Kaori Muro

Mitochondrial DNA Mutations in Focal Segmental Glomerulosclerosis Lesions

Glomerular epithelial cells are primary pathogenic sites in focal segmental glomerulosclerosis (FGS) lesions. Glomerular epithelial cells are regarded as terminally differentiated cells that do not proliferate. These characteristics are also noted for neurons and muscular cells, which are major sites of mitochondrial DNA (mtDNA) mutation accumulation. Screening for mtDNA mutations was performed with renal biopsy specimens from patients with primary FGS and patients with IgA nephropathy (as subjects with secondary FGS and as control subjects). mtDNA extracted from kidney biopsy specimens was amplified with appropriate primer pairs for study of the mtDNA point mutations 3243A-->G, 3271T-->C, 8344A-->G, and 8993T-->G/C, as well as the common deletion (a 4977-bp deletion spanning mtDNA nucleotide pairs 8469 to 13447). In situ amplification of both total mtDNA and the common deletion was also performed. Two patients with FGS demonstrated the 3243A-->G point mutation; 12 patients with FGS and seven patients with IgA nephropathy accompanied by glomerulosclerotic lesions exhibited the common deletion in their kidney tissue. No patient demonstrated the mtDNA mutations 3271T-->C, 8344A-->G, or 8993T-->G/C. The degree of heteroplasmy for the 3243A-->G point mutation was >85%; however, the heteroplasmy for the common deletion was <1%. As determined with in situ PCR, normal mtDNA was mainly distributed in the tubular epithelium and mtDNA with the common deletion was mainly distributed among glomerular epithelial cells. In conclusion, it is suggested that mtDNA mutations are distributed in glomerular epithelial cells among some patients with primary FGS or secondary FGS with IgA nephropathy. These mutations may be related to glomerular epithelial cell damage.

Predominance of type-2 immune response in idiopathic membranous nephropathy. Cytoplasmic cytokine analysis.

Background: The Th1/Th2 paradigm is proving increasingly useful in the understanding of infectious diseases and many autoimmune diseases. Th1 cells predominantly produce interleukin-2 (IL-2) and interferon-γ (IFN-γ) and are instrumental in initiating delayed-type hypersensitivity and activating macrophages. Th2 cells secrete other cytokines, such as IL-4, IL-5, IL-10 and IL-13 that trigger B-cell activation and immunoglobulin synthesis. It has been shown that in patients with membranous nephropathy, there may be a predominance of Th2, because of the presence of IgG, particularly IgG4, which belongs to a subclass of the type-2 immune response, and complement deposits in glomeruli. In this study, we investigated the immunoresponse of helper T cells, i.e. Th predominance in patients with idiopathic membranous nephropathy. Methods: We used flow cytometry to assess the levels of circulating Th cells in patients with idiopathic membranous nephropathy (n = 8) and in normal individuals (n = 23) based on the expression of intracellular type-1 and type-2 cytokines. Because the production of each of these cytokines has a specific time course, we observed the cytokine synthesis at 3, 6, 9 and 12 h after stimulation. Results: The percentages of IL-2+/CD4+ cells from patients with idiopathic membranous nephropathy were significantly lower than those from normal individuals at 6, 9 and 12 h, with the difference becoming more significant over time. IFN-γ+/CD4+ cells and IL-4+/CD4+ cells were not significantly different between the two groups. In patients with idiopathic membranous nephropathy, the percentages of IL-10+/CD4+ cells were significantly higher than those in normal individuals at each point in time. Conclusion: Increased IL-10-producing Th cells may lead to suppression of delayed-type hypersensitivity and activate suppressor cells and IgG4 synthesis, resulting in idiopathic membranous nephropathy.

Specific T-cell receptor usage with cytokinemia in Henoch-Schönlein purpura nephritis associated with Staphylococcus aureus infection

Abstract. Hirayama K, Kobayashi M, Muro K, Yoh K, Yamagata K, Koyama A (Department of Internal Medicine, Institute of Clinical Medicine, University of Tsukuba; and Department of Nephrology, Tokyo Medical University Kasumigaura Hospital, Ibaraki, Japan) Specific T‐cell receptor usage with cytokinemia in Henoch–Schönlein purpura nephritis associated with Staphylococcus aureus infection (Original article). J Intern Med 2001; 249: 289–295.

Usage of T cell receptor variable segments of the beta-chain in IgA nephropathy

Background: We previously reported that glomerulonephritis associated with Staphylococcus aureus infection (SAGN) showed an increased usage of T cell receptor Vβ 5.3 and 8 in peripheral lymphocytes and mesangial IgA and IgG depositions. To elucidate the immunological mechanisms and pathogenesis of IgA nephropathy, we analyzed the usage of TCR Vβ in both peripheral blood lymphocytes (PBLs) and renal infiltrating T cells from IgA-N patients. Methods: In 38 patients with IgA nephropathy and controls, the usage of TCR Vβ in PBLs were analyzed using monoclonal antibodies against Vβ 3.1, Vβ 5.1, Vβ 5.2 + 5.3, Vβ 5.3, Vβ 6.7, Vβ 8, Vβ 12.1, and Vβ 13.1 + 13.3 with three-color flow cytometry. Furthermore, we examined immunohistochemically renal biopsy specimens using antibodies against Vβ 5.3 and Vβ 8. Results: The percentages of DR+CD4+CD8– cells, CD45RO+CD4+ cells, and CD45RO+CD4+DR+ cells in PBLs from IgA nephropathy were significantly higher than controls. The percentages of TCR Vβ 5.3 positive cells and TCR Vβ 8 positive cells in PBLs from patients were 1.3 ± 0.1 and 3.1 ± 0.2%, and both were significantly higher than controls. The percentage of renal interstitial TCR Vβ 5.3 expression was significantly higher than that in PBLs. However, there was no significant difference between the TCR Vβ 8 expression in the interstitium and that in PBLs. Conclusions: TCR Vβ 5.3 and 8 usage and broad CD4+ T cell activation have occurred in IgA nephropathy. These changes were similar but weak compared with formerly reported SAGN.

The usage of variable regions of the human T cell receptor in patients with IgA nephropathy

In order to clarify the usage of variable regions of the human T cell receptor (TCRVP) in IgA nephropathy, we analyzed lymphocyte subsets in peripheral blood from patients with IgA nephropathy. We obtained peripheral lymphocytes from patients with IgA nephropathy (group A, n=38), Henoch-Schonlein purpura nephritis (group B, n = lo), superantigen-related nephritis (group C, n= 19), and normal controls (group D, n = 39). Lymphocyte subsets were analyzed on FACScan, using the following biotin, phycoerythrin or FITC-labeled monoclonal antibodies: