Actin binding proteins,actin cytoskeleton and spermatogenesis – Lesson from toxicant models

Actin cytoskeleton is crucial to support spermatogenesis in the mammalian testis. However, the molecular mechanism(s) underlying changes of actin cytoskeletal organization in response to cellular events that take place across the seminiferous epithelium (e.g., self-renewal of spermatogonial stem cells, germ cell differentiation, meosis, spermiogenesis, spermiation) at specific stages of the epithelial cycle of spermatogenesis remain largely unexplored. This, at least in part, is due to the lack of suitable study models to identify the crucial regulatory proteins and to investigate how these proteins work in concert to support actin dynamics. Much of the information on the role of actin binding proteins in the literature, namely the actin bundling proteins, actin nucleation proteins and motor proteins, are either findings based on genetic models or morphological analyses. While this information is helpful to delineate the function of these proteins to support spermatogenesis, they are not helpful to identify the regulatory signaling proteins, the signaling pathways and the cascade of events to modulate actin cytoskeleton dynamics. Recent studies based on the use of toxicant models, both in vitro and in vivo, however, have bridged this gap by identifying putative regulatory and signaling proteins of actin cytoskeleton. Herein, we summarize and critically evaluate these findings. We also provide a hypothetical model by which actin cytoskeletal dynamics in Sertoli cells are regulated, which in turn supports spermatid transport across the seminiferous epithelium, and at the blood-testis barrier (BTB) during the epithelial cycle of spermatogenesis.     

高压氧对精索静脉曲张家兔附睾组织学影响的实验研究

目的研究高压氧(HBO)对精索静脉曲张(VC)家兔附睾组织学的影响,探讨精索静脉曲张导致男性不育的发病机制。方法采用HBO干预精索静脉曲张家兔模型,对睾丸、附睾的重量,附睾不同节段精子活力,附睾的组织形态以及精液参数进行研究。结果与未行HBO干预的VC模型比较,HBO 干预后的VC模型精液质量明显改善,睾丸、附睾重量明显增加(P值均<0．01),左侧附睾尾部Ⅱ级以上精子明显增加(Ⅱ级57．43％±6．48％,Ⅲ级2．91％±1．51％)(P<0．01,<0．05)。附睾上皮结构改善,腔内精子密度增加。结论 VC可导致附睾病理损害、精子成熟和获能障碍。慢性缺血、缺氧及其微循环障碍是导致附睾病理损害、精子成熟和获能障碍的主要机制。HBO可有效抑制VC致附睾的病理损害进程。     

基因重组细胞生长肽和精氨酸拮抗环孢霉素A的睾丸毒性

目的 探讨基因重组细胞生长肽（bFGF)和精氨酸对环孢霉素A（CsA)在血和睾丸中药物浓度及其睾丸毒性的影响。方法 将80只大鼠随机分为4组：N组,正常对照组;A组,CsA;B组,CsA 精氨酸;C组,CsA bFGF。每天腹腔注射给药,连续3周后取血及睾丸测定CsA浓度,按抗精子发生效应积分评定法作生精功能的评价,测定曲细精管直径及作病理学检查。结果 血药浓度B组明显高于A组（P＜0．05）;A,B,C组睾丸的发生明显障碍,而B,C组比A组的精子发生障碍要轻（P＜0．05）,曲细精管的直径A,B、C组明显小于N组（P＜）．05）,但B组大于A组（P＜0．01）。结论 CsA影响睾丸的生精功能,精氨酸和bFGF能明显降低CsA对睾丸的毒性作用,精氨酸还能提高血中CsA的浓度。     

