本文選題：精索靜脈曲張 + 精子特異性鈣通道　； 參考：《青島大學(xué)》2016年博士論文

【摘要】：第一章精索靜脈曲張患者精子Cat Sper1的差異表達目的:通過檢測精索靜脈曲張患者成熟精子特異性鈣通道Cat Sper1的表達,探討Cat Sper1與精索靜脈曲張之間的關(guān)系,為精索靜脈曲張導(dǎo)致不育的發(fā)病機制提供新的研究方向。方法:健康志愿者20例,精索靜脈曲張患者110例。通過手淫法獲取精液,進行計算機精子運動軌跡分析,應(yīng)用Percoll非連續(xù)梯度離心法對精液進行優(yōu)化分離,應(yīng)用半定量逆轉(zhuǎn)錄酶-聚合酶鏈式反應(yīng)(RT-PCR)以及蛋白質(zhì)印跡法(Western Blot)分別檢測精子中Cat Sper1 m RNA及蛋白表達情況。結(jié)果:與對照組比較,精索靜脈曲張組精液量、液化時間無明顯差異(P0.05),而精子快速前向運動以及前向運動比率(a+b(%))、精子活率以及密度均明顯降低,差異具有統(tǒng)計學(xué)意義(P0.01)。根據(jù)精子運動活力分組,精索靜脈曲張精子活力異常組(3組)Cat Sper1 m RNA及蛋白表達含量均低于對照組(1組)以及精索靜脈曲張精子活力正常組(2組)(P0.01)。與1組比較,2組精子Cat Sper1 m RNA及蛋白表達輕微下降,但差異無統(tǒng)計學(xué)意義(P0.05)。根據(jù)精索靜脈曲張臨床程度分組,精索靜脈曲張臨床I、II、III度各組精子Cat Sper1 m RNA及蛋白表達含量差異不明顯(P0.05)。精索靜脈曲張單側(cè)發(fā)病與雙側(cè)發(fā)病精子Cat Sper1 m RNA及蛋白表達含量差異不明顯(P0.05)。根據(jù)精索靜脈曲張患者靜脈是否存在返流分組,返流組精子Cat Sper1 m RNA及蛋白表達含量均低于對照組以及無返流組(P0.01)。與對照組比較,無返流組精子Cat Sper1 m RNA及蛋白表達差異不明顯(P0.05)。結(jié)論:精子特異性鈣通道Cat Sper1 m RNA及蛋白在精索靜脈曲張患者精子細胞中表達下降,其可能為精索靜脈曲張導(dǎo)致不育的“二次打擊”靶點之一。精子活力低下以及靜脈血液返流與Cat Sper1的表達下降存在一定相關(guān)性,這將為精索靜脈曲張的診斷及治療提供新的思路。第二章CatSper1在實驗性精索靜脈曲張大鼠精子中的差異表達及左卡尼汀對其的保護性研究目的:檢測精索靜脈曲張大鼠附睪精子細胞精子特異性鈣通道Cat Sper1的表達情況,觀察左卡尼汀對大鼠精子Cat Sper1的影響。方法:70只雄性大鼠被隨機分為7組,每組10只。分別為對照組(A組),精索靜脈曲張組(B組),精索靜脈曲張+生理鹽水組(C組),精索靜脈曲張+小、中、大劑量左卡尼汀組(D,E,F組)和F+延長喂養(yǎng)14d組(G組)。手術(shù)結(jié)扎部分左腎靜脈制備精索靜脈曲張大鼠模型。12周后,C組給予生理鹽水1 ml/d,D、E、F組分別用左卡尼汀小(0.05g·kg-1·d-1)、中(0.1 g·kg-1·d-1)、大(0.2 g·kg-1·d-1)劑量灌胃35d,G組在F組基礎(chǔ)上延長灌胃14d,處死大鼠。進行精液參數(shù)精子運動軌跡分析,應(yīng)用RT-PCR以及Western Blot分別檢測精液中Cat Sper1 m RNA及蛋白表達情況。結(jié)果:與A組比較,B組精子a+b(%)、精子活率及密度均不同程度下降(P0.01),B組精子Cat Sper1 m RNA及蛋白表達含量明顯下降(1.44±0.67 VS 0.71±0.38;1.87±0.67 VS 0.84±0.42,P0.01)。與C組比較,應(yīng)用左卡尼汀后,各組精子密度無明顯增加,精子a+b(%)、精子活率以及精子Cat Sper1 m RNA及蛋白表達含量明顯增高,尤其大劑量組F、G組增高更加明顯(P0.01)。與F組比較,G組延長應(yīng)用2周左卡尼汀,精子Cat Sper1 m RNA及蛋白表達量無明顯增加(P0.05)。結(jié)論:精子特異性鈣通道Cat Sper1在精索靜脈曲張大鼠精子細胞中表達下降。應(yīng)用左卡尼汀后,Cat Sper1的表達含量及精子a+b(%)及精子活率增加,從而提高精索靜脈曲張患者的生育能力。第三章Cat Sper1對精子線粒體呼吸與能量代謝的影響目的:應(yīng)用Cat Sper1多克隆抗體抑制Cat Sper1的功能,檢測精子線粒體呼吸功能及能量合成能力,觀察Cat Sper1對精子線粒體呼吸與能量代謝的影響。方法:健康志愿者20例,手淫法獲取精液,檢測精液參數(shù)均符合WHO標準。經(jīng)過上游法處理后每份精液分為A、B兩組,分別與Earles液以及50μg/ml抗Cat Sper1多克隆抗體共孵育。在1h,2h,4h后分別檢測兩組精子精液參數(shù)、細胞線粒體呼吸控制率RCR及精子細胞ATP含量。結(jié)果:與A組比較,B組精子各時間點a+b(%)尤其是a(%)均明顯下降(P0.01);1h后B組精子a(%)即明顯下降,2h,4h后精子下降緩慢,與1h相比無明顯差異(P0.05)。與A組比較,B組精子各時間點態(tài)3值、呼吸控制率RCR以及精子細胞ATP含量均明顯下降(P0.01);B組精子中,2h、4h節(jié)點精子態(tài)3值、呼吸控制率RCR以及精子細胞ATP含量與1h無明顯差異(P0.05)。結(jié)論:阻斷精子特異性鈣通道Cat Sper1,精子細胞線粒體呼吸功能降低,精子生成ATP的能力下降,精子活力降低。這將為精索靜脈曲張引起精子Cat Sper1表達下降,從而導(dǎo)致不育的機制尋找可能的理論依據(jù)。
[Abstract]:Chapter 1 the differential expression of Cat Sper1 in spermatozoa in patients with varicocele Objective: To explore the relationship between Cat Sper1 and varicocele by detecting the expression of specific calcium channel Cat Sper1 of spermatozoa in patients with varicocele, and to provide a new direction for the pathogenesis of sterility caused by varicocele. Method: healthy volunteers In 20 cases and 110 cases of varicocele, spermatozoa were obtained by masturbation, and the sperm movement trajectory was analyzed. The semen was optimized by Percoll discontinuous gradient centrifugation. Semi quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and protein imprinting (Western Blot) were used to detect Cat Sper1 in sperm respectively. M RNA and protein expression. Results: compared with the control group, there was no significant difference in semen volume and liquefaction time in the varicocele group (P0.05), but the rate of sperm motility and forward movement (a+b (%)), sperm motility and density decreased significantly (P0.01). According to sperm motility group, spermatic vein The expression of Cat Sper1 m RNA and protein in the abnormal sperm motility group (3 groups) were lower than the control group (1 groups) and the normal group of spermatic vein varicose sperm (2 groups) (P0.01). Compared with the 1 group, the expression of Cat Sper1 m RNA and protein expression in the 2 groups decreased slightly, but the difference was not significant (P0.05). According to the clinical degree of varicocele, the group of spermatozoa was divided into groups, The difference of Cat Sper1 m RNA and protein expression in spermatozoa was not significant (P0.05) in I, II, and III degree of spermatic varicocele. The difference of M RNA and protein expression in spermatic varicocele unilateral and bilateral sperm Cat Sper1 m RNA and protein expression was not significant. The expression