本文選題：特發(fā)性非梗阻性無(wú)精子癥 + 缺氧誘導(dǎo)因子α　； 參考：《青島大學(xué)》2017年博士論文

【摘要】：研究目的:特發(fā)性非梗阻性無(wú)精子癥(idiopathic azoospermia,IA)是男性不育中病因不明、治療困難的一種病癥,主要病理機(jī)制是睪丸生精功能障礙,一直是該領(lǐng)域的研究熱點(diǎn)。缺氧誘導(dǎo)因子(hypoxia-inducible factors,HIFs)是一類(lèi)在氧穩(wěn)態(tài)失衡狀態(tài)下激活的重要轉(zhuǎn)錄因子,而睪丸在正常生理狀態(tài)下系低氧器官,有研究顯示,在HIF-α基因敲除的小鼠中會(huì)出現(xiàn)睪丸生精功能衰竭。本研究擬探討HIF-α在IA患者睪丸生精功能障礙中的作用。研究方法:本研究按照WHO標(biāo)準(zhǔn)的診斷流程進(jìn)行病史采集和各項(xiàng)檢查,制定嚴(yán)格的入組篩選流程,共募集30例IA患者作為實(shí)驗(yàn)組,30例OA患者作為對(duì)照組,所有入組患者均實(shí)施經(jīng)皮睪丸穿刺抽吸術(shù)(testicular sperm aspiration,TESA)。觀察TESA標(biāo)本HE染色病理變化,分別計(jì)數(shù)實(shí)驗(yàn)組和對(duì)照組的睪丸支持細(xì)胞,并應(yīng)用免疫組化方法檢測(cè)支持細(xì)胞異常增生情況。應(yīng)用western blotting方法檢測(cè)兩組TESA標(biāo)本的抗氧化蛋白超氧化物歧化酶(Superoxide Dismutase,SOD)、血紅素加氧酶(Hemeoxygenase-1,HO-1)、HIF-1α/2α、緊密連接蛋白Zonula occluden-1(ZO-1)、Claudin和Occludin的表達(dá)變化,并進(jìn)一步檢測(cè)HIF-α的靶點(diǎn)調(diào)節(jié)分子鐵代謝相關(guān)蛋白二價(jià)金屬離子轉(zhuǎn)運(yùn)體1(Divalent metal transporter 1,DMT1)、ferroportin1(FPN1)和鐵調(diào)節(jié)蛋白1(Iron regulatory protein1,IRP1)的表達(dá)變化。然后在體外培養(yǎng)的小鼠睪丸支持細(xì)胞(TM4)系中,應(yīng)用western blotting方法檢測(cè)不同濃度雙酚A、氮芥處理后上述蛋白的表達(dá)變化,并評(píng)價(jià)了抗氧化劑姜黃素和N-乙酰半胱氨酸(Nacetyl-L-cysteine,NAC)的保護(hù)作用。研究結(jié)果:1、參考WHO 2000年《男性不育標(biāo)準(zhǔn)化診療手冊(cè)》(2000 WHO Manual for the Standardized Investigation,Diagnosis and Management of the Infertile Male)所述標(biāo)準(zhǔn)的OA和IA診斷流程,經(jīng)過(guò)嚴(yán)格篩選,對(duì)招募的220位無(wú)精子癥志愿者進(jìn)行分類(lèi),將符合上述分類(lèi)診斷標(biāo)準(zhǔn)者分別納入實(shí)驗(yàn)組(IA組,共30例)和對(duì)照組(OA組,共30例)。兩組病人在年齡、體重指數(shù)、就診時(shí)不育時(shí)間方面均無(wú)統(tǒng)計(jì)學(xué)差異。2、IA和OA組患者病因分析表明,IA組病人均無(wú)明確病因,且無(wú)其他適用的疾病診斷,為特發(fā)性無(wú)精子癥;OA組病人無(wú)精子癥病因則包括附睪炎、附睪結(jié)核、先天性發(fā)育異常、醫(yī)源性病因等多種類(lèi)型。3、IA和OA組患者體格檢查表明:兩組患者全面體格檢查未發(fā)現(xiàn)明顯異常,第二性征發(fā)育良好,陰莖發(fā)育正常;IA組30例患者雙側(cè)睪丸、附睪均可于陰囊內(nèi)捫及,但I(xiàn)A組患者睪丸容積較OA組患者變小,且質(zhì)地明顯變軟,OA組患者雙側(cè)睪丸查體未及異常,所有OA患者的附睪均可捫及明顯膨脹、飽滿,另有5例OA患者未能觸及輸精管結(jié)構(gòu)。陰囊超聲檢查輔助估算睪丸容積大小,IA組患者睪丸容積明顯減小,僅不到OA組患者睪丸容積的一半(6.2m L vs.15.1m L,P0.001)。4、血清卵泡刺激素(Follicle Stimulating Factor,FSH)、黃體生成素(luteinizing hormone,LH)、催乳素(Prolactin,PRL)和睪酮(Testosterone,TT)水平檢測(cè)表明:IA組患者血清FSH(16.40±5.60 IU/L vs.3.64±1.53 IU/L,P0.001)、LH(10.48±2.20 IU/L vs.4.49±1.59 IU/L,P0.001)水平較OA組均顯著升高,而IA和OA兩組患者的血清PRL(10.59±2.00 IU/L vs.10.55±1.91 IU/L,P0.05)、TT(11.33±2.26 nmol/L vs.10.65±2.10 nmol/L,P0.05)水平無(wú)明顯差異。5、IA組和OA組患者睪丸穿刺組織的HE染色病理分析表明:IA組典型的病理改變包括:(1)生精小管內(nèi)生殖細(xì)胞數(shù)量顯著減少,且往往精子發(fā)生紊亂,幾乎不見(jiàn)成熟的精子細(xì)胞;(2)生精小管腔內(nèi)充滿脫落的細(xì)胞,含有少量生殖細(xì)胞的生精小管與僅有支持細(xì)胞的生精小管同時(shí)存在,支持細(xì)胞數(shù)量明顯增多;(3)生精小管管腔明顯縮小;(4)睪丸間質(zhì)細(xì)胞未見(jiàn)明顯異常改變。OA組患者睪丸穿刺組織的典型病理改變包括:(1)精子發(fā)生的各個(gè)階段都存在,可見(jiàn)成熟的精子細(xì)胞,但各階段生殖細(xì)胞正常的排列順序消失;(2)生精小管結(jié)構(gòu)、管腔直徑正常,但是中心腔消失并充滿脫落的細(xì)胞;(3)睪丸間質(zhì)細(xì)胞未見(jiàn)明顯異常改變。