Distribution of killer cell immunoglobulin-like receptor genes in the mestizo population from Venezuela.

This study represents the first report on the distribution of KIR genes in 205 unrelated healthy mestizo Venezuelan individuals. Genotyping analysis showed that all KIR genes are present in this population. Frequency of inhibitory killer cell immunoglobulin-like receptors (KIRs) exceeded 0.69, except for KIR2DL2 (0.29) and 2DL5 (0.37). Activating KIRs showed low frequencies (0.11-0.29), except for KIR2DS4 (0.68). Forty-five different KIR genotypes were identified, with a predominance of three genotypes found in 50.7% of the population of which 25.9% were individuals homozygous for haplotype A. The frequencies of KIR genes reflect the ethnic admixture existing in the mestizo Venezuelan population.

Genetic variability of Apolipoprotein E in different populations from Venezuela.

The genetic variation at the Apolipoprotein E locus (APOE) is an important determinant of plasma lipids and has been implicated in various human pathological conditions. The objective of the present study was to estimate the distribution of APOE alleles in five Venezuelan communities: two Amerindian tribes (Bari and Yucpa), one Negroid population from Curiepe, one Caucasoid population from Colonia Tovar and the Mestizo urban population living in Caracas. The APOE*3 allele was the most common allele in all populations studied. However, a significant increase in the APOE*2 allele frequency in the Mestizo (18.96%) and Negroid (16.25%) populations was found. Similar to results reported in other Native American populations we have found that the APOE*2 allele is completely absent in the Bari and Yucpa Amerindians. Frequencies found in the Colonia Tovar population are in agreement with those reported in the population of Germany, indicating a high degree of relatedness. The results support the notion that the distribution of the APOE alleles shows ethnic variability.

Is the CCR5-59029-G/G genotype a protective factor for cardiomyopathy in Chagas disease?

Investigated were two CCR5 gene polymorphisms, the CCR5 Delta 32 deletion and the pCCR5 59029 A-->G promoter point mutation, in 107 ethnically mixed Venezuelan patients serologically positive for Trypanosoma cruzi (34 asymptomatic, 38 arrhythmic, 35 cardiomyopathic). No difference in the distribution of CCR5 Delta 32 among asymptomatic and symptomatic patients was found. We have observed an increase of the 59029-G phenotype among asymptomatic compared with symptomatic chagasic patients (68% vs. 58%), in agreement with previously reported data (57% vs. 31%). This frequency difference, although not statistically significant, is more marked when the 59029-G allele is present in homozygous form. However, a similar distribution of the G/G genotype is present among asymptomatic patients and patients with heart failure. Because it has been reported that the 59029G/G genotype associates with lower CCR5 expression, 37% of our T. cruzi-infected patients with heart failure are genetically predisposed to express low levels of CCR5 on the surface of CD8(+) T cells, contrary to what would be expected if an inflammatory response is required for severe cardiac damage. If confirmed, the possible protection that might be conferred by the G/G genotype may be due to the existence of other genes in linkage disequilibria.

HLA class I and class II allele and haplotype distribution in the Venezuelan population.

Population studies represent an integral part, and a necessary link in a complex chain of host-pathogen interactions, disease pathogenesis, and major histocompatibility complex polymorphism. HLA class I and class II allele and haplotype distributions among Venezuelan mestizos were determined. Genes of Mongoloid, Negroid, and Caucasoid origin have created a distinctive human leukocyte antigen (HLA) genetic profile in this hybrid mestizo population that will influence HLA and disease association studies. The predominant HLA-B DQA1 DQB1 DRB1 haplotype is HLA-B44 DQA1*0201 DQB1*0201 DRB1*0701 (5.3%). It is noteworthy that the HLA-A3 B7 DR2 and the HLA-A1 B8 DR3 linkage groups, which are part of conserved or ancestral haplotypes, the last one associated with a wide range of autoimmune diseases and immune abnormalities in apparently healthy subjects, show low incidence among Venezuelan mestizos. This fact may be useful for future HLA and disease association studies and for localization of genes involved in immune regulation associated with these haplotypes.

