IGF-1在非梗阻性无精子症睾丸组织中的表达

目的 研究胰岛素样生长因子-1(IGF-1)在非梗阻性无精子症睾丸组织中的表达并探讨其意义.方法 采用免疫组化EnVision法对32例非梗阻性无精子症人睾丸组织石蜡标本和13例人正常睾丸组织中的IGF-1的表达进行检测,通过统计学方法 分析其表达情况.结果 非梗阻性无精子症睾丸组织HE染色显示生精小管中无或仅少量精子,生精小管的生精上皮脱落、排列紊乱,生精小管壁部分有玻变萎缩.32例非梗阻性无精子症睾丸组织中IGF-1阳性表达4例,阴性表达28例,阳性率12.5%;13例正常睾丸组织中IGF-1阳性表达10例,阴性表达3例,阳性率76.9%;差异有显著性(P<0.01).结论 (1)IGF-1在睾丸组织中表达减少或缺失与生精功能障碍密切相关.(2)非梗阻性无精子症睾丸组织生精小管的生精上皮脱落、排列紊乱,生精阻滞.     

转录因子E2F1在非梗阻性无精症睾丸组织中的表达

目的旨在探讨E2F1核转录因子在人类睾丸组织中的表达与生精功能的关系。方法选取32例人类非梗阻性无精子症睾丸组织石蜡标本，采用HE染色观察曲细精管病理变化，同时用免疫组化SP法检测睾丸组织中核转录因子E2F1的表达;对照组选取13例人类正常睾丸组织。通过分析病理组HE染色，比较E2F1阳性染色范围及程度，评价E2F1核转录因子与生精功能的关系。结果非梗阻性无精子症睾丸组织HE染色显示精曲小管中无或仅少量精子，精曲小管的生精上皮脱落、排列紊乱，精曲小管壁部分有玻变、纤维变，生精细胞萎缩。32例非梗阻性无精子症睾丸组织中E2F1阳性表达8例，阴性表达24例，阳性率25%;13例正常睾丸组织中E2F1的阳性表达9例，阴性表达4例，阳性率69.23%;x2=7.694，差异有显著性（P〈0.01）。结论（1）E2F1核转录因子表达缺失与睾丸生精功能障碍密切相关;（2）非梗阻性无精子症睾丸组织精曲小管的生精上皮脱落、排列紊乱，生精阻滞。     

无精子症和生精功能正常者睾丸组织中血红素加氧酶的表达与差异性研究

目的:对人类睾丸组织中表达血红素加氧酶(HO)的细胞进行定位;通过测定原发性无精子症及梗阻性无精子症患者睾丸组织中血红素加氧酶1(HO-1)的表达量与正常睾丸组织中HO-1表达量的差异性,来探讨其与无精子症发病的相关性。方法:应用免疫组化方法对人类睾丸组织中表达HO的细胞进行定位;采用逆转录-荧光定量PCR(FQ-PCR)方法定量检测无精子症患者与正常人睾丸组织HO-1及HO-2基因水平的表达量;应用W est-ern印迹检测各组之间HO蛋白水平表达量。结果:在正常睾丸组织,HO-1主要表达在支持细胞上;而HO-2在支持细胞和各级生精细胞中均有表达;FQ-PCR结果显示非梗阻性无精子症患者睾丸组织HO-1、HO-2的表达量均显著低于正常组及梗阻性无精子症组(P<0.05),差异具有统计学意义。而梗阻性无精子症患者睾丸组织表达HO-1、HO-2的量与正常组相比无显著性差异。W estern印迹结果显示HO-1蛋白水平的表达量差异与基因水平一致。而HO-2的蛋白水平在各组之间表达没有显著性差异。结论:非梗阻性无精子症患者睾丸组织中HO的表达量显著性降低,且HO-1无论是蛋白水平还是基因水平的差异一致。HO-1可以通过抗炎、抗氧化、抗凋亡的机制保护睾丸组织免受各种应激的损伤,从而维护正常的生精功能。可见,HO-1的减少可能与生精功能低下相关,这可能是非梗阻性无精子症的发病机制之一。     