6、IA組和OA組患者睪丸穿刺組織中支持細(xì)胞計(jì)數(shù)表明:IA組生精小管每單位橫截面的睪丸支持細(xì)胞數(shù)量要明顯多于OA組(38.0±3.5 vs.19.7±2.6,P0.001),IA組生精小管周長(zhǎng)與OA組無(wú)明顯差異(422.15±18.33μm vs.433.17±25.61μm,P0.05);IA組患者睪丸組織中的支持細(xì)胞分布密度(Sertoli cell density,SCD)顯著高于OA組(0.093±0.008 vs.0.044±0.005,P0.001)。7、IA組和OA組患者睪丸支持細(xì)胞增殖活性評(píng)價(jià):IA組和OA組的睪丸支持細(xì)胞均呈現(xiàn)胞漿內(nèi)支持細(xì)胞標(biāo)記物抑制素α(Inhibinα)染色強(qiáng)陽(yáng)性,而且在連續(xù)組織切片上,可觀察到部分睪丸支持細(xì)胞呈現(xiàn)細(xì)胞核細(xì)胞增殖活性標(biāo)記物Ki-67染色強(qiáng)陽(yáng)性;進(jìn)一步計(jì)算Ki-67陽(yáng)性指數(shù)(Ki-67 Labelling Index,Ki-67 LI),結(jié)果提示IA組睪丸支持細(xì)胞的Ki-67 LI高于OA組3倍以上(48%±6%vs.13%±5%,P0.001)。8、免疫印記法檢測(cè)IA組和OA組患者睪丸穿刺組織中應(yīng)激反應(yīng)蛋白HO-1的表達(dá)變化,結(jié)果提示IA組與OA組之間無(wú)明顯變化;但I(xiàn)A組的抗氧化蛋白SOD表達(dá)水平明顯下降,IA組比OA組降低了75.2%±16.2%(P0.05)。提示IA患者睪丸組織內(nèi)存在著抗氧化能力不足。9、免疫印記法檢測(cè)IA組和OA組患者睪丸穿刺組織中HIF-1α的表達(dá)變化:IA組明顯上調(diào),升高達(dá)OA組的2.44±0.94倍(P0.01);而IA組的HIF-2α表達(dá)水平顯著下調(diào),比OA組降低了49%±11.3%(P0.01)。10、免疫印記法檢測(cè)IA組和OA組患者睪丸穿刺組織中鐵代謝相關(guān)蛋白的表達(dá)變化:IA組的鐵轉(zhuǎn)入蛋白DMT1+IRE表達(dá)水平要比OA組高出29%±8.3%(P0.05);IA組的鐵轉(zhuǎn)出蛋白FPN1表達(dá)水平相對(duì)比OA組存在升高趨勢(shì),但是兩組間FPN1的表達(dá)水平差異無(wú)統(tǒng)計(jì)學(xué)意義;鐵調(diào)節(jié)蛋白IRP1在IA組患者睪丸穿刺組織中表達(dá)水平較OA組升高22%±5.2%(P0.05)。11、免疫印記法檢測(cè)IA組和OA組患者睪丸穿刺組織中緊密連接蛋白的表達(dá)變化:IA組ZO-1和Claudin蛋白表達(dá)水平較OA組均顯著降低,分別為OA組的62.3%和43.5%(P0.05);Occludin蛋白表達(dá)水平降低最明顯,IA組只有OA組的7.2%(P0.05)。12、體外培養(yǎng)小鼠睪丸支持細(xì)胞(TM4)系,分別應(yīng)用不同濃度(10-7、10-6、10-5、10-4、10-3mol/L)的雙酚A(Bisphenol A,BPA)和氮芥(nitrogen mustard,NM)處理后,在藥物濃度低于10-5mol/L時(shí)TM4細(xì)胞存活率均沒(méi)有明顯變化,但在藥物濃度高于10-4mol/L時(shí)TM4細(xì)胞存活率明顯下降,BPA處理組TM4細(xì)胞存活率分別僅為對(duì)照組的57.8%、24.9%,氮芥處理組分別是對(duì)照組的62.2%、38.6%(P0.05)。13、BPA和氮芥分別處理TM4細(xì)胞后,免疫印跡法觀察HIF-1α和HIF-2α的表達(dá)變化:較低濃度10-6mol/L BPA處理TM4細(xì)胞24h后,BPA處理組細(xì)胞裂解液中的HIF-2α表達(dá)水平明顯上調(diào),是對(duì)照組的1.41倍(P0.05),而HIF-1α的表達(dá)有升高趨勢(shì),但無(wú)統(tǒng)計(jì)學(xué)意義;10-6mol/L氮芥處理TM4細(xì)胞24h后,氮芥處理組的HIF-1α和HIF-2α表達(dá)水平均明顯上調(diào),分別是對(duì)照組的2.71倍和1.72倍(P0.05)。較高濃度10-5mol/L BPA處理TM4細(xì)胞24h后,BPA處理組細(xì)胞裂解液中的HIF-1α和HIF-2α表達(dá)水平均明顯下調(diào),分別僅為對(duì)照組的39%和32.2%(P0.05);較高濃度10-5mol/L氮芥處理TM4細(xì)胞24h后,氮芥處理組的HIF-1α和HIF-2α表達(dá)水平也均明顯下調(diào),分別為對(duì)照組的33.6%和37.7%(P0.05)。14、BPA和氮芥分別處理TM4細(xì)胞后,免疫印跡法觀察鐵代謝蛋白的表達(dá)變化:10-6mol/L和10-5mol/L BPA處理TM4細(xì)胞24h后,BPA處理組細(xì)胞裂解液中鐵轉(zhuǎn)入蛋白DMT1+IRE表達(dá)水平均明顯上調(diào),分別為對(duì)照組的1.83倍和1.72倍(P0.05);10-6mol/L和10-5mol/L氮芥處理TM4細(xì)胞24h后,氮芥處理組細(xì)胞裂解液中的DMT1+IRE表達(dá)水平也分別上調(diào)1.75倍和1.82倍(P0.05)。上述不同濃度的BPA和氮芥分別處理TM4細(xì)胞24h后,藥物處理組的鐵轉(zhuǎn)出蛋白FPN1表達(dá)水平雖有上升趨勢(shì),但差別無(wú)統(tǒng)計(jì)學(xué)意義。鐵調(diào)節(jié)蛋白IRP1在BPA和氮芥處理組表達(dá)水平均明顯上調(diào),10-6mol/L和10-5mol/L BPA處理組的IRP1表達(dá)水平分別是對(duì)照組的1.92倍和1.5倍(P0.05),10-6mol/L和10-5mol/L氮芥處理組的IRP1表達(dá)水平分別是對(duì)照組的2.28倍和1.87倍(P0.05)。