EXPRESSION OF CANNABINOID RECEPTOR 1 IN MOUSE TESTES

Marijuana smoke and cannabinoids adversely affect male reproductive function in human and rodent through the cannabinoid receptors. To understand the possible function of cannabinoid receptor 1 (CB1) in spermatogenesis, expression of CB1 in testis during the postnatal development was examined in mice. Semiquantitative RT-PCR analysis revealed that testicular CB1 mRNA level was relatively high at 1 week post partum (p.p.). Following decrease during prepubertal development (2 weeks p.p.) and CB1 mRNA level re-increased during puberty (4 weeks p.p.) and reached the peak in adult testis. At 1 week p.p., some spermatogonia and Leydig cells showed strong immunoreactivity of CB1. At 2 weeks p.p., CB1 immunoreactivity was largely found in the primary spermatocytes as well as spermatogonia, and Leydig cells showed a weak signal. In adult testis, strong immunoreactivity was found in Leydig cells and luminal epithelia of seminiferous tubule. Germ cells including spermatozoa were positive for CB1 immunoreactivity. On Western blot, multiple forms of CB1 proteins were detected in testes, suggesting oligomerization of CB1. Ubiquitous, but spatiotemporal difference in expression of CB1 in soma and germ line during postnatal development of testis suggests functional involvement of CB1 signaling in steroidogenesis, spermatogenesis and fertilization.     

CISPLATIN-INDUCED GERM CELL APOPTOSIS IN MOUSE TESTES

The purpose of this study was to investigate whether exposure of male mice to cisplatin induces apoptosis in male germ cells and the possible role of apoptosis in cisplatin-induced testicular damage. Forty-eight male BALB/c mice were divided into cisplatin and control groups. The mice from the cisplatin group received a single intraperitoneal injection of cisplatin of either 1, 5, or 10 mg/kg. The control group received a single intraperitoneal injection of saline alone. The testes were removed on days 1, 3, and 7 after cisplatin administration, respectively. Following histological examination, apoptotic indices (AIs) were measured within seminiferous tubules of the mouse testes by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay. A low incidence of spontaneous apoptosis was observed in controls, particularly in spermatogonia and spermatocytes of the mouse testes. After cisplatin administration, both increased AIs and decreased spermatozoa and spermatids were found in the seminiferous tubules of the mouse testes. Cisplatin-induced apoptosis was found in spermatogonia, spermatocytes, and spermatids of the mouse testes. In comparison to the control values, AIs increased 2.6- to 6.8-fold in cisplatin-treated mouse testes. AIs reached the highest level on day 1 following 1 mg/kg, on day 3 following 5 mg/kg, and on day 7 following treatment of 10 mg/kg cisplatin. The study showed that cisplatin-induced germ cell apoptosis in the mouse testes was related to both the dose response and the time course of response. It is suggested that cisplatin-induced germ cell apoptosis may result in decreased spermatogenesis, and the higher dose of cisplatin may delay the occurrence of apoptosis in the mouse testes.     

INFLUENCE OF ACETAZOLAMIDE ON AQP1 GENE EXPRESSION IN TESTIS AND ON SPERM COUNT/MOTILITY IN EPIDIDYMIS OF RATS

Acetazolamide (Ace) is a putative inhibitor of carbonic anhydrase (CA), an enzyme that catalyzes the equilibration of carbon dioxide and carbonic acid and plays a key role in HCO 3 - and water reabsorption and acid secretion. Aquaporins (AQPs) are channel-forming membrane glycoproteins that mediate water reabsorption by the renal tubules and other organs of mammals. AQP1 and CAII or CAIV share many common biological properties. Previous studies have shown that AQP1 and CA are located at the same sites in cells of the male reproductive tract. In the present study, Ace at a dose of 40 mg/kg/d &#50 14, administered per os, suppressed AQP1 gene expression and inhibited CA activity in rat testis. On day 7 of treatment the epididymal sperm motility was significantly reduced, while on day 14 a decrease in sperm count occurred. Ace caused a marked downregulation of AQP1 gene expression; significant suppression occurred on days 7 and 14. Moreover, CA activity was totally blocked throughout the treatment period. The present findings suggest that the reduction of rat sperm motility and count by Ace can be attributed to its capacity to downregulate AQP1 water channel gene expression.     

