青海湖裸鲤性腺发育的组织学研究

采用石蜡组织切片法,对青海湖裸鲤的性腺发育进行组织学研究,系统地描述了各期卵巢和精巢的形态结构、特征及变化。结果表明:(1)青海湖裸鲤性腺的发育可分为六个时期,卵母细胞从第3时相发育到第4时相基本同步。第Ⅳ期末卵巢,第4时相卵母细胞的卵径大小比较整齐,卵径平均值为2.3mm,第4时相卵母细胞占切面上卵数的85%以上,占切片面积的96%以上,第2、3时相卵母细胞已很少;(2)产后卵巢的组织结构逐步由第Ⅵ期回复到第Ⅱ期,再由第Ⅱ期到第Ⅲ期向第Ⅳ期过渡;(3)性成熟个体有68.12%的雌鱼和83.9%的雄鱼以第Ⅳ期性腺越冬,另有21.01%的雌鱼和10.7%的雄鱼以第Ⅲ期性腺越冬,还有10.87%的雌鱼和5.4%的雄鱼以第Ⅱ期性腺越冬。根据青海湖裸鲤各季节性腺发育情况,作者认为青海湖裸鲤已达到性成熟的个体并不是每年都参与繁殖活动,存在生殖间断性。这反映了青海湖裸鲤的繁殖习性对高原寒冷多变气候的适应性。还对卵黄核和核仁在卵黄形成中的作用以及青海湖裸鲤的产卵类型和生殖间断性进行了讨论。     

纳木错裸鲤性腺发育的组织学研究

采用常规的组织切片方法,对西藏特有鱼类纳木错裸鲤的性腺发育进行了组织学研究,系统地描述了各期精巢和卵巢的形态结构、特征及其变化.尤其论述了其卵巢中卵细胞的卵黄核破碎与分解的特点、卵膜的结构、核仁排出物在卵黄形成过程中的作用,以及卵粒重吸收的过程.根据各期卵巢中卵母细胞的组成情况,认为纳木错裸鲤已达性成熟的个体并不是每年都参与繁殖活动是对高原极端、多变气候环境的一种生态适应;阐明了纳木错裸鲤属于分批同步产卵鱼类.       

短盖巨脂鲤卵巢发育组织学研究

通过对短盖巨脂鲤各个生长时期卵巢组织学研究以及成熟卵超微结构观察,获得短盖巨脂鲤生长发育过程中卵巢发育规律;同时对卵母细胞核仁排出物与核质关系及在卵黄形成中的作用等问题作了初步探讨;并根据卵巢的卵母细胞组成确定了其产卵类型。     

藏北3种裸鲤同工酶的电泳分析及物种分化的探讨

对藏北高原３种裸鲤的乳酸脱氢酶（ＬＤＨ）、苹果酸脱氢酶（ＭＤＨ）和脂酶（ＥＳＴ）进行电泳分析的结果表明,３种裸鲤酶谱均表现出种间的差别,而且在同一种群个体之间也存在着明显的分化,但无性别差异。３种裸鲤被检测的３种同工酶均有沉默基因表达的现象,重复基因ＬＤＨ－Ａ^2、ＬＤＨ－Ｂ^2、s-MDH-A^2和m-MDH-B^2也在部分个体中表达。遗传距离分析表明,色林错裸鲤（Ｇ.selincuoensis)与错鄂裸鲤（Ｇ．cuoensis)之间较之于与纳木错裸鲤（Ｇ．namensis)有更近的亲缘关系。与其他四倍体鱼类相比,裸鲤鱼类同工酶在重复基因和沉默基因上都有较高的表达频率,这种情况说明裸鲤鱼类目前可能还外于多倍化后进化的早期过程并早于胭脂鱼类所处的相应时期,这与裂腹鱼类起源较晚以及青藏高原业已存在的恶劣环境条件直接相关。     

