中国对虾卵黄蛋白原合成部位的研究

十足目甲壳类卵巢中卵黄的来源,在过去的几十年研究中一直存在争议,内源性合成和外源性合成都有报道。本文以雌性中国对虾为实验材料,根据组织学观察确定其发育阶段,试验虾可以分为:卵巢未发育期;卵黄发生前期;初级卵黄发生期和次级卵黄发生期4个时期。从处于不同卵巢发育期对虾的卵巢和肝胰腺中提取总RNA,用RT-PCR的方法初步探讨不同发育期的中国对虾卵巢和肝胰腺中卵黄蛋白原mRNA的表达,确定是否存在卵黄蛋白原的合成功能。结果发现,从卵巢未发育期到次级卵黄发生期的卵巢和肝胰腺中都有卵黄蛋白原mRNA的表达,说明二者为卵黄蛋白的合成部位。     

Pesticide—induced reproductive dysfunction in Indian fishes

Numerous studies are available reporting the effects of pesticides on reproductive activity in Indian fishes. The majority of these reports deals with histopathological changes in gonads and endocrine glands involved in the regulation of reproduction following treatment with different pesticides. Pesticides are reported to cause degenerative changes in gonads and arrest gametogenic processes either by acting directly on the gonads or by interfering with the secretory activity of the hypothalamo-hypophyseal-gonadal/thyroid axis that regulates various reproductive events. Secretion of hormones such as gonadotropin-releasing hormone (GnRH), gonadotropin, growth hormone, adrenocorticotropic hormone (ACTH), testosterone, estrogens, 17,20β-dihydroxyprogesterone and thyroid hormones are in general lowered, leading to cessation of gametogenesis, vitellogenesis, oocyte maturation, ovulation, spermiation, etc. Adverse effects of pesticides have also been demonstrated on fecundity, fertilization, hatching, and postembryonic development. The effects are highly variable and depend on the nature, dose, and mode of application of the pesticides.     

不同倍性虹鳟卵黄蛋白原的组织定位

采用分步沉淀法提纯虹鳟卵黄蛋白,以同发育期二倍体虹鳟为对照,用免疫组化法对不同发育阶段的三倍体虹鳟性腺、肝脏、血液和肠道进行卵黄蛋白原(Vg)的细胞化学定位研究.试验结果表明:所纯化蛋白为与卵黄蛋白原具有相似免疫原性的卵黄脂磷蛋白,呈雌性特异性.性腺发育Ⅲ～Ⅴ期的二倍体雌性虹鳟,血液和肝细胞呈卵黄蛋白原阳性,Ⅳ～Ⅴ期卵巢呈卵黄蛋白原阳性,各发育期肠组织呈卵黄蛋白原阴性;性腺发育至Ⅰ～Ⅴ期的三倍体雌雄虹鳟及雄性二倍体虹鳟,其性腺、肝脏、血液和肠道组织呈卵黄蛋白原阴性.二倍体虹鳟外源性卵黄蛋白原在肝细胞内合成,经血液运送至卵巢,最终在卵母细胞中形成卵黄颗粒.由于三倍体雌性虹鳟卵巢发育受阻,无滤泡细胞分化,生殖细胞与体细胞的互作缺失,不能分泌足量的17β-E2以诱导Vg的合成,因而肝脏的Vg阴性并不能说明肝脏不具备合成Vg的能力,从卵黄发生的角度分析,卵黄蛋白原的缺乏不是导致三倍体虹鳟雌性不育的原因,而是其卵泡败育的结果.     

Effects of the herbicide LASSO MTX (alachlor 42% W/V) on biometric parameters and liver biomarkers in the common carp (Cyprinus carpio)

The aim of the study was to evaluate the effect of subchronic exposure to the herbicide LASSO MTX (alachlor 42% W/V) on biometric parameters and important liver biomarkers in the common carp (Cyprinus carpio). One year old fish were exposed for 28 days to LASSO MTX added to the tank water at concentrations of 240 and 2400 μg L⁻¹. The exposure did not affect fish biometric parameters. Glutathione-S-tranferase (GST) activity in liver (hepatopancreas) remained unchanged in exposed fish when compared to controls. However, significant induction of total cytochrome P 450 (CYP 450), ethoxyresorufin-O-deethylase (EROD) activity and elevated glutathione (GSH) in liver of exposed fish were detected.     

脊尾白虾翻译控制肿瘤蛋白基因TCTP克隆及自噬调控相关基因在卵巢发育期的表达

为深入了解脊尾白虾(Exopalaemon carinicauda)卵巢发育和卵黄蛋白原发生的分子机制,本研究克隆得到脊尾白虾翻译控制肿瘤蛋白基因TCTP,并结合自噬调控基因TCTP、Hif-1α、Beclin1和Bcl-2在卵巢发育期的表达水平,阐释脊尾白虾卵黄蛋白原合成过程中的分子调控特征。研究显示,脊尾白虾TCTP基因c DNA全长为732 bp,编码168个氨基酸,具有典型的TCTP1和TCTP2功能域以及PKC和TKⅡ等mTOR信号通路相关的磷酸化位点。同时发现,甲壳动物TCTP普遍缺乏在其他动植物中高度保守的C末端的cys残基。进化分析显示,脊尾白虾TCTP与中华绒螯蟹(Eriocheir sinensis)亲缘关系最近。4种自噬调控基因在脊尾白虾卵巢发育期的表达结果显示,肝胰腺TCTP基因从增殖期到产后恢复期呈递减趋势;肝胰腺Hif-1α、Beclin1和Bcl-2基因表达趋势相似,即从增殖期到小生长期高表达,大生长期极显著下降,表达量最低(P0.01),这与外源性卵黄蛋白原的合成趋势大致相反。这些自噬调控基因可能通过自噬作用共同调节外源性卵黄蛋白原的合成。卵巢TCTP基因在小生长期表达量最高;卵巢Hif-1α基因从增殖期到产后恢复期持续升高,这与内源性卵黄蛋白原表达趋势相似;卵巢Beclin1基因在大生长期表达量最高,与脊尾白虾卵巢Ec R表达趋势相似,与卵巢Bcl-2基因表达趋势相反,这些自噬调控基因可能通过自噬作用共同促进内源性卵黄蛋白的合成。本研究表明,自噬调控基因TCTP、Hif-1α、Beclin1和Bcl-2在脊尾白虾卵巢发育时期相互协调共同作用,可能通过自噬作用调节脊尾白虾卵黄蛋白原的合成和卵巢发育。     

Disrupted sexual cycles in female grass carp (Ctenopharyngodon idella) raised in tropical conditions

In the Ivory Coast, grass carp (Ctenopharyngodon idella) have been integrated in the tilapia-based polyculture in order to increase pond productivity. In the tropical conditions prevailing there, female cycles seemed disrupted. We describe oogenesis in these conditions using histological observations, monitoring of individual cycles with intraovarian biopsies, endocrinal monitoring (development of a specific enzyme-linked immunosorbent assay (ELISA) for vitellogenin (Vg), measurement of plasma oestradiol-17β (E2) and testosterone (T)) and comparison with grass carp raised in Poland. We noted that oogenesis was blocked in all females at the migrating germinal vesicle stage, precluding ovulation or spawning without artificial induction. High rates of atypical post-vitellogenic oocytes (translucent, not filled with yolk granules) were observed in many females. Female individual cycles also displayed atypical features: cycles were sometimes (10% of females) blocked at the beginning of vitellogenesis (BV), for females displaying abnormally low E2 (0.5 ng/ml) and Vg (30 μg/ml) levels compared to “normal” females (1.4 ng/ml and 223 μg/ml, respectively). The duration of the cycles was highly variable among females (a few days to several weeks). Sexual cycles were unsynchronised (all the ovarian stages could be found for all seasons) and the number of females at the end of vitellogenesis (EV) was low (<40% most of the year). These characteristics raise problems for artificial induction of spawning in small-scale hatcheries: a large stock of broodfish is required as is regular checking of female broodstock with intraovarian biopsies to select responsive females.     

Levels of Vitellogenin in Male Japanese Medaka (Oryzias latipes) Exposed to the Chemotherapeutics: Oxytetracycline,Romet-30®, and Copper Sulfate

ABSTRACT

Japanese medaka (Oryzias latipes) were exposed to oxytetracycline and Romet-30^® at daily doses via feed of 0.06 mg, 0.11 mg, and 0.28 mg per gram body weight for 11 days, and at daily doses of 0.05 mg, 0.10 mg, and 0.15 mg per gram body weight for 5 days, respectively. In addition, fish were exposed to copper sulfate in their aquaria water at 1.85 mg/L for 1 h each day (over a span of 1 day, 7 consecutive days, or 1 day in every 3 to 5 days for a total of seven doses). Vitellogenin levels were measured in fish using a sandwich enzyme-linked immunosorbent assay (ELISA). Vitellogenin levels were not induced from exposure to either oxytetracycline or Romet-30® (P > 0.05); however, copper sulfate for 1 h on 1 day showed a decrease in vitellogenin from the control fish and other treatment groups (P = 0.007). Under these test conditions, oxytetracycline, Romet-30, and copper sulfate were not found to act in medaka as estrogenic compounds by inducing vitellogenin.     

