碱度对尼罗罗非鱼血清渗透压、离子浓度及离子转运酶活力的影响

为了探讨鱼类在碱水环境中的渗透调节机制, 将尼罗罗非鱼从淡水直接转入2 g·L^–1 和4 g·L^–1NaHCO3 碱水中进行急性胁迫试验, 分别检测胁迫后0、3、6、12、24、48、72、96 和192 h 时血清渗透压、血清Na⁺、K⁺、Cl^–和HCO3^–浓度以及鳃、肾和肠中离子转运酶碳酸酐酶(CAⅡ、CAⅣ)、碳酸氢钠协同转运载体(SLC4A4)、Cl^–/HCO3^–离子交换体(SLC26A6)活力变化。结果显示: 不同碱度胁迫下, 尼罗罗非鱼血清渗透压、离子浓度以及鳃、肾、肠中离子转运酶活力均与碱胁迫浓度呈正相关。随时间推移, 血清渗透压、离子浓度呈现先上升、后下降的变化趋势, 24 h 达到峰值; 鳃、肾和肠中CAⅡ、CAⅣ、SLC4A4、SLC26A6 活力均呈现先短时间降低、后升高、再降低并趋于稳定的趋势。研究表明, 尼罗罗非鱼具有一定的碱环境适应能力, CAⅡ、CAⅣ、SLC4A4、SLC26A6 参与碱胁迫下离子转运、渗透压平衡调节。     

盐度胁迫对尼罗罗非鱼鳃氯细胞调节变化的影响

采用扫描电镜和免疫组化技术,研究了尼罗罗非鱼(Oreochromis niloticus)鳃中氯细胞的分布,及其不同盐度(0、10、20、30)胁迫对氯细胞数目和形态变化的影响.扫描电镜结果表明:氯细胞分布在鳃丝的鳃小片基部,根据其表面开口长度,可分为Ⅰ型(＞6.5μm)、Ⅱ型(3.2～6.5μm)和Ⅲ型(＜3.2 μm)3种亚型;不同盐度下氯细胞总数目变化趋势为盐度10＜盐度20＜盐度0＜盐度30;从盐度0转移到盐度10后,氯细胞总数目减少,主要是由于Ⅰ型氯细胞数目显著下降;盐度20中的氯细胞数量高于盐度10,但不显著;盐度30中的氯细胞数量随Ⅲ型氯细胞数量的提高而显著增加.免疫组织化学进一步证实了不同盐度条件下Na+-K+-ATPase免疫反应性细胞均分布在鳃丝的鳃小片基部.本研究结果表明,尼罗罗非鱼可通过改变鳃氯细胞数量和形态结构来适应环境中的盐度变化,推测Ⅰ型氯细胞和Ⅲ型氯细胞分别在低盐、高盐适应中起着重要作用.     

“灭非灵”对尼罗罗非鱼鳃、肝和肾组织结构的影响

通过"灭非灵"对尼罗罗非鱼(Oreochromis niloticus)(体重为34.65±5.69 g)的急性毒性试验及对鳃、肝、肾的组织学研究,从组织学角度探讨了"灭非灵"对尼罗罗非鱼的致死机理。结果表明:"灭非灵"对尼罗罗非鱼的24、48、72和96 h-LC50分别为0.148、0.103、0.048和0.032 mg·L-1;其组织病理学损伤表现为鳃小片萎缩、卷曲、坏死、脱落和融合,鳃间隙分泌大量的粘液细胞;肝细胞肿大,空泡化,细胞界限模糊,细胞核固缩;肾细胞肿大,充血;"灭非灵"对3种组织的损伤程度为鳃肝脏肾脏,3种组织的损伤很有可能是造成尼罗罗非鱼死亡的主要原因。     

皱纹盘鲍外套膜、鳃和足粘液细胞的类型与分布

以阿新兰和过碘酸雪夫氏反应(AB．PAS)染色法显微观察皱纹盘鲍(1taliotis discus hannai)的外套膜、鳃和足的粘液细胞。根据所显示颜色的不同,可将粘液细胞分为Ⅰ～Ⅳ4种类型：分别呈红色、蓝色、紫红色和蓝紫色。外套触手和外套膜上皮的粘液细胞以Ⅱ型为主,Ⅳ型较少,密度不均,多为近圆形细胞,大型和小型细胞均有分布。鳃轴和鳃叶上皮的粘液细胞密度较大,以Ⅱ型和Ⅰ型为主,Ⅲ型和Ⅳ型较少,形态有杯形、近圆形或棒状等,多为中型及小型细胞。足的上皮粘液细胞较少,均为Ⅱ型,但局部上皮的细胞含有许多棕色颗粒。     

