男性精浆中IL-6 和sICAM-1 与免疫性不育的关系研究

目的:探究男性精浆中白细胞介素-6(IL-6)和可溶性细胞黏附分子-1(sICAM-1)与免疫性不育的关系。方法:选择2014年6月至2015年12月我院收治不育症患者189例及100例健康体检者为研究对象,根据患者精子混合抗球蛋白反应实验(MAR)结果将不育症患者分为免疫性不育组(88例)和非免疫性不育组(101例);免疫不育组患者按照精液白细胞过氧化物酶染色情况分为免疫性阳性白细胞组(WBC≥1×10~6/mL)36例和免疫性阴性白细胞阴性组(1×10~6/m L)52例,分析并比较各组间精子质量、IL-6和sICAM-1水平。结果:免疫性不育组与非免疫性不育组患者精液精子向前运动比率、存活率均低于对照组(P0.05);免疫性不育组与非免疫性不育组精液各项参数均不存在差异(P0.05);免疫性不育组患者IL-6、sICAM-1水平均高于非免疫性不育组及对照组,差异有统计学意义(P0.05);非免疫性不育组IL-6、sICAM-1水平均高于对照组,差异有统计学意义(P0.05)。相关性分析显示,精子被Ig G黏附数比例与研究对象精液IL-6、sICAM-1水平呈正相关关系(r=0.438,0.561;P0.05);免疫性阳性白细胞组患者精液IL-6与sICAM-1水平均高于免疫性阴性白细胞组(P0.05)。结论:免疫性不育症患者精子质量下降,男性精液中IL-6和sICAM-1水平表达越高,MAR阳性率越高,男性免疫性不育发生的可能性越大。     

MHC DQA基因的分子进化研究——Ⅱ.基于核苷酸替换和SCU偏移的系统发育的分析

根据23个等位基因不同外显子(EN)的核苷酸(NT)替换及同义密码子使用谱(SCU)偏移研究了哺乳类7种动物MHC DQA座位的系谱发育关系。发现:(1)在大时间尺度内,MHC DQA基因进化速率(1.0×10~(-9),其中EN2为1.3×10~(-9)9NT/位点/年)与一般核基因相似;鼠类DQA基因的NT替换速率大致是其他哺乳类的2倍;(2)证实DQA等位基因多态性是在哺乳动物分化以后才逐渐形成的;推测牛类也存在与绵羊DQA2即OLA-DQA(c17-2)对等的具有最近共同祖先的DQA2基因座位有待发现;HLA-DQA2系谱与HLA-DQA1祖先的分化年代在20～12Mya(百万年前),HLA-DQA1各等位基因分化时间在24Mya至1Mya以内,因此产生HLA-DQA2座位的基因重组发生在HLA-DQA1产生少数几个等位基因之后;(3)基于SCU分化度的进化树从一个新的角度体现了DQA基因的系统发育关系,并显示HLA-DQA2具有特殊的SCU,提示在DQA基因的进化中产生了SCU的分化,SCU系统树在进化上有重要价值和特殊意义。改进了基因的SCU分化度和SCU相似系数的估算方法。     

用PCR-RFLP方法研究藏族HLA-DQA1和-DQB1基因多态性

应用目前HLA研究领域中成熟的,有效的PCR-RFLP基因分型技术,从DNA水平对藏族健康群体进行了HLA-DQA1(49人)和-DQB1(49人)基因分型,这在国内外属首次。所采用的PCR-RFLP基因分型技术是在HLA-DQA1和-DQB1各等位基因全部序列已知的情况下,对其第2个外显子碱基序列扩增进而进行RFLP分析的方法。这种方法得到的RFLP的所有片段都是已知序列,因而精确度很高,同时为发现新的等位基因提供了成熟而有效的分析方法。研究结果表明,在藏族DQA1的8个等位基因,DQA1*0301的基因频率最高(36.74％)。DQA1*0601(4.08％)、*0103(4.08％)和*0401(5.10％)最低。在DQB1的16个等位基因中,OQB1*0302(16.33％)、*0303(15.31％)和*0602(15.31％)为最常见,没有观察到*0504。统计分析表明,在DQA1各等位基因分布上,藏族与新疆汉族、北方汉族、上海汉族十分相近;与维吾尔族和哈萨克族也没有明显差异。在OQB1各等位基因的分布上,藏族与汉族、维族、哈族之间略有差异,而汉族、维族、哈族之间也存在一些差异。     