Strong association between major histocompatibility complex class I antigens and immune aberrations among healthy Venezuelans.

Immune reactivity indicators studied among 55 unrelated Venezuelan mestizo subjects included lymphoproliferative response to polyclonal mitogen (PHA, Con A, PwM) stimulation, NK cell activity, and enumeration of peripheral blood mononuclear cells. HLA-A, -B, -C, -DR, and -DQ antigens were determined by serologic typing. A strong association between impairment of the parameters studied and MHC class I antigens A11 and A3 was found. Subjects with decreased suboptimal (0.5 micrograms/ml) PHA response as well as decreased optimal (0.5 micrograms/ml) Con A response showed high incidence of HLA-A11 antigen (RR = 81, p = 0.001, pc = 0.021 and RR = 54, p = 0.0029, respectively). Subjects with decreased suboptimal (0.5 micrograms/ml) PHA response HLA-A11- with only one exception, were either HLA-A1+ or HLA-A3+. These antigens belong to the same CREG, share public epitopes, and have low incidence in the Venezuelan mestizo population. Six of 10 persons with decreased CD16 subset count (5.17% +/- 0.23% vs 11.69% +/- 0.44%) had HLA-A3 antigen (RR = 17, p = 0.001, pc = 0.021). The data indicate a possible contribution of HLA-A11,A3, molecules through their private and/or public determinants to immune response aberrations which under certain conditions may result in development of diseases.

Unique HLA-DR/DQ associations revealed by family studies in Warao Amerindians. Haplotype and homozygosity frequencies.

HLA-A, Cw, B and A, Cw, B, DR genotypes have been assigned, respectively, to 318 and 175 Warao Amerindians belonging to 73 sibships, who were tested with International Histocompatibility Workshop reagents. Only 33% of the theoretically possible three-loci and 7% of the possible four-loci haplotypes were found, with 10 and 6 of them accounting, respectively, for 75% of the total observed. This limited haplotype variability, expected in an inbred population, was not accompanied by either an increased or a decreased frequency of homozygous individuals, as demonstrated by population and family analysis. Inheritance of five HLA loci haplotypes in 20 families showed the expected distribution of parental haplotypes in sibling pairs. The study revealed DR2sh (DRw16), DR4 and DRw6 in association with DQw7, and DRw8 with DQw4, and significant positive linkage disequilibria between Bw62 CW1, B51 DRw16, B39 DR4, Bw62 DRw6, and DRw8 DQw4. The DR2-DQw7 and DRw6-DQw7 associations and the first three paired loci disequilibria mentioned are described for the first time in Amerindians and have not yet been found among Japanese, Negroid, or Caucasoid populations.

Immunogenetics of human American cutaneous leishmaniasis. Study of HLA haplotypes in 24 families from Venezuela.

Twenty-four families with one or multiple cases of localized cutaneous leishmaniasis (LCL) from an endemic region with the highest incidence of LCL in Venezuela were typed from HLA-ABC, DR, DQ antigens and complement factors. The parental HLA haplotypes segregated at random among healthy and affected siblings but in backcross families significantly higher frequencies of HLA-A28 (p = 0.0018), -Bw22 (p = 0.0122), or -DQw8 (p = 0.0364) were present in affected compared to healthy siblings. HLA-B15 showed a higher frequency (p = 0.0062) among the latter group. Haplotypes Bw22CF31 (p = 0.0076) and Bw22DRw11DQw7 (p = 0.0163) were also significantly more frequent in affected compared to healthy siblings and A2Cw- (p = 0.0445) among the latter. No HLA genetic linkage with a putative LCL susceptibility gene(s) could be demonstrated in this study. A case/control comparison of 26 unrelated LCL patients (one proband from each family) and healthy individuals of the same ethnic origin confirmed the association of HLA-Bw22 (pc = 0.048) and -DQw3 (pc = 0.036) with LCL. The relative risk reached 12.5 for Bw22 and 4.25 for DQw3 with ethiologic factors of 0.17 and 0.64, respectively. HLA-DQw3 apparently makes the major contribution as a genetic risk factor for LCL at the population level.

