Creatine Kinase Enzyme Activity Research Articles

Protective effects of exercise against sepsis-induced energy metabolism dysfunction in skeletal muscle of rats

We evaluated effects of aerobic physical preconditioning on general performance and energy metabolism in skeletal muscle of septic rats. Forty-eight 10-wk-old male Wistar rats were randomly assigned to either Untrained or Trained groups. Aerobic exercise training protocol (AETP) consisted of an 8-week treadmill program. After AETP, performance was evaluated by graded treadmill and functional ambulation testing. Afterwards animals from both groups were randomly assigned to Sham or CLP surgery (cecal ligation and perforation), resulting in the following groups: Sham untrained (ShamU), CLP untrained (CLPU), Sham trained (ShamT), and CLP trained (CLPT). Two days after surgery, animals repeated the ambulation test, and were euthanized after this. Diaphragm, soleus and plantaris muscles were harvested. Mitochondrial electron transport chain enzyme (METC) and creatine kinase (CK) activity were measured. AETP led to significant improvement in performance of distance run and in skeletal muscle function of the Trained group. Forty-eight hours after surgery the CLPT group was able to maintain similar muscle performance as Sham groups. Dysfunction was shown in the diaphragm in METC complexes I and II-III and in locomotive soleus muscles in complex I; CK enzyme activity was significantly increased in sedentary CLPU group in soleus and plantaris muscle, but in the diaphragm there was only a tendency (p=0.07). CLPT animals that were submitted to AETP avoided all these negative results. Taken together our results provide evidence of the positive effects obtained with an aerobic physical preconditioning program on METC and CK enzyme activity related to the diaphragm and locomotive muscles mitigating sepsis-induced energy metabolism dysfunction.

Biochemical characterization of preovulatory and cystic ovarian follicular fluid of sow

Aim: The present study was designed to biochemically characterize ovarian cyst follicular fluid of sows and to relate possible changes in relation to preovulatory ovarian follicular fluid of sows. Materials and Methods: Ovaries were collected from adult and non-pregnant slaughtered sow (Hampshire X local cross). A total of 16 pairs of ovaries were investigated, and they were classified according to their pathological and physiological status into two categories i.e. preovulatory and cystic follicle (diameter >25 mm with turbid appearance). Fluid was aspirated from the follicles and stored at −20°C prior to assaying. Follicular fluid samples were analyzed for glucose, total protein, cholesterol, triglycerides, blood urea nitrogen (BUN), urea, creatinine, uric acid. The activities of the enzymes aspartate transaminase, alanine transaminase (ALT), alkaline phosphatase (ALP), creatine kinase (CK) and gamma-glutamyl transpeptidase (GGT) were also estimated by spectrophotometric methods. The data were statistically analyzed using the SPSS software. Results: As follicles became cystic, the concentrations of glucose, cholesterol, triglyceride, total protein and albumin showed a significant (p<0.05) increased while BUN decreased significantly (p<0.05). The enzyme activities of ALT, ALP and CK increased significantly (p<0.05), but the activity of GGT significantly (p<0.05) decreased. Conclusion: Results obtained showed that there was a significant change in most of the cystic follicular fluid metabolites.

Chronic Hypoxia Enhances Expression and Activity of Mitochondrial Creatine Kinase and Hexokinase in the Rat Ventricular Myocardium

Background: Creatine kinase (CK) and hexokinase (HK) play a key role in myocardial energy homeostasis. We aimed to determine CK and HK expression and enzyme activity in the left (LV) and right (RV) ventricles of rats adapted for 3 weeks to normobaric hypoxia (10 % O₂) either continuously (CNH) or intermittently with 1-h or 16-h normoxic episode per day. Methods: The Real-Time RT-PCR, Western blot, and enzyme-coupled assays were used. In addition, the effect of CNH on the HK co-localization with mitochondria, which can inhibit apoptosis, was assessed using immunofluorescence techniques. Results: CK and HK activities increased in the LV during all hypoxic adaptations, which was consistent with elevated protein levels of mitochondrial mtCKs, cytosolic CKB, HK1, and HK2 isoforms. Enzyme activities also increased in the hypoxic RV, but only CKB protein was elevated. No effect of CNH on HK1 or HK2 co-localization with mitochondria was observed. Conclusion: Up-regulation of mtCKs and HK isoforms may stimulate the respiratory chain and help to maintain energy homeostasis of chronically hypoxic myocardium and prevent oxidative stress. In this way, CK and HK enzymes can possibly participate in the establishment of ischemia-resistant phenotype of chronically hypoxic hearts.