免不1号和2号治疗免疫性不育雄鼠的组织学和免疫组织化学研究

用中药复方免不1号,免不2号治疗免疫性不育雄鼠,观察睾丸,附睾组织学和免疫组化的变化。用精子抗原免疫昆明种雄性小白鼠,建立免疫性不育动物模型。同时分别饲喂中药复方免不1号,免不2号,醋酸强的松,生理盐水;从组织学和免疫组化等方面观察免疫性不育症的变化。结果显示免疫性不育雄鼠血清,精囊液抗精子抗体高,睾丸间质,睾丸曲细精管界膜,精原细胞,附睾管上皮细胞免疫复合物沉积多,睾丸每曲细精管精子和晚期精子细胞减少,中药免不1号和2号能降低抗精子抗体,清除免疫复合物的沉积,恢复曲细精管精子和晚期精子细胞数。结果表明：免不1号和2号通过调节全身免疫系统,清除循环和局部的抗精子抗体,免疫复合物,提高精子和精子细胞数,从而提高小鼠的受孕率。     

sp32／OY—TES-1在生精细胞中的分布及其抗体的血清学检测

了解sp32／OY—TES-1蛋白在生精细胞中的分布及抗sp32／OY-TES-1抗体在不育症患者血清中的出现情况。用重组表达的OY-TES-1麦芽糖融合蛋白免疫新西兰白兔,制备sp32／OY-TES-1多克隆抗体,该多克隆抗体经纯化鉴定后用于检测人精子和大鼠正常睾丸组织;用OY-TES-麦芽糖融合蛋白进行酶联免疫吸附（ELISA）试验,检测不育症患者和正常人血清中抗sp32／OY-TES-1抗体,     

抗中性粒细胞胞浆抗体、抗核抗体联合检测对类风湿关节炎 的诊断价值

目的:探讨抗中性粒细胞胞浆抗体(ANCA)与抗核抗体(ANA)联合检测对类风湿关节炎的临床意义。方法:采用IIF法对82例RA患者(RA组)、74例非RA自身免疫疾病患者(非RA组)和52例健康体检者(正常对照组)的血清ANCA和ANA谱进行了检测分析,并用ELISA法进行抗丝氨酸蛋白酶3(PR3)、抗髓过氧化物酶(MPO)、ANA谱的定量检测。结果:RA组82例患者中,64例ANCA阳性,阳性率为78.08%,其中核周型(PANCA)37例,阳性率为45.1%,胞浆型(CANCA)27例,阳性率为32.9%;非RA组74例患者中有7例ANCA阳性率分别为9.4%;正常对照组50例中没有一例ANCA阳性。利用Elisa法对患者血清进行检测,分别能够特异的检测到PR3、MPO、抗双链DNA抗体(抗ds-DNA抗体)、抗ss-A等抗体、抗SS-A抗体、抗PM-SCL抗体的存在。结论:联合ANCA、ANA检测有助于提高类风湿关节炎的诊断。     

