Kaposi肉瘤组织内人疱疹病毒8型的K14.1基因型研究

目的:确定人疱疹病毒8 型K14.1基因等位基因型在Kaposi肉瘤中的分布,及与Kaposi肉瘤临床分型的关系.方法:使用酚-氯仿-异戊醇法对32例Kaposi肉瘤石蜡包埋组织进行病毒DNA提取,使用PCR法扩增K14.1基因片段,然后确定其等位基因型.结果:32 例中有27 例人疱疹病毒8型感染为阳性,阳性率为84.38%,其中5例艾滋病相关型KS患者HHV-8感染均为阳性,感染率100% ;在分析的27 例HHV-8病毒株中,24例为P等位基因型(包括5例艾滋病相关型KS患者皮损),3例为M等位基因型(均来自经典型KS).结论:本研究中,Kaposi肉瘤感染的人疱疹病毒8型的K14.1等位基因型以P等位基因型为主.     

新疆Kaposi肉瘤组织内人类8型疱疹病毒DNA的检测

目的 了解Kaposi肉瘤组织内人类8型疱疹病毒(HHV-8)在Kaposi肉瘤发病中的作用。方法 应用荧光原位聚合酶链反应(PCR)方法对20例新疆Kaposi肉瘤组织进行HHV-8DNA检测,Kaposi肉瘤组织损害包括结节12份、斑块6份、斑片2份。同时和20份非Kaposi肉瘤组织,包括纤维瘤组织18份,血管瘤组织2份进行对照。结果 20例新疆Kaposi肉瘤组织中有17例检测到HHV-8DNA(阳性率为85%),而20例非Kaposi肉瘤组织HHV-8DNA均为阴性。两组阳性率经校正χ²检验,χ²=26.2,P<0.001.实验中HHV-8DNA片段主要集中于Kaposi肉瘤组织的梭形瘤细胞和血管内皮细胞。除此之外,在Kaposi肉瘤表皮角质形成细胞内也检测到HHV-8DNA片段,但DNA片段的荧光强度明显弱于梭形瘤细胞和血管内皮细胞中的荧光强度。结论 在新疆Kaposi肉瘤的发生中,HHV-8起着一定的作用,可能参与了Kaposi肉瘤的发病,但不是Kaposi肉瘤发生的唯一病因。     

福建地区经典型Kaposi肉瘤感染病毒的亚型研究

目的:研究福建地区经典型Kaposi肉瘤感染的病毒亚型。方法:收集经典型Kaposi肉瘤患者临床资料及石蜡包埋组织标本24例,利用QIAamp~?DNA FFPE Tissue试剂盒提取患者皮损蜡块组织中的DNA,设计扩增人类疱疹病毒(HHV)-8(ORF-K1、T0.7/K12、ORF-K14.1/K15、ORF26及ORF75)基因的引物,巢式PCR扩增提取的DNA,然后进行双向测序,再采用SnapGene软件对测序结果进行确认,从而确定HHV-8各病毒亚型的分布情况。结果:24例福建地区经典型Kaposi肉瘤均来源于汉族人,皮损好发于老年人双下肢,以结节为主,患者感染HHV-8不同基因扩增阳性率:ORF26 100.00%、ORF-K133.33%、T0.7/K12 29.17%、ORF-K14.1/K15 33.33%、ORF75 25.00%。序列比对分析显示24例经典型Kaposi肉瘤患者感染HHV-8 ORF26的病毒亚型均为C亚型。结论:福建地区经典型Kaposi肉瘤感染HHV-8的基因型主要为ORF26,且均为C亚型。     

PCR检测Kaposi肉瘤患者血清人8型疱疹病毒

1994年Chang等从艾滋病患者的Kaposi肉瘤组织中分离到了两种独有的序列^[1],称为人8型疱疹病毒(HHV-8),目前认为可能是Kaposi肉瘤的病因.国内已有作者对新疆经典型Kaposi肉瘤病理组织内进行过HHV-8的PCR及原位PCR研究^[1-3],现对10例新疆经典型Kaposi肉瘤患者血清进行了HHV-8DNAPCR检测.     

