甲状腺素对大鼠心肌细胞收缩功能及钙瞬变的影响

目的：研究甲状腺素对单个心肌细胞收缩功能及钙瞬变的影响。方法：SD大鼠60只随机分为对照组（30只）、甲状腺素组（30只）。通过给予甲状腺素喂养后,测量血流动力学指标,随后分离单个心室细胞,采用可视化动缘探测系统同步检测大鼠心肌细胞收缩和钙瞬变的变化并与对照组比较。结果：（1）甲状腺素治疗组左室收缩压（LVSP）和最大缩短和复长速率（±dp／dtmax）较对照组明显提高（P〈0．05）,左室舒张末压（LVEDP）较对照组明显下降（P〈0．05）。（2）甲状腺素治疗组单个心肌细胞收缩功能指标最大收缩幅度（PTA）、最大缩短和复长速率（±dL／dtmax）较对照组明显增加（P〈0．05）,钙瞬变指标fura-2荧光强度变化（△FFI）明显增强、Ca2＋离子达峰值时程（TTPCa）、舒张期Ca2＋减少50％时程（T50DCa）较对照组明显缩短（P〈0．05）。结论：甲状腺素可改善大鼠心脏功能并增强单个心肌细胞的收缩功能及钙瞬变幅度,在单细胞水平上为探讨甲状腺素治疗慢性心衰的发生机制提供了实验依据。     

小剂量过氧化氢对大鼠心肌细胞钙瞬变及凋亡的作用

目的:探讨小剂量过氧化氢导致的氧化应激对大鼠心肌细胞钙瞬变及细胞凋亡的作用。方法:解剖取出成年大鼠心脏,应用langendorff方法分离心肌细胞,加入fluo-3荧光指示剂后,应用不同浓度的过氧化氢作用于心肌细胞,在共聚焦显微镜下测定心肌细胞内钙瞬变。分离并培养新生大鼠心肌细胞,观察过氧化氢处理心肌细胞后其形态的变化,从而评价小剂量过氧化氢对心肌细胞的凋亡作用。结果:应用0.125 mmol/L、0.25 mmol/L及0.375 mmol/L的过氧化氢作用于心肌细胞后,心肌细胞内钙瞬变幅度明显升高,并呈时间剂量依赖性。在培养的大鼠原代心肌细胞中加入0.25 mmol/L的过氧化氢后,心肌细胞发生凋亡的形态变化。结论:小剂量过氧化氢可开放心肌细胞L-钙通道,明显增加心肌细胞内钙瞬变,并导致心肌细胞凋亡。     

白细胞介素-2对大鼠心肌收缩功能的影响涉及一氧化氮机制

为深入研究细胞因子白细胞介素-2(interleukin-2,IL-2)对心肌收缩功能的影响及其可能机制,本实验采用酶解分离成年大鼠心室肌细胞模型,用视频跟踪计算机系统记录测定单个心室肌细胞收缩反应并用双波长荧光系统检测细胞[Ca2+]i.心肌细胞收缩参数包括最大收缩幅度(dL)、细胞最大收缩速度(+dL/dtmax)、细胞最大舒张速度(-dL/dtmax)和舒张末期细胞长度.结果显示,IL-2(2-1000U/ml)浓度依赖性地抑制心肌细胞dL、±dL/dtmax和舒张末期细胞长度;用一氧化氮(ni-tric oxide,NO)合酶抑制剂L-NAME(100mmol/L)和可溶性鸟苷酸环化酶(sGC)抑制剂ODQ(10mmol/L)可减弱IL-2对心肌细胞收缩的抑制作用,iNOS抑制剂Aminoguanidine(100mmol/L)对IL-2的作用则无明显影响.200 U/ml的IL-2可降低单个心室肌细胞电刺激诱导的钙瞬变幅度;ODQ(10mmol/L)可明显抑制IL-2对心肌细胞钙瞬变的作用.以上结果提示IL-2对大鼠心肌细胞收缩功能具有直接抑制作用,其机制可能通过刺激NOS活性,增加NO的生成,激活可溶性鸟苷酸环化酶(sGC)从而导致细胞内Ca2+含量降低所致.     