黄芪注射液对氯化镉致睾丸和附睾毒性的拮抗作用

目的研究黄芪注射液拮抗氯化镉诱导的大鼠睾丸和附睾毒性作用及其机制。方法将50只SpragueDawley雄性大鼠随机分成5组:低浓度黄芪组(A1)、高浓度黄芪组(A2)、环磷酰胺组(CP)、氯化镉组(Cd)和对照组(C)。预先连续7d分别给A1组和A2组大鼠腹腔注射黄芪注射液5g·kg-1·d-1和10g·kg-1·d-1,CP组和Cd组以等量蒸馏水腹腔注射作对照。之后连续21d给A1组、A2组和Cd组大鼠同时腹腔注射氯化镉溶液(0.2mg·kg-1·d-1)。染镉后第22天处死大鼠,观察睾丸脏器系数、精子头计数、每日精子生成量、附睾尾精子计数和畸形率以及睾丸和附睾病理学;检测血清和睾丸组织总超氧化物歧化酶(T-SOD)和丙二醛(MDA)。结果A2组睾丸脏器系数、睾丸精子头计数、每日精子生成量和附睾精子计数[(5.68±1.19)、(49±9)×106/g、(10.2±2.3)×106/g、(47.5±22.5)×106/ml]明显高于Cd组[(3.11±0.16)、(39±11)×106/g、(5.1±0.3)×106/g、(10.9±2.4)×106/ml](P<0.05或P<0.01);A2组大鼠精子畸形率[(7.04±0.12)%]明显下降,与Cd组[(17.81±1.55)%]相比,差异有非常显著性(P<0.01);A2组血清及睾丸组织MDA含量和T-SOD活力与Cd组比较差异均有非常显著性(P<0.01)。结论黄芪注射液可用于防治氯化镉的生殖毒性作用,其作用机制可能与黄芪清除氧自?     

大鼠睾丸移植模型的建立及植睾损伤机制的研究

目的：探讨影响植睾生精细胞发育的内在因素及胶质细胞源性神经营养因子（GDNF）mRNA在植睾丸组织中的表达。方法：应用Cuff管技术建立大鼠原位异体睾丸移植模型，并将实验动物分为6组，在光镜和电镜下分别观察移植物的形态学和超微结构变化，应用放免法和RT—PCR技术检测实验各组T、FSH、LH水平和GDNFmRNA的表达量。结果：术后7d，同种移植组植睾组织在光镜下可见间质内大量淋巴细胞浸润，曲细精管内生精细胞层次紊乱，支持细胞数目减少并出现萎缩，电镜下可见支持细胞之间构成的紧密连接出现断裂；血清T值下降而FSH、LH值反馈性升高，GDNFmRNA的表达量较正常对照组减少。术后14d，同种移植组植睾损害进一步加剧，且GDNFmRNA的表达量明显下降。结论：同种移植植睾组织中支持细胞数目的减少和萎缩及其胞突构成的紧密连接出现断裂可能是植睾生精功能不良的主要原因；GDNFmRNA表达水平可作为睾丸生精功能状态指标之一。     

Klinefelter syndrome and TESE-ICSI

Until few years ago, Klinefelter syndrome with a homogenous 47,XXY karyotype was considered a model of absolute male sterility. We will discuss first the potential fertility following Testicular Sperm Injection, then the physiopathology of spermatogenic failure and the origin of focal spermatogenesis and risk of aneuploidy in offspring, and third the advantage of searching spermatozoa earlier instead of adult age. The rate of positive sperm extraction seems to be better for younger patients. During childhood, there is a low rate of spermatogonia. The spermagonia, which completes the spermatogenesis, seems resulting from a rare clone of 46,XY gonia, having lost their extra X chromosome. Several arguments suggest that this focal spermatogenesis decreases with age. In addition, androgen treatment, frequently prescribed in case of Klinefelter syndrome, carries a risk of decreasing focal spermatogenesis by lowering gonadotropins. The question arises if it is necessary to expect the sperm cryopreservation before introducing androgen treatment. Further studies are necessary to determine the best age of sperm retrieval in case of Klinefelter syndrome.     

生殖激素受体与精子发生关系的研究进展

精子发生是一个非常复杂的过程,尤其受生殖激素调控。生殖激素与其受体结合在精子的发生、分化、成熟中起着至关重要的作用。因此了解精子发生过程中生殖激素受体的变化对精子发生障碍研究有着重要的价值。本文对生殖激素受体的基因突变及多态性与精子发生的关系进行综述,以期为男性不育的诊治提供帮助。     

内源性抗氧化剂PRDX4及其在生殖系统作用的研究进展

过氧化物还原酶4（PRDX4）是过氧化物还原酶家族成员,是一种细胞内源性抗氧化剂,可锚定在内质网调节蛋白质氧化折叠,也可分泌到细胞外基质发生抗氧化作用。在女（雌）性生殖系统,氧化应激影响卵巢的功能状态,定位在颗粒细胞及卵母细胞的体细胞型PRDX4（PRDX4s）通过抗氧化应激机制促进卵泡发育成熟、抑制卵母细胞老化,并参与多囊卵巢综合征（PCOS）和卵巢功能减退的病理过程。在睾丸组织内同时表达两种PRDX4亚型,PRDX4s和睾丸特异型PRDX4（PRDX4t）。PRDX4通过抑制内质网应激、清除胞质和细胞外基质活性氧簇（ROS）保护睾丸组织结构与细胞功能,参与调节精子发生和精子形成,抑制生精细胞凋亡。综述PRDX4的结构与功能,及其在调节配子发生中的作用。     