壬基酚对围产期雄性子代SD大鼠早期发育的影响

目的:研究壬基酚(NP)对围产期雄性子代整体发育及睾丸组织发育的影响。方法:对母鼠孕期第7~20 d染毒NP(0,50,100,200 mg/kg),妊娠第20 d分别处死各组孕鼠,观察雄性子代整体发育,并对睾丸进行组织病理学观察和电镜观察。结果:NP染毒组孕鼠血清NP浓度和雌二醇浓度明显上升(P<0.05),子代胎鼠的死亡率随NP染毒剂量组浓度后升高而增加(P<0.05)。100 mg/kg和200 mg/kg NP染毒组雄性胎仔体重及肛殖距显著小于对照组(P<0.05)。睾丸组织病理学切片光镜观察,实验组和对照组未见明显的差异。电镜观察可见200 mg/kg NP染毒组精母细胞有核溶解现象。结论:母鼠孕期染毒壬基酚能够影响子代雄性SD大鼠整体发育和睾丸的早期发育。     

EXPRESSION OF BCL-2 AND BAX IN RHESUS MONKEY TESTIS DURING GERM CELL APOPTOSIS INDUCED BY TESTOSTERONE UNDECANOATE

Apoptosis occurs spontaneously during spermatogenesis and can be induced by androgen withdrawal. However, the molecular events governing apoptosis have not been characterized. To study the molecular mechanism of apoptosis induced by a high dose of testosterone undecanoate (TU), the authors examined the temporal changes in proapoptotic Bax and antiapoptotic Bcl-2 in TU-treated monkey testes. Apoptotic cells were identified in tissue sections by in situ end labeling of fragmented DNA. The results showed that a great deal of the apoptotic cells occurred in the testes on day 30 after TU injection and that the dominant apoptotic germ cells are spermatocytes and spermatids. The expression of Bcl-2 and Bax was assessed by immunohistochemical method and Western blot. As compared with that of normal testes, the levels of Bcl-2 protein increased significantly from 7 to day 14 while that of Bax protein was almost unchanged in the testes from day 7 up to day 60 after TU treatment. Bcl-2 was localized to the spermatids in the normal testes and temporarily distributed in both the cytoplasm and nucleus of those cell types susceptible to TU-induced apoptosis on day 14 after TU injection. Therefore, it is suggested that Bax may not play a role in initiating germ cell apoptosis induced by TU injection and that the evaluation in Bcl-2 expression may represent a survival mechanism for the remaining germ cell.     

羟基酪醇对肥胖大鼠睾丸线粒体氧化损伤的保护作用

目的 研究羟基酪醇(hydroxytyrosol,HT)对高脂膳食诱发的大鼠睾丸线粒体氧化损伤的保护作用.方法 雄性SD大鼠48只,随机分出8只为基础对照组,给予基础饲料喂养.其余40只给予高脂喂养8w成功诱导出16只肥胖模型,将肥胖模型大鼠随机分为高脂对照(high-fat,HF)和HT两组,继续喂高脂饲料5w,同时HT组灌胃给予HT(25 mg/kgb w),实验结束时,称体重,杀动物并提取睾丸组织,检测线粒体氧化损伤指标.结果 与基础组比,HF组大鼠睾丸组织ROS和MDA含量增高,总抗氧化能力和总SOD活性下降,线粒体膜电位下降,与HF组相比,HT组ROS和MDA含量降低,总抗氧化能力和总SOD活性增强,膜电位水平升高.结论 HT对高脂饮食诱导肥胖大鼠睾丸线粒体的氧化损伤具有一定的保护作用.     

