Fermentation of Ganoderma lucidum and Raphani Semen with a probiotic mixture attenuates cyclophosphamide-induced immunosuppression through microbiota-dependent or -independent regulation of intestinal mucosal barrier and immune responses.

BACKGROUND: Probiotic fermentation is a promising strategy for improving the nutritional and functional properties of traditional Chinese medicines (TCMs). Ganoderma lucidum and Raphani Semen are famous TCMs that have been shown to help alleviate immune system disorders. However, few studies have experimentally investigated the effects of probiotic-fermented G.lucidum and Raphani Semen on the immune system. PURPOSE: We established the in vitro fermentation of G. lucidum and Raphani Semen with a probiotic mixture (Bifidobacterium longum, Lactobacillus acidophilus, and l. fermentum) (GRFB), investigated its ameliorating effect against cyclophosphamide (CTX)-induced immunosuppression, and explored its possible mechanisms. METHODS: First, the different components in GRFB were identified by high-performance liquid chromatography. Second, its immune-stimulatory activities were evaluated in CTX-treated mice. Lastly, its possible in vitro and in vivo mechanisms were studied. RESULTS: Probiotic fermentation of G. lucidum and Raphani Semen altered some of its chemical constituents, potentially helping improve the ability of GRFB to alleviate immunosuppression. As expected, GRFB effectively ameliorated CTX-induced immunosuppression by increasing the number of splenic lymphocytes and regulating the secretion of serum and ileum cytokines. GRFB supplementation also effectively improved intestinal integrity in CTX-treated mice by upregulating tight junction proteins. It also protects against CTX-induced intestinal dysbiosis by increasing the abundance of beneficial bacteria and reducing the abundance of harmful bacteria. GRFB could directly promote intestinal immunity but not systemic immunity in vitro, suggesting a microbiota-dependent regulation of GRFB. Interestingly, cohousing CTX-induced immunosuppressed mice with GRFB-treated mice promoted their symptoms recovery. Enhanced CTX-induced immunosuppression by GRFB in vitro depended on the gut microbiota. Remarkably, a Kyoto Encyclopedia of Genes and Genomes analysis showed that the GRFB-reprogrammed microbiota was significantly enriched in DNA damage repair pathways, which contribute to repairing the intestinal mucosal barrier. CONCLUSION: This is the first study to suggest that compare with unfermented G. lucidum and Raphani Semen, GRFB can more effectively promote intestinal immunity and manipulate the gut microbiota to promote immunostimulatory activity and repair immunosuppression-induced intestinal barrier damage by biotransforming G.lucidum and Raphani Semen components.

Effects of different doses of omega-3 polyunsaturated fatty acids on gut microbiota and immunity.

BACKGROUND: Omega-3 polyunsaturated fatty acids (PUFAs) play beneficial roles in metabolism and health. Little is known about the effects of different doses of omega-3 PUFAs on gut microbiota. OBJECTIVE: In this study, we focus on the effects of different doses of omega-3 PUFAs on gut microbiota and immunity. DESIGN: BALB/c mice was first treated with ceftriaxone sodium for 7 days, and then they received saline or different doses of omega-3 PUFAs (30, 60 and 90 mg omega-3 PUFAs) via daily gavage for 21 days. Alterations of cecum microbiota; the tight junction proteins, zonula occludens 3 (ZO3) and occludin, in the ileal wall; serum lipopolysaccharide (LPS); Interleukin-10 (IL-10), interleukin-1ß (IL-1ß), and Tumour Necrosis Factor α (TNF-α) ; mucus SIgA levels were measured. RESULTS: Compared with the ceftriaxone sodium administration group, significant increases in bacterial richness and diversity were observed in the 60- and 90-mg omega-3 PUFA groups, while only a slight increase was observed in the 30-mg omega-3 PUFA group. A higher percentage of several genera, including Lactobacillus, Helicobacter, and Ruminococcus, and a lower percentage of Bacteroides, Clostridium, and Prevotella were observed in the 60- and 90-mg omega-3 PUFA groups when compared with those in the 30-mg group. The expression of ZO3 and occludin proteins increased in 60- and 90-mg omega-3 PUFA groups compared with the natural recovery group. The mucus SIgA and serum IL-10 levels were increased, and serum levels of LPS, IL-1ß, and TNF-α were decreased in the 60- and 90-mg omega-3 PUFA groups when compared with those in the ceftriaxone sodium-treated group. CONCLUSION: Different doses of omega-3 PUFAs have different therapeutic effects on the intestinal microbiota. The 60- and 90-mg omega-3 PUFA supplementation had better recovery effects on the gut microbiota and immunity than those of the 30 mg omega-3 PUFAs supplementation.

