Effects Of Selenomethionine On Gene Expression And Lactation Ability Of Female Mice Mammary Tissues

With the improvement of the living standards of Chinese residents,people’s demand for the output and quality of milk and products are increasing day by day.How to improve the quality of products and their yield has become the focus of lactation research at present.The mammary gland is the site of milk synthesis and secretion in mammals,and its growth and development process is a multifactorial regulation.Exogenous signal molecules such as amino acids have been found to regulate protein synthesis and proliferation of mammary epithelial cells.Selenomethionine(SeMet),an amino acid from organic selenoprotein,has been found to play an important role in improving the quality of livestock and poultry meat,but whether it has a regulatory effect on the growth and development of mammary gland tissue is still unclear.Therefore,SeMet was used to study the proliferation,lactation ability and related signaling pathways of HC11 cells,and explore the effects of SeMet on mammary gland development,lactation growth and related functional genes and signaling pathways in female mice.In this experiment,lactating ICR females were fed a maintenance diet supplemented with 0,5,10,20,40 and 80 mg/kg SeMet for 18 days.From the day of delivery,the litter weight was measured every3 days,and the female mouse was measured every 6 days.The lactation of the 0 mg/kg and 10 mg/kg(the best effect)groups was measured daily,and the mammary tissue growth and lactation increase of the females were analyzed based on these data.Paraffin sections were taken from the mammary glands of female mice and HE staining was performed to observe the morphology and development of mammary glands.The results showed that SeMet could dose-dependently promoted body weight growth of female mice and indirectly promoted body weight growth in pups in a dose-dependent manner;SeMet also promoted milk yield;the morphological observation of mammary gland showed that SeMet also maintained the developmental status of mammary gland and lactation performance.To investigate the gene expression status of SeMet-stimulated mammary gland development in mice,we performed eukaryotic reference transcriptome sequencing to uncover the effect of SeMet on the gene expression profile of mammary gland tissue in lactating female mice.A total of 258 differentially expressed genes were screened in the mammary tissues of SeMet-treated mice compared with control mice,including 176 highly expressed genes and 82 lowly expressed genes.The differentially expressed genes were subjected to GO and KEGG enrichment analysis,variable shear analysis,etc.The highly expressed genes were mainly closely related to the functions of substance metabolism and transport such as CTP metabolism and lipid transport,tissue development,extracellular matrix interaction and signaling.The main pathways involved were amino acid metabolism,nucleotide metabolism,cholesterol metabolism,glyoxylate and dimethyl carboxylic acid metabolism,ABC transporter protein,ECM receptor interaction,cytokine-cytokine receptor interaction,neuroactive ligand-receptor interaction,etc.;the genes with low expression level in mammary gland tissue of SeMet group were mainly related to the immune system,cellular autophagy and other functions,and the main pathways involved were IL-17,5-hydroxytryptaminergic synapses,Epstein-Barr virus infection,graftversus-host disease,inflammatory bowel disease,allogeneic rejection,the MAPK signaling pathway,etc.The results suggest that SeMet stimulates a large amount of anabolic metabolism in mammary tissue to support lactation synthesis;meanwhile,the reduction of disease-causing signaling pathways and viral and inflammatory factors indicates that SeMet may play a key role in maintaining mammary gland health and expression of immunity-related proteins in lactation.From these,11 differentially expressed genes were selected and validated by q RT-PCR analysis of mammary gland tissues and cells,and the analysis results were consistent with the sequencing results,indicating that the results of differentially expressed genes obtained from this sequencing screening are accurate and credible.To investigate the effects of SeMet on the synthesis of milk protein and milk fat and cell proliferation and key signaling molecules related to lactation in mouse mammary epithelial cells HC11,HC11 cells were treated with gradients of 0,2.5,5,10,20 and 40 μM concentrations and analyzed using apporoaches including Westernblotting,lipid droplet staining,triglyceride(TGs)assay kit,Ed U and Cell Counting Kit-8(CCK-8)to detect β-casein protein expression,TGs synthesis and lipid droplet formation,cell proliferation,as well as m TOR protein phosphorylation and downstream S6K1 protein phosphorylation levels.We also selected mammary gland tissues from control(0 mg/kg)and SeMet groups(10 mg/kg)for Westernblotting to verify the protein expression,and the obtained results were consistent with the results at cellular level.These results demonstrate that SeMet could dosedependently promote milk synthesis function of HC11 cells and promote cell proliferation through activating the m TOR-S6K1 signaling pathway.In summary,the above results indicate that SeMet supplementation can maintain healthy mammary tissue growth and lactation performance by increasing the expression of functional genes related to mammary tissue metabolism and transport,tissue development,and signaling transduction,and inhibiting the expression of functional genes related to disease and cell autophagy to promote the growth of mammary tissue and increase lactation.SeMet promotes milk synthesis and proliferation of HC11 cells by activating the lactation-related m TOR-S6K1 signaling pathway.