两种海马鳃组织和消化道的黏液细胞类型及分布

运用阿新兰(AB,pH 2.6)和过碘酸雪夫氏(PAS)反应染色方法,对三斑海马(Hippocampus trimaculatus)和日本海马(H.japonicus)鳃组织与消化道中的黏液细胞类型及分布进行了研究。染色结果显示:两种海马的鳃组织和消化道中均含有黏液细胞,日本海马的鳃组织中含有Ⅰ型和Ⅳ型黏液细胞,三斑海马的鳃组织中含有Ⅰ型、Ⅲ型和Ⅳ型黏液细胞。两种海马消化道各部位的黏液细胞类型和数量有明显差异:日本海马的食道中Ⅰ型细胞最多,而三斑海马的食道中Ⅳ型细胞最多;日本海马的前肠中只含有Ⅰ型细胞,而三斑海马的前肠中含有Ⅰ型、Ⅲ型和Ⅳ型细胞,其中Ⅰ型细胞含量最多;日本海马的中肠中含有Ⅰ型、Ⅲ型和Ⅳ型细胞,其中Ⅲ型细胞含量最多,而三斑海马中肠中只含有Ⅰ型细胞;日本海马与三斑海马的后肠中都分布有Ⅰ型、Ⅱ型、Ⅲ型和Ⅳ型细胞,两者不同的是,日本海马的后肠中Ⅲ型细胞含量最多,三斑海马的后肠中Ⅳ型细胞含量最多。     

尼罗罗非鱼性腺发育的研究

湖南地区生长于池塘环境的尼罗罗非鱼(Tilapia nilotica),性成熟日龄是110—130天,雄性比雌性普遍早熟20天。精细胞的发生能够完成由精原细胞到精子的全部发育过程,同样,卵细胞的发生也能完成由卵原细胞到卵子的全部发育过程。初级卵母细胞处于Ⅲ时相阶段时,可由包被卵周的滤泡细胞分泌产生放射膜,但放射膜不在动物性极形成受精孔,也无精孔细胞的分化,证实尼罗罗非鱼是属于非受精孔受精类型。初级卵母细胞由Ⅲ时相发育到Ⅳ时相是非同步性的,产后卵巢的组织学结构为第Ⅳ期,卵巢系数在繁殖季节可出现三次高峰,证实尼罗罗非鱼是属于多次产卵类型。通过对精巢组织学的研究,发现尼罗罗非鱼的第Ⅰ期精巢是自然两性嵌合体。       

海洋浮游藻类细胞质膜氧化还原活性与细胞外CA活性的关系

海洋浮游藻类除通过吸收和释放分子与离子来改变其环境的化学成分外,还可通过细胞外表面一些酶的作用引起质膜外化学物质变化。在这方面,海洋浮游藻类一个主要的细胞外表面酶-碳酸酐酶(CA),在经胰蛋白酶处理从细胞质膜上释放出来后,仍保留其催化活性。当细胞外表面CA(简称细胞外CA)具活性时,可催化质膜外HCO_3~-与CO_2的相互转化,为Rubisco(磷酸核酮糖羧化酶)提供一稳定的CO_2流量环境,以维持正常的光合作用。     

