RESVERATROL EFFECT ON RENAL OSMOPROTECTION: MODULATION OF COX-2 EXPRESSION

ERJAVEC, LUCIANA; CASALI, CECILIA; PARRA, LEANDRO; RECABARREN, MANUEL; FERNÁNDEZ TOME, MARIA DEL CARMEN

Mendoza

Congreso; LVIII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research (SAIB); 2022

SAIB

Resveratrol (RSV) is a polyphenol naturally present in several plants. Nowadays it is sold as anover-the-counter dietary supplement due to its antioxidant, anti-inflammatory and antitumoral effects.Paradoxically, it was documented that RSV may also present pro-oxidizing and pro-proliferative effects.Likewise, RSV effects on renal tissue are still controversial, as some injury models described beneficialeffects, while others observed nephrotoxicity. Many molecular targets were described to explain RSVeffects, one of them is ciclooxigenase 2 (COX2). Most of the articles exploring RSV effect on COX2 usedcancer or inflammation models and found that it inhibited COX2 expression or activity. In the kidney, thisprotein plays an important role in osmoprotection of tubular cells subjected to physiologicalhyperosmolarity. We demonstrated that renal epithelial cell line MDCK undergoes an adaptive processduring the first 24h of hyperosmolarity, in which the transcription of the osmoprotective gene COX2 isactivated, among others. After 48h, cells are already adapted and begin to differentiate, acquiring apolarized morphology. Considering that many articles found that RSV can inhibit COX2, in this work weevaluate RSV effect on adaptation and differentiation mechanisms, focusing particularly on COX2 role. Todo this, MDCK cells were pretreated with different concentrations of RSV (1-25 μM) and cultured inhyperosmolar media (~512 mOsm/kg H2O) for 24 and 48h. Cells were harvested to obtain cell number andviability. Cell cycle, immunofluorescence (IF), western blot and RT-PCR analysis were performed. We foundthat RSV decreased cell number in a concentration-dependent manner at 24 and 48h. Cell cycle analysisrevealed that RSV increased S-phase and Sub-G0 population. Additionally, treated cells did not reachtypical epithelium morphology. COX2 activity was not increased by RSV, but mRNA and protein levels weresurprisingly upregulated, and IF revealed protein accumulation in cytoplasmic granules. To investigate thepathways leading to this increase, we evaluated TonEBP, NF-κB and MAPKs pathways, which are activatedby hyperosmolarity; and SIRT1 implication, a target of RSV. TonEBP target genes mRNA did not showsignificant changes under RSV treatment, while NF-κB target gene mRNA showed an increase similar tothat of COX2 mRNA. NF-κB IF revealed an increase in its nuclear localization. Regarding MAPKs, the onlytreatment that blocked COX2 protein expression was ERK1/2 selective inhibitor. SIRT1 specific inhibitiondid not change COX2 expression. These results show that RSV pretreatment caused harmful effects inhyperosmolar cultures of MDCK cells, affecting cell number and cell cycle and impeding cells to reachdifferentiated phenotype; in contrast with our hypothesis, Rsv did not decrease COX2 expression butsignificantly upregulated the protein which was not active. Herein we report a possible nephrotoxic effectof Rsv.