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。     

睾丸生精功能障碍的促生精治疗

正睾丸内精子的产生主要受生殖内分泌激素的调控,高浓度睾酮(Testosterone,T)的生精上皮及支持细胞环境是精子发生的必须条件。在生理情况下,下丘脑、垂体及睾丸的内分泌激素形成完整的反馈调控系统,促进和调控精子的发生、成熟[1]。睾丸生精功能障碍是指因各种原因导致睾丸内生精上皮(或生精组织)生精能力下降,生精细胞部分或全部发育阻滞,形成少精子或无精子,导致男性不育。目前临床对睾丸生精功能障碍的治疗方法主要包括药物治疗、手术治疗和辅助生殖。药物治疗在临床上被广泛使用,某些药物也确实     

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.     

原因不明少精子症睾丸生精细胞增殖与凋亡变化的研究

目的　探讨原因不明少精子症睾丸生精细胞增殖与凋亡的病理生理机制。　方法　 4 1例原因不明少精子症患者行睾丸穿刺活检 ,所取睾丸组织采用免疫组化技术观察生精细胞增殖细胞核抗原 (PCNA)及 p16的表达 ;采用原位 3′羟基末端标记法 (ISEL)观察生精细胞凋亡。 5例正常睾丸作为对照。　结果　原因不明少精子症患者睾丸生精细胞PCNA增殖指数 (2 1.6± 13.1) % ,明显低于对照组的 (45 .4± 5 .7) % ,P <0 .0 0 1;p16蛋白表达阳性率 (18.5± 8.9) % ,明显高于对照组的 (7.2± 2 .4 ) % ,P <0 .0 1;生精细胞凋亡指数 (1.7± 1.2 ) % ,明显高于对照组的 (0 .2± 0 .1) % ,P <0 .0 1,差别均有显著性意义。　结论　原因不明少精子症患者精子减少的主要原因之一是生精细胞增殖能力减少及凋亡增加造成增殖与凋亡平衡失调 ,此种失衡可能与 p16蛋白的异常表达有关。     

Fas和FasL 系统在非梗阻性无精子症睾丸中的表达

目的：为探讨睾丸生精功能障碍与细胞凋亡的关系，研究Fas和FasL系统在非梗阻性无精子症睾丸支持、间质和生精细胞中的表达。方法：对20例非梗阻性无精子症患者行睾丸开放性活检，常规病理检查，按Johnson评分法评价精子发生和发生障碍的程度；采用免疫组化SABC法对睾丸支持、间质和生精细胞进行Fas和FasL表达的检测。结果：睾丸活检生精功能评为8分有14例，3分有2例，6、5、4和2分各有1例。在20例非梗阻性无精子症睾丸的间质、支持和生精细胞均有Fas和FasI。的表达；而支持细胞Fas和FasL的阳性和强阳性表达率明显高于间质和生精细胞。结论：非梗阻性无精子症的睾丸支持、间质及生精细胞Fas和FasL的高表达与精子生成障碍是一致的，非梗阻性无精子症可能与生殖细胞过度凋亡密切相关。     

血清抑制素B与睾丸生精功能

目的　探讨血清抑制素B与睾丸生精功能的关系。方法　将男性不育症病人分为原发性无精子症、严重少精子症和梗阻性无精子症 3组 ,另设正常对照组 ,分别测定其抑制素B、FSH及精子密度。结果　上述 4组的抑制素B浓度分别为 4 7.36± 32 .85 ,2 0 0 .92± 86 .4 4 ,2 82 .4 0± 98.2 1,2 36 .79± 10 7.70。结论　血清抑制素B可直接反映睾丸的生精功能状态 ,因而是判断男性生育力更有效的指标     

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。     

生精细胞移植与睾丸功能研究

精原干细胞移植是将供体睾丸细胞转移到受体睾丸中的一项新技术。来自于转基因或变异型供体的一定数量的生精细胞 ,通过显微注射技术进入到受体睾丸的精曲小管中。注射后 ,精原干细胞可以在受体睾丸中生长发育 ,启动精子发生 ,产生具有受精能力的精子。这项技术将容许科学家们作以下研究 :①研究精子发生的基础 ;②为不育男性提供了一种使精子发生重新产生的方法 ;③从遗传学方面控制精原干细胞而发展转基因动物。生精细胞移植与睾丸功能研究@商学军 @黄宇烽     

