Treadmill Running Changes Endothelial Lipase Expression: Insights from Gene and Protein Analysis in Various Striated Muscle Tissues and Serum

Endothelial lipase (EL) is an enzyme capable of HDL phospholipids hydrolysis. Its action leads to a reduction in the serum high-density lipoprotein concentration, and thus, it exerts a pro-atherogenic effect. This study examines the impact of a single bout exercise on the gene and protein expression of the EL in skeletal muscles composed of different fiber types (the soleus—mainly type I, the red gastrocnemius—mostly IIA, and the white gastrocnemius—predominantly IIX fibers), as well as the diaphragm, and the heart. Wistar rats were subjected to a treadmill run: 1) t = 30 [min], V = 18 [m/min]; 2) t = 30 [min], V = 28 [m/min]; 3) t = 120 [min], V = 18 [m/min] (designated: M30, F30, and M120, respectively). We established EL expression in the total muscle homogenates in sedentary animals. Resting values could be ordered with the decreasing EL protein expression as follows: endothelium of left ventricle > diaphragm > red gastrocnemius > right ventricle > soleus > white gastrocnemius. Furthermore, we observed that even a single bout of exercise was capable of inducing changes in the mRNA and protein level of EL, with a clearer pattern observed for the former. After 30 min of running at either exercise intensity, the expression of EL transcript in all the cardiovascular components of muscles tested, except the soleus, was reduced in comparison to the respective sedentary control. The protein content of EL varied with the intensity and/or duration of the run in the studied whole tissue homogenates. The observed differences between EL expression in vascular beds of muscles may indicate the muscle-specific role of the lipase.

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Identification of Nesfatin-1 in Human and Murine Adipose Tissue: A Novel Depot-Specific Adipokine with Increased Levels in Obesity

Endocrinology ◽

10.1210/en.2009-1358 ◽

2010 ◽

Vol 151 (7) ◽

pp. 3169-3180 ◽

Cited By ~ 178

Author(s):

Manjunath Ramanjaneya ◽

Jing Chen ◽

James E. Brown ◽

Gyanendra Tripathi ◽

Manfred Hallschmid ◽

...

Keyword(s):

Adipose Tissue ◽

Protein Expression ◽

Food Deprivation ◽

Inflammatory Cytokines ◽

Energy Homeostasis ◽

Culture Media ◽

Nonesterified Fatty Acid ◽

High Fat ◽

Gene And Protein Expression ◽

The Impact

Nesfatin-1 is a recently identified anorexigenic peptide derived from its precursor protein, nonesterified fatty acid/nucleobindin 2 (NUCB2). Although the hypothalamus is pivotal for the maintenance of energy homeostasis, adipose tissue plays an important role in the integration of metabolic activity and energy balance by communicating with peripheral organs and the brain via adipokines. Currently no data exist on nesfatin-1 expression, regulation, and secretion in adipose tissue. We therefore investigated NUCB2/nesfatin-1 gene and protein expression in human and murine adipose tissue depots. Additionally, the effects of insulin, dexamethasone, and inflammatory cytokines and the impact of food deprivation and obesity on nesfatin-1 expression were studied by quantitative RT-PCR and Western blotting. We present data showing NUCB2 mRNA (P < 0.001), nesfatin-1 intracellular protein (P < 0.001), and secretion (P < 0.01) were significantly higher in sc adipose tissue compared with other depots. Also, nesfatin-1 protein expression was significantly increased in high-fat-fed mice (P < 0.01) and reduced under food deprivation (P < 0.01) compared with controls. Stimulation of sc adipose tissue explants with inflammatory cytokines (TNFα and IL-6), insulin, and dexamethasone resulted in a marked increase in intracellular nesfatin-1 levels. Furthermore, we present evidence that the secretion of nesfatin-1 into the culture media was dramatically increased during the differentiation of 3T3-L1 preadipocytes into adipocytes (P < 0.001) and after treatments with TNF-α, IL-6, insulin, and dexamethasone (P < 0.01). In addition, circulating nesfatin-1 levels were higher in high-fat-fed mice (P < 0.05) and showed positive correlation with body mass index in human. We report that nesfatin-1 is a novel depot specific adipokine preferentially produced by sc tissue, with obesity- and food deprivation-regulated expression.

