Effect of prenatal exposure to diethylstilbestrol on gubernacular development in fetal male mice

AIM: To study the effect of prenatal exposure to diethylstilbestrol (DES) and the role of actin and proliferating cell nuclear antigen (PCNA) on testicular gubernaculum development in fetal male Kunming mice. METHODS: Pregnant mice were randomly assigned to 6 groups and injected with DES subcutaneously from gestational day 9 (E9) to day 17 (E17) at doses of 0, 25, 50, 100, 200 microg.kg-1.d-1 in 0.2 mL dimethyl sulfoxide (DMSO). On E17 they were sacrificed and fetuses quickly removed for fixation. Male fetuses were sliced on serial coronal plane. Histological changes were observed under the light microscope (LM) and ultrastructural changes with the scanning and transmission electron microscopes (SEM and TEM). The expression intensity of actin and PCNA in the gubernacula was quantitated by immunohistochemistry. RESULTS: The mortality of the fetuses was higher in the DES-treated groups than that in the DMSO and saline controls (P<0.05). Under LM the gubernacula were seen to be poorly developed with smaller bulbs. On SEM the bulbs lose the clear demarcation between the mesenchymal inner core and the muscular outer layer and looked like a small cone instead of the normal cylindrical appearance. On TEM there were some smaller disordered myofibrils and sparse cytoplasmic organelles in the gubernacular muscular cells of the treated groups. The expression intensity of actin and PCNA in the gubernacula was significantly weaker in the treated groups than that in the DMSO and saline controls (P<0.05). CONCLUSION: DES induces underdevelopment of the gubernacula in a dose-dependent manner in fetal male mice and down regulates the actin and PCNA expression.     

Early breast development in girls after prenatal exposure to non-persistent pesticides

Contemporary American and European girls experience breast development at earlier ages compared with 15-20 years ago. Alterations in BMI alone cannot account for these changes. Several currently used pesticides possess endocrine disrupting properties and may interfere with reproductive development, but human data are sparse. We examined girls whose mothers worked in greenhouses in the first trimester of pregnancy to assess the long-term effects of prenatal pesticide exposure on puberty. Mothers were prenatally categorized as exposed or unexposed to pesticides. We studied the offspring of these greenhouse workers, and evaluated the anthropometry, pubertal staging in the girls, and blood samples were drawn at 3 months of age (n = 90) and again once at school age (6-11 years, n = 83). No clinical and biochemical differences were found between the exposed and unexposed girls at 3 months of age. Mean onset of B2+ was 8.9 years (95% CI: 8.2; 9.7) in prenatally exposed girls, compared with 10.4 years (9.2; 17.6) in the unexposed (p = 0.05), and 10.0 (9.7-10.3) years in a Danish reference population (p = 0.001). Exposed girls had higher serum androstenedione levels (geometric means: 0.58 vs. 0.79 nmol/L, p = 0.046) and lower Anti-Müllerian Hormone (AMH) compared with the unexposed (geometric means: 16.4 vs. 21.3 pmol/L, p > 0.05) and the reference group (20.2 pmol/L, p = 0.012). Levels of testosterone, estradiol, prolactin, FSH, LH, SHBG, DHEAS, DHT, Inhibin A and Inhibin B did not differ between the groups. In conclusion, our findings suggest that prenatal exposure to currently approved pesticides may cause earlier breast development in girls. This association appeared not to be because of changes in gonadotropins, but rather to higher androgen levels, which indirectly may increase oestrogens through aromatization. In addition, lower serum AMH levels indicated a reduced pool of antral ovarian follicles. The long-term consequences of our findings with regard to establishment of future reproductive function still remain unknown.     