2010年浙江省部分地区急性出血性 结膜炎暴发疫情病原学研究

为查明引起2010年浙江省急性出血性结膜炎(AHC)暴发疫情的病因,并对病原进行分子溯源。本研究采用荧光RT-PCR方法直接从患者眼拭子样本中检测肠道病毒(EV)和柯萨奇病毒A24变异株(CA24v)核酸;用Hep-2细胞分离病毒,对阳性分离物提取病毒核酸后进行VP1全基因和3C蛋白酶区(3C)扩增和测序,同源性与进化分析。结果13份眼拭子样本中EV和CA24v核酸均阳性8份,分离到CA24v6株。选取4株病毒测序,获得VP1全长均为915个核苷酸(nt),3C区全长495nt,VP1和3C区均没有nt插入和缺失。2010年浙江4株CA24v分离株之间在3C区和VP1区核苷酸和氨基酸(aa)高度同源,2010年浙江CA24v分离株与原型株EH24/70在3C区的nt和aa同源性分别为85.2%～85.8%和96.2%～96.7%,与2002～2008年浙江、云南和广东CA24v株的同源性分别为93.4%～96.2%和96.7%～99.3%;浙江2010年CA24v株在3C区进化树的GⅣ基因亚型C4分枝上(GⅣ-C4),在VP1基因进化树的人类肠道病毒C组(EV-C)CA24v分枝上。研究表明引起2010年浙江省急性出血性结膜炎暴发流行的病原为CA24v,GⅣ基因亚型,与引起2002～2008年浙江AHC流行的CA24v株(GⅣ)具有密切的亲缘关系,推测CA24v病毒自2002年以来一直在本地低强度循环,2010年又导致了浙江AHC的暴发。     

海洋浮游藻类无机碳利用机理的研究

为了认识海洋浮游藻类在碳充足和碳受限条件下对水体中溶解无机碳 (DIC)的利用方式与可能机理 ,对 13种海洋浮游藻类在不同pH和CO2 浓度及不同DIC条件下细胞外碳酸酐酶 (CA)的活性进行了分析测定。结果显示 :13种藻中 ,只有Amphidiniumcarterae和Prorocentrumminimum在碳充足条件下具细胞外CA活性。Melosirasp .、Phaeodactylumtricornutum、Skeletonemacostatum、Thalassiosirarotula、Emilianiahuxleyi和Pleurochrysiscarterae则在碳受限条件下才具细胞外CA活性。Chaetoceroscompressus、Glenodiniumfoliaceum、Coccolithuspelagicus、Gephrocapsaoceanica和Heterosigmaakashiwo即使在碳受限条件下也未检测到细胞外CA活性。应用封闭系统中pH漂移技术和阴离子交换抑制剂 4′4′ diisothiocyanatostilbene_2 ,2_disulfonicacid (DIDS)等的研究表明 ,Coc.pelagicus和G .oceanica可通过阴离子交换机制进行HCO-3 的直接利用。H .akashiwo没有潜在的HCO-3 直接利用或细胞外CA催化的HCO-3 利用     

遮目鱼幼鱼鳃线粒体丰富细胞的形态结构及其在不同盐度下的变化

该实验通过普通光学显微镜、透射电子显微镜和扫描电子显微镜的方法,研究不同盐度条件下(盐度0、10、20、27、35)广盐性海水鱼类遮目鱼(Chanos chanos)幼鱼鳃器官结构和鳃上线粒体丰富细胞分布及结构的变化。鳃线粒体丰富细胞呈椭圆形或卵圆形,内含有大量线粒体,细胞核较大。在不同盐度条件下,遮目鱼幼鱼出现两种鳃线粒体丰富细胞:一种是具有顶端小窝、线粒体体积较大的A型线粒体丰富细胞;另一种是单独存在、线粒体体积较小的B型线粒体丰富细胞。随着盐度降低,A型线粒体丰富细胞及其线粒体数量减少、体积减小,电子密度降低,顶端开口变小甚至关闭。盐度降至淡水条件下,鳃小片肿胀、脱落,鳃小片上增生出具有大面积平滑或波状的顶端开口的B型线粒体丰富细胞。结果表明,在高渗环境下,A型线粒体丰富细胞较为丰富和发达,其结构特征适应了离子分泌的功能,为海水型线粒体丰富细胞;在低渗环境下,B型线粒体丰富细胞较为丰富,其结构特征适应了离子吸收的功能,为淡水型线粒体丰富细胞。不同结构类型鳃线粒体丰富细胞的存在使得广盐性海水鱼类可以适应较广的盐度范围变化。     

