Mechanisms Of KISS-54 And Isl-1 Regulating Insulin Synthesis And Secretion

It has been documented that LIM homeodomain transcription factor Isl-1(Insulin gene enhancer binding protein 1)directly binds to insulin promoter and promotes its transcription.Isl-1 plays an important role in insulin secretion and islet development.KISS-54(kisspeptin54)is a peptide hormone that affects insulin secretion.However,we still do not know whether there are interactions between KISS-54 and Isl-1 in regulating insulin secretion.In the present study,mouse pancreas and mouse pancreatic islet p cell line NIT cells are used to investigate whether Isl-1 mediates the effect of KISS-54 on insulin secretion.The important findings in our study include the following points:1.Immunofluorescence double staining results showed that KISS-54 receptor G protein coupled receptor 54(GPR54)and Isl-1 were coexpressed in the pancreatic β cell.Western blot results showed that Isl-1 and GPR54 were highly expressed in pancreatic islet and NIT cells.2.Isl-1 protein level and insulin level were analysed in NIT cells which were added with KISS-54,and the results showed that KISS-54 significantly inhibited Isl-1 expression and insulin secretion.In addition,the insulin degradation was detected in NIT cells after 10 nM KISS-54 treatment.The results showed that KISS-54 had no obvious effect on insulin degradation in different time points.These data indicate that KISS-54 inhibits insulin secretion through the related signaling pathway but not insulin degradation.3.In order to investigate the effect of KISS-54 on the expressions of the transcription factors related to insulin secretion in NIT cells,MafA、Ngn3、Pdx1、Pax4 and Nkx6.1 mRNA levels were analysed by Real-time PCR after 10 nM KISS-54 treatment for 3 h.The results showed that KISS-54 treatment had no significant effect on mRNA levels of these factors,with exception that the MafA mRNA level significantly increased.These results infer that the KISS-54 specifically inhibits Isl-1 expression.4.We used Isl-1 IKO(Inducible knockout)mice to investigate whether the effect of KISS-54 on insulin secretion is Isl-1 dependent.The results showed that Isl-1 IKO had no evident influence on the mouse weight,pancreas weight and daily food intake,but the Isl-1 IKO mice displayed glucose and insulin tolerance impairment.5.Our results have showed that KISS-54 treatment resulted in a decline in insulin mRNA level in wild-type mice,but this kind of effect was not obvious in IKO mice.Moreover,KISS-54 treatment decreased insulin levels after 3 h of treatment in control islets,but no significant effect of KISS-54 treatment on insulin secretion in IKO islets was seen.These suggest that the inhibitory effect of KISS-54 on insulin secretion relies on Isl-1.6.The in vitro results showed that KISS-54 inhibited insulin synthesis and secretion through PKC-ERK signaling pathway.7.Further,we assayed the feed-back regulation of insulin on kisspeptin synthesis and secretion,and the in vitro and in vivo results showed that insulin inhibited kisspeptin synthesis and secretion dependent on the JAK-PI3K signaling pathway.In conclusion,our study suggest that Isl-1 mediates the effect of KISS-54 on insulin secretion and KISS-54 inhibits the Isl-1 expression and insulin secretion through PKC-ERK pathway in pancreatic islet β cell.However,insulin inhibits kisspeptin production and secretion through JAK-PI3K signaling pathway.These results are important in understanding the mechanisms regulating insulin secretion and for providing potential molecular target for the treatment of diabetes.