The α′ subunit of β-conglycinin and the A1–5 subunits of glycinin are not essential for many hypolipidemic actions of dietary soy proteins in rats

This study examined the effects of dietary soy protein (SP) lacking different storage protein subunits and isoflavones (ISF) on the abdominal fat, blood lipids, thyroid hormones, and enzymatic activities in rats. Weanling Sprague-Dawley rats (8 males and 8 females/group) were fed diets containing either 20 % casein without or with supplemental isoflavones or alcohol-washed SP isolate or SP concentrates (SPC) prepared from 6 different soy bean lines for 8 weeks. Feeding of diets containing SPC regardless of their subunit compositions significantly lowered relative liver weights, blood total, free, and LDL cholesterol in both genders (P < 0.05) and also reduced serum free fatty acids (FFA) and abdominal fat in females (P < 0.05) compared to the casein or casein + ISF diets. Dietary SPC significantly elevated the plasma free triiodothyronine (T3) in both genders and total T3 in females compared to the casein diet (P < 0.05). The SPC lacking β-conglycinin α' and either the glycinin A1-3 or A1-5 subunits increased total T3 in males and reduced plasma enzymatic activities of creatine kinase and lactate dehydrogenase compared to casein or casein + ISF diet (P < 0.05). Soy isoflavones were mainly responsible for the hypocholesterolemic effects and increased plasma free T3, whereas reduction in FFA, abdominal fat, liver weight and increased plasma total T3 were the effects of the soy proteins. Neither the α' subunit of β-conglycinin nor the A1-5 subunits of glycinin are essential for the hypolipidemic properties of soy proteins.

Expressão do Mg+2, CK, AST e LDH em equinos finalistas de provas de enduro

Nos últimos anos, o equino atleta vem sendo cada vez mais requerido. Dessa forma, as exigências por alto desempenho têm fomentado o interesse pelo estudo das afecções relacionadas com a fisiopatologia de diversas enfermidades dos equinos. A relação entre o íon magnésio e o exercício físico tem recebido atenção significativa visto que este íon está intimamente relacionado ao tecido muscular estriado esquelético. Além disso, dentre as principais estratégias para a detecção e acompanhamento clínico de lesões musculares, destacam-se a avaliação das atividades das enzimas creatino quinase (CK), lactato desidrogenase (LDH) e aspartato aminotransferase (AST). A busca pelo estabelecimento de parâmetros que se relacionam entre si é um fator determinante na compreensão de alterações fisiológicas encontradas diante do esforço em equinos atletas. Desta forma, o presente trabalho teve como objetivo determinar como as concentrações sanguíneas do íon magnésio e as atividades enzimáticas das enzimas CK, LDH e AST comportaram-se em equinos Puro Sangue Árabe finalistas de provas de enduro de 90km e relacionar as possíveis alterações com o tipo de esforço físico desempenhado pelos animais. Foram avaliadas a atividade enzimática das enzimas CK, LDH, AST e a concentração do íon magnésio no exercício em relação ao repouso de 14 equinos clinicamente hígidos da raça Puro Sangue Árabe, sendo 9 machos e 5 fêmeas, com idades variando entre 6 a 12 anos, submetidos a treinamento para enduro e participantes de provas de 90 km. Pode-se observar que as variáveis acima mencionadas sofreram aumento com diferença estatística em relação ao repouso. O exercício físico de enduro determinou a ocorrência de alterações nas atividades enzimáticas das enzimas CK (p≤0,001), LDH (p=0,0001), AST (p=0,0007) e na concentração do íon magnésio (p=0,0004), no exercício em relação ao repouso (p≤0,05). Fato que determinou alteração de permeabilidade das células musculares estriadas esqueléticas, sugerindo o estabelecimento de um processo inflamatório agudo. Devido à expressão da atividade enzimática da CK (p≤0,001), por sua especificidade em relação à ocorrência de danos na musculatura estriada esquelética, juntamente com o íon magnésio (p=0,0004) que participa de várias reações celulares. Houve alterações na concentração de proteína plasmática total (p=0,0009) e hematócrito (p=0,0001), entre os momentos avaliados. Portanto estes resultados servem como valores de referência de equinos finalistas de provas de enduro de 90 km, auxiliando na prevenção da ocorrência de possíveis danos musculares e processos inflamatórios severos.

