电磁脉冲辐射对小鼠生精细胞超微结构的改变

目的　探讨电磁脉冲辐射对小鼠生精细胞超微结构的改变。　方法　二级昆明雄性小鼠 1 3 8只 ,随机分为辐射组和对照组。辐射组小鼠分别接受 8× 1 0 3V/m、2×1 0 4 V/m、6× 1 0 4 V/m电磁脉冲 5次重复全身照射 ,于照射后 6h和 1、3、7、1 4、2 8、90 d观察生精细胞超微结构的改变。　结果　电磁脉冲照射后 6h～ 2 8d,精原细胞、精母细胞和精子细胞均出现核膜间隙增宽、线粒体肿胀或灶性空化、内质网扩张 ,而精原细胞还出现异染色质凝集或边移 ,核膜局灶性溶解 ,糖原颗粒减少 ;精子细胞高度水肿、核膜膨出或灶性溶解、线粒体高度肿胀、内质网显著扩张 ;精子头部形态异常、染色质浓缩不良、顶体形态结构异常等。　结论　电磁脉冲照射可引起小鼠各级生精细胞超微结构的明显损伤 ,精原细胞、精子细胞和精子是电磁脉冲辐射最敏感的靶细胞     

睾丸移植物间质细胞4种基因的表达分析

目的 研究新生小鼠睾丸组织异体异位移植后,4种在睾丸间质细胞中起重要作用的基因表达情况,为异体异位睾丸组织移植模型用于科研及临床的可行性提供进一步实验数据.方法 将162只1～2d昆明小鼠的睾丸移植到54只7～12w去势雄性免疫缺陷小鼠背部;在移植后9个时间段(3d和1～8w)取出移植物;选取4种在睾丸间质细胞中表达或高表达的基因3β -hsd、1hr、p450scc和star,采用聚合酶链反应,对发育不同阶段移植物中4种基因的表达情况进行分析,并与正常小鼠相应各年龄段睾丸中的基因表达情况相比较.结果 在9个时间段取出的移植物中,所测定4种基因的表达趋势与在正常小鼠睾丸中所见基本相同.结论 新生小鼠睾丸组织异体异位移植到免疫缺陷小鼠体内后,间质细胞4种受试基因的表达趋势与在正常小鼠中的表现基本相同.     

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     

Fas配体阳性睾丸细胞和环孢素A对移植胰岛细胞存活起协同保护作用

目的　探讨Fas配体 (FasL)阳性睾丸细胞与胰岛细胞共移植后联用环孢素A(CsA)对移植胰岛细胞存活的协同保护作用。　方法　将同种大鼠胰岛细胞与睾丸Sertoli细胞同侧或异侧共移植于 4 1只糖尿病SD大鼠受体肾包膜下。实验大鼠分 7组 ,术后酌用CsA ,观察各组大鼠移植物存活情况。　结果　单纯胰岛细胞移植 (对照组 )后胰岛细胞的平均存活期为 (4 6± 1 1)d ,加用CsA存活期明显延长至 (2 1 8± 4 7)d(P <0 0 1)。与 1× 10 7个睾丸细胞同侧共移植的胰岛细胞平均存活期超过 (5 7 5± 4 0 )d ,但如移植前先封闭睾丸细胞表达的FasL后 ,移植的胰岛细胞平均存活期缩短为(5 8± 2 6 )d。胰岛细胞与 1× 10 5个睾丸细胞分别共移植于两侧肾包膜下 ,术后联用CsA ,胰岛细胞的平均存活期超过 (5 5 0± 6 5 )d ,与 1× 10 7个睾丸细胞同侧共移植的存活期相近 ,但比对照组或CsA组则显著延长 (P <0 0 1)。当胰岛细胞与 1× 10 6个睾丸细胞分别共移植且不用CsA时 ,胰岛细胞存活期平均仅为 (11 5± 3 1)d ,但仍较对照组延长 (P <0 0 5 )。　结论　表达FasL的睾丸细胞与CsA联用后可通过不同机制抑制胰岛细胞移植排斥反应而起到全身的协保护作用。     

