall extracts are able to attenuate inflammation through pathways similar to that of live bacteria.

Stability of fungal immunomodulatory protein, FIP-gts and FIP-fve, in IFN-γ production.

Abstract Fungal immunomodulatory proteins (FIPs) are a group of novel proteins, purified from medicinal fungi or edible mushrooms that possess immunomodulatory properties. FIP-gts and FIP-fve have been isolated and purified from Ganoderma tsugae and Flammunlina velutipes, respectively. The evaluation of FIP immunomodulatory activity was based on their ability to stimulate human peripheral blood lymphocytes to release interferon-gamma (IFN-γ). We found that FIP-gts exhibited better immunomodulatory activity than FIP-fve. Activities were both greatly reduced with duration of heating. For digestibility, FIP-fve was more resistant than FIP-gts to digestive enzymes in simulated gastric fluid and simulated intestinal fluid. IFN-γ production is only detectable in dimers of FIP-gts as opposed to polymer of FIP-fve. These results suggest that FIP-gts and FIP-fve have activities that are stable and have a strong potential of being applied to food or pharmaceutical products for commercial development.

Lactobacillus casei MYL01 modulates the proinflammatory state induced by ethanol in an in vitro model

Accumulating studies have suggested that probiotics have beneficial effects on liver injury but the underlying mechanism has remained unclear. Toll-like receptors (TLR) expressed on immune cells and hepatocytes recognize bacterial components that are translocated from the gut into the portal vein. To date, it has been demonstrated that ethanol alone, without microbial components, is able to activate TLR, leading to promotion of proinflammatory cytokine production. Because the enhanced signaling of TLR triggers persistent inflammation, we hypothesized that development of hepatocyte TLR tolerance to repetitive stimulation plays an important role in protecting the liver from hypergeneration of proinflammatory cytokines. In this study, we showed that Lactobacillus casei MYL01 modulated the proinflammatory state induced by ethanol and investigated in detail the mechanism underlying the observation that L. casei MYL01 gave rise to TLR tolerance toward ethanol stimulation. The effects of L. casei MYL01 in the attenuation of ethanol-induced liver damage were due to enhancement of IL-10 production, which limited the proinflammatory process. Furthermore, better defense of hepatocytes against ethanol challenge by treatment of L. casei MYL01 was attributed to previous induction of toll interacting protein (TOLLIP) and suppressor of cytokine signaling (SOCS)1 and SOCS3 expression via activation of TLR1, TLR2, TLR6, and TLR9, an action that cross-regulated ethanol-TLR4-nuclear factor κB signal transduction events. This finding might help establish an in vitro platform for selecting hepatoprotective probiotic strains in terms of ethanol-induced liver damage.

Differential effects of grape juice on gastric emptying and renal function from cisplatin-induced acute adverse toxicity

Grape skin and seeds contain large amounts of phytochemicals such as polyphenols, resveratrol, and proanthocyanidins, which possess antioxidant activities. Cisplatin is widely used in the treatment of cancer. High doses of cisplatin have also been known to produce acute adverse effects. The aim of this study was to investigate the protective effects of antioxidant properties of whole grape juice (with skin and seeds) on cisplatin-induced acute gastrointestinal tract disorders and nephrotoxicity in Wistar rats. Gastric emptying is significantly increased in whole grape juice-pretreated rats when compared to cisplatin treatment alone. The expression of ghrelin mRNA of stomach is increased in rats with whole grape juice. However, pretreatment with whole grape juice did not reduce renal function markers in acute renal toxicity. No significant changes were recorded in the oxidative stress/antioxidant status parameters of any study group. In contrast, pretreatment with whole grape juice slightly improved tubular cell vacuolization, tubular dilatation, and cast formation in renal tubules. These results show that consumption of whole grape juice induces somewhat beneficial effects in preventing cisplatin-mediated dyspepsia but does not offer protection against cisplatin-induced acute renal toxicity.