Comparative biochemical studies of myotoxic phospholipase A(2) from Bothrops venom

Venoms from Bothrops jararacussu, Bothrops asper, Bothrops atrox, Bothrops pirajai, Bothrops moojeni, Bothrops alternatus and Bothrops (Bothriopsis) bilineata were fractionated using a simplified procedure based on ion-exchange chromatography on CM-Sepharose at pH 8.0 or reverse phase HPLC. The resulting elution profiles showed important differences in the myotoxin content of these venoms. The venoms from B. alternatus, B. atrox and Bothriopsis bilineata did not contain the major myotoxin found in the other venoms. The amino acid sequence of the first 50 residues of the N-terminal region of the PLA(2)-like myotoxins showed a homology of 90-96% with other bothropic myotoxins. All of the myotoxins isolated induced rat paw edema, increased the level of plasma creatine kinase and produced myonecrosis together with polymorphonuclear cell infiltration.

Myotoxic effects of mastoparan from Polybia paulista (Hymenoptera, Epiponini) wasp venom in mice skeletal muscle

In a previous study, we showed that the Polybia paulista wasp venom causes strong myonecrosis. This study was undertaken to characterize the myotoxic potency of mastoparan (Polybia-MPII) isolated from venom (0.25 mu g/mu l) and injected in the tibial anterior (TA) muscle (i.m.) of Balb/c mice. The time course of the changes was followed at muscle degenerative (3 and 24 h) and regenerative (3, 7, and 21 days) periods (n = 6) after injection and compared to matched controls by calculation of the percentage of cross-sectional area affected and determination of creatine kinase (CK) activity (n = 10). The results showed that although NIP was strongly myotoxic, its capacity for regeneration was maintained high. Since the extent of tissue damage was not correlated with the CK serum levels, which remained very low, we raised the hypothesis that the enzyme underwent denaturation by the peptide. Evidence suggested that MP induced the death of TA fibers by necrosis and apoptosis and had the sarcolemma as its primordial target. Given its amphiphilic polycationic nature and based on the vast spectrum of functions attributed to the peptide, we suggest that MP interaction with cell membrane impaired the phosphorylation of dystrophin essential for sarcolemma mechanical stability, and disturbed Ca2+ mobilization with obvious implications on sarcoplasmic reticulum and mitochondrial functioning. (c) 2007 Elsevier Ltd. All rights reserved.

Efeito protetor após sessões de exercício excêntrico: comparação entre membros superiores e inferiores

O objetivo foi comparar a magnitude do efeito protetor (EP) contra o dano muscular (DM) induzido por uma sessão de exercícios excêntricos (EEM) entre os extensores do joelho e os flexores do cotovelo. Doze sujeitos do gênero masculino foram divididos em 2 grupos, braços (GB) e pernas (GP), e realizaram 2 sessões de EEM. Foram coletados 3 marcadores de DM, sendo eles, pico de torque isométrico (PTI), creatina quinase (CK) e percepção subjetiva de dor (PSD), antes, imediatamente após (com exceção da CK) e 48 horas após cada sessão de EEM. Foi encontrada queda significante de PTI e aumento significante de CK e PSD tanto imediatamente e 48 horas após a primeira sessão de EEM para o GB. No GP houve aumento significante de CK 48 horas após os EEM e da PSD imediatamente após os EEM, decorrentes da primeira sessão. No GB, a segunda sessão apenas provocou queda de PTI imediatamente após os EEM, enquanto no GP houve aumento significante apenas na PSD imediatamente após a segunda sessão de EEM. Apenas a CK apresentou EP para ambos os grupos. Pudemos concluir que o EP foi maior para o GB em comparação com o GP. Esse fenômeno pode ter ocorrido em detrimento da existência de um EP prévio para o GP, uma vez que este membro realiza contrações excêntricas intensas com maior freqüência no dia-a-dia, quando comparados com os GB.

Padronização de um protocolo experimental de treinamento periodizado em natação utilizando ratos Wistar

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Psychological, biochemical and physiological responses of Brazilian soccer players during a training program

Aims of the study. - The aim of this study is to investigate the behavior of the physiological, biochemical and psychological parameters in Brazilian soccer players during a training program.Materials. - Fifteen athletes were evaluated at the beginning (T1), in the middle (T2) and at the end (T3) of the training program. on the first day, at 7:30 am, before the blood collecting at rest for the determination of serum creatine kinase (CK), serum creatinine and serum urea, the athletes had their psychological parameters assessed by the profile of mood state questionnaire (POMS). After 90 min, they performed a 250-m sprint. on the second day at 8:30 am, the athletes had their alactic anaerobic performance measured and, after 40 min, they completed the aerobic test. Friedman test was used to verify the behavior of overtraining markers.Results. - There was a decrease in vigor score in T3 (p=0.01) compared with T1 and T2. In T3 (p=0.01), the athletes also showed an increase in serum creatinine levels compared to T1. Furthermore, in the same period, we verified a diminishing in the team performance.Conclusion. - The training program developed between T2 and T3 led to the fall of the vigor score, the increase in serum creatinine concentrations and the diminishing in team performance. (c) 2007 Elsevier Masson SAS. All rights reserved.

