孤儿核受体Rev-erbα在小鼠骨髓间充质干细胞成骨分化中的作用

背景:孤儿核受体Rev-erbα已被证明在骨代谢过程中发挥着重要作用,但其在调节骨髓间充质干细胞成骨分化过程中的具体机制尚不明确。目的:研究孤儿核受体Rev-erbα是否参与小鼠骨髓间充质干细胞成骨分化能力的调节。方法:以全骨髓贴壁法进行小鼠骨髓间充质干细胞的分离培养,构建携带Rev-erbα基因的慢病毒载体并转染至体外培养的第3代骨髓间充质干细胞,转染后48 h采用Real Time PCR法检测Rev-erbα基因水平的表达。实验分为3组:实验组骨髓间充质干细胞转染过表达Rev-erbα基因及EGFP基因的慢病毒载体,阳性对照组骨髓间充质干细胞转染含EGFP基因的空病毒载体,设立不含病毒原液的阴性对照组,分别进行成骨诱导并在第0,7,14天采用Real Time PCR法检测成骨分化相关指标碱性磷酸酶、骨桥蛋白、骨钙素的表达变化。结果与结论:①携带Rev-erbα基因的慢病毒成功转染到骨髓间充质干细胞中并稳定表达;②随着成骨诱导时间延长,成骨相关指标碱性磷酸酶、骨桥蛋白表达增加,但组间差异无显著性意义,骨钙素表达增加,但实验组明显低于阳性对照组和对照组(P<0.05);③结果表明,Rev-erbα转染后骨髓间充质干细胞成骨分化能力降低,成骨晚期标志物骨钙素的表达受抑制,说明Rev-erbα在骨髓间充质干细胞成骨分化的晚期阶段有重要作用。     

酪蛋白激酶2相互作用蛋白1调控骨质疏松大鼠骨髓间充质干细胞的成骨能力北大核心

背景:目前对酪蛋白激酶2相互作用蛋白1(casein kinase 2-interaction protein-1,CKIP-1)分子机制的体外研究主要集中在基因敲除小鼠来源成骨细胞或骨髓间充质干细胞,鲜见报道骨质疏松模型大鼠来源骨髓间充质干细胞中CKIP-1表达的研究。目的:探讨下调CKIP-1基因前后骨质疏松状态骨髓间充质干细胞成骨分化能力的变化。方法:用维甲酸诱导雌性SD大鼠骨质疏松模型,采用全骨髓贴壁法体外培养骨质疏松组、正常组大鼠骨髓间充质干细胞。成骨诱导后进行茜素红染色、碱性磷酸酶染色及实时定量RT-PCR检测骨桥蛋白、Runx2 mRNA的相对表达,实时定量RT-PCR检测成骨诱导过程中2组细胞中CKIP-1的动态表达;通过基因转染沉默CKIP-1基因,成骨诱导后进行茜素红染色、碱性磷酸酶染色及实时定量RT-PCR检测骨桥蛋白、Runx2 mRNA的相对表达。结果与结论:①与正常组相比,骨质疏松组骨髓间充质干细胞茜素红染色钙结节定量、碱性磷酸酶活性及骨桥蛋白、Runx2的mRNA水平降低(P<0.05),骨质疏松组骨髓间充质干细胞中CKIP-1基因动态表达水平总体偏高;②与未下调CKIP-1的骨质疏松组骨髓间充质干细胞相比,下调CKIP-1基因表达后,骨质疏松组骨髓间充质干细胞茜素红染色钙结节定量、碱性磷酸酶活性及骨桥蛋白、Runx2的mRNA水平明显升高(P<0.05);③结果表明,维甲酸诱导的骨质疏松大鼠骨髓间充质干细胞成骨能力降低,下调CKIP-1基因可以部分提高其成骨分化能力。     

