RESUMO
One of the major quantitative trait loci (QTLs) associated with arthritis in crosses between B10.RIII and RIIIS/J mice is the Cia5 on chromosome 3. Early in the congenic mapping process it was clear that the locus was complex, consisting of several subloci with small effects. Therefore, we developed two novel strategies to dissect a QTL: the partial advanced inter-cross (PAI) strategy, with which we recently found the Cia5 region to consist of three loci, Cia5, Cia21 and Cia22, and now we introduce the QTL-chip strategy, where we have combined congenic mapping with a QTL-restricted expression profiling using a novel microarray design. The expression of QTL genes was compared between parental and congenic mice in lymph node, spleen and paw samples in five biological replicates and in dye-swapped experiments at three time points during the induction phase of arthritis. The QTL chip approach revealed 4 genes located in Cia21, differently expressed in lymph nodes, and 14 genes in Cia22, located within two clusters. One cluster contains six genes, differently expressed in spleen, and the second cluster contains eight genes, differently expressed in paws. We conclude the QTL-chip strategy to be valuable in the selection of candidate genes to be prioritized for further investigation.
Assuntos
Artrite Experimental/genética , Artrite Experimental/imunologia , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Locos de Características Quantitativas , Animais , Cruzamento , Mapeamento Cromossômico , Linfonodos/metabolismo , Camundongos , Camundongos Congênicos , Análise em Microsséries , Baço/metabolismoRESUMO
We investigated the role of the major histocompatibility complex (MHC) region in the specificity of autoimmunity by analysing specifically the development of sialadenitis, but also insulitis, nephritis and autoantibody production in autoimmune-prone nonobese diabetic (NOD) mice where the MHC H2g7 haplotype had been exchanged for the H2q (NOD.Q) or H2p (NOD.P) haplotype. The exchange of H2 haplotype did not affect the frequency of sialadenitis because the H2q and H2p congenic NOD strains developed sialadenitis with the same incidence as NOD. However, the severity of sialadenitis varied among the strains, as NOD.Q >NOD >NOD.P. At 11-13 weeks of age, the NOD.Q (H2q) female mice developed more severe sialadenitis compared to NOD.P (H2p) (P=0.038). At 20 weeks, the NOD (H2g7) female mice showed more severe sialadenitis than NOD.P (P=0.049). This is in contrast to the development of insulitis in the present strains, because the incidence of insulitis was almost completely inhibited by the replacement of the H2g7 haplotype of NOD. The incidence of insulitis in NOD.Q was 11-22%, compared to 75% in NOD, which correlated well with lower titres of anti-glutamic acid decarboxylase (anti-GAD) antibodies in NOD.Q compared to NOD (P=0.009). However, the introduction of the H2q haplotype into the NOD strain instead directed the autoimmune response towards the production of lupus types of autoantibodies, because the incidence of antinuclear antibodies (ANA) in NOD.Q was 89% compared with 11% in NOD.P and 12% in NOD mice, which in turn correlated with a high incidence of nephritis in NOD.Q compared to NOD. Consequently, we show that different haplotypes of MHC are instrumental in directing the specificity of the spontaneous autoimmune inflammation.
Assuntos
Diabetes Mellitus Tipo 1/imunologia , Complexo Principal de Histocompatibilidade/imunologia , Nefrite/imunologia , Sialadenite/imunologia , Animais , Anticorpos Antinucleares/genética , Anticorpos Antinucleares/imunologia , Apoptose/genética , Apoptose/imunologia , Diabetes Mellitus Tipo 1/genética , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Predisposição Genética para Doença , Haplótipos/genética , Haplótipos/imunologia , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Complexo Principal de Histocompatibilidade/genética , Masculino , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos NOD , Nefrite/genética , Sialadenite/genética , Estatísticas não ParamétricasRESUMO
Identification of genes controlling complex diseases has proven to be difficult; however, animal models may pave the way to determine how low penetrant genes interact to promote disease development. We have dissected the Cia5/Eae3 susceptibility locus on mouse chromosome 3 previously identified to control disease in experimental models of multiple sclerosis and rheumatoid arthritis. Congenic strains showed significant but small effects on severity of both diseases. To improve the penetrance, we have now used a new strategy that defines the genetic interactions. The QTL interacted with another locus on chromosome 15 and a partial advanced intercross breeding of the two congenic strains for eight generations accumulated enough statistical power to identify interactions with several loci on chromosome 15. Thereby, three separate loci within the original QTL could be identified; Cia5 affected the onset of arthritis by an additive interaction with Cia31 on chromosome 15, whereas the Cia21 and Cia22 affected severity during the chronic phase of the disease through an epistatic interaction with Cia32 on chromosome 15. The definition of genetic interactions was a prerequisite to dissect the Cia5 QTL and we suggest the partial advanced intercross strategy to be helpful also for dissecting other QTL controlling complex phenotypes.