Behavioral effects of chronic exposure to low concentrations of halothane during development in rats

Long-term behavioral effects of chronic exposure to low concentrations of halothane were evaluated in rats exposed to low (12.5 ppm) concentrations from day 2 of conception until either 30 (halothane-30) or 60 (halothane-60) days after birth. Rats similarly treated but not exposed to halothane served as controls. When these rats were tested for radial arm maze exploration as adults (1 yr old) both exposure groups showed significant deficits compared with controls. The halothane-treated rats entered significantly fewer arms before reentering an arm (entries-to-repeat). At 55 days of age, in the spontaneous alternation test, response speed was significantly slower than controls in both halothane-30 and halothane-60 rats. This effect was not seen in rats more than 55 days old. Replicating previous results, the halothane-60 rats showed deficits in learning a light-dark discrimination. This deficit was not seen with halothane-30 rats, indicating that continued halothane exposure during the 30- through 60-day period was necessary for inducing a noticeable long-term learning deficit. The results show that chronic exposure of rats to low concentrations of halothane during development results in subsequent behavioral alteration, and that termination of halothane exposure at 30 days of age rather than at 60 days of age avoids some of the signs of behavioral impairment.     

环孢菌素A逆转利多卡因对七氟醚后处理心肌保护的作用

Objective To study the effects of lidocaine on sevoflurane postconditioning-induced cardioprotection.Methods Ischemic status was kept for 40 rain in isolated perfused rat hearts followed by 1 h of reperfusion.Sevoflurane(3%) was administered at the beginning of reperfusion for 15 rain with or without lidocaine (20 μg/ml) perfusion.The direct mitochondrial permeability transition pore (MPTP) inhibitor Cyclosporin A (CsA,0.2 μmol/L) was co-administered in the presence or absence of lidocaine.LVDP,LVEDP,+dp/dtmax were recorded and infarct size was measured with TTC staining.Results Sevoflurane postconditioning significantly improved the recovery of ischernic myocardial function and decreased the infarct size of rat hearts (P＜0.05),which was abolished by lidocaine perfusion.The inhibition of lidocaine on sevoflurane posteonditioning effect was reversed by CsA.Conclusion Sevoflurane postconditioning effectively protects myocardium against ischemia/reperfusion injury,and higher concentration of lidocaine inhibits this protective effect by opening MPTP.     

Reproductive and teratogenic effects of lidocaine in Sprague-Dawley rats

The reproductive and teratogenic effects of lidocaine were studied in 155 Sprague-Dawley rats chronically implanted with osmotic minipumps. Three different lidocaine doses were used as follows: a low dose (100 mg X kg-1 X day-1); an intermediate dose (250 mg X kg-1 X day-1); and a high dose (500 mg X kg-1 X day-1). In the low and intermediate dose groups, lidocaine was administered for two weeks before mating and throughout pregnancy to evaluate reproductive and teratogenic effects. In the high-dose group, lidocaine was administered from days 3 to 17 of pregnancy to evaluate teratogenic effects. On day 21, cesarean section was performed and all of the 1,040 offspring, including those from control and positive control (retinoic acid) groups, were preserved and subsequently examined microscopically to detect external, visceral, and skeletal abnormalities. None was found in lidocaine-treated groups; reproductive indices also were normal. The only treatment effect was a reduction in mean fetal weight in the high-dose group. In a subsequent experiment this was found to be secondary to slightly delayed fetal development. It is concluded that lidocaine is devoid of significant adverse reproductive and teratogenic effects in Sprague-Dawley rats.     

