Butyrate administration strengthens the intestinal epithelium and improves intestinal dysbiosis in a cholestasis fibrosis model.

AIM: Intestinal dysfunction in cirrhosis patients is linked to death by bacterial infections. Currently, there is no effective therapy for this complication. This study aims to evaluate butyrate, a novel postbiotic, on the intestinal inflammatory response, tight junction proteins and the microbiota in the cholestasis model. METHODS AND RESULTS: Wistar rats underwent 15 days of bile duct ligation (BDL). We administered butyrate at a concentration of 1%. The BDL group did not receive treatment. The results showed that butyrate could significantly reduce pro-inflammatory cytokines (IL-17A, IFN-γ, TNF-α) in the ileum and colon while promoting IL-10 expression in the colon. Moreover, it significantly promotes tight junction protein (cld-1, occludin and ZO-1) expression in the ileum. A similar effect was observed in the colon except for ZO-1. Additionally, butyrate limited taxa diversity loss and promoted probiotic genera expansion such as Lachnospira, Prevotella and Lactobacillus. The increase in Turicibacter and Clostridiaceae distinguished the BDL group. CONCLUSIONS: Butyrate is effective in regulating the inflammatory response, tight junction proteins and limits bacterial diversity loss. SIGNIFICANCE AND IMPACT OF THE STUDY: This research reveals that butyrate could represent an interesting postbiotic metabolomic intervention for intestinal epithelium dysfunction in liver disease.

Immune checkpoint expression on peripheral cytotoxic lymphocytes in cervical cancer patients: moving beyond the PD-1/PD-L1 axis.

Immune checkpoint therapy to reverse natural killer (NK) and T cell exhaustion has emerged as a promising treatment in various cancers. While anti-programmed cell death 1 (PD-1) pembrolizumab has recently gained Food and Drug Administration (FDA) approval for use in recurrent or metastatic cervical cancer, other checkpoint molecules, such as T cell immunoreceptor with immunoglobulin (Ig) and immunoreceptor tyrosine-based inhibition motif (ITIM) domains (TIGIT) and T cell immunoglobulin and mucin-domain containing-3 (Tim-3), have yet to be fully explored in this disease. We report expression of TIGIT, Tim-3 and PD-1 on subsets of peripheral blood NK (CD56dim/neg CD16bright/dim/neg and CD56bright CD16dim/neg ) and T cells. The percentages of these cells were increased in women with cervical cancer and pre-malignant lesions. PD-1+ NK and T cells were likely to co-express TIGIT and/or Tim-3. These cells, with an apparently 'exhausted' phenotype, were augmented in patients. A subset of cells were also natural killer group 2 member D (NKG2D)- and DNAX accessory molecule 1 (DNAM-1)-positive. PD-1int and PD-1high T cells were notably increased in cervical cancer. Soluble programmed cell death ligand 1 (PD-L1) was higher in cancer patient blood versus healthy donors and we observed a positive correlation between sPD-L1 and PD-1+ T cells in women with low-grade lesions. Within the cancer group, there were no significant correlations between sPD-L1 levels and cervical cancer stage. However, when comparing cancer versus healthy donors, we observed an inverse association between sPD-L1 and total T cells and a correlation between sPD-L1 and CD56dim NK cells. Our results may show an overview of the immune response towards pre-cancerous lesions and cervical cancer, perhaps giving an early clue as to whom to administer blocking therapies. The increase of multiple checkpoint markers may aid in identifying patients uniquely responsive to combined antibody therapies.

New amino acid changes in drug resistance sites and HBsAg in hepatitis B virus genotype H.

Long-term treatment with retrotranscriptase (RT) inhibitors eventually leads to the development of drug resistance. Drug-related mutations occur naturally and these can be found in hepatitis B virus (HBV) carriers who have never received antiviral therapy. HBsAg are overlapped with RT domain, thus nucleot(s)ide analogues (NAs) resistance mutations and naturally-occurring mutations can cause amino acid changes in the HBsAg. Twenty-two patients with chronic hepatitis B were enrolled; three of them were previously treated with NAs and 19 were NAs-naïve treated. HBV reverse transcriptase region was sequenced; genotyping and analysis of missense mutations were performed in both RT domain and HBsAg. There was predominance of genotype H. Drug mutations were present in 18.2% of patients. Classical lamivudine resistance mutations (rtM204V/rtL180M) were present in one naïve-treatment patient infected with genotype G. New amino acid changes were identified in drug resistance sites in HBV strains from patients infected with genotype H; rtQ215E was present in two naïve-NAs treatment patients and rtI169M was identified in a patient previously treated with lamivudine. Mutations at sites rt169, rt204, and rt215 resulted in the Y161C, I195M, and C206W mutations at HBsAg. Also, new amino acid changes were identified in B-cell and T-cell epitopes and were more frequent in HBsAg compared to RT domain. In conclusion, new amino acid changes at antiviral resistance sites, B-cell and T-cell epitopes in HBV genotype H were identified in Mexican patients.