小鼠间充质干细胞三系分化中SOX9与Ⅱ型胶原mRNA的定量

背景：研究表明,软骨中的主要成分Ⅱ型胶原的基因-Col2a1在软骨细胞中的表达与SOX9 的浓度呈剂量依赖正相关关系。 目的：通过成骨、成软骨、成脂肪诱导干细胞分化,分析3种分化过程及不同时期的SOX9与Ⅱ型胶原 mRNA含量的变化,探讨SOX9在不同时空分布的表达规律及与Ⅱ型胶原的相关关系。 方法：取4周龄昆明小鼠骨髓间充质细胞,体外培养得到间充质干细胞并传达至第3代,对间充质干细胞进行流式细胞仪鉴定细胞表型,共分3组每组设3个时间段,通过成骨、成软骨、成脂肪3种诱导培养液对3组细胞进行诱导,另设不进行诱导的细胞作为对照组。分别在诱导3,7,14 d后收集提取细胞的总RNA,通过RT-PCR进行SOX9与Ⅱ型胶原的mRNA定量检测,同时对诱导后的细胞进行染色、免疫荧光染色,观察其分化状态及相关统计分析。 结果与结论：第3代骨髓间充质干细胞生长良好,流式细胞仪鉴定细胞表型证实为干细胞,对诱导后细胞进行染色、免疫荧光染色结果证实细胞分化为骨、软骨、脂肪细胞。经RT-PCR检测,在3组诱导分化细胞中SOX9 mRNA含量由高到低分别是成软骨、成骨、成脂肪,Ⅱ型胶原 mRNA含量由高到低分别是成软骨、成脂肪、成骨。在成软骨分化中SOX9在3,7 d表达不断升高,14 d呈下降趋势。Ⅱ型胶原在3,7,14 d均逐渐升高。在成骨分化中SOX9 mRNA含量随着时间推移而增加,而Ⅱ型胶原则随着时间推移而不断降低。在成脂肪分化中SOX9 mRNA表达与对照组比较差异无显著性意义(P > 0.05);而Ⅱ型胶原的表达没有规律可循,时间点的延伸及检测未观察到。结果提示,SOX9在软骨分化中作用优于成骨、成脂肪组,且软骨分化中SOX9与Ⅱ型胶原存在相关性,可能在软骨分化的早期Ⅱ型胶原随着SOX9的变化而变化;且软骨分化和成骨分化过程中SOX9可能起到了一个互相协调促进平衡的关键作用。     

反转录病毒介导基质金属蛋白酶特异性组织抑制剂1基因转染骨髓间充质干细胞的成骨能力

背景：间充质干细胞成骨分化早期是成骨性细胞外基质细胞外间质的堆积,其主要成分均是基质金属蛋白酶特异性组织抑制剂1(tissue Inhibitor of Metalloproteinases 1,TIMP-1)作用的基质金属蛋白酶作用底物,通过增加TIMP-1的表达,是否可以抑制基质金属蛋白酶的活性,进而影响间充质干细胞的成骨分化能力？ 目的：观察反转录病毒介导TIMP-1基因转染骨髓间充质干细胞的成骨能力。 方法：设计In-Fusion引物扩增TIMP-1基因片段,与酶切后线性化的pBABE-Puro反转录病毒表达载体进行位点同源重组,构建TIMP-1基因反转录病毒表达载体,RT-PCR和测序验证,并使用GP-293细胞进行包装。通过密度梯度法分离人下颌骨来源骨髓间充质干细胞,流式细胞仪验证其表面标记。将TIMP-1反转录病毒包装液感染骨髓间充质干细胞并筛选稳定表达TIMP-1细胞株,RT-PCR和蛋白电泳验证其表达,并使用成骨诱导液诱导10 d,观察其对成骨能力的影响,RT-PCR验证其成骨相关基因的表达。 结果与结论：经测序证实,RT-PCR所获得的TIMP-1基因 cDNA 序列与NM_003254.2(624 bp)序列完全一致;通过表面标记鉴定证实成功分离了人骨髓间充质干细胞,并具良好的成骨、成脂能力;RT-PCR和Western Blot验证反转录病毒载体成功介导TIMP-1转染骨髓间充质干细胞,RT-PCR表明成骨诱导10 d时,TIMP-1基因感染组成骨增加,RUNX-2和COL1 mRNA表达增加。提示TIMP-1基因可通过反转录病毒成功转染人骨髓间充质干细胞,并能促进其成骨。     

