Nkx6.1 decline accompanies mitochondrial DNA reduction but subtle nucleoid size decrease in pancreatic islet β-cells of diabetic Goto Kakizaki rats (original) (raw)
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Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2010
3D morphology of mitochondrial network 4Pi microscopy 3D image analysis Pancreatic β-cell Type-2 diabetes Morphological diagnostic Insulin production in pancreatic β-cells is critically linked to mitochondrial oxidative phosphorylation. Increased ATP production triggered by blood glucose represents the β-cells' glucose sensor. Type-2 diabetes mellitus results from insulin resistance in peripheral tissues and impaired insulin secretion. Pathology of diabetic β-cells might be reflected by the altered morphology of mitochondrial network. Its characterization is however hampered by the complexity and density of the three-dimensional (3D) mitochondrial tubular networks in these cell types. Conventional confocal microscopy does not provide sufficient axial resolution to reveal the required details; electron tomography reconstruction of these dense networks is still difficult and time consuming. However, mitochondrial network morphology in fixed cells can also be studied by 4Pi microscopy, a laser scanning microscopy technique which provides an ∼ 7-fold improved axial resolution (∼ 100 nm) over conventional confocal microscopy. Here we present a quantitative study of these networks in insulinoma INS-1E cells and primary β-cells in Langerhans islets. The former were a stably-transfected cell line while the latter were transfected with lentivirus, both expressing mitochondrial matrix targeted redoxsensitive GFP. The mitochondrial networks and their partial disintegration and fragmentation are revealed by carefully created iso-surface plots and their quantitative analysis. We demonstrate that β-cells within the Langerhans islets from diabetic Goto Kakizaki rats exhibited a more disintegrated mitochondrial network compared to those from control Wistar rats and model insulinoma INS-1E cells. Standardization of these patterns may lead to development of morphological diagnostics for Langerhans islets, for the assessment of β-cell condition, before their transplantations.
Mitochondrial dysfunction in pancreatic β-cells
2012
In pancreatic b cells, mitochondria play a central role in coupling glucose metabolism to insulin exocytosis, thereby ensuring strict control of glucose-stimulated insulin secretion. Defects in mitochondrial function impair this metabolic coupling, and ultimately promote apoptosis and b cell death. Various factors have been identified that may contribute to mitochondrial dysfunction. In this review we address the emerging concept of complex links between these factors. We also discuss the role of the mitochondrial genome and mutations associated with diabetes, the effect of oxidative stress and reactive oxygen species, the sensitivity of mitochondria to lipotoxicity, and the adaptive dynamics of mitochondrial morphology. Better comprehension of the molecular mechanisms contributing to mitochondrial dysfunction will help drive the development of effective therapeutic approaches.
Metabolism, 1997
Considerable interest has recently focused on the possible role of alterations in mitochondrial activity and mutations in the mitochondrial genome for the development of non-insulin-dependent diabetes: Our study aimed at investigating the normal mitochondrial respiratory chain activity of nonpurified and purified islet cells to further explor e whether some diabetic states are associated with alterations of mitochondrial oxidative processes. For this purpose, pancreatic islets were isolated from Wistar rats. Unpurified islet cells were obtained in the presence of trypsin and DNAse, and purified I~ and non-I~ cells were prepared by autofluorescence-activated sorting Using a flowcytometer. Intact ceil respiration and substrate oxidation in digitonin-permeabiiized cells were measured polarographically with a Clark oxygen electrode in a micro-water-jacketed cell.