English

This experiment was conducted to evaluate the effects of dietary turmeric rhizome powder (TP) on performance, blood metabolite, immune system, antioxidant status, and relative weight of organs in pre and post heat stressed broilers. Two hundred and sixty-four (264) day-old male Arian broiler chicks were randomly allotted to three dietary treatments containing 0, 0.4 or 0.8% turmeric powder. Each diet was fed to four replicates of 22 birds each. Heat stress was applied for 5 h (33&deg;C) from 28 to 42 days. Two birds from each replicate were randomly selected and blood samples were taken to collect the serum, plasma and whole blood. These were slaughtered to determine the relative weight of organs pre (28 days PHS) and after heat stress (42 days AHS). TP did not affect feed intake, body weight, feed conversion ratio (FCR), production index, and protein and energy efficiency ratio of broilers. Blood cholesterol and low density lipoprotein (LDL) decreased and blood high density lipoprotein (HDL) increased when fed TP in PHS and AHS birds. TP did not affect serum triglyceride, protein, antibody production against sheep red blood cell (SRBC), IgG, IgM and Newcastle vaccination (ND) in PHS and AHS. TP suppressed the enzyme activity of lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) of heat stressed birds. The enzyme activity of creatine kinase (CK) in AHS and superoxide dismutase (SOD) in PHS were not affected by TP diets. TP diets increased blood activity of glutathione peroxidase (GPx) and SOD and decreased blood thiobarbituric acid reactive substances (TBARS) index. The H:L ratio, relative weight of bursa of fabircus and abdominal fat decreased in birds fed TP diets. TP diets did not alter percentage of bone Ash, and P of heat stress broilers; however, it increased bone Ca of heat stressed broilers. It can be concluded that the supplementation of TP to diets, decreased the activity of dehydrogenase enzyme, stressor index, and improved the antioxidant status without affecting performance and the immune system of heat stressed broilers. Key words:&nbsp;Turmeric powder, immune and antioxidant system, blood metabolite, broiler

SERUM CHEMISTRY AND ANTIBODIES AGAINST PATHOGENS IN ANTARCTIC FUR SEALS, WEDDELL SEALS, CRABEATER SEALS, AND ROSS SEALS

Information on health parameters, such as antibody prevalences and serum chemistry that can reveal exposure to pathogens, disease, and abnormal physiologic conditions, is scarce for Antarctic seal species. Serum samples from Antarctic fur seals (Arctocephalus gazella, n=88) from Bouvetøya (2000-2001 and 2001-2002), and from Weddell seals (Leptonychotes weddellii, n=20), Ross seals (Ommatophoca rossii, n=20), and crabeater seals (Lobodon carcinophagus, n=9) from the pack-ice off Queen Maud Land, Antarctica (2001) were analyzed for enzyme activity, and concentrations of protein, metabolites, minerals, and cortisol. Adult Antarctic fur seal males had elevated levels of total protein (range 64-99 g/l) compared to adult females and pups (range 52-79 g/l). Antarctic fur seals had higher enzyme activities of creatine kinase, lactate dehydrogenase, and amylase, compared to Weddell, Ross, and crabeater seals. Antibodies against Brucella spp. were detected in Weddell seals (37%), Ross seals (5%), and crabeater seals (11%), but not in Antarctic fur seals. Antibodies against phocine herpesvirus 1 were detected in all species examined (Antarctic fur seals, 58%; Weddell seals, 100%; Ross seals, 15%; and crabeater seals, 44%). No antibodies against Trichinella spp., Toxoplasma, or phocine distemper virus (PDV) were detected (Antarctic fur seals were not tested for PDV antibodies). Antarctic seals are challenged by reduced sea ice and increasing temperatures due to climate change, and increased anthropogenic activity can introduce new pathogens to these vulnerable ecosystems and represent a threat for these animals. Our data provide a baseline for future monitoring of health parameters of these Antarctic seal species, for tracking the impact of environmental, climatic, and anthropogenic changes in Antarctica over time.

