白介素-1β对高糖刺激的人肾小管上皮细胞转分化的影响

目的探讨炎症细胞因子白介素-1β（interleukin-1βIL-1β）对高糖刺激的人肾小管上皮细胞转分化的影响。-方法体外培养人肾近曲小管上皮细胞株（HKCs）,随机分为正常对照组（5.5 mmol/L normal glucose）;高糖组（30 mmol/L high glucose）;高糖＋IL-1β（5ng/ml）组。分别于处理后24h、48h、72h收集细胞,采用免疫细胞化学染色和Western蛋白印迹法检测细胞角蛋白-18（cytokeratin-18 CK-18）、α-平滑肌肌动蛋白（α-smooth muscle actinα-SMA）水平。结果高糖能够诱导肾小管上皮细胞α-SMA蛋白的合成增加,而肾小管上皮细胞的标志物CK-18的表达逐渐减少;IL-1β与高糖同时刺激可使肾小管上皮细胞α-SMA蛋白表达进一步增多,而其自身标志物CK-18的表达则明显下降。结论炎症因子IL-1β能增强高糖对肾小管上皮细胞转分化的作用。     

大黄素对TGF-β1诱导的肾小管上皮细胞间质转分化的影响

目的:探讨大黄素对TGF-β1诱导的人肾小管上皮细胞(HK-2)间质转分化的影响。方法:不同浓度大黄素分别作用于TGF-β1诱导HK-2细胞24 h和48 h,通过细胞增殖实验确定最佳大黄素最佳给药浓度。TGF-β1诱导HK-2细胞24 h后收集细胞用于免疫印迹Western blot和实时荧光定量PCR(RT-PCR)分析。Western印迹法分别检测纤维化相关蛋白Collagen IV的表达,和肾小管上皮细胞向间充质细胞转分化关键蛋白α-SMA和E-Cadherin的表达;RT-PCR法检测肾小管上皮细胞向间充质细胞转分化关键蛋白α-SMA的表达。结果:由细胞增殖实验结果表明40μM大黄素是最佳给药浓度。Western结果表明,与模型组相比,大黄素组下调纤维化相关蛋白Collagen IV的表达,大黄素组与模型组蛋白差异有统计学意义(P0.05)。与模型组相比,大黄素组下调α-SMA蛋白表达水平,而上调E-Cadherin蛋白表达,差异有统计学意义(P0.05)。RT-PCR结果表明,与模型组相比,大黄素组降低α-SMA mRNA的含量,大黄素组与模型组α-SMA mRNA含量差异有统计学意义(P0.05)。结论:大黄素可通过抑制TGF-β1诱导的HK-2细胞间质转分化,从而发挥延缓肾间质纤维化的过程。     

Wnt/β-catenin信号途径在高糖诱导肾小管上皮细胞转分化中的作用

目的探讨Wnt/β-catenin信号途径在高糖诱导肾小管上皮细胞转分化中的作用。方法体外培养人近端肾小管上皮细胞（HKC）,分为正常糖组、甘露醇对照组及高糖组。采用免疫细胞化学观察β-连环蛋白（β-catenin）表达情况;Westernblot检测Wnt4、β-catenin、E-钙粘蛋白（E-cadherin）和α-平滑肌肌动蛋白（α-SMA）表达水平;逆转录-聚合酶链反应检测Wnt4和β-cateninmRNA表达水平。结果高糖组较正常糖及渗透浓度对照组Wnt4蛋白及mRNA、α-SMA蛋白表达增高,E-cadherin表达降低,β-catenin总蛋白及mRNA水平无明显变化,细胞浆及核内蛋白表达增强。高糖刺激肾小管上皮细胞Wnt4及核β-catenin蛋白表达呈时间依赖性,于高糖刺激后12h增强,24h达到高峰。结论Wnt/β-catenin信号通路可能参与了高糖介导的肾小管上皮细胞转分化过程。     

慢病毒介导miR-4258-siRNA表达对波动高糖诱导肾小管上皮细胞转分化的影响

该实验构建了慢病毒介导(lentivirus,LV)的miR-4258-siRNA,探讨干扰miR-4258对肾小管上皮细胞(HK-2)转分化的影响。将HK-2转分化细胞分为正常组、阴性对照组和干扰组;荧光定量PCR方法检测miR-4258的表达情况;免疫印迹检测Fibronectin、E-cadherin和α-SMA的表达情况。结果显示,miR-4258-siRNA-LV可显著降低miR-4258的表达水平,干扰组的Fibronectin和α-SMA的相对表达量较阴性对照组显著下调;而E-cadherin的相对表达量较阴性对照组明显上调。研究说明,通过慢病毒介导的miR-4258-siRNA-LV下调miR-4258的表达,可抑制波动高糖诱导的HK-2细胞转分化过程。     