黄芪注射液对镉诱导大鼠精子畸形的拮抗作用

目的 :观察黄芪注射液拮抗氯化镉诱导的大鼠精子畸形作用。　方法 :将 30只SD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 组大鼠腹腔黄芪注射液 5g/ (kg·d)和 10g/ (kg·d) ,Cd组和CP组大鼠分别腹腔注射等量蒸馏水作对照 ,之后连续 2 1dA1组、A2 组和Cd组大鼠分别同时腹腔注射氯化镉溶液 [0 .2mg/ (kg·d) ]。CP组大鼠给予 5 0mg/ (kg·d)腹腔注射染镉后第 2 2d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。　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 .6 8±1.19)、(4 9.0 1± 8.78)× 10 6/g、(10 .2 5± 2 .30 )× 10 6/ (g·d)、(4 7.5 1± 2 2 .5 1)× 10 6/ml]明显高于Cd组 [(3.11± 0 .16 )、(37.5 9± 10 .6 3)× 10 6/g、(5 .31± 0 .32 )× 10 6/ (g·d)、(10 .89± 2 .4 5 )× 10 6/ml](P <0 .0 5或P <0 .0 1) ;A2 组大鼠精子畸形率 [(7.0 4± 0 .12 ) % ]明显下降 ,与Cd组 [(17.81± 1.5 5 ) % ]相比 ,差异有极显著性 (P <0 .0 1)。　结论 :黄芪注射液可拮抗氯化镉对生殖系统的损害作用 ,对防护     

邻苯二甲酸二-(2-乙基)己酯致小鼠隐睾睾丸和附睾的组织病理学改变

目的 :探讨邻苯二甲酸二 (2 乙基 )己酯 (DEHP)引起的小鼠隐睾睾丸和附睾的组织病理学改变。　方法 :妊娠KM小鼠 4 0只 ,随机分成 5组 ,分别为正常对照组 8只、玉米油对照组 8只、己烯雌酚 (DES)组 8只、DEHP低剂量组 [DEHP 10 0mg/ (kg·d) ]9只和DEHP高剂量组 [DEHP 5 0 0mg/ (kg·d) ]7只。自妊娠第 12d开始到分娩后 3d ,分别持续经口给予DEHP 10 0mg/ (kg·d)、5 0 0mg/ (kg·d)和DES 10 0 μg/ (kg·d)及玉米油 ,观察仔代雄小鼠的隐睾发生率及隐睾睾丸和附睾的组织病理学改变。　结果 :DEHP 5 0 0mg/ (kg·d)组染毒小鼠的隐睾发生率显著增高 ,睾丸和附睾的体积明显减小、重量减轻 ;睾丸生精上皮发育明显异常 ,精曲小管变薄、萎缩 ,间质细胞异常增生 ,电镜下其隐睾精曲小管上皮和间质细胞均出现明显的超微结构改变。同时附睾管腔中的精子数显著减少甚至缺乏。　结论 :高剂量 [5 0 0mg/ (kg·d) ]DEHP可能具有与DES类似的作用 ,是一种诱发隐睾的重要因子。小鼠在孕期及哺乳期接触DEHP后可引起雄性仔鼠性分化异常 ,诱导隐睾发生、睾丸生精上皮损害和生精过程障碍 ,从而对雄性仔鼠生育力产生不利影响。以上作用存在明确的量 效关系。     

