High-dose atorvastatin therapy progressively decreases skeletal muscle mitochondrial respiratory capacity in humans.

BACKGROUNDWhile the benefits of statin therapy on atherosclerotic cardiovascular disease are clear, patients often experience mild to moderate skeletal myopathic symptoms, the mechanism for which is unknown. This study investigated the potential effect of high-dose atorvastatin therapy on skeletal muscle mitochondrial function and whole-body aerobic capacity in humans.METHODSEight overweight (BMI, 31.9 ± 2.0) but otherwise healthy sedentary adults (4 females, 4 males) were studied before (day 0) and 14, 28, and 56 days after initiating atorvastatin (80 mg/d) therapy.RESULTSMaximal ADP-stimulated respiration, measured in permeabilized fiber bundles from muscle biopsies taken at each time point, declined gradually over the course of atorvastatin treatment, resulting in > 30% loss of skeletal muscle mitochondrial oxidative phosphorylation capacity by day 56. Indices of in vivo muscle oxidative capacity (via near-infrared spectroscopy) decreased by 23% to 45%. In whole muscle homogenates from day 0 biopsies, atorvastatin inhibited complex III activity at midmicromolar concentrations, whereas complex IV activity was inhibited at low nanomolar concentrations.CONCLUSIONThese findings demonstrate that high-dose atorvastatin treatment elicits a striking progressive decline in skeletal muscle mitochondrial respiratory capacity, highlighting the need for longer-term dose-response studies in different patient populations to thoroughly define the effect of statin therapy on skeletal muscle health.FUNDINGNIH R01 AR071263.

Genetically increasing flux through ß-oxidation in skeletal muscle increases mitochondrial reductive stress and glucose intolerance.

Elevated mitochondrial hydrogen peroxide (H2O2) emission and an oxidative shift in cytosolic redox environment have been linked to high-fat-diet-induced insulin resistance in skeletal muscle. To test specifically whether increased flux through mitochondrial fatty acid oxidation, in the absence of elevated energy demand, directly alters mitochondrial function and redox state in muscle, two genetic models characterized by increased muscle ß-oxidation flux were studied. In mice overexpressing peroxisome proliferator-activated receptor-α in muscle (MCK-PPARα), lipid-supported mitochondrial respiration, membrane potential (ΔΨm), and H2O2 production rate (JH2O2) were increased, which coincided with a more oxidized cytosolic redox environment, reduced muscle glucose uptake, and whole body glucose intolerance despite an increased rate of energy expenditure. Similar results were observed in lipin-1-deficient, fatty-liver dystrophic mice, another model characterized by increased ß-oxidation flux and glucose intolerance. Crossing MCAT (mitochondria-targeted catalase) with MCK-PPARα mice normalized JH2O2 production, redox environment, and glucose tolerance, but surprisingly, both basal and absolute insulin-stimulated rates of glucose uptake in muscle remained depressed. Also surprising, when placed on a high-fat diet, MCK-PPARα mice were characterized by much lower whole body, fat, and lean mass as well as improved glucose tolerance relative to wild-type mice, providing additional evidence that overexpression of PPARα in muscle imposes more extensive metabolic stress than experienced by wild-type mice on a high-fat diet. Overall, the findings suggest that driving an increase in skeletal muscle fatty acid oxidation in the absence of metabolic demand imposes mitochondrial reductive stress and elicits multiple counterbalance metabolic responses in an attempt to restore bioenergetic homeostasis.NEW & NOTEWORTHY Prior work has suggested that mitochondrial dysfunction is an underlying cause of insulin resistance in muscle because it limits fatty acid oxidation and therefore leads to the accumulation of cytotoxic lipid intermediates. The implication has been that therapeutic strategies to accelerate ß-oxidation will be protective. The current study provides evidence that genetically increasing flux through ß-oxidation in muscle imposes reductive stress that is not beneficial but rather detrimental to metabolic regulation.

