Quercetin attenuates avermectin-induced cardiac injury in carp through inflammation, oxidative stress, apoptosis and autophagy.

As an important antibiotic, avermectin (AVM) has been widely used in China, but its unreasonable application has caused serious harm to the water environment. In view of the various pharmacological effects of quercetin (QUE), such as anti-inflammatory and antioxidant, the scientific hypothesis that "QUE may cause carp poisoning by inhibiting AVM" was proposed in this study. However, its protective effect in AVM -induced heart damage has not been reported. QUE reduced the symptoms of AVM toxicity and decreased the levels of creatine kinase, lactate dehydrogenase, and creatine kinase in the serum of carp. By histological observation, QUE was found to significantly reduce cardiac fiber swelling in carp. A DHE fluorescence probe study showed that QUE was able to inhibit AVM -induced accumulation of reactive oxygen species (ROS) in carp myocardium. We found that QUE significantly increased the intracellular antioxidant enzymes CAT, T-AOC and GSH enzyme activity and reduced intracellular MDA content. In addition, QUE significantly increased il-10 and tgf-ß1 expression, and significantly down-regulated tnf-α, il-6, il-1ß and inos expression. Tunel assay showed that QUE attenuated AVM -induced apoptosis, significantly decreased the transcript levels of pro-apoptosis-related genes, and increased the expression of anti-apoptosis-related genes. We also detected the protein expression of LC3 in the AVM group and QUE + AVM group, and found that the expression of LC3 was significantly increased in both groups compared with the Control group, but after adding QUE, the expression of LC3 was significantly decreased compared with the AVM group. In addition, the transcript levels of p62 and atg5 were also detected by qPCR. QUE significantly increased the expression of p62 and decreased the expression of atg5, suggesting that QUE could attenuate AVM -induced cardiac autophagy in carp. This study will provide preliminary evidence of the principle of QUE attenuating AVM -induced myocardial injury in carp from four aspects, including oxidative stress, inflammatory response, apoptosis and autophagy, and provide a theoretical basis for its prevention and treatment.

Effects of the combination of vitamins C and E supplementation on oxidative stress, inflammation, muscle soreness, and muscle strength following acute physical exercise: meta-analyses of randomized controlled trials.

Background:The combined supplementation of vitamins C and E potentially can mitigate oxidative stress (OS) and accelerate recovery following exercise. However, there is little evidence and a lack of consensus on the effects of these vitamins for this purpose. The objective of this systematic review was to summarize the evidence on the effects of the combined supplementation of vitamins C and E in OS, inflammatory markers, muscle damage, muscle soreness, and musculoskeletal functionality following acute exercise. Methods: The search was carried out from inception until March 2021, on MEDLINE, EMBASE, Cochrane CENTRAL, Web of Science, and SPORT Discus. We included placebo-controlled randomized clinical trials (RCTs) that evaluated the effects of combined supplementation of vitamins C and E in OS, inflammatory markers, muscle damage, muscle soreness, and muscle strength following a single bout of exercise. Random-effect meta-analyses were used to compare pre to post-exercise mean changes in subjects who received supplementation with vitamins C and E or placebo versus controls. Data are presented as standard mean difference (SMD) and 95% confidence interval (95% CI). Results: Eighteen RCTs, accounting for data from 322 individuals, were included. The use of vitamins attenuated lipid peroxidation (SMD= -0.703; 95% CI= -1.035 to -0.372; p < 0.001), IL-6 (SMD= -0.576; 95%CI= -1.036 to -0.117; p = 0.014), and cortisol levels (SMD= -0.918; 95%CI= -1.475 to -0.361; p = 0.001) immediately, and creatine kinase levels 48 h following exercise (SMD= -0.991; 95%CI= -1.611 to -0.372; p = 0.002). Supplementing the combination of vitamins had no effects on protein carbonyls, reduced/oxidized glutathione ratio, catalase, interleukin-1Ra, C-reactive protein, lactate dehydrogenase, muscle soreness, and muscle strength. Conclusion: Prior supplementation of the combination of vitamins C and E attenuates OS (lipid peroxidation), the inflammatory response (interleukin-6), cortisol levels, and muscle damage (creatine kinase) following a session of exercise.

Abamectin causes cardiac dysfunction in carp via inhibiting redox equilibrium and resulting in immune inflammatory response and programmed cell death.

