Metformin alleviates neuronal and renal related stress signals in diabetic C57BL/6 mice.
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Date
2019
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Abstract
In recent years, diabetes has become more prevalent due to modern, demanding, and sedentary lifestyle patterns. This disease is characterised by insulin resistance and associated molecular complications. Metformin (MF) is a popular antidiabetic agent that has effects beyond glycaemic control such as regulation of metabolic molecular pathways. However, the exact mechanisms against hyperglycaemic induced end organ damage remain elusive. This study aimed to investigate the protective effects of MF in the brain and kidney in vivo, by exploring dysregulated pathways related to mitochondrial function, oxidative stress, ER stress, inflammation, and apoptosis.
This study established a diabetic mouse (C57BL/6) model (Ethics no: AREC/057/016) through intraperitoneal multiple low-dose STZ (50 mg/kg BW) injections (10 days). Blood sugar levels of 7-16mmol/L were considered diabetic, and the 15 day treatment period (MF, 20 mg/kg BW per day, oral gavage) was inducted thereafter. Fasting (12 hr) plasma OGTT revealed MF significantly lowered blood glucose levels in diabetic mice. All mice experiments were performed by Dr. N. Naicker. Post-sacrifice (isoflurane), the investigator (author) of the work in the presented in this thesis assisted in harvesting Whole brain and kidney tissue and performed all downstream protein and mRNA analyses. Diabetic mice exhibited heightened oxidative stress by protein carbonylation, and diminished antioxidant responses in both the brain and kidney compared to normoglycaemic mice. Metformin significantly reduced protein carbonylation, increased GSTA4 expression in the brain; and Nrf2 and GPx mRNA levels in the kidney, alleviating oxidative stress. Further, MF improved mt activity, and decreased the HIF-1 expression in the kidney through upregulation of AMPK, and Sirt1 expression. In addition, MF induced epigenetic changes in mice brain through miR-148a repression and concomitant increases in PGC-1α, Sirt1, and Sirt3 protein and gene expressions, thus regulating mt biogenesis. Mitochondrial chaperone proteins HSP60, HSP70 and LonP1 in diabetic mice brain were upregulated through a MF-induced miR-132 repression mechanism. Regulation of the UPR by PERK-eIF2α inhibition after MF-treatment attenuated ER stress in diabetic mice brain and kidney tissue. Moreover, renal injury associated with diabetes was attenuated by MF through decreased CHOP expression, downstream to ER stress. This finding was supplemented by inhibition of Bax, cyt-c, and ultimately the intrinsic apoptotic pathway.
MiR-141 modulates expression of PP2A, a phosphoesterase that regulates phosphorylation of tau protein. In hyperglycaemic mice there was increased miR-141 expression with concomitant PP2A downregulation in the brain. Treatment with MF exerted epigenetic regulation by downregulating miR-141 expression, concomitantly increasing PP2A and subsequent downregulation of tau protein phosphorylation at Ser396. Additionally, MF inhibited proinflammatory NLRP3
inflammasome and related components by regulation of the PP2A/ NF-κB cascade. Neuroplasticity was increased by increased BDNF overexpression by MF in diabetic mice.
Herein we show that MF exerts protective mechanistic effects in the brain and kidney over an acute experimental period. We highlight that anti-oxidant and Sirt1 modulation are at the forefront of renal cell defence to metabolic stress. Neuroinflammatory and epigenetic therapeutic targets of MF are revealed through miRNA regulatory mechanisms, integrating the mechanisms of diabetic neuronal and renal damage.
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Doctoral Degree. University of KwaZulu-Natal, Durban.