青春期己烯雌酚暴露对大鼠睾丸发育及功能的影响

目的 探讨青春期己烯雌酚(diethylstilbestrol,DES)暴露对SD(Sprague-Dawley)大鼠睾丸发育及功能的影响.方法 35日龄雄性SD大鼠90只,随机分为DES 0.01、0.1、1.0、10.0 μg·kg-1·d-14个实验组和1个对照组(编码为Bda、BDb、BDc、BDd和BC组,每组18只).于青春期,即出生后第36天(postnatal day 36,PND 36)至PND 49,实验组每日皮下注射相应剂量的DES,共14 d,对照组仅注射溶媒.于青春期晚期(PND 50)、性成熟后(PND 64)和成年期(PND 130)分3批(每批6只)处死各组大鼠取材,测定睾丸重量,观察比较睾丸组织形态学变化,分析PND 130大鼠附睾尾精子质量.结果 PND 50时,BC、Bda、BDb、BDc和BDd组单侧睾丸重量分别为(1.26±0.13)、(1.23±0.20)、(0.99±0.15)、(0.85±0.23)和(0.60±0.04)g,其中BDb、BDc和BDd组均较BC组减轻(P＜0.05);与BC组比较,BDb组仅有少数生精小管生精上皮中的细胞数目稍减少,BDc和BDd组生精小管发育较差、生精上皮中细胞数目减少、精子发生阻滞、间质细胞发育幼稚,其程度随DES暴露剂量增加而加重.PND 64时,BC、Bda、BDb、BDc和BDd组单侧睾丸重量分别为(1.54±0.14)、(1.55±0.17)、(1.52±0.11)、(1.37±0.14)和(0.88±0.15)g,其中BDc和BDd组均较BC组减轻(P＜0.05);BDc和BDd组睾丸组织形态学改变与PND 50时类似,但较PND 50时有所改善.PND 130时,各实验组与对照组比较,单侧睾丸重量差异无统计学意义(P＞0.05),睾丸组织形态学改变未见明显差异;BC、Bda、BDb、BDc和BDd组大鼠附睾尾精子密度分别为(71.00±14.85)、(69.00±23.98)、(67.00±13.52)、(31.67±12.94)和(18.83±6.68)×106/ml,其中BDc和BDd组精子密度均较BC组明显降低(P＜0.01);与BC组比较,BDd组精子活动率下降(P＜0.01),BDb、BDc和BDd组A级精子比例降低(P＜0.05),BDd组B级精子比例降低(P＜0.01).结论 青春期小剂量DES(0.01μg·kg-1·d-1×14 d)暴露对SD大鼠睾丸发育及功能无明显影响,大剂量DES(1.0～10.0 μg·kg-1·d-1×14 d)暴露对大鼠睾丸发育及功能具有明显的近期(PND 50和PND 64)和较远期(PND 130)毒性作用,该毒性作用随DES暴露剂量增加而加重,随鼠龄增长而逐渐减退,其机制可能与间质细胞和支持细胞的发育及功能受损密切相关.     

Multiple head injuries in rats: effects on behavior

BACKGROUND: Evidence suggests that mild head injuries in humans can result in cumulative damage. No investigation to date has considered the effects of multiple subacute mild head injuries in an animal model. METHODS: Forty-one male Long-Evans hooded rats were trained in a Morris water maze. All animals were fitted with a hollow intracranial screw. Concussions were generated using a fluid percussion device. Animals were then evaluated in the water maze until performance returned to baseline. Control animals received no concussions. The remaining animals were randomized to receive one, two, or three concussions. Animals were allowed to return to baseline after each concussion and were then killed. Motor performance was evaluated on a balance beam both before and after concussions. RESULTS: After one concussion, 85% of animals showed performance deviation from baseline as measured by time to reach the platform, returning to baseline within a mean of 14.0 trials. After two concussions, 48% of animals showed deviation, with a mean return to baseline of 6.8 trials. After three concussions, 25% of animals showed deviation, with a mean return to baseline of 2.3 trials. Of postconcussive animals, 42% developed new inconsistent baseline levels of performance. Balance beam performance was unaffected. CONCLUSION: Multiple concussions cause immediate transient impairment in spatial recognition and have extended effects on baseline performance in rats. Motor performance is not affected.     

Biphasic effects of postnatal exposure to diethylhexylphthalate on the timing of puberty in male rats

