Metformin inhibits TGF-beta 1-induced MCP-1 expression through BAMBI-mediated suppression of MEK/ERK1/2 signalling

Nephrology (Carlton). 2019 Apr;24(4):481-488. doi: 10.1111/nep.13430.

Abstract

Aims: Metformin is a biguanide derivative widely used for the treatment of type 2 diabetes mellitus. Recent evidence demonstrates that this anti-hyperglycaemic drug exerts renal protective effects, yet the mechanisms remain poorly understood. monocyte chemoattractant protein 1 (MCP-1) has been recognized as a key mediator of renal fibrosis in chronic kidney diseases, including diabetic nephropathy. This study aimed to investigate the effects of metformin on transforming growth factor beta 1 (TGF-β1)-induced MCP-1 expression and the underlying mechanisms in rat renal tubular epithelial cells.

Methods: Rat renal tubular epithelial cell line NRK-52E cells were stimulated with TGF-β1 and/or metformin. The messenger RNA (mRNA) of MCP-1 and bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) was evaluated by real-time quantitative polymerase chain reaction. MCP-1 protein was measured by enzyme linked immunosorbent assay (ELISA). Total and phosphorylated extracellular signal-regulated kinases 1/2 (ERK1/2) was evaluated by western blot. Down- and upregulation of BAMBI were achieved by RNA interference targeting BAMBI and lentiviral vector-mediated overexpression of the BAMBI gene, respectively. Cell viability was analysed using Cell Counting Kit 8 (CCK-8) reagents.

Results: Stimulation with TGF-β1 resulted in the increased expression of MCP-1 and decreased expression of BAMBI in NRK-52E cells. Metformin inhibited the expression of MCP-1 in NRK-52E cells. Pretreatment with metformin suppressed upregulation of MCP-1 and downregulation of BAMBI, as well as phosphorylation of ERK1/2 induced by TGF-β1. U0126, a specific inhibitor for mitogen-activated and extracellular signal-regulated kinase kinases 1/2 (MEK-1/2), completely blocked TGF-β1-induced MCP-1 expression. Knockdown of the BAMBI gene promoted phosphorylation of ERK1/2 and TGF-β1-induced expression of MCP-1. Overexpression of BAMBI inhibited phosphorylation of ERK1/2 and TGF-β1-induced upregulation of MCP-1.

Conclusion: In rat renal tubular epithelial cells, metformin prevents TGF-β1-induced MCP-1 expression, in which BAMBI-mediated inhibition of MEK/ERK1/2 might be involved.

Keywords: BAMBI; MCP-1; TGF-β1; metformin; renal tubular epithelial cell.

MeSH terms

  • Animals
  • Cell Line
  • Chemokine CCL2 / genetics
  • Chemokine CCL2 / metabolism*
  • Epithelial Cells / drug effects*
  • Epithelial Cells / enzymology
  • Epithelial Cells / pathology
  • Extracellular Signal-Regulated MAP Kinases / metabolism*
  • Fibrosis
  • Kidney Tubules / drug effects*
  • Kidney Tubules / enzymology
  • Kidney Tubules / pathology
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Metformin / pharmacology*
  • Mitogen-Activated Protein Kinase Kinases / metabolism*
  • Phosphorylation
  • Rats
  • Signal Transduction / drug effects
  • Transforming Growth Factor beta1 / pharmacology*

Substances

  • Bambi protein, rat
  • Ccl2 protein, rat
  • Chemokine CCL2
  • Membrane Proteins
  • Transforming Growth Factor beta1
  • Metformin
  • Extracellular Signal-Regulated MAP Kinases
  • Mitogen-Activated Protein Kinase Kinases