ISSN:
1440-1797
Source:
Blackwell Publishing Journal Backfiles 1879-2005
Topics:
Medicine
Notes:
Background: The IgA1 molecule, which is predominantly deposited in glomeruli of IgA nephropathy, has the characteristic O-glycan side chain in its hinge region. Several lines of evidence have demonstrated that underglycosylated IgA1 acquires self-aggregation1 and binding affinities to the extra-cellular matrix,1,2 resulting in deposition in the glomeruli.3,4 However, there is little information concerning actual biological activities of the underglycosylated IgA1 deposited in glomeruli, which lead to mesangial cell proliferation and/or matrix expansion.Aim: To clarify the influence of O-glycan side chain in the hinge of IgA1 on its biological activity in mesangial cells, we performed a comprehensive gene expression profiling analysis of human cultured mesangial cells stimulated by enzymatically underglycosylated IgA1 using cDNA array.Methods: IgA1 was purified by affinity chromatography using anti IgA column followed by Jacalin column. Asialo/agalacto IgA1 was obtained by enzyme digestion using sialidase and β-galactosidase. Heat aggregated IgA1 was obtained by incubation at 63°C for 150 min. Cultured human mesangial cells were stimulated for 3 h by non-treated IgA1, heat aggregated IgA1 or asialo/agalacto IgA1, respectively, and total RNA was obtained. Only the enzyme stimulation was performed as a negative control for asialo/agalacto IgA1. After DNase treatment, isotope labelled probes were prepared by reverse transcription and hybridized to the Atlas Human 1.2 Array (CLONTECH, Palo Alto, CA, USA) according to the manufacturer’s protocol. A total of 1176 arrayed genes were quantitatively evaluated using BAS.Results: Expression profiles of 1176 genes were obtained. In all experiments, none of the three negative controls on the cDNA array was detected. The expression profiles of mesangial cells by each IgA1 stimulation resembled one another, but they were widely different from that of mononuclear cells. On the whole, 10% of the genes expressed in the mesangial cells were up-regulated by asialo/agalacto IgA1 stimulation compared to that by enzyme stimulation. The results suggested that under-O-glycosylated IgA1 has a biological function in human mesangial cells in vitro. In good agreement with previous studies,5 heat aggregated IgA1 stimulation up-regulated IL-6 and TNF-α expressions in mesangial cells compared to by non-treated IgA1. Asialo/agalacto IgA1 stimulation also up-regulated these molecules, suggesting its pathogenetic role in IgA nephropathy. However, since a similar tendency was revealed by enzyme stimulation alone, further study may be necessary to establish these observations.Conclusion. In the present study, we investigated the comprehensive expression profiles of mesangial cells stimulated by various types of IgA1 using cDNA array. This approach may serve as a useful database not only to evaluate the biological activities of under-O-glycosylated IgA1 but also to elucidate the pathogenesis of IgA nephropathy.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1046/j.1440-1797.2001.00018.x
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