Shao-Wen Weng

Mitochondrial DNA Coding and Control Region Variants as Genetic Risk Factors for Type 2 Diabetes

Both the coding and control regions of mitochondrial DNA (mtDNA) play roles in the generation of diabetes; however, no studies have thoroughly reported on the combined diabetogenic effects of variants in the two regions. We determined the mitochondrial haplogroup and the mtDNA sequence of the control region in 859 subjects with diabetes and 1,151 normoglycemic control subjects. Full-length mtDNA sequences were conducted in 40 subjects harboring specific diabetes-related haplogroups. Multivariate logistic regression analysis with adjustment for age, sex, and BMI revealed that subjects harboring the mitochondrial haplogroup B4 have significant association with diabetes (DM) (odds ratio [OR], 1.54 [95% CI 1.18–2.02]; P < 0.001), whereas subjects harboring D4 have borderline resistance against DM generation (0.68 [0.49–0.94]; P = 0.02). Upon further study, we identified an mtDNA composite group susceptible to DM generation consisting of a 10398A allele at the coding region and a polycytosine variant at nucleotide pair 16184–16193 of the control region, as well as a resistant group consisting of C5178A, A10398G, and T152C variants. The OR for susceptible group is 1.31 (95% CI 1.04–1.67; P = 0.024) and for the resistant group is 0.48 (0.31–0.75; P = 0.001). Our study found that mtDNA variants in the coding and control regions can have combined effects influencing diabetes generation.

Peripheral blood mitochondrial DNA content and dysregulation of glucose metabolism.

OBJECTIVE The aim of this study was to examine the potential influence of insulin resistance (IR), hyperglycemia and oxidative stress on leucocytes mitochondrial DNA (mtDNA) content. RESEARCH DESIGN AND METHOD One hundred twenty-five T2DM, 101 IFG and 70 normal subjects were enrolled in this study. The quantity of relative mtDNA content was measured by a real-time PCR and corrected by simultaneous measurement of the nuclear DNA. Parameters of lipid peroxidation, thiobarbituric acid reactive substance (TBARS), and total free thiols as antioxidative status were measured from serum samples. IR was assessed by homeostasis model assessment in the non-diabetic groups. Relationships among different variables were analyzed by general linear model correlation. RESULTS In all subjects, after correcting for age, sex and BMI, there were progressive increases of leucocyte mtDNA copy number, TBARS, and total reduced thiols with progressive dysregulation of glucose metabolism (normal vs. IFG vs. T2DM). Furthermore, correlation between mtDNA content and glucose dysregulation persisted after sequential correction for age, sex, BMI and TBARS. The independent predictor of mtDNA content by regression analysis was hyperglycemia. In non-diabetic group, influence of family history of diabetes on mtDNA content turned to non-significant after correcting for fasting plasma glucose (FPG). Correlation study revealed that mtDNA content was correlated with FPG (P<0.001), but not IR. CONCLUSION Our results indicate that hyperglycemia, not IR, is associated with an increase of leucocyte mtDNA copy number in cases of glucose dysregulation.

Tissue-specific differences in mitochondrial DNA content in type 2 diabetes

BACKGROUND To investigate whether the effect of hyperglycemia on mitochondrial DNA (mtDNA) content is tissue-specific. METHOD We compared the mtDNA contents in leg muscle, blood vessel, and peripheral leucocytes in seventeen patients with type 2 diabetes (T2DM) with those of seven controls. We measured 8-hydroxydeoxyguanosine (8-OHdG) expression in the muscles and thiobarbituric acid reactive substance (TBARS) in sera to evaluate oxidative stress. Immunohistochemical detection of peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC1-α), mitochondrial transcription factor A (Tfam), and apoptosis were performed in the muscle tissue. RESULTS The mtDNA copy number was highest in muscle tissue, followed by blood vessel tissue, and lowest in leucocytes in both the diabetic and control subjects. The diabetic patients had less mtDNA content in the muscle than the controls (2.86±0.33 vs. 3.20±0.14, P=0.025), but more mtDNA content in the leucocytes (2.25±0.26 vs. 1.98±0.06, P=0.04). In both groups, there was a positive correlation between muscle tissue mtDNA content and the expression of 8-OHdG. Patients with T2DM had significantly increased 8-OHdG and TUNEL labeling index and non-significant increases in the expression of PGC1-α and Tfam. CONCLUSION Oxidative stress stimulates mitochondrial biogenesis but induces a greater degree of apoptosis in diabetic patients, resulting in a decrease in muscle tissue mtDNA content.

Mitochondrial DNA variants in the pathogenesis of type 2 diabetes - relevance of asian population studies.

