Vectorial secretion of CTGF as a cell-type specific response to LPA and TGF-β in human tubular epithelial cells

Abstract

BackgroundIncreased expression of the pro-fibrotic protein connective tissue growth factor (CTGF) has been detected in injured kidneys and elevated urinary levels of CTGF are discussed as prognostic marker of chronic kidney disease. There is evidence that epithelial cells lining the renal tubular system contribute to uptake and secretion of CTGF. However, the role of different types of tubular epithelial cells in these processes so far has not been addressed in primary cultures of human cells.ResultsTubular epithelial cells of proximal and distal origin were isolated from human kidneys and cultured as polarized cells in insert wells. The pro-fibrotic stimuli lysophosphatidic acid (LPA) and transforming growth factor β (TGF-β) were used to induce CTGF secretion.LPA activated CTGF secretion in proximal tubular cells when applied from either the apical or the basolateral side as shown by immunocytochemistry. CTGF was secreted exclusively to the apical side. Signaling pathways activated by LPA included MAP kinase and Rho kinase signaling. TGF-β applied from either side also stimulated CTGF secretion primarily to the apical side with little basolateral release.Interestingly, TGF-β activation induced different signaling pathways depending on the side of TGF-β application. Smad signaling was almost exclusively activated from the basolateral side most prominently in cells of distal origin. Only part of these cells also synthesized CTGF indicating that Smad activation alone was not sufficient for CTGF induction. MAP kinases were involved in apical TGF-β-mediated activation of CTGF synthesis in proximal cells and a subset of epithelial cells of distal origin. This subpopulation of distal tubular cells was also able to internalize recombinant apical CTGF, in addition to proximal cells which were the main cells to take up exogenous CTGF.ConclusionsAnalysis of polarized human primary renal epithelial cells in a transwell system shows that vectorial secretion of the pro-fibrotic protein CTGF depends on the cell type, the stimulus and the signaling pathway activated. In all conditions, CTGF was secreted mainly to the apical side upon TGF-β and LPA treatment and therefore, likely contributes to increased urinary CTGF levels in vivo. Moreover, CTGF secreted basolaterally may be active as paracrine pro-fibrotic mediator.

Dual inhibition of Src family kinases and Aurora kinases by SU6656 modulates CTGF (connective tissue growth factor) expression in an ERK-dependent manner

“…The link between ERK and CTGF seems to be dependent on the stimulus and cell type, as it is not observed under all conditions (e.g., Samarakoon et al, 2009). In tubular epithelial cells, however, ERK activation has been shown by us and others as one of the important signaling pathways of CTGF expression (Phanish et al, 2005;Zuehlke et al, 2012). Next we addressed the question whether activation of ERK was a direct consequence of inhibition of Aurora kinases.…”

Connective tissue growth factor induces tubular epithelial to mesenchymal transition through the activation of canonical Wnt signalingin vitro

“…The experiment has revealed that CTGF expression was increased in tubular epithelial cells in multiple forms of CKD. 17,18 However, CTGF, a 36-38-kDa small molecule, can filter across glomerular basement membrane and be present in the tubular fluid in pharmacologically active concentrations. Therefore, tubular epithelial cells not only express and secrete CTGF but are also exposed to CTGF from the glomerulus.…”

The pan-deacetylase inhibitor panobinostat affects angiogenesis in hepatocellular carcinoma models via modulation of CTGF expression

“…Because CTGF can be both secreted and membrane bound, western blot analysis of the cell lysate may not reflect the total protein amount produced when compared to mRNA expression. While in mesangial cells it is bound to the cell surface (49), it has been shown that in hepatocellular carcinoma cells, a fraction of the produced protein remains cell-bound (38). This may explain why our mRNA and protein expression data do not necessarily correlate.…”