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Uremia Impacts VE-Cadherin and ZO-1 Expression in Human Endothelial Cell-to-Cell Junctions

Toxins ◽

10.3390/toxins10100404 ◽

2018 ◽

Vol 10 (10) ◽

pp. 404 ◽

Cited By ~ 8

Author(s):

Rayana Maciel ◽

Regiane Cunha ◽

Valentina Busato ◽

Célia Franco ◽

Paulo Gregório ◽

...

Keyword(s):

Protein Expression ◽

Intercellular Junctions ◽

Serum Levels ◽

Endothelial Damage ◽

Cell Junctions ◽

Cell Junction ◽

Uremic Serum ◽

Gene And Protein Expression ◽

The Impact

Endothelial dysfunction in uremia can result in cell-to-cell junction loss and increased permeability, contributing to cardiovascular diseases (CVD) development. This study evaluated the impact of the uremic milieu on endothelial morphology and cell junction’s proteins. We evaluated (i) serum levels of inflammatory biomarkers in a cohort of chronic kidney disease (CKD) patients and the expression of VE-cadherin and Zonula Occludens-1 (ZO-1) junction proteins on endothelial cells (ECs) of arteries removed from CKD patients during renal transplant; (ii) ECs morphology in vitro under different uremic conditions, and (iii) the impact of uremic toxins p-cresyl sulfate (PCS), indoxyl sulfate (IS), and inorganic phosphate (Pi) as well as of total uremic serum on VE-cadherin and ZO-1 gene and protein expression in cultured ECs. We found that the uremic arteries had lost their intact and continuous endothelial morphology, with a reduction in VE-cadherin and ZO-1 expression. In cultured ECs, both VE-cadherin and ZO-1 protein expression decreased, mainly after exposure to Pi and uremic serum groups. VE-cadherin mRNA expression was reduced while ZO-1 was increased after exposure to PCS, IS, Pi, and uremic serum. Our findings show that uremia alters cell-to-cell junctions leading to an increased endothelial damage. This gives a new perspective regarding the pathophysiological role of uremia in intercellular junctions and opens new avenues to improve cardiovascular outcomes in CKD patients.

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Topical application of Chinese herbal medicine DAEP relieves the osteoarthritic knee pain in rats

Chinese Medicine ◽

10.1186/s13020-019-0278-1 ◽

2019 ◽

Vol 14 (1) ◽

Cited By ~ 2

Author(s):

Wing Sum Siu ◽

Wai Ting Shum ◽

Wen Cheng ◽

Chun Wai Wong ◽

Hoi Ting Shiu ◽

...

Keyword(s):

Knee Joint ◽

Protein Expression ◽

Knee Pain ◽

Topical Application ◽

Weight Ratio ◽

Treadmill Running ◽

Morphologic Change ◽

Chinese Herbal ◽

Iκb Kinase ◽

Gene And Protein Expression

Abstract Background The potential adverse effects of conventional oral pharmacotherapy of osteoarthritis (OA) restrict their long-term use. Topical application of a Chinese herbal paste for relieving OA knee pain can be effective and safe. However, evidence-based scientific research is insufficient to support its application worldwide. The aim of this study was to investigate the in vivo efficacy of a topical Chinese herbal paste on relieving OA knee pain and its underlying mechanism. Methods An OA rat model was developed by anterior cruciate ligament transection (ACLT) followed by treadmill running. A herbal paste including Dipsaci Radix, Achyranthis Bidentatae Radix, Eucommiae Cortex and Psoraleae Fructus, named as DAEP, was applied topically on the knee joint of the rats (DAEP). The rats without DAEP treatment served as Control. Rats with surgery but without ACLT, treadmill running and DAEP treatment acted as Sham. The morphologic change of the knee joint was observed radiographically. Nociception from the knee of the rats was assessed using Incapacitent test and CatWalk gait system. The therapeutic mechanism was investigated by analyzing the gene and protein expression of inflammatory markers via qPCR and Western blot, respectively. Results Radiographic images showed less destruction at the posterior tibial plateau of the DAEP group compared with the Control after 2 weeks of treatment. The static weight ratio and the gait parameters of the Control were reduced significantly via Incapacitance test and CatWalk gait analysis, respectively. DAEP treatment increased the Print Area and Maximum Intensity significantly compared with the Control. DAEP significantly suppressed the upregulation of gene expression of interleukin (IL)-6, tumor necrosis factor (TNF)-α, and inducible nitric oxide synthase (iNOS). Conclusions DAEP exhibited its effect via the nuclear factor (NF)-κB pathway by suppressing the phosphorylation of IκB kinase αβ (p-IKKαβ) and cyclooxygenase-2 (COX-2) protein expression. This study provides scientific evidence to support the clinical application of the Chinese herbal paste on reliving OA pain.