人类疱疹病毒8型ORF26基因亚型与Kaposi肉瘤的相关性研究

目的 明确Kaposi肉瘤患者感染的人类疱疹病毒8型（HHV-8）ORF26基因亚型分类，初步探讨其与Kaposi肉瘤不同临床分型及侵袭性的相关性。方法 对32例Kaposi肉瘤石蜡包埋组织进行HHV-8 DNA抽提、扩增、双向测序，使用DNAStar软件、Clustal W软件和PHYLIP软件包对测序结果进行系统发生学分析，从而确定HHV-8 ORF26基因亚型，最后运用Fisher确切概率法对结果进行统计学分析。结果 32例Kaposi肉瘤中有30例HHV-8阳性，阳性率为93.75%，其中6例艾滋病相关型患者HHV-8均阳性。30例HHV-8阳性患者中，17例为HHV-8 ORF26 A亚型，13例为C亚型。不同亚型间Kaposi肉瘤患者有无黏膜损害及临床分型的分布差异均无统计学意义（P > 0.05）。结论 Kaposi肉瘤患者感染HHV-8 ORF26亚型属于A亚型和C亚型，不同亚型与黏膜损害及临床分型无关。     

新疆Kaposi肉瘤患者感染的人类疱疹病毒8型ORF75基因多态性研究

目的通过对人类疱疹病毒8型(HHV-8)ORF75基因多态性研究,初步探讨其与新疆Kaposi肉瘤的相关性。方法对25例新疆Kaposi肉瘤患者石蜡包埋组织进行HHV-8 DNA抽提、扩增及双向测序,使用CLUSTALW软件和PHYLIP软件包对ORF75基因进行核苷酸多态性分析。结果①25例新疆Kaposi肉瘤患者有21例HHV-8感染阳性,阳性率为84%,其中7例艾滋病相关型Kaposi肉瘤(AIDS-KS)患者HHV-8感染均为阳性。②新疆Kaposi肉瘤患者HHV-8 ORF75基因核苷酸具有多态性,存在462、618、647三个主要突变位点。结论人类疱疹病毒8型ORF75基因多态性与新疆Kaposi肉瘤具有相关性。     

人疱疹病毒8型ORF75基因亚型与Kaposi肉瘤的相关性研究

目的 探讨人类疱疹病毒8型(HHV-8)ORF75基因亚型,与Kaposi肉瘤不同临床分型及侵袭性的相关性.方法 对25例新疆Kaposi肉瘤石蜡包埋组织进行HHV-8 DNA抽提、扩增及双向测序,使用Clustal W软件和PHYLIP软件包对测序结果进行发生学分析,从而确定HHV-8 ORF75基因哑型.结果 25例Kaposi肉瘤中,21例HHV-8阳性,阳性率为84%,其中7例AIDS相关型Kaposi肉瘤患者HHV-8均阳性.21例HHV-8阳性患者中,18例为HHV-8 ORF75 A亚型,3例为C亚型;不同亚型间Kaposi肉瘤患者有无黏膜损害及临床分型的分布差异均无统计学意义(P＞0.05).结论 新疆Kaposi肉瘤患者感染HHV-8 ORF75亚型属于A亚型和C亚型,HHV-8 ORF75不同亚型可能与新疆Kaposi肉瘤黏膜损害及临床分型无关.     

人类疱疹病毒8型基因型的研究进展

人类疱疹病毒8型广泛存在于各种临床表型的Kaposi肉瘤中,其基因型分布呈现种族与地域特异性.了解Kaposi肉瘤患者中HHV-8基因型特征及分布情况,探讨其与Kaposi肉瘤临床可能的相关性和演变及传播等具有重要意义.并对HHV-8病毒体及其基因组特征,K1和K15基因位点的生物学功能.HHV-8基因型演变呈现地域与种族特异性进行概述.     

人疱疹病毒8型ORF26单核苷酸多态性研究

目的 探讨人疱疹病毒8型(HHV-8)ORF26的单核苷酸多态性,分析其与Kaposi肉瘤不同临床分型及黏膜侵袭性的相关性.方法 Kaposi肉瘤患者32例,其中经典型26例,艾滋病相关型6例.使用酚-氯仿-异戊醇方法对Kaposi肉瘤石蜡包埋组织进行HHV-8 DNA抽提,采用巢式PCR方法扩增ORF26基因并双向测序,使用DNAStar软件和Clustal W软件分析ORF26基因的单核苷酸多态性.运用Fisher确切概率法对结果进行统计学分析.结果 ORF26基因研究发现,32例Kaposi肉瘤患者中HHV-8阳性30例,6例艾滋病相关型HHV-8均为阳性.30例患者的病毒株中,HHV-8 ORF26基因SNP主要集中在981T/C( 12例)、1086C/T(12例)、1139A/C(12例);HHV-8 ORF26基因单核苷酸多态性在不同临床分型或有无黏膜损害的Kaposi肉瘤之间的差异无统计学意义.结论 HHV-8 ORF26基因单核苷酸多态性可能与Kaposi肉瘤不同临床分型和黏膜侵袭性无关.     

鳞状细胞癌组织中HHV-8 K15等位基因型研究

目的 探讨皮肤鳞状细胞癌和食道鳞状细胞癌患者癌组织中人类疱疹病毒8型（HHV-8）K15等位基因型与肿瘤的发病关系。方法 序列特异性引物-巢式PCR技术检测40例皮肤鳞状细胞癌、40例食道鳞状细胞癌石蜡包埋组织中HHV-8 K15基因,并确定HHV-8 K15等位基因型,运用χ2检验对结果进行统计学分析。结果 9例皮肤鳞状细胞癌（22.5%）石蜡包埋组织中检测到HHV- 8 K15P型,8例食道鳞状细胞癌（20%）石蜡包埋组织中检测到 HHV- 8K15P型,两种肿瘤均未检测出K15M基因型。HHV-8在皮肤鳞状细胞癌与食道鳞状细胞癌感染率组间比较差异无统计学意义（P > 0.05）。结论 鳞状细胞癌中感染HHV-8K15P基因为K15P等位基因型,其发生发展可能与HHV-8的感染有一定的关系。     

Kaposi肉瘤组织人类疱疹病毒8型免疫组化检测

目的．探讨免疫组化检测Kaposi肉瘤（KS）组织人类疱疹病毒8型（HHV-8）的可行性及其诊断意义。方法采用免疫组化方法分别检测58例KS组织和40例化脓性肉芽肿组织中HHV-8的表达。结果①HHV-8在KS真皮瘤体中的表达阳性率为94．83％（55／58）,在化脓性肉芽肿表达均阴性,在KS组织和血管瘤组织瘤体中的表达差异有统计学意义（P〈0．05）。②HHV-8在KS组织表皮中表达阳性率为6．90％（4／58）,血管内皮细胞中的表达阳性率为91．38％（53／58）,真皮梭形细胞中表达阳性率为94．83％（55／58）。⑧结节期KS组织中表达HHV-8阳性的细胞所占比例较斑片期及斑块期明显增多,其差异有统计学意义（P〈0．05）。结论免疫组化方法检测HHV-8在KS的诊断中具有重要意义。     

新疆经典型Kaposi肉瘤29例血清HHV-8 DNA的套式PCR检测

目的了解新疆经典型Kaposi肉瘤(Kaposi’ssarcoma,KS)患者血清中人类8型疱疹病毒(humanherpesivirus8,HHV8)感染的情况,以探讨HHV8感染与经典型KS发病之间的关系。方法采用套式PCR技术检测29例新疆经典型KS及68例正常对照的血清中HHV8感染的情况。结果25例KS(86.2%)检测到HHV8的DNA片断;正常对照中14例(20.6%)检测到HHV8的DNA序列,差异有显著性(χ2=36.412,P<0.001)。结论HHV8在新疆经典型KS的发病中发挥着非常重要的作用,但可能不是发病的唯一原因。     

新疆Kaposi肉瘤组织内EBV,HHV—8双重感染的调查

应用ＰＣＲ方法，地２０例新疆Ｋａｐｏｓｉ肉瘤病理组织进行了ＥＢＶ和ＨＨＶ－８双重杂的调查，结果：２０例Ｋａｐｏｓｉ肉瘤病理组织中１４例检出ＨＨＶ－８ＤＮＡ（７０％），ＥＢＶ均为阴性。正常皮肤对照；１０例这两种疱疹类病毒均为阴性，作者认为新疆Ｋａｐｏｓｉ肉瘤的发生与ＥＢＶ的相关性很小，但明显与ＨＨＶ－８感染有关，但是否ＨＨＶ－８感染就是新疆Ｋａｐｏｓｉ肉瘤发生的决定因素，仍需进一步研究。     

HHV-8/KSHV during the development of Kaposi's sarcoma: evaluation by polymerase chain reaction and immunohistochemistry

The human gamma-herpes virus-8 (HHV-8) was first described in AIDS-related Kaposi's sarcoma (KS) tumour samples. In this study, we report comparative studies on paraffin-embedded biopsies of AIDS-related KS (AKS) and endemic KS (EKS) with regard to HHV-8 content as evaluated using polymerase chain reaction (PCR) and immunohistochemistry. DNA was extracted either using Chelex-100 or using Qia-gene kit and was evaluated with the help of a semiquantitative PCR assay. The PCR detection of HHV-8 was more sensitive to the Chelex method than to Qia-gene. The threshold for PCR test sensitivity with the help of serial dilution of DNA was at the level of five plasmid ORF-26 regions, and DNA from 25 body cavity-based lymphoma-1 cells. The results expressed as virus load/actin unit showed progressively higher HHV-8 levels in late (nodular) cases, compared to those in early (patch/plaque) stages. Evaluation of HHV-8 DNA levels in tumour tissues, thus, indicates a correlation between virus load and KS stage. Double immunostaining of spindle cells (SC) in KS biopsies for CD34 and HHV-8/latency-associated nuclear antigen (LANA) showed an increase in double-positive SC in the lesions of nodular AKS and EKS cases, compared to that in plaque and patch stages. However, 10-15% of CD34+/LANA- SC cells were observed during the development from patch to nodular cases of AKS and EKS. Our results indicate that PCR analysis is a simple and sensitive diagnostic method for HHV-8 evaluation in KS tissues, processed for conventional histopathology.     

