Fig 2: FGF21 inhibites apoptosis in flaps.a TUNEL assay performed to assess the level of DNA damage in the ischemic random skin flaps (scan bar, 15 μm) (nuclei, blue; damaged DNA, green). b Quantification of percentage of TUNEL positive cells in dermal layer. c Expression of CASP3 detected by IHC in flaps. d Quantification of integral absorbance of CASP3 protein. e, f, g Western blotting performed for expression of Bax, CYC, CASP3, which was corrected by GAPDH as internal control. h, i, j Quantization of optical density values of Bax, CYC, CASP3 expression in the flaps. Significance: *p < 0.05 and **p < 0.01 vs. Control group. Data were expressed as means ± SEM (n = 6 per group).

Fig 3: TMZ − POH induces cell death via mitochondrial pathway.(A) A549, H460 and H520 cells were treated with 100 μM TMZ, POH, TMZ + POH, TMZ − POH or DMSO respectively for 48 h and subjected to apoptosis assay using Annexin-V &PI staining. Three independent experiments were performed and the results statistically analyzed as means ± SD (B). (C) Above cells were treated with 100 μM TMZ − POH for 48 h with or without presence of z-Vad-Fmk (Z-VAD), and cell viability was detected by MTT assay. Two independent experiments were performed. (D) Lysates from drug-treated A549 and H520 cells (DMSO, TMZ, POH, TMZ + POH and TMZ − POH) were subjected to western blot using following antibodies against PARP (full and cleaved PARP), cleaved CASP9, cleaved CASP7, pro-CASP3, cleaved CASP3, Cytochrome C (CYCS) and ACTB. cl = cleaved; ns = no significance. The results shown are means ± SD; ***p < 0.001.

Fig 4: Schematic presentation probable protective mechanism of Ursodeoxycholic acid (UDCA) against As(III)-mediated hepatic injury by activating the Nrf2 pathway. (A) Protein-protein interaction (PPI) network between Nrf2-mediated oxidative stress pathway related proteins (SOD1, CAT, Gpx, HO-1, NQO-1, Keap1, Nrf2) and p53 mediated apoptosis pathway related proteins (Caspase-3, 9, Bax, Bcl-2, p53, CYCS) by STRING. (B) The green arrow indicated the pathological events involved within As(III)-exposed LO2 cells. The red lines denoted the activity restricted by UDCA.

Fig 5: 6-Gingerol induces apoptosis. (A) Flow cytometry analysis using annexin V and PI staining in MDA-MB-231 and MCF-7 cells after treatment with 6-gingerol for 48 h. (B) Graphical representation of the percentage of cells undergoing phases of apoptosis after treatment with 200 µM of 6-gingerol in breast cancer cells. The values are presented as means ± SEM of three independent experiments performed in triplicate (n = 3). *** p < 0.001 (Student’s t-test). (C) qPCR analysis of expression of intrinsic apoptosis genes. The representative expression of the BAX, BCL-2, CYCS, and CASP9 mRNA is shown; Cp values were then normalized to GAPDH mRNA. Controls are set to 100. ** p < 0.01 and *** p < 0.001 (ANOVA test), and & p < 0.01 vs. control. # p < 0.001 vs. control. (D) Western blotting analysis of MDA-MB-231 and MCF-7 cells after 48 h treatment with 100 and 200 µM 6-gingerol showing the expression of BAX, BCL-2, BCL-xL, C-caspase 9, and caspase 9. (E) The representative expression of proteins was determined by densitometry and normalized to GAPDH. Controls are set to 100. Data were confirmed after repeating the experiment three times. *** p < 0.001 (ANOVA test). # p < 0.001 vs. control.