Raghavakaimal, Sreekumar

Age‐related mitochondrial dysfunction is paralleled by reductions in gene transcripts and expression of proteins involved in cellular ATP production. Short‐term endurance exercise has been shown to enhance muscle protein synthesis in the elderly. We determined the impact of long‐term endurance exercise on gene expression and protein abundance of numerous proteins involved in cellular energetics. Vastus lateralis tissue was biopsied from 11 young sedentary (YS), 10 old sedentary (OS), 11 young trained (YT), and 11 old trained (OT) subjects. Trained subjects vigorously exercised ¡Ý 1 hr/d, ¡Ý five d/wk for ¡Ý 4 years. Large‐scale measurement of relative protein abundance was performed using isobaric tags for relative and absolute quantitation (iTRAQ) reagents and tandem mass spectrometry. Gene transcript levels were measured by high‐density oligonucleotide microarrays. Ingenuity Pathway Analysis was used to determine canonical pathways that were altered by age. Of the 33 proteins involved in cellular ATP production 27 were less abundant in OS vs. YS, while only 4 of these proteins were less abundant in OT vs. YT. Genes involved in glycolysis and oxidative phosphorylation were underexpressed in OS vs. YS, with no effects of age in OT vs. YT. CONCLUSIONS: Long‐term vigorous endurance exercise ameliorates the detrimental effects of old age on muscle gene and protein expression in humans.

Significance We have identified a lethal subtype of gastric cancer (GC) that is characterized by high levels of clonal heterogeneity and TP53 (tumor protein P53) mutation. We have also uncovered key novel mutations in the targetable NRG1 (neuregulin-1) and ERBB4 (V-Erb-B2 avian erythroblastic leukemia viral oncogene homolog 4) ligand-receptor pair and identified BRCA2 (breast cancer 2, early onset) mutations as new genetic markers to predict better survival for GC. Our study represents a novel approach for GC personalized medicine and identified novel clinical actionable therapies for GC therapy.

OBJECTIVE— We determined whether reduced insulin sensitivity, mitochondrial dysfunction, and other age-related dysfunctions are inevitable consequences of aging or secondary to physical inactivity. RESEARCH DESIGN AND METHODS— Insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp and ATP production in mitochondria isolated from vastus lateralis biopsies of 42 healthy sedentary and endurance-trained young (18–30 years old) and older (59–76 years old) subjects. Expression of proteins involved in fuel metabolism was measured by mass spectrometry. Citrate synthase activity, mitochondrial DNA (mtDNA) abundance, and expression of nuclear-encoded transcription factors for mitochondrial biogenesis were measured. SIRT3, a mitochondrial sirtuin linked to lifespan-enhancing effects of caloric restriction, was measured by immunoblot. RESULTS— Insulin-induced glucose disposal and suppression of endogenous glucose production were higher in the trained young and older subjects, but no age effect was noted. Age-related decline in mitochondrial oxidative capacity was absent in endurance-trained individuals. Although endurance-trained individuals exhibited higher expression of mitochondrial proteins, mtDNA, and mitochondrial transcription factors, there were persisting effects of age. SIRT3 expression was lower with age in sedentary but equally elevated regardless of age in endurance-trained individuals. CONCLUSIONS— The results demonstrate that reduced insulin sensitivity is likely related to changes in adiposity and to physical inactivity rather than being an inevitable consequence of aging. The results also show that regular endurance exercise partly normalizes age-related mitochondrial dysfunction, although there are persisting effects of age on mtDNA abundance and expression of nuclear transcription factors and mitochondrial protein. Furthermore, exercise may promote longevity through pathways common to effects of caloric restriction.

OBJECTIVE— Type 2 diabetes has become a global epidemic, and Asian Indians have a higher susceptibility to diabetes than Europeans. We investigated whether Indians had any metabolic differences compared with Northern European Americans that may render them more susceptible to diabetes. RESEARCH DESIGN AND METHODS— We studied 13 diabetic Indians, 13 nondiabetic Indians, and 13 nondiabetic Northern European Americans who were matched for age, BMI, and sex. The primary comparisons were insulin sensitivity by hyperinsulinemic-euglycemic clamp and skeletal muscle mitochondrial capacity for oxidative phosphorylation (OXPHOS) by measuring mitochondrial DNA copy number (mtDNA), OXPHOS gene transcripts, citrate synthase activity, and maximal mitochondrial ATP production rate (MAPR). Other factors that may cause insulin resistance were also measured. RESULTS— The glucose infusion rates required to maintain identical glucose levels during the similar insulin infusion rates were substantially lower in diabetic Indians than in the nondiabetic participants (P < 0.001), and they were lower in nondiabetic Indians than in nondiabetic Northern European Americans (P < 0.002). mtDNA (P < 0.02), OXPHOS gene transcripts (P < 0.01), citrate synthase, and MAPR (P < 0.03) were higher in Indians irrespective of their diabetic status. Intramuscular triglyceride, C-reactive protein, interleukin-6, and tumor necrosis factor-α concentrations were higher, whereas adiponectin concentrations were lower in diabetic Indians. CONCLUSIONS— Despite being more insulin resistant, diabetic Indians had similar muscle OXPHOS capacity as nondiabetic Indians, demonstrating that diabetes per se does not cause mitochondrial dysfunction. Indians irrespective of their diabetic status had higher OXPHOS capacity than Northern European Americans, although Indians were substantially more insulin resistant, indicating a dissociation between mitochondrial dysfunction and insulin resistance.