小睾丸组织超微结构变化与性激素相关性研究

目的 :探讨小睾丸组织超微结构变化和性激素改变及其相关性。　方法 :对 8例小睾丸病人和 12例健康成人血清进行性激素测定 ,光镜及电镜观察小睾丸组织的超微结构变化。　结果 :小睾丸病人和正常对照组血清性激素FSH、LH、T分别为 (2 1.0 5± 9.15 )IU/Lvs (6 74± 3 5 2 )IU/L、(2 2 .88± 6 .2 5 )IU/Lvs (6 6 0± 1 4 8)IU/L、(0 .30± 0 .0 4 )nmol/Lvs (17 5 5± 9 2 5 )nmol/L ,两者相比差异有显著性 (P <0 .0 1) ;精曲小管直径和管壁厚度分别为 (37.33± 6 .80 )、(10 .30± 1.82 ) μm ,与正常组的 (198 4 6± 2 9 84 )、(2 95± 0 2 0 ) μm相比 ,差异有极显著性 (P <0 .0 1) ;组织超微结构变化显著。　结论 :小睾丸组织精曲小管、生精上皮、支持细胞、界膜、间质细胞及血管均发生严重的病理改变 ,其形成可能与遗传和免疫反应有关     

睾丸细针穿刺吸液细胞学检查诊断阻塞性无精子症

目的 :观察睾丸细针穿刺吸液 ( FNA)细胞学检查的效果 ,为诊断阻塞性无精子症提供新的诊断方法。方法 :2 86例无精子症患者采用睾丸 FNA细胞学检查结合精浆生化指标测定及输精管造影对睾丸生精功能及阻塞部位进行诊断 ;以 42例精子密度在正常范围 ( 2 5～ 86× 1 0 6 / ml)的成年男性作为对照组。 2 4例做钳穿活检进行自身对照。结果 :( 1 )双侧输精管未触及者 58例 ,睾丸 FNA细胞学检查生精功能正常 2 6例 (可见较多生精细胞、精子细胞及精子 )、生精功能低下 2 4例、无生精功能 8例 ,精浆果糖在正常值范围 ,而肉毒碱及α-糖苷酶明显低于正常值范围 ;( 2 ) 3 2例睾丸 FNA细胞学检查见较多精子 ,精液沉渣涂片未见生殖细胞 ,其中 6例精浆果糖、肉毒碱及 α-糖苷酶明显低于正常值范围 ,结合输精管造影确诊为射精管阻塞 ,其余 2 6例精浆果糖在正常值范围 ,而肉毒碱及α-糖苷酶明显低于正常值范围 ,确诊为附睾尾部阻塞性无精子症 ;( 3 )睾丸生精功能极度低下或无生精功能 1 96例 ,其中 1 60例仅见各级生精细胞、精子细胞和支持细胞 (睾丸生精功能阻滞 ) ,3 6例仅见支持细胞 (唯支持细胞综合征 ) ,精浆果糖、肉毒碱及 α-糖苷酶均在正常值范围 ,为非阻塞性无精子症。结论 :睾丸 FNA细胞学检查可作为阻塞性无?     

Ultrastructure of the seminiferous tubules in oligoasthenoteratozoospermic men associated with varicocele

Varicocele is associated with venous reflux that may cause increased heat and interstitial pressure within the testes, with variable pathological effects on spermatogenesis. This study aimed to study the ultrastructural testicular changes in the seminiferous tubules of 20 infertile severe oligoasthenoteratozoospermia (OAT) men associated with varicocele and five patients with obstructive azoospermia without varicocele as controls. They were subjected to testicular biopsy which was evaluated by transmission electron microscopy. Ultrastructurally, the seminiferous epithelium in the testicular biopsies of infertile severe OAT men associated with varicocele was variably affected in the form of thickening of the peritubular connective tissue, vacuolation of Sertoli cell and germ cell cytoplasm, presence of degenerated and apoptotic cells among the germinal epithelium, altered spermatids and abnormal spermatozoa. It is concluded that varicocele in severe OAT men is associated with ultrastructural changes in the seminiferous tubule.     