睾丸精子获取和单精子卵胞浆内注射术治疗非梗阻性无精子症不育

目的　探讨非梗阻性无精子症患者外科获取睾丸精子的方法和意义。　方法　 4 9例非梗阻性无精子症患者行开放睾丸活检和诊断性睾丸精子获取术 (TESE) ,诊断性TESE有精子者至少 3个月后行单精子卵胞浆内注射 (ICSI)治疗。　结果　 12例 (2 4 .9% )诊断性TESE中发现精子 ,其中 3例为生精减少 ,2例为生精阻滞 ,7例为Sertoli细胞综合征。睾丸体积、血FSH水平和睾丸病理类型不能准确预测是否有精子。 8例行ICSI治疗 ,7例 (87.5 % )再次TESE获得睾丸精子行显微注射 ,3例获得临床妊娠。　结论　非梗阻性无精子症患者有必要行诊断性TESE确定睾丸内是否存在精子 ,获取睾丸精子结合ICSI可以有效治疗非梗阻性无精子症不育。     

梗阻性和非梗阻性无精子症患者低温保存的睾丸精子的质量

睾丸精子提取术获得的精子，采用卵细胞浆内注射进行体外受精，常用于治疗不育症。７３例梗阻性无精症和４２例非梗阻性无精症患者，在诊断性活检的时候，同时进行睾丸精子提取，所有的梗阻性无精症患者和１５例非梗阻性无精症患者提取的精子低温保存。冰冻前测量精子的数量、活动能力、形态和成活率，解冻后复查精子的成活率和活动能力。１７对夫妻共进行了２０个循环的卵细胞浆内注射体外受精，分别测定梗阻性和非梗阻性无精症患者的受精率、细胞分裂率和妊娠率。睾丸活检时发现，与梗阻性无精症相比，非梗阻性无精症精子的数量和形态都比…     

无精子症患者睾丸穿刺中评价支持细胞意义

目的 评价无精子症患者睾丸穿刺中检测支持细胞指数的意义。方法 采用Johnson评分法评价睾丸活检组织生精情况；采用瑞-姬染色，人工判读睾丸穿刺液涂片中各期生精细胞和支持细胞，并对其进行分类计数。结果 睾丸活检组织随着Johnson评分降低，支持细胞形态结构越不规则；睾丸穿刺中生精正常组9例，支持细胞指数范围均在20％-40％之间；生精障碍组15例，其中11例支持细胞指数范围在80％～150％，4例支持细胞指数范围在30％以下；生精阻滞组9例，支持细胞指数范围均在150％～500％之间，1例为15％。结论 生精正常组与生精障碍组、生精阻滞组的支持细胞指数差异明显，无精子症患者睾丸穿刺报告中支持细胞指数具有重要意义。     

无精子症患者睾丸系统针吸检查与组织病理检查的关系

目的探讨将睾丸系统针吸检查(SFNA)与组织病理检查相结合来预测睾丸中精子存在的可能性。方法67例无精子症患者先接受睾丸切开活检术及组织病理学检查,根据活检结果将患者分为梗阻性和非梗阻性无精子症(NOA),然后进行睾丸系统针吸检查及湿片镜检来发现精子的存在。结果67例患者中梗阻性无精子症12例,非梗阻性无精子症55例。所有患者的睾丸组织病理类型分为以下几种:正常精子生成、精子生成低下、生精阻滞、唯支持细胞综合征和混合型损害。行睾丸SFNA后,49％的NOA患者中发现精子,其中组织病理类型为精子生成低下的患者具有最高的精子获取率(95％)。结论对于无精子症患者而言,睾丸系统针吸检查是在睾丸中检测精子存在的有效方法;通过该方法检测到精子的可能性与睾丸组织病理类型有密切关系。     

钙粘附分子CDH18、PCDH17在人无精子症睾丸中的表达

目的:评价钙粘附家族分子CDH18和PCDH17在人无精子症及正常睾丸组织中的表达及意义。方法:常规病理组织学切片观察人非梗阻性无精子症及正常睾丸组织支持细胞-生精细胞紧密连接改变情况,随后应用包含有多个人钙粘附家族分子在内的cDNA微矩阵芯片对正常人及无精子症睾丸组织中差异表达情况进行研究;依据杂交结果设计Western蛋白印迹验证。结果:37.5%的无精子症睾丸组织出现明显的支持细胞-生精细胞紧密连接失常;部分钙粘附分子表达出现明显改变,其中CDH18及PCDH17表达下调。结论:钙粘附家族分子CDH18及PCDH17可能在无精子症的发生与进展过程中起一定的作用,该作用可能与支持细胞-生精细胞紧密连接失常有关。     