15、10-6mol/L和10-5mol/L BPA或氮芥分別處理TM4細(xì)胞24h后,免疫印跡法檢測(cè)TM4細(xì)胞裂解液中的應(yīng)激反應(yīng)蛋白HO-1和抗氧化蛋白SOD表達(dá)水平均沒(méi)有明顯變化。16、分別用1、2、5、10μmol/L姜黃素處理TM4細(xì)胞24h后,在5μmol/L時(shí)細(xì)胞存活率略有下降,為對(duì)照組的94.5%,但無(wú)統(tǒng)計(jì)學(xué)意義;10μmol/L姜黃素可引起TM4細(xì)胞存活率明顯下降,為對(duì)照組的81.9%(P0.05)。應(yīng)用5μmol/L姜黃素與BPA共孵育,未能觀察到姜黃素對(duì)TM4細(xì)胞存活率的保護(hù)作用;強(qiáng)抗氧化劑N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)與BPA共孵育,也未能觀察到其對(duì)細(xì)胞存活率的保護(hù)作用。研究結(jié)論:1.特發(fā)性非梗阻性無(wú)精子癥患者睪丸穿刺組織中存在支持細(xì)胞異常增生,抗氧化能力下降,提示特發(fā)性非梗阻性無(wú)精子癥患者睪丸支持細(xì)胞存在著數(shù)量和功能異常。2.特發(fā)性非梗阻性無(wú)精子癥患者睪丸穿刺組織中,以及藥物處理后的體外培養(yǎng)小鼠睪丸支持細(xì)胞中,均觀察到了缺氧誘導(dǎo)因子-α蛋白表達(dá)異常。3.缺氧誘導(dǎo)因子α的下游調(diào)控分子緊密連接蛋白和鐵代謝相關(guān)蛋白表達(dá)異常,提示疾病狀態(tài)下血-睪屏障破壞和鐵穩(wěn)態(tài)失衡,這可能是缺氧誘導(dǎo)因子α調(diào)控異常參與特發(fā)性非梗阻性無(wú)精子癥患者睪丸生精功能障礙的發(fā)病機(jī)制。
[Abstract]:Objective: idiopathic non obstructive azoospermia (idiopathic azoospermia, IA) is a disease with unknown etiology and difficult treatment in male infertility. The main pathological mechanism is testicular spermatogenesis dysfunction, which has always been a hot topic in this field. Hypoxia inducible factor (hypoxia-inducible factors, HIFs) is a class of imbalance in oxygen homeostasis. The important transcription factors are activated and the testicles are hypoxic organs in normal physiological state. Studies have shown that testicular spermatogenesis failure in HIF- alpha knockout mice. This study intends to explore the role of HIF- alpha in the Gao Wansheng sperm dysfunction in patients with IA. Methods: This study was conducted in accordance with the diagnostic process of the WHO standard. A total of 30 IA patients were recruited as the experimental group and 30 patients with OA were used as the control group. All the patients were treated with percutaneous testicular puncture aspiration (testicular sperm aspiration, TESA). The pathological changes of HE staining in TESA specimens were observed, and the testicular support of the experimental and control groups was counted. Western blotting method was used to detect the antioxidant protein superoxide dismutase (Superoxide Dismutase, SOD), heme oxygenase (Hemeoxygenase-1, HO-1), HIF-1 alpha /2 alpha, and tightly connexin Zonula occluden-1 (TESA). Expression changes, and further detection of HIF- alpha targets to regulate the expression of molecular iron metabolism related protein two valence metal ion transporter 1 (Divalent metal transporter 1, DMT1), ferroportin1 (FPN1) and iron regulatory protein 1 (Iron regulatory protein1, IRP1), but then in the mouse testis support cells (TM4) in vitro Tern blotting method was used to detect the changes of the expression of the above protein after the treatment of different concentrations of bisphenol A and nitrogen mustard, and the protective effects of the antioxidant curcumin and N- acetylcysteine (Nacetyl-L-cysteine, NAC) were evaluated. 