沙柳河青海湖裸鲤早期资源发生量及时空分布

为了掌握青海湖裸鲤(Gymnocypris przewalksii)资源补充状况, 于2019年5月26日至9月2日逐日在青海湖第二大支流沙柳河开展了青海湖裸鲤早期资源丰度时空变化特征的调查研究, 共采集青海湖裸鲤鱼卵3386粒, 仔鱼4690尾。调查发现, 亲鱼自5月底开始洄游, 鱼卵6月初出现, 6月底至7月初达到高峰, 随后逐渐下降, 至8月中旬基本消失。仔鱼数量自7月初呈波动式上升, 8月达到高峰, 9月初逐渐消失。经估算, 沙柳河鱼卵径流量为25.58×106粒, 仔鱼径流量为62.00×106尾。鱼卵仔鱼空间分布为从河口往上丰度依次递减, 断面水平分布为右岸>左岸>中心。Kruskal-Wallis检验表明, 昼夜鱼卵丰度存在显著差异(P<0.05); 昼夜仔鱼丰度存在极显著性差异(P<0.01), 鱼卵仔鱼漂流高峰期均集中在夜间。鱼卵丰度与流速呈显著正相关(P<0.05), 仔鱼丰度与流速呈极显著正相关(P<0.01), 与径流量日上涨率呈极显著正相关(P<0.01)。研究报道了沙柳河青海湖裸鲤早期资源现状, 填补了该水域青海湖裸鲤早期资源研究的空缺, 为青海湖裸鲤上溯亲本资源量和入湖幼鱼资源量估算提供了数据支撑, 可为青海湖裸鲤资源管理与保护提供科学依据。     

硒拮抗超氧阴离子导致的心肌线粒体膜损伤

由黄嘌呤和黄嘌呤氧化酶体系产生的超氧阴离子作用于心肌线粒体后,其膜脂双层内产生了脂类自由基。在一定时间内,脂类自由基与自旋捕捉剂形成的加合物的ＥＳＲ信号强度随着孵育时间的增加而加强。１．０μｍｏｌ／Ｌ硒代蛋氨酸（Ｓｅ－Ｍｅｔ）或２．３μｍｏｌ／ＬｍＮａ２ＳｅＯ３可显著清除并抑制脂类自由基的产生。在的影响下,荧光探针ＤＰＨ在膜脂双层中的荧光寿合和膜脂流动性发生了明显改变。一定浓度的Ｓｅ－Ｍｅｔ或Ｎａ２ＳｅＯ３可明显拮抗的上述影响,前者的作用更为显著。     

西藏朗错秋季裸鲤属营养生态位及种间食物关系

西藏朗错海拔4300 m,是典型的高原湖泊之一。湖内具有兰格湖裸鲤和拉孜裸鲤共存的现象。本研究采用碳、氮稳定同位素技术分析了2018年秋季兰格湖裸鲤和拉孜裸鲤的食物组成和营养生态位,探讨了两种裸鲤的种间食物关系,包括饵料相似性和重叠系数。结果表明: 兰格湖裸鲤主要摄食蜉蝣类幼虫、摇蚊类幼虫、着生藻类、钩虾类和浮游生物;拉孜裸鲤主要摄食蜉蝣类幼虫、着生藻类、摇蚊类幼虫、钩虾类和水生维管束植物。两种裸鲤的营养生态位存在重叠,兰格湖裸鲤的δ¹³C值变幅(CR=1.60)、δ¹⁵N值变幅(NR=1.89)和凸多边形,总面积(TA=1.80)均小于拉孜裸鲤,差值分别为0.09、0.57和1.29。两种裸鲤的饵料相似性系数(94.2％)和重叠系数(91.5%)均较高,表明秋季西藏朗错两种裸鲤可能存在激烈的种间食物竞争关系。     