922例无精子症患者的病因诊断及其意义

目的探讨无精子症患者的病因。方法对922例无精子症患者按WHO标准进行病因诊断分析。结果922例无精子症患者中先天性异常445例（48．3％）、特发性无精子症206例（22．3％）、梗阻性无精子症198例（21．5％）、医源性疾病26例（2．8％）、获得性睾丸损伤23例（2．5％）、全身性疾病10例（1．1％）、精索静脉曲张8例（0．9％）、内分泌病因6例（0．7％）;206例特发性无精子症患者睾丸活检结果,睾丸生精功能低下38例（18．4％）、生精阻滞69例（33．5％）、唯支持细胞综合征（SCOS）90例（43．7％）、生精小管透明变性9例（4．4％）。结论先天性异常、特发性无精子症和梗阻性无精子症是无精子症患者的主要病因,SCOS是特发性无精子症患者睾丸生精障碍的主要病理类型。     

显微切割睾丸活检在非梗阻性无精子症中的应用

一直以来,学者们认为非梗阻性无精子症因睾丸生精功能受损,导致精液中无精子,而无法生育自己的后代。但随着卵胞质内单精子注射技术的问世,近十几年来涌现出多种睾丸取精术(包括开放性睾丸活检、细针穿刺抽吸、显微切割睾丸活检等)。之后,大量研究表明非梗阻性无精子症患者睾丸中仍存有局部的生精灶,即使是Klinefelter综合征,也可成功取出精子。2010年欧洲泌尿外科学会(EAU)指南明确推荐非梗阻性无精子症采用开放性睾丸活检或显微切割睾丸活检取精。与开放性睾丸活检相比,显微切割睾丸活检的取精成功率高且并发症少,本文就其取精前预测指标、手术操作方法、取精成功率及术后并发症进行综述。     

Men with nonobstructive azoospermia have Leydig cell hypertrophy but not hyperplasia

PURPOSE: We determined whether testicular histology in men with spermatogenic failure due to nonobstructive azoospermia shows true Leydig cell hyperplasia. MATERIALS AND METHODS: Testicular biopsy specimens from 17 patients evaluated for infertility were retrospectively analyzed. Interstitial, tubular and Leydig cell volume were quantitatively evaluated. The total volume and number of Leydig cells per testicle were then calculated. RESULTS: In 10 patients with obstructive azoospermia testicular histology showed normal spermatogenic function, while 7 had nonobstructive azoospermia. Average testicular volume plus or minus standard deviation was significantly larger in those with obstructive versus nonobstructive azoospermia (18.0 +/- 7.0 versus 9.3 +/- 8.7 cc, p = 0.025). Interstitial versus tubular volume was 32% of the total testis in the obstructive and 63% in the nonobstructive groups (p = 0.003). Although Leydig cell volume was proportionally greater in men with nonobstructive versus obstructive azoospermia (13.3% versus 0.05%, p = 0.045), there was no significant difference in the average number of Leydig cells per testicle (3.96 x 10 and 6.17 x 10, respectively, p = 0.16). The average volume of individual Leydig cells was significantly greater in men with the nonobstructive condition (253.0 +/- 98.7 versus 174.0 +/- 57.7 microm., p = 0.045). CONCLUSIONS: These results suggest that men with nonobstructive azoospermia and those with normal spermatogenesis have an equivalent number of Leydig cells. However, the Leydig cells are hypertrophic and occupy a larger proportion of total testis volume in men with nonobstructive azoospermia. Therefore, patients with spermatogenic failure show Leydig cell hypertrophy but not hyperplasia.     

Mast cells in testicular biopsies of azoospermic men

This work aimed at identifying mast cells in testicular biopsies from 10 normal fertile controls, 20 patients with obstructive azoospermia and 70 patients with nonobstructive azoospermia. The biopsies were stained with haematoxylin and eosin stain for tubular-modified Johnson score and with toluidine blue stain for mast cells. Two populations of mast cells, peritubular and interstitial, were demonstrated in all sections with varied counts. Testicular sections with Sertoli cell only and spermatogenic arrest patterns demonstrated a significant increase in both peritubular and interstitial mast cells compared with other groups, whereas obstructive azoospermia demonstrated a nonsignificant increase compared with the controls. Mast cell count was significantly correlated negatively with Johnson score for both peritubular (P = 0.001) and interstitial (P = 0.001) populations. Whether these results could be a cause or an effect, a special role might be assigned to mast cells in the pathogenesis of disturbed spermatogenesis.     

