Mitochondrial defects in pancreatic beta-cell dysfunction and neurodegenerative diseases: Pathogenesis and therapeutic applications.

Mitochondria malfunction is linked to the development of ß-cell failure and a variety of neurodegenerative disorders. Pancreatic ß-cells are normally configured to detect glucose and other food secretagogues in order to adjust insulin exocytosis and maintain glucose homeostasis. As a result of the increased glucose level, mitochondria metabolites and nucleotides are produced, which operate in concert with cytosolic Ca2+ to stimulate insulin secretion. Furthermore, mitochondria are the primary generators of adenosine triphosphate (ATP), reactive oxygen species (ROS), and apoptosis regulation. Mitochondria are concentrated in synapses, and any substantial changes in synaptic mitochondria location, shape, quantity, or function might cause oxidative stress, resulting in faulty synaptic transmission, a symptom of various degenerative disorders at an early stage. However, a greater understanding of the role of mitochondria in the etiology of ß-cell dysfunction and neurodegenerative disorder should pave the way for a more effective approach to addressing these health issues. This review looks at the widespread occurrence of mitochondria depletion in humans, and its significance to mitochondria biogenesis in signaling and mitophagy. Proper understanding of the processes might be extremely beneficial in ameliorating the rising worries about mitochondria biogenesis and triggering mitophagy to remove depleted mitochondria, therefore reducing disease pathogenesis.

Therapeutic efficacy of Clompanus pubescens leaves fractions via downregulation of neuronal cholinesterases/Na+-K+ATPase/IL-1 ß, and improving the neurocognitive and antioxidants status of streptozotocin-induced diabetic rats.

The increasing global burden of diabetes mellitus has called for the search for a therapeutic alternative that offers better activities and safety than conventional chemotherapy. Herein, we evaluated the neuroprotective and antioxidant properties of different fractions (ethyl acetate, N-butanol and residual aqueous) of Clompanus pubescens leaves in streptozotocin (STZ)-induced diabetic rats. Our results revealed a significant elevation in the levels of blood glucose, pro-inflammatory cytokines, lipid peroxidation, neuronal activities of acetylcholinesterase, butyrylcholinesterase, nitric oxide, epinephrine, norepinephrine, and Na+/K+-ATPase in diabetic non treated rats. In addition, decreased levels of enzymatic and non-enzymatic antioxidants were observed. Treatment with different fractions of C. pubescens leaves resulted in significant reversal of the biochemical alteration and improved the neurocognitive deficit in STZ induced diabetic rats. However, the ethyl-acetate fraction demonstrated higher activities than the other fractions and was characterized for its phytoconstituents, revealing the presence of Gallic acid (713.00 ppm), catechin (0.91 ppm), ferulic acid (0.98 ppm), rutin (59.82 ppm), quercetin (3.22 ppm) and kaempferol (4.07 ppm). Our molecular docking analysis revealed that these compounds exhibited different binding affinities and potentials for targeting BChE/AChE/ IL-1 ß/Na+ -K+ -ATPase. However, only Kampferol and ferulic exhibited good drug-like, ADMET, and permeability properties suitable for use as a neuronal drug target agent. Hence, the ethyl-acetate fraction of C. pubescens leaves could be considered as a source of promising bioactive metabolite for the treatment and management of cognitive impairments related to type II diabetes mellitus.

The Perlman syndrome DIS3L2 exoribonuclease safeguards endoplasmic reticulum-targeted mRNA translation and calcium ion homeostasis.

DIS3L2-mediated decay (DMD) is a surveillance pathway for certain non-coding RNAs (ncRNAs) including ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), small nuclear RNAs (snRNAs), and RMRP. While mutations in DIS3L2 are associated with Perlman syndrome, the biological significance of impaired DMD is obscure and pathological RNAs have not been identified. Here, by ribosome profiling (Ribo-seq) we find specific dysregulation of endoplasmic reticulum (ER)-targeted mRNA translation in DIS3L2-deficient cells. Mechanistically, DMD functions in the quality control of the 7SL ncRNA component of the signal recognition particle (SRP) required for ER-targeted translation. Upon DIS3L2 loss, sustained 3'-end uridylation of aberrant 7SL RNA impacts ER-targeted translation and causes ER calcium leakage. Consequently, elevated intracellular calcium in DIS3L2-deficient cells activates calcium signaling response genes and perturbs ESC differentiation. Thus, DMD is required to safeguard ER-targeted mRNA translation, intracellular calcium homeostasis, and stem cell differentiation.