Definition of DW8.2 by primary and secondary mixed lymphocyte cultures.

Using HLA-DW8 homozygous typing cells (HTC) of different ethnic origin it is possible to identify three subgroups of the DW8/DRW8 product (Mickelson et al., 1983). To further characterize the DW8.2 subgroup defined by HTCs of Amerindian origin we have now generated bulk PLTs within members of one extended Amerindian family and within selected HTCs of Caucasian, Oriental, and Amerindian origin. A panel of 61 DRW8 positive and negative donors of the three ethnic groups was used to test 15 different PLTs. Our results demonstrate that it is possible to generate DW8.1, 8.2, or 8.3 sensitized lymphocytes which distinguish in secondary cultures between each of the three subgroups of the DW8/DRW8 products. Of 40 DRW8 cells tested, 100% Caucasians typed as DW8.1, 100% Amerindians were 8.2; 75% Orientals were DW8.3; 8.3% were DW8.2, and 16.6% could not be classified within any of these subgroups. DRW8 individuals of mixed ethnic origin typed as either DW8.1 or DW8.2 and one DRW8 homozygous donor behaved as heterozygous 8.1/8.2. These results confirm the subdivision of the DW8/DRW8 product and explain the poor correlation and unexpected responses reported in MLC with DW8 HTCs and DRW8 donors of different ethnic origin.

Family studies of the HLA system in acute post-streptococcal glomerulonephritis.

Eighteen families (67 siblings) of index cases with acute post-streptococcal glomerulonephritis (APSGN) were typed for HLA-A,B,C,DR antigens. Twenty cases of clinical nephritis and 10 cases of asymptomatic disease with detected among the sibships. In eight families with more than one affected individual comprising 18 sib pairs random segregation of paternal and maternal HLA haplotypes was found (0.5 less than p less than 0.06), but some antigens (CW1, DR3) showed deviation from the expected 1:1 ratio in affected and nonaffected siblings in backcross families. We had previously noticed the existence of Mendelian recessive ratios in APSGN but in the absence of clear evidence for a dominant or recessive mode of inheritance for a putative APSGN susceptibility gene(s), pedigree data were analyzed twice for linkage with HLA using the two genetic models. The data obtained, although not sufficient to reject the hypothesis of linkage, provide no support for it. Comparison of the frequency of 61 HLA antigens among 42 unrelated APSGN patients and 109 controls, showed that HLA-DRW4 is more frequent among the former (pc = 0.0500).

HLA class II DRB1, DQB1, DPB1 polymorphism and cardiomyopathy due to Trypanosoma cruzi chronic infection.

Trypanosomiasis is an important cause of cardiomyopathy in endemic rural areas of Latin America. Previous studies have suggested participation of HLA molecules in the immune response regulation of T. cruzi infection, and association of HLA antigens with heart damage. One hundred and eleven unrelated T. cruzi antigen-seropositive individuals were tested for HLA class II alleles by the polymerase chain reaction and sequence specific oligonucleotide (PCR-SSO) method. Patients were classified in 3 groups according to clinical and electrocardiographic characteristics: asymptomatics (group A), with arrhythmia (group B), and with overt congestive heart failure (group C). Statistical analysis confirmed the significant increment of the DRB1*01 DQB1*0501 haplotype (p = 0.03) previously reported by our laboratory in patients with cardiomyopathy. The DPB1*0401 allele frequency is also significantly increased in patients with heart disease (groups B + C) (p = 0.009) while DPB1*0101 frequency is higher among the asymptomatic group (p = 0.04) compared with individuals of group C. The DPB1*0401 allele in homozygous form or in combination with allele DPB1*2301 or 3901, was found present more often in patients of groups B and C. Thus, the combination of two of these three alleles, sharing specific sequence motifs in positions 8, 9, 76, and 84-87 confers a relative risk of 6.55 to develop cardiomyopathy in seropositive patients (p = 0.041). Furthermore, 32% of the cardiomyopathics have either DRB1*01 DQB1*0501 and/or DPB1*0401/*0401, 0401/*2301, or* 0401/*3901 compared with 9% of the seropositive asymptomatics (OR = 5.0; p = 0.006).