Allethrin induces oxidative stress,apoptosis and calcium release in rat testicular carcinoma cells (LC540)

Over the years, pyrethroids, including D-allethrin, are widely used for domestic and agricultural purposes and are found to be toxic to many organ systems including the male reproductive system. However, the molecular mechanisms of allethrin toxicity are not well understood. In this study, we demonstrate that allethrin exhibited a dose-dependent cytotoxicity on Leydig cell carcinoma cells (LC540) and isolated primary Leydig cells with an IC₅₀ of 125 μM and 59 μM respectively. Cytotoxicity was associated with generation of reactive oxygen species, increased lipid peroxidation and alterations in antioxidant enzyme status. Morphological analyses of LC540 cells treated with allethrin revealed the presence of apoptotic bodies. Using flow cytometry, we observed that the number of cells that displayed early apoptotic features and entering into G₀ phase of cell cycle increased significantly with loss of mitochondrial membrane potential. The levels of p53 mRNA and cleaved PARP-1 protein were increased, whereas BCL-2, pro-Caspase-3 and PARP-1 were decreased. Allethrin induced apoptosis was associated with voltage gated calcium channel mediated intracellular calcium release. Results of our study demonstrate that allethrin toxicity in the male reproductive tract may involve Leydig cell apoptotic death.     

The Anti-Oxidant Effects of Ginger and Cinnamon on Spermatogenesis Dys-function of Diabetes Rats

Background

Diabetes rats have been linked to reproductive dysfunction and plant medicine has been shown to be effective in its treatment. Antioxidants have distinctive effects on spermatogenesis, sperm biology and oxidative stress, and changes in anti-oxidant capacity are considered to be involved in the pathogenesis of chronic diabetes mellitus. Ginger and cinnamon are strong anti-oxidants and have been shown to reduce oxidative stress in the long-term treatment of streptozotocin (STZ)-induced diabetes in animal models. The present study examined the influence of combined ginger and cinnamon on spermatogenesis in STZ-induced diabetes in male Wistar rats.

Materials and Methods

Animals (n = 80) were allocated randomly into eight groups, 10 each: Group 1: Control rats given only 5cc Normal saline (0.9% NaCl) daily;Group2: rats received ginger (100mg/kg/rat) daily; Group 3: rats received cinnamon (75mg/kg) daily; Group 4: rats received ginger and cinnamon, (100mg/kg/rat ginger and 75mg/kg cinnamon) daily; Group 5: Diabetic control rats received only normal saline. Group 6: Diabetic rats received 100mg/kg/day ginger; Group 7: Diabetic rats received 75mg /kg/ day cinnamon; Group 8: Diabetic rats received ginger and cinnamon (100mg/kg/day and 75mg/kg /day). Diabetes was induced with 55 mg/kg, single intra-peritoneal injection of STZ in all groups. At the end of the experiment (56th day), blood samples were taken for determination of testosterone, LH,FSH, total anti-oxidant capacity, and levels of malondialdehyde, SOD, Catalase and GPX. All rats were euthanized, testes were dissected out and spermatozoa were collected from the epididymis for analysis.

Results

Sperm numbers, percentages of sperm viability and motility, and total serum testosterone increased in ginger and cinnamon and combined ginger and cinnamon treated diabetic rats compared with control groups. Serum testosterone, LH and FSH were higher compared to control group and also serum anti-oxidants (TAC, SOD, GPX and catalase) all were increased at the end of treatment. Combined ginger and cinnamon showed more intense increase in all parameters compare to ginger and cinnamon alone. Most of the results were significant (P<0.05).

Conclusion

We concluded that combined ginger and cinnamon have significant beneficial effects on the sperm viability, motility, and serum total testosterone, LH,FSH and serum anti-oxidants'' level and could be effective for maintaining healthy sperm parameters and male reproductive function in diabetics.