性成熟期前小鼠生殖细胞发育的超微结构研究

[目的]观察小鼠性成熟期前不同发育阶段睾丸中生精细胞、支持细胞、睾丸间质细胞的超微结构特征,为探讨3种生殖细胞在发育过程中的形态学结构变化及相互关系提供实验依据. [方法]取生后5 d、10 d、20 d雄性小鼠睾丸,制片后经透射电镜观察并拍片记录. [结果]生后5～7 d,生精小管管腔小,细胞排列呈单层,仅有精原细胞和幼稚的支持细胞,睾丸问质细胞极少.生后10 d,小管管腔增大,细胞排列呈2～3层,精原细胞出现增殖分化,支持细胞、睾丸间质细胞开始发育.生后20 d,小管细胞层数增多,出现各级生精细胞和胞质问桥及变态发育中的精子;支持细胞、间质细胞发育成熟. [结论]生精细胞的发育具有明显的规律性,不同发育阶段表现出不同的细胞组合,且间质细胞、支持细胞的结构成熟先于生精细胞的分化,表明生精细胞的发育受闻质细胞、支持细胞的共同影响.     

ACTIVATION OF ENDOTHELIAL NITRIC OXIDE SYNTHASE IN CONTRALATERAL TESTIS DURING UNILATERAL TESTICULAR TORSION IN RATS

There are controversies about the injury of the contralateral testis during unilateral testicular torsion (UTT). An autonomic reflex arc between bilateral testes has been proposed. The authors focused on the involvement of nitric oxide (NO) in the contralateral testis during UTT. Eight-week-old male Wistar rats underwent unilateral torsion (1 h)-detorsion (up to 24 h). NO synthase (NOS) activity was detected as NADPH-diaphorase activity after fixation by paraformaldehyde. N -nitro- L -Arginine methyl ester (L-NAME, 20 mg/kg) was injected intravenously to the other group of rats. To evaluate the testicular injury, proteolysis of &#102 -fodrin production was detected by Western blotting. Apoptosis of the germ cells was evaluated by TUNEL. Long-term effect on spermatogenesis was evaluated by flow cytometry at 60 days after UTT. Transient activation of NOS was detected following the proteolysis of &#102 -fodrin in the contralateral testis. L-NAME inhibited these alterations. NADPH-diaphorase activity and eNOS immunoreactivity were co-localized in the endothelial cells. These reactions were not observed in other organs. There was neither enhanced apoptosis nor deteriorated spermatogenesis in the contralateral testis during and 60 days after UTT. In the contralateral testis, eNOS-derived NO regulates the vasomotor function against unilateral testicular torsion, whereas it acts slightly cytotoxic. These results suggest the possible involvement of a testis-specific neurovasomotor reflex between the bilateral testes.     

乳酸菌对感染肠上皮细胞通透性及紧密连接蛋白表达的影响

目的:研究乳酸菌对侵袭性大肠杆菌(EIEC) 感染肠上皮细胞(Caco-2)后,对其通透性和紧密连接(TJ)蛋白表达的影响. 方法:建立Caco-2 单层细胞感染模型.实验分为正常组、感染组、乳酸菌组和庆大霉素组.用电阻仪测定单层细胞的跨膜电阻值(TER),用高效液相法(HPLC)测定甘露醇透过率的变化,采用免疫组化法观察TJ相关蛋白,如Claudin-1蛋白,Occludin蛋白, JAM-1蛋白,ZO-1蛋白的分布和结构变化. 结果:EIEC感染Caco-2细胞后,单层细胞的TER值和甘露醇的透过率随时间延长而升高,而经乳酸菌处理后,TER值升高趋缓(P＜0.05),甘露醇的透过率降低(P＜0.05);EIEC感染后,相邻Caco-2细胞间TJ结构遭到破坏,TJ相关蛋白的表达亦减少,而乳酸菌处理后可使EIEC引起的TJ结构受损减轻、相关蛋白的表达增多. 结论:乳酸菌黏附于肠上皮细胞Caco-2后,可抑制EIEC破坏单层细胞的完整性,并改善TJ的结构变化和相关蛋白的表达分布.     

