Fig 2: Casp9 HZ mice are protected from Cis-induced AKI, apoptosis, and inflammation.(A) Experiment setup. Wild-type (WT) and Casp9 HZ (HZ) mice were euthanized 3 days after Cis. CTRL mice were injected with phosphate-buffered saline (PBS). (B to D) The levels of renal Casp9 mRNA (B), BUN (C), and renal Havcr1 mRNA (D) in CTRL and Cis-treated mice. (E) Representative images and quantification of tubulointerstitial damage in hematoxylin and eosin sections in CTRL and Cis-injected kidneys of WT and Casp9 HZ (HZ) mice. Scale bar, 20 µm. (F) Western blot image and quantification of pro-CASP9 (CASP9), cleaved CASP9 (cCASP9), BAX, BCL-2, and cleaved CASP3 (cCASP3) in CTRL and Cis-injected kidneys of WT and Casp9 HZ (HZ) mice. GAPDH was used for loading CTRL. (G) Relative expression [quantitative reverse transcription polymerase chain reaction (qRT-PCR)] of Il1b, Csf2, Tnfa, Cxcl10, and Icam1 in CTRL and Cis-injected Cis kidneys of WT and Casp9 HZ (HZ) mice. CTRL WT (n = 6), CTRL HZ (n = 6), Cis WT (n = 6), and Cis HZ (n = 6). Data are presented as means ± SEM. *P < 0.05 and **P < 0.01.

Fig 3: Lower inflammation and kidney fibrosis in Casp9 HZ mice subjected to UUO injury.(A) Experimental design. (B) Western blot image and quantification of pro-CASP9 (CASP9), cleaved CASP9 (cCASP9), BAX, BCL2, and cleaved CASP3 (cCASP3) in CTRL and UUO kidney from WT and Casp9 HZ (HZ) mice. GAPDH was used for loading CTRL. (C) Relative expression (qRT-PCR) of Il1b, Csf2, Tnfa, and Cxcl10 in CTRL and UUO kidney from WT and Casp9 HZ (HZ) mice. (D) Western blot image and quantification of LC3-II, cGAS, and STING in CTRL and UUO kidney from WT and Casp9 HZ (HZ) mice. GAPDH was used for loading CTRL. (E) Representative images and quantification of tubulointerstitial fibrosis by Sirius red staining in CTRL and UUO kidney from WT and Casp9 HZ (HZ) mice. Scale bar, 20 µm. (F) Relative transcript expression (qRT-PCR) of Col1a1, Col1a3, Fn1, and Vim in CTRL and UUO kidney from WT and Casp9 HZ (HZ) mice. CTRL WT (n = 6), CTRL HZ (n = 6), UUO WT (n = 6), and UUO HZ (n = 7). Data are presented as means ± SEM. *P < 0.05 and **P < 0.01.

Fig 4: Deficiency of transforming growth factor-ß-inducible early gene-1 (TIEG1) decreases infarct size, and alters the expression of molecules in the Pten/Akt and Bcl-2/Bax signalling pathways in myocytes and endothelial cells at the mRNA and protein level. (A) Representative Masson's trichrome staining of the heart to show the infarct zone 28 days post-myocardial infarction (MI). (B) Quantification of infarct zone in heart tissue. #P<0.01 vs. the WT group. (C) Quantitative analysis of mRNA expression of TIEG1, Pten, Akt, Bcl-2, Bax, caspase-3 in cardiomyocytes (CMs). #P<0.01 vs. the WT group. (D) Western blot analysis of the altered expression of Pten/Akt and Bcl-2/Bax signalling pathways in CMs. (E) Quantification of TIEG1, Pten, Akt, Bcl-2, Bax and caspase-3 expession in CMs, n=3. #P<0.01 vs. the WT group. (F) Quantitative analysis of mRNA expression of TIEG1, Pten, Akt, Bcl-2, Bax, caspase-3 in endothelial cells. #P<0.01 vs. the WT group. (G) Western blot analysis of the altered expression of Pten/Akt and Bcl-2/Bax signalling pathways in endothelial cells. (H) Quantification of TIEG1, Pten, Akt, Bcl-2, Bax and caspase-3 expession in endothelial cells, n=3. #P<0.01 vs. the WT group.

Fig 5: METLL3 overexpression increases proliferative capacity and decreases apoptosis upon myocardial infarction (MI) in rats.A Echocardiographic data of left ventricular function, n = 5. B mRNA levels of a-MHC, ß-MHC, ANP, and BNP in rats from indicated groups, n = 5. C Representative images of non-infarcted left ventricular myocardium sections stained with wheat germ agglutinin (WGA, top), and hematoxylin-eosin (HE, bottom) from indicated groups. Scale bar, 500 µm. D Representative images of LV sections stained with Masson (top), and picrosirius red (bottom) from different groups. Scale bar, 50 µm. E Protein expression of METTL3, BAX, Bcl2, C3, and CC3 in the AC16 from indicated groups, n = 3. F, G Representative immunofluorescence images of non-infarcted left ventricular myocardium sections labeled with EDU (F) or TUNEL (G) (a-actinin, green; EDU or TUNEL, red; DAPI, blue. Scale bars, 50 µm) and the corresponding quantitative analysis, n = 3. The results are expressed as the mean ± SD.