This study aims to investigate the effects of environmentally relevant concentrations of abamectin on the cardiac function of carp and the potential mechanisms. Here, male carp were exposed to abamectin, and cardiac function-related enzymatic markers were examined. Cardiac histopathology, redox equilibrium, inflammation, and cell death were evaluated. Abamectin exposure caused cardiac dysfunction by upregulating lactate dehydrogenase (LDH), aspartate aminotransferase (AST), creatine kinase (CK), creatine Kinase MB isoenzyme (CK-MB) and white blood cells (WBCs), and decreasing red blood cells (RBCs) and hemoglobin (Hb). DHE staining and biochemical assays revealed that abamectin caused ROS release and oxidative stress by inhibiting Nrf2-ARE pathway. Histopathological and real-time fluorescence quantitative PCR (RT-qPCR) assays revealed that abamectin caused myocardial fiber swelling and inflammatory cell infiltration, enhanced pro-inflammatory cytokines tumor necrosis factor-α (Tnf-α), interleukin-1 beta (Il-1ß), and Il-6 levels and attenuated anti-inflammatory cytokines Il-10 and transforming growth factor beta 1 (Tgf-ß1) through activating NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome and nuclear factor kappa-B (NF-κB) pathway. Tunel staining showed that abamectin triggered cardiac apoptosis via activating p53-mediated mitochondrial apoptosis with elevated bcl2-associated X (Bax), reduced B-cell lymphoma-2 (Bcl-2), and activated Caspase-9 and Caspase-3. Immunoblot analysis revealed that abamectin activated autophagic flow by inhibiting mammalian target of rapamycin (mTOR), resulting in the conversion of LC3B from LC3-I to LC3-II, elevation of autophagy protein 5 (Atg5), and reduction of p62. Overall, abamectin caused cardiac dysfunction in carp via inhibiting redox equilibrium and resulting in immune inflammatory response and programmed cell death.

Shenlian extract attenuates myocardial ischaemia-reperfusion injury via inhibiting M1 macrophage polarization by silencing miR-155.

CONTEXT: Shenlian extract (SL) is a combination of Salvia miltiorrhiza Bge. (Labiatae) and Andrographis paniculata (Burm. F.) Wall. Ex Nees (Acanthaceae) extracts, which promote blood circulation and clear endogenous heat toxins. Myocardial ischaemia-reperfusion injury (MI/RI) is aggravated myocardial tissue damage induced by reperfusion therapy after myocardial infarction. OBJECTIVES: This study explores the effect of SL on MI/RI and the underlying mechanism. MATERIALS AND METHODS: Primary peritoneal macrophages (pMACs) were treated with LPS and SL (5, 10 or 20 µg/mL) for 24 h. The myocardial ischaemia-reperfusion (MI/R) model was established after administration of different doses of SL (90, 180 or 360 mg/kg). Myocardial tissue injury was assessed by methylthiazolyl tetrazolium (TTC) staining and levels of creatine kinase (CK), lactate dehydrogenase (LDH) and superoxide dismutase (SOD) in mice. The double immunofluorescence staining of iNOS/F4/80 and CD86/F4/80 was used to detect macrophage M1 polarization. The levels of miR-155, inflammatory factors and chemokines were detected by qRT-PCR or ELISA. CD86, iNOS, SOCS3, JAK2, p-JAK2, STAT3 and p-STAT3 proteins expressions in macrophages were analyzed by western blotting. Conditioned medium transfer systems were designed to unite M1 macrophages with H/R cardiomyocytes, and cell apoptosis was detected by TUNEL staining, western blotting or immunohistochemistry. RESULTS: SL reduced apoptosis, diminished CK and LDH levels, raised SOD concentration and decreased infarct size in the MI/R model. Meanwhile, SL decreased miR-155 level, inhibited M1 macrophage polarization and inflammation. Furthermore, SL promoted SOCS3 expression and blocked JAK2/STAT3 pathway in vitro. CONCLUSIONS: SL may be a promising TCM candidate for MI/RI. The underlying mechanisms could be associated with inhibition of M1 macrophage polarization via down-regulating miR-155.

The Effect of Years-Long Exposure to Low-Dose Colchicine on Renal and Liver Function and Blood Creatine Kinase Levels: Safety Insights from the Low-Dose Colchicine 2 (LoDoCo2) Trial.

BACKGROUND AND OBJECTIVE: The Low-Dose Colchicine-2 (LoDoCo2) trial showed that 2-4 years exposure to colchicine 0.5 mg once daily reduced the risk of cardiovascular events in patients with chronic coronary artery disease. The potential effect of years-long exposure to colchicine on renal or liver function and creatine kinase (CK) has not been systematically evaluated and was investigated in this LoDoCo2 substudy. METHODS: Blood samples drawn from 1776 participants at the close-out visit of the LoDoCo2 trial were used to measure markers of renal function (creatinine, blood urea nitrogen [BUN]), liver function (alanine aminotransferase [ALT], γ-glutamyl transferase [GGT], bilirubin and albumin), and CK. Renal and liver function as well as hyperCKemia (elevated CK) were categorized to the degree of elevation biomarkers as mild, mild/moderate, moderate/severe, and marked elevations. RESULTS: In total, 1776 participants (mean age 66.5 years, 72% male) contributed to this analysis, with a median exposure to trial medication of 32.7 months. Compared with placebo, colchicine was not associated with changes in creatinine and BUN but was associated with elevations in ALT (30 U/L vs. 26 U/L; p < 0.01) and CK (123 U/L vs. 110 U/L; p < 0.01). Most elevations in ALT and CK were mild in both treatment groups. There were no moderate to marked ALT elevations (> 5-10 × upper limit of normal [ULN]) in both treatment groups, and 6 (0.7%) colchicine-treated vs. 2 (0.2%) placebo-treated participants had moderate to marked CK elevations (> 5-10 × ULN). CONCLUSION: In chronic coronary artery disease, 2-4 years of exposure to colchicine 0.5 mg once daily was associated with small elevations in ALT and CK, but was not associated with changes in renal function. TRIAL REGISTRATION: https://www.anzctr.org.au ; ACTRN12614000093684, 24 January 2014.

Ameliorative effect of flavocoxid on cyclophosphamide-induced cardio and neurotoxicity via targeting the GM-CSF/NF-κB signaling pathway.

Cyclophosphamide (Cyclo) is a chemotherapeutic agent used as an immunosuppressant and as a treatment for many cancerous diseases. Many previous pieces of literature proved the marked cardio and neurotoxicity of the drug. Thus, this research provides evidence on the alleviative effect of flavocoxid on the cardiac and brain toxicity of cyclophosphamide in mice and determines its underlying mechanisms. Flavocoxid (Flavo) is a potent antioxidant and anti-inflammatory agent that inhibits the peroxidase activity of cyclooxygenase (COX-1 and COX-2) enzymes and 5-lipooxygenase (5-LOX). Flavo was administered orally (20 mg/kg) for 2 weeks, followed by Cyclo (100 mg/kg, i.p.) on day 14. Higher heart and brain weight indices, serum lactate dehydrogenase (LDH), creatine kinase (CK-MB), and nitric oxide (NO) were mitigated following Flavo administration. Flavo modulated oxidative stress biomarkers (malonaldehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD)), tumor necrosis factor-α (TNF-α), and interleukin (IL)-1ß. Additionally, cardiac troponin I (cTn-I), nuclear factor kappa B (NF-κB), brain amyloid precursor protein (APP), and granulocyte macrophage colony-stimulating factor (GM-CSF) were decreased by Flavo administration. Moreover, Flavo ameliorated heart and brain histopathological changes and caspase-3 levels. Collectively, Flavo (20 mg/kg) for 14 days showed significant cardio and neuroprotective effects due to its antioxidant, anti-inflammatory, and antiapoptotic activities via modulation of oxidative stress, inflammation, and the GM-CSF/NF-κB signaling pathway.

Ginkgolide B targets and inhibits creatine kinase B to regulate the CCT/TRiC-SK1 axis and exerts pro-angiogenic activity in middle cerebral artery occlusion mice.