胰岛素样生长因子Ⅰ转染对脂肪干细胞TWEAK/Fn14信号通路的影响

背景：胰岛素样生长因子Ⅰ具有诱导间充质干细胞向软骨细胞分化的能力,通过基因转染的方式将胰岛素生长因子Ⅰ转染入脂肪干细胞或许能够更好的促进脂肪干细胞向软骨细胞分化。 目的：探讨胰岛素样生长因子Ⅰ基因对脂肪间充质干细胞体外定向成软骨分化能力以及对TWEAK/Fn14信号通路的影响。 方法：构建慢病毒载体pLVX-IGF-I-IRES-ZsGreenl基因并转染第3代人脂肪间充质干细胞,诱导细胞向软骨分化。同时将pLVX-IRES-ZsGreenl转染的脂肪间充质干细胞设为绿色荧光蛋白/脂肪间充质干细胞组,单纯脂肪间充质干细胞设为对照组。 结果与结论：与绿色荧光蛋白/脂肪间充质干细胞组和对照组相比,胰岛素样生长因子Ⅰ/脂肪间充质干细胞组细胞中TWEAK mRNA表达水平降低,而胰岛素样生长因子Ⅰ,Col2al和Sox9 mRNA的表达水平增加,Col2a1表达水平增加,基质金属蛋白酶3以及TWEAK的表达降低。提示pLVX-IGF-I-IRES-ZsGreenl慢病毒转染脂肪间充质干细胞后可获得胰岛素样生长因子Ⅰ的高效表达,且能下调细胞TWEAK基因及蛋白表达,可促进体外培养的脂肪间充质干细胞转化为软骨细胞。 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

miR-889-3p靶向Runx2抑制脂肪间充质干细胞的成骨分化

背景：研究表明miR-889-3p可参与调控多种细胞的增殖分化过程,但是其对脂肪间充质干细胞成骨分化的影响尚不明确。目的：探讨miR-889-3p对脂肪间充质干细胞成骨分化的调节作用。方法：体外分离、培养SD大鼠脂肪间充质干细胞,转染miR-889-3p模拟物(miR-889-3p mimic)、miR-889-3p抑制剂(miR-889-3p inhibitor)和阴性对照(miR-NC)并诱导成骨分化。RT-PCR和Western blot检测成骨诱导14 d后的碱性磷酸酶活性、Runx2、骨钙素、骨桥素、Osterix等成骨相关mRNA和蛋白的表达。将野生型Runx2、突变型Runx2分别与miR-889-3p模拟物和miR-NC共转染脂肪间充质干细胞,通过双荧光素酶报告基因检测miR-889-3p与Runx2的靶向关系。结果与结论：(1)miR-889-3p表达水平随成骨诱导时间延长而逐渐减低;(2)成骨诱导第7天和第14天miR-889-3p inhibitor组的矿化结节数目、大小、颜色深浅明显超过miR-889-3p mimic组和miR-NC组(P <0.05);...     

重组慢病毒载体转染羊骨髓间充质干细胞及成骨基因表达量的变化

背景：碱性成纤维细胞生长因子具有促进骨髓基质细胞分裂增殖作用,而骨形态发生蛋白2在诱导新骨形成方面有重要的研究意义。目的：分析骨形态发生蛋白2和碱性成纤维细胞生长因子基因单独和联合转染对体外培养的骨髓间充质干细胞的增殖和骨向分化的影响,比较Ⅰ型胶原蛋白、骨钙蛋白、骨桥蛋白等非特异性成骨基因于转染前后骨髓间充质干细胞相对表达量的差别,为构建组织工程骨的种子细胞做理论依据。方法：构建碱性成纤维细胞生长因子、骨形态发生蛋白2慢病毒载体,分别转染羊骨髓间充质干细胞,得到碱性成纤维细胞生长因子组、骨形态发生蛋白2组、联合组、对照组细胞,提取RNA后采用实时定量PCR的方法检测Ⅰ型胶原蛋白、骨钙蛋白、骨桥蛋白等非特异性成骨基因的mRNA水平相对表达量的变化。结果与结论：对照组、碱性成纤维细胞生长因子、骨形态发生蛋白2及联合组4组间非特异性成骨基因表达量存在明显差异(P < 0.05),且碱性成纤维细胞生长因子和骨形态发生蛋白2具有交互作用(P < 0.05),碱性成纤维细胞生长因子、骨形态发生蛋白2联合组的Ⅰ型胶原蛋白和骨钙蛋白基因表达量明显高于其他3组,且差异有显著性意义(P < 0.05);但是骨桥蛋白基因中差异无显著性意义(P > 0.05)。体外实验结果显示联合转染组中的Ⅰ型胶原蛋白、骨钙蛋白、骨桥蛋白等非特异性成骨基因mRNA水平相对表达量较多,该组细胞的成骨功能最强,适合作为组织工程骨的种子细胞。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

