Skeletal Muscle Mitochondrial Functions, Mitochondrial DNA Copy Numbers, and Gene Transcript Profiles in Type 2 Diabetic and Nondiabetic Subjects at Equal Levels of Low or High Insulin and Euglycemia

Abstract

We investigated whether previously reported muscle mitochondrial dysfunction and altered gene transcript levels in type 2 diabetes might be secondary to abnormal blood glucose and insulin levels rather than an intrinsic defect of type 2 diabetes. A total of 13 type 2 diabetic and 17 nondiabetic subjects were studied on two separate occasions while maintaining similar insulin and glucose levels in both groups by 7-h infusions of somatostatin, low-or high-dose insulin (0.25 and 1.5 mU/kg of fat-free mass per min, respectively), and glucose. Muscle mitochondrial DNA abundance was not different between type 2 diabetic and nondiabetic subjects at both insulin levels, but the majority of transcripts in muscle that are involved mitochondrial functions were expressed at lower levels in type 2 diabetes at low levels of insulin. However, several gene transcripts that are specifically involved in the electron transport chain were expressed at higher levels in type 2 diabetic patients. After the low-dose insulin infusion, which achieved postabsorptive insulin levels, the muscle mitochondrial ATP production rate (MAPR) was not different between type 2 diabetic and nondiabetic subjects. However, increasing insulin to postprandial levels increased the MAPR in nondiabetic subjects but not in type 2 diabetic patients. The lack of MAPR increment in response to high-dose insulin in type 2 diabetic patients occurred in association with reduced glucose disposal and expression of peroxisome proliferator-activated receptor-␥ coactivator 1␣, citrate synthase, and cytochrome c oxidase I. In conclusion, the current data supports that muscle mitochondrial dysfunction in type 2 diabetes is not an intrinsic defect, but instead a functional defect related to impaired response to insulin. Diabetes 55:3309 -3319, 2006

Isolation of Mitochondria from Minimal Quantities of Mouse Skeletal Muscle for High Throughput Microplate Respiratory Measurements

“…These gentler homogenization and resuspension techniques leads to a high percentage of mitochondria with intact membranes (92.5± 2.0%), as assessed from citrate synthase activity following the isolation procedure. Our findings are consistent with Asmann et al 17 , who report between 90-95% intact mitochondrial preparations following isolation from human skeletal muscle. It is important to note that measuring citrate synthase activity is better suited for measuring physical integrity of the mitochondrial membranes and should not be used to assess functional integrity of the isolated mitochondria.…”

High-fat diets cause insulin resistance despite an increase in muscle mitochondria

“…It has been hypothesized that skeletal muscle insulin resistance is mediated by a mitochondrial deficiency that limits fat oxidation and results in accumulation of intramyocellular lipids (2). This concept is based on the finding that muscles of insulin-resistant individuals generally contain Ϸ30% less mitochondria than those of insulin-sensitive control subjects (3)(4)(5)(6)(7). This phenomenon has also been referred to as mitochondrial dysfunction (1, 2), although a detailed evaluation has provided evidence that the remaining mitochondria function normally (36).…”

“…The mechanism by which a decrease in mitochondria is proposed to cause insulin resistance is accumulation of intramyocellular lipids caused by a decrease in the capacity to oxidize fat (2). This hypothesis is based on the finding that type 2 diabetics and insulin-resistant individuals with impaired glucose tolerance have Ϸ30% less mitochondria in their muscles than insulinsensitive control subjects (3)(4)(5)(6)(7). In support of this concept, recent studies have reported that raising serum free fatty acids (FFA) by a high-fat diet in humans (8), or by feeding mice or rats high-fat diets (8)(9)(10), results in decreases in skeletal muscle peroxisome proliferator-activated receptor ␥ coactivator-1␣ (PGC-1␣) mRNA (8)(9)(10) and the mRNA levels of various mitochondrial constituents (8).…”

Reduction of non-esterified fatty acids improves insulin sensitivity and lowers oxidative stress, but fails to restore oxidative capacity in type 2 diabetes: a randomised clinical trial

“…Insulin has been shown to increase specific phosphorylation sites of ATP synthase by ∼50% in lean and healthy, but not insulinresistant, individuals [13]. Some investigators suggest that the insulin-resistant state may limit glucose uptake or substrate oxidation or reduce substrate transport into mitochondria during hyperinsulinaemia [11,12]. Reduced substrate transport results in diminished production of tricarboxylic acid (TCA) cycle intermediates and lower production of NADH and FADH 2 , which drives mitochondrial oxygen consumption and ATP synthesis.…”

“…Intravenous insulin infusion stimulates in vivo unidirectional flux through muscle ATP synthase (fATP) [7,9,10] as well as ex vivo maximal ATP production rate in non-diabetic humans [11,12], indicating that insulin stimulates maximal mitochondrial oxidative capacity. This effect seems to be blunted or absent in patients with type 2 diabetes [7,12], FDRs [8] and patients with type 1 diabetes [9].…”