ROLE OF PROSAPOSIN IN THE MALE REPRODUCTIVE SYSTEM: EFFECT OF PROSAPOSIN INACTIVATION ON THE TESTIS,EPIDIDYMIS, PROSTATE,AND SEMINAL VESICLES

The effect of testosterone replacement therapy on serum prostate-specific antigen (PSA) was investigated in 11 patients with Klinefelter syndrome. Significant increases in serum testosterone level and prostate volume were observed after testosterone replacement therapy. However, serum PSA level did not change after testosterone replacement therapy. It would appear that serum PSA is not influenced by exogenous testosterone in patients with Klinefelter syndrome.     

原发性高血压病患者血清中HMGB1的表达

[目的]通过检测高迁移率组蛋白B1(High mobilitygroup protein B1,HMGB1)在原发性高血压病患者血清中的表达水平,探讨其表达差异在其发病中的意义。[方法]采用酶联免疫吸附法(ELISA)检测50例高血压不伴靶器官损害、62例高血压伴靶器官损害以及45例健康体检者血清中HMGB1的浓度。[结果]高血压病患者血清HMGB1水平均显著高于健康对照组,高血压伴靶器官损害组患者血清中HMGB1的水平明显高于高血压不伴靶器官损害组。[结论]从3组患者血清HMGB1的表达差异,推测HMGB1可能作为一种重要的炎症介质参与了高血压病的发生与发展过程。     

微创睾丸穿刺活检术的配合及护理

目的提高微创睾丸穿刺术的护理技术水平,减少并发症。方法从2007年7月～2009年10月共210例无精子症患者应用微创睾丸穿刺活检取曲细精管。结果210例患者均成功取出曲细精管。结论做好心理护理、术前准备、术中密切配合、术后的健康宣教,能有效地预防微创睾丸穿刺活检术后并发症的发生。     

NDRG1在前列腺癌中的表达及意义

目的探讨NDRG1在前列腺癌中的表达及其意义,以及NDRG1的表达与前列腺癌分化的关系。方法收集2002年1月～2011年12月广州市第一人民医院手术切除的前列腺癌标本154例,其中伴转移16例(骨转移标本9例,淋巴结转移的标本7例)。所有患者术前未行放疗和化疗。观察前列腺癌病理形态特征,并用免疫组织化学EnVison法检测NDRG1的表达。结果 NDRG1在前列腺癌中低表达,肿瘤的分化程度越低表达越低(p<0.05),在前列腺癌转移的患者中NDRG1的表达明显下降。结论 NDRG1在前列腺癌中低表达,并且随着肿瘤的发生发展,表达量明显降低。NDRG1可能对前列腺癌起着抑制作用,提示该基因可望作为一个候选的转移抑制基因和早期预测肿瘤转移的分子生物学指标之一,并对前列腺癌的预防和治疗提供一定的帮助。     

THYROID HORMONES: THEIR ROLE IN TESTICULAR STEROIDOGENESIS

Thyroid hormones are important for growth and development of many tissues. Altered thyroid hormone status causes testicular abnormalities. For instance, juvenile hypothyroidism/neonatal transient hypothyroidism induces macroorchidism, increases testicular cell number (Sertoli, Leydig, and germ cells) and daily sperm production. Triiodothyronine (T3) receptors have been identified in sperm, developing germ cells, Sertoli, Leydig, and peritubular cells. T3 stimulates Sertoli cell lactate secretion as well as mRNA expression of inhibin- &#102, androgen receptor, IGF-I, and IGFBP-4. It also inhibits Sertoli cell mRNA expression of Müllerian inhibiting substance (MIS), aromatase, estradiol receptor, and androgen binding protein (ABP) and ABP secretion. T3 directly increases Leydig cell LH receptor numbers and mRNA levels of steroidogenic enzymes and steroidogenic acute regulatory protein. It stimulates basal and LH-induced secretion of progesterone, testosterone, and estradiol by Leydig cells. Steroidogenic factor-1 acts as a mediator for T3-induced Leydig cell steroidogenesis. Although the role of T3 on sperm, germ, and peritubular cells has not yet been completely studied, it is clear that T3 directly regulates Sertoli and Leydig cell functions. Further studies are required to elucidate the direct effect of T3 on sperm, germ, and peritubular cells.     