17ß-Estradiol Directly Lowers Mitochondrial Membrane Microviscosity and Improves Bioenergetic Function in Skeletal Muscle.

Menopause results in a progressive decline in 17ß-estradiol (E2) levels, increased adiposity, decreased insulin sensitivity, and a higher risk for type 2 diabetes. Estrogen therapies can help reverse these effects, but the mechanism(s) by which E2 modulates susceptibility to metabolic disease is not well understood. In young C57BL/6N mice, short-term ovariectomy decreased-whereas E2 therapy restored-mitochondrial respiratory function, cellular redox state (GSH/GSSG), and insulin sensitivity in skeletal muscle. E2 was detected by liquid chromatography-mass spectrometry in mitochondrial membranes and varied according to whole-body E2 status independently of ERα. Loss of E2 increased mitochondrial membrane microviscosity and H2O2 emitting potential, whereas E2 administration in vivo and in vitro restored membrane E2 content, microviscosity, complex I and I + III activities, H2O2 emitting potential, and submaximal OXPHOS responsiveness. These findings demonstrate that E2 directly modulates membrane biophysical properties and bioenergetic function in mitochondria, offering a direct mechanism by which E2 status broadly influences energy homeostasis.

Myotonic Dystrophy Type 1 Management and Therapeutics.

OPINION STATEMENT: Myotonic dystrophy (DM1) is the most common form of adult muscular dystrophy. It is a multisystem disorder with a complex pathophysiology. Although inheritance is autosomal dominant, disease variability is attributed to anticipation, a maternal expansion bias, variable penetrance, somatic mosaicism, and a multitude of aberrant pre-mRNA splicing events. Patient presentations range from asymptomatic or mild late onset adult to severe congenital forms. Multiple organ systems may be affected. Patients may experience early cataracts, myotonia, muscle weakness/atrophy, fatigue, excessive daytime sleepiness, central/obstructive apnea, respiratory failure, cardiac arrhythmia, insulin resistance, dysphagia, GI dysmotility, cognitive impairment, Cluster C personality traits, and/or mood disorders. At present, there is no curative or disease-modifying treatment, although clinical treatment trials have become more promising. Management focuses on genetic counseling, preserving function and independence, preventing cardiopulmonary complications, and symptomatic treatment (e.g., pain, myotonia, hypersomnolence, etc.). Currently, there is an increasing international consensus on monitoring and treatment options for these patients which necessitates a multidisciplinary team to provide comprehensive, coordinated clinical care.

Constitutive activation of CaMKKα signaling is sufficient but not necessary for mTORC1 activation and growth in mouse skeletal muscle.

Skeletal muscle loading/overload stimulates the Ca²âº-activated, serine/threonine kinase Ca²âº/calmodulin-dependent protein kinase kinase-α (CaMKKα); yet to date, no studies have examined whether CaMKKα regulates muscle growth. The purpose of this study was to determine if constitutive activation of CaMKKα signaling could stimulate muscle growth and if so whether CaMKKα is essential for this process. CaMKKα signaling was selectively activated in mouse muscle via expression of a constitutively active form of CaMKKα using in vivo electroporation. After 2 wk, constitutively active CaMKKα expression increased muscle weight (~10%) and protein content (~10%), demonstrating that activation of CaMKKα signaling can stimulate muscle growth. To determine if active CaMKKα expression stimulated muscle growth via increased mammalian target of rapamycin complex 1 (mTORC1) signaling and protein synthesis, [³H]phenylalanine incorporation into proteins was assessed with or without the mTORC1 inhibitor rapamycin. Constitutively active CaMKKα increased protein synthesis ~60%, and this increase was prevented by rapamycin, demonstrating a critical role for mTORC1 in this process. To determine if CaMKKα is essential for growth, muscles from CaMKKα knockout mice were stimulated to hypertrophy via unilateral ablation of synergist muscles (overload). Surprisingly, compared with wild-type mice, muscles from CaMKKα knockout mice exhibited greater growth (~15%) and phosphorylation of the mTORC1 substrate 70-kDa ribosomal protein S6 kinase (Thr³89; ~50%), demonstrating that CaMKKα is not essential for overload-induced mTORC1 activation or muscle growth. Collectively, these results demonstrate that activation of CaMKKα signaling is sufficient but not necessary for activation of mTORC1 signaling and growth in mouse skeletal muscle.