Heavy-intensity aerobic exercise affects the isokinetic torque and functional but not conventional hamstrings:quadriceps ratios

Running exercises are frequently related to muscular injuries, which may be a result of muscular imbalance. The present study aimed to verify the effects of heavy-intensity continuous running exercise on the functional and conventional hamstrings: quadriceps ratios, and also in the knee flexors and extensors EMG activity in active non-athletic individuals. Sixteen active males performed maximal isokinetic concentric and eccentric knee flexions and extensions at 60 degrees s(-1) and 180 degrees s(-1). In another session, the same procedure was conducted after a continuous running exercise at 95% onset of blood lactate accumulation. Torque and electromyographic ratios were calculated from peak torque and integrated electromyographic activity (knee flexor and extensors). Creatine kinase was measured before and 24 h after running exercise. Eccentric torque (knee flexion and extension) decreased significantly after running only at 180 degrees s(-1) (p < 0.05). No differences were found for the conventional torque ratios (p > 0.05), however, the functional torque ratios at 180 degrees s(-1) decreased significantly after running (p < 0.05). No effects on the electromyographic activity and electronnyographic ratios were found (p > 0.05). Creatine kinase increased slightly 24 h after running (p < 0.05). Heavy-intensity continuous running exercise decreased knee flexor and extensor eccentric torque, and functional torque ratios under fast velocities (180 degrees s(-1)), probably as result of peripheral fatigue. (C) 2008 Elsevier Ltd. All rights reserved.

Muscle damage slows oxygen uptake kinetics during moderate-intensity exercise performed at high pedal rate

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Parâmetros bioquímicos de equinos submetidos à simulação de prova de enduro recebendo dietas com adição de óleo de soja

Com o objetivo de verificar o efeito da adição de níveis crescentes de óleo no concentrado sobre a atividade plasmática das enzimas creatina quinase (CK), aspartato aminotransferase (AST) e lactato desidrogenase (LDH) como indicativo de metabolismo energético, foram fornecidas dietas experimentais compostas de cinco níveis de óleo (controle, 6, 12, 18 e 24% do concentrado). Utilizaram-se 20 equinos da raça Árabe, peso médio de 400 kg, submetidos a prova de enduro de 80 km em esteira rolante. O enduro foi dividido em quatro anéis de 20 km, com duração média de 1 hora e dez minutos. A adição de óleo e a distância percorrida tiveram efeitos sobre as variáveis AST, CK e LDH, que apresentaram as respectivas expressões: AST (7,045-0,2292x+0,007991x2+0,008517z- 0,0003282xz), CK (8,06-,07020x+0,05546x2-0,001262x3+0,01204z+0,0006207xz) e LDH (6,624-0,3522x+0,03448x2-0,0008382x3+0,02401z-0,0007489xz) . O óleo é uma importante e bem aproveitada fonte de energia para equinos em exercício, pois sua adição na dieta de animais submetidos a prova de enduro promoveu alteração metabólica que favorece a produção de energia. O metabolismo animal poupou suas reservas energéticas oriundas da glicose, favorecendo a utilização do óleo. A menor atividade plasmática das enzimas AST, CK e LDH com a adição de óleo nas dietas indica direcionamento do metabolismo energético para a β-oxidação. Como apresentam várias isoenzimas, as enzimas estudadas atuam amplamente no metabolismo energético, favorecendo a constante reposição de ATP ao longo do exercício.

Dietary macronutrients, endocrine functioning and intermediary metabolism in broiler chickens - Pair wise substitutions between protein, fat and carbohydrate

Three pens of male broiler chicks were raised under standard conditions and fed from 7 to 42 days of age three isocaloric diets each with 15.8; 19.6 and 19.5% of CP; and 51, 51, and 44% of CHO; and 6.5; 3.0 and 7.7% of fat, and designated as the low protein (LowCP), low lipid (LowL) and low carbohydrate (LowCHO) diets, respectively. Body weights and feed intake were monitored weekly and blood samples were collected at the same time for posterior analysis of hormone and metabolite content. Chickens fed the LowCP diet were characterized by a reduced body weight gain and feed intake and poorer feed conversion efficiency compared to those fed the LowL and LowCHO diets, which were very similar in this respect. Plasma corticosterone and glucose levels and creatine kinase activity were not significantly changed by diet composition. LowCP chickens were characterised by the lowest plasma T-4 and uric acid levels (indicative for reduced protein breakdown and lower protein ingestion) but highest plasma triglyceride levels (congruent with their higher fat deposition) compared to the LowL and LowCHO chickens. LowL chickens had on average higher plasma T-3 and free fatty acid levels compared to the LowCP and LowCHO chickens.In conclusion, a limited substitution of carbohydrate for fat in iso-nitrogenous, iso-energetic diets has no pronounced effects on plasma hormone and metabolite levels, except for the elevation in T-3 (may enhance glucose uptake) and free fatty acid levels in the plasma of the chickens fed the LowL diet. The protein content of the diet has a greater impact on zootechnical performance, and underlying endocrine regulation of the intermediary metabolism compared to the dietary lipid and CHO fraction. (C) 2003 Elsevier B.V. All rights reserved.