Type 2 diabetes is characterized by muscle insulin resistance. Nondiabetic first-degree relatives of type 2 diabetic patients have also been reported to have insulin resistance. A polygenic basis for pathogenesis of type 2 diabetes has been proposed. A gene expression profile was evaluated in the skeletal muscle of patients with type 2 diabetes while not on treatment for 2 weeks and after 10 days of intensive insulin treatment. Comparison of gene expression pattern with age-, sex-, and BMI-matched people with no family history of diabetes was performed using a microarray technique (Hu6800 arrays; Affymetrix, Santa Clara, CA). Only those gene transcripts showing ≥1.9-fold changes and an average difference in fluorescence intensity of ≥1,000 in all subjects are reported. Insulin sensitivity (SI) was measured using an intravenous glucose tolerance test. Of 6,451 genes surveyed, transcriptional patterns of 85 genes showed alterations in the diabetic patients after withdrawal of treatment, when compared with patterns in the nondiabetic control subjects. Insulin treatment reduced the difference in patterns between diabetic and nondiabetic control subjects (improved) in all but 11 gene transcripts, which included genes involved in structural and contractile functions, growth and tissue development, stress response, and energy metabolism. These improved transcripts included genes involved in insulin signaling, transcription factors, and mitochondrial maintenance. However, insulin treatment altered the transcription of 29 additional genes involved in signal transduction; structural and contractile functions; growth and tissue development; and protein, fat, and energy metabolism. Type 2 diabetic patients had elevated circulating insulin during the insulin-treated phase, although their blood glucose levels (98.8 ± 6.4 vs. 90.0 ± 2.9 mg/dl for diabetic vs. control) were similar to those of the control subjects. In contrast, after withdrawal of treatment, the diabetic patients had reduced SI and elevated blood glucose (224.0 ± 26.2 mg/dl), although their insulin levels were similar to those of the nondiabetic control subjects. This study identified several candidate genes for muscle insulin resistance, complications associated with poor glycemic control, and effects of insulin treatment in people with type 2 diabetes.

Abstract Mycobacterium avium complex (MAC) causes chronic lung disease in immunocompetent people and disseminated infection in patients with AIDS. MAC is intrinsically resistant to many conventional antimycobacterial agents, it develops drug resistance rapidly to macrolide antibiotics, and patients with MAC infection experience frequent relapses or the inability to completely eradicate the infection with current treatment. Treatment regimens are prolonged and complicated by drug toxicity or intolerances. We sought to identify biochemical pathways in MAC that can serve as targets for novel antimycobacterial treatment. The cytochrome P450 enzyme, CYP51, catalyzes an essential early step in sterol metabolism, removing a methyl group from lanosterol in animals and fungi, or from obtusifoliol in plants. Azoles inhibit CYP51 function, leading to an accumulation of methylated sterol precursors. This perturbation of normal sterol metabolism compromises cell membrane integrity, resulting in growth inhibition or cell death. We have cloned and characterized a CYP51 from MAC that functions as a lanosterol 14α-demethylase. We show the direct interactions of azoles with purified MAC-CYP51 by absorbance and electron paramagnetic resonance spectroscopy, and determine the minimum inhibitory concentrations (MICs) of econazole, ketoconazole, itraconazole, fluconazole, and voriconazole against MAC. Furthermore, we demonstrate that econazole has a MIC of 4 μg/ml and a minimum bacteriocidal concentration of 4 μg/ml, whereas ketoconazole has a MIC of 8 μg/ml and a minimum bacteriocidal concentration of 16 μg/ml. Itraconazole, voriconazole, and fluconazole did not inhibit MAC growth to any significant extent.

Cumulative mtDNA damage occurs in aging animals, and mtDNA mutations are reported to accelerate aging in mice. We determined whether aging results in increased DNA oxidative damage and reduced mtDNA abundance and mitochondrial function in skeletal muscle of human subjects. Studies performed in 146 healthy men and women aged 18–89 yr demonstrated that mtDNA and mRNA abundance and mitochondrial ATP production all declined with advancing age. Abundance of mtDNA was positively related to mitochondrial ATP production rate, which in turn, was closely associated with aerobic capacity and glucose tolerance. The content of several mitochondrial proteins was reduced in older muscles, whereas the level of the oxidative DNA lesion, 8-oxo-deoxyguanosine, was increased, supporting the oxidative damage theory of aging. These results demonstrate that age-related muscle mitochondrial dysfunction is related to reduced mtDNA and muscle functional changes that are common in the elderly.