FGF6 and FGF9 regulate UCP1 expression independent of brown adipogenesis.

Uncoupling protein-1 (UCP1) plays a central role in energy dissipation in brown adipose tissue (BAT). Using high-throughput library screening of secreted peptides, we identify two fibroblast growth factors (FGF), FGF6 and FGF9, as potent inducers of UCP1 expression in adipocytes and preadipocytes. Surprisingly, this occurs through a mechanism independent of adipogenesis and involves FGF receptor-3 (FGFR3), prostaglandin-E2 and interaction between estrogen receptor-related alpha, flightless-1 (FLII) and leucine-rich-repeat-(in FLII)-interacting-protein-1 as a regulatory complex for UCP1 transcription. Physiologically, FGF6/9 expression in adipose is upregulated by exercise and cold in mice, and FGF9/FGFR3 expression in human neck fat is significantly associated with UCP1 expression. Loss of FGF9 impairs BAT thermogenesis. In vivo administration of FGF9 increases UCP1 expression and thermogenic capacity. Thus, FGF6 and FGF9 are adipokines that can regulate UCP1 through a transcriptional network that is dissociated from brown adipogenesis, and act to modulate systemic energy metabolism.

Retinol binding protein 3 is increased in the retina of patients with diabetes resistant to diabetic retinopathy.

The Joslin Medalist Study characterized people affected with type 1 diabetes for 50 years or longer. More than 35% of these individuals exhibit no to mild diabetic retinopathy (DR), independent of glycemic control, suggesting the presence of endogenous protective factors against DR in a subpopulation of patients. Proteomic analysis of retina and vitreous identified retinol binding protein 3 (RBP3), a retinol transport protein secreted mainly by the photoreceptors, as elevated in Medalist patients protected from advanced DR. Mass spectrometry and protein expression analysis identified an inverse association between vitreous RBP3 concentration and DR severity. Intravitreal injection and photoreceptor-specific overexpression of RBP3 in rodents inhibited the detrimental effects of vascular endothelial growth factor (VEGF). Mechanistically, our results showed that recombinant RBP3 exerted the therapeutic effects by binding and inhibiting VEGF receptor tyrosine phosphorylation. In addition, by binding to glucose transporter 1 (GLUT1) and decreasing glucose uptake, RBP3 blocked the detrimental effects of hyperglycemia in inducing inflammatory cytokines in retinal endothelial and Müller cells. Elevated expression of photoreceptor-secreted RBP3 may have a role in protection against the progression of DR due to hyperglycemia by inhibiting glucose uptake via GLUT1 and decreasing the expression of inflammatory cytokines and VEGF.

CISD2 Haploinsufficiency Disrupts Calcium Homeostasis, Causes Nonalcoholic Fatty Liver Disease, and Promotes Hepatocellular Carcinoma.

CISD2 is located within the chromosome 4q region frequently deleted in hepatocellular carcinoma (HCC). Mice with Cisd2 heterozygous deficiency develop a phenotype similar to the clinical manifestation of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). Cisd2 haploinsufficiency causes a low incidence (20%) of spontaneous HCC and promotes HBV-associated and DEN-induced HCC; conversely, 2-fold overexpression of Cisd2 suppresses HCC in these models. Mechanistically, Cisd2 interacts with Serca2b and mediates its Ca2+ pump activity via modulation of Serca2b oxidative modification, which regulates ER Ca2+ uptake and maintains intracellular Ca2+ homeostasis in the hepatocyte. CISD2 haploinsufficiency disrupts calcium homeostasis, causing ER stress and subsequent NAFLD and NASH. Hemizygous deletion and decreased expression of CISD2 are detectable in a substantial fraction of human HCC specimens. These findings substantiate CISD2 as a haploinsufficient tumor suppressor and highlights Cisd2 as a drug target when developing therapies to treat NAFLD/NASH and prevent HCC.

Mitochondrial dysfunction leads to impairment of insulin sensitivity and adiponectin secretion in adipocytes.