Phthalate esters such as di(2-ethylhexyl)phthalate (DEHP), which are commonly found in cosmetics and in flexible plastics distributed by the food, construction, and medical products industries, have been classified as anti-androgens. High-dose DEHP exposure in utero is associated with decreased androgen levels. However, when administered after birth, low doses of DEHP (eg, 10 mg/kg body weight) may stimulate androgen production. In the present study, the potential of phthalate exposure to advance or delay the timing of puberty was assessed. Male Long-Evans rat pups were chronically subjected to low or high doses of DEHP, with the androgen-driven process of preputial separation serving as an index of pubertal timing. Rats were treated with 0, 10, 500, or 750 mg/kg body weight DEHP for 28 days starting at day 21 postpartum. The average age at which the animals completed preputial separation was measured in each group. The age of preputial separation was 41.5 +/- 0.1 days postpartum in controls (vehicle). The 10 mg/kg DEHP dose advanced pubertal onset significantly to 39.7 +/- 0.1 days postpartum, whereas the 750 mg/kg DEHP dose delayed pubertal onset to 46.3 +/- 0.1 days postpartum. The 10 mg/kg DEHP dose also significantly increased serum testosterone (T) levels (3.13 +/- 0.37 ng/mL) and seminal vesicle weights (0.33 +/- 0.02 g) compared with control serum T (1.98 +/- 0.20 ng/mL) and seminal vesicle weight (0.26 +/- 0.02 g), while the 750 mg/kg dose decreased serum T (1.18 +/- 0.18 ng/mL) as well as testes and body weights. Direct action of the DEHP metabolite, monoethylhexylphthalate (MEHP), on Leydig cell steroidogenic capacity was investigated in vitro. MEHP treatment at a low concentration (100 microM) increased luteinizing hormone-stimulated T production, whereas 10 mM concentrations were inhibitory. In conclusion, data from the present study indicate that DEHP has a biphasic effect on Leydig cell function, with low-dose exposure advancing the onset of puberty. High doses of DEHP, which are anti-androgenic, may also be outside the range of real environmental exposure levels.     

The effects of tumescent solutions containing lidocaine and epinephrine on skin flap survival in rats

The effects of tumescent solutions consisting of lidocaine and epinephrine on skin flap survival in rats were studied. Dorsal skin flaps of rats were infiltrated using lidocaine (1%) with epinephrine in concentrations of 1:100,000, 1:200,000, 1:400,000, and 1:800,000 prior to elevating flaps of the different experimental groups. The solutions were applied intradermally or subcutaneously, and the flaps were raised "immediately" or "delayed" after injection in the different groups. Control flaps were infiltrated by lidocaine (1%) only. The survival of the flaps was assessed on the seventh day after the operation. As a result, the flaps showed higher necrosis rates in the groups injected by lidocaine with epinephrine in concentration of 1:100,000 and 1:200,000 than of the other experimental or all control groups (P < 0.01). In conclusion, lidocaine with epinephrine in concentrations of 1:400,000 and 1:800,000 was found safe on skin flap survival for tumescent technique in rats.     

Gestational exposure to atrazine: effects on the postnatal development of male offspring

Atrazine is an herbicide used worldwide to control grasses and weeds. Previous studies have shown that, depending on atrazine's administered dose, exposure of male rats during the early postnatal or peripubertal periods can result in alterations in endocrine function. The gestational period is particularly vulnerable to environmental agents; however, the possible effects of atrazine exposure during this period have received only limited attention. Herein we examine the dose effects of atrazine exposure during Sprague-Dawley rat gestation on the postnatal development of male offspring. Pregnant dams were treated by oral gavage with atrazine at 0 to 100 mg/kg/d from gestational day 14 to parturition. Thereafter, neither the pups nor the dams received atrazine. Atrazine had no effect on the number of live births per dam. Neonatal pup survival was affected, however, with increased pup death seen at doses of 10 mg/kg/d and higher. There was no effect of atrazine on the testosterone concentration within the testes of newborn pups. Anogenital distance, an androgen-dependent process, decreased from the control level at the 75 and 100 mg/kg/d doses, with the decrease reaching significance at 100 mg/kg/d. Preputial separation, also an androgen-dependent process, was delayed significantly compared with that in controls in response to the 50 and 100 mg/kg/d doses. At postnatal day 60, serum testosterone concentrations were reduced significantly from controls in the 50 to 100 mg/kg/d groups. However, these decreases had little effect on seminal vesicle or ventral prostate weights. These results, taken together, are suggestive of antiandrogenic effects of gestational atrazine exposure on male offspring, although for most parameters, the doses used in this study are unlikely to be experienced under any but experimental conditions.     