ANCA与SLE临床表现的相关性及其意义

目的:探讨系统性红斑狼疮(Systemic Lupus Erythematosus,SLE)患者抗中性粒细胞胞浆抗体(Anti-neutrophil Cytoplasmic Antibodies,ANCA)与肾炎及其他临床表现和实验室检查的相关性及其意义.方法:采用前瞻性研究收集77例系统性红斑狼疮患者,用间接免疫荧光法(IIF)检测患者血清ANCA、ELISA法检测ANCA抗原,检测其他免疫学指标如抗核抗体、抗dsDNA抗体等.结果:77例SLE患者中ANCA阳性28(36.4%)例,ANCA阳性组浆膜炎、肾损害、神经精神症状、皮肤血管炎、抗dsDNA抗体阳性、抗Sm抗体阳性、补体下降以及血清IgG升高的发生率明显高于阴性组(P＜0.05).52例LN患者中,25例(48.1%)ANCA阳性,其中P-ANCA阳性者22例(88%).3例(12%)为a-ANCA均出现在RPGN,无一例出现c-ANCA.非LN组25例患者中,仅3例(12%)p-ANCA阳性,且均为抗-MPO.正常对照组无一例ANCA阳性.77例SLE患者中,14例(18.2%)为抗-MPO;13例(16.9%)为抗LF,且只见于DPGN、FPGN和RPGN伴有新月体形成者;10例(13%)为抗-CG,但在非狼疮肾炎患者未检测到抗-LF及抗-CG.在各种临床表现中,抗-MPO与肾脏和皮肤表现有关;而抗-LF与肾脏、关节炎及浆膜炎有关;抗-CG可见于各种临床表现.结论:ANCA可作为评价SLE疾病及鉴别血管炎和狼疮肾炎的一个重要指标.     

自身免疫性疾病患者抗2GP1抗体和抗t-PA抗体的相关性

目的:研究自身免疫性疾病患者抗2GP1抗体水平和抗t-PA抗体水平之间的关系。方法:用酶联免疫吸附法(ELISA)检测原发性抗磷脂综合症和红斑狼疮患者(32个狼疮样抗凝物阳性,32个狼疮样抗凝物阴性)与40例健康对照的IgG类抗2GP1和抗t-PA抗体的水平,用竞争ELISA的方法研究抗2GP1抗体与t-PA的互作。结果:64个病人中有8个IgG类抗2GP1和抗t-PA抗体共存,并且这两种抗体的共存和血栓病史显著相关(P=0.02)。我们还发现两个来自病人的抗2GP1抗体能和t-PA交叉互作。但是,在这些病人群体中未发现这两种抗体显著相关。结论:病人体内可能存在能和t-PA交叉互作的抗2GP1抗体。     

成年女性系统性红斑狼疮血清中抗黄体抗体与月经异常 的相关性研究

目的:探讨成年女性系统性红斑狼疮(SLE)血清中抗黄体抗体与月经异常的相关性。方法:收集SLE患者及正常对照者的临床资料,详细记录入选研究对象的月经及生育情况,同时记录抗核抗体(ANA)等自身抗体的测定结果。采用酶联免疫吸附试验(ELISA)法检测患者血清中抗黄体抗体。结果:在入选的69例成年女性SLE患者中,26(38%)例患者抗黄体抗体阳性。而在40例健康对照者中,仅2(5%)例出现抗黄体抗体阳性。在入选的69例SLE患者中,29(42%)例患者的月经正常,40(58%)例患者出现月经异常。在40例健康对照者中,31(78%)例患者的月经正常,仅9(21%)例患者出现月经异常。有月经异常的SLE患者中抗黄体抗体的阳性率显著高于无月经异常的SLE患者(66%vs 18%,P<0.01),而在健康对照者中则未见有显著性差异。结论:SLE患者中抗黄体抗体的阳性率为38%,并可能与SLE的月经异常相关。     

富亮氨酸胶质瘤失活1蛋白抗体脑炎的病理特征及临床分析（附1 例案例报道）

目的:探索富亮氨酸胶质瘤失活1蛋白(LGI1)抗体相关自身免疫性脑炎的临床特点及治疗。方法:报道l例LG I1抗体阳性相关自身免疫性脑炎的临床资料,并结合相关文献讨论该病的临床病理特点。结果:老年男性,亚急性起病,反复多次发作并进行性加重,以近记忆下降、癫痫、认知和睡眠障碍为主要表现;头颅MRI示脑萎缩;LGI1抗体阳性。结论:本病患者具有认知功能、睡眠障碍及癫痫等,血清和脑脊液中抗LGI1抗体阳性,但无低钠血症,头颅影像学检查正常;急性发作期给予免疫抑制剂治疗后可获良好效果。     

Specific amino acid residues in the second hypervariable region of HLA-DQA1 and DQB1 chain genes promote the Ro (SS-A)/La (SS-B) autoantibody responses