A study of HLA-DR2 associated HLA-Dw/LD specificities.

HLA-Dw2 and Dw12 are both associated with HLA-DR2; however, these specificities accounts for only 86% (161/188) of the DR2+ haplotypes in our North American Caucasian panel. In an attempt to identify new DR2 associated antigenic clusters, we have generated four primed lymphocyte (LD) typing (PLT) reagents in haploidentical familial combination against DR2+ Dw blank haplotypes. These reagents were positively restimulated by 11 of 16 DR2+ Dw blank cells tested, with good discrimination from Dw2 and Dw12+ cells, thus identifying a new antigenic cluster provisionally termed LD-MN2. We have compared the LD-MN2 specificity with the specificity LD-5a defined by two DR2+ HTCs, BAS and REM, (Layrisse, Caracas) which have been included in the pre-1984 Workshop Cluster DB9. Although none of our DR2+ cells gave typing responses to these two HTCs defining LD-5a, PLT studies did indicate an interrelationship between these specificities and with the specificity tb24 defined with the HTC, FJO (Betuel). The LD-5a HTCs, four LD-5a heterozygous cells, and two additional HTCs (WJR-Hansen, Seattle and FJO/tb24--Betuel, Lyon) significantly restimulated the anti-MN2 PLT reagents, though usually not as strongly as the MN2+ cells. MN2+ cells primed against the LD-5a HTCs were restimulated by only the LD-5a+ cells. Dw2+ cells primed against FJO were restimulated by some, but not all MN2+ cells. These results suggest that MN2, tb24, and LD-5a share some determinants, not shared with most cells which type as Dw2 and Dw12, though differing by other stimulatory determinants. These studies emphasize the necessity of studying new antigenic clusters by both PLT and HTC methodologies as well as testing different ethnic groups.

Extended HLA haplotypes in a Carib Amerindian population: the Yucpa of the Perija Range.

Eleven MHC loci haplotypes have been defined among a Carib speaking Amerindian population; the Yucpa, inhabiting the northern section of the Perija Range, on the limits between Colombia and Venezuela. This tribe has been known with the name of "Motilones mansos" and is located close to the Chibcha-Paeze speaking Bari or "Motilones bravos." Seventy-three full blooded Yucpa living at the villages of Aroy, Marewa, and Peraya, were selected using a genealogy previously collected by an anthropologist and tested for Bf-C4AB complement allotypes and by serology, high resolution PCR-SSO and SBT typing for HLA class 1 and class 2 alleles. Combinations of 6 HLA-A, 6 HLA-B, 5 HLA-C, 1 Bf, 3 C4AB, 3 DQA1, 3DQB1 and 2 DPA1 and 2 DPB1 alleles present in this population originate 17 different haplotypes, 3 of which represent 63% of the haplotypic constitution of the tribe. The presence of 13 individuals homozygous for 11-loci haplotypes corroborates the existence of the following allelic combinations: DRB1*0411 DQA1*03011 DQB1*0302 DPA1*01 DPB1*0402 with HLA-A*6801 C*0702 B*3909 BfS C4 32 (f = 0.3372) or with A*0204 C*0702 B*3905 (f = 0.1977) and a third haplotype which differs only in DRB1*0403 and A*2402 (f = 0.0930). The results demonstrate the isolation of the tribe and the existence of high frequencies of a reduced number of "Amerindian" ancestral and novel class 1 and class 2 alleles (B*1522, DRB1*0807) with significant linkage disequilibria. These results will be useful to test the hypothesis that differentiation of Amerindian tribal groups will have to rely on haplotypes and micropolymorphism rather than allelic lineage frequencies due to the uniformity shown thus far by the putative descendants of the original Paleo-Indians.