miR-21对TGF-β1诱导的肾小管上皮细胞间质转分化的影响

目的:观察miR-21在转化生长因子β1(TGF-β1)诱导的人肾小管上皮细胞(HK-2细胞)上皮间质转分化(EMT)中的作用,并探讨miR-21参与调控HK-2细胞EMT的可能靶点。方法:体外培养的HK-2细胞分为6组:正常对照组、转化生长因子β1(TGF-β1)模型组、miR-21 mimic阴性组、miR-21 mimic组、miR-21 inhibitor阴性组和miR-21 inhibitor组。细胞经4 ng/ml TGF-β1处理建立EMT模型,检测miR-21和EMT相关指标的表达变化,利用基因转染技术,将miR-21 mimic质粒或miR-21 inhibitor质粒转染经TGF-β1处理的HK-2细胞,使细胞过表达或抑制表达miR-21,在此基础上观察细胞EMT相关指标的变化以及磷酸酯酶(PTEN)基因的影响。结果:(1)与正常组相比,模型组的miR-21含量显著升高(P0.05),上皮表型标志物E-cadherin的mRNA和蛋白表达水平均显著降低(P0.01),间质表型标志物α平滑肌肌动蛋白(α-SMA)mRNA和蛋白水平也显著升高(P0.05,P0.01);(2)转染miR-21 mimic后,与miR-21 mimic阴性对照组相比,miR-21含量显著升高(P0.01),PTEN、E-cadherin的mRNA和蛋白水平显著降低(P0.05,P0.01),α-SMA mRNA和蛋白水平显著升高(P0.05,P0.01);转染miR-21 inhibitor后,与miR-21 inhibitor阴性对照组相比,miR-21含量显著降低(P0.01),PTEN、E-cadherin的mRNA和蛋白含量显著升高(P0.05,P0.01),α-SMA mRNA、蛋白水平显著降低(P0.05,P0.01)。结论:miR-21在TGF-β1诱导的HK-2细胞EMT发生中具有重要作用,并且可能通过靶基因PTEN参与EMT相关分子的表达调控。     

氰酸盐诱导氧化应激促进肾小管上皮细胞上皮–间充质转化

该研究探讨氰酸盐(cyanate)诱导肾小管上皮细胞氧化应激损伤和促进肾纤维化的作用。氰酸盐作用HK-2肾小管上皮细胞后, CCK8法检测其对细胞活力的影响;倒置显微镜观察细胞形态的改变; DCFH-DA法检测细胞ROS水平;细胞免疫荧光和Western blot分别检测E-cadherin、Fibronectin、α-SMA的表达; Western blot检测TGF-β的表达水平。结果显示, 2 mmol/L氰酸盐明显下调HK-2细胞的活力(P<0.05),细胞形态变为长梭形。氰酸盐作用24 h后, HK-2细胞内ROS水平呈浓度依赖性升高。免疫荧光和Western blot结果均显示,氰酸盐作用24 h后, HK-2的Fibronectin、α-SMA表达升高, E-cadherin表达下降; TGF-β的表达水平随氰酸盐浓度升高而上调(P<0.05)。以上结果表明,氰酸盐诱导肾小管上皮细胞产生过量ROS,上调TGF-β水平促进细胞上皮–间充质细胞转化(epithelia-mesenchymal transition, EMT)。     

lncRNA BANCR 在晶体上皮细胞上皮-间质转化，增殖、凋亡及自噬的作用

目的:探索长链非编码RNA BANCR(lncRNA BANCR)在人晶状体上皮细胞FHL24中对上皮-间质转化的作用,并进一步探究了其调控晶体上皮细胞增殖、凋亡及自噬的作用及相关分子机制。方法:运用qReal-time PCR检测TGF-β诱导对FHL24细胞内EMT相关标志物α-SMA,E-cadherin,Coll I,ZO1及BANCR m RNA相对表达量。细胞中转染BANCR。Western印迹检测各组细胞中EMT相关标志蛋白及LC3Ⅱ/Ⅰ的蛋白表达。MTT法检测各组细胞的增殖,凋亡情况。结果:与正常对照组比较,TGF-β诱导组细胞中BANCR,α-SMA, Coll I,ZO1 m RNA的相对表达量明显增加,而E-cadherin m RNA显著下降,差异均有统计学意义(t=-5.031,-7.145,-9.023,-6.012, 5.097均P0.05),以上因子的蛋白表达趋势相同,差异均有统计学意义(均P0.05)。si RNA-BANCR TGF-β诱导组细胞中E-cadherin m RNA相对表达量比si RNA TGF-β诱导组显著增加(t=-9.98, P0.05);α-SMA,Coll I及ZO1 m RNA相对表则显著减少(t=9.003; 27.738; 19.620, P0.05)。抑制BANCR后细胞增殖活力48、72 h时细胞活性显著降低(t=5.032, 9.041,均P0.05),细胞凋亡率显著升(t=16.772,P0.001)。自噬标志蛋白LC3-II/LC3-I比例增加(P 0.05)。结论:长链非编码BANCR参与了晶体上皮细胞的抑制其上皮-间质转化,抑制BANCR可抑制晶体上皮细胞增殖、增加凋亡及自噬的发生。     