of R1 m RNA and protein were lower than that in the control group and no reflux group (P0.01). Compared with the control group, there was no significant difference in the expression of Cat Sper1 m RNA and protein in the non reflux group (P0.05). Conclusion: sperm specific calcium channel Cat Sper1 m and protein expression in spermatozoa of patients with varicocele may be spermatic vein One of the "two hits" target of infertility is one of the targets of "two hits". There is a certain correlation between low sperm motility and venous blood reflux and the decrease of expression of Cat Sper1. This will provide new ideas for the diagnosis and treatment of varicocele. The differential expression of CatSper1 in experimental spermatic varicocele spermatozoa and Levocarnitine in experimental spermatic vein second The purpose of the study was to detect the expression of sperm specific calcium channel Cat Sper1 in the epididymal spermatozoa of spermatic varicocele rats, and to observe the effect of levocarnitine on Cat Sper1 in rat sperm. Methods: 70 male rats were randomly divided into 7 groups, 10 in each group, respectively, in the control group (group A), the varicocele group (group B) and spermatic cord static. Pulse varicose + physiological saline group (group C), varicocele + small, medium, high dose levocarnitine group (D, E, F) and F+ lengthening 14d group (group G). After ligating part of the left renal vein to prepare the varicocele rat model of the left renal vein for.12 weeks, group C was given 1 ml/d, D, E. ), large (0.2 g. Kg-1. D-1) dose gavage 35d. Group G extended 14d in group F on the basis of group F and killed rats. The sperm motility trajectory of semen was analyzed. RT-PCR and Western Blot were used to detect Cat Sper1 and protein expression in semen. (P0.01) the expression of Cat Sper1 m RNA and protein in group B decreased significantly (1.44 + 0.67 VS 0.71 + 0.38; 1.87 + 0.67 VS 0.84 + 0.42, P0.01). Compared with the C group, the sperm density was not significantly increased, sperm a+b (%), sperm viability, and spermatozoon Cat and protein expression levels were significantly increased, especially in large dose groups The increase in the G group was more obvious (P0.01). Compared with the F group, the expression of Cat Sper1 m RNA and protein in the sperm was not significantly increased in the G group for 2 weeks. Conclusion: the expression of the sperm specific calcium channel Cat Sper1 in the spermatozoa of the varicocele rat was decreased. The sperm viability increased, thus improving the fertility of the patients with varicocele. Third the effects of Cat Sper1 on the mitochondrial respiration and energy metabolism of spermatozoa aim: to use the Cat Sper1 polyclonal antibody to inhibit the function of Cat Sper1, to detect the mitochondrial respiratory function and energy synthesis ability, and to observe the mitochondrial respiration and energy of the sperm of Cat Sper1. Methods: 20 healthy volunteers, 20 cases of masturbation, the semen were obtained and the semen parameters were all conformed to the standard of WHO. After the upstream process, each semen was divided into A, B two, and Earles solution and 50 mu g/ml anti Cat Sper1 polyclonal antibody respectively. After 1h, 2h and 4H respectively, the parameters of sperm semen were detected in two groups, and the cell mitochondrial respiration was detected respectively. Control rate RCR and sperm cell ATP content. Results: compared with group A, a+b (%), especially a (%) in each time point of group B decreased significantly (P0.01), and a (%) of sperm in group B decreased obviously after 1h, 2h, and the sperm decreased slowly after 4H. Compared with the group, the 3 value of each time point of sperm, respiratory control rate and sperm cells were compared with those of the group. The content of TP decreased significantly (P0.01), in group B, the sperm state of 2h, 4H node 3, respiratory control rate RCR and sperm cell ATP content were not significantly different from 1H (P0.05). Conclusion: blocking spermatozoa specific calcium channel Cat Sper1, sperm cell mitochondrial respiratory function decreased, spermatogenesis ATP capacity decreased, sperm vitality decreased. This will be spermatocele. The mechanism of Cat Sper1 expression in the spermatozoa decreased, resulting in the mechanism of infertility.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：R697.24