Adipocytes play an integrative role in the regulation of energy metabolism and glucose homeostasis in the human body. Functional defects in adipocytes may cause systemic disturbance of glucose homeostasis. Recent studies revealed mitochondrial abnormalities in the adipose tissue of patients with type 2 diabetes. In addition, patients with mitochondrial diseases usually manifest systemic metabolic disorder. However, it is unclear how mitochondrial dysfunction in adipocytes affects the regulation of glucose homeostasis. In this study, we induced mitochondrial dysfunction and overproduction of reactive oxygen species (ROS) by addition of respiratory inhibitors oligomycin A and antimycin A and by knockdown of mitochondrial transcription factor A (mtTFA), respectively. We found an attenuation of the insulin response as indicated by lower glucose uptake and decreased phosphorylation of Akt upon insulin stimulation of adipocytes with mitochondrial dysfunction. Furthermore, the expression of glucose transporter 4 (Glut4) and secretion of adiponectin were decreased in adipocytes with increased ROS generated by defective mitochondria. Moreover, the severity of insulin insensitivity was correlated with the extent of mitochondrial dysfunction. These results suggest that higher intracellular ROS levels elicited by mitochondrial dysfunction resulted in impairment of the function of adipocytes in the maintenance of glucose homeostasis through attenuation of insulin signaling, downregulation of Glut4 expression, and decrease in adiponectin secretion. Our findings substantiate the important role of mitochondria in the regulation of glucose homeostasis in adipocytes and also provide a molecular basis for the explanation of the manifestation of diabetes mellitus or insulin insensitivity in a portion of patients with mitochondrial diseases such as MELAS or MERRF syndrome.

The role of cytochrome c oxidase subunit Va in non-small cell lung carcinoma cells: association with migration, invasion and prediction of distant metastasis.

BACKGROUND: Lung cancer is one of the most lethal malignancies worldwide, but useful biomarkers of lung cancer are still insufficient. The aim of this study is to identify some membrane-bound protein(s) associated with migration and invasion in human non-small cell lung cancer (NSCLC) cells. METHODS: We classified four NSCLC cell lines into high and low migration/invasion groups by Transwell and Matrigel assays. Using two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), we identified 10 membrane-associated proteins being significantly overexpressed in the high migration/invasion group. The expression of the target protein in the four NSCLC cell lines was then confirmed by reverse transcription polymerase chain reaction (RT-PCR), western blot and immunostaining. RNA interference technique was applied to observe the influence of the target protein on migration and invasion. Gelatin zymography was also performed to evaluate the activities of matrix metalloproteinase (MMP)-2 and MMP-9. Expression condition of the target protein on surgical specimens was further examined by immunohistochemical staining and the clinicopathologic data were analyzed. RESULTS: We identified a mitochondria-bound protein cytochrome c oxidase subunit Va (COX Va) because of its abundant presence found exclusively in tumorous areas. We also demonstrated that migration and invasion of NSCLC cells decreased substantially after knocking down COX Va by siRNA. Meanwhile, we found a positive correlation between COX Va expression, Bcl-2 expression and activities of MMP-2 and MMP-9 in NSCLC cells. Immunohistochemical staining of surgically resected lung adenocarcinomas in 250 consecutive patients revealed that strong COX Va expression was found in 54.8% (137/250) of patients and correlated positively with the status of lymph node metastasis (P = 0.032). Furthermore, strong COX Va expression was associated with the presence of distant metastasis (P = 0.033). CONCLUSIONS: Our current study showed that COX Va may play a role in migration and invasion of NSCLC cells and can be used as a biomarker to predict aggressiveness of NSCLC.

Chronic hepatitis C virus infection increases mortality from hepatic and extrahepatic diseases: a community-based long-term prospective study.

BACKGROUND: The study aimed to evaluate the risk of hepatitis C virus (HCV) infection on hepatic and extrahepatic deaths. METHODS: A cohort of 23 820 adults aged 30-65 years old were enrolled during 1991-1992. The seromarkers hepatitis B surface antigen (HBsAg), anti-HCV, and serum HCV RNA levels at study entry were tested. The vital status was ascertained through computerized linkage with national death certification profiles from 1991 to 2008. RESULTS: There were 19,636 HBsAg-seronegatives, including 18,541 anti-HCV seronegatives and 1095 anti-HCV seropositives. Among anti-HCV seropositives, 69.4% had detectable serum HCV RNA levels. There were 2394 deaths that occurred during an average follow-up period of 16.2 years. Compared with anti-HCV seronegatives, anti-HCV seropositives had higher mortality from both hepatic and extrahepatic diseases, showing multivariate-adjusted hazard ratio (95% confidence interval) of 1.89 (1.66-2.15) for all causes of death; 12.48 (9.34-16.66) for hepatic diseases; 1.35 (1.15-1.57) for extrahepatic diseases; 1.50 (1.10-2.03) for circulatory diseases; 2.77 (1.49-5.15) for nephritis, nephrotic syndrome, and nephrosis; 4.08 (1.38-12.08) for esophageal cancer; 4.19 (1.18-14.94) for prostate cancer; and 8.22 (1.36-49.66) for thyroid cancer. Anti-HCV seropositives with detectable HCV RNA levels had significantly higher mortality from hepatic and extrahepatic diseases than anti-HCV seropositives with undetectable HCV RNA. CONCLUSIONS: Monitoring HCV RNA in anti-HCV seropositives is essential for the prediction of mortality associated with hepatitis C.