Histopathological changes induced by Hemiscorpius lepturus scorpion venom in mice

Envenomation by Hemiscorpius lepturus (H. lepturus) is associated with local necrosis, followed by systemic manifestations. In this work the LD50 of H. lepturus venom were determined by subcutaneous (SC) injection in white Balb/c mice (5 mg/kg). Histopathological alterations in organs such as kidney, heart, liver, lungs, stomach and intestine were determined in 3, 6, 12 and 24 h following experimental (SC) envenoming injection of one LD50 of the venom in Balb/c mice. Histological studies showed degenerative changes in the kidney with disorganized glomeruli and necrotic tubular in 3 h and reached to its climax in 6 h. Myocardium showed massive myocytolysis with interstitial necrosis in 3 h and reached to its peak after 6 h past envenoming. Bowels showed edema of lamina propria and slight villous necrosis. The enzymatic activities of creatine kinase (CK) and lactate dehydrogenase (LDH) were significantly increased in the serum in 9 h. No necrotic lesion observed in lungs and liver. The results indicate that the venom of H. lepturus is a highly cytotoxic, and induces massive tissue damages in specific organs, starting from the heart and kidney as the first target in 3 h and ends to the bowels in 6 h post envenomation.

Evaluation of cardiac subacute toxicity of ciprofloxacin in rats using serum biochemical parameters.

Cardiac subacute toxicity induced by ciprofloxacin (1-cyclopropyl-6-fluoro-1,4-dihydro-4-oxo-7-(1-piperazinyl)-3-quinoline carboxylic acid, CAS 86393-32-0) a relatively new quinolone carboxylic acid derivative with an extensive antibacterial spectrum was investigated in healthy rats using serum biochemical parameters. Toxicological evaluation was performed in serum samples following the administration of the therapeutic dose regimens of the compound. Cardiac subacute toxicity was evaluated by measuring serum enzyme activity of creatine kinase, creatine kinase isoenzyme-MB (muscular-brain), lactate dehydrogenase and aspartate aminotransferase. The serum biochemical parameters indicated that the cardiac subactue toxicity of ciprofloxacin was: Cipro 1.2 (250 mg) = Cipro 2.4 (500 mg) < Cipro 4.3 (750 mg).

Diferentes tempos de eletroestimulação neuromuscular (eenm) de média frequência (kotz) em cães

O objetivo desta pesquisa foi empregar a estimulação elétrica neuromuscular (EENM) de média frequência no músculo quadríceps femoral de cães com atrofia muscular induzida, avaliar o ganho de massa muscular e comparar a EENM sob diferentes tempos de tratamento. Foram utilizados oito cães, pesando entre 15 e 25kg e distribuídos aleatoriamente em dois grupos denominados de GI (30minutos) e GII (60minutos). Para a indução da atrofia muscular, a articulação do joelho direito foi imobilizada por 30 dias por transfixação percutânea tipo II. Após a retirada do aparelho de imobilização, foi realizada a EENM nos cães dos grupos GI e GII três vezes por semana, com intervalo mínimo de 48 horas entre cada sessão, pelo período de 60 dias. Foram mensuradas a perimetria da coxa, goniometria dos joelhos, atividade da enzima creatina-quinase (CK) e morfometria das fibras musculares do vasto lateral em cortes transversais colhido mediante a biópsia muscular. Não houve diferença quanto aos valores da perimetria da coxa e atividade da enzima CK. A goniometria revelou significância (P&lt;0,05) nos grupos GI e GII entre os tempos zero e 30 minutos. Os grupos GI e GII apresentaram aumento significativo (P&lt;0,05) da área de secção quando comparados com o dia zero e noventa. Pode-se concluir que a EENM de média frequência ocasiona hipertrofia do músculo vasto lateral em cães após atrofia muscular induzida. A EENM com duração de 60minutos (GII) promove um maior ganho de massa muscular em relação ao GI.

Extrapolation From Mouse to Man

Most scientists are becoming aware that KO mice often develop cellular compensatory mechanisms that can ‘muddy’ the predicted results. This problem is often compounded because many physiologists and molecular biologists do not understand how different even the mouse and rat cardiovascular systems are, let alone mouse versus man. —C. Gibbs (2003) Despite the above caveat, the impressive new technology described by Gupta et al1 in this issue of Circulation: Cardiovascular Imaging should help to resolve a number of issues relative to the hypothesis that the failing heart is energy starved and that depressed high-energy phosphate flux through the creatine kinase (CK) system is a causative factor.2,3 It will allow longitudinal measurements of CK kinetics during the progression to failure in wild-type and transgenic mice with systematically varied amounts of each component of the CK system. Despite years of research, the importance of the several CK isoforms in myocardial bioenergetics is not clear. CK substrates (total creatine, total adenylates) and enzyme activity typically are depressed in heart failure in both murine and human hearts, contributing to the hypothesis that the failing heart is energy starved. However, knockout of myofibrillar (MM) CK and of both MM and sarcomeric mitochondrial CK isoform has surprisingly little effect: Animals are viable, and cardiac function is relatively normal. Thus, there is no obligatory creatine phosphate (PCr) shuttle where high-energy phosphates generated by mitochondrial oxidative phosphorylation must be transported to the myofibrils by way of PCr. Nevertheless, the evolutionary preservation of the complex myocardial CK system speaks to its importance, and the correlation between derangements in this system and contractile dysfunction is intriguing. Article see p 42 It appears that despite the massive difference in size and resting rate between murine and human hearts, the rate of ATP synthesis (≈3 μmol/g per second) …

Measurement of creatine kinase reaction rate in human brain using magnetization transfer image‐selected in vivo spectroscopy (MT‐ISIS) and a volume 31P/1H radiofrequency coil in a clinical 3‐T MRI system

High-energy phosphate metabolism, which allows the synthesis and regeneration of adenosine triphosphate (ATP), is a vital process for neuronal survival and activity. In particular, creatine kinase (CK) serves as an energy reservoir for the rapid buffering of ATP levels. Altered CK enzyme activity, reflecting compromised high-energy phosphate metabolism or mitochondrial dysfunction in the brain, can be assessed using magnetization transfer (MT) MRS. MT (31)P MRS has been used to measure the forward CK reaction rate in animal and human brain, employing a surface radiofrequency coil. However, long acquisition times and excessive radiofrequency irradiation prevent these methods from being used routinely for clinical evaluations. In this article, a new MT (31)P MRS method is presented, which can be practically used to measure the CK forward reaction rate constant in a clinical MRI system employing a volume head (31)P coil for spatial localization, without contamination from the scalp muscle, and an acquisition time of 30 min. Other advantages associated with the method include radiofrequency homogeneity within the regions of interest of the brain using a volume coil with image-selected in vivo spectroscopy localization, and reduction of the specific absorption rate using nonadiabatic radiofrequency pulses for MT saturation. The mean value of k(f) was measured as 0.320 ± 0.075 s(-1) from 10 healthy volunteers with an age range of 18-40 years. These values are consistent with those obtained using earlier methods, and the technique may be used routinely to evaluate energetic processes in the brain on a clinical MRI system.