Quantitative testicular biopsy in spinal cord injured men: comparison to fertile controls.

Spermatogenic abnormalities have been reported in the majority of spinal cord injured men on routine testicular biopsy. However, given the interim advances in their urological and rehabilitative care, a quantitative assessment of the germinal epithelium after spinal cord injury and comparison of these parameters to normal controls are warranted. Incisional testicular biopsy was performed in 14 spinal cord injured men. Quantitative micrometric techniques were applied to assess spermatogenesis and the results were compared to a normative data base of testicular biopsies previously obtained from a group of 15 fertile volunteers. From a minimum of 10 randomly selected round seminiferous tubules per subject the mean number of Sertoli cells, mature spermatids, tubular diameter and tubular wall thickness were determined in both groups and statistically analyzed. In the spinal cord injury group the mean number of spermatids per tubule was significantly lower and the mean number of Sertoli cells per tubule was significantly higher than in fertile controls (p less than 0.05). Moreover, the mean Sertoli cell-to-spermatid ratio per seminiferous tubule was significantly higher in the spinal cord injury group and discriminated between spinal cord injured men and controls, with a sensitivity of 93% and specificity of 100% (p less than 0.0001). Half of the spinal cord injury group showed a mean tubular spermatid density of less than 10. Compared to the fertile population, spinal cord injured men show significant differences in quantitative parameters of the germinal epithelium that may contribute to the reproductive dysfunction.     

Quantitative parameters of seminiferous epithelium in secretory and excretory oligoazoospermia

Testicular biopsy specimens from infertile men (sperm count, less than 10(6)/ml) were evaluated on 1-micron thick sections, and counts of stem cells and differentiated spermatogonia, primary spermatocytes, early and late spermatids, and Sertoli cells were compared to counts in six fertile men. Biopsy specimens were also compared for the appearance of seminiferous tubule wall, blood vessels, and interstitium. Infertile men were grouped according to the following diagnoses: hypospermatogenesis (n = 5), spermatocyte arrest of spermatogenesis (n = 5), and obstruction of the genital tract (n = 7). A low productivity of spermatogenesis in cases of hypospermatogenesis appeared to be due to an exaggerated degeneration of primary spermatocytes and to a yield of abnormal spermatids. A block of meiosis in spermatocyte arrest was associated with a degeneration of primary spermatocytes and with a reduced number of staminal spermatogonia. Abnormal spermiogenesis was observed in cases of obstruction of the genital tract and was associated with an increase in stem cell spermatogonia. A thickening of seminiferous tubule and blood vessel walls could be responsible for the limited functional capacity of Sertoli cells, causing altered spermiogenesis in cases of excretory azoospermia. A severe primitive failure of Sertoli cells in secretory oligoazoospermia could account for a deranged maturation and degeneration of premeiotic and postmeiotic germ cells.     

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。     

Quantitative （stereological） study of incomplete spermatogenic suppression induced by testosterone undecanoate injection in rats

Aim: To evaluate the key lesions in spermatogenesis suppressed partially by testosterone undecanoate(TU) treatment. Methods: Adult male SD rats were treated with vehicle or TU (19 mg/kg) injection (i.m.) every 15 days for 130 days. The numbers of all types of cells (nuclei) in the seminiferous tubules and the interstitial tissue were estimated using a contemporary stereological tool, the optical disector. Results: In response to TU treatment, the numbers of non-type B spermatogonia, type B sperrnatogonia and late elongated spermatids per testis were reduced to 51%, 66% and 14% of the controls, respectively. The conversion ratios from type B spermatogonia to early spermatocytes and pachytene spermatocytes were not significantly affected and the ratios to the later germ cell types fell to 51%-65% of the controls. Less than 1.0 % of immature round spermatids were seen sloughing into the tubule lumen, 4.0% of elongated spermatids retained in the seminiferous epithelium, and about half of the elongated spermatid nuclei appreciably malformed. Leydig cells were atrophied but their number and the peritubular myoid cell number per testis were unchanged. Conclusion: Double inhibition of spermatogenesis (i.e. inhibition at spermiation and spermatogonial conversion to type B spermatogonia), a scenario seen in the monkey and human following gonadotrophin withdrawal, was not sufficiently effective for a complete spermatogenic suppression in the rat after TU treatment, probably due to ineffective inhibition of the Leydig cell population and therefore the intra-testicular test-osterone levels. (Asian J Androl 2004 Dec; 6: 291-297)     

