本文選題：睪丸 + 缺血再灌注��； 參考：《中國人民解放軍醫(yī)學(xué)院》2015年博士論文

【摘要】：目的基于睪丸扭轉(zhuǎn)建立兔子睪丸缺血再灌注損傷模型,結(jié)合氧化應(yīng)激指標(biāo)及細(xì)胞凋亡因子檢測(cè)探索睪丸缺血預(yù)處理、后處理及聯(lián)合處理對(duì)睪丸缺血再灌注損傷的保護(hù)作用及潛在調(diào)控機(jī)理。方法選擇雄性新西蘭大白兔30只,隨機(jī)分為5個(gè)組：A組：對(duì)照組,暴露右側(cè)精索,不作缺血處理并關(guān)閉；B組：缺血再灌注組(I/R),使用無創(chuàng)血管夾閉右側(cè)精索致睪丸缺-60min,再灌注3d；C組：預(yù)處理組,缺血再灌注前夾閉精索3次(缺血5min/次+灌注5min/次),夾閉右側(cè)精索致缺血60min,再灌注3d；D組：后處理組,夾閉右側(cè)精索致缺血60min后,進(jìn)行3次缺血再灌注(再灌注5s/次+再缺血5s/次),再灌注3d；E組：聯(lián)合處理組,進(jìn)行缺血再灌注前夾閉精索3次(缺血5min/次+灌注5min/次),夾閉右側(cè)精索致缺血60min后,進(jìn)行3次缺血再灌注(再灌注5s/次+再缺血5s/次),再灌注3d。造模完成后,采用3%巴比妥鈉麻醉兔子后采集血液并分離血清測(cè)定血清睪酮含量；接著分離睪丸組織,按照缺血側(cè)及健側(cè)分組后一部分進(jìn)行裂解后按照MDA、PC、NO, SOD、 MPO、GSH-Px酶聯(lián)免疫吸附檢測(cè)試劑盒所述步驟進(jìn)行檢測(cè)分析表達(dá)水平變化,一部分組織樣本采用Western blot法進(jìn)行檢測(cè)分析Bcl-2,Bax表達(dá)水平,其余組織樣本采用10%中性福爾馬林固定后進(jìn)行HE染色和TUNEL細(xì)胞凋亡檢測(cè)。結(jié)果與正常對(duì)照A組相比,缺血B組、缺血預(yù)處理C組、缺血后處理D組及聯(lián)合處理E組后血清睪酮表達(dá)水平顯著降低(p0.01)；缺血B組、缺血預(yù)處理C組、缺血后處理D組及聯(lián)合處理E組術(shù)前與術(shù)后比較發(fā)現(xiàn),血清睪酮表達(dá)水平明顯降低(p0.05)。與正常對(duì)照A組及缺血B組健側(cè)相比,缺血B組缺血側(cè)MDA、PC、 NO、SOD、MPO、GSH-Px表達(dá)水平顯著升高(p＜0.01)；與缺血B組缺血側(cè)相比,缺血預(yù)處理C組、缺血后處理D組及聯(lián)合處理E組缺血側(cè)MDA、PC、NO、SOD、 MPO、GSH-Px表達(dá)水平顯著降低(p0.01),且三種處理方式之間無統(tǒng)計(jì)學(xué)差異。病理染色結(jié)果顯示,與正常對(duì)照A組及缺血B組健側(cè)相比,缺血B組缺血側(cè)睪丸組織內(nèi)大部分生精小管結(jié)構(gòu)被破壞,生精小管內(nèi)各級(jí)生精細(xì)胞結(jié)構(gòu)消失,可見凋亡的生精細(xì)胞,間質(zhì)及管腔內(nèi)有淡伊紅色水腫液滲出,凋亡指數(shù)明顯升高p0.01),Johnsen評(píng)分顯著降低(p0.01)；與缺血B組缺血側(cè)相比,缺血預(yù)處理C組、缺血后處理D組及聯(lián)合處理E組缺血側(cè)睪丸組織生精小管結(jié)構(gòu)完整,細(xì)胞層次清楚,排列整齊,大部分管腔內(nèi)有精子生成,部分小管管腔內(nèi)有少量淡伊紅色水腫液,凋亡指數(shù)明顯降低(p0.01),Johnsen評(píng)分顯著增加(p0.01),且三種處理方式之間無統(tǒng)計(jì)學(xué)差異。同樣地,與正常對(duì)照A組和缺血B組健側(cè)相比,缺血B組缺血側(cè)Bcl-2/Bax比值顯著降低(p0.01)；與缺血B組缺血側(cè)相比,缺血預(yù)處理C組、缺血后處理D組及聯(lián)合處理E組缺血側(cè)Bcl-2/Bax比值明顯增加并趨近正常水平p0.01)。結(jié)論缺血預(yù)處理、后處理及聯(lián)合處理可通過調(diào)控氧化應(yīng)激指標(biāo)及細(xì)胞凋亡水平明顯改善兔睪丸缺血再灌注損傷,且三種處理方式對(duì)睪丸缺血再灌注損傷的保護(hù)作用相同。不僅為睪丸缺血再灌注損傷的潛在保護(hù)機(jī)理研究提供了重要方法參考,而且為睪丸缺血再灌注損傷的臨床治療提供了潛在開發(fā)藥物,具有重要的臨床應(yīng)用價(jià)值。
[Abstract]:Objective to establish a rabbit testis ischemia-reperfusion injury model based on testicular torsion and to explore the protective effect and potential regulation mechanism of testis ischemic preconditioning, post-processing and combined treatment on testis ischemia reperfusion injury combined with oxidative stress index and apoptosis factor. Methods 30 male New Zealand white rabbits were selected and divided into 5 randomly selected rabbits. Group A: the control group, the control group, exposed to the right spermatic cord, without ischemic treatment and closure; group B: ischemia reperfusion group (I/R), using noninvasive blood vessels to close the right spermatic cord to -60min and reperfusion 3D; group C: preconditioning group, 3 times before ischemia and reperfusion (ischemia 5min/ + perfusion 5min/), and clipping the right spermatic cord ischemia 60min, reperfusion Group 3D, group D: post treatment group, after occlusion of the right spermatic cord ischemia 60min, 3 times of ischemia reperfusion (reperfusion 5s/ + re ischemia 5s/), and reperfusion 3D, E group: combined treatment group, before ischemia-reperfusion of the clipping spermatic cord (ischemia 5min/ + perfusion 5min/), clipping the right spermatic ischemia 60min, 3 times ischemia reperfusion (RE) After perfusion of 5s/ + ischemia 5s/) and reperfusion of 3D. after reperfusion, the blood was collected and serum testosterone content was measured by 3% barbiturate sodium anaesthetized rabbits. Then the