Probiotic fermentation of Ganoderma lucidum fruiting body extracts promoted its immunostimulatory activity in mice with dexamethasone-induced immunosuppression.

Ganoderma lucidum is a legendary traditional Chinese medicine with various bioactivities. This study was conducted (a) to explore the in vitro fermentation of the water extracts of G. lucidum fruiting body with Lactobacillus acidophilus and Bifidobacterium breve and (b) to investigate the effect of fermentation broth (GLFB) on dexamethasone (DEX)-induced immunosuppressed mice. Our results demonstrated that probiotic fermentation of G. lucidum fruiting body extracts underwent structural changing of major ganoderic acid components, such as ganoderic acid A (GA) into GC2, and this fermentation process involves changing of several metabolic pathways in the probiotic strains. GLFB could significantly improve the immunity, intestinal integrity, and gut microbiota dysbiosis in DEX-treated mice, and the immunostimulatory activity of GLFB was found closely related to its direct regulation on the expansion of CD4+ T cells in Peyer's patches of mice. These data implied that probiotic fermentation of G. lucidum fruiting body extracts promoted its immunostimulatory activity via biotransformation of components such as GA. This research provides a theoretical support for the development and application of G. lucidum fermentation by probiotics.

Oral Supplements of Combined Bacillus licheniformis Zhengchangsheng® and Xylooligosaccharides Improve High-Fat Diet-Induced Obesity and Modulate the Gut Microbiota in Rats.

Gut dysbiosis induced by high-fat diet (HFD) may result in low-grade inflammation leading to diverse inflammatory diseases. The beneficial effects of probiotics and prebiotics on obesity have been reported previously. However, their benefits in promoting human health and the underlying mechanisms still need to be further characterized. This study is aimed at understanding how probiotic Bacillus licheniformis Zhengchangsheng® (BL) and prebiotic xylooligosaccharides (XOS) influence the health of a rat model with HF (60 kcal %) diet-induced obesity. Five groups of male Sprague Dawley (SD) rats were fed a normal fat diet (CON) or an HFD with or without BL and XOS supplementation for 3 weeks. Lipid profiles, inflammatory biomarkers, and microbiota composition were analyzed at the end of the experiment. Rats fed an HFD exhibited increased body weight and disordered lipid metabolism. In contrast, combined BL and XOS supplementation inhibited body weight gain and returned lipid metabolism to normal. Furthermore, BL and XOS administration changed the gut microbiota composition and modulated specific bacteria such as Prevotellaceae, Desulfovibrionaceae, and Ruminococcaceae. In addition, supplements of combined BL and XOS obviously reduced the serum LPS level, which was significantly related to microbial variations. Our findings suggest that modulation of the gut microbiota as a result of probiotic BL and prebiotic XOS supplementation has a positive effect on HFD-induced obesity in rats.

Traditional Herbal Medicine-Derived Sulforaphene LFS-01 Reverses Colitis in Mice by Selectively Altering the Gut Microbiota and Promoting Intestinal Gamma-Delta T Cells.

Sulforaphene (LFS-01) is a natural compound derived from traditional herbal medicine. Here, we show that oral administration of LFS-01 is able to dramatically alter the skewed gut microbiota and reverse colitis in model mice associated with an increase of intestinal γδT cells. Through 16S rDNA sequencing, we showed that LFS-01 can selectively suppress enteric pathogens such as Escherichia-Shigella and Helicobacter whereas the protective strains including Lactobacillus and Lachnospiraceae were significantly expanded after LFS-01 treatment. Interestingly, we demonstrated that LFS-01 administration can significantly promote the IL-17+γδT cells in model mice in response to the expanded Lactobacillus. We verified that the intracellular components of Lactobacillus can stimulate the growth of IL-17+γδT cells upon preincubation. The increased IL-17A after LFS-01 treatment in turn recovers the disrupted occludin subcellular location and protects the epithelial barrier in the colon of model mice. Remarkably, LFS-01 does not show apparent toxicity to animals and we demonstrated that LFS-01 also exerts strong protective effects in TNBS-induced colitis rats. Therefore, LFS-01 holds great promise for the treatment of inflammatory bowel disease (IBD) and warrants translation for use in clinical trials. Our work provided a new avenue for the treatment of IBD based on the strategy of harnessing intestinal symbiosis.