In order to define the HLA-DR and DQ alleles, as well as the specific DQA1 and DQB1 chain genes involved in the anti-Ro/La autoantibody responses, RFLP analysis and sequence-specific oligonucleotide typing was carried out on 58 Caucasians and 48 American blacks with SLE or Sj?gren's syndrome and anti-Ro antibodies. Among both Caucasian and black patients, the highest relative risk for the anti-Ro response (both with and without accompanying anti-La) was conferred by heterozygosity for the DQw2.1 (in linkage disequilibrium with HLA-DR3) and DQw6 (a subtype of DQw1) alleles compared with either 269 normal race-matched controls or 80 anti-Ro negative SLE/Sj?gren's syndrome patients. Analysis of individual DQA1 and DQB1 chain alleles revealed that DQA1*0501 and DQB1*0201 were most frequent, followed by DQA1 and DQB1 alleles comprising DQw6. In patients not possessing DQw2.1 and/or DQw6 alleles, HLA-DQB1*0302 and HLA-DQA1*0401 (especially in blacks) were significantly increased. Nucleotide sequence analysis of these associated alleles showed that 100% of patients with anti-Ro had a glutamine residue at position 34 of the outermost domain of the DQA1 chain and/or a leucine at position 26 of the outermost domain of the DQB1 chain. Patients with anti-Ro plus La were more likely to have all four of their DQA1/DQB1 chains containing these amino acid residues than either anti-Ro-negative SLE patients or controls. These data implicate specific amino acid residues on both DQA1 and DQB1 chains located in the floor of the Ag binding cleft of the HLA-DQA1:B1 heterodimer and further suggest a role for "gene dosage" in the anti-Ro (+/- La) autoantibody response.     

新疆吐鲁番地区维吾尔族HLA-DQA1等位基因多态性研究

目的：探讨新疆吐鲁番地区维吾尔族HLA—DQA1等位基因多态性。方法：采用聚合酶链反应一序列特异性引物技术,检测107例新疆吐鲁番地区维吾尔族健康个体的HLA—DQA1等位基因频率,分析HLA—DQA1基因多态性,并将所得结果与国内其他民族的同类资料进行比较。结果：新疆吐鲁番地区维吾尔族健康个体中共检出10个等位基因,以DQA1＊0104（O．3727）、DQA1＊0501（0．2636）、DQA1＊0103（0．1318）的频率较高,DQA1＊0401（0．0045）、DQA1＊0601（0．0091）的频率较低。结果与国内其他民族的同类资料进行比较,等位基因分布频率上有共同点,也有一定的独特性。结论：新疆吐鲁番地区维吾尔族HLA—DQA1基因,与国内其他民族群体的资料比较,总体检验有显著差异（p〈0．01）,显示人类的遗传系统既有共性亦有各自的特点,显示华人群体遗传背景的复杂性,结果为人类学研究和HLA—DQA1基因相关的疾病提供了较为重要的信息.     

HLA class II DR and DQ genotypes and haplotypes associated with rheumatic fever among a clinically homogeneous patient population of Latvian children

The HLA system is being paid more and more attention because it is very significant in polymorphous immunological reactions. Several studies have suggested that genetic susceptibility to rheumatic fever (RF) and rheumatic heart disease (RHD) is linked to HLA class II alleles. We hypothesized that HLA class II associations within RHD may be more consistent if analysed amongst patients with a relatively homogeneous clinical outcome. A total of 70 RF patients under the age of 18 years were surveyed and analysed in Latvia. HLA genotyping of DQA1, DQB1 and DRB1 was performed using PCR with amplification with sequence-specific primers. We also used results from a previous study of DQB1 and DRB1 genotyping. In the RF patients, HLA class II DQA1*0401 was found more frequently compared to DQA1*0102. In the RF homogeneous patient groups, DQA1*0402 has the highest odds ratio. This is also the case in the multivalvular lesion (MVL) group, together with DQA1*0501 and DQA1*0301. In the chorea minor patients, DQA1*0201 was often found. Significant HLA DQA1 protective genotypes were not detected, although DQA1 genotypes *0103/*0201 and *0301/*0501 were found significantly and frequently. In the distribution of HLA DRB1/DQA1 genotypes, *07/*0201 and *01/*0501 were frequently detected; these also occurred significantly often in the MVL group. The genotype *07/*0201 was frequently found in Sydenhamn's chorea patients that had also acquired RHD, but DRB1*04/DQA1*0401 was often apparent in RF patients without RHD. In the distribution of HLA DQA1/DQB1 genotypes, both in RF patients and in the homogeneous patient groups, the least frequent were *0102/*0602-8. The genotype DQA1*0501 with the DQB1 risk allele *0301 was often found in the MVL group. The genotype *0301/*0401-2 was frequently found in the RF and Sydenhamn's chorea patient groups. The haplotype *07-*0201-*0302 was frequently found in RF and homogeneous patient groups, including the MVL group. In addition, haplotypes *04-*0401-*0301 and *04-*0301-*0401-2 were frequent amongst patients with Sydenhamn's chorea. The protective alleles DQA1*0102 and DQB1*0602-8 in the haplotype DRB1*15 were less frequently found in RF patients. The results of the present study support our hypothesis and indicate that certain HLA class II haplotypes are associated with risk for or protection against RHD and that these associations are more evident in patients in clinically homogeneous groups.     