1, reference WHO 2000 < standard treatment manual for male infertility > (2000 WHO Manual for the Standardized) Ion, Diagnosis and Management of the Infertile Male) the standard OA and IA diagnostic process. After strict screening, the recruited 220 azoospermia volunteers were classified into the experimental group (IA group, 30 cases) and the control group (OA group, a total of 30 cases). The two groups of patients were in age, body mass index, There was no statistical difference in the time of infertility at the time of diagnosis.2. The cause analysis of the patients in group IA and OA showed that the patients in group IA had no definite cause, and no other suitable diagnosis of the disease, and it was idiopathic azoospermia. The causes of azoospermia in group OA include epididymitis, epididymal tuberculosis, congenital dysplasia, iatrogenic etiology, and other types of.3, IA, and OA. The physical examination showed that there were no obvious abnormalities in the physical examination of the two groups, the second sex sign developed well, and the penis developed normally. In group IA, 30 cases of testis and epididymis were palpable in the scrotum, but the volume of testicle in group IA was smaller than that in group OA, and the texture of the patients was softer than that in the group of OA. All the patients in group OA had no abnormalities in the bilateral testis body, all O The epididymis of the patients with A could be palpably inflated and plump, and 5 patients with OA failed to touch the vas deferens. The scrotal ultrasonography assisted the estimation of the size of the testis, and the testicular volume in the IA group decreased significantly, only half of the OA patients' testicular volume (6.2m L vs.15.1m L, P0.001).4, and serum follicle stimulating hormone (Follicle Stimulating). FSH), luteinizing hormone (LH), prolactin (Prolactin, PRL) and testosterone (Testosterone, TT) level test showed that the serum FSH (16.40 + 5.60 IU/L vs.3.64 + 1.53) in the IA group was significantly higher than that in the group of two groups (10.59 + 2. (10.59 + 2.). There was no significant difference between 00 IU/L vs.10.55 + 1.91 IU/L, P0.05), TT (11.33 + 2.26 nmol/L vs.10.65 + 2.10 nmol/L, P0.05),.5. The pathological analysis of testicular biopsy in IA group and OA group showed that the typical pathological changes included: (1) the number of germ cells in the seminiferous tubules decreased significantly, and the sperm was