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自己免疫性糸球体腎炎モデルマウスの解析 : MRL/MpJマウス由来の第1染色体テロメア領域に原因遺伝子を求めて

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Title: 自己免疫性糸球体腎炎モデルマウスの解析 : MRL/MpJマウス由来の第1染色体テロメア領域に原因遺伝子を求めて
Other Titles: Research on autoimmune glomerulonephritis model mouse : Candidate gene on the telomeric region of chromosome 1 derived from MRL/MpJ
Authors: 市居, 修1 Browse this author →KAKEN DB
Authors(alt): Ichii, Osamu1
Issue Date: 25-Mar-2009
Abstract: Autoimmune glomerulonephritis, a complication of systemic autoimmune disease, is caused by renal glomerular deposition of immune complexes circulating through the body. Previous studies have reported that the telomeric region of chromosome 1 in some disease model mice contains several loci that are associated with autoimmune glomerulonephritis susceptibility. However, it is unclear whether these loci originated inform the same genes. In order to detect autoimmune glomerulonephritis at an early stage and provide subsequent therapy, it is essential to identify the candidate genes and aggravating factors of this disease. In this study, we created an MRL/MpJ-type congenic mouse that carries the telomeric region of chromosome 1 and is susceptible to autoimmune disease. We confirmed the application of this congenic strain in autoimmune glomerulonephritis models, pathology analyses, and the search for candidate genes or aggravating factors of this disease. We created an MRL/MpJ-type congenic mouse (B6.MRLc1) which was homozygous in the telomeric region of chromosome 1 (82-100 cM) [B6.MRLc1(82-100)] by applying serial backcrosses between C57BL/6 and (C57BL/6 × MRL/MpJ)F1. The B6.MRLc1(82-100) mice showed mild glomerular damage from 6 months of age. Their glomerular basement membranes were hypertrophic and contained IgG depositions. Glomerular damage in the B6.MRLc1(82-100) mice was more severe in the females; their glomerulus deteriorated with age. Furthermore, splenomegaly, which occurs due to proliferation of CD3- or B220-positive cells, was observed in the spleen of the B6.MRLc1(82-100) mice. Serum levels of blood urea nitrogen (BUN) and the anti-double strand DNA (dsDNA) antibody in the B6.MRLc1(82-100) mice were found to have increased with age and were also significantly higher than those of C57BL/6 at 6 months of age. From these findings, it became evident that the B6.MRLc1(82-100) mice had developed autoimmune glomerulonephritis and the candidate gene derived from MRL/MpJ was localized on chromosome 1 (82-100 cM). We named the loci in the B6.MRLc1(82-100) strain associated with autoimmune glomerulonephritis susceptibility "Mag" (MRL autoimmune glomerulonephritis). In order to reduce the number of regions of these loci, we produced 3 new B6.MRLc1 strains, namely, B6.MRLc1(82-86), B6.MRLc1(92-100), and B6.MRLc1(100), and assessed their pathology. Of these 3 strains, only the B6.MRLc1(92-100) strain showed glomerular damage with immune complex deposition in addition to elevated spleen weights, serum BUN and anti-dsDNA antibody concentrations, and urinary albumin excretions. These results confirm the localization of autoimmune glomerulonephritis candidate genes on chromosome 1 (92-100 cM). For the selection of the candidate gene, relative mRNA expression levels of Crp (pentraxin-related C-reactive protein), Apcs (serum amyloid P-component), Fcgr2b (low-affinity Fc receptor for IgG IIB), and Fcgr3, which are major immune-associated genes present in chromosome 1 (92-100 cM), were analyzed in the major organs. A high expression of Fcgr3 and an elevated Fcgr3/Fcgr2b ratio were observed in the kidney, spleen, and thymus of the B6.MRLc1(92-100) strain. Both the inhibitory and active Fc gamma receptors coded by Fcgr2b and Fcgr3, namely, FcγRIIB and FcγRIII, respectively, were localized on glomerular mesangial regions and dendritic cells in the lymphoid organs. The FcγRIII expressions of the B6.MRLc1(92-100) strains in these regions were stronger than those of C57BL/6. From these findings, the inhibitory and active FcγR genes were considered to be candidate genes. It was strongly suggested that the imbalance between those genes had a great impact on the pathogenesis of autoimmune glomerulonephritis. Female mice showed a higher incidence of systemic lupus erythematosus (SLE), which is the