Constitutively active CaMKKα stimulates skeletal muscle glucose uptake in insulin-resistant mice in vivo.

In insulin-sensitive skeletal muscle, the expression of constitutively active Ca(2+)/calmodulin-dependent protein kinase kinase α (caCaMKKα) stimulates glucose uptake independent of insulin signaling (i.e., Akt and Akt-dependent TBC1D1/TBC1D4 phosphorylation). Our objectives were to determine whether caCaMKKα could stimulate glucose uptake additively with insulin in insulin-sensitive muscle, in the basal state in insulin-resistant muscle, and if so, to determine whether the effects were associated with altered TBC1D1/TBC1D4 phosphorylation. Mice were fed a control or high-fat diet (60% kcal) for 12 weeks to induce insulin resistance. Muscles were transfected with empty vector or caCaMKKα plasmids using in vivo electroporation. After 2 weeks, caCaMKKα protein was robustly expressed. In insulin-sensitive muscle, caCaMKKα increased basal in vivo [(3)H]-2-deoxyglucose uptake approximately twofold, insulin increased glucose uptake approximately twofold, and caCaMKKα plus insulin increased glucose uptake approximately fourfold. caCaMKKα did not increase basal TBC1D1 (Ser(237), Thr(590), Ser(660), pan-Thr/Ser) or TBC1D4 (Ser(588), Thr(642), pan-Thr/Ser) phosphorylation. In insulin-resistant muscle, caCaMKKα increased basal glucose uptake approximately twofold, and attenuated high-fat diet-induced basal TBC1D1 (Thr(590), pan-Thr/Ser) and TBC1D4 (Ser(588), Thr(642), pan-Thr/Ser) phosphorylation. In cell-free assays, CaMKKα increased TBC1D1 (Thr(590), pan-Thr/Ser) and TBC1D4 (Ser(588), pan-Thr/Ser) phosphorylation. Collectively, these results demonstrate that caCaMKKα stimulates glucose uptake additively with insulin, and in insulin-resistant muscle, and alters the phosphorylation of TBC1D1/TBC1D4.

Forgiveness of others and health: do race and neighborhood matter?

OBJECTIVES: This study examines the relationship between interpersonal forgiveness and health for older Blacks and Whites. We outline a series of arguments concerning the following: (a) how forgiveness can affect health, (b) how forgiveness may be more protective for Blacks, and (c) how the relationship between forgiveness and health may vary by neighborhood deterioration. METHOD: Two waves (2001 and 2004) of the Religion, Aging, and Health Survey provided data from a nationally representative elderly sample of 436 Blacks and 500 Whites. Measures included sociodemographics, forgiveness, and three dimensions of health: self-reported health, alcohol use, and chronic conditions. We employ both longitudinal and cross-sectional analyses. RESULTS: Results suggest that forgiveness of others was protective of health for Blacks but not Whites. Moreover, among Blacks, we found the following: (a) forgiveness was positively associated with self-reported health over time, (b) forgiveness was negatively associated with alcohol use and number of chronic conditions, and (c) forgiveness interacted with neighborhood deterioration such that the beneficial effects of forgiveness for self-reported health did not extend to those living in run-down neighborhoods. DISCUSSION: Race and neighborhood were shown to be important for understanding the forgiveness-health connection. Forgiveness was associated with better health for Blacks but not Whites, consistent with McCullough's evolutionary framework (McCullough, M. E. (2008). Beyond revenge: The evolution of the forgiveness instinct. San Francisco, CA: Jossey-Bass), forgiveness was beneficial in some settings but had a deleterious impact in more noxious environments. This study suggests that researchers should give more consideration to race and social context in attempting to more fully understand the relationship between forgiveness and health.