ANTAGONISM OF THE MYOTOXIC EFFECTS OF BOTHROPS-JARARACUSSU VENOM AND BOTHROPSTOXIN BY POLYANIONS

The effects of heparin and other polyanions on the myotoxicity of Bothrops jararacussu venom and purified bothropstoxin (BthTX) were investigated. The release rate of creatine kinase (CK) from isolated extensor digitorum longus muscle and the plasma CK activity of mice were used to quantify the results. The myotoxic effects of B. jararacussu venom or BthTX were inhibited by preincubation of these agents with one of the following: a heterogeneous heparin preparation (designated 'heparin'), low mol. wt heparin (H-4500) or dextran sulfates (DS-8000 and DS-500,000). Non-sulfated dextran (D-40,000) and two chondroitin sulfates were ineffective. The antimyotoxic effects of the polyanions are ascribed to their forming inactive acid-base complexes with the basic myotoxins of Bothrops venoms. Gel-filtration experiments in Sephadex provided direct evidence for complex formation between heparin and BthTX. Intravenous (i.v.) administration of H-4500 or DS-8000 opposed the increase in plasma CK activity induced by a subsequent i.m. injection of venom or BthTX. In contrast, pretreatment with i.v. heparin or DS-500,000 enhanced the venom-induced increase in plasma CK activity. This effect was not observed (1) when the animals were treated with a polyvalent antivenom, which inhibits the coagulation and local stasis induced by Bothrops venoms, and (2) when BthTX, which has no thrombotic or hemorrhagic properties, was the myotoxic agent. The potentiation of the venom-induced increase in plasma CK activity by heparin and DS-500,000 is ascribed to improved washout of the CK released from damaged fibers, because of the anticoagulant properties of the drugs.

Environmental cadmium exposure and metabolic responses of the Nile tilapia, Oreochromis niloticus

The contamination of water by metal compounds is a worldwide environmental problem. This study was undertaken to evaluate the impact of short-term cadmium exposure on metabolic patterns of the freshwater fish Oreochromis niloticus. The fish were exposed to 320, 640, 1280 and 2560 mug/l sublethal concentrations of Cd++ (CdCl2) in water for 7 days. The specific activities of the enzymes phosphofructo kinase (PFK-E.C.2.7.1.11.), lactate dehydrogenase (LDH-E.C.1.1.1.27.) and creatine kinase (CK-E.C.2.7.3.2.) were decreased in white muscle after cadmium treatments, indicating decreases in the capacity of glycolysis in this tissue. Cadmium exposure induced increased glucose concentration in white muscle of fish. on the other hand, cadmium exposure at sublethal concentrations increased phosphofructo kinase and LDH in red muscle of fish. Cadmium significantly decreased total protein concentrations in liver and white muscle regardless of tissue glycogen levels. The data suggest that cadmium acts as a stressor, leading to metabolic alterations similar to those observed in starvation. (C) 2001 Elsevier B.V. Ltd. All rights reserved.

Acclimation of broiler chickens to chronic high environmental temperature

The present study was conducted to evaluate the zootechnical parameters and age related changes in physiological responses of broiler chickens exposed to hot environment from early age onwards. The broiler chickens were exposed to high temperature (30 degreesC) at 15 d of age and maintained to Day 38 or maintained under thermoneutral environment (control).No significant decrease in feed consumption (FC) and body weight (BW) gain was observed in high temperature group after 7 d of exposure, but in the subsequent period, heat exposure lowered BW and FC, compared to control group. However, the weight gain was not significantly changed after 24 d of exposure, and the feed efficiency was not affected throughout the experimental period.The venous pCO(2) pressure was only significantly decreased by temperature after 24 d of heat exposure. The glucose, non-esterified fatty acid (NEFA), triglyceride (TG), glucose, lipid peroxidation (LPO), creatine kinase (CK), and corticosterone were not influenced by the temperature treatment. The significant decrease in uric acid and increase in lactate concentration due to high temperature were observed respectively at 28 and 35 d of age. The concentrations of triiodothyronine (T-3) and thyroxine (T-4) were changed oppositely at 28 d of age, as T-3 was decreased and T-4 was elevated by high temperature. However, the concentration of T-4 in plasma was decreased whereas T-3 was not changed at 38 d of age. The relationships between the blood parameters were changed due to the temperature treatment, suggesting that not only absolute values but also their interrelationships have to be considered when studying the effects of a particular treatment on physiological functioning.These results suggest the growth and physiological responses of broiler chickens, exposed to high temperature from early age onwards, differed at different stages of acclimation. The process of heat acclimation is related to the mode of heat exposure imposed and is not only reflected in the changes in the absolute concentrations, but also in the correlations among the blood indices.