The effects of maternal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on testicular steroidogenesis in infantile male rats

Exposure of adult male animals to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) decreases serum androgen concentrations. Reduction in androgen levels after maternal exposure has also been reported, but these results have not been reproduced. We have earlier shown that TCDD stimulates rather than inhibits testosterone synthesis in the prenatal rat testis. The aim of the present study was to elucidate in utero-induced effects of TCDD on testicular steroidogenesis in the 14-day-old infant rats. At that time the foetal Leydig cell population is still the prevailing source of androgens. Pregnant Sprague-Dawley dams were given a single oral dose of TCDD (0, 0.04, 0.2, or 1.0 microg/kg) on day 13 of pregnancy. On postnatal day 14, the body weight of male offspring was reduced after exposure to 1.0 microg/kg TCDD (from 33.9 +/- 1.66 g to 31.6 +/- 2.67 g). Relative testis weight, plasma testosterone, luteinizing hormone and follicle-stimulating hormone levels remained unaltered in all exposure groups. Moreover, in ex vivo incubations, testosterone and cAMP production was not affected. StAR protein level in the freshly isolated testes was increased in the 0.2 microg/kg group, and seminiferous cord diameter in the 0.04 microg/kg group. The present study confirms our earlier findings in in utero TCDD-exposed foetal testis indicating that maternal TCDD exposure does not negatively influence the developmental testosterone production of foetal type Leydig cells in rats.     

Effects of prenatal flutamide on testicular development, androgen production and fertility in rats

INTRODUCTION: To investigate the effects of blocking prenatal androgen with the anti-androgen flutamide on testicular weight (TW), seminiferous tubular diameter (STD), testicular biopsy score (TBS), testicular testosterone (TT), and fertility. MATERIALS AND METHODS: Ten pregnant Wistar albino rats were injected with flutamide (100 mg/kg) on gestational days 16 and 19. Rats injected with the solvent were used as controls. Male pups from these dams were evaluated for testicular descent on postnatal day 22. The ability of each male to produce offspring was tested when the animals were aged 130 days. The rats were killed when 180 days old, and the testes were removed. RESULTS: Sixteen of the 24 male rats (66.6%) exhibited cryptorchidism (6 unilateral, 10 bilateral) in the flutamide-treated group. Three of the rats with normally descended testicles were fertile, but none of the cryptorchid rats was fertile in flutamide-treated group. The average TW, mean STD, TBS and TT levels of the flutamide-treated rats was significantly lower than in the solvent-injected rats. However, there was no significant difference in these values between descended and undescended testes in flutamide-treated rats. CONCLUSION: Blocking of prenatal androgen with flutamide interferes with testicular development by inhibiting testicular descent, and also effects testicular morphology and function in both the descended and undescended testes of rats.     

Different duration of crowding and noise exposure effects on exploratory behavior,cellular immunity and HSP70 expression in rats

This study examined the effects of different duration of stress exposure on cellular immunity and heat shock protein (HSP) 70 expression in rats. The different durations of crowding and noise were used as different stress modes. Observation of exploratory behavior in an open‐field test was used to indicate stress level. The expression of HSP70 and T‐cell subsets in blood samples were measured in the different groups of stressed rats. The results showed that there were significant reductions in the percentages of CD3⁺ and CD4⁺ T cells in the stressed groups. The lowest point was at 2 weeks from stress. There were significant increases in the percentage of CD8⁺ in 2 day, 1 week and 2 week stress groups compared with the control group. The CD4⁺/CD8⁺ ratio was also remarkably lower in rats of the 2 day, 1 week and 2 week stress groups compared with the control group. HSP70 expression rose significantly after stress and reached a maximum after 2 weeks of stress. Open field test activity showed higher vertical movement scores at 2 weeks stress compared with the control group. Grooming scores at 2 days stress was obviously more than that of the control and there were higher grooming scores at 2 weeks and 3 weeks stress compared to the 2 day stress group. Those results showed that the different duration stress exposure affects both exploratory behavior and immune function. The results also showed that not only immune function but also HSP70 expression was mainly dependent on the duration of stress. Copyright © 2006 John Wiley & Sons, Ltd.     

Histomorphometric analysis of the epiphyseal growth plate in rats after prenatal alcohol exposure.

