Comparative Analysis of Human Primary Mesangial Cell Clones: Implications for Experimental Design

Background: Mesangial cells (MCs) not only form the central stalk of the glomerulus but are of importance for glomerular cross talk and function. The MCs are central in several glomerular diseases such as IgA nephropathy and diabetic kidney disease and are often used in vitro to reveal molecular mechanisms of onset and progression. Human primary MCs (HMCs) are considered the best option for in vitro studies, but over the years we have observed that there are differences between HMCs from different suppliers (and hence donors) in their phenotype and response to stimuli and/or treatment Methods: Commercially available HMCs from two different suppliers were cultured and characterized by their expression of mesangial markers (qPCR, western blot, immunofluorescence, mass spectrometry), response to PDGF-BB (proliferation assay, mass spectrometry) and angiotensin 2 (contractility assay). In addition to the commonly used MC markers we put together a list of 144 MC markers, compiled based on 12 recent articles containing omics data, and these markers were used combination with the proteomic data set. Results: Both clones (HMCv1 and HMCv2) expressed the common MC markers and in the proteomic data set, 70 out of the 144 MC markers were found for both. HMCv1 and 2 and responded with increased proliferation in response to stimulation with PDGF-BB. HMCv1 contracted significantly in response to angiotensin 2, but not HMCv2. Proteomic analysis of HMCv1 and 2 after stimulation with PDGF-BB revealed that HMCv1 was much more responsive than HMCv2 in terms of more differentially regulated proteins and pathways. Conclusion: In conclusion, we found that there were similarities as well as differences between the two clones of HMCs. Both HMCv1 and HMCv2 behaved as MCs in that they expressed the major MC markers and responded to PDGF-BB stimulation. However, HMCv1 was much more responsive to stimulation with PDGF-BB and angiotensin 2 than HMCv2. These differences could have a major impact on the outcome of experiments performed using these cells, and therefore we suggest to carefully examine the response of the chosen HMC clone to relevant stimulants before conducting experiments on HMCs. In addition, out of our list of 144 MC markers, the clones were found to express 70 and we suggest that these markers can be used as a signature for MCs. Funding: Private Foundation Support