miR-302对脂肪间充质干细胞向成脂和成骨分化的调控

背景：间充质干细胞具有自我更新能力,在一定条件下能够分化为一定谱系的细胞,但很多机制至今未明。 目的：探究miR-302b对脂肪间充质干细胞向成脂和成骨分化的调控作用。 方法：miR-302模拟物转染作用于脂肪间充质干细胞进行成骨成脂诱导,对照组转染miR-302阴性对照模拟物miR-NC。采用碱性磷酸酶染色及活性分析、茜素红染色、油红O染色和萃取实验观察miR-302上调对脂肪间充质干细胞成骨和成脂分化的影响,以及Western blot检测miR-302上调后成骨分化转录因子Runx2和成骨早期标志物碱性磷酸酶在脂肪间充质干细胞中的表达。 结果与结论：①碱性磷酸酶沉淀物在miR-302过表达细胞中产生的量均明显少于对照组,进一步发现miR-302过表达实验组碱性磷酸酶活性明显低于对照组(P < 0.05)。②miR-302过表达明显抑制了矿物质沉积钙结节的形成,miR-302上调实验组橘红色的钙结节明显少于对照组。③miR-302过表达实验组油红O染色阳性的细胞数明显高于对照组,进一步表明实验组细胞萃取得到的油红O吸光度值明显上升(P < 0.05)。④成骨诱导第6天时成骨分化转录因子Runx2和成骨早期标志物碱性磷酸酶在miR-302过表达的细胞中都有不同程度的下降。⑤以上结果表明miR-302的上调能够抑制脂肪间充质干细胞的成骨分化,同时促进其向成脂分化。miR-302在间充质干细胞向成脂和成骨分化平衡发挥了双向调控作用。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

间充质干细胞体外成骨及成脂肪分化中sortilin基因表达的研究

目的:研究间充质干细胞(MSC)体外成骨、成脂肪分化时sortilin基因表达变化。方法:从骨髓中分离培养人间充质干细胞,流式细胞术鉴定其表型,加入成骨和成脂肪诱导剂,RT-PCR检测成骨标志骨桥蛋白(Op)和成脂肪标志脂蛋白脂肪酶(LpL)表达,半定量RT-PCR分析sortilin基因随诱导时间的表达变化。结果:①体外分离培养的人间充质干细胞能向成骨细胞和脂肪细胞分化,分别表达Op和LpL。②在体外成骨分化过程中,sortilin基因表达第1d开始上调,持续约1周后恢复至原水平;在培养3d时,sortilin基因表达明显上调,与未诱导组相比,差异明显(P＜0.01)。③体外成脂肪分化时,sortilin基因表达无明显的改变(P＞0.05)。结论:sortilin基因可能参与间充质干细胞成骨分化的调节,而与成脂肪分化无关,通过调控sortilin基因的表达,有可能为成骨障碍性疾病的治疗,提供新的思想和策略。     

脂多糖干预卵巢切除小鼠骨髓间充质干细胞的成骨分化能力

背景：骨平衡被打破造成的绝经后骨质疏松往往是由炎症因子的升高造成的,在牙周组织中,炎症致病菌可产生破坏宿主组织的毒性因子,如脂多糖、菌毛、凝集素等。 目的：通过建立卵巢切除小鼠骨质疏松的模型,观察脂多糖刺激下骨髓间充质干细胞成骨分化能力的改变,初步探讨雌激素缺乏患者易患牙周炎的细胞分子生物学机制,为绝经后妇女牙周炎的防治提供实验基础。 方法：30只雌性昆明小鼠随机均分为卵巢切除组和对照组。取两组小鼠的骨髓间充质干细胞进行成骨诱导,同时加入不同质量浓度脂多糖(0,10,100 μg/L)刺激,检测碱性磷酸酶染色、茜素红染色以及碱性磷酸酶活性;RT-PCR之后检测成骨相关分子Runx2、Ocn、Alp的表达。 结果与结论：成骨诱导7 d后可表达碱性磷酸酶,21 d后茜素红染色阳性,可形成矿化结节。检测发现成骨相关分子Ocn、Runx2、Alp的mRNA在成骨诱导后均有表达;3种目的基因在100 μg/L脂多糖作用下表达显著下降(P < 0.05),卵巢切除小鼠骨髓间充质干细胞在脂多糖作用下3种基因表达较对照组下降显著(P < 0.05)。卵巢切除小鼠骨髓间充质干细胞的成骨分化能力下降;脂多糖使骨髓间充质干细胞的成骨分化能力下降,卵巢切除组小鼠骨髓间充质干细胞成骨分化能力在脂多糖作用下明显减弱,这可能是雌激素缺乏导致牙周炎易感性增加的细胞分子学机制之一。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