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2 聶宇星;顯微鏡下精索靜脈曲張低位結(jié)扎術(shù)與傳統(tǒng)精索靜脈高位結(jié)扎手術(shù)預(yù)后對比與分析[D];寧夏醫(yī)科大學(xué);2015年

3 卞建強;精索靜脈曲張三種微創(chuàng)治療方法的對比觀察[D];新鄉(xiāng)醫(yī)學(xué)院;2015年

4 柯鑫文;茶多酚對精索靜脈曲張大鼠睪丸組織氧化應(yīng)激和細胞凋亡的影響[D];山西醫(yī)科大學(xué);2015年

5 武政華;茶多酚對精索靜脈曲張大鼠睪丸生精細胞凋亡的影響[D];山西醫(yī)科大學(xué);2015年

6 孫廣海;精索靜脈曲張對青年影響及治療效果研究[D];安徽醫(yī)科大學(xué);2015年

7 陳喬;毓麟茶對精索靜脈曲張不育癥（腎虛血瘀型）精液質(zhì)量的影響[D];安徽中醫(yī)藥大學(xué);2015年

8 李勉洲;精索靜脈曲張患者精漿蛋白質(zhì)組學(xué)的初步研究[D];青島大學(xué);2015年

9 李靜;顯微鏡與腹腔鏡手術(shù)治療精索靜脈曲張療效與安全性的試驗序貫Meta分析[D];山西醫(yī)科大學(xué);2016年

10 徐順龍;顯微鏡下外環(huán)下精索靜脈結(jié)扎術(shù)中提出與不提出睪丸的對比研究[D];吉林大學(xué);2016年

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