Mitochondrial dysfunction involves defective insulin secretion by pancreatic beta-cells, and insulin resistance in insulin-sensitive tissues such as muscle and adipose tissue. Mitochondria are recognized as the most important cellular source of energy, and the major generator of intracellular reactive oxygen species (ROS). Intracellular antioxidative systems have been developed to cope with increased oxidative damage. In case of minor oxidative stress, the cells may increase the number of mitochondria to produce more energy. A mechanism called mitochondrial biogenesis, involving several transcription factors and regulators, controls the quantity of mitochondria. When oxidative damage is advanced beyond the repair capacity of antioxidative systems, then oxidative stress can lead to cell death. Therefore, this organelle is central to cell life or death. Available evidence increasingly shows genetic linkage between mitochondrial DNA (mtDNA) alterations and type 2 diabetes (T2D). Based on previous studies, the mtDNA 16189 variant is associated with metabolic syndrome, higher fasting insulin concentration, insulin resistance index and lacunar cerebral infarction. These data support the involvement of mitochondrial genetic variation in the pathogenesis of T2D. Importantly, phylogeographic studies of the human mtDNAs have revealed that the human mtDNA tree is rooted in Africa and radiates into different geographic regions and can be grouped as haplogroups. The Asian populations carry very different mtDNA haplogroups as compared to European populations. Therefore, it is critically important to determine the role of mtDNA polymorphisms in T2D.

Expression of the multidrug resistance protein MRP and the lung-resistance protein LRP in nasal NK/T cell lymphoma: further exploring the role of P53 and WT1 gene

Aims: Nasal NK/T‐cell lymphoma (NKTL) is relatively common in the adult men of Asia. Many patients with nasal NKTL have poor response to therapy. Some of them show P‐glycoprotein over‐expression. To investigate the expression of other multidrug resistance proteins (MDR) and possible regulatory factors in nasal NKTL, the clinical and pathological features are described. Methods: Thirty Chinese adults with nasal NKTL are presented. Immunohistochemical study was carried out to detect multidrug resistance‐associated protein 1 (MRP) and lung resistance‐related protein (LRP). The association between possible regulatory proteins (P53 and WT1) and MDR were explored. In situ hybridisation for Epstein–Barr virus (EBV) detection, polymerase chain reaction assay for T‐cell receptor gene and direct sequencing for the P53 gene were performed. Results: Seven (23.3%) and eight (26.7%) patients showed immunoreactivity of MRP and LRP, respectively. Positive staining for both markers was identified in 6.7% (2 cases). The EBV was detected in most cases (97%). Twenty‐six (86.7%) cases expressed positive nuclear staining of P53. However, of the cases analysed for P53 mutation, none showed a mutaion at the hot spots studied. WT1 protein was not detected in the nasal NKTL. Conclusion: Our study reports expression of MRP and LRP in nasal NKTL. The over‐expression of P53 is probably associated with high incidence of EBV infection and unlikely a regulatory protein for the expression of MRP and LRP. Further studies are necessary to validate the association between P53 mutation and expression of MRP and LRP in nasal NKTL.

Gly482Ser polymorphism in the peroxisome proliferator-activated receptor γ coactivator-1α gene is associated with oxidative stress and abdominal obesity.

The objective of the study was to o investigate the relationship of the Gly482Ser (G482S) polymorphism in the peroxisome proliferator-activated receptor gamma coactivator-1alpha (PPARGC1A) gene and type 2 diabetes mellitus (T2DM), obesity, and oxidative status in Chinese adults. We enrolled 276 T2DM patients and 1049 nondiabetic subjects aged at least 35 years. The G482S variant was detected using polymerase chain reaction and restriction enzyme digestion. The levels of thiobarbituric acid reactive substance, an indicator of lipid peroxidation, were measured in plasma samples. The homeostasis model assessment-estimated insulin resistance (HOMA-IR) index was determined for nondiabetic subjects. P values were adjusted for age, sex, and body mass index by using a generalized linear model. In this series, there was no association between G482S polymorphism and T2DM and obesity (body mass index >25 kg/m(2)). However, the plasma fasting insulin levels and HOMA-IR indices were significantly higher in nondiabetic subjects harboring the variant (S/S) genotype than in those with the heterozygous (G/S) genotype. With regard to the effect of the different genotypes on body fat distribution, overweight nondiabetic subjects harboring the S/S or G/S genotype had a significantly higher waist-to-hip ratio than those with the wild-type (G/G) genotype. Furthermore, subjects with the S/S genotype had significantly higher serum thiobarbituric acid reactive substance levels than those with the G/G genotype; the diabetic group mainly contributed to this significant association (P < .001). In overweight, nondiabetic Chinese adults, G482S polymorphism in the PPARGC1A gene is associated with hyperinsulinemia, HOMA-IR indices, and abdominal obesity. Furthermore, in hyperglycemia, the S482 allele is related to increased oxidative stress.

Analysis of traditional and nontraditional risk factors for peripheral arterial disease in elderly type 2 diabetic patients in Taiwan.