often disorganized. Mature spermatids; (2) the cells filled with exfoliated tubules were filled with shedding cells, the seminiferous tubules containing a small amount of germ cells existed at the same time as the seminiferous tubules only supporting cells, and the number of supporting cells increased significantly; (3) the cavity of the spermatogenic tubule was obviously reduced; (4) the testicular cells did not significantly alter the canon of the testicular tissue in the.OA group. Pathological changes include: (1) all stages of spermatogenesis exist, mature spermatocyte can be seen, but the sequence of normal arrangement of germ cells in each stage disappears; (2) spermatogenic tubule structure, the diameter of the lumen is normal, but the central cavity disappears and fills the shedding cells; (3) there is no obvious abnormal change in.6, IA and OA patients. The count of support cells in the pellet puncture tissue showed that the number of testis supporting cells per unit cross section of the IA group was more than that of the OA group (38 + 3.5 vs.19.7 + 2.6, P0.001). There was no significant difference between the perimeter of the seminiferous tubules in the IA group and the OA group (422.15 + 18.33 mu m vs.433.17 + 25.61 m, P0.05), and the support cells in the testicular tissue of the IA group were densely distributed. Sertoli cell density (SCD) was significantly higher than that of OA group (0.093 + 0.008 vs.0.044 + 0.005, P0.001).7, and the proliferation activity of testis support cells in the IA group and OA group was evaluated. Ki-67 positive index (Ki-67 Labelling Index, Ki-67 LI) of Ki-67 positive index (Ki-67 Labelling Index) was further calculated. The results suggested that the Ki-67 LI of testis support cells in IA group was more than 3 times higher than that of OA group (48% + 5%). The expression of stress response protein HO-1 expression in the tissue showed no significant changes between the IA group and the OA group, but the expression level of the antioxidant protein SOD in the IA group decreased obviously, and the IA group decreased by 75.2% + 16.2% (P0.05) than the OA group. It suggested that the testicular tissue in the IA patients was less than the antioxidant capacity.9, and the immunoimprint method was used to detect the testicles of IA group and OA group. The changes in the expression of HIF-1 alpha in the puncture tissue: the IA group was obviously up-regulated and up to 2.44 + 0.94 times (P0.01) in the OA group, while the HIF-2 alpha expression level in the IA group was significantly lower than that in the OA group by 49% + 11.3% (P0.01).10. The expression of iron metabolism related proteins in the testis tissue of the IA and OA groups