Transient Receptor Potential Vanilloid Subtype 1 Inhibits Inflammation and Apoptosis via the Release of Calcitonin Gene-Related Peptide in the Heart after Myocardial Infarction.

OBJECTIVE: A high mortality rate occurs with silent myocardial infarction (MI), particularly in aging and diabetic populations due to defects in the transient receptor potential vanilloid (TRPV1)-positive sensory nerve function. We have previously shown that TRPV1 deficiency markedly enhances post-MI inflammation and remodeling. However, the mechanisms remain unknown. The objective of this study was to clarify whether calcitonin gene-related peptide (CGRP) release was associated with the protective role of TRPV1 against postmyocardial inflammation and apoptosis. METHODS: TRPV1 gene knockout (TRPV1KO) and wild-type (WT) mice were subjected to left anterior descending ligation or sham operation. The concentration of CGRP in the myocardium was measured at 30 min, 1, 6 and 24 h post-MI. Mice received saline vehicle, CGRP or the CGRP antagonist CGRP8-37 before ligation. Inflammation was evaluated by ELISA assay and histological staining. Apoptosis was assessed by Western blot and TUNEL assay. RESULTS: Post-MI, both TRPV1KO and WT mice displayed elevated CGRP levels in myocardium when compared to sham controls. However, the levels of CGRP were significantly lower in TRPV1KO mice than in WT mice at 30 min after MI. Exogenous CGRP downregulated the levels of tumor necrosis factor-α and interleukin-6 expression in TRPV1KO mice post-MI. Moreover, exogenous CGRP decreased the neutrophil infiltration in TRPV1KO mice, whereas inhibition of CGRP by CGRP8-37 increased the neutrophil infiltration in WT mice. Western blotting data indicated that CGRP attenuated caspase-3 and caspase-9 expression, and enhanced Bcl-2 expression in TRPV1KO mice post-MI. CGRP8-37 upregulated caspase-3 and caspase-9 expression and downregulated Bcl-2 expression in WT mice. CONCLUSION: Our data suggest a protective role of TRPV1 activation against inflammation and apoptosis in mice post-MI, possibly through CGRP release. These findings elucidate a neurogenic mechanism in mice post-MI, which may participate in sensory neurotransmitter-mediated protection in TRPV1 activation.

Calcitonin Gene-Related Peptide Improves Hypoxia-Induced Inflammation and Apoptosis via Nitric Oxide in H9c2 Cardiomyoblast Cells.

OBJECTIVES: The aim of this work was to investigate whether calcitonin gene-related peptide (CGRP) plays a protective role in cardiomyocytes against hypoxia-induced inflammation and apoptosis via an NO-mediated pathway. METHODS: H9c2 cardiac cells were exposed to hypoxia for 2 h to establish a model of myocardial hypoxic-ischemic injury. The cells were pretreated with either CGRP or nitric oxide synthase (NOS) inhibitor (L-NAME) before being exposed to hypoxia for 30 min. Cell viability was analyzed using a cell counter kit 8 (CCK-8). The levels of IL-6 and TNF-α were determined by the corresponding enzyme-linked immunosorbent assay. The expression levels of several apoptosis proteins (p53, caspase-3, cytochrome C) and NOS were detected by Western blot assays. An NO kit was used to evaluate the production of NO. RESULTS: Pretreatment of H9c2 cardiac cells with CGRP for 30 min prior to exposure to hypoxia markedly improved cell viability (83.57 ± 3.21 vs. 62.83 ± 8.30%, p < 0.001); the same effect was observed following pretreatment with the NOS inhibitor L-NAME (89.34 ± 5.95 vs. 75.01 ± 5.61%, p < 0.01). Pretreatment with CGRP also significantly attenuated the inflammatory responses induced by hypoxia, as evidenced by decreases of the levels of both IL-6 (193.21 ± 13.54 vs. 293.38 ± 56.49%, p < 0.001) and TNF-α (207.71 ± 44.27 vs. 281.46 ± 64.88%, p < 0.001). Additionally, CGRP significantly decreased the hypoxia-induced overexpression of the apoptotic proteins (p53: 0.27 ± 0.10 vs. 0.87 ± 0.30, p < 0.001; caspase-3: 0.65 ± 0.15 vs. 0.98 ± 0.26, p < 0.001; cytochrome C: 1.51 ± 0.39 vs. 2.80 ± 0.69, p < 0.001) and enhanced the expression of both endothelial NOS (eNOS; 0.59 ± 0.24 vs. 0.37 ± 0.14, p < 0.05) and phosphorylated eNOS (0.60 ± 0.13 vs. 0.40 ± 0.07, p < 0.05). Furthermore, the application of both L-NAME and CGRP attenuated the hypoxia-induced expression of inducible NOS (iNOS; p < 0.05) and enhanced a hypoxia-mediated decrease in NO (p < 0.01). Interestingly, the expression levels of cell apoptosis (p < 0.05), iNOS and eNOS (p < 0.05) were decreased with L-NAME and CGRP cotreatment following 2 h of acute hypoxia, but the apoptotic factors (p < 0.05) were increased compared with only CGRP pretreatment. CONCLUSION: CGRP protects cardiomyocytes from hypoxia-induced inflammation and apoptosis by modulating NO production.

Complete genome analysis of Bacillus velezensis TS5 and its potential as a probiotic strain in mice.

Introduction: In recent years, a large number of studies have shown that Bacillus velezensis has the potential as an animal feed additive, and its potential probiotic properties have been gradually explored. Methods: In this study, Illumina NovaSeq PE150 and Oxford Nanopore ONT sequencing platforms were used to sequence the genome of Bacillus velezensis TS5, a fiber-degrading strain isolated from Tibetan sheep. To further investigate the potential of B. velezensis TS5 as a probiotic strain, in vivo experiments were conducted using 40 five-week-old male specific pathogen-free C57BL/6J mice. The mice were randomly divided into four groups: high fiber diet control group (H group), high fiber diet probiotics group (HT group), low fiber diet control group (L group), and low fiber diet probiotics group (LT group). The H and HT groups were fed high-fiber diet (30%), while the L and LT groups were fed low-fiber diet (5%). The total bacteria amount in the vegetative forms of B. velezensis TS5 per mouse in the HT and LT groups was 1 × 109 CFU per day, mice in the H and L groups were given the same volume of sterile physiological saline daily by gavage, and the experiment period lasted for 8 weeks. Results: The complete genome sequencing results of B. velezensis TS5 showed that it contained 3,929,788 nucleotides with a GC content of 46.50%. The strain encoded 3,873 genes that partially related to stress resistance, adhesion, and antioxidants, as well as the production of secondary metabolites, digestive enzymes, and other beneficial nutrients. The genes of this bacterium were mainly involved in carbohydrate metabolism, amino acid metabolism, vitamin and cofactor metabolism, biological process, and molecular function, as revealed by KEGG and GO databases. The results of mouse tests showed that B. velezensis TS5 could improve intestinal digestive enzyme activity, liver antioxidant capacity, small intestine morphology, and cecum microbiota structure in mice. Conclusion: These findings confirmed the probiotic effects of B. velezensis TS5 isolated from Tibetan sheep feces and provided the theoretical basis for the clinical application and development of new feed additives.