ACS患者动脉粥样斑块稳定性相关细胞因子变化的临床意义

目的：探讨急性冠脉综合征（ACS）病人血清中的细胞因子基质金属蛋白酶-1（MMP-1）、金属蛋白酶组织抑制剂-1（TIMP1）、sICAM-1、sVCAM-1水平与冠状动脉粥样硬化斑块稳定性的相关性及临床意义。方法：用ELISA检测ACS病人血清中sICAM-1、sVCAM-1、MMP-1、TIMP-1水平,并与正常对照组进行比较分析。结果：ACS组血清sICAM-1、sVCAM-1、MMP-1、TIMP-1水平均非常明显地高于正常对照组（P〈0.01）。结论：外周血中sICAM-1、sVCAM-1、MMP-1、TIMP-1水平增高与ACS相关,能反映冠脉粥样硬化斑块的稳定情况,可作为评价冠状动脉粥样硬化斑块稳定性与病变严重程度的一个参考指标。     

HLA Class II (DRB,DQA1 and DQB1) Allele and Haplotype Frequencies in the Patients with Pemphigus Vulgaris

Introduction  Pemphigus vulgaris (PV), an autoimmune disease affecting the skin and mucous membranes, is associated with some human leukocyte antigen (HLA) class II alleles and haplotypes. Materials and Methods  In order to evaluate the association of HLA-DR and DQ alleles and haplotypes in Iranian non-Jewish patients with PV, 52 patients with PV and 180 normal subjects as control group were investigated in this study. Results and Discussion  HLA-DRB1*04, -DRB1*1401, -DRB4, -DQA1*0104, -DQA1*03011, -DQB1*0302, and -DQB1*0502 alleles have been significantly increased in our patients group. Moreover, the haplotypes HLA-DRB1*04/-DQA1*03011/-DQB1*0302 and HLA-DRB1*1401/-DQA1*0104/-DQB1*0502 increased significantly in our patients. In contrast, the following alleles decreased significantly in our patients: HLA-DRB1*15, -DRB1*0301, -DRB1*07, -DRB1*11, -DRB5, -DQA1*0101, -DQA1*0103, -DQA1*201, -DQA1*05, -DQB1*0201, -DQB1*0301, -DQB1*06011, and -DQB1*0602. In addition, HLA-DRB1*15/-DQA1*0103/-DQB1*06011, HLA-DRB1*0301/-DQA1*05011/-DQB1*0201, HLA-DRB1*07/-DQA1*0201/-DQB1*0201, and HLA-DRB1*11/-DQA1*05/-DQB1*03011 decreased significantly in our patients. Genetic factors are involved in the occurrence of PV; HLA-DRB1*04 and -DRB1*1401 alleles and the related haplotypes are suggestive to be two major PV susceptibility factors in our population study.     

Molecular analysis of HLA class II polymorphism in Croatians

Abstract: HLA-class II polymorphisms have been studied in a population of 141 unrelated healthy Croatians using PCR amplification, followed by non-radioactive oligonucleotide hybridization. Thirty one DRB1, 8 DQA1, 13 DQB1 and 16 DPB1 alleles were found in the tested population. DRB1*1601, 0701, 1501, 0101 and 1104 are the most frequent alleles at the DRB1 locus. At the DQA1 locus two alleles predominate: DQA1*0501 and 0102, while the most frequent DQB1 allele is *0301. Analysis of HLA-DPB1 polymorphism showed that, as in other Europeans, DPB1* 0401 is the most frequent allele. Four different two locus haplotypic associations (DRB1-DRB3, DRB1-DRB5, DRB1-DQB1 and DQA1-DQB1) as well as three locus DRB1-DQA1-DQB1 haplotypic associations were assigned on the basis of known linkage disequilibria. Several unusual two-locus associations have been observed: DRB1*0301-DRB3* 0202, DRB1*1501-DRB5*02, DRB1*1601-DRB5*0101, DRB1*1502-DRB5*0101, DQA1*0103-DQB1*0503 and DQA1*0501-DQB1*0302. Among 236 examined DRB1-DQA1-DQB1 haplotypic combinations, the most frequent was DRB1*1601-DQA1*0102-DQB1*0502 that was found with statistically significant higher frequency than in other Europeans. Twenty-eight distinct probable haplotypes were observed just once, suggesting that the main characteristic of Croatian population is great heterogeneity of haplotypes. This study will serve as a reference for further anthropology studies, HLA and disease associations studies and for donor/recipient matching in organ and bone marrow transplantation.     