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Meprin-α in chronic diabetic nephropathy: interaction with the renin-angiotensin axis

AJP Renal Physiology ◽

10.1152/ajprenal.00037.2005 ◽

2005 ◽

Vol 289 (4) ◽

pp. F911-F921 ◽

Cited By ~ 23

Author(s):

Roy Mathew ◽

Stephen Futterweit ◽

Elsa Valderrama ◽

Antonio A. Tarectecan ◽

John E. Bylander ◽

...

Keyword(s):

Diabetic Nephropathy ◽

Kidney Disease ◽

Protein Expression ◽

Protein Content ◽

Ace Inhibitor ◽

Inhibitor Therapy ◽

Treated Water ◽

Renin Angiotensin ◽

Gene And Protein Expression ◽

The Impact

Meprin (MEP) A is a metalloendopeptidase that is present in the renal proximal tubule brush-border membrane (BBM) and that colocalizes with angiotensin-converting enzyme (ACE). The MEP β-chain gene locus on chromosome 18 has been linked to a heightened risk of diabetic nephropathy (DN) in patients with type 2 diabetes. This study evaluated 1) whether MEP-α and MEP-β gene and protein expression are altered in db/db mice before the onset of DN and 2) the role of MEP-α in the pathogenesis of DN and the impact of the renin-angiotensin system on this interaction in two experimental models of diabetes. MEP-α and MEP-β gene and protein expression were evaluated in db/db mice, 13–14 wk of age, compared with lean C57BLKS/J littermate animals. A treatment study was then performed in which db/db mice and controls were assigned to one of three groups: control (C) water, no therapy; ACE inhibitor therapy, enalapril (EN)-treated water, 50 mg/l; ANG II receptor type 1 blocker (ARB) therapy, losartan (LOS)-treated water, 500 mg/l. Treatment was started at 8 wk of age and continued for 52 wk. Male Sprague-Dawley rats with diabetes for 52 wk following a single dose of streptozocin (STZ; 60 mg/kg) were also studied. At 13.5 wk of age, MEP-α and MEP-β kidney mRNA abundance and protein expression were significantly lower in db/db mice compared with lean controls, with greater changes in MEP-β ( P < 0.05). In the treatment study, EN ameliorated and LOS exacerbated DN in db/db mice. BBM MEP A enzymatic activity and MEP-α protein content were lower in db/db mice vs. control nonobese mice at 52 wk ( P < 0.02). EN-treated db/db mice showed increased MEP A activity, MEP-α content in BBM, decreased urinary MEP-α excretion, and enhanced BBM staining for MEP-α protein vs. C and LOS-treated db/db mice. In nonobese mice, EN and LOS treatment had no effect on MEP-α expression. In rats with STZ-induced diabetes for 52 wk, urinary MEP-α excretion was increased and MEP A activity and MEP-α protein content per milligram of BBM protein were decreased compared with age-matched control animals ( P < 0.05). These results indicate that db/db mice manifest decreased MEP-α and MEP-β gene and protein expression, before the development of overt kidney disease. Moreover, in db/db mice with DN and rats with STZ-diabetes, there was an inverse relationship between renal MEP-α content and the severity of the renal injury. Treatment with an ACE inhibitor was more effective than ARB in ameliorating DN in db/db mice, a change that correlated with alterations in urinary excretion and BBM content of MEP-α. MEP-α may play a role in the pathogenesis of DN and the benefits of ACE inhibitor therapy on the progression of diabetic kidney disease may be related, in part, to its impact on renal MEP-α expression.

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Endothelial cells respond to hyperglycemia by increasing the LPL transporter GPIHBP1

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00007.2014 ◽

2014 ◽

Vol 306 (11) ◽

pp. E1274-E1283 ◽

Cited By ~ 12

Author(s):

Amy Pei-Ling Chiu ◽

Fulong Wang ◽

Nathaniel Lal ◽

Ying Wang ◽

Dahai Zhang ◽

...