Genetic and immunologic aspects of type 1 diabetes mellitus

Prediction of type 1 diabetes mellitus (IDDM) and its identification in preclinical period is one of the central problems in modern medicine. They are based comprehensive genetic, immunologic and metabolic evaluations. We observed four hundred seven first-degree relatives of patients with IDDM (240 families in which one of the children or one of the parents had IDDM) have been included in the study. The study of HLA-DQA1, HLA-DQB1 polymorphic alleles and DRB1 genes and their combinations. The genetic study included searching HLA loci (HLA-DQA1, HLA-DQB1 polymorphic alleles and DRB1 genes) loci. To evaluate the genetic risk two approaches we used: first--carrying predisposing HLA-DQ alleles and DRB1-genes and it's combination (mainly associated in Russian population was DRB1*04-DQB1*0302, DRB1*04-DQA1*0301, DQA1*0301-DQB1*0302, DQA1*0301-DQB1*0302 and four susceptible alleles in A- and B- chains (Asp 57-, Arg 52+)) and second--IBD (identity by descent), in Russian population HLA-identical for 2 haplotypes sibs had risk of development of IDDM of 18%, for 1 haplotype--3%, for 0 haplotype-0.9%. The antibodies (ICA, IAA) prevalence rate has not depended on availability of predisposing HLA-DQ alleles and DRB1-genes and haploidentity of normal sibs and sibs with IDDM. However, GADA prevalence rate in groups having high predisposed alleles has been noticed as significantly higher (28.6%) comparing with 7.7% in groups that had no predisposing alleles (p < 0.05). The comparison of antibodies prevalence rate to sibs HLA-identity has shown the significant increase or GADA prevalence rate in group of siblings identical for one haplotype comparing with non-identical sibs (27.3% and 0% respectively, p < 0.001).     

新疆维吾尔族、哈萨克族人群HLA-DQA1、-DQB1两基因座多态性的研究及与25个种族或民族的比较分析

应用ＰＣＲ－ＲＦＬＰ基因分型技术,首次对我国新疆地区维吾尔族和哈萨克族２个少数民族群体的ＨＬＡ－ＤＱＡ１、－ＤＱＢ１两个基因座的多态性进行了研究。结果显示,在ＤＱＡ１８个等位基因中,维族和哈族均表现为ＤＱＡ１＊０３０１最常见。最少见的ＤＱＡ１等位基因,在维族中为ＤＱＡ１＊０４０１和＊０６０１,而在哈族中为ＤＱＡ１＊０６０１;在ＤＱＢ１１６个等位基因中,ＤＱＢ１＊０２０１和＊０３０１在维族和哈族中均表现为最常见。在维族中未观察到ＤＱＢ１＊０５０２、＊０５０３２和＊０５０４,在哈族中未观察到ＤＱＢ１＊０５０３２、＊０５０４和＊０６０５等位基因。统计分析表明,维族和哈族ＤＱＡ１、ＤＱＢ１各等位基因的分布无显著性差异,说明维、哈族间具有较密切的亲缘关系。在以２７个种族或民族的ＨＬＡ－ＤＱＡ１、－ＤＱＢ１两基因座基因频率构建的分子系统树上,维族和哈族独立于其他群体而聚类,独处一支。维族和哈族接近蒙古人种,而离高加索人种较远。     

HLA-DQA1*0301-associated susceptibility for autoimmune polyglandular syndrome type II and III.

