RESUMEN
Chronic lymphocytic leukemia (CLL) is a malignant disorder of B cell origin, with low incidence in Asian populations. In this study we investigated the HLA-class I A and B allele frequencies in 87 Iranian CLL patients and 64 healthy controls using sequence specific primer-polymerase chain reaction (SSP-PCR) technique. Our results showed increased frequencies of HLA-A11:01 (p=0.02) and HLA-B35:01 (p=0.002) alleles and HLA-A11:01/B35:01 haplotype (p=0.036) and decreased frequencies of HLA-A01:01 (p=0.02), HLA-A26:01 (p=0.03), HLA-B65:01 (p=0.03) and HLA-B53:01 (p<0.00001) alleles in CLL patients compared to the control group. Classification of the patients into non-progressive and progressive groups did not reveal significant differences for the frequency of any of the HLA-A and -B alleles or haplotypes between these two subtypes. Comparison between patients with immunoglobulin heavy chain variable region genes (IGHV) mutated (n=56) and unmutated (n=31) subtypes showed a significant increase in HLA-A32:01 (p=0.05) and HLA-A33:01 (p=0.05) alleles in IGHV unmutated patients compared to IGHV mutated patients. Similarly, a higher frequency of HLA-B52:01 (p=0.037) alleles was observed in CD38(+) compared with CD38(-) patients. Our results obtained from an Iranian population indicate that CLL is associated with distinct HLA class I alleles and haplotypes some of which are linked to disease prognostic factors.
Asunto(s)
Linfocitos B/inmunología , Etnicidad , Antígenos HLA-A/genética , Antígenos HLA-B/genética , Leucemia Linfocítica Crónica de Células B/inmunología , ADP-Ribosil Ciclasa 1/metabolismo , Adulto , Progresión de la Enfermedad , Femenino , Frecuencia de los Genes , Genotipo , Haplotipos , Prueba de Histocompatibilidad , Humanos , Irán , Leucemia Linfocítica Crónica de Células B/genética , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
B-cell chronic lymphocytic leukemia (B-CLL) results from clonal expansion of phenotypically mature but functionally immature B-lymphocytes. The incidence of this type of leukemia is low in Asian countries, whereas it is the most frequent type of leukemia in the West. Previous investigations mainly conducted in Western populations have demonstrated non-random rearrangement of certain immunoglobulin variable region heavy (VH) and/or light (VL) chain genes in different groups of B-CLL patients. Little is known about the profile of VH gene expression in Asian patients. In the present study, we determined the frequency of VH gene family usage in 59 Iranian patients with B-CLL. VH gene family of patients was determined by reverse transcriptase-polymerase chain reaction using VH1-VH7 family specific primers. The most frequently expressed VH gene family was found to be VH3 (45.8%) followed by VH4 (32.2%), VH1 (18.6%), VH5 (1.7%) and VH6 (1.7%), with no expression of VH2 and VH7 gene families. The results indicate a lower representation of the VH1 and VH2 gene families and a higher representation of the VH4 gene family in Iranian B-CLL patients compared to Western patients, suggesting involvement of ethnic and/or environmental factors in B-CLL disease initiation.
Asunto(s)
Cadenas Pesadas de Inmunoglobulina/metabolismo , Región Variable de Inmunoglobulina/metabolismo , Leucemia Linfocítica Crónica de Células B/metabolismo , Adulto , Anciano , Progresión de la Enfermedad , Femenino , Citometría de Flujo , Expresión Génica , Reordenamiento Génico de Cadena Pesada de Linfocito B , Humanos , Inmunofenotipificación , Irán , Leucemia Linfocítica Crónica de Células B/inmunología , Masculino , Persona de Mediana EdadRESUMEN
Hemophilia A patients treated with human coagulating factor VIII (FVIII) may develop inhibitory antibodies (inhibitors). Characterization of the inhibitors at the clonal level may help exploring new therapeutic strategies. We have generated lymphoblastoid cell lines (LCLs) producing anti-FVIII antibodies from peripheral blood lymphocytes of hemophilia A patients with high inhibitor titers. We fused the anti-FVIII-positive LCLs with a heteromyeloma, to produce FVIII specific hybridomas. We determined the specificity, isotype, idiotypic and immunoglobulin (Ig) variable region heavy (VH) chain gene family profiles of the secreted antibodies (Ab) by ELISA, immunoblotting and RT-PCR. We established eight hybridomas which produced high titers of anti-FVIII Ab. All hybridomas secreted IgM Ab, associated with either kappa(5/8) or lambda(3/8) light chain. Analysis of the expressed VH genes by RT-PCR revealed that the hybridomas utilized only the VH1 (63%) or the VH3 (37%) gene families. Among the cross-reactive idiotypes (CRIs) we tested, only the VH1 and VK3b-associated CRIs were expressed by 3 hybridomas. Immunoblotting of thrombin-digested FVIII demonstrated distinct patterns of reactivity of the monoclonal Ab (MAb) secreted by the hybridomas, which recognized either the A2 domain of the Fvm heavy chain, or the light chain, or both. Our findings suggest that: a) the isotype of the anti-FVIII Ab secreted by LCLs and hybridoma clones (IgM) differs from that of anti-FVIII Ab in vivo, which are predominantly IgG4: this suggests a negative selection of the isotype-switched FVIII-specific B-cells in the periphery of these patients; b) the anti-FVIII Ab have a biased representation of the VH1 gene family, and c) somatic mutations in the VH genes coding for FVIII specificity occur in the anti-FVIII Ab response, as evidenced by lack of expression of the VH-associated CRI.
Asunto(s)
Factor VIII/inmunología , Hemofilia A/inmunología , Hibridomas/inmunología , Autoanticuerpos/sangre , Secuencia de Bases , Western Blotting , Reacciones Cruzadas , Cartilla de ADN , Ensayo de Inmunoadsorción Enzimática , Humanos , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The frequency of expression of immunoglobulin (Ig) variable region heavy (VH ) chain gene products was studied in 43 Iranian patients with mutiple myeloma (MM). The expressed VH gene families and associated cross-reactive idiotypes (CRI) were analysed by immunoblotting and ELISA, using peptide-induced polyclonal antibodies specific for VH 1-VH 6 gene families and monoclonal antibodies (MAb) recognising CRI linked to theVH 1, VH 3, VH 4 and VH 6 gene families. The results revealed that the VH 3 family (60. 5%) was the most predominant gene family. In contrast, no paraproteins were encoded by genes from the VH 2 gene family and only 2.3% were encoded by the VH 5 family. The panel of paraproteins tested rarely expressed the probed VH -associated CRI. Our results suggest that: 1-The Ig VH genes, may not be randomly expressed in the malignant plasma cells from Iranian patients with MM. 2- Some of the genes seem to be negatively selected or highly mutated, as evidenced by the lack of expression of the probed CRI.