青海湖裸鲤和黄河裸裂尻鱼感染多子小瓜虫的病理学比较研究

为探讨不同裂腹鱼类感染多子小瓜虫后的病理学差异, 利用多子小瓜虫对青海湖裸鲤指名亚种(Gymnocypris przewalskii przewalskii)和黄河裸裂尻鱼(Schizopygopsis pylzovi Kessler)实施感染实验, 并对2种鱼进行了深入的病理学研究。研究结果显示: (1) 2种鱼的死亡量均呈现先激增后明显回落的趋势, 青海湖裸鲤死亡高峰在感染后第3至第4天, 黄河裸裂尻鱼死亡高峰在第3至第5天, 青海湖裸鲤的死亡比黄河裸裂尻鱼急剧。(2)感染后2种鱼体表均出现大量肉眼可见的白点。青海湖裸鲤皮肤黏液分泌量明显增加, 体表形成胶状黏液层, 黏液层中见不同细胞期小瓜虫包囊。黄河裸裂尻鱼鳍出现蛀鳍现象, 皮肤出现细菌感染样溃烂。(3)解剖发现, 感染组青海湖裸鲤和黄河裸裂尻鱼肝脏发生病理改变呈淡黄色, 胆有不同程度肿大。(4)组织切片和电镜观察显示, 小瓜虫在鳃部位的寄生导致青海湖裸鲤和黄河裸裂尻鱼鳃丝黏连, 鳃小片和鳃丝上皮细胞萎缩、脱落, 鳃丝结构被严重破坏。小瓜虫在2种鱼皮肤的寄生使寄生部位组织突起, 周围组织塌陷。青海湖裸鲤表皮下层出现空隙, 表皮结构被严重破坏。黄河裸裂尻鱼皮肤表皮细胞出现空泡化病理改变, 失去原有紧密结构, 表皮层和固有层间界限变模糊。综上所述, 小瓜虫的感染对青海湖裸鲤和黄河裸裂尻鱼的鳃和皮肤组织造成严重损伤, 但2种鱼表现的症状和造成的组织损伤类型有明显差异, 这与2种鱼长期适应不同水体环境密切相关。     

青海湖裸鲤体腔寄生蠕虫群落研究

青海湖裸鲤体腔为三种寄生蠕虫寄生,即裂头绦虫裂头蚴,舌状绦虫裂头蚴和对盲囊缄虫的三期幼虫,这几种蠕虫不同大小的青海湖裸鲤中数量差异很大,将其看作一个组分群落,并根据其发生数量进行模糊聚类和极点排序,结果将其分为二个亚组分群落,即体长小于１５０ｍｍ的青海湖裸鲤体腔蠕虫亚组分群落和体长大于１５０ｍｍ的个体的体腔蠕虫亚组分群落。这与宿主的行为和食性改变有密切关系,舌状绦虫与另两种蠕虫间有显著负协调关系。     

花斑裸鲤胚胎/仔鱼型血红蛋白基因家族成员鉴定与时序表达研究

为了探讨花斑裸鲤(Gymnocypris eckloni)血红蛋白时序转换 利用花斑裸鲤全基因组数据鉴定胚胎/仔鱼型血红蛋白基因家族成员, 并通过整胚原位杂交方法, 检测花斑裸鲤胚胎/仔鱼型血红蛋白基因在胚胎发育不同阶段的表达及定位。结果表明, 花斑裸鲤基因组中共鉴定到5个胚胎/仔鱼型血红蛋白基因, 分别为hbae1、hbae4、hbae5、hbbe1和hbbe3, 与斑马鱼(Danio rerio)相比, 花斑裸鲤基因组缺少hbae3和 hbbe2 基因, 暗示第四轮全基因组复制事件后所经历的小规模基因删除事件在花斑裸鲤特异性血红蛋白基因形成中发挥了重要作用。整胚原位杂交结果显示, hbae1基因在胚胎发育的120h至432h内持续表达, hbbe1基因在96h开始表达持续至432h, hbbe3基因杂交信号出现在胚胎发育120h至384h内, 在胚胎发育全过程中未能观察到hbae4和hbae5基因的杂交信号。杂交信号主要位于胚胎正中轴、后部侧向中胚层、背主动脉腹侧区、尾部造血区及卵黄。正义探针作为阴性对照, 在胚胎发育阶段均无任何杂交信号。花斑裸鲤具有与其他鱼类不同的胚胎/仔鱼型血红蛋白基因家族成员及血红蛋白转换表达特征; hbae1、hbbe1和hbbe3基因在花斑裸鲤早期胚胎发育过程中发挥重要作用, 而hbae4和hbae5基因的生物学功能可能有所弱化。     