靶向沉默HIF-1α信号通路抑制百草枯中毒诱导的肺泡上皮细胞上皮间质转化

目的:探讨HIF-1α信号通路在百草枯(paraquat,PQ)诱导大鼠Ⅱ型肺泡上皮细胞上皮间质转化(Epithelial-mesenchymal transition,EMT)中的作用机制。方法:使用20μmol/L浓度的百草枯溶剂对大鼠Ⅱ型肺泡上皮RLE-6TN细胞干预24 h,随后在倒置光学显微镜观察各组细胞形态学变化;用real-time PCR与Western blot法检测RLE-6TN细胞中HIF-1α、上皮表型标记蛋白E-cadherin及间质表型标记蛋白Vimentin的表达,Transwell侵袭实验检测各处理组细胞侵袭能力的改变;使用HIF-1α靶向si RNA抑制其表达后,进一步采用RT-PCR和Western blot检测HIF-1α、E-cadherin和Vimentin的表达水平,Transwell法检测细胞侵袭能力变化。结果:体外百草枯溶液可显著诱导大鼠Ⅱ型肺泡上皮细胞RLE-6TN细胞HIF-1α表达升高和上皮间质转化的发生,同时细胞的体外侵袭能力也增强。靶向沉默HIF-1α基因后,百草枯诱导的上皮间质转化过程被逆转,同时细胞侵袭能力显著减弱。结论:百草枯通过调控HIF-1α信号通路来诱导RLE-6TN细胞上皮间质转化的发生,进而促进肺纤维化的形成。     

α-晶状体蛋白在高糖培养人RPE细胞中的表达机制研究

目的：研究高糖体外培养环境下人RPE细胞的α晶状体蛋白的表达变化,并排除渗透影响因素下与正常糖浓度组进行比较。方法：对人RPE细胞（ARPE-19）在D-葡萄糖为5.5mM条件下培养和传代3周,然后按培养条件分为3组,即5.5mM葡萄糖,33mM葡萄糖,5.5mM葡萄糖＋27.5mM甘露醇,再分别培养12h,24h,48h后进行总蛋白和RNA提取。进而通过Western blotting对αA晶状体蛋白,αB晶状体蛋白以及凋亡相关蛋白bax,bcl-2进行检测,并通过RT-PCR检测αA-,αB-两种晶状体蛋白的mRNA水平。结果：相对于正常糖浓度组,高糖环境下αA晶状体蛋白,αB晶状体蛋白均表达明显上调,渗透变化对蛋白表达没有明显影响。高糖培养环境下,随着时间延长,bcl-2表达量明显下降,而bax表达量呈上升趋势,αA-,αB-两种晶体蛋白的mRNA变化趋势与蛋白水平一致。结论：高糖培养环境下人视网膜色素上皮细胞的αA-,αB-两种晶状体蛋白均明显表达上调。     

p~(38)MAPK在IL-18诱导肾小管上皮细胞转分化中的作用

目的:白细胞介素18(IL-18)可诱导肾小管上皮细胞转分化,本研究探讨其是否是通过p38MAPK途径而起作用。方法:应用不同浓度的p38MAPK通路特异性阻断剂SB203580(0、5、10、20μmol/L)预孵育人近端肾小管上皮细胞(HK-2细胞)30min后,加入IL-18(100ng/ml)共培养24、48、72h。应用RT-PCR法检测α-平滑肌肌动蛋白(α-SMA)mRNA的表达水平;应用ELISA法测定细胞浆中α-SMA蛋白质含量。结果:SB203580呈剂量依赖性地抑制IL-18诱导的HK-2细胞α-SMA基因表达(P0.05)。结论:p38MAPK通路是调控IL-18诱导肾小管上皮细胞转分化的主要信号通路之一。     

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.     

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.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.