Dual modulatory effects of diosmin on calcium oxalate kidney stone formation processes: Crystallization, growth, aggregation, crystal-cell adhesion, internalization into renal tubular cells, and invasion through extracellular matrix

Diosmin is a natural flavone glycoside (bioflavonoid) found in fruits and plants with several pharmacological activities. It has been widely used as a dietary supplement or therapeutic agent in various diseases/disorders. Although recommended, evidence of its protective mechanisms against kidney stone disease (nephrolithiasis/urolithiasis), especially calcium oxalate (CaOx) monohydrate (COM) that is the most common type, remained unclear. In this study, we thus systematically evaluated the effects of diosmin (at 2.5–160 nM) on various stages of kidney stone formation processes, including COM crystallization, crystal growth, aggregation, crystal-cell adhesion, internalization into renal tubular cells and invasion through extracellular matrix (ECM). The results showed that diosmin had dose-dependent modulatory effects on all the mentioned COM kidney stone processes. Diosmin significantly increased COM crystal number and mass during crystallization, but reduced crystal size and growth. While diosmin promoted crystal aggregation, it inhibited crystal-cell adhesion and internalization into renal tubular cells. Finally, diosmin promoted crystal invasion through the ECM. Our data provide evidence demonstrating both inhibiting and promoting effects of diosmin on COM kidney stone formation processes. Based on these dual modulatory activities of diosmin, its anti-urolithiasis role is doubtful and cautions should be made for its use in kidney stone disease

Identification and characterization of ARID1A-interacting proteins in renal tubular cells and their molecular regulation of angiogenesis

Abstract Background Defects and deficiency of AT-rich interactive domain-containing protein 1A (ARID1A) encoded by a tumor suppressor gene ARID1A have recently been suggested to get involved in angiogenesis, a crucial process in carcinogenesis. However, molecular mechanisms of ARID1A deficiency to induce angiogenesis in kidney cancer remain underinvestigated. Methods We performed large-scale identification of ARID1A protein interactors in renal tubular epithelial cells (RTECs) using immunoprecipitation (IP) followed by nanoLC-ESI-LTQ-Orbitrap tandem mass spectrometry (MS/MS). Their roles in angiogenesis were investigated using various assays. Results A total of 74 ARID1A-interacting proteins were identified. Protein–protein interactions analysis revealed that these identified proteins interacted directly or indirectly with ARID1A. Among them, the direct interaction between ARID1A and β-actin was validated by IP and reciprocal IP followed by Western blotting. Small interfering RNA (siRNA) was used for single and double knockdowns of ARID1A and ACTB. Semi-quantitative RT-PCR demonstrated that deficiency of ARID1A, but not ACTB, significantly affected expression of angiogenesis-related genes in RTECs (VEGF and FGF2 were increased, whereas PDGF and EGF were decreased). However, the knockdowns did not affect TGFB1 and FGF1 levels. The quantitative mRNA expression data of VEGF and TGFB1 were consistent with the secreted levels of their protein products as measured by ELISA. Only secreted products derived from ARID1A-deficient RTECs significantly increased endothelial cells (ECs) migration and tube formation. Some of the other carcinogenic features could also be confirmed in the ARID1A-deficient RTECs, including increased cell migration and chemoresistance. Double knockdowns of both ARID1A and ACTB did not enhance the effects of single ARID1A knockdown in all assays. Conclusions We report herein a large dataset of the ARID1A-interacting proteins in RTECs using an IP-MS/MS approach and confirm the direct interaction between ARID1A and β-actin. However, the role of ARID1A deficiency in angiogenesis is independent of β-actin