Figure 3.

MiR‐24‐3p modulates the expression of genes that regulate milk protein synthesis in mammary epithelial cells. The expression levels of factors involved in the PI3K/AKT/mTOR (AKT, mTOR, S6K1, and 4E‐BP1) and JAK/STAT (STAT5) pathways that are associated with milk protein synthesis were assessed using quantitative real‐time PCR (qRT‐PCR) in MAC‐T cells at 24 hr after transfection with miR‐24‐3p mimics (mimics)/mimics NC (mimics NC, a) and miR‐24‐3p inhibitor (inhibitor)/inhibitor NC (inhibitor NC, b). Meanwhile, the expression of κ casein (CSNK), one of the key proteins that can be detected in MAC‐T cells, was also assessed in the cells. The data are shown as the relative expression levels normalized to the internal control, β‐actin. *p <  0.05, **p < 0.01. The phosphorylation levels of AKT at Ser473, mTOR at Ser2481, STAT5 at Tyr694 were detected simultaneously. The data are shown as the relative expression levels normalized to the loading controls, β‐actin. The horizontal dashed line represents the normalized level of their corresponding negative controls (b,d). Representative WB images of the expression of AKT, AKT phosphorylated at Ser473, mTOR, mTOR phosphorylated at Ser2481, STAT5 and STAT5 phosphorylated at Tyr694 are shown (c,e). AKT: protein kinase B; JAK: Janus kinase; MAC‐T: mammary epithelial cell; mTOR: mammalian target of rapamycin; NC: negative control; PCR: polymerase chain reaction; PI3K: phosphatidylinositol‐3‐kinase; STAT: signal transducer and activators of transcription