HLA class II allele and haplotype frequencies in Koreans based on 107 families

We have investigated the distribution of HLA class II alleles and haplotypes in 107 Korean families (207 parents and 291 children) for the HLA-DRB1, DRB3/B4/B5, DQA1, DQB1 and DPB1 loci. Numbers of alleles observed for each locus were DRB1: 25, DQA1: 14, DQB1: 15, and DPB1: 13. Only two to three alleles were observed for the DRB3 (*0101, *0202, *0301), DRB4 (*0103, * 0103102 N), and DRB5 (*0101, *0102) loci. These alleles showed strong associations with DRB1 alleles: DRB3*0101 with DRB1*1201, *1301 and *1403; DRB3*0301 with DRB1*1202 and *1302; DRB3*0202 with DRB1*0301, *1101, *1401 and *1405; DRB5*0101 and *0102 were exclusively associated with DRB1*1501 and *1502, respectively. The seven most common DRB1-DQB1 haplotypes of frequencies > 0.06 accounted for 52% of the total haplotypes. These haplotypes were exclusively related with the seven most common DRB1-DRB3/B4/B5-DQA1-DQB1 haplotypes: DRB1*1501-DRB5*0101-DQA1*0102-DQB1*0602 (0.085), DRB1*0405-DRB4*0103-DQA1*0303-DQB1*0401 (0.082), DRB1*09012-DRB4*0103-DQA1*0302-DQB1*03032 (0.082), DRB1*0101-DQA1*0101-DQB1*0501 (0.075), DRB1*0701-DRB4*0103-DQA1*0201-DQB1*0202 (0.065), DRB1*0803-DQA1*0103-DQB1*0601 (0.065), and DRB1*1302-DRB3*0301-DQA1*0102-DQB1*0604 (0.065). When these haplotypes were extended to the DPB1 locus, much diversification of haplotypes was observed and only one haplotype remained with a frequency of > 0.06: DRB1*0405-DRB4*0103-DQA1*0303-DQB1*0401-DPB1*0501 (0.062). Such diversification would have resulted from cumulated events of recombination within the HLA class II region, and the actual recombination rate observed between the HLA-DQB1 and DPB1 loci was 2.3% (10/438 informative meioses, including 2 recombinants informative by analysis of TAP genes). Comparison of the distribution of DRB1-DQB1 haplotypes with other populations revealed that Koreans are closest to Japanese people. However, Koreans share a few haplotypes with white people and Africans, which are rare in Japanese: DRB1*0701-DQB1*0202 and DRB1*1302-DQB1*0609. The results obtained in this study will provide useful information for anthropology, organ transplantation and disease association studies.     

Polymorphism of human leukocyte antigen-DRB1, -DQB1, and -DPB1 genes of Shandong Han population in China

