Excessive iron accumulation mediated oxidative stress and β-cell dysfunction via perturbations of insulin secretion in MIN6 cells

Type 2 diabetes mellitus (T2DM) is the predominant form of diabetes, with prevalence increasing steadily both in developed and developing countries. Defects in both insulin action and insulin secretion are known to contribute to the development of T2DM. Reactive oxygen species (ROS) mediated cellular oxidative damage has been implicated in cellular dysfunction in various organs and disease states. Exposure to high level of glucose during hyperglycaemia has been co-related to ROS generation and oxidative stress in β-cells. In the initial series of experiments MIN6 pseudoislets were cultured under standard conditions and exposed to a range of iron concentrations (20 μM – 100 μM; signifying low to high iron exposure) at predefined glucose levels (5.5 mM and 11 mM) in a static incubation experiment. These series of experiments were categorised into two type of timelines in which experiments were carried. These timelines were at 24 and 48 h and 48 and 72 h. Within the timeline of 24 & 48 h, insulin secretion & content, expression of SNAP-25, mitochondrial oxygen consumption rate (OCR), mitochondrial membrane potential (Δψm), and protein carbonylation were estimated. In addition, antioxidant activity (cellular/non-cellular), cytotoxicity, and cellular ferritin were assessed using the shell of nanocarriers composed of (i) potato protein (PP) - potato dextrin (PD), (ii) PP - modified citrus pectin (MCP) used to formulate Fe and hesperetin in the presence/absence of EGCG. These experiments were followed by assessments of cellular iron content, lipid peroxidation, and cell viability at 48 & 72 h. Our results suggest that presence of 100 μM iron exerted the most detrimental effect on MIN6 β-cell viability and this alteration was consistent with the data from cellular iron uptake analysis, lipid peroxidation, protein carbonyl, OCR, Δψm, SNAP-25, and insulin secretion and its content. All nanocarrier formulations coated antioxidant especially MCP demonstrated higher stability and robustness compared to dextrin control and the ability to resist the inhibitory effect of a potent iron inhibitor highlight its potential as an iron delivery vehicle. These data indicate that excessive iron and glucose accumulation, and consequent oxidative stress results in cellular membrane and mitochondrial damage and disruption. The perturbations in mitochondria functionality correlate with diminishing of MIN6 β-cell insulin secretion, suggesting a mechanistic role for iron overload in the development and progression of type 2 diabetes. These nanoformulations may be potentially used as a model to counter oxidative stress in pancreatic β-cell.

Level dc:type.qualificationlevel

PhD thesis

Grantor dc:publisher.institution

University of Westminster

Year dc:date.issued

2020

Author dc:creator

• Blesia, V.

Identifier

oai:westminsterresearch.westminster.ac.uk:qzvvw

OAI identifier oai:identifier

oai:westminsterresearch.westminster.ac.uk:qzvvw