Comparative analysis of commercial human primary mesangial cell, implications for experimental design.

Abstract

Background: Mesangial cells (MCs) are involved in several glomerular diseases such as IgA nephropathy and diabetic kidney disease. In vitro work on human MCs is mainly conducted on primary MCs. However, cells from different donors could be significantly different, thus potentially affecting the outcome of the experiments.

Method: We have compared commercially available primary human MCs from two different sources: HMCv1 and HMCv2. The cells were characterized using qPCR, western blot, and immunofluorescence. Response to PDGF-BB was assessed with proliferation assays, proteomics, and qPCR. Response to angiotensin II was assessed through contractility assay and response to IL-1β, diabetic milieu and TGFβ1 with qPCR.

Results: Cells from both sources expressed mesangial markers. HMCv1, but not HMCv2, showed significant contractility in response to angiotensin II. Both HMCv1 and 2 significantly increased their proliferation rate in response to PDGF-BB. Proteomics revealed a stronger response to PDGF-BB for HMCv1 in respect to HMCv2, though similar pathways were regulated in both. IL-1β stimulus was stronger in HMCv1 in terms of increased expression of IL6 and CCL2/MCP1 mRNA. Diabetic milieu increased expression of IL-6 for both HMCv1 and 2, but significantly higher for HMCv1. TGFβ1 gave similar results in terms of IL-6 expression for cells from both sources. In addition, a list of 144 potential mesangial markers was compiled, that can be used for identification of MCs in omics data.

Conclusion: This study shows that there are broad differences between sources of primary human MCs. The potential differences between clones of primary MCs need to be carefully considered when conducting in vitro experiments.