Epigallocatechin-3-gallate plays more predominant roles than caffeine for inducing actin-crosslinking, ubiquitin/proteasome activity and glycolysis, and suppressing angiogenesis features of human endothelial cells

Gallemit, Pauline Edenn Mendoza; Yoodee, Sunisa; Malaitad, Thanyalak; Thongboonkerd, Visith

doi:10.1016/j.biopha.2021.111837

Cited by 14 publications

(8 citation statements)

References 71 publications

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“…The cells were maintained and treated as aforementioned. After 24-h incubation with or without secretome derived from control or COM-treated HK-2 or MDCK cells, BHK-21 cells were subjected to F-actin and immunofluorescence stainings as described previously [ 33 , 34 ]. Briefly, the cells were fixed with 4% paraformaldehyde at 25 °C for 15 min followed by permeabilization with 0.1% Triton X-100 in PBS at 25 °C for 15 min.…”

Section: Methodsmentioning

confidence: 99%

Calcium oxalate crystal-induced secretome derived from proximal tubular cells, not that from distal tubular cells, induces renal fibroblast activation

2023

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Background Kidney stone disease (KSD) is commonly accompanied with renal fibrosis, characterized by accumulation and reorganization of extracellular matrix (ECM). During fibrogenesis, resident renal fibroblasts are activated to become myofibroblasts that actively produce ECM. However, such fibroblast–myofibroblast differentiation in KSD remained unclear. Our present study thus examined effects of secreted products (secretome) derived from proximal (HK-2) vs. distal (MDCK) renal tubular cells exposed to calcium oxalate monohydrate (COM) crystals on activation of renal fibroblasts (BHK-21). Methods HK-2 and MDCK cells were treated with 100 µg/ml COM crystals under serum-free condition for 16 h. In parallel, the cells maintained in serum-free medium without COM treatment served as the control. Secretome derived from culture supernatant of each sample was mixed (1:1) with fresh serum-free medium and then used for BHK-21 culture for another 24 h. Results Analyses revealed that COM-treated-HK-2 secretome significantly induced proliferation, caused morphological changes, increased spindle index, and upregulated fibroblast-activation markers (F-actin, α-SMA and fibronectin) in BHK-21 cells. However, COM-treated-MDCK secretome had no significant effects on these BHK-21 parameters. Moreover, level of transforming growth factor-β1 (TGF-β1), a profibrotic factor, significantly increased in the COM-treated-HK-2 secretome but not in the COM-treated-MDCK secretome. Conclusions These data indicate, for the first time, that proximal and distal tubular epithelial cells exposed to COM crystals send different messages to resident renal fibroblasts. Only the secretome derived from proximal tubular cells, not that from the distal cells, induces renal fibroblast activation after their exposure to COM crystals. Such differential effects are partly due to TGF-β1 secretion, which is induced by COM crystals only in proximal tubular cells.

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Section: Methodsmentioning

confidence: 99%

Calcium oxalate crystal-induced secretome derived from proximal tubular cells, not that from distal tubular cells, induces renal fibroblast activation

2023

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“…Transwell migration assay was performed as described previously [ 29 , 30 ] to examine the migratory response of EA.hy926 ECs to angiogenic factors secreted from MDCK renal cells. Prior to cell seeding, the membrane insert (8-µm-pore size) of the Transwell plate (0.33 cm 2 culture area/well) (Corning Costar) was pre-coated with matrigel (BD Biosciences) and incubated at 37 °C for 1 h. EA.hy926 cells (1 × 10 5 in 200 µl serum-free DMEM) were seeded onto the pre-coated membrane insert in upper chamber of each well.…”

Section: Methodsmentioning

confidence: 99%

“…ECs tube formation assay [ 29 , 30 ] was performed to evaluate the ability of EA.hy926 ECs to form capillary/mesh-like tubes on basement membrane matrix in response to the angiogenic factors secreted from MDCK renal cells. Prior to cell seeding, each well of the 96-well plate (Corning Costar) was pre-coated with 50 μl matrigel (BD Biosciences) at 37 °C for 1 h. EA.hy926 cells (5 × 10 4 cells/well in 100 μl CM harvested from the siRNA-transfected MDCK cells) were seeded into the pre-coated well and incubated at 37 °C with 5% CO 2 for 24 h. Thereafter, the capillary/mesh-like tubes were imaged by using Nikon Eclipse Ti-S inverted phase-contrast light microscope (Nikon).…”

Section: Methodsmentioning

confidence: 99%

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

Yoodee,

Peerapen,

Plumworasawat

et al. 2023

J Transl Med

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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.

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“…After 24-h incubation with or without 100 µM caffeine, the cells were washed with PBS and then extracted by 100 µl ATP extraction buffer (25 mM Tricine, 100 μM EDTA, 1 mM DTT, and 1% Triton X-100). After centrifugation at 1000 × g at 4 °C for 5 min, the supernatant (extracted intracellular compartment) was collected for ATP measurement using the luminescence-based protocol [25] , [26] . The intracellular ATP level in each sample was determined from the standard curve, normalized by protein amount, and then reported as pmol/mg protein unit.…”

Section: Methodsmentioning

confidence: 99%

Caffeine causes cell cycle arrest at G0/G1 and increases of ubiquitinated proteins, ATP and mitochondrial membrane potential in renal cells

Kanlaya,

Subkod,

Nanthawuttiphan

et al. 2023

Computational and Structural Biotechnology Journal

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Epigallocatechin-3-gallate plays more predominant roles than caffeine for inducing actin-crosslinking, ubiquitin/proteasome activity and glycolysis, and suppressing angiogenesis features of human endothelial cells

Cited by 14 publications

References 71 publications

Calcium oxalate crystal-induced secretome derived from proximal tubular cells, not that from distal tubular cells, induces renal fibroblast activation

Calcium oxalate crystal-induced secretome derived from proximal tubular cells, not that from distal tubular cells, induces renal fibroblast activation

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

Caffeine causes cell cycle arrest at G0/G1 and increases of ubiquitinated proteins, ATP and mitochondrial membrane potential in renal cells

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