title: Detection of cytokine mRNA-expressing cells in peripheral blood of patients with IgA nephropathy using non-radioactive in situ hybridization.
abstract: IgA nephropathy (IgA-N) is considered to be an immune-mediated disorder and several immunological abnormalities have been observed. In the present study, we optimized non-radioactive in situ hybridization and applied this technique to evaluate the degree of expression of various cytokine mRNAs in peripheral blood mononuclear cells (PBMC) taken from patients with IgA-N on cytospin preparation. Using this method, together with image analysis, we examined the expression of mRNA in cells which secrete cytokines, such as IL-2, interferon-gamma (IFN-gamma), IL-4, IL-5 and IL-6. The expression of these mRNAs was clearly observed in monocytes and lymphocytes at a single-cell level. Compared with healthy adults, the expression of IL-4 mRNA, IL-5 mRNA and IL-6 mRNA, but not IL-2 mRNA or IFN-gamma mRNA, was significantly enhanced. Our results indicate that non-radioactive in situ hybridization method is a powerful technique for analysis of cytokine mRNAs in PBMC at a cellular level. Our results also suggest that mRNA expression of IL-4, IL-5 and IL-6, produced by Th2 cells, is increased in patients with IgA-N. The abnormal regulatory process involved in cytokine expression may play an important immunopathologic role in IgA-N.
authors.affiliation: Second Department of Internal Medicine, Nagasaki University School of Medicine, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Ichinose
authors.forename: M
authors.initials: M
authors.lastname: Miyazaki
authors.forename: T
authors.initials: T
authors.lastname: Koji
authors.forename: A
authors.initials: A
authors.lastname: Furusu
authors.forename: Y
authors.initials: Y
authors.lastname: Ozono
authors.forename: T
authors.initials: T
authors.lastname: Harada
authors.forename: M
authors.initials: M
authors.lastname: Shin
authors.forename: P K
authors.initials: PK
authors.lastname: Nakane
authors.forename: K
authors.initials: K
authors.lastname: Hara
mesh_terms: D000328:Adult; D016207:Cytokines / Q000235:genetics*; D004076:Digoxigenin; D005404:Fixatives; D005922:Glomerulonephritis, IGA / Q000097:blood*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D017403:In Situ Hybridization; D015850:Interleukin-6 / Q000235:genetics; D007963:Leukocytes, Mononuclear / Q000737:chemistry*; D015345:Oligonucleotide Probes; D012333:RNA, Messenger / Q000096:biosynthesis*
chemical_list: D016207:Cytokines; D005404:Fixatives; D015850:Interleukin-6; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D004076:Digoxigenin