Assuntos
Artrite/genética , Epistasia Genética , Predisposição Genética para Doença , Locos de Características Quantitativas , Animais , Cruzamentos Genéticos , Feminino , Marcadores Genéticos , Camundongos , Mapeamento Físico do Cromossomo , Fatores de TempoRESUMO
The nonobese diabetic mouse is highly susceptible not only to diabetes but to several autoimmune diseases, and one might suspect that these are controlled by a shared set of genes. However, based on various gene-segregation experiments, it seems that only a few loci are shared and that each disorder is influenced also by a unique set of genes.
Assuntos
Doenças Autoimunes/genética , Camundongos Endogâmicos NOD/genética , Animais , Doenças Autoimunes/imunologia , Feminino , Predisposição Genética para Doença , Variação Genética/imunologia , Complexo Principal de Histocompatibilidade/genética , Complexo Principal de Histocompatibilidade/imunologia , Masculino , Camundongos , Camundongos Endogâmicos NOD/imunologia , Locos de Características Quantitativas/imunologiaRESUMO
The nonobese diabetic (NOD) mouse spontaneously develops autoimmune-mediated diseases such as diabetes and Sjögren's syndrome. To investigate whether NOD genes also promote autoimmune-mediated arthritis we established a NOD strain with an MHC class II fragment containing the A(q) class II gene predisposing for collagen induced arthritis (NOD.Q). However, this mouse was resistant to arthritis in contrast to other A(q) expressing strains such as B10.Q and DBA/1. To determine the major resistance factor/s, a genetic analysis was performed. (NOD.Q x B10.Q)F1 mice were resistant, whereas 27% of the (NOD.Q x B10.Q)F2 mice developed severe arthritis. Genetic mapping of 353 F2 mice revealed two loci associated with arthritis. One locus was found on chromosome 2 (LOD score 9.8), at the location of the complement factor 5 (C5) gene. The susceptibility allele was from B10.Q, which contains a productive C5 encoding gene in contrast to NOD.Q. The other significant locus was found on chromosome 1 (LOD score 5.6) close to the Fc-gamma receptor IIb gene, where NOD carried the susceptible allele. An interaction between the two loci was observed, indicating that they operate on the same or on interacting pathways. The genetic control of arthritis is unique in comparison to diabetes, since none of these loci have been identified in analysis of diabetes susceptibility.
Assuntos
Antígenos CD/genética , Artrite Reumatoide/genética , Colágeno , Complemento C5/genética , Diabetes Mellitus Tipo 1/genética , Receptores de IgG/genética , Animais , Cruzamentos Genéticos , Modelos Animais de Doenças , Feminino , Antígenos H-2 , Imunidade Inata/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NODRESUMO
To identify chromosomal regions containing susceptibility loci for systemic lupus erythematosus (SLE), we performed genome scans in families with multiple SLE patients from Iceland, a geographical and genetic isolate, and from Sweden. A number of chromosomal regions showed maximum lod scores (Z) indicating possible linkage to SLE in both the Icelandic and Swedish families. In the Icelandic families, five regions showed lod scores greater than 2.0, three of which (4p15-13, Z=3.20; 9p22, Z=2.27; 19q13, Z=2.06) are homologous to the murine regions containing the lmb2, sle2 and sle3 loci, respectively. The fourth region is located on 19p13 (D19S247, Z=2.58) and the fifth on 2q37 (D2S125, Z=2.06). Only two regions showed lod scores above 2.0 in the Swedish families: on chromosome 2q11 (D2S436, Z=2. 13) and 2q37 (D2S125, Z=2.18). The combination of both family sets gave a highly significant lod score at D2S125 of Z=4.24 in favor of linkage for 2q37. This region represents a new locus for SLE. Our results underscore the importance of studying well-defined populations for genetic analysis of complex diseases such as SLE.
Assuntos
Cromossomos Humanos Par 2/genética , Lúpus Eritematoso Sistêmico/genética , Animais , Feminino , Ligação Genética , Marcadores Genéticos , Genótipo , Humanos , Islândia , Escore Lod , Masculino , Camundongos , Linhagem , SuéciaRESUMO
We have previously reported linkage of systemic lupus erythematosus to chromosome 2q37 in multicase families from Iceland and Sweden. This locus (SLEB2) was identified by linkage to the markers D2S125 and D2S140. In the present study we have analyzed additional microsatellite markers and SNPs covering a region of 30 cM around D2S125 in an extended set of Nordic families (Icelandic, Swedish, and Norwegian). Two-point linkage analysis in these families gave a maximum lod score at the position of markers D2S2585 and D2S2985 (Z = 4.51, PIC = 0.65), by applying a "model-free" pseudo-marker linkage analysis. Based on multipoint linkage analysis in the Nordic families, the most likely location of the SLEB2 locus is estimated to be in the interval between D2S125 and the position of markers D2S2585 and D2S2985, with a peak multipoint lod score of Z = 6.03, assuming a dominant pseudo-marker model. Linkage disequilibrium (LD) analysis was performed using the data from the multicase families and 89 single-case families of Swedish origin, using the same set of markers. The LD analysis showed evidence for association in the single-case and multicase families with locus GAAT3C11 (P < 0.0003), and weak evidence for association was obtained for several markers located telomeric to D2S125 in the multicase families. Thirteen Mexican families were analyzed separately and found not to have linkage to this region. Our results support the presence of the SLEB2 locus at 2q37.
