Additional file 1: of Naringin ameliorates the high glucose-induced rat mesangial cell inflammatory reaction by modulating the NLRP3 Inflammasome

Key summary points. Diabetic kidney disease (DKD) is one of the most serious chronic complications of diabetes mellitus (DM), is a strong risk factor for cardiovascular diseases, and is a major cause of end stage kidney disease. The pathogenesis of DKD is complex and there are no effective measures to treat it currently.The aim of this study was to investigate the expression of the NLRP3-inflammasome under high glucose conditions, the effects of naringin during these conditions, and elucidate the role of naringin in the pathogenesis of DKD. Our results confirmed that naringin can regulate the NLRP3-Caspase-1-IL-1β / IL-18 signaling pathway by the NLRP3 inflammasome, which can improve DKD by playing an anti-inflammatory role.This study provides new insights into the nephroprotective mechanism of naringin to improve DKD by anti-inflammatory responses. (DOCX 12 kb

The chemical structure of naringin.

<p>The molecular formula of naringin is C27H32O14 and the molecular weight is 580.53.</p

Naringin inhibited high glucose-induced proliferation in HBZY-1 cells.

<p>The proliferation of HBZY-1 cells was determined by MTT assay. Data were expressed as means ± SD, n = 5.^a-f Means with different superscripts are significantly different (P< 0.05).</p

Naringin mitigated changes of pathomorphology and kidney injury biochemical indexes.

<p>(A) The pathological changes of renal tissues were investigated by PAS staining(400×). The glomerular injury and tubular injury scores were shown. (B) Body weight and percentage change in weight of rats. (C) Food intake per rat per day. (D) The ratios of kidney weight/ body weight were calculated. The concentrations of BUN (E), Cr(F), and UP(G) were detected. Results represent three independent experiments. Data were expressed as means ± SD, n = 6.^a-d Means with different superscripts are significantly different (P< 0.05).</p

Naringin restrained oxidative stress injury by activating Nrf2 antioxidant pathway.

<p>The concentration of MDA (A), the activity of SOD(B) and GSH-Px(C) in kidney tissues were detected. Data were expressed as means ± SD, n = 6. (D) The ROS production in HBZY-1 cells was evaluated by flow cytometry and ROS generation rates were shown. Data were expressed as means ± SD, n = 3. (E) The protein levels of Nrf2 and HO-1 in kidney tissues or HBZY-1 cells were detected by western blot. Results represent three independent experiments. (F) The DNA binding activities of Nrf2 in kidney tissues were assessed by EMSA assay. Data were expressed as means ± SD, n = 5. (G) The HO-1 activities in kidney tissues were determined. Data were expressed as means ± SD, n = 6.^a-d Means with different superscripts are significantly different (P< 0.05).</p

Naringin suppressed NF-κ B signaling pathway activation.

<p>(A) The protein levels of p-I κ B α, I κ B α and NF-κ B in kidney tissues and HBZY-1 cells were determined by western blot. Results represent three independent experiments. (B) The distribution change of NF-κ B in HBZY-1 cells was observed by immunofluorescence assay. Results represent three independent experiments. (C) The DNA binding activities of NF-κ B in kidney tissues were assessed by EMSA assay. Data were expressed as means ± SD, n = 5.^a-c Means with different superscripts are significantly different (P< 0.05).</p

Naringin inhibited collagen production and renal interstitial fibrosis.

<p>(A) The collagen deposition and renal interstitial fibrosis was observed by Masson’s staining(200×). The percentages of fibrosis area were shown. (B) The expression of collagen I in kidney tissues was determined by immunohistochemical staining(400×). (C) The expressions of collagen I, MMP-2, TIMP-1 and TGF-β1 in kidney tissues were detected by western blot. Results represent three independent experiments. Data were expressed as means ± SD, n = 6.^a-c Means with different superscripts are significantly different (P< 0.05).</p