二价汞、铅、镉氯化物对脑磷脂平板双分子层膜电特性的影响

本文报道了重金属毒物对脑磷脂双分子层膜电性质的影响.二价汞、铅、镉氯化物的存在,使膜电阻下降,膜的通透性增加.并且随着浓度的增加,膜平均寿命下降,击穿电压减小,说明膜机械脆性增加,膜容易破裂.而其它二价的Mg~(2 )、Ca~(2 )、Cu~(2 )、Mn~(2 )氯化物并无此作用.可以认为重金属毒物对生命的毒害作用中包括了对膜脂双层的直接影响.     

竹红菌甲素对红细胞膜内脂双层的微扰

本文以荧光探针为手段,以人红细胞膜为材料,测量了膜偏振度的改变,荧光探针能量转移,荧光峰的蓝移和甲素激发峰的分裂。结果表明在有竹红菌甲素存在时,红细胞膜偏振度增加,探针荧光强度减小,荧光峰蓝移。甲素浓度增加时,上述现象更加明显,即它们之间有正的相关关系。同时,甲素激发光谱的a带发生分裂。据此,我们认为甲素对红细胞膜内脂双层产生明显微扰,甲素与红细胞膜间存在着相互作用。在甲素浓度较大时,它主要是渗入到红细胞膜脂双层的深层部位(膜脂肪酸链的12—16位)。     

Organization of lipids in sarcoplasmic reticulum membrane and Ca2+-dependent ATPase activity.

The organization of lipids in sarcoplasmic reticulum membrane was studied with a variety of stearic spin labels and a phosphatidylcholine spin label. The ESR spectra of the spin-labeled membranes consisted of two components, one due to labels in lipid bilayer structure and the other due to more immobilized labels. The relative intensity of the immobilized component increased when the lipid content of the membrane was decreased by treatment with phospholipase A [EC 3.1.1.4] and subsequent washing with bovine serum albumin. Membrane containing 30% of the intact phospholipid, i.e.0.15 mg of phospholipid per mg of protein, showed a spectrum consisting only of the immobilized component (the overall splitting ranged from 58.5 G to 60.5 G). The immobilized component was ascribed to lipids complexed with protein. The fraction of lipids in the two different organizations was determined from the ESR spectrum. The activity of the Ca2+-Mg2+ dependent ATPase [ATP phosphohydrolase, EC 3.6.1.3] was found to increase almost linearly with the lipid bilayer content in the membrane, whereas phosphoenzyme formation was almost independent of the bilayer content. This indicated that the bilayer structure is necessary for the ATPase to attain its full transport activity.     

A general mechanism for drug promiscuity: Studies with amiodarone and other antiarrhythmics

Amiodarone is a widely prescribed antiarrhythmic drug used to treat the most prevalent type of arrhythmia, atrial fibrillation (AF). At therapeutic concentrations, amiodarone alters the function of many diverse membrane proteins, which results in complex therapeutic and toxicity profiles. Other antiarrhythmics, such as dronedarone, similarly alter the function of multiple membrane proteins, suggesting that a multipronged mechanism may be beneficial for treating AF, but raising questions about how these antiarrhythmics regulate a diverse range of membrane proteins at similar concentrations. One possible mechanism is that these molecules regulate membrane protein function by altering the common environment provided by the host lipid bilayer. We took advantage of the gramicidin (gA) channels’ sensitivity to changes in bilayer properties to determine whether commonly used antiarrhythmics—amiodarone, dronedarone, propranolol, and pindolol, whose pharmacological modes of action range from multi-target to specific—perturb lipid bilayer properties at therapeutic concentrations. Using a gA-based fluorescence assay, we found that amiodarone and dronedarone are potent bilayer modifiers at therapeutic concentrations; propranolol alters bilayer properties only at supratherapeutic concentration, and pindolol has little effect. Using single-channel electrophysiology, we found that amiodarone and dronedarone, but not propranolol or pindolol, increase bilayer elasticity. The overlap between therapeutic and bilayer-altering concentrations, which is observed also using plasma membrane–like lipid mixtures, underscores the need to explore the role of the bilayer in therapeutic as well as toxic effects of antiarrhythmic agents.     

