| Research background and purpose:With the rapid development of modern industry,several heavy metal pollutants that directly endanger human health have been found in soil,water,and air.Cadmium(Cd)is widely used in various industrial production,including the manufacturing of nuclear reactors,metal electroplating and welding,mining,and nickel cadmium battery manufacturing.The industrial "three wastes" generated in these industrial production are discharged into the natural environment,causing pollution of water,air,and soil.Cd is considered to be one of the most deadly heavy metal poisons due to its hazardous properties such as high toxicity,transferability,and durability.Reproductive organs such as the uterus,testis,and epididymis are the main target organs for cadmium accumulation in the body.Studies have found that oxidative stress is the main biological mechanism of testicular cadmium damage.Delphinidin(Dp)is a water-soluble natural pigment with multiple biological functions such as antioxidant,anti-inflammatory,and cancer prevention.Studies have confirmed that delphinidin can prevent various oxidative stress induced damage by activating the Nrf2 signaling pathway,but whether delphinidin can alleviate cadmium induced reproductive damage in rats has not been reported.Therefore,the main purpose of this study is to explore the improvement effect of delphinidin on cadmium induced reproductive toxicity in rats and its biological mechanism.Materials and Methods:Twenty four Wistar male rats were chosen as experimental animals in this study.Experiments the rats were randomly divided into four groups and gavaged every day continuously.They were as follows: control group gavaged with normal saline,Dp group gavaged with 400 mg / kg.BW delphinidin solution,Cd group gavaged with 5mg / kg.BW cadmium chloride solution,Dp + Cd group gavaged with 400 mg / kg.BW delphinidin solution + 5 mg / kg.BW cadmium chloride solution,in which the Dp + Cd group was gavaged with cadmium chloride solution one hour after the delphinidin solution was gavaged.A model of subacute poisoning by cadmium in rats was established by continuous administration for 28 days.The body weights were weighed on the 7th,14 th,21st,and 28 th day.Experimental animals euthanasia after sacrifice,body,testis,and epididymis weights were recorded;Blood was collected for four oxidative indicators including propylene glycol(MDA),superoxide dismutase(SOD),glutathione peroxidase(GSH-Px),catalase(CAT)and two sex hormone tests including testosterone(testosterone),follicle stimulating hormone(FSH);One cauda epididymis was used for sperm analysis,and one testis was stored at-70 ° C for glutathione(GSH),two oxidative indicators of total antioxidant capacity(T-AOC)and Western blot analysis of gene protein expression;On the other side,the testis and epididymis were fixed in 4% paraformaldehyde solution and used for subsequent H & E staining of paraffin sections to observe the pathological changes,TUNEL assay and immunohistochemical analysis.Result:1.HE staining observation of paraffin sections showed that the rat testis was severely shrunken under the effect of Cd chloride,the basement membrane and spermatogenic cells were sloughed off in the seminiferous tubules,vacuoles and spermatogenic cell arrangement were appeared disordered,and the number of spermatozoa in the epididymis was reduced.Delphinidin significantly alleviated Cd induced pathological alterations in tissue structure and decreased sperm count.The results of sperm count under the microscope showed that the number of spermatozoa in the epididymis was decreased under the effect of Cd chloride,and this phenomenon was significantly improved by delphinidin.2.Cadmium causes severe oxidative stress,through the detection of indicators of oxidative stress,delphinidin was able to downregulate the increase in MDA caused by cadmium and upregulate five antioxidant indicators: SOD,GSH-Px,T-AOC,CAT,and GSH.3.TUNEL apoptosis assay and immunohistochemical staining for Ki67 protein in testis the results of the experiment showed that Cd caused germ cell apoptosis and inhibited germ cell proliferation.Delphinidin significantly alleviated this spermatogenic failure.4.The results of Western immunoblotting assay showed that cadmium was able to significantly downregulate the expression of Nrf2,HO-1,which were the downstream pathway proteins of Nrf2,indicating that cadmium exposure could inhibit Nrf2 signaling pathway in germ cells.The Dp + Cd group significantly upregulated the protein expression of the related antioxidant genes HO-1 and Nrf2,indicating that delphinidin can activate the nrf2 / HO-1 signaling pathway in the testes and alleviate the toxicity induced by Cd induced oxidative stress in reproductive organs.Conclusion:The results of this study indicate that cadmium can cause reproductive toxicity in rats.In male rats,cadmium causes testicular tissue damage and reduced sperm production.This study confirmed that delphinidin has an improvement effect on cadmium induced reproductive damage,spermatogenesis disorders,and sexual hormone production disorders.Delphinidin significantly improved the pathological characteristics of testicular and epididymal tissues in cadmium poisoned rats.In the model of cadmium induced reproductive toxicity in rats,delphinidin not only improved the inhibition of cadmium on the proliferation of testicular germ cells,but also inhibited the apoptosis of testicular germ cells,which suggests that delphinidin can improve the reproductive damage caused by cadmium;Regulate serum reproductive hormones in cadmium poisoned rats and significantly reverse the decrease in sperm count induced by cadmium,suggesting that delphinidin can improve spermatogenesis caused by cadmium induced sexual hormone production disorders in rats;At the same time,delphinidin upregulates SOD,GSH-PX,T-AOC,CAT,GSH,and downregulates MDA in the redox system,suggesting that delphinidin protects the reproductive system of rats from cadmium damage through antioxidant stress;Delphinidin upregulates the expression of Nrf2 pathway related proteins,suggesting that activating the Nrf2 signaling pathway is a therapeutic target for delphinidin to improve cadmium induced reproductive toxicity.This study provides new directions and ideas for the mechanism research and therapeutic targets of natural plant compounds to improve cadmium induced reproductive damage. |
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