Role of methyl ethyl ketone-extracellular signal-regulated protein kinase signaling pathway in breast iodine uptake during lactation

Objective Balb/c mouse mammary gland primary cells were cultured, and the expression of methyl ethyl ketone (MEK), extracellular signal-regulated protein kinase (ERK) and Na +/I- symporter (NIS) in mouse mammary gland cells was observed after stimulation with different concentrations of l7β-estradiol (E2); after application of MEK signaling pathway inhibitor U0126, the stimulation effect of E2 on mice lactating mammary cell ERK and NIS mRNA transcription level and protein expression were studied, and the role of MEK-ERK signaling pathway in iodine uptake of lactating mammary gland cell was observed. Methods Balb/c mouse mammary gland primary cells of normal primary generation were sub cultured, and then the stimulation effect of different concentrations of E2 (0.000 8, 0.004 0, 0.020 0, 0.100 0, 0.500 0 mg/L), mouse mammary gland cells MEK, ERK and mRNA NIS expression were observed; MEK inhibitor U0126 (20 μmmol/L) was applied and after 30 minutes E2 (0.1 mg/L) was administered. Cells were sub grouped to inhibitor U0126 + E2 group, E2 group and control group, cultured for 24 h, and samples were collected. Mouse mammary primary cell MEK, ERK and NIS mRNA expression levels were detected by real-time quantitative PCR; Western blotting was used to detect primary mouse mammary cell phosphorylated ERK, total ERK and NIS protein expression levels. Results In the E2 stimulating test, with increasing of E2 concentration, the expression of ERK and NIS mRNA in breast cancer cells increased gradually (F = 28.23, 18.37, all P < 0.05). In the MEK inhibition test, NIS mRNA expression level of E2 group (1.90 ± 1.36) was higher than that of U0126 + E2 inhibitor group (0.90 ± 0.39, P < 0.05), and that of control group (0.76 ± 0.18, P < 0.05), the differences were statistically significant(t = 3.218, 2.737). In total ERK protein expression, ERK protein expression of E2 group was the highest. ERK protein expression of E2 group (1.62 ± 0.30) compared with that of U0126 + E2 inhibitor group (1.19 ± 0.32) and control group (1.25 ± 0.27), the differences were statistically significant respectively (t = 2.401, 2.246, all P < 0.05). After ERK phosphorylation, ERK phosphorylated protein expression of E2 group (0.97 ± 0.02) was higher than that of U0126 + E2 inhibitor group (0.76 ± 0.18, t =-2.840, P < 0.05). NIS protein expression of E2 group (0.49 ± 0.08) was higher than that of U0126 + E2 inhibitor group (0.37 ± 0.04) and the control group (0.41 ± 0.03), and the differences were statistically significant (t = 3.286, 2.294, all P < 0.05). Conclusions E2 has raised the MEK, ERK, NIS mRNA and protein expression of lactating mammary gland cells. When MEK-ERK signaling pathway is activated, ERK and NIS mRNA and protein expression show a rising trend; when this pathway is inhibited, ERK and NIS mRNA and protein expression show a significant decreasing trend. We speculate that MEK-ERK pathway is an important signal transduction pathway of iodine uptake in lactating mammary gland. Key words: Lactating; Mammary gland cells; 17β-estradiol; Methyl ethyl ketone; Extracellular signal-regulated protein kinase; Na + /I- symporter; Signaling pathway