Dataset Information

Human mesenchymal stromal cells broadly modulate high glucose-induced inflammatory responses of renal proximal tubular cell monolayers.

ABSTRACT:

Background

Renal proximal tubular epithelial cells (RPTEC) are dysfunctional in diabetic kidney disease (DKD). Mesenchymal stromal cells (MSC) may modulate DKD pathogenesis through anti-inflammatory mediators. This study aimed to investigate the pro-inflammatory effect of extended exposure to high glucose (HG) concentration on stable RPTEC monolayers and the influence of MSC on this response.

Methods

Morphologically stable human RPTEC/TERT1 cell monolayers were exposed to 5?mM and 30?mM (HG) D-glucose or to 5?mM D-glucose?+?25?mM D-mannitol (MAN) for 5?days with sequential immunoassays of supernatants and end-point transcriptomic analysis by RNA sequencing. Under the same conditions, MSC-conditioned media (MSC-CM) or MSC-containing transwells were added for days 4-5. Effects of CM from HG- and MAN-exposed RPTEC/MSC co-cultures on cytokine secretion by monocyte-derived macrophages were determined.

Results

After 72-80?h, HG resulted in increased RPTEC/TERT1 release of interleukin (IL)-6, IL-8, monocyte chemoattractant protein (MCP)-1 and neutrophil gelatinase-associated lipocalin (NGAL). The HG pro-inflammatory effect was attenuated by concentrated (10×) MSC-CM and, to a greater extent, by MSC transwell co-culture. Bioinformatics analysis of RNA sequencing data confirmed a predominant effect of HG on inflammation-related mediators and biological processes/KEGG pathways in RPTEC/TERT1 stable monolayers as well as the non-contact-dependent anti-inflammatory effect of MSC. Finally, CM from HG-exposed RPTEC/MSC transwell co-cultures was associated with attenuated secretion of inflammatory mediators by macrophages compared to CM from HG-stimulated RPTEC alone.

Conclusions

Stable RPTEC monolayers demonstrate delayed pro-inflammatory response to HG that is attenuated by close proximity to human MSC. In DKD, this MSC effect has potential to modulate hyperglycemia-associated RPTEC/macrophage cross-talk.

SUBMITTER: Islam MN

PROVIDER: S-EPMC6862760 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Publications

Human mesenchymal stromal cells broadly modulate high glucose-induced inflammatory responses of renal proximal tubular cell monolayers.

Islam Md Nahidul MN Griffin Tomás P TP Sander Elizabeth E Rocks Stephanie S Qazi Junaid J Cabral Joana J McCaul Jasmin J McMorrow Tara T Griffin Matthew D MD

Stem cell research & therapy 20191119 1

<h4>Background</h4>Renal proximal tubular epithelial cells (RPTEC) are dysfunctional in diabetic kidney disease (DKD). Mesenchymal stromal cells (MSC) may modulate DKD pathogenesis through anti-inflammatory mediators. This study aimed to investigate the pro-inflammatory effect of extended exposure to high glucose (HG) concentration on stable RPTEC monolayers and the influence of MSC on this response.<h4>Methods</h4>Morphologically stable human RPTEC/TERT1 cell monolayers were exposed to 5 mM and ...[more]

PMID: 31744554

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Project description:Epithelial-mesenchymal transition (EMT) occurs in several disease states, including renal fibrosis and carcinogenesis. Myofibroblasts produced from EMT of renal tubular cells are responsible for the deposition of extracellular matrix components in a large portion of renal interstitial fibrosis. Transforming growth factor-beta (TGF-beta) plays an essential role in the EMT of renal tubular cells, but the molecular mechanism governing this process remains largely unknown. In this study, we found that RGC-32 (response gene to complement 32) is critical for TGF-beta-induced EMT of human renal proximal tubular cells (HPTCs). RGC-32 is not normally expressed in the HPTCs. However, TGF-beta stimulation markedly activates RGC-32 while inducing an EMT, as shown by the induction of smooth muscle alpha-actin (alpha-SMA) and extracellular matrix proteins collagen I and fibronectin, as well as the reduction of epithelial marker E-cadherin. TGF-beta function is mediated by several signaling pathways, but RGC-32 expression in HPTCs appears to be mainly regulated by Smad. Functionally, RGC-32 appears to mediate TGF-beta-induced EMT of HPTCs. Blockage of RGC-32 using short hairpin interfering RNA significantly inhibits TGF-beta induction of myofibroblast marker gene alpha-SMA while repressing the expression of E-cadherin. In contrast, overexpression of RGC-32 induces alpha-SMA expression while restoring E-cadherin. RGC-32 also inhibits the expression of another adherens junction protein, N-cadherin, suggesting that RGC-32 alone induces the phenotypic conversion of renal epithelial cells to myofibroblasts. Additional studies show that RGC-32 stimulates the production of extracellular matrix components fibronectin and collagen I. Mechanistically, RGC-32 induces EMT via the activation of other transcription factors such as Snail and Slug. RGC-32 knockdown inhibits the expression of Snail and Slug during TGF-beta-induced EMT. Taken together, our data demonstrate for the first time that RGC-32 plays a critical role in TGF-beta-induced EMT of renal tubular cells.

Dataset Information

Human mesenchymal stromal cells broadly modulate high glucose-induced inflammatory responses of renal proximal tubular cell monolayers.

Background

Methods

Results

Conclusions

Publications

Human mesenchymal stromal cells broadly modulate high glucose-induced inflammatory responses of renal proximal tubular cell monolayers.

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