most common autoimmune disease. Further, similar sex-related differences were observed in the B6.MRLc1(82-100) strain; glomerulonephritis was more severe in the female mice than in the male mice. Since sex hormones were considered to be one of the many definitive factors responsible for these sex-related differences, we castrated sex organs of both sexes of the B6.MRLc1(82-100) strain and assessed the effects of gonadectomy in inducing autoimmune glomerulonephritis. In sham operated groups, the female mice showed significantly higher values than the male mice with regard to the glomerular damage scores, the percentage of IgG- and CD3-positive glomerulus, the spleen weights, and the serum levels of total IgG and anti-dsDNA antibodies. The effects of gonadectomy in the male mice were more apparent than those in the female mice. The male gonadectomy group showed significantly higher values than the male sham operated group with regard to the glomerular damage scores, the percentage of IgG- and CD3-positive glomerulus, the urinary albumin excretions, the spleen weights, and the CD3-positive areas in the spleen. Interestingly, CD3-positive cells were observed in both the thymic cortex and the thymic medulla, in all animals except the males in the sham operated groups. On analysis of the active Fcgr3/Fcgr2b ratio in the kidney, the male gonadectomy groups and the female sham operated groups were found to have significantly higher values of the active Fcgr3/Fcgr2b ratio than the male sham operated groups. These results suggested that the pathological sex-related differences in autoimmune glomerulonephritis were strongly affected by the inhibitory roles of male sex hormones. The elevated levels of expression of inflammatory cytokines, chemokines, and their receptors were recently clarified by the array analysis using humor or biopsy samples from patients. These factors are attracting researchers as the targets for disease controls. We attempted to identify the aggravating factors for autoimmune glomerulonephritis in the B6.MRLc1(82-100) strain by studying the expressions of inflammatory mediators by comprehensive PCR array analysis. In the kidney of the B6.MRLc1(82-100) strain, several genes of chemokine ligands and their receptors showed remarkably high expression levels. The expressions of the Cxcl5 gene [chemokine (C-X-C motif) ligand 5] in the B6.MRLc1(82-100) strain were 72-fold higher than those in C57BL/6; the expressions of this gene increased over time in the B6.MRLc1(82-100) strain. The CXCL5 protein coded by the Cxcl5 gene was mainly localized on glomerular podocytes. In contrast, the gene that showed the highest expression in the kidney of the B6.MRLc1(82-100) strain was Il1f6 (Interleukin 1 family member 6), which is a member of the IL-1 family; these expressions were 186-fold higher than those of C57BL/6. Furthermore, Il1f6 gene expressions increased over time in the kidneys of the B6.MRLc1(82-100) strains. The expressions of the Il1f6 gene and its protein were observed in the kidneys from the distal tubules to the collecting ducts. The dilated tubules in the kidneys of the B6.MRLc1(82-100) strains, in particular, showed a higher level of expression of this protein. Interestingly, the number of renal tubules containing the IL-1F6 protein was in proportion to the number of renal tubules containing protein casts as indices for proteinuria. These results suggest that the Cxcl5 and Il1f6 genes, which were highly expressed in the kidney of the B6.MRLc1(82-100) strain, play important roles as aggravating factors of glomerular damage and tubulointerstitial lesions. Thus, by research on disease model mice, it was concluded that the quantitative changes in candidate genes derived form mouse genome, sex hormones, and local aggravating factors in mouse kidney were closely related to the pathogenesis of autoimmune glomerulonephritis. These findings will provide fundamental knowledge that will enable early diagnosis and subsequent therapy of autoimmune glomerulonephritis in the fields of medicine and veterinary science.
Conffering University: 北海道大学
Degree Report Number: 甲第9038号
Degree Level: 博士
Degree Discipline: 獣医学
Type: theses (doctoral)
Appears in Collections:学位論文 (Theses) > 博士 (獣医学)

Submitter: 市居 修

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