白细胞介素4在输注供体肝脏非实质细胞诱导免疫耐受中的作用

目的　探讨皮肤移植受体在输注供体肝脏非实质细胞 (NPC)诱导免疫耐受过程中 ,白细胞介素 4(IL 4)所起的作用。方法　将 2× 1 0 7个C3H/He (C3H)小鼠的肝脏非实质细胞通过尾静脉输入C57BL/ 6(B6)小鼠的体内 ,48h后B6小鼠腹腔注射环磷酰胺 2 0 0mg/kg ,1 8d后接受C3H小鼠皮片的移植 ,分别于NPC细胞输注前及输注后 7、1 8、30、60d采血监测IL 4水平的动态变化。结果　 1 5只接受皮片移植的小鼠 ,其皮片存活时间显著延长 ,皮片平均存活时间 (70± 1 7.2 )d ,正常对照组其移植皮肤的平均存活时间仅 (1 0± 0 .4)d ,用Elisa法检测NPC输注后不同时期IL 4水平的动态变化发现 ,B6小鼠体内血清IL 4水平逐渐升高 ,在长期耐受的小鼠中 ,其水平增加尤其显著 ,甚至成倍的提高。结论　IL 4水平的升高对于诱导和保持免疫耐受起着重要的作用     

AAPH体外诱导小鼠睾丸间质细胞氧化应激模型建立与评价

目的:利用偶氮二异丁脒盐酸盐(AAPH)作为刺激源,建立体外小鼠睾丸间质细胞(LCs)氧化应激损伤模型,并进行生理功能评价. 方法:实验一将小鼠睾丸间质细胞(TM3)随机分为6组,分别加入0、1、5、10、50和100 mmol/L AAPH,分别作用4、8、24 h.测定TM3细胞活性(MTS法)、衰老情况(β-半乳...     

交变磁场照射对小鼠睾丸生殖功能的影响

目的:探讨交变磁场的物理作用与生物学效应的关系,研究磁场对小鼠睾丸生殖功能的影响。方法:30只ICR雄性小鼠随机均分为正常对照组、X线照射组、弱磁场(1000Hz)组、强磁场(2000Hz)1h组和强磁场2h组。照射后7d处死小鼠,分析附睾精子活率,对睾丸组织切片行HE染色观察睾丸病理组织变化,以Johnsen评分标准,评定睾丸变化积分。结果:精子活率正常对照组为(42.37±10.24)%,X线照射组为(39.00±12.35)%,弱磁场组为(36.00±17.28)%,强磁场1h组为(10.72±5.67)%,强磁场2h组为(4.44±2.87)%,强磁场1h组和2h组与正常对照组比较,精子活率明显下降(P<0.01)。睾丸病理变化Johnsen评分随着交变磁场的加大和时间延长积分越低,睾丸损伤越严重。结论:强、弱磁场均引起小鼠睾丸功能破坏,导致生精小管损伤,间质、间质细胞损坏,基膜增厚,腔内生精细胞排列紊乱,凋亡、坏死和脱落增多,导致无精子发生。     

小鼠单侧睾丸损伤后环孢素A对Fas系统的影响

目的 :研究昆明小鼠 (KM小鼠 )单侧注射冰乙酸致睾丸损伤后环孢素A(CsA)对对侧睾丸生精功能和Fas系统表达的影响。　方法 :6 0只KM小鼠随机分为 4组 :A组为对照组 ,B组为单侧睾丸损伤组 ,C组为单侧睾丸损伤后 6h切除损伤睾丸组 ,D组为单侧睾丸损伤后 6h内开始腹腔注射CsA组。 4周后取对侧附睾尾 ,计数精子及其活率 ,对侧睾丸作石蜡切片苏木精 伊红染色和免疫组化链霉素抗生物素蛋白过氧化物酶连结 (SP)法检测Fas和FasL的表达。　结果 :D组附睾尾精子和活率计数显著高于B组 (P <0 .0 5 ) ,D组的FasL和Fas较B组显著降低 (2 4 .3± 7.0vs37.8± 5 .8和 17.8± 4 .3vs32 .4± 3.6 ,P <0 .0 5 )。　结论 :KM小鼠单侧睾丸损伤后CsA可以通过抑制Fas和FasL的表达 ,降低生精细胞凋亡 ,维持生精功能的稳定     

Isolation method of Leydig cells from mature male Djungarian hamsters (Phodopus sungorus) and their steroidogenic activity in vitro

For studies addressing the functions of Leydig cells, isolated cells are often better suited than intact animals. Here, the isolation procedure of Leydig cells from adult male Djungarian hamsters (Phodopus sungorus) is described. Cells were isolated using a procedure involving enzymatic dissociation and Percoll-gradient centrifugation. For each experiment, approximately 4.4 x 10(6) Leydig cells from six animals were obtained. The cells showed high steroidogenic responsiveness to physiological (ovine luteinizing hormone (oLH) and human chorionic gonadotropin (hCG)) and nonphysiological (forskolin) stimuli in vitro. Approximately 98% of cells were viable as assessed by trypan blue exclusion, and the purity varied from 80 to 95% as tested by 3 beta-hydroxysteroid dehydrogenase activity. Leydig cells were also identified by a bright yellow halo under phase-contrast microscopy. They contained numerous lipid droplets and showed round nuclei and prominent nucleoli. The cells responded to oLH, hCG and forskolin with an increased testosterone production in a dose-dependent manner. Dose-response curves in these studies suggest that Leydig cells of Djungarian hamsters undergo desensitization, probably due to down regulation of their LH/CG receptors.     

Biochemical and histochemical studies on the pituitary-testicular axis in obese (C57B1/6J-ob/ob) mice

Male C57Bl/6J-ob/ob mice (4 months old) and their homozygous lean controls were compared with respect to pituitary LH secretion and functional parameters of purified Leydig cells in vitro. Compared with controls, obese mice showed reductions in the following parameters: Plasma testosterone levels (reduced by 57%), hCG-stimulated testosterone formation in vitro (by 31%), conversion of progesterone to androgens by Leydig cells (by 39%), and GnRH-stimulated LH secretion (by 26%). Lipid accumulation and a 37% decrease in naphthylesterase activity in the Leydig cells as well as hyperplasia of pituitary gonadotrophs were observed histochemically in obese mice. The changes in testicular endocrine function in obese mice are interpreted as consequences of pituitary dysfunction.     

Experimental cryptorchidism in the adult mouse. III. Qualitative and quantitative electron microscopic morphology of Leydig cells.

The morphology of Leydig cells of control and 28-day-old cryptorchid mice was studied by electron microscopy and stereologic techniques. Leydig cell profiles of control mice were larger in section when compared to cryptorchid mice, but no differences were observed in the distribution of organelles in Leydig cells in the two groups. Quantitatively, the absolute volumes of smooth endoplasmic reticulum (SER), rough endoplasmic reticulum (RER), mitochondria, lysosomes, multivesicular bodies, peroxisomes, cytoplasmic matrix, nucleus, lipid droplets, membrane whorls, ribosomal aggregates, and annulate lamellae per Leydig cell were reduced significantly after 28 days of cryptorchidism. However, the absolute volumes of these organelles per testis were not significantly different between control and cryptorchid mice, due to the increase in Leydig cell number per testis in the cryptorchid testis, compared to the controls, except that the absolute volume of Golgi per Leydig cell was not significantly different between control and cryptorchid rats, but the absolute volume of Leydig cell Golgi was significantly lower in control rats. Based on these results, we conclude that, morphologically, a 28-day cryptorchid mouse Leydig cell clearly approximates a "half unit" of a control Leydig cell.     