Quantitative morphology of the testicular tubular epithelium in the water buffalo (Bubalus bubalis).

Ultrastructural features and morphometric evaluations of water buffalo seminiferous epithelium are reported for the 6 phases of the spermatogenic cycle. The relative Sertoli cell volume varies between 30% (phase 4) and 39% (phase 8), the calculated volume of a Sertoli cell between 7118 microns3 and 8968 microns3 (phase 4). Smooth ER is the organelle that exhibits the most prominent changes in Sertoli cells during the spermatogenic cycle: it occupies about 6% in phase 3 and 21% in phase 4. All spermatogenic cells of the same clone present cytoplasmic bridges among them. From preleptotene (about 470 microns3) to late diplotene (about 2300 microns3) the volume of a primary spermatocyte increases nearly 5-fold; their nuclear volumes increase 3.5-fold in the same period. Secondary spermatocytes are found only in phase 4 of the cycle. Due to partial cell necrosis and autolysis late maturation phase spermatids display not more than 25% of the size of early cap phase spermatids. 63% of all numerically possible germ cells disappear from the seminiferous epithelium during spermatogenesis. Particularly heavy cell loss is observed in phase 4 and involves the spermatogonial fraction as well as cells during the second meiotic division.     

Bilateral prepubertal testicular biopsies predict significance of cryptorchidism-associated mixed testicular atrophy, and allow assessment of fertility

INTRODUCTION: Mixed atrophy of the testis (MAT), a frequent finding in biopsies of formerly cryptorchid and/or infertile patients, is defined as the synchronous occurrence of both seminiferous tubules containing germ cells and Sertoli cell only-tubules in variable proportions. In tubules containing germ cells, different types of abnormalities in spermatogenesis may be seen. The presence of adult spermatids in the biopsy, even in small numbers, correlates with successful spermatozoa retrieval for "in vitro" fertilization techniques. Currently, it is unknown whether precursor lesions of MAT can be identified in cryptorchid patients during childhood. MATERIAL AND METHODS: Eighteen formerly cryptorchid adults who had undergone testicular biopsies in childhood had a repeat testicular biopsy to evaluate infertility. In prepubertal biopsies, abnormalities of the testicular parenchyma were classified into types I (slight alterations), II (marked germinal hypoplasia), and III (severe germinal hypoplasia). In postpubertal biopsies, the percentage of tubules containing germ cells and Sertoli cell only-tubules were estimated, as well as the presence of complete spermatogenesis. Abnormalities in spermatogenesis were classified into lesions of the adluminal or basal compartments of seminiferous tubules. RESULTS: Comparison between prepubertal and postpubertal biopsies revealed that most specimens developing from type III lesions presented with incomplete spermatogenesis (P<0.0001) and more severe lesions of the germinal epithelium (P=0.049). DISCUSSION: Type III lesions correlated with MAT characteristics that confer a worse prognosis for in vitro fertilization. Thus, MAT characteristics may be predicted in prepubertal cryptorchid patients, allowing a fertility prognosis. The pathogenesis of these lesions, and their possible inclusion into the spectrum of the testicular dysgenesis syndrome, are discussed.     