Renal nitric oxide production in rat pregnancy: role of constitutive nitric oxide synthases.

Functional studies show that increased renal nitric oxide (NO) mediates the renal vasodilation and increased glomerular filtration rate that occur during normal pregnancy. We investigated whether changes in the constitutive NO synthases (NOS), endothelial (eNOS) and neuronal (nNOS), were associated with the increased renal NO production in normal midterm pregnancy in the rat. In kidneys from midterm pregnant (MP: 11-13 days gestation), late-term pregnant (LP: 18-20 days gestation), and similarly aged virgin (V) rats, transcript and protein abundance for eNOS and the nNOSα and nNOSß splice variants, as well as the rate of L-arginine-to-L-citrulline conversion, were determined as a measure of NOS activity. At MP, renal cortical abundance of the total eNOS protein and phosphorylated (Ser(1177)) eNOS was reduced, and L-arginine-to-L-citrulline conversion in the cortical membrane fraction was decreased; these declines were also seen in LP. There were no changes in the eNOS transcript. In contrast, L-arginine-to-L-citrulline conversion in the soluble fraction of renal cortex increased at MP and then declined at LP. This MP increase was ablated by S-methylthiocitrulline, a nNOS inhibitor. Using Western blotting, we did not detect a change in the protein abundance or transcript of the 160-kDa nNOSα, but protein abundance and transcript of the nNOSß were increased at MP in cortex. Collectively, these studies suggest that the soluble nNOSß is responsible for the increased renal cortical NO production during pregnancy.

Transforming growth factor-beta following skeletal muscle strain injury in rats.

Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine implicated in inflammatory processes, wound healing, and fibrosis. In muscle diseases (i.e., dystrophy and inflammatory myopathy) and in animal models of muscle injury (i.e., produced by cardiotoxin, laceration, and eccentric contractions), increased TGF-beta was associated with muscle fibrosis and healing. Although TGF-beta transcript abundance was increased following injury, many studies presume that TGF-beta protein was also active as evident by increases in collagen transcript abundance. The purpose was to determine whether TGF-beta protein is present and active 48 h following injury. Using female rats, muscle strains were produced by stretching (50 stretches) the plantar flexor muscles. Forty-eight hours following injury, the medial gastrocnemius was removed and compartmentalized into five equal segments. Damaged myofibers with intracellular concanavalin A staining were counted. The percentage of damaged myofibers was significantly greater in the distal-most segment. TGF-beta was assessed by using immunohistochemistry, RT-PCR, and immunoblot analysis. Immunohistochemistry revealed the presence of TGF-beta1 in areas of myofiber injury, whereas TGF-beta2 was not detected. Increases in TGF-beta1 and TGF-beta2 transcript abundance following strain injury were documented by RT-PCR analysis. Increases in TGF-beta1 and TGF-beta2 precursor abundance were observed following strain injury by using immunoblot analysis but there was no change in active TGF-beta abundance. Although there was no correlation between the amount of cellular injury and TGF-beta transcript and protein abundance, elevated levels of TGF-beta1 and TGF-beta2 precursor proteins were present in strain-injured skeletal muscles 48 h after injury.

Effects of exercise and creatine on myosin heavy chain isoform composition in patients with Charcot-Marie-Tooth disease.