In the present study, polymerase chain reaction-sequence-based typing (PCR-SBT) was used to analyze human leukocyte antigen (HLA)-DRB1, -DQB1, and -DPB1 alleles of 98 unrelated healthy Shandong Han individuals. A total of 60 alleles, in which 28 in DRB1, 15 in DQB1 and 17 in DPB1 were found. Among the 28 detected DRB1 alleles, DRB1*150101, DRB1*070101, DRB1*090102, DRB1*120201, and DRB1*080302 were commonly observed, with frequencies of 16.3%, 11.2%, 10.2%, 8.2%, and 5.6%, respectively. The most predominant DQB1 allele was DQB1*030101/0309 with the frequency of 20.4%, followed by DQB1*0201/0202 (14.8%), DQB1*0602 (14.3%), DQB1*030302 (12.2%), and DQB1*060101/060103 (10.7%). Of the 17 detected DPB1 alleles, DPB1*0501 was the most frequent allele with the frequency of 37.2%. DPB1*020102 (18.4%), DPB1*040101 (11.2%), DPB1*0402 (7.1%), and DPB1*1701 (6.6%) were also very frequent alleles. A total of 53 estimated DRB1-DQB1 two-locus haplotypes were observed in Shandong Han population, of which DRB1*150101-DQB1*0602 was the most predominant, followed by DRB1*090102-DQB1*030302, DRB1*070101-DQB1*0201/0202 DRB1*120201-DQB1*030101/0309, and DRB1*080302- DQB1*060101/060103. The distribution of the HLA class II alleles and haplotypes frequencies as well as the dendrogram showed that the Shandong Han population belongs to the northern group of Chinese. The data have implications for anthropological studies and disease associations.     

The distribution of HLA class II susceptible/protective haplotypes could partially explain the low incidence of type 1 diabetes in continental Italy (Lazio region)

HLA class II is the primary susceptibility gene to type 1 diabetes and the analysis of HLA class II association could help to clarify the relative weight of genetic contribution to the incidence of the disease. Here we present an extensive typing for HLA class II alleles and their haplotypes in a homogenous population of type 1 diabetic patients (n=134) and controls (n=128) and in simplex (n=100) and multiplex families (n=50) from continental Italy (Lazio region). Among the various haplotypes tested, the DRB1*0301-DQA1*0501-DQB1*0201 was the most frequent found in type 1 diabetic patients and was transmitted in 82% of affected siblings, whereas DRB1*0402-DQA1*0301-DQB1*0302 appeared to have the highest odds ratio (10.4), this haplotype was transmitted in 96.3% of affected siblings, followed by DRB1*0405-DQA1*0301-DQB1*0302, DRB1*0405-DQA1*0301-DQB1*0201, DRB1*0401-DQA1*0301-DQB1*0302 and DRB1*0404-DQA1*0301-DQB1*0302. The following haplotypes showed a significant decreased transmission to diabetic siblings: DRB1*0701-DQA1*0201-DQB1*0303, DR2-DQA1*01-DQB1*0602, DR5-DQA1*0501-DQB1*0301. We suggest that the HLA DR/DQ haplotype/genotype frequencies observed could in part explain the low incidence of type 1 diabetes registered in Lazio region (8.1/100.000/year), for a number of reasons: i) the low frequency, in the general control population, of the most susceptible haplotypes and genotype for type 1 diabetes DRB1*0301-DQA1*0501-DQB1*0201 (14%), and DR4-DQA1*0301-DQB1*0302 (9%) and DRB1*0301-DQA1*0501-DQB1*0201/DR4-DQA1*0301-DQB1*0302 (0.8%) compared to other countries characterised by high incidence rate of the disease, Sardinia and Finland, respectively; ii) a significant lower ratio, in the control population, between the susceptible DRB1*0301-DQA1*0501-DQB1*0201 and the neutral DRB1*0701-DQA1*0501-DQB1*0201 haplotypes compared to the Sardinian population; iii) the high frequency of protection haplotypes/genotypes as the DR5-DQA1*0501-DQB1*0301, and DR5-DQA1*0501-DQB1*0301/DR5-DQA1*0501-DQB1*0301 very common in the control population of Lazio region and the DRB1*1401-DQA1*0101-DQB1*0503 haplotype.     

HLA class IIDRB, DQA1 and DQB1 polymorphisms in the Polish population from Wielkopolska

HLA DRB1, DQA1 and DQB1 alleles were determined by DNA PCR-SSO typing in a sample of 99 individuals originating from Wielkopolska (midwestern Poland). A high number of alleles (38 DRB1, 8 DQA1 and 14 DQB1) was detected at each locus, many of them presenting notable frequencies in this population. The three HLA loci are thus characterized by very high heterozygosity levels (93% for DRB1, 85% for DQA1, and 88% for DQB1), which confirms the results found for other European populations. A total of 6 DRB1-DQA1-DQB1 haplotypes are detected with an estimated frequency higher than 5%, namely, DRB1*1501-DQA1*0102-DQB1*0602, DRB1*0701-DQA1*0201-DQB1*0201, DRB1*0101-DQA1*0101-DQB1*0501, DRB1*1101-DQA1*0501-DQB1*0301, DRB1*03011-DQA1*0501-DQB1*0201, and DRB1*1301-DQA1*0103-DQB1*0603. A genetic distance analysis between the Polish and other world populations tested for HLA class II indicates that the Wielkopolska community is close to geographically close, rather than linguistically related populations from Europe. More generally, a good agreement between genetics and geography is found for DRB1 and DQB1 polymorphisms in Europe, suggesting that these two loci are highly informative for assessing historical relationships among humans.     