of the IA and OA groups was detected by the immuno imprint method: the iron transfer of the IA group to the protein The expression level of 1+IRE was 29% + 8.3% higher than that of the OA group (P0.05), and the expression level of iron transfer protein FPN1 in the IA group was higher than that in the OA group, but the expression level of FPN1 in the two groups was not statistically significant; the expression of iron regulatory protein IRP1 in the testis tissue in the IA group was 22% + 5.2% (P0.05).11, immune imprinting method The expression of tight connexin in IA group and OA group was detected. The expression level of ZO-1 and Claudin protein in group IA was significantly lower than that in group OA, 62.3% and 43.5% (P0.05) in group OA, respectively, and the expression level of Occludin protein was the most obvious. The IA group was only 7.2% of OA group (P0.05), and in vitro culture of mouse testis support cells After the treatment of Bisphenol A (Bisphenol A, BPA) and nitrogen mustard (nitrogen mustard, NM) with different concentrations (10-7,10-6,10-5,10-4,10-3mol/L), there was no significant change in the survival rate of TM4 cells when the drug concentration was lower than 10-5mol/L, but the survival rate of TM4 cells decreased significantly when the drug concentration was higher than that of 10-4mol/L. Only 57.8% of the control group, 24.9%, nitrogen mustard treatment group was 62.2% of the control group, 38.6% (P0.05).13, BPA and nitrogen mustard treated TM4 cells respectively. The expression of HIF-1 A and HIF-2 alpha was observed by immunoblotting: after the low concentration 10-6mol/L BPA treated TM4 cell 24h, the expression of HIF-2 alpha in the cell lysate of BPA treatment group was obviously up. The expression of HIF-1 alpha was 1.41 times (P0.05), but the expression of HIF-1 alpha was higher, but it was not statistically significant. After 24h of 10-6mol/L mustard treated TM4 cells, the expression level of HIF-1 alpha and HIF-2 alpha in the treatment group of nitrogen mustard were obviously up, which were 2.71 times and 1.72 times of that of the control group (P0.05). The expression level of HIF-1 alpha and HIF-2 alpha in the liquid was obviously down, only 39% and 32.2% of the control group (P0.05), while the high concentration of 10-5mol/L nitrogen mustard treated TM4 cells 24h, and the expression level of HIF-1 A and HIF-2 alpha in the treatment group of nitrogen mustard were also down significantly, respectively, 33.6% and 37.7% (P0.05).14 of the control group, and BPA and nitrogen mustard treated TM4 cells respectively. The expression of iron metabolic protein was observed by the immunoblotting method: after 10-6mol/L and 10-5mol/L BPA treated TM4 cell 24h, the expression level of iron to protein DMT1+IRE in the cell lysate of BPA treatment group were up to 1.83 and 1.72 times (P0.05), respectively. 