It is not known whether myosin heavy chain (MHC) content changes in response to exercise training or creatine supplementation in subjects with Charcot-Marie-Tooth disease (CMT). Based on previous data, we hypothesized that resistance exercise and creatine would increase the percentage of type I MHC composition in the vastus lateralis muscle and that myosin isoform changes would correlate with improved chair rise-time in CMT subjects. To test this hypothesis, 18 CMT subjects were randomly assigned to either a placebo or creatine group. All subjects performed a 12-week, home-based, moderate-intensity resistance training program. Chair rise-time was measured before and after the training program. Muscle biopsies were obtained from the vastus lateralis before and after the 12-week program. Gel electrophoresis showed a significant decrease (approximately 30%) in MHC type I in CMT subjects given creatine supplementation when compared with placebo. There was a nonsignificant increase in both MHC type IIa (approximately 23%) and MHC type IIx (approximately 7%) in CMT subjects given creatine. Reduced MHC type I content and increased MHC type IIa content correlated with faster chair rise-times (i.e., improved muscle performance). The training-induced change in MHC IIa content was inversely correlated with chair rise-time in CMT subjects given creatine. When the two subject groups were combined, there was a linear, negative relationship between the change in MHC type IIa content and chair rise-time after training and a positive relationship between the training-induced change in MHC type I content and chair rise-time. These data suggest that improved function (chair rise-time) was associated with a lower level of MHC type I and increased MHC type IIa composition. Furthermore, the data are consistent with the hypothesis that creatine supplementation alters MHC composition in CMT patients undergoing resistance training and that MHC changes associated with creatine supplementation can improve muscle function.

Neurointensivists' opinions about death by neurological criteria and organ donation.

INTRODUCTION: Neurointensivists are at the front line of treatment of patients who progress to death by neurological criteria (DNC). Although some of these patients will become organ donors, there has not been a systematic evaluation of the opinions and resources available to neurointensivists in regard to these important issues. METHODS: We conducted a survey of neurointensivists regarding controversial issues in the declaration of DNC, procedures for discussing death and approaching donor families, and participation in donation after cardiac death (DCD). RESULTS: The majority of centers described by the respondents had all five most commonly accepted ancillary tests to determine DNC (61%). Radionuclide blood flow studies are the most frequently reported test used (64%). Younger physicians are more likely to use trans-cranial Doppler exams (TCD) than their older counterparts (41% versus 28%, p<0.001). Discussions about DNC with the family are most often presided by the attending physician, and donation requests are most commonly initiated by organ procurement organization (OPO) representatives, but there is significant variation from center to center. Nine out of 10 physicians in our survey reported that they are likely to participate in DCD. CONCLUSION: Despite this enthusiasm, there is no clear consensus on many of the issues surrounding DCD, including how long after cardiac cessation recovery should begin. We believe that this study will serve as a springboard for more discussion about the diagnosis of DNC, the role of physicians in organ requests and donor management, and the procurement of organs through DCD.

Potential role for Id myogenic repressors in apoptosis and attenuation of hypertrophy in muscles of aged rats.

Aging attenuates the overload-induced increase in myogenic regulatory transcription factor (MRF) expression and the extent of muscle enlargement. To identify whether mRNA levels of repressors of the MRFs are greater in overloaded muscles from aged animals, overload was achieved in plantaris muscle of aged (33 mo; n = 14) and adult (9 mo; n = 17) rats. After 14 days, plantaris muscles in the overloaded limb were ~25% and 6% larger in adult and aged rats, respectively, compared with the contralateral limb. Hypertrophied muscles of adult rats had significantly greater levels of mRNA and protein levels for myogenin and MyoD compared with control muscles, but neither MRF increased with overload in muscles of aged rats. Muscles of aged rats had greater Id mRNA (150-700%) and protein repressor (200-6,000%) levels compared with adult rats. BAX and caspase 9 protein levels were 9,500% and 300% greater, respectively, in both control and hypertrophied muscles of aged rats compared with young adult rats. These data are consistent with the hypothesis that aging increases Id transcripts that activate apoptotic pathways involving BAX. This may contribute to sarcopenia by attenuating MRF protein levels in muscles of old animals.