Promoting angiogenesis in the ischemic penumbra is a well-established method of ischemic stroke treatment. Ginkgolide B (GB) has long been recognized for its neuroprotective properties following stroke. As previously reported, it appears that stroke-induced neurogenesis and angiogenesis interact or are dependent on one another. Although the pharmacodynamic effect of GB on cerebral blood flow (CBF) following ischemic stroke has been reported, the molecular mechanism underlying this effect remains unknown. As such, this study sought to elucidate the pharmacodynamic effects and underlying mechanisms of GB on post-stroke angiogenesis. To begin, GB significantly increased the proliferation, migration, and tube formation capacity of mouse cerebral hemangioendothelioma cells (b.End3) and human umbilical vein endothelial cells (HUVEC). Additionally, GB significantly improved angiogenesis after oxygen-glucose deprivation/reperfusion (OGD/R) in endothelial cells. The dynamics of CBF, brain microvascular neovascularization and reconstruction, and brain endothelial tissue integrity were examined in middle cerebral artery occlusion (MCAO) mice following GB administration. Through label-free target detection techniques, we discovered for the first time that GB can specifically target Creatine Kinase B (CKB) and inhibit its enzymatic activity. Additionally, we demonstrated through network pharmacology and a series of molecular biology experiments that GB inhibited CKB and then promoted angiogenesis via the CCT/TRiC-SK1 axis. These findings shed new light on novel therapeutic strategies for neurological recovery and endothelial repair following ischemic stroke using GB therapy.

D-limonene (5 (one-methyl-four-[1-methylethenyl]) cyclohexane) diminishes CCl4-induced cardiac toxicity by alleviating oxidative stress, inflammatory and cardiac markers.

Background: The cardiovascular crisis is advancing rapidly throughout the world. A large number of studies have shown that plant polyphenols affect major mechanisms involved in cardiovascular events through their action on the antioxidant system, signaling, and transcription pathways. D-limonene, a monocyclic monoterpene obtained from citrus fruits, is reported to possess many pharmacological activities.Methods: The experiment was designed to determine the protective effect of D-limonene against cardiac injury induced by CCl4 in Wistar rats. Rats were treated with two doses of D-limonene against cardiac injury induced by CCl4. Serum toxicity markers, cardiac toxicity biomarker enzymes, inflammatory mediators, anti-oxidant armory, lipid peroxidation, lipid profile, and histology were done.Results: CCl4 intoxication resulted in a substantial rise in FFA, TC, TG, PL, LDL, VLDL, and a reduction in HDL, restoring these changes with the administration of D-limonene at a dosage of 200 mg/kg. CCl4 administration also resulted in lipid oxidation and decreased antioxidant activity. At the same time, D-limonene at a dosage of 200 mg/kg body weight inhibited LPO and restored in vivo antioxidant components to normal. CCl4 intoxication also resulted in a significant increase in inflammatory markers like IL-6, TNF-α, high sensitivity Corticotropin Releasing Factor (Hs-CRF), and biomarkers of cardiac toxicity like alanine aminotransferase (ALT), lactate dehydrogenase (LDH), creatine kinase (CK), creatine kinase MB (CKMB), and Troponin I & troponin-t activities. D-limonene reversed all these changes to normal. Histology further confirmed our obtained results.Conclusion: These findings indicate that D-limonene can ameliorate cardiac injury at a 200 mg/kg body weight dosage. Henceforth, D-Limonene intervenes in mediating CCl4 induced toxicity by various signaling pathways.

Protective Effect of Amino Acids on the Muscle Injury of Aerobics Athletes after Endurance Exercise Based on CT Images.