Êxpression of inducible nitric oxide synthase (iNOS) in the azoospermic human testis

Azoospermia, which is the absence of spermatozoa in the ejaculate, is not a rare cause of male infertility. Inducible nitric oxide synthase (iNOS) is a calcium-independent NOS, which is present in the testis and involved in spermatogenesis, and apoptosis of Sertoli and germ cells. Twenty idiopathic infertile men presenting nonobstructive azoospermia were enrolled in this study, and testicular sperm extraction procedures were performed. Tissue extracts were dissected, and the fluid samples were investigated to determine the presence of spermatozoa. Histologic evaluation of the spermatozoa-present samples revealed that seminiferous tubules were normal and were lined by Sertoli cells and spermatogenic cells. However, in the spermatozoa-absent samples, the diameter of the seminiferous tubules was small, and Sertoli-cell-only syndrome was determined in most of the tubules. iNOS expression was very weak in Sertoli cells, germ cells, and in Leydig cells in the spermatozoa-present group. In the spermatozoa-absent group, the immunostaining was very intense in Sertoli and Leydig cells. Electron microscopy findings were supported the histologic results. In conclusion, complete germ cell loss and intense expression of iNOS in the Sertoli and Leydig cells in the spermatozoa-absent groups of azoospermic human testis suggest an essential role of iNOS in spermatogenesis.     

Histological alterations in Leydig cells and macrophages in azoospermic men

The study aimed to compare the histological features of Leydig cells and macrophages in the testicular interstitium of obstructive versus nonobstructive azoospermia. Thirty‐nine azoospermic men undergoing testicular sperm extraction during intracytoplasmic sperm injection were allocated into obstructive azoospermia group (GI) and nonobstructive azoospermia group (GII) which was subdivided into Sertoli cell‐only syndrome (GIIA), germ cell arrest (GIIB) and hypospermatogenesis (GIIC) subgroups. Serum LH, FSH and testosterone levels were measured. Ultrastructural changes and the mean number of CD68‐positive cells were estimated in the different groups. In GIIA, Leydig cells' processes came in contact with macrophages and showed smooth endoplasmic reticulum dilatation. In GIIB, Leydig cells showed apoptotic changes. Macrophages were commonly encountered in their vicinity demonstrating large number of lysosomes. In GIIC, Leydig cells showed euchromatic nuclei. Macrophages showed expulsion of their lysosomal contents in the interstitium surrounded by apoptotic bodies. The mean count of total CD68‐positive macrophages was higher in cases of obstructive azoospermia with nonsignificant differences compared to nonobstructive azoospermia groups. Significant increase in FSH level was detected in GIIA compared to GI. It is concluded that structural interactions might take place between Leydig cells and macrophages in the interstitial tissue of azoospermic men.     

Expression of endothelial nitric oxide synthase in the Sertoli cells of men with infertility of various causes

OBJECTIVE: To investigate how endothelial nitric oxide (eNOS) expression in the seminiferous tubules might be related to spermatogenesis, by examining eNOS expression in testicular tissue of patients infertile from various causes. PATIENTS AND METHODS: The study included five fertile men with a normal sperm concentration, nine patients with obstructive azoospermia, 20 with varicocele testes and eight with idiopathic azoospermia (Sertoli cell-only syndrome). Testicular biopsy specimens were examined by immunohistochemistry for eNOS protein expression, in addition to a routine pathological assessment. eNOS protein was detected using an eNOS monoclonal antibody. A Sertoli cell staining index (SSI) was defined as the ratio of stained Sertoli cells per total number of Sertoli cells, and was compared among the groups. RESULTS: eNOS was localized to Sertoli cells in the seminiferous tubules and Leydig cells in the interstium; although some degenerating germ cells stained, normal germ cells did not. The SSI was significantly lower in patients with Sertoli cell-only syndrome than in either fertile men or patients with obstructive azoospermia or varicocele. However, the SSI did not correlate significantly with the Johnsen score. CONCLUSION: The expression of eNOS in Sertoli cells may depend on the existence of germ cells and be associated with germ cell development.     

