SMCR7/MID49 Antibody - #DF12044

Figure 4 miR-27b reduces mitochondrial fission by targeting Mff. (A, B), RT-qPCR detection of miR-27b and Mff mRNA expression levels in each cell group. (C–F), Quantitative analysis of Mff, Drp1, Mid49, and Mid51 protein expression levels by Western blot. (G), Representative Western blot bands for each group. (H), Dual-luciferase reporter assay verifying the targeting relationship between miR-27b and Mff; upper panel shows wild-type (WT) and mutant-type (MT) Mff 3’UTR sequence diagrams. (I), Transmission electron microscopy observation of mitochondrial ultrastructure in each cell group, scale bar=500 nm. (J), Confocal laser scanning microscopy observation of MFF expression and subcellular localization in each cell group, scale bar=50 μm. #P

Figure 4 Sfrp5 decreased apoptosis and improved mitochondrial dysfunction. (A) Cardiomyocyte apoptosis determined by TUNEL staining. Scale bars = 25 μm. (B) Statistical analysis of TUNEL-positive cells (values are presented as mean ± SD, n = 6 per group). (C) Protein expression of Bax and Bcl-2 in mouse hearts at 14 days after MI. (D) Quantitative analysis of the relative Bcl-2/Bax ratio (n = 5 per group). (E) Macrophage infiltration determined by immunohistochemistry. Scale bars = 50 μm. (F) Statistical analysis of F4/80-positive cells (values are presented as mean ± SD, n = 6 per group). (G) IL1β mRNA expression (n = 6 per group). β-Actin was used as a loading control (values are presented as mean ± SD, n = 6 per group). (H) Quantitative analysis of the NADH oxidase activity level (values are presented as mean ± SD, n = 6 per group). (I) Quantitative analysis of the ATP level (values are presented as mean ± SD, n = 6 per group). (J) Quantitative analysis of the NAD+/NADH ratio (values are presented as mean ± SD, n = 5–6 per group). (K) Mitochondrial morphology was detected by TEM. Scale bars = 1 μm. (L) Quantitative analysis of the size of the mitochondria (values are presented as mean ± SD, n = 6 per group). (M) Quantitative analysis of the mitochondrial number/total area (values are presented as mean ± SD, n = 6 per group). (N) Protein expression of mitochondrial fusion proteins (MFN1 and MFN2) and mitochondrial fission proteins (p-Drp1Ser616, p-Drp1Ser616/Drp, Mid49, MFF, and Fis1) in mouse hearts at 14 days after MI (n = 5 per group). (O) Quantitative analysis of the relative MFN1 protein expression. Tubulin was used as a loading control. (P) Quantitative analysis of the relative MFN2 protein expression. Tubulin was used as a loading control. (Q) Quantitative analysis of the relative p-Drp1Ser616. Tubulin was used as a loading control. (R) Quantitative analysis of the relative p-Drp1Ser616/Drp. (S) Quantitative analysis of the relative Mid49 protein expression. Tubulin was used as a loading control. (T) Quantitative analysis of the relative MFF protein expression. Tubulin was used as a loading control. (U) Quantitative analysis of the relative Fis1 protein expression. Tubulin was used as a loading control. *p < 0.05 AAV9-NC-MI mice at 14 days after MI vs. AAV9-Sfrp5-MI mice at 14 days after MI. **p < 0.01 AAV9-NC-MI mice at 14 days after MI vs. AAV9-Sfrp5-MI mice at 14 days after MI. ***p < 0.001 AAV9-NC-MI mice at 14 days after MI vs. AAV9-Sfrp5-MI mice at 14 days after MI.

Fig. 5.| HIIT and MICT ameliorated mitochondrial fission and fusion in the hippocampus of APP/PS1 transgenic mice. The levels of DRP1 (a), FIS1 (b), MFF (c),MID49 (d), MID51 (e), MFN1 (f), MFN2 (g) and OPA1 (h) were detected in the hippocampus.

Figure 3 Mitochondrial changes in activated microglia (M1). (a) Schematic diagram of intracellular mitochondrial fusion and fission process in microglia after OGD/R. (b) Western blot assay of mitochondrial fusion protein (Opa1 and Mfn1), TOM20, and cytochrome c in mitochondria of M0 and M1 microglia (n = 3). Results are displayed in a form of mean ± SD; ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001. (c) Western blotting findings of mitochondrial fission protein including MFF, Fis1, Mid49, and Mid51 in mitochondria of M0 and M1 microglia (n = 3). Data presented are mean ± SD; ∗∗P < 0.01 and ∗∗∗P < 0.001. (d) Mitochondrial function in M1 microglia and M0 microglia was investigated by determining ATP (n = 3), mitochondria membrane potential (n = 3), and ROS (n = 3). The relative ratio of intracellular ATP was determined by calculating the ratio of level of ATP in M0-BV2 and M1-BV2 cells to level of ATP in M0-BV2 cells. Results are displayed in a form of mean ± SD. ∗∗P < 0.01 and ∗∗∗P < 0.001. (e) Morphology of intracellular mitochondria in BV2 cells visualized under TEM, scale bar: 1.0 μm.