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TGF-beta- and angiotensin-II-induced mesangial matrix protein secretion is mediated by protein kinase C
Background:
Glomerulonephritis is characterized by the accumulation of extracellular matrix protein within the glomerulus. This process, when allowed to proceed unimpeded, leads to glomerulosclerosis and eventually to cessation of glomerular filtration. There is evidence that protein kinase C (PKC) activation plays an important role in mediating at least some of the effects of TGF-beta in vascular smooth-muscle cells. The current study was undertaken to determine whether PKC activity is required for both TGF-beta and angiotensin II (Ang II) to induce mesangial cell matrix protein secretion.
Methods:
PKC was inhibited by two separate methods, and [3H]thymidine incorporation was assessed in both the presence and the absence of PKC inhibition. Conditioned medium from cells stimulated with TGF-beta or Ang II was collected and analysed for secreted matrix proteins and sulphated proteins by SDS-polyacrylamide gel electrophoresis and western blotting.
Results:
Twenty-four-hour incubation of rat mesangial cells with phorbol-12-myristate-13-acetate (PMA) reduced total PKC activity to basal levels. Both TGF-beta and Ang II were mitogenic in mesangial cells, and chronic PMA pre-incubation inhibited this DNA synthesis. TGF-beta-and Ang-II-induced sulphated protein secretion into conditioned medium was markedly attenuated in PKC-downregulated cells. Secretion of the specific matrix proteins laminin and fibronectin by mesangial cells stimulated with either TGF-beta or Ang II was also diminished in PKC-downregulated cells and in cells pre-incubated with the specific PKC inhibitor, chelerythrine. There was no evidence of generalized cell toxicity or decreased non-specific protein synthesis caused by these PKC inhibitors.
Conclusions:
PKC is a key intermediary in the process by which TGF-beta and Ang II cause DNA synthesis and mesangial cell matrix protein production. Thus, PKC inhibitors deserve further study as potential therapeutic agents for a variety of glomerular diseases.
Insights
Protein kinase C (PKC) is crucial for TGF-beta and Angiotensin II (Ang II) to stimulate mesangial cell matrix production. Inhibiting PKC may offer a therapeutic strategy for glomerular diseases.
Area of Science:
- Nephrology
- Cell Biology
- Biochemistry
Background:
- Glomerulonephritis involves extracellular matrix accumulation, leading to glomerulosclerosis and kidney failure.
- Protein kinase C (PKC) activation is implicated in mediating transforming growth factor-beta (TGF-beta) effects.
- The study investigates PKC's role in TGF-beta and angiotensin II (Ang II)-induced mesangial cell matrix secretion.
Purpose of the Study:
- To determine if PKC activity is essential for TGF-beta and Ang II to induce matrix protein secretion in mesangial cells.
- To explore the potential of PKC inhibitors as therapeutic agents for glomerular diseases.
Main Methods:
- PKC was inhibited using two methods, and [3H]thymidine incorporation was measured.
- Secreted matrix and sulphated proteins were analyzed via SDS-PAGE and Western blotting after TGF-beta or Ang II stimulation.
Main Results:
- PKC downregulation by phorbol-12-myristate-13-acetate (PMA) inhibited TGF-beta and Ang II-induced DNA synthesis.
- TGF-beta and Ang II-induced sulphated protein secretion was significantly reduced in PKC-downregulated cells.
- Secretion of laminin and fibronectin was diminished in PKC-downregulated cells and with chelerythrine treatment, with no observed toxicity.
Conclusions:
- PKC acts as a key mediator for TGF-beta and Ang II in driving DNA synthesis and matrix protein production in mesangial cells.
- PKC inhibitors warrant further investigation as potential treatments for various glomerular diseases.