| Retinol-binding protein4(RBP4) is adipocyte-derived secreted adipokines and many investigation shows that elevated RBP4expression level was closely positive related to insulin resistance and type Ⅱ diabetes mellitus. However, the research was limited to correlation analysis at present, the exact mechanisms are unknown. To clarify the mechanism of RBP4knockdown on insulin signal pathways, We detected the RBP4gene relative expression in different aged pig tissues and each phase of the procine adipocyte differentition, and compared it with Solexa sequencing data; Analysis the RBP4and PI3K/Akt pathways gene expression in different differentiation stages and different aged pig fat tissues. RBP4lentivirus particles were packaged to infect porcine preadipocytes. Then pig adipocytes were activated by insulin or induced adipocytes model of insulin resistance and deal with specific inhibitor of insulin pathways. RBP4interference efficiency and the gene expression of each treatment groups in PI3K/Akt pathways were examined by Real time-qPCR and Western blotting. Several results are as follows:(1) RBP4expressed in each tissues of3d old and180d old pigs, highest expressed in hepatic tissue and adipose tissue followed; The tend of RBP4expression is risen and then fell, last boost during procine adipocyte differentiation.(2) Compared with the young pigs, PI3K/Akt pathways related gene expression were reduced, it opposited with RBP4gene expression in180d old pig. With an increase in RBP4gene expression, the PI3K/Akt pathways related expression reduced in adipocyte differentiation at6,8,10d.(3) Packaging the Scramble and RBP4lentivirus by calcium phosphate method, the lentivirus titer is0.6×106KIU/L and0.6×107KIU/L, respectively. To infect porcine preadipocytes with lentivirus, resulting that RBP4mRNA and protein expressions were suppressed more than60%(P<0.01).(4) Furthermore, under insulin stimulation, RBP4knockdown significantly increased the levels of PI3K/Akt pathways related gene mRNA and phosphorylation protein expression. When RBP4knockdown could anesis the cells insulin resistance. Deal the adipocyte with specific inhibitor of PI3K, we find that RBP4knockdown could remit the insulin pathway’s inhibition effect to some degree.Collectively, knockdown of RBP4increased the insulin sensitivity through upregulated PI3K/Akt pathways related factors’expression and phosphorylation in porcine adipocytes. This research will provide a new idea to treat insulin resistance related diseases. |