During the training process, the aerobics athletes gradually increase their technical movements, the appreciation of the movements has been gradually improved, and the injuries of the athletes themselves have also gradually become serious. Based on CT image analysis, we study the protective effect of amino acids on aerobics athletes' muscle injury after endurance exercise. There are three major substance metabolism disorders in patients with muscle sclerosis, which are mainly manifested as decreased glucose tolerance and insulin resistance. Some patients develop muscle-derived diabetes. At the same time, the synthesis of lipids such as cholesterol and apolipoproteins decreases, the production of ketone bodies increases and the body uses more ketones for energy. The BCAA/AAA factor refers to the branched-chain amino acid/aromatic amino acid (BCAA/AAA) value. In amino acid metabolism, plasma albumin decreased significantly, the ratio of amino acids was unbalanced, and BCAA/AAA decreased, which was more likely to induce muscular encephalopathy. Using computer tomography (CT) to study the protective effect of amino acids on muscle injury, 32 aerobics athletes were randomly divided into an intervention group (Ig) and a control group (CG), each with 16 people. After 64-slice spiral CT scanning of muscles and three-dimensional reconstruction, the intervention group and the control group participated in aerobic endurance training 3 weeks in advance to establish a muscle microinjury model. The intervention group took the preprepared BCAA, while the control group did not take it. After three weeks of training, there will be one hour and three hours of aerobics competition. We need to detect changes in blood glucose (BS), creatine kinase (SCK), lactate dehydrogenase (LD), alanine (ALA), and alanine aminotransferase (AA) before and after exercise and 1 hour after exercise and record AVS athletes' pain analysis table. We successfully established the muscle injury model, letting all athletes' VAS score in 6-8 points; after 1 hour of exercise, the measurement results were the same as those of 2 hours. Therefore, after endurance training, the blood glucose content of the intervention group gradually decreased and returned to the original level after 2 hours of exercise, while the control group was lower than the level of exercise after 2 hours of exercise; the content of alanine in the two groups decreased more after 2 hours of exercise; the results of serum creatine kinase in the intervention group were higher than those in the control group after exercise. In the intervention group, lactate dehydrogenase increased rapidly at 2 hours after exercise; the alanine aminotransferase in the intervention group increased after exercise, but there was no significant change in the control group. It is also concluded that the longer the exercise time and the more energy consumption, the more effective the branched-chain amino acids supplement will be. The obtained imaging data can provide a more intuitive and accurate basis for the scientific selection of athletes, and amino acids can promote the synthesis of hormones, accelerate the synthesis of proteins and other products, reduce the content of creatine kinase in the blood, and protect the rapid recovery of muscle damage.

Selected root plant supplementation reduces indices of exercise-induced muscle damage: A systematic review and meta-analysis.

This systematic review and meta-analysis examined the effects of selected root plants (curcumin, ginseng, ginger and garlic) on markers of muscle damage and muscular performance measures following muscle-damaging protocols. We included 25 studies (parallel and crossover design) with 353 participants and used the PEDro scale to appraise each study. Forest plots were generated to report on standardised mean differences (SMD) and p-values at 24 and 48 hours following the muscle-damaging protocols. The meta-analysis showed that the supplemental (SUPP) condition showed significantly lower levels of indirect muscle damage markers (creatine kinase, lactate dehydrogenase and myoglobin) and muscle soreness at 24 hours and 48 hours (p < 0.01) than the placebo (PLA) condition. The inflammatory markers were significantly lower for the SUPP condition than the PLA condition at 24 hours (p = 0.02), although no differences were identified at 48 hours (p = 0.40). There were no significant differences in muscular performance measures between the SUPP and PLA conditions at 24 hours and 48 hours (p > 0.05) post-exercise. According to our qualitative data, a number of studies reported a reduction in oxidative stress (e.g., malondialdehyde, superoxide dismutase) with a concomitant upregulation of anti-oxidant status, although other studies showed no effects. Accordingly, selected root plants minimised the level of several biomarkers of muscle damage, inflammation and muscle soreness during periods of exercise-induced muscle damage. However, the benefits of these supplements in ameliorating oxidative stress, increasing anti-oxidant status and accelerating recovery of muscular performance appears equivocal, warranting further research in these outcome measures.

The protective effect of chrysin against oxidative stress and organ toxicity in rats exposed to propetamphos.

The aim of this study was to investigate the protective efficacy of chrysin against propetamphos exposure. For this purpose, 2 to 3-month-old 40 male Wistar Albino rats were used. These animals were randomly assigned to four groups. The animals in the control group received the vehicle substance (corn oil) alone. Groups 2, 3 and 4 were administered with 50 mg/kg.bw/day of chrysin (in corn oil), 10 mg/kg.bw/day of propetamphos (in corn oil), and 10 mg/kg.bw/day of propetamphos plus 50 mg/kg.bw/day of chrysin, respectively, for 28 days. Some oxidative stress/lipid peroxidation parameters (MDA, SOD, CAT, GSH-Px, NO, glutathione) and serum biochemical parameters (triglyceride, cholesterol, creatinine, BUN, creatine phosphokinase, ALT, ALP and pseudocholinesterase) were analyzed in tissue/blood samples. Also, histopathological findings were observed. According to the data obtained, no significant alteration had occurred in these parameters and the histological findings in the group given chrysin alone, when compared to the control group. Significant unfavorable alterations were detected in the oxidative stress/lipid peroxidation/antioxidant status parameters, all biochemical parameters and histopathological findings of the group that received propetamphos alone. In the group that was given both chrysin and propetamphos, remedial/recovery alterations were observed in the oxidative stress/lipid peroxidation/antioxidant status values, serum biochemical parameters and histopathological findings, such that the values and histopathological findings showed partly similarity to those of the control group. In result, it is suggested that chrysin may provide protection against propetamphos exposure and propetamphos-induced organ damage in rats at a certain level.