白细胞介素-2降低缺氧/复氧心肌细胞对细胞内钙的处理能力

目的 :研究在正常和缺氧 /复氧过程中白细胞介素 2 (IL 2 )对心肌细胞收缩和细胞内钙的处理能力的影响。方法 :采用酶解分离大鼠心室肌细胞化学缺氧模型 ,用视频跟踪系统和细胞内双波长钙荧光系统检测单个心肌细胞收缩和细胞内钙的变化。结果 :①缺氧过程中 ,心肌细胞收缩被抑制 ,钙瞬变幅度降低、静息钙水平增高 ,咖啡因诱导的钙释放减少 ,但对细胞膜L -型钙通道活性无明显影响 ;复氧期间 ,各指标不能恢复到对照水平。②IL 2 (2× 10 5U/L)抑制心肌细胞收缩 ,使钙瞬变幅度降低、静息钙水平增高 ,使咖啡因诱导的钙释放减少。③在缺氧期间加入IL 2 (2× 10 5U/L)后 ,复氧期间各参数回复均减慢。结论 :缺氧时同时存在IL 2 ,可加剧复氧时心肌细胞收缩功能和钙处理能力的降低 ,这可能与心肌细胞肌浆网内贮钙释放减少有关。     

β2肾上腺素能受体过表达后大鼠心肌细胞收缩功能的改变及机制

目的:探讨大鼠心肌细胞过表达2-AR后心肌细胞收缩功能的改变及其可能机制.方法:采用胶原酶消化法分离培养大鼠心肌细胞,转染携带β2-A目的基因的重组腺病毒,通过免疫印迹方法检测细胞β2-AR蛋白表达的变化,通过ELISA方法检测细胞中cAMP水平的改变,采用单个细胞动态边缘检测系统测定细胞收缩功能的变化.结果:与正常心肌细胞相比,β2-AR的转染增加了细胞上β2-AR蛋白的含量(P＜0.05),并促进胞内cAMP水平的增加(14.76 3.15 pmol/ml vs 9.3 1.4pmol/ml,P＜0.05);进而增强了心肌细胞的基础收缩(8.203±2.596%vs 5.472±2.918%,P＜0.01),但不改变最大收缩(9.128±2.852%vs 9.366±2.646%).结论:β2-AR的过表达能够增加细胞内cAMP水平并改善心肌细胞的收缩功能.     

DMA增加正常大鼠心肌细胞钙瞬变和收缩

实验观察了钠氢交换或钠钙交换抑制剂 5 (N ,N 二甲基 )氨氯吡咪 (DMA)对正常和心肌肥厚大鼠分离心室肌细胞钙瞬变和细胞收缩的影响。通过负载荧光染料Fura 2 /Am ,应用离子影像分析系统 (IonImagingSystem)同步测定离体大鼠心肌细胞钙瞬变和细胞长度。结果表明 :DMA 10 μmol/L分别使钙瞬变和细胞缩短从对照组的 2 0 9.6 0± 5 4.96和 3.0 7± 0 .97μm增加到 2 38.5 0± 80 .41和 4.0 7± 1.0 2 μm (P <0 .0 5 ,n =7)。应用特异性反向钠钙交换阻断剂KB R7943可完全阻断DMA的激动作用。DMA还可使尼卡地平抑制L 型钙通道后的钙瞬变和细胞收缩增加。在肥厚心肌细胞 ,DMA表现出相同的药理作用 ,但对钙瞬变和细胞缩短的刺激作用更强。结果表明 :DMA可通过反向钠钙交换途径增加正常和肥厚大鼠心肌细胞钙瞬变和细胞收缩 ,且对肥厚心肌细胞的影响比对正常心肌细胞大。     

间歇运动对心梗大鼠梗死周边区单个心肌细胞钙瞬变和收缩功能的影响

目的:探讨间歇运动对心梗大鼠缺血心肌细胞钙瞬变和收缩功能改变的影响。方法:3月龄SD雄性大鼠24只,适应性喂养1周后随机分为假手术组(S)、心梗组(MI)、心梗+运动组(ME),每组8只。MI组结扎左冠状动脉前降支制备心梗模型;S组只穿线不结扎;ME组术后一周开始间歇训练,运动方式为1周适应性运动(10 m/min×30 min/d),然后先以10 m/min×10 min,再以15 m/min×6 min和25 m/min×4 min依次交替运功,60 min/d,每周5 d连续8周。训练结束后次日,腹腔麻醉并分离心肌细胞。采用单细胞可视化动缘探测系统(IonOptix)测定[Ca^2+]i amplitude、[Ca^2]+i荧光比率(Ratio)、Departure velocity、TTP、TTP50%、TTB50%、Return velocity以及Ratio amplitude等钙瞬变指标和±dl/dtmax、SL、PTA、SL shortening%等心肌细胞收缩指标。结果:与S组相比,MI组[Ca^2+]i amplitude、[Ca^2+]i Ratio amplitude,Departure velocity和Return velocity均显著下降(P<0.01),TTB50%、TTP和TTP50%均显著增加(P<0.01),心肌细胞肌节SL Shortening%、PTA、±dl/dtmax均显著减少(P<0.01);与MI组相比,ME组Ratio amplitude、[Ca^2+]i amplitude、Return velocity和Departure velocity均显著提高(P<0.01),TTB50%、TTP和TTP50%均显著缩短(P<0.01,P<0.05),心肌细胞肌节SL Shortening%、PTA、±dl/dtmax均显著提高(P<0.01,P<0.05)。结论:间歇运动可同步改善MI大鼠梗死周边区心肌活细胞的钙瞬变异常和心肌细胞的收缩功能。     