Extended HLA haplotypes among the Bari Amerindians of the Perija Range. Relationship to other tribes based on four-loci haplotype frequencies.

Extended HLA haplotypes among Bari Amerindians living at the Perija Range on the limits between Colombia and Venezuela have been defined using serology for class I, electrophoresis and immuno-fixation for Bf and C4, and PCR-SSO for class II loci typing. Haplotypes were assigned based on family studies and gene frequencies were calculated using a subset of less related subjects selected from the genealogy. No rare class III variants were observed, but the characteristic low HLA diversity of isolated Amerindians populations present also in the Bari extends to Bf and C4. Thus there were only 22 different haplotypes segregating in families when nine loci were considered. All of them except three carried Bf*S, C4A*3, C4B*1. The null allele C4A*Q0 reached a frequency of 0.147 and was predominantly present in A24 Cw7 B39 DRB1*0411 haplotypes. In contrast to what has been reported using HLA alleles or class I haplotype frequencies and other isolated South American tribes, genetic distance estimates based on A-Cw-B-DR haplotype frequencies show a closer relationship between the two linguistically but geographically distant Venezuelan tribes, the Bari and the Warao, as compared to two culturally different Brazilian populations. The information reported here will be useful for identifying ancestral haplotypes in native peoples of America, for population comparison, and for discussing the differential influence of MHC haplotype diversity and population survival when similar data on other Amerindian tribes becomes available.

HLA-C(*)03 is a risk factor for cardiomyopathy in Chagas disease.

Previous studies have shown the effect of class 1 as detected by serology or class 2 HLA genes by oligotyping upon susceptibility or resistance to the cardiomyopathy that develops in approximately one third of the Trypanosoma cruzi chronically infected patients. Low and intermediate resolution DNA typing of class 1 alleles was performed in a sample of 113 serologically positive individuals with and without cardiomyopathy. A polymerase chain reaction-sequence-specific oligonucleotide probe method using primers and probes from the British Society of Histocompatibility and Immunogenetics as modified for the VII Latin American Histocompatibility Workshop by D. Middleton, and LiPA kits from Innogenetics were used. Several alleles (A(*)11, A(*)31, B(*)15, B(*)35, B(*)45, B(*)49, B(*)51, and C(*)03) showed increased frequencies among patients with cardiac damage versus the asymptomatic group, but only the last one remained significant after correction of the p value (OR = 5.8, p(c) = 0.03). HLA-C(*)03 showed linkage disequilibrium with B(*)40 and B(*)15 and although both haplotypes were increased in cardiopathic patients compared with asymptomatic individuals, the difference is not significant. These results suggest that the HLA-C*03 allele could confer susceptibility to the development of cardiomyopathy among Venezuelan T. cruzi seropositive individuals and contrast with the protective effect conferred by the HLA B40 Cw3 haplotype among Chilean chagasic patients. Further studies will be needed to confirm the role of this allele on the cardiomyopathy of Chagas disease.

[High- and low-risk molecular sequences in autoimmune diseases. An analysis of type I diabetes in Latin America]. / Secuencias moleculares de alto y bajo riesgo en enfermedades autoinmunes. Un análisis de la diabetes tipo I en Latinoamérica.