膜融合剂对脂质体及血影膜膜流动性的影响

本文用荧光探针ANS,DPH与A研究了几种膜融合剂对脂质体与血影膜流动性的影响.蔗糖使PS脂质体的脂双层流动性降低,探针越是在极性区流动性越小,说明蔗糖主要作用于脂双层的极性区;蔗糖也使血影膜流动性降低,此作用是可逆的.油酸甘油脂(GMO)使PS脂质体的流动性增加,且越是在疏水区内部,流动性增加得越大,说明GMO主要是作用于脂双层的非极性区:GMO也使血影膜流动性增加,此作用是不可逆的.二甲亚砜(DMSO)对血影膜的作用,两种不同荧光探针不一样,对DPH的作用出现双相让,低浓度与高浓度的作用结果分别与蔗糖和GMO的作用一致.     

Membrane fusogenic activity of the Alzheimer's peptide A beta(1-42) demonstrated by small-angle neutron scattering

Amyloid-β peptide (Aβ) is considered a triggering agent of Alzheimer's disease. In relation to a therapeutic treatment of the disease, the interaction of Aβ with the cell membrane has to be elucidated at the molecular level to understand its mechanism of action. In previous works, we had ascertained by neutron diffraction on stacked lipid multilayers that a toxic fragment of Aβ is able to penetrate and perturb the lipid bilayer. Here, the influence of Aβ(1-42), the most abundant Aβ form in senile plaques, on unilamellar lipid vesicles of phospholipids is investigated by small-angle neutron scattering. We have used the recently proposed separated form factor method to fit the data and to obtain information about the vesicle diameter and structure of the lipid bilayer and its change upon peptide administration. The lipid membrane parameters were obtained with different models of the bilayer profile. As a result, we obtained an increase in the vesicle radii, indicating vesicle fusion. This effect was particularly enhanced at pH 7.0 and at a high peptide/lipid ratio. At the same time, a thinning of the lipid bilayer occurred. A fusogenic activity of the peptide may have very important consequences and may contribute to cytotoxicity by destabilizing the cell membrane. The perturbation of the bilayer structure suggests a strong interaction and/or insertion of the peptide into the membrane, although its localization remains beyond the limit of the experimental resolution.     

Low molecular channel-former protein from the mitochondrial membrane

Individual low molecular weight protein component was isolated by gel-filtration method from the inner mitochondrial membrane. This membrane component increases the conductivity of bilayer lipid membranes (BLM) in the presence of K+ and Ca2+-ions. This phenomenon may be explained by the formation of single conductivity channels. The voltage - current characteristics of this channel is nonlinear, which may be the result of asymmetrical operation of the channel - former in the polarized membrane, in spite of equal concentration of protein on both sides of the membrane.     

Elastic deformation and failure of lipid bilayer membranes containing cholesterol.

Giant bilayer vesicles were reconstituted from several lipids and lipid/cholesterol (CHOL) mixtures: stearolyloleoylphosphatidylcholine (SOPC), bovine sphingomyelin (BSM), diarachidonylphosphatidylcholine (DAPC), SOPC/CHOL, BSM/CHOL, DAPC/CHOL, and extracted red blood cell (RBC) lipids with native cholesterol. Single-walled vesicles were manipulated by micropipette suction and several membrane material properties were determined. The properties measured were the elastic area compressibility modulus K, the critical areal strain alpha c, and the tensile strength tau lys, from which the failure energy or membrane toughness Tf was calculated. The elastic area expansion moduli for these lipid and lipid/cholesterol bilayers ranged from 57 dyn/cm for DAPC to 1,734 dyn/cm for BSM/CHOL. The SOPC/CHOL series and RBC lipids had intermediate values. The results indicated that the presence of cholesterol is the single most influential factor in increasing bilayer cohesion, but only for lipids where both chains are saturated, or mono- or diunsaturated. Multiple unsaturation in both lipid chains inhibits the condensing effect of cholesterol in bilayers. The SOPC/CHOL system was studied in more detail. The area expansion modulus showed a nonlinear increase with increasing cholesterol concentration up to a constant plateau, indicating a saturation limit for cholesterol in the bilayer phase of approximately 55 mol% CHOL. The membrane compressibility was modeled by a property-averaging composite theory involving two bilayer components, namely, uncomplexed lipid and a lipid/cholesterol complex of stoichiometry 1/1.22. The area expansion modulus of this molecular composite membrane was evaluated by a combination of the expansion moduli of each component scaled by their area fractions in the bilayer. Bilayer toughness, which is the energy stored in the bilayer at failure, showed a maximum value at approximately 40 mol% CHOL. This breakdown energy was found to be only a fraction of the available thermal energy, implying that many molecules (approximately 50-100) may be involved in forming the defect structure that leads to failure. The area expansion modulus of extracted RBC lipids with native cholesterol was compared with recent measurements of intact RBC membrane compressibility. The natural membrane was also modeled as a simple composite made up to a compressible lipid/cholesterol matrix containing relatively incompressible transmembrane proteins. It appears that the interaction of incompressible proteins with surrounding lipid confers enhanced compressibility on the composite structure.     

