Does Insulin, as Choice Agent, Recover The Diabetes-Induced Germ Cell Cycle Arrest during Spermatogenesis; An Experimental Trial

Background: Diabetes is majorly sub categorized into: diabetesmellitus or type I (DM) and type II. The diabetes mellitus isassociated with insulin disarrangement, and is phenotypicallymanifested with hyperglycemia. Thus, in the case of DM, theinsulin-therapy has been accepted as a routin medication withhigh efficiency on cellular and tissue metabolism. It has been illustratedthat, the cyclins (especially cyclin D۱) and the kinases(especially CDK-۴) in association with kinase inhibitor p۲۱ areable to fairly maintain and/or inhibit the cell cycle arrest duringmitosis and meiosis of germ cells. Considering the potential effectof diabetes-induced oxidative stress and metabolic failureagainst DNA integrity, the present study was aimed to uncoverthe cross-link between two mentioned majorities with cell cyclemachinery via focusing on the role of cyclin D۱, CDK-۴ and theinhibitor p۲۱ (as kinase inhibitor) and p۵۳ as mediator elementbetween p۲۱-induced apoptosis.Materials and Methods: Thirty mature Wistar rats were dividedinto ۳ groups: Control, DM-induced (by single injectionof ۵۰ mg/kg of sterptozotocin, intraperitonealy) and insulintreatedDM-induced (DMI). After ۵۶ days, the histological alterations,the tubular differentiation (TDI), and spermiogenesis(SPI) indices were assessed by H&E staining. Moreover, theTUNEL staining was performed to estimate the apoptois index.The expressions of p۵۳, p۲۱, cyclin-D۱ and cdk-۴ were evaluatedusing RT-PCR and immunohistochemistry staining (IHC)techniques, respectively. Finally, to reduce the bias problems/errors the positive reactions of TUNEL and IHC staining techniqueswere re-evaluated by pixel based frequency analyzes andthe positive cells distribution percentage per mm۲ of testiculartissue were counted and compared between groups.Results: The animals in DM-sole group exhibited diminishedTDI and SPI ratios by representing the enhanced percentageof tubules with negative germ cell differentiation as well asspermiogenesis. However, the insulin remarkably (P<۰.۰۵) improvedTDI and SPI ratios. The DMI animals represented noremarkable improvement in cdk-۴ expression, while they exhibitedenhanced cyclin D۱ mRNA and protein contents. Nostatistical changes were revealed in mRNA level of p۲۱ in DMIgroup, whereas significantly lower p۲۱+ cells were revealed inthe same group versus DM-sole animals. Finally, the animals inthe DMI group showed diminished expression of p۵۳ as well asapoptotic cellularity compared to DM-sole group.Conclusion: To wrap it up, the present study illustrated that,the insulin-therapy was remarkably ameliorated DNA damageand p۵۳ overexpression in testicular tissue. Meanwhile simultaneouslythe insulin could not potentially rainforce the cell cycle machinery. Because, the expression of cdk-۴ did not altersignificantly after insulin administration. As a logic theory,despite of insulin-induced reduction in p۲۱ protein content, theappropriate cdk-۴ expression was not found in the germ line,leading to lame cell cycle progression. Taking together, it couldbe concluded that the insulin, potentially inhibits the apoptosisratio, while it is not able to directly/effectively impact the cellcycle machinery, at least in the case of cyclin D۱-cdk-۴-relatedpathway. More analyses are needed to find the other possibleeffects of insulin on cell cycle progression in diabetic cases.

کلیدواژه ها:

Diabetes Mellitus ، Insulin ، Germ Cell ، Cell Arrest