Induction of hyperacute rejection of skin allografts by CD8+ lymphocytes

BACKGROUND: Second-set rejection is generally regarded as a phenomenon mainly mediated by humoral cytotoxic antibodies, although a few discordant data have been presented. In the reported experiments, we have taken advantage of the absence of production of specific cytotoxic alloantibodies contrasting with the normal development of transplantation cellular immunity, in two murine models: chimeric mice and RAG mice. METHODS: Chimeras (BALB/c-->CBA) were obtained by transplantation of 2x10(7) fetal liver cells from BALB/c (H-2d) mice to lethally irradiated CBA (H-2k) mice. After hyperimmunization with third-party C57/ BL6 (B6) (H-2b) skin transplants and with injections of 2x10(7) B6 spleen cells, antibody production, and skin graft survival were analyzed. To identify further the factors or cells responsible for accelerated rejection of B6 skin transplants in hyperimmunized chimeras, transfer experiments were carried out involving the injection of serum, whole spleen cells, spleen T cells, spleen CD8+ T cells or spleen CD4+ T cells from chimeras into BALB/c mice that had received 6 Gy irradiation. The recipient mice were then grafted with B6 skin. Similarly, the immunodeficient RAG mice were used to construct a model of recipient animals with anti-H-2d hyperimmunized B6 T cells in the total absence of antibody. RESULTS: In chimeras, anti-B6 cytotoxic antibodies were not detectable in any of hyperimmunized chimeric mice, yet accelerated rejection of B6 skin transplant occurred: a graft survival of 8.6+/-0.5 days (d), comparable to 8.9+/-0.8 d survival in CBA control mice subjected to the same hyperimmunization procedure, and significantly shorter than that in nonhyperimmunized (BALB/c-->CBA) chimeras (11.6+/-0.5 d) or in non-hyperimmunized CBA control mice (12.1+/-0.6 d). High titers of anti-B6 cytotoxic antibodies were present in the serum of hyperimmunized CBA control mice. In transfer experiments, the graft survival was over 14 d in mice treated with irradiation alone, with irradiation + serum or with irradiation + CD4+ T cells. It was significantly shorter in mice treated with irradiation + whole spleen cells, with irradiation + T cells or with irradiation + CD8+ T cells (8.9+/-0.8 d). Similarly, in immunodeficient RAG mice, reconstitution of the T cell compartment with T cells from hyperimmunized B6 mice led to accelerated rejection of BALB/c skin allografts (11.4+/-1.1 d vs. 18.8+/-0.8 d when T cells were provided by nonimmunized mice). In a second transfer of cells from these reconstituted RAG mice into naive RAG mice, CD8+ T cells were shown to induce accelerated rejection of skin allografts (12.0+/-0.6 d) whereas CD4+ T cells were much less efficient (16.5+/-0.1 d). CONCLUSION: These data indicate that T cells, and especially the CD8+ subset, can be responsible for second-set rejection in the absence of anti-donor antibodies in chimeric and RAG mouse models. These sensitized CD8+ T cells are also likely to play an important role in normal mice, in addition to that of cytotoxic antibodies.     

The pituitary-testicular axis in the streptozotocin diabetic male rat: evidence for gonadotroph, Sertoli cell and Leydig cell dysfunction

The pituitary-testicular axis was investigated in the streptozotocin diabetic male rat to determine the relationship between hormonal alterations and Leydig cell morphology. Male Wistar rats (60 days of age) were made diabetic with a single intravenous injection (65 mg/kg body weight) of streptozotocin and were studied with non-diabetic controls at 7–11, 14, 21, and 28 days after treatment. The observations on these animals were compared to those from diabetic rats (28 days) treated with 1–2 U protamine zinc insulin/100 mg body weight. Rats with untreated diabetes had hyperglycaemia and weighed less than controls. Untreated rats also had low plasma testosterone levels associated with decreased prostate and seminal vesicle weights. Insulin treatment partially or completely prevented these abnormalities. Studies of testosterone production y perfused testis suggested Leydig cell insensitivity to LH as one possible cause of decreased androgen blood levels in diabetic rats. Lipid droplets not found in Leydig cells of normal rats were seen in diabetic animals. Plasma FSH levels were also low in untreated diabetic rats but were normal in insulin     

Alloantigen-specific prolongation of allograft survival in recipient mice treated by alloantigen immunization following ultraviolet-B irradiation