Structure, dynamics, and membrane topology of stannin: a mediator of neuronal cell apoptosis induced by trimethyltin chloride

Organotin compounds or alkyltins are ubiquitous environmental toxins that have been implicated in cellular death. Unlike other xenobiotic compounds, such as organomercurials and organoleads, alkyltins activate apoptotic cascades at low concentrations. Trimethyltin (TMT) chloride is amongst the most toxic organotin compounds, and is known to selectively inflict injury to specific regions of the brain. Stannin (SNN), an 88-residue mitochondrial membrane protein, has been identified as the specific marker for neuronal cell apoptosis induced by TMT intoxication. This high specificity of TMT makes SNN an ideal model system for understanding the mechanism of organotin neurotoxicity at a molecular level. Here, we report the three-dimensional structure and dynamics of SNN in detergent micelles, and its topological orientation in lipid bilayers as determined by solution and solid-state NMR spectroscopy. We found that SNN is a monotopic membrane protein composed of three domains: a single transmembrane helix (residues 10-33) that transverses the lipid bilayer at approximately a 20 degrees angle with respect to the membrane normal; a 28 residue unstructured linker, which includes a conserved CXC metal-binding motif and a putative 14-3-3zeta binding domain; and a distorted cytoplasmic helix (residues 61-79) that is partially absorbed into the plane of the lipid bilayer with a tilt angle of approximately 80 degrees from the membrane normal. The structure and architecture of SNN within the lipid environment provides insight about how this protein transmits toxic insults caused by TMT across the membrane.     

Effects of potassium on lipid-protein interactions in light sarcoplasmic reticulum

This study shows the effect of K+ on phospholipid-protein interactions in light sarcoplasmic reticulum (LSR) as measured by 31P NMR. In the presence of 110 mM K+, a substantial effect of the membrane protein on the behavior of the phospholipids was detected. Subtracting the spectrum of the LSR lipid extract from the spectrum of the intact LSR membrane produced a difference spectrum of much greater breadth than the normal phospholipid bilayer powder pattern. This powder pattern is indicative of a phospholipid domain considerably more motionally restricted than the phospholipids in a normal phospholipid bilayer. The apparent axially symmetric powder pattern is consistent with axial diffusion. In a reconstituted membrane containing the calcium pump protein at a lipid/protein ratio much less than in the light sarcoplasmic reticulum, the broad component was more prominent. The relative resonance intensity of the broad component appeared to be proportional to the lipid/protein ratio of the membrane. In 10 mM K+, no broad powder pattern is observed in the corresponding difference spectrum. Thus, in the absence of potassium, the membrane protein has much less influence on the phospholipid of the membrane, as measured by 31P NMR. In addition to the effects of K+ on the membrane structure of the sarcoplasmic reticulum, K+ modulated the function of the calcium pump. The rate of calcium-dependent ATP hydrolysis increased in light sarcoplasmic reticulum when [K+] increased from 10 to 110 mM. The rate of calcium transport was also stimulated by an increase in K+.