Fig 2: mVNS regulated myocardial pyroptosis by suppressing OGDHL expression and improving mitochondrial damage in vivo. (A) Western blot analysis of protein level of OGDHL (GAPDH as an internal control) and quantification of OGDHL activity in the myocardium of rats after I/R or mVNS treatment, n = 6 per group. (B) Western blot analysis of OGDHL expression in heart tissue transfected OGDHL, n = 6 per group. GAPDH as an internal control. (C-J) Rats were transfected with AAV9-ctrl or AAV9-cTNT-OGDHL three week before I/R surgery and mVNS tratment. (C) Evans blue and TTC staining was used to detect the infarct size of I/R rats. The area of normally supplied blood was blue, the AAR was red and that of INF was pale. INF/AAR reflects the level of dead myocardium., n = 6 per group, Scale bar: 3 mm. (D) Representative echocardiographic images showed heart function 24 h after I/R and quantitative analysis of LVEF and LVFS among the different groups, n = 6 per group. (E) TUNEL analysis for myocardial pyroptosis. Green, TUNEL-positive nuclei; blue, DAPI-stained nuclei, n = 6 per group. Scale bar: 50 μm. (F) Immunohistochemical observation of NLRP3-positive expression, n = 6 per group. Scale bar: 40 μm. (G) Western blot analysis of protein levels of NLRP3, Cleaved Caspase-1, Caspase-1, Total GSDMD and Cleaved GSDMD-N in myocardium and quantification, n = 6 per group, GAPDH as an internal control. (H) ELISA was used to measure the levels of IL-1β and IL-18 in myocardium, n = 6 per group. (I) Representative images of mitochondrial ultrastructure of myocardium with TEM, n = 6 per group. Scale bar: 2 μm. (J) Representative images of mtROS production in myocardium and quantification of relative mtROS mean intensity, n = 6 per group. Scale bar: 50 μm. Data are mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; NS means not significant between groups. All P values were obtained by one-way ANOVA

Fig 3: QHG improves myocardial markers and serum inflammatory factors in sepsis, but this effect can be reversed by MasR/PI3K-AKT-mTOR pathway inhibitors (n = 3). (A) The levels of serum myocardial markers cTnT and BNP. (B) The levels of serum inflammatory factors TNF-α and IL-1β. Data are presented as mean ± SEM. *P < 0.05, ns: no significant difference.

Fig 4: MiR‐145‐5p overexpression alleviates DOX‐induced cardiac injury. (A) A representative cardiac tissue sections stained with H&E (scale bar = 50 μm) and Masson's Trichrome (scale bar = 20 μm). Histopathological annotations: (arrow) cardiomyocyte degeneration; (triangle) inflammatory cell infiltration; (rectangle) fibrotic collagen deposition (blue in Masson). (B) Quantification of collagen volume. (C) Serum CK‐MB and (D) c‐TnT levels. (E) LVEF and (F) LVFS. (G) Quantification of serum NT‐proBNP levels. N = 10 animals per group. Data are presented as the mean ± standard deviation. ***p < 0.001.

Fig 5: MiR‐145‐5p is downregulated and SOX9 is upregulated in DOX‐induced cardiotoxicity. (A) Viability of H9C2 cells treated with DOX (1, 3, 5, 7, and 10 μM) for 24 h detected by MTT. N = 3 individual experiments. (B) The morphology of H9C2 cells treated with 5 μM DOX for 24 h observed by light microscopy (scale bar = 200 μm). Arrows indicate cell shrinkage. N = 3 individual experiments. (C) LVEF and (D) LVFS in Control group and DOX group. N = 10 animals per group. (E) Serum CK‐MB and (F) c‐TnT levels in Control group and DOX group. N = 10 animals per group. (G) MiR‐145‐5p expression in rat hearts of Control group and DOX group detected by RT‐qPCR. N = 10 animals per group. (H) SOX9 protein expression in control group and DOX group measured by Western blot. N = 10 animals per group. (I) MiR‐145‐5p (mRNA) and SOX9 (mRNA) expression levels in DOX‐treated (5 μM, 24 h) H9C2 cells detected by RT‐qPCR. N = 3 individual experiments. (J) Serum miR‐145‐5p levels and (K) SOX9 mRNA levels measured by RT‐qPCR in healthy controls, cancer patients before DOX chemotherapy (pre‐DOX), and cancer patients 48 h after final DOX infusion (post‐DOX). N = 10 per group. (L) Correlation analysis of miR‐145‐5p and SOX9 levels in 10 DOX‐treated patients. N = 10. Data are presented as the mean ± standard deviation. ***p < 0.001.