Keyword(s):

Endothelial Cells ◽

Protein Expression ◽

Binding Sites ◽

Therapeutic Strategy ◽

Density Lipoprotein ◽

Streptozotocin Diabetes ◽

Gene And Protein Expression ◽

Lipoprotein Binding ◽

Rate Limiting

In diabetes, when glucose uptake and oxidation are impaired, the heart is compelled to use fatty acid (FA) almost exclusively for ATP. The vascular content of lipoprotein lipase (LPL), the rate-limiting enzyme that determines circulating triglyceride clearance, is largely responsible for this FA delivery and increases following diabetes. Glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein [GPIHBP1; a protein expressed abundantly in the heart in endothelial cells (EC)] collects LPL from the interstitial space and transfers it across ECs onto the luminal binding sites of these cells, where the enzyme is functional. We tested whether ECs respond to hyperglycemia by increasing GPIHBP1. Streptozotocin diabetes increased cardiac LPL activity and GPIHBP1 gene and protein expression. The increased LPL and GPIHBP1 were located at the capillary lumen. In vitro, passaging EC caused a loss of GPIHBP1, which could be induced on exposure to increasing concentrations of glucose. The high-glucose-induced GPIHBP1 increased LPL shuttling across EC monolayers. GPIHBP1 expression was linked to the EC content of heparanase. Moreover, active heparanase increased GPIHBP1 gene and protein expression. Both ECs and myocyte heparan sulfate proteoglycan-bound platelet-derived growth factor (PDGF) released by heparanase caused augmentation of GPIHBP1. Overall, our data suggest that this protein “ensemble” (heparanase-PDGF-GPIHBP1) cooperates in the diabetic heart to regulate FA delivery and utilization by the cardiomyocytes. Interrupting this axis may be a novel therapeutic strategy to restore metabolic equilibrium, curb lipotoxicity, and help prevent or delay heart dysfunction that is characteristic of diabetes.

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Inducible isoform of HSP70 is constitutively expressed in a muscle fiber type specific pattern

AJP Cell Physiology ◽

10.1152/ajpcell.1991.261.5.c774 ◽

1991 ◽

Vol 261 (5) ◽

pp. C774-C779 ◽

Cited By ~ 108

Author(s):

M. Locke ◽

E. G. Noble ◽

B. G. Atkinson

Keyword(s):

Muscle Fiber ◽

Fiber Type ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Constitutive Expression ◽

Muscle Fiber Types ◽

Type I ◽

Fiber Types ◽

Nylon Membrane ◽

White Gastrocnemius

The most prominent group of stress or heat-shock proteins (HSPs) has an Mr of approximately 70,000 and is collectively referred to as the HSP70 family. The extent of stress inducibility and subcellular location of the various HSP70 isoforms differ, but all appear to be involved with ATP-dependent stabilization or solubilization of proteins. One isoform, termed the inducible isoform of HSP70 (HSP72i), is normally absent in unstressed cells. In a previous study, we detected a protein corresponding in Mr and pI to HSP72i in unstressed rat muscle. Therefore, it was of interest to determine if this expression in unstressed muscle cells is general or confined to specific muscle fiber types. To answer this question we have employed various rat hindlimb muscles that differ in fiber type proportion from predominantly type I (soleus) to predominantly type IIB (white gastrocnemius). Proteins from muscle homogenates were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, blotted to a nylon membrane, probed with a monoclonal antibody for HSP72i, and visualized using an alkaline phosphatase-conjugated secondary antibody. Immunoblot analyses demonstrate the constitutive expression of HSP72i in rat muscles comprised primarily of type I muscle fibers (soleus), but not in muscles comprised primarily of type IIB fibers (white gastrocnemius). In muscles of mixed fiber type, HSP72i content is roughly proportional to the percentage of type I fibers. These results substantiate that unstressed rat muscles express the inducible HSP72 isoform and demonstrate that its constitutive expression is proportional to the type I muscle fiber composition.

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Ultrasound Tissue Characterization (UTC) of the Achilles Tendon in Pre- and Post-Pubertal Dancers

Journal of Dance Medicine & Science ◽

10.12678/1089-313x.24.2.51 ◽

2020 ◽

Vol 24 (2) ◽

pp. 51-58

Author(s):

Nili Steinberg ◽

Michal Pantanowitz ◽

Aviva Zeev ◽

Myriam Stern ◽

Gali Dar

Keyword(s):