甜菜碱对蛋鸡血清卵黄前体物水平的影响

以20周龄伊莎蛋鸡为试验对象,研究日粮添加甜菜碱对蛋鸡卵黄前体物极低密度脂蛋白VLDL和卵黄磷蛋白VTG含量的影响.结果表明日粮中添加600 mg·kg-1甜菜碱促产蛋效果最佳,全程平均产蛋率比对照组提高8.67%(P＜0.05),料蛋比下降9.02%(P＜0.05).日粮添加600mg·kg-1甜菜碱使50和70周龄蛋鸡血清VLDL含量分别提高34.61%(P＜0.05)和53.41%(P＜0.03),VTG含量提高33.74%(P＜0.01)和39.14%(P＜0.01);腹脂LPL酶活性分别下降31.0%(P＜0.01)和44.83%(P＜0.01);血清FSH水平分别提高29.40%(P＜0.05)和19.85%(P＜0.01);LH水平分别提高45.49%(P＜0.01)和26.69%(P＜0.01).结果表明,甜菜碱对蛋鸡血清卵黄前体物VLDL和VTG含量有显著影响,它是通过影响血清卵黄前体物含量来提高蛋鸡生产性能.     

Endocrine disruption and metabolic changes following exposure of Cyprinus carpio to diethyl phthalate

Diethyl phthalate (DEP) enter into aquatic environment from industries manufacturing cosmetics, plastic and many commercial products and can pose potential fish and human health hazard. This experiment evaluated effects of DEP in adult male (89 g) common carp (Cyprinus carpio) by exposing them to fractions of LC₅₀ (1/500-1/2.5) doses with every change of water for 28 days. Vitellogenin induction metabolic enzymes, somatic indices and bioaccumulation were studied on 7th, 14th, 21st and 28th day. The 96th hour LC₅₀ of DEP in fingerlings was found to be 48 mg/L. Compared to control, except increase (P < 0.01) in alkaline phosphatase activity (EC 3.1.3.1) and liver size, there was decrease (P < 0.01) in activity of acid phosphatase (EC 3.1.3.2), aspartate aminotransferase (EC 2.6.1.1), alanine aminotransferase (EC 2.6.1.2) and testiculosomatic index following exposure to 1, 5 and 20 ppm DEP. Significant (P < 0.01) dose dependant vitellogenin induction was observed with exposure of fish to 0.1, 1 and 5 ppm DEP. The bioaccumulation of DEP in testis, liver, brain, gills and more importantly in muscle tissues of fish increased significantly (P < 0.01) with increase of dose from 1 to 5 ppm. Significant interaction (P < 0.01) of dose and duration of exposure indicated that exposure period of a week to two was sufficient to bring about changes in quantifiable parameters studied. Fish exposed to 20 ppm DEP became lethargic and discolored during onset of the 4th week. This is the first report describing metabolic changes and vitellogenin induction following exposure of C. carpio to DEP dose that is as low as 1/500th fraction of LC₅₀.     

One-tenth dose of LC50 of 4-tert-butylphenol causes endocrine disruption and metabolic changes in Cyprinus carpio

Of the huge annual worldwide production (500,000 MT in 1997) of alkylphenol polyethoxylates (APEs) that are widely used as nonionic surfactants and anti-oxidants in variety of products, 60% ends up in water bodies. They undergo biodegradation to form octyl-, butyl-, and nonyl-phenols. This experiment evaluated effects of 4-tert-butyl phenol (4-TBP) in Cyprinus carpio, a projected candidate species in sewage fed fisheries. The 96th h LC₅₀ of 4-TBP was found to be 6.9 mg/L. Fishes were treated with 1/10th (0.69 mg/L), 1/5th (1.38 mg/L), and 1/3rd (2.3 mg/L) dose of LC₅₀. Whereas there was significant (P < 0.01) decrease in alkaline phosphatase [EC 3.1.3.1] and aspartate aminotransferase [EC 2.6.1.1] activity; alanine aminotranferase [EC 2.6.1.2] and acid phosphatase [3.1.3.2] (except decrease at 1/10th dose of LC₅₀) activity, vitellogenin production in muscle and hepatic- and reno-somatic indices were increased compared to control. With all the dose levels tested, testicular-somatic index (testis size) was reduced (P < 0.01) and histo-architectural changes in testicular and liver tissue were found even in group given 1/3rd dose of LC₅₀.