维生素A缺乏对小鼠胚胎Hox基因表达的影响

目的：观察维生素Ａ（ＶＡ）缺乏对小鼠胚胎ＨｏｘＣ４（３．５）和ＨｏｘＤ１０（４．５）基因表达量的影响。方法：初断乳的昆明小鼠,雌鼠４０ 只均分为正常对照组（Ａ）和维生素Ａ缺乏组（Ｂ）,分别饲含ＶＡ４０００ ＩＵ／ｋｇ 饲料和ＶＡ ０ ＩＵ／ｋｇ 饲料。雄鼠饲以普通饲料。在饲养的第１７ 周按雌∶雄＝ ２∶１ 合笼交配。在怀孕的第１２ 天和第１４ 天,分别对孕鼠颈椎脱臼处死,立刻剖腹取出胎鼠,迅速置于液氮中速冻后置于－ ７０℃冰箱保存。用地高辛标记的探针,采用原位杂交方法探测小鼠胎儿ＨｏｘＣ４（３．５）和ＨｏｘＤ１０（４．５）基因ｍ ＲＮＡ含量。结果：ＶＡ缺乏时,ｍ ＲＮＡ的含量明显减少。结论：ＶＡ缺乏导致Ｈｏｘ 基因表达量下降,从而影响小鼠胚胎的正常发育。     

4种药物致小鼠肝脏毒性的基因表达谱聚类分析

目的在基因表达谱水平上观察红霉素、四环素、环磷酰胺、异环磷酰胺4种药物对Balb/c小鼠肝脏毒性效应。方法利用本室构建的小鼠毒理基因芯片和4种药物致Balb/c小鼠肝毒性动物模型,观察在药物作用不同时相点和剂量点上小鼠肝脏基因表达谱变化,结合层次聚类等手段对差异表达基因的生物学功能进行初步信息分析。结果共筛选出差异表达基因302个,其中282个参加了聚类分析。结果表明,环磷酰胺和异环磷酰胺组归为一类,红霉素和四环素归为另一类,不同时相和剂量组存在不同的聚类特征,一些差异表达基因在各药物组呈现共同上调或下调趋势。所有差异基因经genecards分析表明涉及脂肪代谢、蛋白质应激合成、抗氧化、炎症发生、细胞周期调控及药物代谢等多种生理功能。结论本研究揭示了与4种药物致Balb/c小鼠肝脏毒性相关的基因表达谱模式和特征基因群,为深入分析药物致小鼠肝毒性效应分子机制提供线索。     

六味地黄汤对衰老小鼠肾脏凋亡相关基因表达的影响

[目的]研究六味地黄汤对衰老小鼠肾皮质凋亡相关基因bcl-2、bax表达的影响。[方法]雄性KM小鼠分为青年对照组、老年对照组和六味地黄汤组。六味地黄汤组小鼠每日灌胃六味地黄汤,其余两组灌胃等量蒸馏水,共8周。应用免疫组织化学方法检测肾皮质bcl-2、bax表达。[结果](1)与青年对照组比较,老年小鼠肾皮质bcl-2表达下降,bax表达明显增加(P﹤0.05);(2)六味地黄汤组小鼠肾皮质bcl-2表达增多,bax表达下降,与老年对照组比较差异有统计学意义(P﹤0.05)。[结论]六味地黄汤可通过影响细胞凋亡相关基因的表达延缓肾脏衰老。     