Dietary macronutrients and performance and plasma hormone and metabolite levels of broiler chickens fat by carbohydrate substitution

The present experiment was aimed to study the effects of an isocaloric substitution of fat by carbohydrate calories while maintaining the same CP content - on some endocrine parameters and key metabolites of the lipid (L), protein (CP) and carbohydrate (CHO) metabolism and muscle cell membrane integrity of broiler chickens. Ninety male broiler chicks (Cobb) were obtained at 1-day old and raised in an environmentally controlled room. Food and water was provided ad libitum. At day 7, the chicks were provided with the experimental diets: Control diet (CON: 20% CP; 9.1% L; 39.6% CHO), Low lipid diet (LowL: 22% CP; 3.36% L; 59.7% CHO), and low carbohydrate diet (LowCHO: 22% CP; 8.09% L; 46.7% CHO). T e chickens fed the CON manifested the best growth rate feed conversion compared to both other groups. The CO chickens showed the highest plasma T-3 Concentrations at 14 days of age as compared to both LowL and LowCHO chickens. At 35 days of age however, LowL chickens were characterized by the highest plasma T-3 levels. Overall, CON birds had the highest plasma T-4 levels (P < 0.0001) compared to LowL and LowCHO chickens. Plasma corticosterone levels were not affected by diet composition. Glucose levels increased towards the end of the experimental period, but there was no overall effect of diet composition on plasma glucose levels. Chickens fed the CON diet were consistently characterized by the highest plasma uric acid levels compared to LowL and LowCHO chickens. Irrespectively of diet, plasma triglyceride levels were high at 14 days of age, then decreased and finally increased again during the last week. This latter increase was most pronounced for the CON chickens and is congruous with their high fat deposition. At 28 days of age, a significant effect of diet was observed, as CON birds had lower plasma free fatty acid (FFA) levels compared to LowL but not to LowCHO chickens. However there was no overall effect of diet composition on plasma FFA levels. Creatine kinase (CK) activities increased significantly with age and were consistently higher in CON chickens. In conclusion, fast growth in broiler chickens is positively correlated with catabolism of protein from dietary or endogenous origin and (muscle) cell membrane disruption as reflected in high plasma uric acid levels and CK activity, respectively. The currently investigated substitution of fat by CHO had no marked effects on endocrine functioning and intermediary metabolism, but this does not exclude that larger substitutions between both nutrients can have an effect.

Clinical pathological data and analyses of mineral elements of cattle affected by epizootic botulism in the State of São Paulo

In order to study laboratorial aspects of beef cow mortality, a syndrome popularly known as ''doenca da vaca caida'', examens were made of blood, cerebrospinal fluid, serum, bone and liver samples from 32 naturally affected 4 to 9 year old cows, 27 belonging to the Nellore breed and 5 were crossbred Nellore, all originating from farms located in municipalities near Botucatu, State of São Paulo. Laboratory determinations were analysed by descriptive statistics and included hematological values, total plasma protein, plasma fibrinogen, cerebrospinal fluid analysis, and concentration measurements of serum calcium, phosphorus, magnesium, sodium, potassium, chloride, total protein, albumin, globulin, alkaline phosphatase, aspartate aminotransferase, gama-glutamyltransferase and creatine kinase activities, included bone ash percentage and concentrations of calcium, phosphorus and magnesium, and also hepatic levels of copper, zinc, iron, manganese and cobalt. In addition, mouse bioassays and complement micro-fixation tests were performed to detect botulinum toxins in liver samples. The results indicated leukocytosis (13,3+/-3,9 x10(3)/mm(3)) with neutrophilia (8,9+/-3,2 x10(3)/mm(3)), hypocalcemia (7,8+/-1,7mg/dl), hypophosphatemia (3,6+/-1,6mg/dl), hypoalbuminemia (2,9+/-0,9g/dl), increased creatine kinase activity (691,0+/-829,7 UI/1), and reduced ash percentage (60,3+/-1,9%) and low phosphorus (17,2+/-0,4%) in bone. The other values were ail within normal limits. The diagnosis of botulism, involving type C and D toxins, was confirmed as the cause of the mortality in the region of study, what is strongly consistent with the other laboratorial findings.

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 a-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.