TmcN is involved in ATP regulation of tautomycetin biosynthesis in Streptomyces griseochromogenes.

The regulatory mechanism of tautomycetin (TMC) biosynthesis remains largely unknown, although it has been of great interest to the pharmaceutical industry. Our previous study showed that intracellular adenosine triphosphate (inATP) level is negatively correlated with secondary metabolite biosynthesis in various Streptomyces spp. In this study, by exogenous treatment of ATP, we also found a negative correlation between TMC biosynthesis and inATP level in Streptomyces griseochromogenes (S. griseochromogenes). However, the underlying mechanism remains unclear. TmcN, a pathway-specific transcriptional regulator of TMC biosynthetic genes, was previously revealed as a large ATP-binding LuxR (LAL) family protein. The predicted amino acid sequence of TmcN shows highly conserved Walker A and B binding motifs, which suggest an ATPase function of TmcN. We therefore hypothesized that the ATPase domain of TmcN may play a role in sensing endogenous pool of ATP, and is thus involved in the ATP regulation of TMC biosynthesis. To test the hypothesis, we first explored the key residue that affects the ATPase activity of TmcN by amino acid sequence alignment and structural simulation. After that, we disrupted tmcN gene in S. griseochromogenes, and the tmcN or site-direct-mutated tmcN were re-introduced to get the complementary and ATPase domain disrupted strains. The transcription level of tmcN, TMC yield, and inATP, as well as the effect of ATP on TMC production of different mutants were evaluated. Deletion of tmcN or site-direct mutation of ATPase domain of TmcN in S. griseochromogenes significantly reduced the TMC production, and it was not affected by exogenous ATP treatment. In addition, a relatively high level of inATP was detected in tmcN deletion and site-direct mutation strains. Our results here suggested that TmcN, especially its ATPase domain, is involved in consuming of endogenous ATP pool and thus plays pivotal role in connecting the primary and secondary metabolite in S. griseochromogenes.

Inhibition effects of scorpion venom extracts (Buthus matensii Karsch) on the growth of human breast cancer MCF-7 cells.

BACKGROUND: To observe the inhibition effects of the Buthus matensii Karsch (BmK) scorpion venom extracts on the growth of human breast cancer MCF-7 cells, and to explore its mechanisms. METHODS: Two common tumor cells (SMMC7721, MCF-7) were examined for the one which wasmore sensitivity to scorpion venom by MTT method. Cell cycle was determined by flow cytometry. Immunocytochemistry was applied to detect apoptosis-related protein Caspase-3 and Bcl-2 levels, while the expression of cell cycle-related protein Cyclin D1 was shown by Western blotting. RESULTS: Our data indicated that MCF-7 was the more sensitive cell line to scorpion venom. The extracts of scorpion venom could inhibit the growth and proliferation of MCF-7 cells. Furthermore, the extract of scorpion venom induced apoptosis through Caspase-3 up-regulation while Bcl-2 down-regulation in MCF-7 cells. In addition, the extracts of scorpion venom blocked the cells from G0/G1 phase to S phase and decreased cell cycle-related protein Cyclin D1 level after drug intervention compared with the negative control group. CONCLUSIONS: These results showed that the BmK scorpion venom extracts could inhibit the growth of MCF-7 cells by inducing apoptosis and blocking cell cycle in G0/G1 phase. The BmK scorpion venom extracts will be very valuable for the treatment of breast cancer.

The anti-proliferative effects and mechanisms of low molecular weight scorpion BmK venom peptides on human hepatoma and cervical carcinoma cells in vitro.

Peptides from scorpion venom have been previously studied for use in the prevention and treatment of various types of cancer in folk medicine. The present study investigated the anti-proliferative effects and mechanisms of the low molecular weight (~3 kDa) BmK scorpion venom peptides (LMWSVP) on human hepatoma (SMMC 7721) and cervical carcinoma (HeLa) cells. The data indicated that LMWSVP inhibited the growth of SMMC 7721 cells, but had no effect on the growth of HeLa cells. SMMC 7721 cells were more sensitive, with a higher affinity, to LMWSVP as compared with HeLa cells. In addition, LMWSVP induced apoptosis of SMMC 7721 cells by upregulating the expression of caspase-3 and downregulating the expression of Bcl-2. These data provide an experimental basis for further purification and application of LMWSVP for use as an anti-tumor drug in clinical trials.