A persistent level of Cisd2 extends healthy lifespan and delays aging in mice.

The CISD2 gene, which is an evolutionarily conserved novel gene, encodes a transmembrane protein primarily associated with the mitochondrial outer membrane. Significantly, the CISD2 gene is located within the candidate region on chromosome 4q where a genetic component for human longevity has been mapped. Previously, we have shown that Cisd2 deficiency shortens lifespan resulting in premature aging in mice. Additionally, an age-dependent decrease in Cisd2 expression has been detected during normal aging. In this study, we demonstrate that a persistent level of Cisd2 achieved by transgenic expression in mice extends their median and maximum lifespan without any apparent deleterious side effects. Cisd2 also ameliorates age-associated degeneration of the skin, skeletal muscles and neurons. Moreover, Cisd2 protects mitochondria from age-associated damage and functional decline as well as attenuating the age-associated reduction in whole-body energy metabolism. These results suggest that Cisd2 is a fundamentally important regulator of lifespan and provide an experimental basis for exploring the candidacy of CISD2 in human longevity.

Increase of reduced nicotinamide adenine dinucleotide fluorescence lifetime precedes mitochondrial dysfunction in staurosporine-induced apoptosis of HeLa cells.

In vivo noninvasive detection of apoptosis represents a new tool that may yield a more definite diagnosis, a more accurate prognosis, and help improve therapies for human diseases. The intrinsic fluorescence of reduced nicotinamide adenine dinucleotide (NADH) may be a potential optical biomarker for the apoptosis detection because NADH is involved in the respiration for the mitochondrial membrane potential (ΔΨ) formation and adenosine-5'-triphosphate (ATP) synthesis, and the depletion of ΔΨ and ATP level is the hallmark of apoptosis. We have previously observed the NADH fluorescence lifetime change is associated with staurosporine (STS)-induced mitochondria-mediated apoptosis. However, its relationship with mitochondrial functions such as ΔΨ, ATP, and oxygen consumption rate is not clear. In this study, we investigated this relationship. Our results indicate that the NADH fluorescence lifetime increased when ΔΨ and ATP levels were equal to or higher than their values of controls and decreased before the depletion of ΔΨ and ATP, and the oxygen consumption rate did not change. These findings suggest that the increased NADH fluorescence lifetime in STS-induced cell death occurred before the depletion of ΔΨ and ATP and activation of caspase 3, and was not simply caused by cellular metabolic change. Furthermore, the NADH fluorescence lifetime change is associated with the pace of apoptosis.

Mitochondrial dysfunction in insulin insensitivity: implication of mitochondrial role in type 2 diabetes.

Abundant evidence has been accumulated to suggest that mitochondrial dysfunction is associated with type 2 diabetes. Research findings from this and other laboratories have supported the notion that impaired mitochondrial function is a cause of insulin insensitivity in myocytes and adipocytes as a result of insufficient supply of energy or defects in the insulin signaling pathway. We demonstrated that inhibition of respiration and oxidative phosphorylation by respiratory inhibitors or knockdown of genes involved in mitochondrial biogenesis can impair the differentiation of preadipocytes and response of adipocytes to insulin. Moreover, defective mitochondria also cause a decrease in adiponectin secretion that leads to decline glucose utilization of other tissues. Besides, it has been elucidated that some environmental factors, pollutants, and mitochondrial toxins are involved in the pathogenesis of type 2 diabetes. Taken together, we suggest that mitochondrial dysfunction plays a role in the pathophysiology of insulin insensitivity, and that activation of mitochondrial biogenesis may be an effective strategy in the prevention or treatment of insulin resistance and type 2 diabetes.

A simple Bayesian mixture model with a hybrid procedure for genome-wide association studies.