Concentrations of Total Proteins and Albumins, and AST, AP, CK and GGT Activities in the Blood Serum Boer and Saanen Goats During Puerperium

The metabolism of proteins in the blood serum in Boer and Saanen goats was investigated during puerperium. Twenty Boer goats (10 primiparous and 10 pluriparous) and 10 Saanen goats (five primiparous and five pluriparous) between 2 and 5 years of age were used in this research. Blood for analysis was taken every fourth day from day 3 until day 40 post-partum. Blood samples were collected by jugular puncture. In the obtained blood serum, the concentration of total proteins (PT) and albumin (ALB), and the activity of enzymes aspartate aminotransferase (AST) [the Enzyme Commission number (EC number) 2. 6. 1. 1.], gamma-glutamyltransferase (GGT) (EC 2. 3. 2. 2.), creatine kinase (CK) (EC 2. 7. 3. 2.) and alkaline phosphatase (AP) (EC 3. 1. 3. 1.) were determined by spectrophotometry. These parameters were in physiological ranges in Boer goats and in Saanen goats, without significant differences according to number of kids per doe. According to the research results of the blood serum in goats during puerperium, there were no significant differences in the concentration of ALB. Boer goats had significant higher (p < 0.05) concentration of PT and enzyme activity of AP, CK and GGT. Saanen goats had only enzyme activity of AST significantly higher (p < 0.05). Enzyme activity of alkaline phosphatase was significant higher (p < 0.05) in pluriparous goats in both breeds than in primiparous. The obtained results may represent a contribution to a better understanding of protein metabolism during puerperium in dairy and meat goats and for diagnostic purposes.

The estimation of post-transplant lymphocele origin using creatine kinase activity

Introduction. The aim of this research was to create a laboratory instrument for the estimation of post-transplant lymphocele origin. It is based on the enzymatic activity of creatine kinase (CK) in the lymphocele content.Material and methods. A total of 120 lymph samples from different retroperitoneal regions were obtained from non-transplanted patients; equal numbers from the iliac region, renal cysts, and the subdiaphragmatic retroperitoneum. Activities of creatine kinase (CK) and γ-glutamyl transpeptidase (GGT) were determined in all samples and statistically analyzed against their activity in serum from patients without surgery.Results. Activities of CK in the pelvis, retroperitoneum, renal cysts, and serum were 5.06, 0.83, 6.48 (P < 0.001), 2.50, 0.73, 3.60 (P < 0.001), 0.02, 0.01, 0.05 (P < 0.001), and 0.66, 0.41, 0.79 μkat/l, respectively. Activities of GGT in the same lymph samples were 0.26, 0.16, 0.36 (P = 0.048), 0.41, 0.25, 0.48 (non-significant), 0.11, 0.07, 1.17 (P = 0.003) and 0.34, 0.24, 0.55 μkat/l, respectively. A graph was constructed relating CK activity to lymph origin.Conclusion. Significantly different CK enzyme activity was observed in different regions of the retroperitoneum. The presented graph is a simple instrument for the estimation of the lymphocele content origin. The method requires percutaneous aspiration of the lymphocele and evaluation of the CK and GGT activity in the sample. From the graph the estimated proportion of renal lymph in the lymphocele can be read directly. This instrument can provide better understanding of post-transplant lymphocele fluid source.

The Virulence of S. Aureus Is Supported by Host Fibrinogen and Its Bacterial Receptor ClfA in the Context of Bacteremia.