Type I diabetes is an autoimmune and a polygenic disease, in which MHC-class II genes contribute to 48% of the disease. The aim of the present study, is to provide a guideline to understanding the molecular association of these genes, through the immunogenetic analysis of 3 Latin american mestizo populations. We included 606 individuals, 349 patients with DMDI and 257 healthy controls coming from 3 geographical areas: Mexico City, Mexico; Caracas, Venezuela and Medellin, Colombia. The results clearly indicate that in mestizo groups, the diabetogenic haplotypes are from mediterranean ancestry, while protection is due to Amerindian genes. It was demonstrated that the relevant sequences for IDDM expression are located to DRB1 and DQB1 loci with a minimal contribution of DQA1 residues. The sequences determining peptide recognition and the induction of TH1 cells mediating the cellular autoimmune response are in positions DRB1-57 and 74 (an aspartic acid and a glutamic acid respectively, confer protection), modulated by D-57 in the DQ, 8 chain. These data show that DRB1-DQB1 haplotypes are central for IDDM expression and open new pathways for the disease management.

HLA-DRB1*0402 haplotypes without DQB1*0302 in Venezuelan patients with pemphigus vulgaris.

The two basic forms of autoimmune intraepidermal blistering diseases, pemphigus vulgaris (PV) and pemphigus foliaceus (PF), affect different layers of the skin, have different symptoms and target different antigens. We have defined human leukocyte antigen (HLA)-DRB1-DQB1 alleles and haplotypes in a case-control study of 66 non-Jewish patients attending a public reference Hospital over the past 10 years. The control group consisted of 101 matched individuals tested also by medium to high-resolution polymerase chain reaction-sequence-specific oligonucleotide with primers and probes from the 12th and 13th International Histocompatibility Workshop. Patients and controls were descendants of three-generation individuals born in the country. Among the patients, 49 had PV, 50% showed predominantly mucosal involvement, 50% showed predominantly the cutaneous clinical phenotype and 17 had PF. Statistically significant HLA-DR frequency differences between patients with PV and controls were found only for DRB1*0402 and DRB1*1401 [odds ratio (OR) = 27.22, confidence interval (CI) 94.7-7.82, P= 1.1 x 10(-14) and OR = 46.56, CI 801.4-2.70 P= 7.5 x 10(-6), respectively]. Both alleles were also increased in the patients with PF compared with the controls (OR = 7.0, P= 0.038 and OR = 21.64, P= 0.009, respectively), but the significance of the difference did not resist Bonferroni correction. Haplotype analysis showed that DRB1*0402 was always present with DQB1*0302 and DRB1*1401 with DQB1*0503, but no independent effect of the DQB1*0302 in the former haplotype was evident. Our results support the hypothesis that the DRB1*0402 without DQB1*0302 is the most relevant HLA-DRB1 allele responsible for the pathogenesis of pemphigus in Venezuelan patients with PV and discard the DQB1*0302 influence observed in other populations.

Human leukocyte antigen class I and MICA haplotypes in a multicase family with Cladophialophora carrionii chromoblastomycosis.

Previous studies carried out in an endemic semiarid region northwest of Venezuela at Falcon State have shown a prevalence of 15.4/1000 of chromoblastomycosis following traumatisms with xenophile vegetation infected with Cladophialophora carrionii. We performed high-resolution DNA typing of human leukocyte antigen (HLA)-A, -B and -C and major histocompatibility complex class I chain related gene A (MICA) alleles and segregation analysis in 49 members of one extended family with 12 affected individuals, who have lived for approximately 70 years in this endemic zone. None of the alleles, haplotypes or genotypes is shared by all the patients. No deviation from the expected HLA haplotype distribution or association of chromoblastomycosis with HLA-A, -B and -C haplotypes was observed. Further, a haplotype-sharing transmission/disequilibria testing of 11 nuclear families did not give enough evidence to claim linkage (P = 0.398), suggesting that genes located in the short arm of chromosome 6 may not be relevant in the immune response toward infection with C. carrionii in this Venezuelan endemic zone. Deleted MICA alleles on HLA-B*4802 haplotypes were present among several members of the extended family, but only two of them were affected.