Shear Stress-Induced Alteration of Epithelial Organization in Human Renal Tubular Cells.

Damien Maggiorani,Audrey Casemayou,Jean-Sébastien Saulnier-Blache,Sandra Grès,Romain Dissard,Marcy Belloy,Julie Bellière,Joost P Schanstra,Bénédicte Buffin-Meyer,Laure Ducasse,Cécile Caubet,Jean-Loup Bascands,Shree Ram Singh

doi:10.1371/journal.pone.0131416

Damien Maggiorani, Audrey Casemayou + Show 11 more

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https://doi.org/10.1371/journal.pone.0131416

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Abstract

Tubular epithelial cells in the kidney are continuously exposed to urinary fluid shear stress (FSS) generated by urine movement and recent in vitro studies suggest that changes of FSS could contribute to kidney injury. However it is unclear whether FSS alters the epithelial characteristics of the renal tubule. Here, we evaluated in vitro and in vivo the influence of FSS on epithelial characteristics of renal proximal tubular cells taking the organization of junctional complexes and the presence of the primary cilium as markers of epithelial phenotype. Human tubular cells (HK-2) were subjected to FSS (0.5 Pa) for 48h. Control cells were maintained under static conditions. Markers of tight junctions (Claudin-2, ZO-1), Par polarity complex (Pard6), adherens junctions (E-Cadherin, β-Catenin) and the primary cilium (α-acetylated Tubulin) were analysed by quantitative PCR, Western blot or immunocytochemistry. In response to FSS, Claudin-2 disappeared and ZO-1 displayed punctuated and discontinuous staining in the plasma membrane. Expression of Pard6 was also decreased. Moreover, E-Cadherin abundance was decreased, while its major repressors Snail1 and Snail2 were overexpressed, and β-Catenin staining was disrupted along the cell periphery. Finally, FSS subjected-cells exhibited disappeared primary cilium. Results were confirmed in vivo in a uninephrectomy (8 months) mouse model where increased FSS induced by adaptive hyperfiltration in remnant kidney was accompanied by both decreased epithelial gene expression including ZO-1, E-cadherin and β-Catenin and disappearance of tubular cilia. In conclusion, these results show that proximal tubular cells lose an important number of their epithelial characteristics after long term exposure to FSS both in vitro and in vivo. Thus, the changes in urinary FSS associated with nephropathies should be considered as potential insults for tubular cells leading to disorganization of the tubular epithelium.

Highlights

Urinary fluid shear stress (FSS) is the friction force resulting from movement of urine on the surface of renal tubular cells
In static conditions (FSS 0), Claudin-2 was detected in both plasma membrane and cytoplasm while ZO-1 was only localized at the plasma membrane majorly as a continuous belt (Fig 1A)
FSS caused the disappearance of Claudin-2 and changes the staining of ZO-1, which remained at the plasma membrane but becomes punctuated and discontinuous (Fig 1A)

Summary

Introduction

Urinary fluid shear stress (FSS) is the friction force resulting from movement of urine on the surface of renal tubular cells. It depends on the urine flow rate and viscosity as well as on the diameter of the renal tubule. FSS-injured cells secrete mediators that stimulate adhesion of monocytes to endothelial cells and their differentiation into inflammatory macrophages [14, 15] suggesting that FSS acts in vivo as a promoter of renal inflammation This combined body of evidence suggests that changes in urinary FSS potentially represent an early aggression for renal tubule cells, thereby playing a role in the progression of CKD [6]

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