大鼠胚胎发育期NBC1 在胰腺的表达与定位

目的:探讨碳酸氢钠协同转运载体(NBC1)在大鼠胰腺胚胎发育期不同阶段核酸、蛋白水平的动态变化以及在腺泡和β细胞的定位表达。方法:采用高密度寡核苷酸芯片对孕12.5 d(E12.5)、E15.5、E18.5、新生和成年胰腺进行基因转录水平分析,用RT-PCR和Western blot分别验证了NBC1核酸和蛋白在E15.5、E18.5、新生和成年时期胰腺中的表达情况,用Double fluorescence immunohistochemistry分析了NBC1在E18.5、新生和成年时期胰腺腺泡和β细胞的定位表达。结果:在大鼠胰腺胚胎发育过程中,NBC1核酸、蛋白在E18.5时特异高表达,新生下降直至成年最低;在腺泡基底侧膜和β细胞膜有强烈的阳性信号,且在成年胰腺中β细胞膜阳性信号较腺泡基底侧膜强。NBC1的表达变化与其功能近似基因的表达趋势相反,而与其协同发挥作用的基因及胰腺特异基因的表达趋势一致。结论:NBC1在胰腺发育过程中不仅与结构形成而且与功能发挥相关。     

Personality endophenotypes for bipolar affective disorder: a family-based genetic association analysis

Genetic analyses of complex conditions such as bipolar disorder (BD) may be facilitated by the use of intermediate phenotypes. Various personality traits are overrepresented in people with BD and their unaffected relatives, and may constitute genetically transmitted risk factors or endophenotypes of the illness. In this study, we administered a battery of seven different personality questionnaires comprising 19 subscales to 31 Caucasian BD families (n = 241). Ten of these personality traits showed significant evidence of heritability and were therefore selected as candidate endophenotypes. In addition, a principal components analysis produced two heritable components (negative affect and appetitive drive), which accounted for a considerable proportion of the variance (29% + 12%) and were also used in the analysis. A family-based quantitative association study was carried out using the orthogonal model from the quantitative transmission disequilibrium tests (QTDT) program. Monte Carlo permutations (M = 500), which allow for non-normal data and provide a global P value, corrected for multiple testing, were used to calculate empirical P values for the within-family component of association. The 3' untranslated region repeat polymorphism of the dopamine transporter gene (SLC6A3) was associated with self-directedness (P < 0.0001) and negative affect (P = 0.010). The short allele of the serotonin transporter gene (SLC6A4) promoter polymorphism showed a trend toward association with higher harm avoidance (P = 0.016) and negative affect (P = 0.028). The catechol-o-methyltransferase val158met polymorphism was weakly associated with the personality traits, 'Spirituality' (P = 0.040) and irritable temperament (P = 0.022). Furthermore, the met allele of the brain-derived neurotrophic factor val66met polymorphism was associated with higher hyperthymic temperament scores. We raise the possibility that the 10R allele of the SLC6A3 repeat polymorphism and the short allele of the SLC6A4 promoter variant constitute risk factors for irritable-aggressive and anxious-dysthymic subtypes of BD, respectively.     

Mapping of equine potassium chloride co-transporter (SLC12A4) and amino acid transporter (SLC7A10) and preliminary studies on associations between SNPs from SLC12A4, SLC7A10 and SLC7A9 and osmotic fragility of erythrocytes

Consensus DNA sequences from human, mouse and/or rat were used to design oligonucleotide primers for equine homologues of exons 16, 17 and 20-23 of potassium chloride co-transporter (SLC12A4) and exons 10, 11 and 3, 4, respectively, for two amino acid transporters (SLC7A10 and SLC7A9). DNA sequences of the PCR products showed high sequence identity to these regions. Equine BAC clones were obtained for SLC12A4 and SLC7A10 and mapped to equine chromosomes ECA3p13 and ECA10p15, respectively, by fluorescence in situ hybridization (FISH). Several single nucleotide polymorphisms (SNP) were found. Substitutions of A/G were found within exon 17 of SLC12A4, within intron 11 of SLC7A10 and within intron 3 of SLC7A9. The SNP associated with SLC7A10 and SLC7A9 were sufficiently polymorphic to investigate associations with erythrocyte fragility among a group of 20 thoroughbred horses. A non-parametric rank-sum test showed a weak association between erythrocyte fragility and the SNP associated with SLC7A10 (P < 0.05).     