Abstract 3188Poster Board III-125The pervasive gram-positive bacterial pathogen Staphylococcus aureus has evolved and maintained a number of proteins that directly engage the host hemostatic system, including factors that directly interact with the host coagulation factor fibrinogen. Here, we investigate the hypothesis that binding of host fibrin(ogen) through the bacterial fibrinogen receptor, clumping factor A (ClfA), significantly contributes to virulence in the context of acute bacteremia. To directly test this view, we compared survival following intravenous injection of ClfA-postive S. aureus into fibrinogen-deficient and control mice. The genetic elimination of fibrinogen resulted in a dramatic prolongation in survival relative to fibrinogen sufficient control animals. To determine if this prolongation in survival was related to the ability of the pathogen to directly bind fibrinogen, we performed identical survival studies in mice expressing a mutant form of fibrinogen that retains normal clotting function but lacks the binding motif for ClfA located on the fibrinogen γ chain (termed Fib γΔ5). A profound survival advantage was observed for Fib γΔ5 mice relative to wild-type animals over a wide bacterial dose range corresponding to an LD50 to an LD100 for control animals. This difference in survival appeared to be linked to ClfA, as control and Fib γΔ5 mice revealed an identical survival profile when infected with ClfA-deficient S. aureus. Analysis of bacterial burden after infection in various organ systems revealed that ClfA-positive S. aureus rapidly marginalized from the circulation and colonized target organs in mice of both genotypes. The highest numbers of colony forming units (cfu) were found in the heart and kidney of challenged mice, but the cfu values observed in both of these tissues were significantly reduced in Fib γΔ5 mice, relative to control animals at 24 and 48 hours after initial infection. The cfu values also tended to be reduced in Fib γΔ5 mice over wild-type controls in the liver, lung, spleen and blood, but this difference did not achieve statistical significance. Complementary studies revealed a comparable systemic inflammatory response in control and Fib γΔ5 animals following initial infection; plasma levels of Il-6 and fibrinogen (an established acute phase reactant) were similarly elevated in both genotypes following infection. Notably, increased histological evidence of cardiac damage secondary to S. aureus bacteremia in wild-type mice relative to Fib γΔ5 animals correlated with significantly higher plasma levels of markers of cardiac damage (i.e., lactose dehydrogenase and muscle creatine kinase enzyme activity) 48 hours after infection. These data suggest that bacterial engagement of host fibrinogen via the bacterial fibrinogen receptor ClfA significantly contributes to S. aureus virulence following intravenous infection. Strategies designed to disrupt this interaction may be of significant clinical benefit to limit disease progression in S. aureus sepsis and could be achieved without necessarily compromising host hemostatic function. DisclosuresNo relevant conflicts of interest to declare.

Development and validation of a sensitive immunoassay for the skeletal muscle isoform of creatine kinase

Creatine kinase (CK) is a marker of muscle damage and pathology present as multiple tissue-specific circulating isoforms. CK is often measured using enzyme activity assays that are unable to distinguish these isoforms. We have developed an immunoassay specific for the MM isoform of CK, found predominantly in skeletal muscle, which uses very small volumes of plasma (1–2 μL). A sandwich enzyme-linked immunosorbent assay (ELISA) for CK-MM was developed using isoform-specific antibodies. Cross-reactivity with CK-BB and MB isoforms was also assessed. The ELISA was validated using plasma samples from a group of athletes, and the measured CK-MM concentrations were correlated with CK enzyme activity assays measured by a contractor using the same samples. The CK-MM ELISA has a limit of detection of 0.02 ng/mL, an IC 50 of 2.3 ng/mL, and 5.8% cross-reactivity with CK-MB. CK-MM concentrations measured using this assay correlate well ( p < 0.0001, Spearman r = 0.89) with enzyme activity assays. The CK-MM-specific ELISA can be used to help assess skeletal muscle damage independent of enzyme activity or interference from other CK isoforms, leading to more precise studies of muscle biology.

Proteomic Approach for Caudal Trauma-Induced Acute Phase Proteins Reveals That Creatine Kinase Is a Key Acute Phase Protein in Amphioxus Humoral Fluid

Elevated creatine kinase (CK) in the circulation was generally regarded to be a passive release from muscle damage. We utilized proteomic methodologies to characterize amphioxus humoral fluid APPs in response to caudal trauma, and found several spots of CK alterations with up-regulation and pI shift. Its amount and enzyme activity showed a dynamic pattern of APP in humoral fluid accompanied with a reduction in enzyme activity of muscle, whereas there was no significant difference in CK amount of muscle and the other tissues and in CK enzyme activity of the other tissues between different time points of sample collection following caudal trauma. In addition, CK phosphorylation regulation during injury was not achieved by monoclonal antibodies separately against phosphothreonine, phosphotyrosine, and phosphoserine. These results suggested that the CK elevation of humoral fluid might be from muscle, being an active response to caudal trauma rather than a passive release from muscle damage. Therefore, CK ability in response to caudal trauma should be highly concerned.