共轭亚油酸对小鼠乳腺脂肪合成相关基因表达的作用研究

目的研究共轭亚油酸(CLA)对小鼠乳腺脂肪合成相关基因表达的影响。方法选取32只泌乳昆明鼠,随机分为对照组(2%花生油);0.5%CLA组(0.5%CLA+1.5%花生油);1.0%CLA组(1%CLA+1%花生油);1.5%CLA组(1.5%CLA+0.5%花生油),每组8只小鼠,从泌乳第4日饲喂至第14日,每日检测母鼠采食量、体重和仔鼠窝重,分析乳成分,采用荧光定量PCR检测对照组和1.5%CLA组母鼠乳腺组织脂肪合成相关基因的表达。结果在饲喂期间各组母鼠体重没有差异,饲料添加1.5%CLA显著减少了母鼠的采食量、仔鼠窝重和乳脂肪含量(P<0.05)。与对照组相比,1.5%CLA组母鼠乳腺组织的脂蛋白脂酶(LPL)、乙酰辅酶A羧化酶(ACACA)、硬脂酰辅酶A去饱和酶(SCD)和脂肪酸合成酶(FASN)的基因表达显著降低(P<0.05),并且转录因子固醇调节元件结合蛋白(SREBF)和过氧化物酶体增殖激活受体(PPARγ)的基因表达也显著下调(P<0.01)。结论饲料中加入1.5%CLA能够减少泌乳母鼠乳脂肪含量,抑制乳腺脂肪合成相关基因的表达。     

邻苯二甲酸二丁酯对F1 代仔鼠睾丸损害的可逆性恢复特性

目的研究和探讨邻苯二甲酸二丁酯(DBP)对F1代仔鼠睾丸损害是否具有可逆性恢复的特性。方法在宫内暴露和哺乳期(妊娠1d至仔鼠出生21d),每日每kg体重0、50、250和500mg DBP灌胃染毒孕鼠(每组10只),从解剖学、病理学等角度观察DBP对不同发育阶段(出生14、21和70d)F1代雄性大鼠的睾丸损害。结果与对照组相比,虽然不同发育阶段大鼠睾丸重量及系数没有明显统计学意义,但与对照组相比较,250、500mg／L组出生14d大鼠生精上皮变薄,Sertoli细胞数量明显减少(分别减少至对照组的84％和86％),并出现空泡样改变;出生21d时,Sertoli细胞数量和形态基本恢复正常,但有少量生精细胞脱落;至出生70d时,个别大鼠睾丸出现曲细精管变性、生精上皮萎缩等不可逆性损害。结论DBP宫内暴露和哺乳期染毒可损害F1代大鼠睾丸Sertili细胞,但随着仔鼠的发育,DBP致Sertoli细胞损害作用具有部分可逆性恢复的特性。     

YB-1在子宫内膜异位症中的表达

目的探讨YB-1在子宫内膜异位症(EMs)患者血清及子宫内膜组织中的表达及意义,以期将YB-1作为EMs早期诊断和治疗的判定指标。方法采用ELISA法检测EMs组与对照组血清中YB-1的表达水平,采用免疫组化SP法检测两组子宫内膜组织中YB-1的表达情况。结果与对照组(9.64±2.37 ug/L)相比,EMs组患者血清YB-1的平均浓度(14.65±3.91 ug/L)较高,二者相比差异有统计学意义(p<0.05)。免疫组化结果显示YB-1在EMs组患者子宫内膜中主要呈中等强度阳性表达。正常子宫内膜中也可见YB-1的少量弱阳性表达,两组子宫内膜组织中YB-1的阳性表达率相比有明显差异(p<0.05)。结论 YB-1在EMs患者血清及子宫内膜组织中高表达,其不仅可作为EMs细胞内的标志物,也有望成为一个理想的血清学标志物。