title: Limiting Hyperinsulinaemia To Prevent Type 2 Diabetes
Creators: name: Shamoon, Muhammad
Publisher: name: The Australian National University
Year: 2024

description: The rising prevalence of type 2 diabetes (T2D) is a major concern globally. This thesis focuses on severe insulin resistant subtype of T2D (SIR-T2D) in which hyperinsulinaemia and obesity, accompanied by insulin resistance, occur prior to onset of diabetes. We hypothesised that interventions aimed at curtailing the development of hyperinsulinaemia would prevent SIR-T2D. Main aims were to determine in a mouse model of SIR-T2D if (i) intermittent fasting (IF) and (ii) deficiency of the system B(0) neutral amino acid transporter AT1 (B0AT1, Slc6a19 gene) could prevent hyperinsulinaemia and T2D. NODk mice, derived from non-obese diabetic (NOD) mouse model of type 1 diabetes (T1D) but protected from T1D due to congenic replacement of Idd1-H2g7 MHC II locus with an autoimmune diabetes low risk Idd1-H2k MHC II locus, were used. Male NODk mice fed a Western diet (WD) develop a SIR-T2D like phenotype characterised by excessive weight gain, hyperinsulinaemia, dyslipidaemia, and severe hyperglycaemia. To investigate IF, male NODk mice were fed chow diet (CD), WD, or WD with 15 h of fasting twice per week (WD+IF) from 6 to10 weeks (short-term (ST) protocol) and 6 to 30 weeks (long-term (LT) protocol) of age. Metabolic characteristics of mice were serially assessed, including i.p.GTT after 3 and 17 weeks on diet, histological examination of pancreases and livers, and the incidence of diabetes by 30 weeks of age. To investigate B0AT1 deficiency, male NODk.Slc6a19 wild type (WT) and KO mice were fed CD or WD from 8 weeks of age for 5 days-WD challenge (WDC protocol), and CD or WD in ST and LT protocols as for the IF studies. Additional readouts included urine amino acid profiles, and on day 4 of WDC protocol, i.p.GTT and oral GTT testing. IF attenuated WD-induced whole body and fat depot weight gains, hyperinsulinaemia and hypertriglyceridaemia, improved glucose tolerance, and prevented SIR-T2D. WD resulted in increased pancreatic fat deposits, enlarged islet size, frequent pseudo-rosette structured islets, presence of intracytoplasmic inclusion bodies in endocrine cells, with some apoptotic endocrine cells. IF lessened islet enlargement and prevented pseudo-rosette formation. Peri-islet and peri-ductal inflammation were present in both WD and WD-IF mice pancreases. Although WD feeding did not elevate serum transaminases, marked hepatic steatosis, hepatocyte ballooning and mild to moderate hepatic lobular inflammation were found, partially mitigated by IF. Finally, with WD-feeding there was not much evidence of renal function impairment. The phenotype of the CD-fed NODk.Slc6a19 KO compared to WT mice included marked aminoaciduria (confirming B0AT1 deficiency), normal body weight, behaviour and appearance at 6 weeks of age, mildly reduced body and fat depot weights at 30 weeks of age, normal glucose tolerance, reduced basal and glucose stimulated insulinaemia, higher basal and glucose-stimulated GLP-1 concentrations and markedly increased plasma FGF-21 concentrations. As for IF, B0AT1 deficiency attenuated WD-induced body and fat depot weight gains, hyperinsulinaemia and hypertriglyceridaemia, improved glucose tolerance, and prevented SIR-T2D. GLP-1 and FGF-21 concentrations remained higher in the KO-WD compared to WT-WD mice, and curtailed WD-induced falls in serum adiponectin. CD-fed KO mice had normal pancreas and liver histological appearances. In WD-fed KO compared to WT mice pancreases, fat deposition, inflammation, pseudo-rosette islet structures and endocrine cell apoptosis were less evident. WD-induced changes in liver were not different between two genotypes. Finally, KO mice had heavier kidneys, but renal function was normal in both genotypes and not adversely affected by WD-feeding. In conclusion, both IF and B0AT1 deficiency attenuated WD-induced hyperinsulinaemia and completely prevented diabetes in WD-fed male NODk mice. IF and pharmacological targeting of B0AT1 are promising interventions for the prevention of SIR-T2D.