Effect Of IR-61 On Voiding Function In STZ-Induced Diabetic Rats

BACKGROUND: Diabetic bladder dysfunction(DBD)is the most common complication of urinary system in diabetic patients,which afflicts nearly half of diabetic patients.The prevalence of DBD in diabetic patients has been estimated to be 43-87%.While DBD does not endanger the patient’s life,it greatly affects the patient’s quality of life.So far,however,there is still a lack of effective treatment for DBD in clinical practice for those patients who fail to respond to glycemic control and behavioral therapy,especially those with advanced DBD who develop detrusor weakness.Therefore,it is still of great clinical significance to develop new treatments for DBD.In a previous study,our research group synthesized a novel small molecule compound — IR-61.IR-61 has been found to have good biocompatibility,can preferentially accumulate in the mitochondria of cells,and can reduce cellular damage caused by oxidative stress.Considering that the occurrence of DBD is largely caused by oxidative stress injury,we hypothesized that IR-61 with potential antioxidant effect could ameliorate DBD by regulating the oxidative stress level of bladder tissue and cells in the hyperglycemic environment and reducing their damage to bladder cells and tissues.In the present study,we investigated the effect of IR-61 on bladder function in streptozotocin(STZ)-induced diabetic rats to determine whether it could alleviate DBD and the possible mechanism of action.Materials and Methods: Female diabetic rat models were induced with STZ and divided into control group,diabetic group and IR-61 treatment group by intraperitoneal injection of IR-61 solution or blank vehicle,respectively.First,we examined the accumulation level of IR-61 in vital organs of diabetic rats by near-infrared fluorescence imaging as well as its tissue localization and subcellular organelle localization by tissue frozen section and cell immunofluorescence staining under confocal microscopy.Then the ameliorative effect of IR-61 on bladder function in diabetic rats was evaluated by filling cystometry.Finally,the bladder tissues of rats in each group were isolated for near-infrared fluorescence imaging,tissue transmission electron microscopy(TEM),tissue section staining and tissue Western blot(WB)detection and analysis of the expression of related protein molecules.RESULTS: NIR fluorescence imaging of diabetic rats showed that IR-61 could highly accumulate in the bladder of diabetic rats,and confocal images demonstrated that it was mainly localized on the mitochondria of BSMCs.Filling cystometry demonstrated that IR-61 could significantly improve voiding function in diabetic rats.The results of histomorphological experiments showed that IR-61 effectively alleviated the pathological damage of bladder smooth muscle(BSM)in diabetic rats.In addition,IR-61 significantly reduced the apoptotic number of BSMCs and the unfavorable expression of mitochondrial apoptotic pathway-related proteins(B lymphocytoma-2 gene,Bcl-2;BCL2-associated X protein,BAX;cytochrome c,Cytochrome C;cleaved cysteine-containing aspartate proteolytic enzyme 9,cleaved-caspase-9)in diabetic rats.At the same time,frozen section staining and tissue transmission electron microscopy results demonstrated that IR-61 significantly reduced reactive oxygen species(ROS)levels in the BSM of diabetic rats and improved the reduction in the number of mitochondria and morphological pathological changes in the BSMCs of diabetic rats.In addition,WB experiments revealed that IR-61 significantly up-regulated the expression levels of nuclear factor 2-related factor 2(Nrf2)and its related antioxidant proteins in BSM of diabetic rats.Together,these results suggest that IR-61 can improve voiding function in DBD rats by protecting the mitochondria of BSMCs from oxidative stress,and this protective effect may be mediated by activation of the Nrf2 pathway.Notably,IR-61 did not improve body weight and blood glucose levels in diabetic rats.CONCLUSION:1.By intraperitoneal injection,IR-61 can preferentially accumulate in the bladder tissue of diabetic rats and localized on the mitochondria of BSMCs..2.IR-61 significantly improved bladder smooth muscle tissue damage and voiding function in diabetic rats,and this protective effect was not achieved by improving body weight and blood glucose levels in diabetic rats.3.IR-61 may protect the mitochondria of BSMCs from oxidative stress damage and inhibit their occurrence of apoptosis by activating the Nrf2 pathway,suggesting that this may be a potential mechanism by which IR-61 protects or slows down the development of DBD in diabetic rats.