10-6mol/L and 10-5mol/ L nitrogen mustard treated the cell cracking of the nitrogen mustard treatment group. The expression level of DMT1+IRE in the liquid was also up up 1.75 and 1.82 times (P0.05). The expression level of iron transferred protein FPN1 in the treatment group was up, but the difference was not statistically significant. The expression level of iron regulatory protein IRP1 in BPA and nitrogen mustard treatment group was up obviously up, 10, 10. The expression level of IRP1 in -6mol/L and 10-5mol/L BPA treatment group was 1.92 times and 1.5 times as much as that of the control group (P0.05). The IRP1 expression level of 10-6mol/L and 10-5mol/L nitrogen mustard treatment group was 2.28 times and 1.87 times (P0.05).15,10-6mol/L and 10-5mol/L BPA or nitrogen mustard respectively. The expression level of stress response protein HO-1 and antioxidant protein SOD in the liquid did not significantly change.16. The cell survival rate of 1,2,5,10 mu mol/L curcumin in TM4 cell 24h decreased slightly at 5 u mol/L, which was 94.5% of the control group, but no statistical significance. The survival rate of TM4 cells decreased significantly by 10 u mol/L curcumin, which was the control group. 81.9% (P0.05). The protective effect of curcumin on the survival rate of TM4 cells was not observed with 5 mu curcumin and BPA, and the strong antioxidant N- acetylcysteine (N-acetyl-L-cysteine, NAC) was incubated with BPA, and the protection of cell survival was not observed. Conclusion: 1. idiopathic non obstructive azoospermia patients There are abnormal proliferation of support cells and decreased antioxidant capacity in the testicular tissue, suggesting that the testicular support cells in the patients with idiopathic non obstructive azoospermia exist in the testicular tissue of the patients with.2. idiopathic non obstructive azoospermia, and the mouse testis support cells in vitro after the drug treatment. The abnormal expression of close connexin and iron metabolism related protein in the downstream regulator of hypoxia inducible factor alpha protein expression.3. hypoxia inducible factor alpha was observed, suggesting the destruction of blood testosterone barrier and iron homeostasis under the condition of disease, which may be the abnormal involvement of hypoxic inducible factor alpha in idiopathic non obstructive azoospermia patients. The pathogenesis of testicular spermatogenesis dysfunction.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R698.2

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