Study of HHV-8 DNA sequences in archival biopsies from lesional skin of Kaposi's sarcoma,various mesenchymal tumors and related reactive conditions

Background: HHV-8 has been identified as the causative agent of Kaposi's sarcoma (KS) and some lymphoproliferative disorders. In addition, there are anecdotal reports on the presence of HHV-8 in other tumors, especially cutaneous epithelial and mesenchymal neoplasms. The aim of the study was to ascertain the value of identification of HHV-8 viral DNA sequences in routinely processed, formalin-fixed, paraffin-embedded tissues for the diagnosis of Kaposi's sarcoma and other mesenchymal tumors.
Methods: The presence of HHV-8 sequences in archival material was studied by nested PCR using specific primers for amplification of a 233-bp long fragment of HHV-8 (ORF 26).
Results: Thirty-three patients with KS (18 classic/sporadic, six post-transplant and nine AIDS-related) and various mesenchymal tumors and related conditions (n = 76) were studied. HHV-8 DNA sequences were detected in 29 of the 33 cases of KS and in one case of multiple eruptive dermatofibroma (MEDF).
Conclusions: Identification of HHV-8 DNA sequences in routinely processed tissue is a useful diagnostic marker for KS. Although other mesenchymal tumors are usually not associated with HHV-8, its presence is not fully specific for KS since HHV-8 sequences were also found in one case of MEDF. Therefore, PCR analysis for the detection of HHV-8 should only be used as an additional diagnostic marker for KS and in the context of other tools such as routine histology.     

Infection of circulating CD34+ cells by HHV-8 in patients with Kaposi's sarcoma.

Human herpesvirus type 8 (HHV-8) has been identified as the most likely candidate to be involved in the development of Kaposi's Sarcoma (KS). HHV-8 has been associated with all forms of KS, primary effusion lymphoma, and multicentric Castleman's disease and detected in various non-neoplastic cells. Its presence in cells of the different hemopoietic lineages has not yet been investigated in a comprehensive and systematic manner. In this study we searched for the presence of HHV-8 in different subpopulations of peripheral blood mononuclear cells (PBMC) from patients with classic and AIDS-associated KS, as well as from HIV-1 sero-positive and sero-negative persons without KS. Thirty-four samples of PBMC were isolated from 30 patients. Subpopulations were isolated with immunomagnetic beads. Polymerase chain reaction for HHV-8 DNA was performed on PBMC and subpopulations with a primer pair selected from ORF26 of the viral genome. Polymerase chain reaction products were subsequently Southern blotted and hybridized. In patients with KS, HHV-8 DNA was detected in nine of 11 (81%) CD19+ cells, four of 11 (36%) CD2+ cells, three of 11 (27%) CD14+ cells, and nine of 11 (81%) of the remaining depleted cell populations (DP) that contain CD34 positive cells. In a subsequent set of experiments HHV-8 DNA was detected in 10 of 12 (83%) CD34 positive cell fractions. All cell subpopulations from the non-KS group were HHV-8 negative, with the exception of one positive B cell sample obtained from an HIV-infected patient. Our data demonstrate that in peripheral blood HHV-8 is detectable not only in CD19+ cells, as previously reported, but also in other cells, including T cells, monocytes, and cells devoid of specific lineage markers. We also show for the first time that CD34+ cells in peripheral blood of KS patients are a predominant HHV-8-harboring population, suggesting that they represent an additional important reservoir for this virus in vivo.     

Human herpesvirus 8 (HHV-8) in familial Kaposi's sarcoma

Human herpesvirus 8 (HHV-8) has been detected in various epidemiological forms of Kaposi's sarcoma (KS). Since familial KS cases are exceedingly rare and the occurrence of familial KS in siblings has thought to depend rather on genetic factors than on a viral factor, familial KS has not been investigated for the presence of HHV-8. To investigate whether HHV-8 is present also in this rare form of KS, we examined tumor biopsies of 2 siblings with familial KS for the presence of HHV-8 specific DNA sequences by a nested PCR protocol. HHV-8 DNA sequences could be detected in KS specimens of both patients. Sequence analysis revealed an identical DNA sequence of HHV-8 in KS tissue of both siblings, but the sequence in our cases differs in one base pair at position 67 from the previously published HHV-8 KS330Bam fragment. The findings indicate that besides the yet poorly defined genetical factors involved in the pathogenesis of KS, HHV-8 may act as a cofactor also in familial KS. In addition, our data demonstrate that HHV-8 is found in all epidemiological forms of KS, including the rarely occurring familial KS. Familial KS may act as a further model to study the interaction of an oncogenic virus with genetic host factors in the context of a neoplastic disorder.