Genome-wide association studies often face the undesirable result of either failing to detect any influential markers at all because of a stringent level for testing error corrections or encountering difficulty in quantifying the importance of markers by their P-values. Advocates of estimation procedures prefer to estimate the proportion of association rather than test significance to avoid overinterpretation. Here, we adopt a Bayesian hierarchical mixture model to estimate directly the proportion of influential markers, and then proceed to a selection procedure based on the Bayes factor (BF). This mixture model is able to accommodate different sources of dependence in the data through only a few parameters. Specifically, we focus on a standardized risk measure of unit variance so that fewer parameters are involved in inference. The expected value of this measure follows a mixture distribution with a mixing probability of association, and it is robust to minor allele frequencies. Furthermore, to select promising markers, we use the magnitude of the BF to represent the strength of evidence in support of the association between markers and disease. We demonstrate this procedure both with simulations and with SNP data from studies on rheumatoid arthritis, coronary artery disease, and Crohn's disease obtained from the Wellcome Trust Case-Control Consortium. This Bayesian procedure outperforms other existing methods in terms of accuracy, power, and computational efficiency. The R code that implements this method is available at http://homepage.ntu.edu.tw/~ckhsiao/Bmix/Bmix.htm.

Biomarkers of exposure, effect, and susceptibility of arsenic-induced health hazards in Taiwan.

Long-term exposure to inorganic arsenic from drinking water has been documented to induce cancers and vascular diseases in a dose-response relationship. A series of molecular environmental epidemiological studies have been carried out to elucidate biomarkers of exposure, effect, and susceptibility for arsenic-related health hazards in Taiwan. Arsenic levels in urine, hair, and nail are biomarkers for short-term (<1 year) internal dose, skin hyperpigmentation and palmoplantar hyperkeratosis are for long-term (many years) internal dose, and percentage of monomethylarsonic acid in total metabolites of inorganic arsenic in urine may be considered as an exposure marker for biologically effective dose. The biomarkers of early biological effects of ingested inorganic arsenic included blood levels of reactive oxidants and anti-oxidant capacity, genetic expression of inflammatory molecules, as well as cytogenetic changes including sister chromatid exchange, micronuclei, and chromosome aberrations of peripheral lymphocytes. Both mutation type and hot spots of p53 gene were significantly different in arsenic-induced and non-arsenic-induced TCCs. The frequency of chromosomal imbalances analyzed by comparative genomic hybridization and the frequency of loss of heterozygosity were significantly higher in arsenic-induced TCC than non-arsenic-induced TCC at specific sites. Biomarkers of susceptibility to arsenic-induced health hazards included genetic polymorphisms of enzymes involved in xenobiotic metabolism, DNA repair, and oxidative stress, as well as serum level of carotenoids. Gene-gene and gene-environment interactions are involved in arsenic-induced health hazards through toxicological mechanisms including genomic instability and oxidative stress.

Biological gradient between long-term arsenic exposure and carotid atherosclerosis.

BACKGROUND: Long-term exposure to ingested arsenic has been documented to induce peripheral vascular disease, ischemic heart disease, and cerebral infarction in a dose-response relationship. This study further examined the biological gradient between ingested inorganic arsenic and carotid atherosclerosis. METHODS AND RESULTS: We studied 199 male and 264 female adult residents from the southwestern area of endemic arseniasis in Taiwan. The extent of carotid atherosclerosis was assessed by duplex ultrasonography. Diabetes mellitus was determined by oral glucose tolerance test, hypertension by mercury sphygmomanometers, and serum lipid profiles by autoanalyzers. Information regarding the consumption of high-arsenic artesian well water, cigarette smoking, and alcohol consumption was obtained through standardized questionnaire interviews. Logistic regression analysis was used to estimate the odds ratio and its 95% CI of carotid atherosclerosis for various risk factors. Three indices of long-term exposure to ingested arsenic, including the duration of consuming artesian well water, the average arsenic concentration in consumed artesian well water, and cumulative arsenic exposure, were all significantly associated with prevalence of carotid atherosclerosis in a dose-response relationship. The biological gradient remained significant after adjustment for age, sex, hypertension, diabetes mellitus, cigarette smoking, alcohol consumption, waist-to-hip ratio, and serum levels of total cholesterol and LDL cholesterol. The multivariate-adjusted odds ratio was 3.1 (95% CI 1.3 to 7.4) for those who had a cumulative arsenic exposure of > or =20 mg/L-years compared with those without exposure to arsenic from drinking artesian well water. CONCLUSIONS: Carotid atherosclerosis is associated with ingested inorganic arsenic, showing a significant biological gradient.