testicular tissue was separated and a part of the ischemic side and the healthy side were divided according to the MDA, PC, NO, SOD, MPO, and GSH-Px enzyme immunoadsorption kit. Western blot method was used to detect the expression level of Bcl-2 and Bax in some tissue samples. The other tissue samples were stained with 10% neutral formalin for HE staining and TUNEL cell apoptosis detection. The results were compared with the normal control A group, ischemic B group, ischemic preconditioning C group, and post ischemic place. Serum testosterone expression levels in group D and group E were significantly decreased (P0.01); ischemic B group, ischemic preconditioning C group, ischemic postconditioning group D group and combined treatment group E group before and after operation showed that serum testosterone expression level decreased significantly (P0.05). The expression level of GSH-Px was significantly higher (P < 0.01). Compared with ischemic side of ischemic B group, ischemic preconditioning C group, D group after ischemic post-processing and combined treatment of E group MDA, PC, NO, SOD, MPO, GSH-Px expression level were significantly reduced, and the three treatments were no difference between the methods. Pathological staining results showed that with normal control group and ischemic group In the ischemic B group, most of the spermatogenic tubule structure in the ischemic testicular tissue was destroyed. The structure of spermatogenic cells in the seminiferous tubules disappeared, the apoptotic spermatogenic cells were seen, the interstitial and the endoplasma and the oedema fluid exuded in the lumen, the apoptosis index was significantly increased by P0.01), the Johnsen score was significantly decreased (P0.01), and the ischemic side of the ischemic B group was in the ischemic side phase. Compared with the ischemic preconditioning group C, the structure of the seminiferous tubules in the ischemia side testis tissue of group D and E group was complete, the cell level was clear and the arrangement was neat, most of the lumen had spermatogenesis, a small amount of light red edema was found in the lumen of the part of the tubules, the apoptosis index decreased significantly (P0.01), and the Johnsen score increased significantly (P0.01), and three Compared with the normal control group A and the ischemic B group, the ischemic side Bcl-2/Bax ratio in the ischemic B group was significantly lower than that in the ischemic B group (P0.01), and compared with the ischemic side of the ischemic group, the ischemic preconditioning C group, the ischemic postconditioning group and the combined treatment of the E group were significantly increased and approached the normal level of the normal level in the E group. 1) conclusion. Conclusion ischemic preconditioning, post-processing and combined treatment can obviously improve the ischemia reperfusion injury of rabbit testis by regulating the oxidative stress index and the level of cell apoptosis, and the protective effect of the three treatments on testis ischemia-reperfusion injury is the same. It not only provides an important study on the potential protection mechanism of testis ischemia reperfusion injury. This method provides a potential development medicine for clinical treatment of testicular ischemia reperfusion injury, and has important clinical application value.

【學(xué)位授予單位】：中國人民解放軍醫(yī)學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R697.22

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