Ethylenediaminetetraacetic acid induces antioxidant and anti-inflammatory activities in experimental liver fibrosis.

BACKGROUND: Experimental liver fibrosis induced by carbon tetrachloride (CCl(4)) is associated with oxidative stress, lipid peroxidation, and inflammation. This work was focused on elucidating the anti-inflammatory and antioxidant effects of ethylenediaminetetraacetic acid (EDTA) in this model of hepatotoxicity. METHODS: Wistar male rats were treated with CCl(4) and EDTA (60, 120, or 240 mg/kg). Morphometric analyses were carried out in Masson's stained liver sections to determine fibrosis index. Coagulation tests prothrombin time (PT) and partial thromboplastin time (PTT) were also determined. Gene expression for transforming growth factor beta (TGF-beta1), alpha1(I) procollagen gene (alpha1 Col I), tumor necrosis factor alpha (TNF-alpha), interleukin-6 (IL-6), and superoxide dismutase (SOD) was monitored by real-time PCR. Antioxidant effect of EDTA was measured by its effects on lipid peroxidation; biological activity of ceruloplasmin (Cp), SOD, and catalase (Cat) were analyzed by zymography assays. RESULTS: Animals with CCl(4)-hepatic injury that received EDTA showed a decrement in fibrosis (20%) and lipid peroxidation (22%). The mRNA expression for TNF-alpha (55%), TGF-beta1 (50%), IL-6 (52%), and alpha1 Col I (60%) was also decreased. This group of animals showed increased Cp (62%) and SOD (25%) biological activities. Coagulation blood tests, Cat activity, and gene expression for SOD were not modified by EDTA treatment. CONCLUSION: This study demonstrates that EDTA treatment induces the activity of antioxidant enzymes, decreases lipid peroxidation, hepatic inflammation, and fibrosis in experimental liver fibrosis induced by CCl(4).

Combinatorial gene therapy induces regression of hepatic encephalopathy.

Capillarization of the sinusoid impedes the clearance of neurotoxic substances in liver fibrosis. These events may result in hepatic encephalopathy. Neurological and hepatic features of rats after bile duct ligation (BDL) supplemented with Manganese (BDL+Mn(2+)) were examined. The 4-week-old BDL rats had elevated levels of ammonia and were concomitantly fed with 1 mg ml(-1) of MnCl(2) in drinking water (BDL/Mn(+2)). Five out of fifteen rats were killed and the serum, liver and brain tissue (striatum and substantia nigra) were recovered. Of the remaining BDL/Mn(+2)-cirrhotic animals (n=10), five were injected with a combination of Adenovirus-human plasminogen activator (Ad-huPA) and Adenovirus-matrix metalloproteinase-8 (Ad-MMP-8) (3 × 10(11)+1.5 × 10(11) vector particles per kg), and five with 4.5 × 10(11) vector particles per kg of Adenovirus-ß-galactosidase (Ad-ß-Gal). This treatment was carried on for 10 days. The BDL/Mn(+2) rats displayed tremor, rigidity and gait abnormalities, which improved notably with combinatorial gene therapy, as well as motor coordination. Liver fibrosis was evidently less after treatment with Ad-huPA+Ad-MMP-8 (25%). In the brain (striatum), Ad-huPA+Ad-MMP-8 treatment rendered higher concentrations of dopamine compared with Ad-ß-Gal-treated encephalopathic rats (210 and 162 ng g(-1) of tissue, respectively). The BDL/Mn(+2) animals and controls treated with Ad-ß-Gal showed abnormal morphology in astrocytes (gliosis) in striatum and substantia nigra, in which expressions of green fibrillar acidic protein and tyrosine hydroxylase were altered. These abnormalities decreased with Ad-huPA+Ad-MMP-8 treatment. Importantly, the latter animals showed an increment in sprouting of nervous fibers in substantia nigra. Combinatorial gene therapy improves neuroanatomical and neurochemical characteristics similar to human hepatic encephalopathy.

Hepatotoxic effects of ethanol and phenformin association in rats.

1. The association of ethanol and phenformin during 6 months in the Sprague-Dawley rat produces an alteration in lactate homeostasis. A basal blood lactate value of 54.13 +/- 15.43 mg/dl was found compared to 23.65 +/- 7.4 mg/dl in control rats. 2. Lactic acid levels increased to 28.8 +/- 7.42 mg/dl and 22.01 +/- 8.08 mg/dl after chronic administration of ethanol or phenformin in Sprague-Dawley rats, respectively. Nevertheless, these were not statistically significant with respect to those of control group. 3. The total hepatic collagen content after chronic administration of phenformin and ethanol was moderately elevated 7.12 +/- 1.85 mg/g of wet tissue, and statistically significant with respect to the control group, 4.77 +/- 1.17 mg/g. Collagen content values in phenformin and ethanol rats did not reveal statistically significant differences. 4. Liver showed histological degenerative changes but not any sign of fibrosis after chronic administration of ethanol and phenformin together.