Changes in serum creatine kinase, leg muscle tightness, and delayed onset muscle soreness after a full marathon race.

BACKGROUND: Muscle tightness (MT) is believed to be an important cause of injury for runners. This study evaluated the change of serum creatine kinase (CK), MT in the leg muscles, and delayed onset muscle soreness after running. METHODS: We evaluated 11 college students who completed a full marathon race. Participants completed a questionnaire on the right quadriceps muscle soreness. The CK activity and MT (iliopsoas, rectus femoris, hamstrings, gastrocnemius, and soleus muscles) were measured. The time points for CK measurements were before; immediately after; and at 1, 2, and 5 days after the race. The time points for MT measurements were the same as for CK except MT was not measured one day after the race. The time points for muscle soreness analysis were before the race and then every morning and night for 5 days after the race. RESULTS: Long-distance running led to significant increases in CK, MT, and muscle soreness. The CK levels peaked day 1 after the race. MT of iliopsoas peaked on day 5; of rectus femoris immediately after the race; and of hamstrings, gastrocnemius, and soleus on day 2. muscle soreness peaked at night on day 1. MT did not decrease to the pre-race levels on day 5. There were no significant changes but CK tended to correlate with the peak of MT of the rectus femoris (r=0.55, P=0.082) and hamstrings (r=0.57, P=0.065). CONCLUSIONS: Long-distance running may cause muscle fiber microdamage that may consequently increase CK, MT, and muscle soreness.

Ex vivo evaluation of erythrocytosis-enhanced platelet thrombus formation using the cone and plate(let) analyzer: effect of platelet antagonists.

Red blood cells (RBC) contribute significantly to haemostasis and thrombosis under oscillatory flow conditions, and erythrocytosis has been associated with increased thrombotic risk. To measure the dynamic influences of RBC on platelets, we used a recently described cone and plate(let) analyzer (CPA), evaluating the effect of haematocrit (Hct) on platelet function in whole blood under arterial flow conditions (1800/s, 2 min, 25 degrees C). Anticoagulated blood, reconstituted to varying haematocrits with autologous RBC, demonstrated a significant increase in adherent platelet aggregate formation at Hct levels >45%. This increase was not affected by pretreatment of blood with 0.05 mmol/l aspirin, but was prevented by antagonists of P2Y1, P2Y12, or P2X1, ADP and ATP receptors, and by converting exogenous ADP to ATP with creatine phosphate/creatine phosphokinase. As negligible platelet granule secretion was measured during CPA analysis, but metabolic inhibition of RBC with sodium azide or glutaraldehyde fixation inhibited erythrocytosis-enhanced increases in platelet aggregate size, adenine nucleotides contributing to shear-induced platelet aggregate formation appear to be derived from erythrocytes. These findings support the use of CPA for ex vivo evaluation of the contribution of RBC to platelet function and its inhibition under physiological shear conditions.

Downhill running in rats: influence on neutrophils, macrophages, and MyoD+ cells in skeletal muscle.

The accumulation of neutrophils and macrophages, as well as the activation of satellite cells, are early events following skeletal muscle injury. We examined the temporal relationship between changes in neutrophils, macrophages, and MyoD protein, a marker of satellite cell activation, after injurious exercise. Male rats ( n=47) performed an intermittent downhill (-16% grade) running (17 m/min) protocol and the solei were obtained at 0, 2, 6, 24, 48, or 72 h post-exercise. Neutrophils, macrophages (ED1 and ED2), and MyoD+ cells were determined in muscle cross sections using immunohistochemistry. Downhill running increased ( P

Plasma creatine kinase activity and glutathione after eccentric exercise.