Atividade sérica das enzimas AST, CK e GGT em cavalos Crioulos

O objetivo deste trabalho foi verificar a influência da idade, sexo, manejo e estado gestacional sobre a atividade sérica das enzimas: aspartato aminotransferase (AST), creatina quinase (CK) e gama-glutamiltransferase (GGT) em cavalos da raça Crioula. Foram utilizados 142 eqüinos, divididos em seis grupos: potros até um ano de idade; cavalos adultos em regime de atividade livre; cavalos adultos em treinamento; machos adultos; fêmeas não gestantes; fêmeas gestantes. O valor da CK foi mais elevado em animais adultos do que em potros, o mesmo tendo ocorrido em animais em atividade livre, comparados a animais em treinamento. Fêmeas não gestantes apresentaram valores das enzimas CK e GGT maiores que os machos; o valor da CK em fêmeas gestantes foi mais elevado do que em fêmeas não gestantes. A idade, o sexo, o manejo e o estado gestacional influenciam a atividade sérica da CK, e o sexo influencia a atividade sérica da GGT.

Creatine kinase activity in patients with diabetes mellitus type I and type II.

Diabetes mellitus can be looked upon as an array of diseases, all of which exhibit common symptoms. While pathogenesis of IDDM (insulin dependant diabetes mellitus) is well understood, the same is not true for diabetes mellitus type II. In the latter case, relative contribution of the two factors (insulin resistance or decreased insulin secretion) varies individually, being highly increased in peripheral tissues and strictly dependant on insulin for glucose uptake. Moreover, in patients with diabetes mellitus type II, disbalance at the level of regulation of glucose metabolism as well as lipid metabolism has been noted in skeletal muscles. It is normal to assume that in this type of diabetes, these changes are reflected at the level of total activity of enzyme creatine kinase. This experimental work was performed on a group of 80 regular patients of Sarajevo General Hospital. Forty of those patients were classified as patients with diabetes type I and forty as patients with diabetes type II. Each group of patients was carefully chosen and constituted of equal number of males and females. The same was applied for adequate controls. Concentration of glucose was determined for each patient with GOD method, while activity of creatine kinase was determined with CK-NAC activated kit. Statistical analysis of the results was performed with SPSS software for Windows. Obtained results point out highly expressed differences in enzyme activity between two populations examined. Changes in enzyme activity are more expressed in patients with diabetes type II. Positive correlation between concentration of glucose and serum activity of the enzyme is seen in both categories of diabetic patients which is not the case for the patients in control group. At the same time, correlation between age and type of diabetes does exist . This is not followed at the level of enzyme activity or concentration of glucose.

Inhibition of mechanosensitive cation channels inhibits myogenic differentiation by suppressing the expression of myogenic regulatory factors and caspase‐3 activity

Mechanosensitive cation channels (MSC) are ubiquitous in eukaryotic cell types. However, the physiological functions of MSC in several tissues remain in question. In this study we have investigated the role of MSC in skeletal myogenesis. Treatment of C2C12 myoblasts with gadolinium ions (MSC blocker) inhibited myotube formation and the myogenic index in differentiation medium (DM). The enzymatic activity of creatine kinase (CK) and the expression of myosin heavy chain-fast twitch (MyHCf) in C2C12 cultures were also blocked in response to gadolinium. Treatment of C2C12 myoblasts with gadolinium ions did not affect the expression of either cyclin A or cyclin D1 in DM. Other inhibitors of MSC such as streptomycin and GsTMx-4 also suppressed the expression of CK and MyHCf in C2C12 cultures. The inhibitory effect of gadolinium ions on myogenic differentiation was reversible and independent of myogenic cell type. Real-time-polymerase chain reaction analysis revealed that inhibition of MSC decreases the expression of myogenic transcription factors MyoD, myogenin, and Myf-5. Furthermore, the activity of skeletal alpha-actin promoter was suppressed on MSC blockade. Treatment of C2C12 myoblasts with gadolinium ions prevented differentiation-associated cell death and inhibited the cleavage of poly (ADP-ribose) polymerase and activation of caspase-3. On the other hand, delivery of active caspase-3 protein to C2C12 myoblasts reversed the inhibitory effect of gadolinium ions on myogenesis. Our data suggest that inhibition of MSC suppresses myogenic differentiation by inhibiting the caspase-3 activity and the expression of myogenic regulatory factors.