It is well documented that ultraviolet (UV) radiation present in sunlight suppresses immune responses. However, the majority of studies documenting the immunosuppressive effects of UV irradiation have been carried out in animals exposed to UV irradiation before immunization. Here, we report that recipient mice exposed to UV irradiation 7 days after immunization with a donor alloantigen exhibited prolongation of allograft survival in an alloantigen-specific manner. Recipient mice (H-2(b)) intravenously immunized with 2 x 10(7) allogeneic spleen cells (H-2(b/d)) 7 days before UV irradiation (40 kJ/m(2)) showed prolonged survival of allografts presenting the alloantigen used for sensitization (H-2(b/d)), but not third-party allografts (H-2(b/k)). Adoptive transfer experiments revealed that CD4(+) T cells in UV-irradiated recipients were responsible for this prolongation. CD4(+) T cells that could transfer the suppression produced large amounts of interleukin (IL)-10, but not IL-4. The effect of UV irradiation on alloantigen-specific immunosuppression was cancelled by administration of an anti-IL-10 monoclonal antibody. These results indicate that UV irradiation given after alloantigen immunization induces alloantigen-specific type 1 regulatory T cell-like regulatory T cells that prolong allograft survival and imply that the difficulties associated with predicting donor-related organ availability in transplantation can be dealt with, given the effectiveness of UV irradiation after immunization.     

Effects of Oestrogens and FSH on LH Stimulation of Steroid Production by Testis Leydig Cells from Immature Rats

Hypophysectomy of immature male rats results after 5 days in a decreased production of testosterone by isolated testis Leydig cells in response to LH. The LH responsiveness of the Leydig cells can be partly restored by treatment of the hypophysectomized rats with FSH. In continuation of previous reports on this subject ( Steroids 28 (1976) 847; and 30 (1978) the following conclusions were derived from the results in the present paper:
1. After hypophysectomy of immature male rats the production of testosterone (T) as well as of 5-pregnenolone (Δ5P) by isolated Leydig cells in response to LH is reduced.
2. Daily administration of FSH after hypophysectomy restores the Δ5P production in response to LH almost completely, but has a much smaller effect on the restoration of T production.
3. Administration of oestradiol benzoate (E₂B) together with FSH has no effect on the restoration of LH-stimulated Δ5P production, but causes a reduction of T production, when compared with Leydig cells from animals treated with FSH only.
4. Treatment of intact immature rats with E₂B results in a decreased production of T and an increased production of Δ5P in isolated Leydig cells.
5. From experiments with labelled pregnenolone it appears that E₂B and diethylstilboestrol (DES) inhibit the 17α-hydroxylase activity of Leydig cells from intact as well as from hypophysectomized rats. This results in a reduced conversion of pregnenolone to C1:)-steroids and in increased production of 3α-hydroxy-5α-pregnan-20-one from δ5P.
6. The observed effects of FSH and E, were similar within a dose range of 100–10000 ng LH per 106 Leydig cells.     

Exposure to diesel exhaust affects the male reproductive system of mice

Several recent reports have suggested that sperm count and quality in normal men are declining. Various environmental chemical compounds may affect the male reproductive system. We propose here that diesel exhaust is an environmental pollutant with the potential to influence male reproductive function. Ultrastructural changes were observed in Leydig cells of mice exposed to diesel exhaust (0.3 mg diesel exhaust particles (DEP)/m3 through the airway, 12 h daily, up to 6 months) and reduction in LH receptor mRNA expression in Leydig cells was observed at a concentration of 1 mg DEP/m3. Daily sperm production per gram of testis dose-dependently decreased with exposure to DE for 6 months; 29%, 36%, and 53% reductions were observed at 0.3, 1.0, and 3.0 mg DEP/m3, respectively. A no-observed-adverse-effect level (NOAEL) was observed with approximately 30 micrograms DEP/m3, which is lower than the WHO-recommended limit.