Assuntos
Cromossomos Humanos Par 2 , Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , Sequência de Bases , Mapeamento Cromossômico , Primers do DNA , Ligação Genética , Genética Populacional , Humanos , Funções VerossimilhançaRESUMO
There is considerable evidence that the development of systemic lupus erythematosus (SLE) has a strong genetic basis. For more than 20 years, much effort has been made to understand the genetics of SLE. Association studies in humans suggest the existence of genetic effects by the alleles encoded in the HLA, deficiencies in the complement genes and the low-affinity variants of Fcgamma-receptors. In mouse models of SLE at least 13 loci have been identified, including the MHC, linked to lupus-related phenotypes such as nephritis and production of autoantibodies. Recently, linkage studies have been performed in human SLE; one investigating a candidate region based on synteny to a murine susceptibility locus and four genome-wide linkage studies in various populations. Linkage was demonstrated to several chromosomal regions, some of which are syntenic to murine lupus susceptibility loci. Interestingly, many of the identified chromosomal regions co-localise with loci implicated in other autoimmune diseases.
Assuntos
Doenças Autoimunes/genética , Lúpus Eritematoso Sistêmico/genética , Alelos , Animais , Doenças Autoimunes/etnologia , Mapeamento Cromossômico , Cromossomos Humanos/genética , Complemento C4a/deficiência , Modelos Animais de Doenças , Doenças em Gêmeos/genética , Etnicidade/genética , Proteína Ligante Fas , Ligação Genética , Predisposição Genética para Doença , Antígenos HLA/genética , Antígenos HLA/imunologia , Humanos , Lúpus Eritematoso Sistêmico/etnologia , Nefrite Lúpica/genética , Complexo Principal de Histocompatibilidade/genética , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos MRL lpr , Receptores de IgG/genética , Especificidade da Espécie , Receptor fas/genéticaRESUMO
OBJECTIVE: To study the contribution of the IL10 gene to the susceptibility to systemic lupus erythematosus (SLE). METHODS: Analysis by fluorescent-semiautomated genotyping of a dinucleotide repeat located in the promoter region of the IL10 locus (microsatellite G). RESULTS: No significant difference was found in the frequencies of the microsatellite alleles of 330 Mexican patients with SLE compared to 368 controls from the same population. Two-point linkage analyses were carried out using 13 Mexican, 13 Swedish, and 8 Icelandic families with 2 or more cases with SLE. No linkage was revealed between IL10 and SLE, using a variety of parameter settings. CONCLUSION: Our results do not support that the IL10 gene contributes to the susceptibility to SLE in the populations we studied.
Assuntos
Interleucina-10/genética , Lúpus Eritematoso Sistêmico/genética , Repetições de Dinucleotídeos/genética , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Lúpus Eritematoso Sistêmico/imunologia , Masculino , México , Repetições de Microssatélites , Regiões Promotoras Genéticas/genéticaRESUMO
Systemic lupus erythematosus is a disease of unknown etiology. Multiple genetic factors are believed to be involved in its pathogenesis. In addition, and due to genetic heterogeneity, these factors and/or their combinations may be different in different ethnic groups, while some might be shared between populations. We have performed genome scans in multicase families from three different population groups, two from Northern Europe, with a high degree of homogeneity, and the third from a recently admixed population of Mexican Mestizos. Although our family material is relatively small, the results presented here show that using family sets from well defined populations are sufficient to detect susceptibility loci for SLE. Our results also reveal the chromosomal regions most likely to contain susceptibility genes for SLE.
Assuntos
Genoma Humano , Lúpus Eritematoso Sistêmico/genética , Etnicidade/genética , Feminino , Técnicas Genéticas , Genética Populacional , Humanos , Islândia/epidemiologia , Indígenas Norte-Americanos/genética , Escore Lod , Lúpus Eritematoso Sistêmico/epidemiologia , Masculino , México/epidemiologia , Suécia/epidemiologia , Estados Unidos/epidemiologia , População Branca/genéticaRESUMO
A set of 391 microsatellite markers (Weber set 6), 85% of which consist of tri- and tetranucleotide repeat markers, were used to design chromosome-specific panels that allowed for a high degree of multiplexing with respect to the fragment size range and fluorophore (FAM, HEX, TET). This marker set has an average coverage of 10.5 cM, with the largest gap being 28.1 cM. The markers were divided into 49 panels, with a maximum degree of multiplexing of 15 markers per panel. The utility of the markers for analysis of DNA from blood, hair, and formalin-fixed archival tissue biopsies was evaluated with respect to amplification efficiency, product yield, and degree of preferential amplification of the shorter allele in heterozygotes. The amplification efficiency was inversely related to repeat length and amplicon length. Based on the analysis of DNA from formalin-fixed biopsies, 51 markers suitable for loss of heterozygosity (LOH) studies were identified. The utility of the marker set for genome scanning, LOH, and forensic analyses is discussed.