Excess Intramyocellular Lipid Does Not Affect Muscle Fiber Biophysical Properties in Mice or People with Metabolically Abnormal Obesity.

Observational studies show correlations between intramyocellular lipid (IMCL) content and muscle strength and contractile function in people with "metabolically abnormal" obesity. However, a clear physiologic mechanism for this association is lacking and causation is debated. We combined immunofluorescent confocal imaging with force measurements on permeabilized muscle fibers from metabolically normal and metabolically abnormal mice and metabolically normal (defined as normal fasting plasma glucose and glucose tolerance) and metabolically abnormal (defined as pre-diabetes and type 2 diabetes) people with overweight/obesity to evaluate relationships among myocellular lipid droplet characteristics (droplet size and density) and biophysical (active contractile and passive viscoelastic) properties. The fiber type specificity of lipid droplet parameters varied between metabolically abnormal and normal mice and among metabolically normal and metabolically abnormal people. However, despite considerable quantities of IMCL in the metabolically abnormal groups, there were no significant differences in peak active tension or passive viscoelasticity between the metabolically abnormal groups and the control group in mice or people. Additionally, there were no significant relationships among IMCL parameters and biophysical variables. Thus, we conclude that IMCL accumulation per se does not impact muscle fiber biophysical properties or physically impede contraction.

Identification of a leucine-mediated threshold effect governing macrophage mTOR signalling and cardiovascular risk.

High protein intake is common in western societies and is often promoted as part of a healthy lifestyle; however, amino-acid-mediated mammalian target of rapamycin (mTOR) signalling in macrophages has been implicated in the pathogenesis of ischaemic cardiovascular disease. In a series of clinical studies on male and female participants ( NCT03946774 and NCT03994367 ) that involved graded amounts of protein ingestion together with detailed plasma amino acid analysis and human monocyte/macrophage experiments, we identify leucine as the key activator of mTOR signalling in macrophages. We describe a threshold effect of high protein intake and circulating leucine on monocytes/macrophages wherein only protein in excess of ∼25 g per meal induces mTOR activation and functional effects. By designing specific diets modified in protein and leucine content representative of the intake in the general population, we confirm this threshold effect in mouse models and find ingestion of protein in excess of ∼22% of dietary energy requirements drives atherosclerosis in male mice. These data demonstrate a mechanistic basis for the adverse impact of excessive dietary protein on cardiovascular risk.

Dietary protein intake and obesity-associated cardiometabolic function.

PURPOSE OF REVIEW: High-protein intake is commonly recommended to help people manage body weight. However, high-protein intake could have adverse health consequences. Here we review the latest findings concerning the effect of high-protein intake on cardiometabolic health. RECENT FINDINGS: Calorie-reduced, high-protein, low-carbohydrate diets lower plasma glucose in people with type 2 diabetes (T2D). However, when carbohydrate intake is not markedly reduced, high-protein intake often does not alter plasma glucose and increases insulin and glucagon concentrations, which are risk factors for T2D and ischemic heart disease. High-protein intake does not alter plasma triglyceride and cholesterol concentrations but promotes atherogenesis in animal models. The effect of high-protein intake on liver fat remains unclear. In population studies, high-protein intake is associated with increased risk for T2D, nonalcoholic fatty liver disease, and possibly cardiovascular diseases. SUMMARY: The relationship between protein intake and cardiometabolic health is complex and influenced by concomitant changes in body weight and overall diet composition. Although a high-protein, low-carbohydrate, reduced-energy diet can have beneficial effects on body weight and plasma glucose, habitual high-protein intake, without marked carbohydrate and energy restriction, is associated with increased cardiometabolic disease risk, presumably mediated by the changes in the hormonal milieu after high-protein intake.

No independent or combined effects of vitamin D and conjugated linoleic acids on muscle protein synthesis in older adults: a randomized, double-blind, placebo-controlled clinical trial.

