Lactation Biology 2

Semen Vaccariae is a traditional Chinese herb that is widely used to increase lactation. However, the underlying changes in gene expression that drive the increased milk production by Semen Vaccariae, especially the specific active monomer, remain an open question. Our lab has successfully separated active monomer dibutyl phthalate (DBP) from Semen Vaccariae to study its role in lactation. A model DT CASY Cell Counter was used to study the effect of increasing concentrations of DBP on proliferation and viability of primary cultured dairy cow mammary epithelial cells (DCMEC) harvested from a lactating cow at d 140 (n = 3). QRT–PCR and Western blot were used to study changes in mRNA and protein of prlr, erα, akt1, socs2, pparγ and elf5at 6, 12, 24, 36, 48, and 72 h. miRNAs (21,125b,143 and 195) and secretion of β–casein and lactose were detected by qRT–PCR analysis and RP-HPLC. Each assay was performed in 5 independent experiments using 3 different treatments and the data were analyzed with SPSS by ANOVA. The results showed that DBP (0.5 mg/ml) increased proliferation and viability of DCMEC significantly (P < 0.05). DBP acted similarly to prolactin (PRL). It increased the expression of prlr, erα, akt1 and elf5, but repressed the expression of pparγ. DBP promoted the expression of socs2 mRNA, but inhibited the expression of socs2 protein. Both DBP and PRL repressed the expression of miRNA–125b, miRNA–143 and miRNA–195 in DCMEC. DBP repressed the expression of miRNA–21, while the influence of PRL on miRNA–21 was uncertain. Both DBP and PRL enhanced the expression of β–casein (P < 0.05) and the secretion of lactose (P < 0.05) significantly. In conclusion, Semen Vaccariae active isomer increased proliferation and secretion of milk components by DCMEC. This is the first demonstration that miRNA expression can be changed by DBP and PRL. Further studies to uncover the lactogenic targets of DBP will help to shed light on the genetic mechanisms of mammary gland development and lactation. This work was supported by the Innovation Team of the Northeast Agricultural University (Grant No. LXT005–1–2).