Loureirin B inhibits the proliferation of hepatic stellate cells and the Wnt/ß-catenin signaling pathway by regulating miR-148-3p.

BACKGROUND: We investigated the activity of loureirin B against liver fibrosis and the underlying molecular mechanisms. METHODS: Hepatic stellate cells (HSCs) from Sprague-Dawley rats were treated with different concentrations of loureirin B. We used the MTT assay to determine HSC proliferation, flow cytometry to analyze apoptosis, and western blot to determine the expressions of Bax, Bcl-2, Wnt1 and ß-catenin. Real-time PCR was used to determine the expressions of Wnt1 and miR-148-3p. RESULTS: The MTT assay showed that loureirin B treatment significantly inhibited the proliferation of HSCs in time- and dose-dependent manners. Loureirin B significantly promoted the apoptosis of HSCs, increased the expression of Bax and decreased the Bcl-2 level. Western blot analysis showed that the expressions of Wnt1 and ß-catenin were obviously lower in the loureirin B treatment group than in the control group. We also found that loureirin B could decrease the Wnt1 mRNA level and increase miR-148-3p expression. Knockdown of miR-148-3p using inhibitor could reverse the effects of loureirin B on the proliferation and apoptosis of HSCs and the expressions of Bax, Bcl-2, Wnt1 and ß-catenin. CONCLUSION: Our results suggest that loureirin B inhibited the proliferation and promoted the apoptosis of HSCs, and suppressed the Wnt/ß-catenin signaling pathway via regulation of miR-148-3p.

[Gut flora and gut-derived endotoxin in minimal hepatic encephalopathy].

OBJECTIVE: To investigate the relationship of gut flora and gut-derived endotoxin with minimal hepatic encephalopathy (MHE). METHODS: Patients with hepatitis B virus-related liver cirrhosis (HBV-LC) were screened for MHE using the number connect test-A (NCT-A) and digital symbol test (DST) and divided into the following groups: HBV-LC with (+) MHE (n = 26) and HBV-liver cirrhosis without (-) MHE (n = 25); in addition, one healthy immediate family member of each patient in the HBV-LC + MHE group was enrolled as a control. Each participant provided fecal and blood samples. PCR amplification and 454 pyrosequencing were used to detect bacterial 16S rRNA in feces. Turbidimetric Limulus amebocyte lysate assay was used to detect level of endotoxin in serum. The significance of inter-group differences was assessed by one-way ANOVA or Student's t-test. RESULTS: The three groups showed different distributions of gut flora. The differences in the microbial communities' members and distributions were related to disease or health status, but not to the patient's genetic makeup or diet. In particular, the HBV-LC + MHE patients showed significantly lower amounts of different bacterial species and abundance of these species than the other two (non-MHE) groups (P less than 0.05). The healthy control family members had a richer diversity of gut flora than their counterparts with HBV-LC + MHE (P less than 0.05). The HBV-LV + MHE patients also had higher serum levels of endotoxin. CONCLUSION: Development of minimal hepatic encephalopathy in patients with HBV-LC may be related to a gut flora disorder or higher levels of endotoxin in serum.

Effect of different treatments and alcohol addiction on gut microbiota in minimal hepatic encephalopathy patients.

Minimal hepatic encephalopathy (MHE) is caused by dysbiosis of gut microbiota, particularly the ammonia-producing bacteria. Given the efficacy of certain treatments on MHE and the connection between alcoholism and MHE, a thorough understanding of how these strategies affect the gut microbiota in patients (alcoholic or non-alcoholic) will facilitate the assessment of their efficacy in the reshaping of gut microbiota. In the present study, a metagenomics approach was adopted to reveal alterations in gut microbiota of 14 MHE patients following treatment with rifaximin alone or rifaximin plus probiotics. Patients were grouped into the alcoholic and non-alcoholic groups to examine differences in terms of their response to treatment. Treatment reduced the overall microbiota diversity and decreased the abundance of certain ammonia-producing bacteria, such as Clostridium, with the treatment of rifaximin plus probiotics presenting a more apparent effect. Non-alcoholic MHE patients responded better to the treatment, as they presented greater reduction in microbiota diversity and a more consistent decline in certain ammonia-producing bacteria genera (such as Clostridium and Streptococcus) belonging to the Firmicutes phylum. In conclusion, treatment with rifaximin alone and rifaximin plus probiotics exhibited a different effect in different MHE patients, decreasing the overall gut microbiota diversity to various extents and reshaping microbiota in different ways. Furthermore, non-alcoholic MHE patients responded better to treatment in microbiota alterations.