Body Mass Index ◽

Achilles Tendon ◽

Body Mass ◽

Tissue Characterization ◽

Lower Percentage ◽

Fibrillar Structure ◽

Type I ◽

Fiber Types ◽

Echo Type ◽

The Impact

This study examined whether maturation status, body physique, and the impact of training are related to the development of Achilles tendon structure in young dancers. Seventy-one pre- and post-menarche dancers (12 to 15 years of age) were recruited. The Achilles tendon of each dancer was examined via ultrasonography tissue characterization (UTC) imaging. The cross-sectional area (CSA) and the fibrillar structure (echo types I to IV) were measured. The participants were screened for anthropometric parameters (weight, height, and leg length) with body mass index (BMI) and BMI percentile calculated; for hours and impact of training; for Tanner pubertal maturation; and for pain in their Achilles tendon (VAS scale). In addition, age and age at onset of menarche were documented. Tendon structure was found to differ between pre- and post-menarche dancers. Post-menarche dancers had a significantly lower percentage of echo type I fibers and a significantly higher percentage of echo type II, III, and IV fibers, with a greater CSA compared to pre-menarche dancers. The tendon structure was found to be correlated with BMI percentile, but no correlations were found with chronologic age or the impact of dance training. Furthermore, ANCOVA showed that BMI had a statistically significant effect on fiber types II and III (p < 0.005) and that the effect of menarche was significant, meaning that pre-menarche dancers had a lower BMI compared with those who were post-menarche. It is concluded that pre- and post-menarche dancers had developed different patterns of Achilles tendon fiber structure. Body mass index was found to be the most significant factor influencing the different tendon structures in young pubertal dancers.

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Hepatoprotective Effects of Different Extracts From Triphala Against CCl4-Induced Acute Liver Injury in Mice

Frontiers in Pharmacology ◽

10.3389/fphar.2021.664607 ◽

2021 ◽

Vol 12 ◽

Author(s):

Xichuan Wei ◽

Chuanhong Luo ◽

Yanan He ◽

Haozhou Huang ◽

Fei Ran ◽

...

Keyword(s):

Liver Injury ◽

Mouse Model ◽

Protein Expression ◽

Acute Liver Injury ◽

Quinone Oxidoreductase ◽

Tnf Α ◽

Gene And Protein Expression ◽

Wide Range ◽

Hepatoprotective Effects ◽

The Impact

Background:Triphala is a traditional polyherbal formula used in Indian Ayurvedic and Chinese Tibetan medicine. A wide range of biological activities have been attributed to Triphala, but the impact of various extraction methods on efficacy has not been determined.Purpose: The study aimed to evaluate Triphala extracts obtained by various methods for their hepatoprotective effects and molecular mechanisms in a mouse model of carbon tetrachloride (CCl4)-induced liver injury.Methods: HPLC fingerprinting was used to characterize the chemical characteristics of Triphala extracts obtained by (a) 0.5 h ultrasonication, (b) 2 h reflux, and (c) 4 h reflux. Hepatoprotective efficacy was evaluated in a mouse model of CCl4-induced liver damage. Serum levels of alanine transaminase (ALT) and aspartate aminotransferase (AST) were measured, as well as the liver antioxidant and inflammatory markers malondialdehyde superoxide dismutase glutathione peroxidase (GSH-Px), TNF-α, and IL-6. Gene and protein expression of Nrf-2 signaling components Nrf-2, heme oxygenase (HO-1), and NADPH Quinone oxidoreductase (NQO-1) in liver tissue were evaluated by real-time PCR and western blotting.Results: Chemical analysis showed a clear difference in content between extracts produced by ultrasonic and reflux methods. The pharmacological analysis showed that all three Triphala extracts reduced ALT, AST, MDA, TNF-α, and IL-6 levels and increased SOD and GSH-Px. Triphala extracts also induced transcript and protein expression of Nrf-2, HO-1, and NQO-1.Conclusion: Triphala extract prevents CCl4-induced acute liver injury. The ultrasonic extract of Triphala was most effective, suggesting that hepatoprotection may be related to the larger tannins via activation of Nrf-2 signaling.

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Endothelial lipase-modified high-density lipoprotein exhibits diminished ability to mediate SR-BI (scavenger receptor B type I)-dependent free-cholesterol efflux

Biochemical Journal ◽

10.1042/bj20031882 ◽

2004 ◽

Vol 382 (1) ◽

pp. 75-82 ◽

Cited By ~ 20

Author(s):

Martin GAUSTER ◽

Olga V. OSKOLKOVA ◽

Josef INNERLOHINGER ◽

Otto GLATTER ◽

Gabriele KNIPPING ◽

...