PURPOSE: This study examined whether plasma total glutathione levels could explain the intersubject variability in the creatine kinase (CK) response to eccentric exercise. We hypothesized that the increase in plasma CK activity after eccentric exercise would be lower for individuals with low plasma total glutathione (<2.5 micromol x L-1) compared with individuals with high total glutathione (>3.8 micromol x L-1), but other indicators of muscle damage would be the same between groups. METHODS: Resting blood samples were obtained over 2 d from 60 subjects and analyzed for plasma total glutathione. Eight subjects who had total glutathione values below 2.5 micromol x L-1 (LG), and nine who had values above 3.8 micromol x L-1 (HG) performed 50 maximal eccentric actions of the elbow flexors. Maximal voluntary isometric contraction (MVC), relaxed arm angle (RANG), and blood samples for CK, myoglobin (Mb), and total glutathione were obtained pre, post (except blood samples), 24, 48, 72, 96, and 120 h after exercise. RESULTS: There was a significant group-by-time interaction in analysis of MVC, RANG, total glutathione, CK, and Mb response to exercise. Although LG showed a smaller CK response to eccentric exercise compared with HG, LG also showed a smaller increase in plasma Mb, a faster recovery of MVC and RANG, and an increase in plasma total glutathione. CONCLUSION: Subjects with low plasma total glutathione levels had a smaller plasma CK and Mb response and a faster recovery from eccentric exercise compared with subjects having high plasma total glutathione levels. We suggest that a blunted inflammatory response in subjects with low plasma glutathione may be one explanation for these findings.

Necrotising myopathy, an unusual presentation of a steroid-responsive myopathy.

OBJECTIVE: To evaluate the clinical features, muscle pathology and response to treatment in patients with a necrotising myopathy, without mononuclear cell infiltrates. BACKGROUND: Mononuclear cell infiltrates in the muscle biopsy specimen are the diagnostic hallmark of the immune-mediated idiopathic inflammatory myopathies (IIM). In patients with the typical clinical features of IIM, absence of these infiltrates in the muscle biopsy specimen casts doubt on the diagnosis and leads to uncertainty about therapeutical strategies. METHODS: A detailed description is given of the clinical, laboratory, and histopathological features of eight patients suspected of having an idiopathic inflammatory myopathy, in whom mononuclear cell infiltrates in their muscle biopsy specimens were lacking. RESULTS: Eight patients (five men, three women, age range 40-69 years) had severe, symmetrical proximal weakness with a subacute onset. There were no skin abnormalities suggesting dermatomyositis. Serum creatine kinase activity was more than 10 times elevated. Repeated muscle biopsy specimens, taken from a symptomatic muscle prior to immunosuppressive treatment showed widespread necrosis, regeneration, and atrophy of muscle fibres, but no mononuclear cell infiltrates. Known causes of necrotising myopathy were excluded. Three patients had a malignancy. Adequately dosed and sustained immunosuppressive treatment eventually resulted in normal or near normal muscle strength in seven patients. One patient showed marked improvement. CONCLUSION: Occasionally, patients who clinically present as an idiopathic inflammatory myopathy may lack mononuclear cell infiltrates in their muscle biopsy specimens. This subacute-onset progressive necrotising myopathy should not deter the clinician from timely and appropriate treatment as we consider this myopathy to be steroid-responsive with a possible immune-mediated pathogenesis.

Inhibition of the yeast V-type ATPase by cytosolic ADP.

The activity of the vacuolar H(+)-ATPase has been characterized in isolated vacuoles of the yeast Saccharomyces cerevisiae by means of the patch-clamp technique. With cytosolic calcium at virtually zero (<10(-9) M), Mg-ATP induced a transient, bafilomycin A(1)-sensitive current corresponding to the flow of positive charges from the cytoplasmic surface to the vacuolar lumen. The Mg-ATP-dependent current reached its maximum amplitude (30+/-8 mA m(-2) with 5 mM Mg-ATP, n=34) within 15-20 s and declined slowly over a period of about 15-20 min even in the continuous presence of Mg-ATP. This decline of pumping activity was independent of the cytosolic KCl concentration, suggesting an inhibitory mechanism different from the high salt-induced dissociation of V(0) and V(1) reported for the V-ATPase of plants and fungi. Cytosolic ADP was found to modulate the pump activity since Mg-ATP-induced pump current was smaller if monitored in the presence of 5 mM ADP and addition of 5 mM ADP in the presence of 5 mM Mg-ATP reduced the pump current by more than 50%. Furthermore, reduction of the cytosolic ADP concentration by the ATP-regenerating system creatine phosphate/creatine kinase partially relieved the endogenous inhibition of the V-ATPase, confirming that interaction of cytosolic ADP with the V-ATPase is the reason for the transient nature of the pump current in yeast vacuoles.