Human antibodies to herpes simplex virus type 1 glycoprotein C are neutralizing and target the heparan sulfate-binding domain

Human antibodies specific for glycoprotein C (gC1) of herpes simplex virus type 1 (HSV-1) neutralized the virus infectivity and efficiently inhibited attachment of HSV-1 to human HaCaT keratinocytes and to murine mutant L cells expressing either heparan sulfate or chondroitin sulfate at the cell surface. Similar activities were observed with anti-gC1 monoclonal antibody B1C1. In addition to HaCaT and L cells, B1C1 antibody neutralized HSV-1 infectivity in simian GMK AH1 cells mildly pre-treated with heparinase III. Human anti-gC1 antibodies efficiently competed with the binding of gC1 to B1C1 antibody whose epitope overlaps a part of the attachment domain of gC1. Human anti-gC1 and B1C1 antibodies extended survival time of mice experimentally infected with HSV-1. We conclude that in HaCaT cells and in cell systems showing restricted expression of glycosaminoglycans, human and some monoclonal anti-gC1 antibodies can target the cell-binding domain of this protein and neutralize viral infectivity.     

Notch1在前列腺癌细胞PC3中对其配体Jagged1表达的调控

目的探讨在前列腺癌细胞PC3中,Notch1和Jagged1对细胞生长的影响,及Notch1对其配体Jagged1表达的调控作用。方法通过siRNA干扰的方法分别抑制Notch1和Jagged1蛋白的表达,用MTT法检测PC3细胞的生长;分别通过siRNA干扰和转染质粒的方法,抑制和促进PC3细胞中Notch1蛋白的表达,用Western blot检测Jagged1蛋白水平,用Real-timePCR检测Jagged1 mRNA水平。结果抑制Notch1和Jagged1蛋白的表达后,PC3细胞的生长减慢;抑制Notch1的表达引起Jagged1的蛋白水平下降而过表达Notch1引起Jagged1的蛋白水平上升,同时,Jagged1蛋白水平与mRNA水平的变化不一致。结论Notch1和Jagged1对前列腺癌细胞PC3的生长有重要影响。Notch1可以调控其配体Jagged1的表达。     

Combined UGT1A1 and UGT1A7 variant alleles are associated with increased risk of Gilbert's syndrome in Taiwanese adults

Gilbert's syndrome (GS) is caused by a reduction in the activity of hepatic bilirubin UDP-glucuronosyltransferase (UGT). This reduction is associated with UGT1A1*28 and UGT1A1*6 polymorphisms. Recent research also showed that carriage of UGT1A1*6 allele were significantly related with UGT1A7*3. Polymerase chain reaction-restriction fragment length polymorphism were utilized to determine UGT1A7 and UGT1A1 genes for 207 patients with GS and 207 gender/age-matched healthy controls. For the 207 healthy controls, linkage disequilibrium was observed between -57UGT1A7 and 622UGT1A7 loci (D' = 1.00 and r(2) = 1.00), -57UGT1A7 and 211UGT1A1 loci (D' = 0.72 and r(2) = 0.36), respectively. A dose-response effect for number of at-risk allele of UGT1A1 and risk for GS was noted (odds ratio (OR) = 8.19 for heterozygous UGT1A1*28 genotype; OR = 124.96 for homozygous UGT1A1*28 genotype; and p for trend <0.05). Patients with combined genotypes carrying UGT1A7 variant alleles and UGT1A1 variant alleles (including UGT1A1*28 and UGT1A1*6) are associated with increased risk of GS (OR = 13.96 for patients with combined genotype carrying at least one variant allele of UGT1A1 and UGT1A7). In conclusion, the -57UGT1A7 (T>G) is highly associated with UGT1A7*3 and moderately associated with 211UGT1A1 (G>A). Certain UGT1A1/UGT1A7 combined genotypes are risk factors of GS.