Keyword(s):

High Density Lipoprotein ◽

Conditioned Medium ◽

Cholesterol Efflux ◽

Free Cholesterol ◽

Binding Capacity ◽

Scavenger Receptor ◽

Density Lipoprotein ◽

High Density ◽

Endothelial Lipase ◽

Type I

Endothelial lipase (EL) is a phospholipase with little triacylglycerol lipase activity. To assess structural and functional properties of EL-HDL (EL-modified high-density lipoprotein), HDL was incubated with conditioned medium from Cos-7 cells infected with adenovirus encoding human EL. After re-isolation of HDL by ultracentrifugation, TLC and HPLC analyses revealed that EL-HDL was markedly depleted in phosphatidylcholine and enriched in lyso-phosphatidylcholine compared with LacZ-HDL (control HDL) incubated with conditioned medium from Cos-7 cells infected with adenovirus encoding β-galactosidase. The EL-HDL was enriched in non-esterified fatty acids and, as revealed by lipid electrophoresis, was more negatively charged than control HDL. The HDL particle size as well as the total cholesterol, free cholesterol and triacylglycerol content of HDL were not significantly altered after EL modification. The ability of EL-HDL to mediate 3H-cholesterol efflux from SR-BI (scavenger receptor B type I) overexpressing Chinese-hamster ovary cells was impaired and markedly lower compared with LacZ-HDL at HDL concentrations of 100 μg/ml and above. Studies with 125I-labelled HDL showed almost unaltered binding affinity (Km values) and a slightly but significantly decreased binding capacity (Bmax values) of EL-HDL to SR-BI, compared with LacZ-HDL. The ATP-binding-cassette transporter A1-dependent cholesterol and phospholipid effluxes were not affected by EL modification. From these results, we concluded that EL modification alters chemical composition and physical properties of HDL, resulting in its decreased binding capacity to SR-BI and a diminished ability to mediate SR-BI-dependent cholesterol efflux.

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Differential apoptosis-related protein expression, mitochondrial properties, proteolytic enzyme activity, and DNA fragmentation between skeletal muscles

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00488.2010 ◽

2011 ◽

Vol 300 (3) ◽

pp. R531-R543 ◽

Cited By ~ 31

Author(s):

Elliott M. McMillan ◽

Joe Quadrilatero

Keyword(s):

Skeletal Muscle ◽

Cytochrome C ◽

Protein Expression ◽

Dna Fragmentation ◽

Fiber Type ◽

Reactive Oxygen Species Generation ◽

Permeability Transition ◽

Type I ◽

Fiber Types ◽

Oxidative Potential

Increased skeletal muscle apoptosis has been associated with a number of conditions including aging, disuse, and cardiovascular disease. Skeletal muscle is a complex tissue comprised of several fiber types with unique properties. To date, no report has specifically examined apoptotic differences across muscles or fiber types. Therefore, we measured several apoptotic indices in healthy rat red (RG) and white gastrocnemius (WG) muscle, as well as examined the expression of several key proteins across fiber types in a mixed muscle (mixed gastrocnemius). The protein content of apoptosis-inducing factor (AIF), apoptosis repressor with caspase recruitment domain (ARC), Bax, Bcl-2, cytochrome c, heat shock protein 70 (Hsp70), and second mitochondria-derived activator of caspases (Smac) were significantly ( P < 0.05) higher in RG vs. WG muscle. Cytosolic AIF, cytochrome c, and Smac as well as nuclear AIF were also significantly ( P < 0.05) higher in RG compared with WG muscle. In addition, ARC protein expression was related to muscle fiber type and found to be highest ( P < 0.001) in type I fibers. Similarly, AIF protein expression was differentially expressed across fibers; however, AIF was correlated to oxidative potential ( P < 0.001). Caspase-3, -8, and -9 activity, calpain activity, and DNA fragmentation (a hallmark of apoptosis) were also significantly higher ( P < 0.05) in RG compared with WG muscle. Furthermore, total muscle reactive oxygen species generation, as well as Ca2+-induced permeability transition pore opening and loss of membrane potential in isolated mitochondria were greater in RG muscle. Collectively, these data suggest that a number of apoptosis-related indices differ between muscles and fiber types. Given these findings, muscle and fiber-type differences in apoptotic protein expression, signaling, and susceptibility should be considered when studying cell death processes in skeletal muscle.

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