成年大鼠心肌细胞分离培养及兴奋-收缩耦联表征

目的：比较两种细胞分离液分离成年大鼠心肌细胞,进一步表征成年大鼠心室肌细胞兴奋-收缩耦联。方法 Langendorff装置进行主动脉逆流灌流,分别用两种细胞分离液分离成年大鼠心肌细胞,无血清培养并进行腺病毒感染。显微镜下观察单个心肌细胞的形态学特点,荧光显微镜下检测病毒感染。采用IonOptix仪器检测心肌细胞肌节收缩-舒张指标以及心肌细胞钙离子摄入-排出指标。结果两种分离液均可获得70％横纹清晰的长杆状心肌细胞,培养可存活7 d以上。腺病毒感染48 h,绿色荧光蛋白持续表达7 d以上。分离液一获得的心肌细胞不能很好地随电场刺激产生收缩,分离液二获得的细胞可用于检测兴奋-收缩耦联特性,心肌细胞肌节缩短分数为11.61％±2.15％,舒张时间为（0.177±0.031） s,钙瞬变幅度为30.79％±9.74％,钙瞬变衰减时间为（0.300±0.074） s。结论两种分离液均可用于分离和培养成年大鼠心肌细胞,并用于腺病毒转染等长时程研究。分离液二更适用于检测成年大鼠心肌细胞的兴奋-收缩耦联特性。     

白细胞介素-2对心肌负性肌力作用机制的探讨

为研究白细胞介素-2（IL-2）对心肌的负性肌力作用的可能机制,本采用酶解分离成年大鼠心室肌细胞,用细胞内双波长荧光系统和膜片钳全细胞记录检测细胞膜钙离子通道和细胞内酸碱度（pHi）及钙水平的变化,分别以fura-2/AM和BCECF/AM作为细胞内钙离子和氢离子荧光指示剂。结果：（1）IL-2(2.5-200U/ml）浓度依赖性地降低单个心室肌细胞电刺激诱导的钙瞬变幅度,使舒张末钙水平升高,选择性κ-阿片受体阻断剂nor-BNI(10nmol/L）可阻断IL-2对心肌细胞内钙的作用;（2）用200U/ml的IL-2灌流10min后,与对照组相比膜片钳全细胞记录的L-型钙电流无明显改变;（3）用200U/ml的IL-2灌流后,与对照组相比Mn^2 对fura-2/AM的淬灭率无明显改变。（4）IL-2(200U/ml）使大鼠心室肌细胞pHi降低,其作用可被选择性κ-阿片受体阻断剂nor-BNI(10nmol/L）所减弱。结论：IL-2引起的心室肌细胞pHi降低可能是其负性肌力作用机制之一,细胞膜上L-型钙离子通道可能不参与IL-2降低心肌收缩力的作用;细胞膜上的阿片受体可能介导IL-2对心肌的负性肌力作用。     

E4031对慢性心衰大鼠离体心脏功能和心肌细胞内静息Ca^2＋水平的影响

目的：研究具有钠钙交换（NCX）激动作用的药物E 4031对慢性心衰大鼠离体心脏功能和心肌细胞内静息Ca2＋水平的影响。方法：通过腹主动脉缩窄建立大鼠慢性心力衰竭模型;利用Langendorff装置进行离体心脏灌流,检测大鼠心功能及E 4031对血流动力学指标的影响;急性分离心衰大鼠心肌细胞,与钙荧光指示剂fluo3/AM共同孵育后,用激光共聚焦显微镜系统观察E 4031对心肌细胞内荧光强度的影响。结果：缩窄大鼠腹主动脉12周后,langendorff离体灌流检测显示大鼠心功能明显降低;在灌流液中加入10μmol/L E 4031可以使心衰大鼠心脏左室发展压（LVDP）和左室收缩/舒张最大速率（±dp/dtmax）提高;与正常组和伪手术组相比,心衰大鼠心肌细胞内静息钙荧光强度明显升高,和10μmol/L E 4031共孵育后,心衰大鼠心肌细胞静息钙荧光强度呈现短期先升后降过程,然后在较低的水平保持稳定。结论：E 4031可以增强慢性心衰大鼠离体心功能,可能与其增强心肌细胞膜NCX活动,稳定细胞内Ca2＋水平有关。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.