Diabetes, 1998
BSA, bovine serum albumin; [Ca 2 + ] i , intracellular free Ca 2 + c o n c e n t r a t i o n ; COX, cytochrome c oxidase; COX2, cytochrome c oxidase subunit II gene; COX4, cytochrome c oxidase subunit IV gene; EtBr, ethidium bromide; K AT P channel, AT Psensitive K + channel; KRH buffer, HEPES-balanced Krebs-Ringer bicarbonate buffer; MIN6 mt cells, mitochondrial DNA-depleted MIN6 cells; mtDNA, mitochondrial DNA; ND1, NADH dehydrogenase subunit 1; np, nucleotide position;
In pancreatic β-cells, mitochondria play a central role in coupling glucose metabolism to insulin secretion. Chronic exposure of β-cells to metabolic stresses impairs their function and potentially induces apoptosis. Little is known on mitochondrial adaptation to metabolic stresses, i.e. high glucose, fatty acids or oxidative stress; being all highlighted in the pathogenesis of type 2 diabetes. Here, human islets were exposed for 3 days to 25 m glucose, 0.4 m palmitate, 0.4 m oleate and transiently to H 2 O 2. Culture at physiological 5.6 m glucose served as no-stress control. Expression of mitochondrion-associated genes was quantified, including the transcriptome of mitochondrial inner membrane carriers. Targets of interest were further evaluated at the protein level. Three days after acute oxidative stress, no significant alteration in β-cell function or apoptosis was detected in human islets. Palmitate specifically increased expression of the pyruvate carriers MPC1 and MPC2, whereas the glutamate carrier GC1 and the aspartate/glutamate carrier AGC1 were down-regulated by palmitate and oleate, respectively. High glucose decreased mRNA levels of key transcription factors (HNF4A, IPF1, PPARA and TFAM) and energy-sensor SIRT1. High glucose also reduced expression of 11 mtDNA-encoded respiratory chain subunits. Interestingly, transcript levels of the carriers for aspartate/glutamate AGC2, malate DIC and malate/oxaloacetate/aspartate UCP2 were increased by high glucose, a profile suggesting important mitochondrial anaplerotic/cataplerotic activities and NADPH-generating shuttles. Chronic exposure to high glucose impaired glucose-stimulated insulin secretion, decreased insulin content, promoted caspase-3 cleavage and cell death, revealing glucotoxicity. Overall, expression profile of mitochondrion-associated genes was selectively modified by glucose, delineating a glucotoxic-specific signature.
Mitochondrial nucleoid visualization in isolated pancreatic β-cells
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2014
ATP/O ratio, reflecting efficiency of ATP synthesis, was higher in shCLS HepaRG cells (+30% p b 0.05) without modification of ATP synthesis rate (using succinate as substrate). Mechanisms involved in these modifications are under investigation. In conclusion, our work clearly demonstrated, for the first time, that the reduction of cardiolipin content regulates directly ATP synthesis in human hepatic cells.
Depletion of mitochondrial DNA alters glucose metabolism in SK-Hep1 cells
American Journal of Physiology-Endocrinology and Metabolism, 2001
Maternally inherited mitochondrial DNA (mtDNA) has been suggested to be a genetic factor for diabetes. Reports have shown a decrease of mtDNA content in tissues of diabetic patients. We investigated the effects of mtDNA depletion on glucose metabolism by use of ρ0 SK-Hep1 human hepatoma cells, whose mtDNA was depleted by long-term exposure to ethidium bromide. The ρ0 cells failed to hyperpolarize mitochondrial membrane potential in response to glucose stimulation. Intracellular ATP content, glucose-stimulated ATP production, glucose uptake, steady-state mRNA and protein levels of glucose transporters, and cellular activities of glucose-metabolizing enzymes were decreased in ρ0 cells compared with parental ρ+ cells. Our results suggest that the quantitative reduction of mtDNA may suppress the expression of nuclear DNA-encoded glucose transporters and enzymes of glucose metabolism. Thus this may lead to diabetic status, such as decreased ATP production and glucose utilization.
Mitochondrial factors in the pathogenesis of diabetes: a hypothesis for treatment
Alternative medicine review : a journal of clinical therapeutic, 2002
A growing body of evidence has demonstrated a link between various disturbances in mitochondrial functioning and type 2 diabetes. This review focuses on a range of mitochondrial factors important in the pathogenesis of this disease. The mitochondrion is an integral part of the insulin system found in the islet cells of the pancreas. Because of the systemic complexity of mitochondrial functioning in terms of tissue and energetic thresholds, details of structure and function are reviewed. The expression of type 2 diabetes can be ascribed to a number of qualitative or quantitative changes in the mitochondria. Qualitative changes refer to genetic disturbances in mitochondrial DNA (mtDNA). Heteroplasmic as well as homoplasmic mutations of mtDNA can lead to the development of a number of genetic disorders that express the phenotype of type 2 diabetes. Quantitative decreases in mtDNA copy number have also been linked to the pathogenesis of diabetes. The study of the relationship of mtDNA t...