EGCG Attenuates Renal Damage via Reversing Klotho Hypermethylation in Diabetic db/db Mice and HK-2 Cells

Abstract

To explore whether epigallocatechin-3-gallate (EGCG) improves renal damage in diabetic db/db mice and high-glucose- (HG-) induced injury in HK-2 cells by regulating the level of Klotho gene promoter methylation. Western blotting was used to detect the protein expression levels of DNA methyltransferase 1 (DNMT1), DNMT3a, DNMT3b, transforming growth factor-β1 (TGF-β1), α-smooth muscle actin (α-SMA), and Klotho. The methylation level of the Klotho gene promoter was detected by pyrosequencing. Chromatin immunoprecipitation was used to detect the binding of the Klotho gene promoter to DNMT1 and DNMT3a. The expression of oxidative stress markers (reactive oxygen species (ROS), superoxide dismutase (SOD), malondialdehyde (MDA), catalase (CAT), and 8-hydroxy-2′-deoxyguanosine (8-OHdG)) and inflammatory cytokines (interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α)) in kidney homogenates was also measured using ELISA. Klotho and DNMT3b protein expression was upregulated, while DNMT1, DNMT3a, TGF-β1, and α-SMA protein expression was downregulated after EGCG treatment. EGCG treatment also reduced the methylation level of the Klotho gene promoter as well as the binding of DNMT3a to the Klotho gene promoter. In addition, EGCG treatment significantly decreased the levels of ROS, MDA, 8-OHdG, IL-1β, IL-6, and TNF-α and increased the levels of CAT and SOD. Under HG conditions, EGCG regulated Klotho gene promoter methylation via DNMT3a and decreased the methylation level of the Klotho gene promoter, thereby upregulating the expression of the Klotho protein to exert its protective effect.

Hyperoside Protects HK-2 Cells Against High Glucose-Induced Apoptosis and Inflammation via the miR-499a-5p/NRIP1 Pathway

“…Western blotting was conducted according to a previous study [ 23 ]. The primary antibodies used in the study are listed as follows: anti-Bax (ab32503, 1/1000), anti-Bcl-2 (ab32124, 1/1000), anti-IL-10 (ab133575, 1/2000), anti-IL-6 (ab233706, 1/1000), anti-IL-1β (ab234437, 1/1000), and anti-GAPDH (ab9485, 1/2500).…”

Influence of the Bioactive Diet Components on the Gene Expression Regulation

“…Bioactive Component Disorders References upregulation of DNMT omega-3 fatty acids: DHA, EPA colorectal cancer [25,26] folic acid colorectal cancer, breast cancer [27,28] methionine lung cancer [29] vitamin A congenital heart defects [30] DNA methyltransferase inhibition kaempferol bladder cancer [31] gallic acid lung cancer and oral cancer [32] epigallocatechin-3-gallate breast cancer, diabetic kidney disease [33,34] β-caroten colorectal cancer [35] sulforaphane breast cancer, cardiomyopathy [36,37] omega-3 fatty acids: EPA hepatocarcinoma [38] vitamin A congenital heart defects [30] histone deacetylase inhibition resveratrol breast cancer, renal cell carcinoma, colorectal cancer [39][40][41][42] apigenin prostate cancer, lung cancer [43,44] luteolin lung cancer, leukemia [45,46] chrysin melanoma [47] cinnamic acid derivatives colon and cervical cancer [48] gallic acid prostate cancer, cardiovascular diseases [49] epigallocatechin- Epigallocatechin-3-gallate (EGCG), a polyphenol member, can reduce global DNA methylation levels, DNA methyltransferase (DNMT) activity, messenger RNA (mRNA), and protein levels of DNMT1, DNMT3a, and DNMT3b. EGCG can directly inhibit the DNMT catalytic site.…”

Heme Oxygenase-1 Induction by Cobalt Protoporphyrin Ameliorates Cholestatic Liver Disease in a Xenobiotic-Induced Murine Model

“…To evaluate the effect of CoPP on oxidative stress, we performed immunofluorescent staining for 8-hydroxy-2′-deoxyguanosine (8-OHdG). This molecule is a widely used marker of oxidative-stress-induced DNA damage [ 17 ]. DDC-fed mice exhibited an increase in the number of 8-OHdG-stained cells compared to control mice ( Figure 2 A,B).…”