Impaired trafficking and intracellular retention of mutant kidney anion exchanger 1 proteins (G701D and A858D) associated with distal renal tubular acidosis

Abstract

Novel compound heterozygous mutations, G701D, a recessive mutation, and A858D, a mild dominant mutation, of human solute carrier family 4, anion exchanger, member 1 (SLC4A1) were identified in two pediatric patients with distal renal tubular acidosis (dRTA). To examine the interaction, trafficking, and cellular localization of the wild-type and two mutant kidney AE1 (kAE1) proteins, we expressed the proteins alone or together in human embryonic kidney (HEK) 293T and Madin-Darby canine kidney (MDCK) epithelial cells. In individual expressions, wild-type kAE1 was localized at the cell surface of HEK 293T and the basolateral membrane of MDCK cells. In contrast, kAE1 G701D was mainly retained intracellularly, while kAE1 A858D was observed intracellularly and at the cell surface. In co-expression experiments, wild-type kAE1 formed heterodimers with kAE1 G701D and kAE1 A858D, and promoted the cell surface expression of the mutant proteins. The co-expressed kAE1 G701D and A858D could also form heterodimers but showed predominant intracellular retention in HEK 293T and MDCK cells. Thus impaired trafficking of the kAE1 G701D and A858D mutants would lead to a profound decrease in functional kAE1 at the basolateral membrane of α-intercalated cells in the distal nephron of the patients with dRTA.     

Mouse fatty acid transport protein 4 (FATP4): characterization of the gene and functional assessment as a very long chain acyl-CoA synthetase

FATP4 (SLC27A4) is a member of the fatty acid transport protein (FATP) family, a group of evolutionarily conserved proteins that are involved in cellular uptake and metabolism of long and very long chain fatty acids. We cloned and characterized the murine FATP4 gene and its cDNA. From database analysis we identified the human FATP4 genomic sequence. The FATP4 gene was assigned to mouse chromosome 2 band B, syntenic to the region 9q34 encompassing the human gene. The open reading frame was determined to be 1929 bp in length, encoding a polypeptide of 643 amino acids. Within the coding region, the exon-intron structures of the murine FATP4 gene and its human counterpart are identical, revealing a high similarity to the FATP1 gene. The overall amino acid identity between the deduced murine and human FATP4 polypeptides is 92.2%, and between the murine FATP1 and FATP4 polypeptides is 60.3%. Northern analysis showed that FATP4 mRNA was expressed most abundantly in small intestine, brain, kidney, liver, skin and heart. Transfection of FATP4 cDNA into COS1 cells resulted in a 2-fold increase in palmitoyl-CoA synthetase (C16:0) and a 5-fold increase in lignoceroyl-CoA synthetase (C24:0) activity from membrane extracts, indicating that the FATP4 gene encodes an acyl-CoA synthetase with substrate specificity biased towards very long chain fatty acids.     

Mouse ZIP1 and ZIP3 genes together are essential for adaptation to dietary zinc deficiency during pregnancy

Subfamily II of the solute-linked carrier 39A superfamily contains three well-conserved zinc transporters (ZIPs1, 2, 3) whose physiological functions are unknown. We generated mice homozygous for knockout alleles of ZIP1 and both ZIP1 and ZIP 3 (double-knockout). These mice were apparently normal when dietary zinc was replete, but when dietary zinc was limited during pregnancy embryos from ZIP1 or ZIP3 knockout mice were two to three times more likely to develop abnormally than those in wildtype mice, and 91% (71/78) of embryos developed abnormally in ZIP1, ZIP3 double-knockout mice. Analysis of the patterns of expression of these genes in mice revealed predominate expression in intestinal stromal cells, nephric-tubular epithelial cells, pancreatic ductal epithelial cells, and hepatocytes surrounding the central vein. This suggests that these zinc transporters function, at least in part, in the redistribution and/or retention of zinc rather than its acquisition from the diet. In conclusion, mutations in the ZIP1 and ZIP3 zinc transporter genes are silent when dietary intake of zinc is normal, but can dramatically compromise the success of pregnancy when dietary intake of zinc is limiting.     