BACKGROUND: Aging is associated with skeletal muscle anabolic resistance (i.e., reduced muscle protein synthesis during anabolic conditions such as hyperaminoacidemia). The results from studies conducted in cell culture systems and animals suggest that both vitamin D and conjugated linoleic acids (CLAs) stimulate muscle protein synthesis. OBJECTIVES: To conduct a randomized, double-blind, placebo-controlled clinical trial to determine the independent and combined effects of dietary vitamin D and CLA supplementation on myofibrillar protein synthesis rates in sedentary older adults. METHODS: Thirty-two sedentary, older adults were randomized to receive either: 1) 2000 IU vitamin D-3 (Vit D) per day; 2) 4000 mg CLA per day; 3) both Vit D (2000 IU/d) and CLA (4000 mg/d); or 4) placebo for 8 wk. Myofibrillar protein synthesis rates were evaluated by using intravenous [ring-2H5]phenylalanine infusion in conjunction with muscle biopsies during basal, postabsorptive conditions and during combined amino acid and insulin infusion before and after the supplementation period. RESULTS: Before the intervention, basal myofibrillar protein synthesis rates were not different among groups (Placebo: 0.033 ± 0.003; Vit D: 0.034 ± 0.002; CLA: 0.029 ± 0.005; Vit D + CLA: 0.038 ± 0.005 %·h-1), and hyperinsulinemia-hyperaminoacidemia increased myofibrillar protein synthesis rates by ∼35%. Compared with placebo, neither Vit D nor CLA nor combined Vit D + CLA supplementation affected the basal myofibrillar protein synthesis rates (placebo: 0.040 ± 0.004%/h; Vit D: 0.044 ± 0.006%/h; CLA: 0.039 ± 0.006%/h; Vit D + CLA: 0.040 ± 0.007%/h) or the hyperinsulinemia-hyperaminoacidemia-induced increase in myofibrillar protein synthesis (percentage increase from basal before and after the interventions: placebo, 30 ± 11 and 36 ± 11; Vit D, 38 ± 8 and 34 ± 10; CLA, 50 ± 14 and 51 ± 16; Vit D + CLA, 29 ± 15 and 35 ± 8). CONCLUSIONS: Vitamin D and/or CLA supplementation, at the doses provided in our study, does not have muscle anabolic effects in sedentary older adults.The study was registered at clinicaltrials.gov (NCT03115775).

Different physiological mechanisms underlie an adverse cardiovascular disease risk profile in men and women.

CVD affect about one-third of the population and are the leading cause of mortality. The prevalence of CVD is closely linked to the prevalence of obesity because obesity is commonly associated with metabolic abnormalities that are important risk factors for CVD, including insulin resistance, pre-diabetes, and type-2 diabetes, atherosclerotic dyslipidaemia, endothelial dysfunction and hypertension. Women have a more beneficial traditional CVD risk profile (lower fasting plasma glucose, less atherogenic lipid profile) and a lower absolute risk for CVD than men. However, the relative risk for CVD associated with hyperglycaemia and dyslipidaemia is several-fold higher in women than in men. The reasons for the sex differences in CVD risk associated with metabolic abnormalities are unclear but could be related to differences in the mechanisms that cause hyperglycaemia and dyslipidaemia in men and women, which could influence the pathogenic processes involved in CVD. In the present paper, we review the influence of a person's sex on key aspects of metabolism involved in the cardiometabolic disease process, including insulin action on endogenous glucose production, tissue glucose disposal, and adipose tissue lipolysis, insulin secretion and insulin plasma clearance, postprandial glucose, fatty acid, and triglyceride kinetics, hepatic lipid metabolism and myocardial substrate use. We conclude that there are marked differences in many aspects of metabolism in men and women that are not all attributable to differences in the sex hormone milieu. The mechanisms responsible for these differences and the clinical implications of these observations are unclear and require further investigation.

Congenital fiber type disproportion caused by TPM3 mutation: A report of two atypical cases.

