Diabetic Nephropathy Treatment by Zn Supplementation in a Murine Model of Type 1 Diabetes Mellitus: Potential Role of Nrf2 Signaling Pathway

Objective  To evaluate the renoprotective effects of zinc (Zn) supplementation in diabetes kidney disease (DKD) and to explore its impact on the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway.

Methods  A total of 12 male OVE26 mice (spontaneous type 1 diabetes mellitus mice) aged 3 months and weighing approximately 24-27 g were selected and randomly assigned to a diabetes mellitus (DM) group and a zinc-treated DM (DM/Zn) group (n = 6 each). In addition, 12 age-matched male FVB mice weighing approximately 27-30 g were selected and randomly assigned to a non-diabetic control (Ctrl) group and a zinc-treated (Zn) group (n = 6 each). Mice in the DM/Zn and Zn groups were given zinc supplementation for 3 months, with each mouse receiving 5 mg/kg of zinc sulfate by gavage every other day. Mice in the DM and Ctrl groups were given the same volume of normal saline. At the end of the experiment, the albumin-to-creatinine ratio (ACR) in urine was used as an indicator to evaluate renal function. Sirius red staining was performed to assess renal fibrosis in each group of mice. Western blotting was performed to determine the expression of fibrotic growth factors, including connective tissue growth factor (CTGF) and transforming growth factor-β1 (TGF-β1), in renal tissue, and the protein expression of Nrf2, an antioxidant substance, and the protein expression levels of its downstream targets, including NAD(P)H quinone dehydrogenase 1 (NQO1), heme oxygenase 1 (HO-1), superoxide dismutase (SOD)-1, SOD-2, and catalase (CAT).

Results 1) Compared to the Ctrl group, the urinary protein secretion levels of mice in the DM group exhibited progressive increase. After 3 months of zinc supplementation treatment, the urinary protein secretion levels of mice in the DM/Zn group decreased Compared to that of mice in the DM group, and the difference was statistically significant (P < 0.05). 2) Compared to that in the Ctrl group, the collagen deposition in the renal tissues of mice in the DM group increased, and the difference was statistically significant (P < 0.05), while no obvious change was observed in mice in the DM/Zn group. Compared to the Ctrl group, mice in the DM group exhibited increased expression levels of CTGF and TGF-β1 in the renal tissues, but the expression levels decreased after zinc supplementation treatment, with the differences being statistically significant (P < 0.05). 3) Compared to that of the Ctrl group, the expression level of Nrf2 in the renal tissues of mice in the Zn and DM groups increased, and the level of Nrf2 in the renal tissues of mice in the DM/Zn group showed a further increase, with the differences being statistically significant (P < 0.05). 4) Compared to those of the Ctrl group, the protein expression levels of Nrf2 downstream target genes, including NQO1 and HO-1, in the renal tissues of mice in the Zn group increased, and the levels of NQO1 and HO-1 in the renal tissues of mice in the DM/Zn group showed a further increase, with the differences being statistically significant (P < 0.05). Compared to those of the mice in the Ctrl group, the protein expressions of Nrf2 downstream target genes, including SOD-1, SOD-2, and CAT of in the renal tissues of the mice in the Zn group increased, while the expression levels of SOD-1, SOD-2, and CAT in the renal tissues of the mice in the DM group decreased, with the differences being statistically significant (P < 0.05). Zn supplementation could completely inhibit these changes (P < 0.05).

Conclusions Zn supplementation has therapeutic effects on DKD and mitigates T1DM-induced renal dysfunction and oxidative injury in mice, which may be associated with the activation of the Nrf2 antioxidant signaling pathway.