Gill morphology during hypercapnia in brown bullhead (Ictalurus nebulosus): role of chloride cells and pavement cells in acid-base regulation

The role of gill chloride cells (CCs) and pavement cells (PVCs) in acid-base regulation was evaluated in brown bullhead catfish (Ictalurus nebulosus) subjected to acute hypercapnia (water Pco₂=15 torr). Chronic (10 day) cortisol treatment was used as a tool to cause CC proliferation to permit a comparison of the regulatory capacities in groups of fish with widely different gill CC populations. Cortisol (4mg kg^?1 day^?1) caused a pronounced increase (170%) in the surface area of CCs exposed to the water based on scanning and transmission electron microscope analysis. The density of PVC apical membrane microvilli was significantly increased (20%) by cortisol treatment. Exposure of either group of fish to hypercapnia caused similar changes in gill epithelial morphology including: (i) a marked reduction in the surface area of exposed CCs (52 and 78% reduction in the control and cortisol-treated fish, respectively); and (ii) pronounced increases in PVC apical membrane microvilli density (21 and 27% in the control and cortisol-treated fish, respectively). The rates of Cl^? uptake (J_incl^?) and Na⁺ uptake (J_inNa⁺) were elevated (150 and 262%, respectively) in the cortisol-treated fish. Regardless of treatment, J_incl^? was markedly reduced to approximately the same levels after 6 h of hypercapnia, J_inNa⁺ was stimulated in the control group and reduced in the cortisol-treated group and thus, after 6 h of hypercapnia, J_inNa⁺ was equal in each group. The similar morphological responses in fish possessing different initial populations suggests that the predominant mechanism of acid-base regulation during hypercapnia, reduction of C1^?/HCO₃^? exchange, is accomplished by removal of the CC-associated C1-/HCO₃^? exchange sites from the water. The increase in PVC microvilli density during hypercapnia suggests a role for the PVC in acid-base regulation.     

Quantitative trait loci mapping and gene network analysis implicate protocadherin‐15 as a determinant of brain serotonin transporter expression

Presynaptic serotonin (5‐hydroxytryptamine, 5‐HT) transporters (SERT) regulate 5‐HT signaling via antidepressant‐sensitive clearance of released neurotransmitter. Polymorphisms in the human SERT gene (SLC6A4) have been linked to risk for multiple neuropsychiatric disorders, including depression, obsessive‐compulsive disorder and autism. Using BXD recombinant inbred mice, a genetic reference population that can support the discovery of novel determinants of complex traits, merging collective trait assessments with bioinformatics approaches, we examine phenotypic and molecular networks associated with SERT gene and protein expression. Correlational analyses revealed a network of genes that significantly associated with SERT mRNA levels. We quantified SERT protein expression levels and identified region‐ and gender‐specific quantitative trait loci (QTLs), one of which associated with male midbrain SERT protein expression, centered on the protocadherin‐15 gene (Pcdh15), overlapped with a QTL for midbrain 5‐HT levels. Pcdh15 was also the only QTL‐associated gene whose midbrain mRNA expression significantly associated with both SERT protein and 5‐HT traits, suggesting an unrecognized role of the cell adhesion protein in the development or function of 5‐HT neurons. To test this hypothesis, we assessed SERT protein and 5‐HT traits in the Pcdh15 functional null line (Pcdh15^av‐3J), studies that revealed a strong, negative influence of Pcdh15 on these phenotypes. Together, our findings illustrate the power of multidimensional profiling of recombinant inbred lines in the analysis of molecular networks that support synaptic signaling, and that, as in the case of Pcdh15, can reveal novel relationships that may underlie risk for mental illness .     

中国荷斯坦牛CVM的基因检测及其与产奶性状的关联分析

脊椎畸形综合征(Complex vertebral malformation, CVM)是由位于牛第3号染色体(BTA3)的SLC35A3基因外显子4的一个单碱基突变(G559T)所致。该致病基因在世界许多国家的荷斯坦牛群中都有一定的比例。文章对北京地区38头优秀种公牛进行分析, 发现了4头携带者, 进而检测了这些携带者公牛的555头女儿的基因型, 其中携带者占检测母牛数的44.0%。此外, 关联分析结果表明, 携带者母牛与非携带者母牛的生产性能之间存在显著差异(P<0.01)。携带者母牛的5个产奶性状育种值均显著高于非携带者, 泌乳持续力和体细胞评分SCS的育种值也比非携带者略高。CVM致病基因可能与BTA3上影响产奶性状的QTL或基因连锁。因此, 建议生产中对CVM携带者进行逐步淘汰