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~(–1)NaHCO_3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HCO_3~–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~–/HCO_3~–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。     

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。     

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。     

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。     

珠江口池养梭鱼鳃的光镜、扫描和透射电镜观察

应用光学显微镜、扫描电镜和透射电镜对珠江口池塘养殖梭鱼Liza haematocheila鳃的组织结构、表面形态特征及鳃小片超微结构进行了观察。结果表明梭鱼具有4对鳃,每个鳃由鳃弓、鳃丝、鳃小片和鳃耙组成。梭鱼鳃丝和鳃小片的表面结构和超微结构与其他硬骨鱼类的基本相似,鳃丝表面分布有众多规则或不规则的环形微嵴、沟、坑、孔等结构。鳃丝分为呼吸区和非呼吸区,呼吸区较为平滑,上皮细胞表面无微嵴,呈皱褶状;非呼吸区分布有沟、坑、孔等结构,上皮细胞有较规则的指纹状微嵴。鳃小片是最主要的呼吸场所,由基膜、上皮细胞、内皮细胞、柱细胞和毛细血管网组成。泌氯细胞主要分布在鳃小片基部,并有开口通往外界。本文还探讨了梭鱼鳃的结构与其功能的密切关系。     

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。     

斑马鱼鳃的光镜和透射电镜观察

应用光学显微镜和透射电镜对斑马鱼(Danio rerio)鳃的组织结构及鳃丝、鳃小片超微结构进行了观察。结果表明,斑马鱼有4对全鳃,鳃耙呈长锥状,鳃丝呈梳状排列在鳃弓上,鳃小片均匀排列在鳃丝两侧。鳃小片由上皮细胞、柱细胞、内皮细胞和毛细血管网组成,鳃小片基部和血管周围分布有泌氯细胞,胞内有丰富的线粒体和排泄小泡,根据线粒体形态特征和细胞质电子密度可将其分为两个亚型。黏液细胞通常与泌氯细胞对生存在,并且有通外的开口。斑马鱼鳃组织结构与其他硬骨鱼鳃结构相似,其结构和功能有密切的关系。     

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     

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 .     

Zinc nutritional status influences ZnT1 and ZIP4 gene expression in children with a high risk of zinc deficiency

IntroductionSubclinical deficiency of zinc is associated with impairment of immune system function, growth, and cognitive development in children. Although plasma zinc is the best available biomarker of the risk of zinc deficiency in populations, its sensitivity for early detection of deficiency is limited. Therefore, we aimed to investigate zinc deficiency among preschool children and its relationship with whole blood gene expression of zinc transporters ZIP4 and ZnT1.Material and methodsThis cross-sectional study included 139 children aged 32–76 months enrolled in philanthropic day-care centers. We performed an anthropometric evaluation, weighed food record and dietary record for dietary assessment, blood sample collection for zinc, and whole blood gene expression analyses of ZnT1 (SLC30A1) and ZIP4 (SLC39A4).ResultsZinc deficiency was observed in 26.6 % of the children despite adequate zinc intake and a phytate:zinc molar ratio < 18. Usual zinc intake did not affect whole blood gene expression of zinc transporters, but zinc status influenced ZnT1 and ZIP4 whole blood mRNA. Children with zinc deficiency exhibited 37.1 % higher ZnT1 expression and 45.3 % lower ZIP4 expression than children with adequate zinc (p < 0.05).ConclusionChildren with plasma zinc deficiency exhibited higher expression of ZnT1 and lower expression of ZIP4 in whole blood mRNA, reinforcing the existence of strong regulation of mineral homeostasis according to the nutritional status, indicating that this analysis may be useful in the evaluation of dietary interventions.