Creatine kinase injection restores contractile function in creatine-kinase-deficient mouse skeletal muscle fibres.

Viable genetically engineered animals generally exhibit adaptations to the altered genotype, which may mask the role of the protein of interest. We now describe a novel method by which the direct effects of the altered genotype can be distinguished from secondary adaptive changes in isolated adult skeletal muscle cells. We studied contractile function and intracellular Ca2+ handling in single skeletal muscle fibres that are completely deficient of creatine kinase (CK; CK-/-) before and after microinjection of purified CK (injected together with the fluorescent Ca2+ indicator indo-1). The mean total CK activity after CK injection was estimated to be approximately 4 mM s-1, which is approximately 5 % of the activity in wild-type muscle fibres. After CK injection, CK-/- fibres approached the wild-type phenotype in several aspects: (a) the free myoplasmic [Ca2+] ([Ca2+]i) increased and force showed little change during a period of high-intensity stimulation (duty cycle, i.e. tetanic duration divided by tetanic interval = 0.67); (b) [Ca2+]i did not decline during a brief (350 ms) tetanus; (c) during low-intensity fatiguing stimulation (duty cycle = 0.14), tetanic [Ca2+]i increased over the first 10 tetani, and thereafter it decreased; (d) tetanic [Ca2+]i and force did not display a transient reduction in the second tetanus of low-intensity fatiguing stimulation. Conversely, tetanic force in the unfatigued state was lower in CK-/- than in wild-type fibres, and this difference persisted after CK injection. Injection of inactivated CK had no obvious effect on any of the measured parameters. In conclusion, microinjection of CK into CK-/- fibres markedly restores many, but not all, aspects of the wild-type phenotype.

The central role of thromboxane and platelet activating factor receptors in ex vivo regulation of endotoxin-induced monocyte tissue factor activity in human whole blood.

Expression of tissue factor (TF) by activated monocytes may initiate thrombotic episodes associated with diseases, such as thrombosis and atherosclerosis. In this study, steps in the regulatory pathways of lipopolysaccharide (LPS)-induced monocyte TF activity and released TNF-alpha in human whole blood were probed for using an array of inhibitors, comprising specific inhibitors of cytosolic phospholipase A(2) (PLA(2)) (AACOCF(3)), secretory PLA(2) (SB-203347), protein kinase (PK) (staurosporine), PKC (GF-109203; BIM), and serine protease (Pefabloc SC), antagonists of thromboxane prostanoid (TP) receptor (R) (SQ-29548), platelet activating factor (PAF) R (BN-52021), leukotriene B(4) R (SC-41930), serotonin R (cyproheptadine), fibronectin/fibrinogen R (RGDS), and finally, creatine phosphate/creatine phosphokinase (CP/CPK) which removes ADP. Whereas when added alone neither of these agents significantly inhibited LPS-induced TF or TNF-alpha, when presented as a reference cocktail comprising all the agents, TF activity and TNF-alpha were reduced by 77% and 49%, respectively. By subsequently testing a series of incomplete inhibitory cocktails equal to the reference except for deleted single agents or combinations of two or three active agents, the inhibitory effect of the reference cocktail could be shown to depend on the presence of the protease inhibitor and the thromboxane A(2) and PAF antagonists.

Tissue tolerance of diclofenac sodium encapsulated in liposomes after intramuscular administration.

In this work the effect of the encapsulation of diclofenac sodium within liposomes on the reduction of the myotoxicity after intramuscular administration in rats was studied. Diclofenac sodium was encapsulated in small unilamellar liposomes obtained from phosphatidylcholine, cholesterol, and alpha-tocopherol (40:10:0.04 mM), and administered by intramuscular injection in the quadriceps femoral muscle of male Wistar rats. After a single dose of 0.2 mg diclofenac formulations the local tissue damage was assessed by plasma creatine kinase (CPK) activity and histological analysis. It was demonstrated that formulations containing free diclofenac produced a higher increase in CPK activity, while those encapsulated in liposomes exhibited CPK activity similar to the control groups. Histopathological analysis of local muscle tissue performed on the third and seventh days following the injection showed intense cellular damage when free drug solution was used, while encapsulation in liposome protected the tissue against the local tissue inflammation.