Figure 5.

Phosphorylated p62 promotes estrogen receptor α (ESR1) expression. The results of (A) the BrdU assay and (B) Ki-67 staining revealed that the proliferation capacity of cells with silenced p62 expression was lower than that of control cells. MCF7 and Ishikawa cells were transfected for 48 h with control siRNA or p62 siRNA and subsequently exposed to 17β-estradiol (100 nM) or a vehicle for 24 h. Results are expressed as means ± standard errors of the mean from three independent experiments. * p < 0.05, ** p < 0.01. (C) The results of proximity ligation assay confirmed the interactions between ESR1, KEAP1, and p62 in human endometrial specimens. Staining was performed with antibodies raised against ESR1, KEAP1, and p62, with IgG serving as negative control. The red dots indicate protein interactions. The nuclei were stained with DAPI, which showed blue color. (D) Immunohistochemical staining of ESR1 (left panel), phosphorylated p62 (right panel) and KEAP1 (lower panel) in uterine epithelial cells of mice fed with a vehicle (upper panel) or hydroxychloroquine (HCQ, 50 mg/kg/day, lower panel) five days a week (magnifications: 10× and 40×). The arrowheads indicate different expression levels of ESR1 and phosphorylated p62.