Congenital fiber type disproportion (CFTD) is a rare congenital myopathy subtype defined by slow type 1 hypotrophy in the absence of any other major structural findings such as rods, central nuclei or cores. Dominant missense changes in slow alpha-tropomyosin coded by TPM3 gene are the main cause of the CFTD. There are only a few reports of recessive loss-of-function mutations in TPM3 causing severe Nemaline Myopathy and CFTD. We present two patients harboring TPM3 mutations. The first is a novel homozygous missense variant with a mild CFTD clinical phenotype inherited in a recessive fashion. The second is a previously reported heterozygous mutation presenting within pronounced early axial involvement and dropped head. This report expands the genotype-phenotype correlation in the TPM3 myopathy showing a recessive mutation causing a mild clinical phenotype and also shows that TPM3 mutations should be part of the investigation in patients with dropped head.

Skeletal Muscle Response to Deflazacort, Dexamethasone and Methylprednisolone.

Glucocorticoids represent some of the most prescribed drugs that are widely used in the treatment of neuromuscular diseases, but their usage leads to side effects such as muscle atrophy. However, different synthetic glucocorticoids can lead to different muscle effects, depending upon its chemical formulation. Here, we intended to demonstrate the muscle histologic and molecular effects of administering different glucocorticoids in equivalency and different dosages. Methods: Seventy male Wistar rats distributed into seven groups received different glucocorticoids in equivalency for ten days or saline solution. The study groups were: Control group (CT) saline solution; dexamethasone (DX) 1.25 or 2.5 mg/kg/day; methylprednisolone (MP) 6.7 or 13.3mg/kg/day; and deflazacort (DC) 10 or 20 mg/kg/day. At the end of the study, the animals were euthanized, and the tibialis anterior and gastrocnemius muscles were collected for metachromatic ATPase (Cross-sectional area (CSA) measurement), Western blotting (protein expression of IGF-1 and Ras/Raf/MEK/ERK pathways) and RT-PCR (MYOSTATIN, MuRF-1, Atrogin-1, REDD-1, REDD-2, MYOD, MYOG and IRS1/2 genes expression) experiments. Results: Muscle atrophy occurred preferentially in type 2B fibers in all glucocorticoid treated groups. DC on 10 mg/kg/day was less harmful to type 2B fibers CSA than other doses and types of synthetic glucocorticoids. In type 1 fibers CSA, lower doses of DC and DX were more harmful than high doses. DX had a greater effect on the IGF-1 pathway than other glucocorticoids. MP more significantly affected P-ERK1/2 expression, muscle fiber switching (fast-to-slow), and expression of REDD1 and MyoD genes than other glucocorticoids. Compared to DX and MP, DC had less of an effect on the expression of atrogenes (MURF-1 and Atrogin-1) despite increased MYOSTATIN and decreased IRS-2 genes expression. Conclusions: Different glucocorticoids appears to cause muscle atrophy affecting secondarily different signaling mechanisms. MP is more likely to affect body/muscles mass, MEK/ERK pathway and fiber type transition, DX the IGF-1 pathway and IRS1/2 expression. DC had the smallest effect on muscle atrophic response possibly due a delayed timing on atrogenes response.

Omega-3 multiple effects increasing glucocorticoid-induced muscle atrophy: autophagic, AMPK and UPS mechanisms.