Spermatogenese

Spermatogenesis takes place within the testicular seminiferous tubules which consist of the peritubular lamina propria and the seminiferous epithelium. The latter is composed of germ cells and somatic Sertoli cells. Sertoli cells trigger germ cell development by mediating follicle-stimulating hormone and androgen hormonal stimuli. Spermatogenesis comprises proliferation of spermatogonia, meiosis of spermatocytes, and differentiation of spermatids into spermatozoa (spermiogenesis). There are six distinct and specific germ cell associations (I–VI). These “stages of spermatogenesis” occur sequentially along the length of a tubule. Different defects in spermatogenesis occur in adjacent seminiferous tubules (mixed atrophy) and are associated with deficits in differentiation of Sertoli cells. Biopsy specimens should be fixed in Bouin’s solution. Diagnosis of preinvasive carcinoma in situ is based on the immunohistochemical demonstration of placental-like alkaline phosphatase (PLAP), which is expressed exclusively in carcinoma in situ cells. Histological evaluation should be performed using a score count system, and the use of histological techniques for protein and mRNA expression. Testicular biopsy should only be performed in accordance with strict indication criteria, and histological evaluation should be carried out in specialist centres, i.e. as recommended by the European Academy of Andrology (EAA).     

Spermatogenesis--physiology and pathophysiology

Spermatogenesis takes place within the testicular seminiferous tubules which consist of the peritubular lamina propria and the seminiferous epithelium. The latter is composed of germ cells and somatic Sertoli cells. Sertoli cells trigger germ cell development by mediating follicle-stimulating hormone and androgen hormonal stimuli.Spermatogenesis comprises proliferation of spermatogonia, meiosis of spermatocytes, and differentiation of spermatids into spermatozoa (spermiogenesis). There are six distinct and specific germ cell associations (I-VI). These "stages of spermatogenesis" occur sequentially along the length of a tubule. Different defects in spermatogenesis occur in adjacent seminiferous tubules (mixed atrophy) and are associated with deficits in differentiation of Sertoli cells. Biopsy specimens should be fixed in Bouin's solution. Diagnosis of preinvasive carcinoma in situ is based on the immunohistochemical demonstration of placental-like alkaline phosphatase (PLAP), which is expressed exclusively in carcinoma in situ cells. Histological evaluation should be performed using a score count system, and the use of histological techniques for protein and mRNA expression. Testicular biopsy should only be performed in accordance with strict indication criteria, and histological evaluation should be carried out in specialist centres, i.e. as recommended by the European Academy of Andrology (EAA).     

Histopathological mapping of open testicular biopsies in patients with unobstructive azoospermia.

OBJECTIVE: To evaluate, in patients with unobstructive azoospermia, the heterogeneity of spermatogenesis within the testes and thus whether there is any region of advanced spermatogenesis. Patients and methods Seventy infertile men (mean age 34 years, SD 5.01) with no varicoceles or testicular atrophy had bilateral open testicular biopsies taken from six different sites. For each biopsy specimen the number of seminiferous tubules and of tubules with sperm maturation were counted (by light microscopy at x 400). The ratio of tubules with active spermatogenesis to the total number was calculated for each biopsy sample. RESULTS: The mean (SD) right and left testicular volumes were 19.82 (7.8) and 18.84 (7.89) mL, respectively; the patients' follicle-stimulating hormone level was 8.34 (1.17) IU/mL. On sextant biopsy spermatozoa were detected in 42 of the 70 patients (60%). The mean (SD) ratio of tubules with spermatozoa was 5.23 (0.8)% for the right and 5.37 (0.76)% for the left testes. There was no statistically significant difference in the ratio of seminiferous tubules positive for spermatozoa at the different biopsy sites in either the right or left testis. Spermatozoa were identified in only one to three biopsy sites in almost half of those with maturation arrest; this ratio increased to 74% in patients diagnosed as having Sertoli-cell-only syndrome with focal spermatogenesis. Conclusion There is no region of the testis that is rich or advanced in spermatogenesis in patients with unobstructive azoospermia. Without multiple testicular biopsy it is possible to miss advanced spermatogenesis in some unobstructed patients. The sextant testis biopsy is a reliable method for detecting the presence and exact location of seminiferous tubules with spermatozoa in patients with unobstructive azoospermia.