AIMS Diagnosing peripheral arterial disease (PAD) and recognizing its associated risk factors in diabetes is important due to high cardiovascular disease and limb loss risk. However, both traditional and nontraditional risk factors have seldom been analyzed in the same diabetic cohort. The aim of this study was to examine the traditional and nontraditional risk factors for PAD in elderly type 2 diabetic patients. METHODS Five hundred and eighty type 2 diabetic subjects aged >or=60 years were cross-sectionally studied. Diagnosis of PAD was by ankle-brachial index (ABI) <0.90 on either leg. The association between traditional and nontraditional risk factors of PAD was analyzed. RESULTS Among the confounders, age, diabetes duration, HDL cholesterol, albuminuria, CKD (chronic kidney disease), hsCRP and insulin use differed between patients with and without PAD. Multiple logistic regression revealed that only CKD, insulin use, albuminuria, elevated hsCRP level (>3mg/l) and low HDL cholesterol were independent risk factors. CONCLUSIONS The findings of this study highlight the importance of monitoring nontraditional risk factors of PAD in diabetes. Implementing effective interventions to improve management of these risk factors may lower the risk for PAD.

Study of insulin resistance in cybrid cells harboring diabetes-susceptible and diabetes-protective mitochondrial haplogroups

AIM This study aims to elucidate the independent role of mitochondria in the pathogenesis of insulin resistance (IR). METHODS Cybrids derived from 143B osteosarcoma cell line and harboring the same nuclear DNA but different mitochondrial haplogroups were studied. Cybrid B4 (the major diabetes-susceptible haplogroup in Chinese population), cybrid D4 (the major diabetes-resistant haplogroup in Chinese population) and cybrid N9 (the diabetes-resistant haplogroup in Japanese population) were cultured in a medium containing 25 mM glucose and stimulated with 0 μM, 0.1 μM, and 1.0 μM insulin. We compared the insulin activation of PI3K-Akt (glucose uptake) and ERK-MAPK (pro-inflammation) signaling pathways, intracellular and mitochondrial oxidative stress (DCF and MitoSOX Red), and their responses to the antioxidant N-acetylcysteine (NAC). RESULTS Upon insulin treatment, the translocation of cytoplasmic GLUT1/GLUT4 to the cell membrane in cybrid D4 and N9 cells increased significantly, whereas the changes in B4 cells were not or less significant. On the contrary, the ratio of insulin-induced JNK and P38 to Akt phosphorylation was significantly greater in cybrid B4 cells than in cybrid D4 and N9 cells. The levels of DCF and MitoSOX Red, which are indicative of the oxidative stress, were significantly higher in the B4 cells in basal conditions and after insulin treatment. Following treatment with the antioxidant NAC, cybrid B4 cells showed significantly reduced insulin-induced phosphorylation of P38 and increased GLUT1/GLUT4 translocation to the cell membrane, suggesting that NAC may divert insulin signaling from pro-inflammation to glucose uptake. CONCLUSIONS Mitochondria play an independent role in the pathogenesis of IR, possibly through altered production of intracellular ROS.

Biphasic Response of Mitochondrial Biogenesis to Oxidative Stress in Visceral Fat of Diet-Induced Obesity Mice

AIMS Studies in skeletal muscle demonstrate a strong association of mitochondrial dysfunction with insulin resistance (IR). However, there is still a paucity of knowledge regarding the alteration of mitochondria in adipose tissue (AT) in the pathogenesis of IR in obesity. We investigated the mitochondrial biogenesis in visceral fat (VF) and subcutaneous fat (SF) in C57BL/6J mice fed a high-fat high-sucrose diet for 12 months. RESULTS Impairment of glucose tolerance and insulin sensitivity developed after 1 month of the diet and was associated with a prompt increase of VF. The VF adipocytes were larger than those in the SF and had increased expressions of HIF-1α and p-NFκB p65. However, the alteration of mitochondrial biogenesis did not occur in the early stage when increased intracellular reactive oxygen species (ROS), mitochondrial oxygen consumption rate, and mitochondrial ROS emerged at the 1st, 2nd and 2nd month, respectively. Until the 6th month, the VF had markedly increased mitochondrial DNA content and expression of PGC-1α, Tfam, ATP5A, and MnSOD. This increase of mitochondrial biogenesis was followed by a generalized decrease at the 12th month and the mitochondrial morphology altered markedly. In the late stage, although mitochondrial ROS decreased, the increased expression of 8-OHdG in VF continued. INNOVATION AND CONCLUSION These data suggest that IR and ROS production occur before the biphasic changes of mitochondrial biogenesis in AT, and the VF plays a more crucial role.

Mitochondrial DNA variants as genetic risk factors for Parkinson disease

Investigation of the relationship between mitochondrial DNA (mtDNA) variants and Parkinson disease (PD) remains an issue awaiting more supportive evidence. Moreover, an affirming cellular model study is also lacking.