Muscle atrophy occurs in many conditions, including use of glucocorticoids. N-3 (omega-3) is widely consumed due its healthy properties; however, concomitant use with glucocorticoids can increase its side effects. We evaluated the influences of N-3 on glucocorticoid atrophy considering IGF-1, Myostatin, MEK/ERK, AMPK pathways besides the ubiquitin-proteasome system (UPS) and autophagic/lysosomal systems. Sixty animals constituted six groups: CT, N-3 (EPA 100 mg/kg/day for 40 days), DEXA 1.25 (DEXA 1.25 mg/kg/day for 10 days), DEXA 1.25 + N3 (EPA for 40 days + DEXA 1.25 mg/kg/day for the last 10 days), DEXA 2.5 (DEXA 2.5 mg/kg/day for 10 days), and DEXA 2.5 + N3 (EPA for 40 days + DEXA 2.5 mg/kg/day for 10 days). Results: N-3 associated with DEXA increases atrophy (fibers 1 and 2A), FOXO3a, P-SMAD2/3, Atrogin-1/MAFbx (mRNA) expression, and autophagic protein markers (LC3II, LC3II/LC3I, LAMP-1 and acid phosphatase). Additionally, N-3 supplementation alone decreased P-FOXO3a, PGC1-alpha, and type 1 muscle fiber area. Conclusion: N-3 supplementation increases muscle atrophy caused by DEXA in an autophagic, AMPK and UPS process.

Neuraminidase-1 mediates skeletal muscle regeneration.

Neuraminidase-1 (NEU1) is the sialidase responsible for the catabolism of sialoglycoconjugates in lysosomes. Congenital NEU1 deficiency causes sialidosis, a severe lysosomal storage disease associated with a broad spectrum of clinical manifestations, which also include skeletal deformities, skeletal muscle hypotonia and weakness. Neu1(-/-) mice, a model of sialidosis, develop an atypical form of muscle degeneration caused by progressive expansion of the connective tissue that infiltrates the muscle bed, leading to fiber degeneration and atrophy. Here we investigated the role of Neu1 in the myogenic process that ensues during muscle regeneration after cardiotoxin-induced injury of limb muscles. A comparative analysis of cardiotoxin-treated muscles from Neu1(-/-) mice and Neu1(+/+) mice showed increased inflammatory and proliferative responses in the absence of Neu1 during the early stages of muscle regeneration. This was accompanied by significant and sequential upregulation of Pax7, MyoD, and myogenin mRNAs. The levels of both MyoD and myogenin proteins decreased during the late stages of regeneration, which most likely reflected an increased rate of degradation of the myogenic factors in the Neu1(-/-) muscle. We also observed a delay in muscle cell differentiation, which was characterized by prolonged expression of embryonic myosin heavy chain, as well as reduced myofiber cross-sectional area. At the end of the regenerative process, collagen type III deposition was increased compared to wild-type muscles and internal controls, indicating the initiation of fibrosis. Overall, these results point to a role of Neu1 throughout muscle regeneration.

The effects of omega-3 fatty acid supplementation on dexamethasone-induced muscle atrophy.

Corticosteroids cause muscle atrophy by acting on proteasomal and lysosomal systems and by affecting pathways related to muscular trophysm, such as the IGF-1/PI-3k/Akt/mTOR. Omega-3 fatty acid (n-3) has been used beneficially to attenuate muscle atrophy linked to sepsis and cachexia; however, its effect on dexamethasone-induced muscle atrophy has not been evaluated. Objectives. We evaluated whether n-3 supplementation could mitigate the development of dexamethasone-induced muscle atrophy. Methods. Two groups of Wistar rats were orally supplemented with n-3 or vehicle solution for 40 days. In the last 10 days, dexamethasone, or saline solution, was administrated establishing four groups: control, dexamethasone, n-3, and dexamethasone + n-3. The cross-sectional areas of muscle fibers, gene expression (MyoD, Myogenin, MuRF-1, and Atrogin-1), and protein expression (Akt, GSK3ß, FOXO3a, and mTOR) were assessed. Results. Dexamethasone induced a significant loss in body and muscle weight, atrophy in type 2B fibers, and decreased expression of P-Akt, P-GSK3ß, and P-FOXO3a. N-3 supplementation did not attenuate the negative effects of dexamethasone on skeletal muscle; instead, it caused atrophy in type 1, 2A, reduced the expression of Myogenin, and increased the expression of Atrogin-1. Conclusion. Food supplements containing n-3 are usually healthful, but they may potentiate some of the side effects of glucocorticoids.