HIFα independent mechanisms in renal carcinoma cells modulate divergent outcomes in fibronectin assembly mediated by hypoxia and CoCl2

Abstract

Fibronectin (FN) is a core matrix protein that assembles to form a dynamic cellular scaffold, frequently perturbed during oncogenic transformation. Tumor hypoxia, characterized by low oxygen concentrations in the microenvironment of most solid tumors has been shown to accelerate FN assembly in fibroblasts and cancer-associated fibroblasts, cell types that produce abundant amounts of FN protein. Nevertheless, FN matrix regulation in epithelial cancer cells during hypoxia remains less well defined. In this study we investigate the assembly of the FN matrix during hypoxia in renal cancer epithelial cells, the cells of origin of renal cell carcinoma (RCC). We show that hypoxia (1% O2) specifically increases matrix disassembly and increases migratory propensity in renal cancer cells. However, HIFα stabilization using hypoxia mimetics, does not recapitulate the effect of hypoxia on FN matrix reorganization or cell migration. Using a combination of knockdown and inhibitor-based approaches, our work characterizes the signaling events that mediate these two disparate changes on the matrix and explores its functional significance on chemotactic cell migration. Our study systematically reexamines the role of hypoxia mimetics as experimental substitutes for hypoxia and provides new findings on HIFα stabilization and the FN matrix in the context of renal cancer.

Fibronectin assembly regulates lumen formation in breast acini

“…The decrease in FN fibrils in response to hypoxia was further confirmed in the three cell lines by immunolabelling FN ( Figure S1A). In accordance with our observations and from previous studies in renal epithelial cells, [27][28][29][30] we observed a two-fold increase in total FN protein in response to hypoxia indicating that the decrease in FN fibrils ( Figures 1A and S1A) is unlikely to be due to insufficient protein ( Figure 1C,D). Increased levels of HIF-1α and HIF-2α confirmed hypoxia response in these cell lines ( Figures 1E and S1B).…”

“…This reasoning has been corroborated by findings in breast cell lines, demonstrating that although FUD treatment did not alter chemotactic migration of MCF10A cells, FUD decreased the TGF‐β1‐mediated cell migration 43 . Although we and others have previously shown that renal epithelial cells exposed to hypoxia disassemble the FN matrix, perhaps involving cell trafficking of the FN protein, it is still unclear how this process of FN disassembly is initiated or mediated 28 . For this reason, we were unable to target a specific mechanistic process to reverse fibril disassembly.…”

“…However, this data is the first observation investigating real-time migration of the breast progression series under hypoxia. While several studies in other cancer types correlate hypoxia to increased cell migration, 28,48,49 limited but significant data in breast cell lines generated using hypoxia mimetics or alternatively by pre-exposing cells to 1% O 2 followed by migration in normoxia have also pointed to similar conclusions. 50,51 In contrast to much of the previous in vitro migration assays but in agreement with our findings, in vivo imaging of breast cancer cells at single-cell resolution, has revealed a persistent slow migration of hypoxic cells compared to their normoxic counterparts.…”

“…43 Although we and others have previously shown that renal epithelial cells exposed to hypoxia disassemble the FN matrix, perhaps involving cell trafficking of the FN protein, it is still unclear how this process of FN disassembly is initiated or mediated. 28 For this reason, we were unable to target a specific mechanistic process to reverse fibril disassembly. However, this data is the first observation investigating real-time migration of the breast progression series under hypoxia.…”

The biological effect of cobalt chloride mimetic-hypoxia on nucleus pulposus cells and the comparability with physical hypoxia in vitro

“…It has a complex mechanism which is still not clear and may be related to the MAPK signaling pathway, phosphatidylinositol signaling pathway and cytokine-cytokine receptor pathway [40,41]. Many studies have shown that cells migrate faster under hypoxia than normoxia, including mesenchymal stem cells and carcinoma cells [42][43][44]. Magdaleno et al [43] compared the migration of renal carcinoma cells under hypoxia and CoCl 2 and found that physical hypoxia promoted migration but CoCl 2 failed to achieve similar effect.…”