No significant differences were reported for the frequency of DR3-DQ2 and DR4-DQ8 haplotypes in a recent study of one of the largest cohorts worldwide of patients with isolated Addison's disease compared to patients with APS II. However, previous studies had suggested that the HLA-DQ genes, especially DQA1*0102, may be a genetic marker for resistance to autoimmune thyroid disease, which is the most frequent disease in APS II or III. Until now, HLA-DQA1 alleles have not been systematically investigated in APS. We determined the HLA-DR and HLA-DQA1 association in 112 unrelated patients with APS II (n = 29), APS III (n = 83) and 184 unrelated patients with single-component diseases without further manifestations of APS: Graves' disease (n = 70), Hashimoto's thyroiditis (n = 53), autoimmune Addison's disease (n = 15), vitiligo (n = 16) and alopecia (n = 30), and 72 healthy controls - German Caucasians - to identify possible predisposing and protective HLA class II alleles in APS. In agreement with previous studies, we detected a significantly higher frequency of DR 3 and/or DR 4 in patients with APS II and III compared to controls. In patients with APS II, we detected a significantly higher frequency of DQA1*0301 and *0501 compared to controls confirming the increased frequency of an extended HLA DRB1-*04-DQA1-*03-DQB-*03 haplotype as previously described. In contrast, only DQA1*0301 was increased in our patients with APS III compared to controls. Moreover, we detected an increased frequency of DQA1*0301 in patients with APS, whereas DQA1*0301 was only significantly elevated in alopecia in patients with single-component diseases without APS. Therefore, our results indicate an association between DQA1*0301 and APS II or III since this allele was otherwise not significantly associated with any of its component diseases except alopecia. Moreover, our data imply that the allele DQA1*0301 is a marker of increased risk for further APS manifestations in patients who suffer from an organ-specific autoimmune disease.     

Analysis of the association of HLA-DQ1 alleles with mutation of the 21-hydroxylase gene in patients with congenital adrenal hyperplasia

In 96 patients with congenital adrenal hyperplasia (CAH) and 50 healthy donors from northwestern Russia the distribution of the HLA-DQA1 alleles and the mutation spectrum and frequency at the CYP21B gene were examined. In the patients with nonclassical (NC) CAH, the distribution of the HLA-DQA1 polymorphic alleles was similar to that in the population sample. In the patients with the salt-wasting form of the disease a statistically significant decrease of the *0401 or *501 major allele frequency was observed. The prevalence of certain HLA-DQA1 genotypes, namely, HLA5, HLA3, and HLA4, was observed in the patients with the NC, salt-wasting (SW), and simple virilizing CAH, respectively. Each clinical group was characterized by a specific spectrum of clinically valuable mutations. An association between the CYP21B mutations most frequently found in case of SW and SV CAH (delB, I2splice, and I172N) and certain HLA-DQA1 alleles was demonstrated. The necessity of more precise clinical diagnostics of the NC CAH cases along with detailed examination of this group for determination of the major mutations typical of the NC CAH cases from northwestern Russia is discussed.     

HLA—DQ分子遗传结构与中国人重症肌无力的相关性

重症肌无力与ＨＬＡⅡ类基因关联性在不同人种和民族中具有不同遗传易感性,为探讨中国人重症肌无力（ＭＧ）与ＨＬＡ０ＤＱ分子关联性,采用聚合酶链式反应－限制性片段长度多态性（ＰＣＲ－ＲＦＬＰ）方法,分析了５０例中国正常人及４９例重症肌无力患者的ＨＬＡ－ＤＱＡ１和－ＤＱＢ１座位的基因型,结果：共检出正常人ＤＱＡ１等位基因８种,ＤＱＢ１等位基因１０种,重症肌无力患者ＤＱＡ１等位基因８种,ＤＱＢ１等位基因９种