Fig. 4: ADM promotes cell autophagy in LPS-exposed Leydig cells.

Semi-quantitative real-time PCR was performed to detect the gene expression of LC3 and Beclin-1 (a) and ATG-5 and p62 (d) normalised by the control group. Western blot was used to evaluate the protein level of LC3 and Beclin-1 (b) and ATG-5 and p62 (e). β-actin was used as internal control. Histogram displaying the densitometric analysis results of LC3 and Beclin-1 (c) and ATG-5 and p62 (f) normalised by the control group. Immunofluorescence staining was used to detect LC3 and Beclin-1 immunoreactivity (g) (scale bar: 10 µm). Fluorescence spectrophotometer was used to analyse the fluorescence intensity (h) normalised by the control group. i TEM showed the ultrastructural feature of intracellular mitochondria and autophagosomes (scale bar: 200 nm). Arrow heads show autophagosomes, and arrows show mitochondria. The normalised levels of gene expression are expressed as ratios of the copy number of mRNA and that of β-actin cDNA. Data were obtained from five independent experiments and expressed as mean ± SD. *P < 0.01 and ^#P < 0.05, compared with the corresponding treatment or control group