We studied the effect of prenatal alcohol exposure on growth in the proximal tibial growth plate in 0- and 15-day-old rats, using histomorphometric methods. Body weight and tibial length were reduced in all alcohol-exposed rats. In 15-day-old rats, these parameters were lower than in the 15-day-old controls, thus showing a persistence of the effects of ethanol. The proximal tibial growth plate showed alterations, principally in 15-day-old rats. The most notable of these was a decrease in growth plate height produced by a significant reduction in hypertrophic zone height. Likewise, there were fewer cells in this zone in alcohol-exposed rats than in controls. This work shows that prenatal ethanol exposure induces growth retardation which may be due to growth plate alterations that might reflect impaired cell function.     

利多卡因对大鼠微栓栓塞致脑损伤的保护作用

目的观察利多卡因对大鼠微栓栓塞致脑损伤的保护作用。方法雄性Wistar大鼠65 只,随机分为5组,对照组(n=8):经右侧颈动脉注射20%葡聚糖(微球载体);微栓1组(n=14)及微栓2组(n=14),分别注射600和900个微球;保护1组(n=12)和保护2组(n=13),分别注射600和900个微球,并于注射微球前30 min经股静脉注射利多卡因1.5 mg/kg负荷剂量,继以2 mg·kg-1·h-1 持续输注直至注射微球后1 h,其余组给予等量生理盐水。另取雄性Wistar大鼠4只,按上述剂量、速率及途径给予利多卡因,用于测定利多卡因血药浓度。注射微球后第1-7天进行脑卒中行为评分; 第8-12天进行水迷宫试验(包括潜伏期和有效搜索策略比率);水迷宫试验后,每组取3只大鼠,取海马制成切片,观察CA1区神经元的病理变化。结果与对照组比较,微栓组和保护组脑卒中行为评分均升高;微栓2组注射微球后第12天潜伏期延长,有效搜索策略比率降低;与微栓1组比较,保护1组在注射微球后第3、4天脑卒中行为评分降低,注射微球后第9、12天有效搜索策略比率较高; 与微栓2组比较,保护2组在注射微球后第1-4、6天脑卒中行为评分降低,注射微球后第12天潜伏期缩短,第9、12天有效搜索策略比率升高(P<0.05或0.01)。与对照组比较,微栓组栓塞侧海马CA1 区的正常神经元计数减少;缺血坏死神经元计数微栓组和保护组均升高(P<0.05或0.01),但保护组少于微栓组(P<0.05)。利多卡因血药浓度为(2.20±0.18)μg/ml。结论利多卡因对大鼠微栓栓塞引起的脑损伤有一定的保护作用。     

Microalbuminuria in adults after prenatal exposure to the Dutch famine

Maternal undernutrition during gestation is associated with an increase in cardiovascular risk factors in the offspring in adult life. The effect of famine exposure during different stages of gestation on adult microalbuminuria (MA) was studied. MA was measured in 724 people, aged 48 to 53, who were born as term singletons in a university hospital in Amsterdam, the Netherlands, around the time of the Dutch famine 1944 to 1945. Twelve percent of people who were exposed to famine in mid gestation had MA (defined as albumin/creatinine ratio >/=2.5) compared with 7% of those who were not prenatally exposed to famine (odds ratio 2.1; 95% confidence interval 1.0 to 4.3). Correcting for BP, diabetes, and other influences that affect MA did not attenuate this association (adjusted odds ratio 3.2; 95% confidence interval 1.4 to 7.7). The effect of famine was independent of size at birth. Midgestation is a period of rapid increase in nephron number, which is critical in determining nephron endowment at birth. Fetal undernutrition may lead to lower nephron endowment with consequent MA in adult life.     

利多卡因对脓毒症大鼠肺损伤的影响

目的 研究利多卡因对脓毒症大鼠肺损伤的影响.方法 选择清洁级SD成年雄性大鼠30只,2月龄,体重250～300 g.采用随机数字表法将大鼠分为三组:假手术组(S组)、盲肠结扎穿孔组(C组)和利多卡因组(L组),每组10只.S组仅打开腹腔后缝合,C组和L组采用盲肠结扎穿孔法(CLP)建立脓毒症模型.L组建立脓毒症模型后即...