Probability of sperm detection in nonobstructive azoospermic men undergoing testicular sperm extraction procedures unrelated to clinical parameters

The study was conducted to evaluate the significance of preoperative clinical parameters for detection of mature testicular sperm cells in nonobstructive azoospermic men. Sixty-five consecutive men with nonobstructive azoospermia underwent testicular sperm extraction procedures. Testicular samples were analyzed histologically with patterns classified as mature spermatogenesis (normal or partial), arrest of spermatogenesis, and Sertoli cell only. Testicular sperm cells were isolated for use in an IVF/ICSI program. Histologic patterns and detection rate of sperm cells were correlated to clinical characteristics. Mature sperm cells were found in all levels of serum FSH. The men were divided into 3 groups based on their clinical characteristics (serum FSH level and testicular size). The distribution of the different testicular histologic patterns, as well as detection rate of sperm cells, was similar in all groups. No correlation was found between serum levels of FSH, LH, prolactin, or testosterone and sperm presence. None of these parameters, nor the testicular size and consistency, can serve as predictive variables of the histological pattern or the presence of mature sperm cells in the testicular biopsies in cases of nonobstructive azoospermia. Until an effective predictive tool is available, a trial of sperm retrieval is recommended for all azoospermic men independent of their clinical characteristics.     

抑制素B βB亚单位在不同病理分型的无精子症患者睾丸组织中的表达

目的:探讨抑制素B(INH B)βB亚单位在不同生精功能状态的人睾丸组织中的表达情况。方法:对83例无精子症患者进行睾丸组织病理检查诊断,根据病理形态的不同分为:唯支持细胞综合征型(n=21);生精功能低下型(n=20);生精阻滞型(n=24);生精功能基本正常型(n=18)。选择上述各型结构完整的睾丸组织,分别应用免疫组化法(SP)对血清INH B βB亚单位在不同生精功能状态的睾丸组织,进行定位研究。结果:各型睾丸组织内均存在血清INH B βB的表达,其分布特点为:间质细胞(Leydig cell)和早期生精细胞多为强阳性表达,呈深棕黄色;支持细胞(Sertoli cell)多为阳性表达;而晚期精子细胞和成熟精子未见表达;生精小管管周类肌细胞呈弱阳性表达。结论:INH B可能是睾丸Sertoli细胞和早期生精细胞的一个联合产物。     

Analysis of histological findings of testicular biopsies in male infertility

Histological findings of testicular biopsy were studied following the Johnsen's score count method in 68 cases of idiopathic male infertility, and the relation between serum levels of luteinizing hormone (LH), follicle stimulating hormone (FSH), and testosterone (T) and histological findings were analyzed in the same cases. In oligozoospermia, there were no cases showing a Johnsen's score lower than 4. The score counts ranged widely from 1 to 9 in azoospermia. The cases with the Johnsen's score count lower than 4 revealed high values of serum LH and FSH and a low level of serum T. There was no relationship between Leydig cell accumulations or thickness of the seminiferous tubular walls and score values. Further examination using ABC (avidin biotin complex) method was carried out to find the localization of FSH and T in the testicular tissues. Immunohistochemical localization FSH was not noted in normal testicular tissues obtained the autopsy cases and testicular biopsy specimens of idiopathic male infertility. The localization of T was found in the Leydig cells and the Sertoli cells of normal and infertile testes. In the cases with the thickness of tubular walls, Sertoli cells were not stained. This fact might indicate that absence of T in Sertoli cells is related to spermatogenetic maturation only with the thickness of seminiferous tubular walls.     

Expression of beta-catenin in human testes with spermatogenic defects

beta-catenin is a multifunctional molecule that functions in intercellular adhesion and signal transduction during assembly of AJs between Sertoli cells as well as between Sertoli cells and germ cells. To assess changes in the testicular beta-catenin in male infertility conditions, testicular tissues from obstructive azoospermia with normal spermatogenesis, spermatogenic arrest (SA) and Sertoli cell-only syndrome (SCO) patients were examined for immunohistochemical localization of beta-catenin. In normal spermatogenic tissue, expression of beta-catenin was largely found in the Sertoli cell-germ cell (primarily spermatocytes) contact areas. Interestingly, perinuclear localization of beta-catenin was found in spermatocytes and spermatids. In spermatogenic arrest, beta-catenin in cell contact areas between Sertoli cells and germ cells was greatly decreased, but perinuclear beta-catenin in spermatocytes was not. In SCO, weak or negligible immunoreactivity of beta-catenin was found in cell contacts between Sertoli cells. Nuclear localization of beta-catenin was found in myotubular cells in all samples. Taken together, altered expression of beta-catenin in cell contacts within the seminiferous epithelia in spermatogenic arrest and SCO suggests that interactions between Sertoli cells and germ cell are crucial for expression of beta-catenin, and thus functional development of AJs in seminiferous epithelia in human testis. It should be also emphasized that perinuclear beta-catenin in germ cells may play a specific role in spermatogenesis.