(Bicarbossilasi®) Administration on the Exercising Horse Metabolism
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Published in IVIS with the permission of the editor Close window to return to IVIS Effect of Thiamine Pyrophosphate (Bicarbossilasi®) Administration on the Exercising Horse Metabolism Laus, F.,1* Faillace, V.,2* Tesei, B.,3 Paggi, E.,1 Serri, E.,1 Marini, C.,1 Marvasi, L.,2 Vullo, C.3 and Spaterna, A.1 * The authors contribute equally to this work. 1 School of Biosciences and Veterinary Medicine, University of Camerino.Via Circonvallazione 93/95, 62024 Matelica (MC), Italy. 2 FarefarmaSrl (www.farefarma.com), Contract Research Organization, Via Ferrari , 9 28045 Invorio (NO), Italy. 3 School of Pharmacy.University of Camerino, Via Madonna delle Carceri 9, 62032, Camerino (MC), Italy. * Corresponding author: Dr. Fulvio Laus, School of Biosciences and Veterinary Medicine, Via Circonvallazione 93/95, 62024 Matelica (MC). Phone: +39 0737 403439, Fax: +39 0737 403402 Email: [email protected] ABSTRACT Thiamine pyrophosphate (TPP) is the phosphorylated and active form of thiamine (Vitamin B1). We hypothesized that the administration of thiamine in its immediately available active form could provide the metabolic pathways for a supplement able to promote the metabolism of ketoacids and to reduce lactate accumulation in exercising horses. Ten horses were conditioned for 20 days to daily standardized exercise. All horses underwent a first “stress test” (ST) consisting in 1,200 meters at maximum speed on track, and were checked before and after for clinical and clinical pathology parameters. After the ST, the same animals were administered TPP (Bicarbossilasi®, Teknofarma S.p.A., Torino, Italy), 1 mg/Kg b.w., I.V., twice daily for seven days. At the end of treatment, a second stress test (STTPP) was performed. Post-exercise serum lactate concentration resulted in significantly lower levels (p< 0.05) after treatment with TPP (ST vs STTPP). These data suggest that supplementation with thiamine in its active form improves glucose metabolism and prevents lactate accumulation in muscle, enhancing aerobic capacity and metabolic pathways of glucose utilization during exercise. Keywords: Horse; Equine; Exercise; Vitamin B; Thiamine; Glucose Metabolism; Lactate. INTRODUCTION of ketoacids and plays an essential role in the oxidative de- Vitamin B plays an important role in energy metabolism and carboxylation of pyruvate, allowing the formation of acetyl is considered a key nutrient in exercising horses (1). Thiamine coenzyme-A under aerobic condition. The latter will enter pyrophosphate (TPP) is the phosphorylated and activated in the Krebs cycle to allow for energy production (1). Since form of thiamine (Vitamin B1). The activation of thiamine pyruvate is derived from glucose, it should be emphasized occurs primarily in the liver with ATP consumption, and that the energy drive from glucose oxidation is highly de- once phosphorylated, the molecule acquires the active form pendent upon TPP (3). as TPP. All living organisms needs thiamine, but only bac- During short and intense efforts, especially in the most teria, fungi, and plants are able to synthesize it. Thiamine is energy demanding tissue such as muscles, an excess of py- an essential nutrient for animals and humans, and must be ruvate is produced as the final product of glycolysis and can obtained from the diet or through metabolic production by be converted to lactic acid (4). The excess of lactic acid is gut microorganisms (2). transferred into the blood and reaches the liver where it is TPP mainly acts as coenzyme for the decarboxylation converted back to pyruvate and then glucose (Cori cycle) (4). Israel Journal of Veterinary Medicine Vol. 72 (2) June 2017 Thiamine Pyrophosphate supplementation in horses 15 Published in IVIS with the permission of the editor Close window to return to IVIS Research Articles If the effort is continuous, active muscles may fail to transfer Breeds of horses included four Sella Italiano, three Anglo- the excessive lactic acid into the blood, which contributes to Arabian, two Murgese and one Tolfetano. All horses were symptoms of “fatigue” due to the alteration of the physiologi- active at the time of the study. The age of the horses ranged cal pH of the cells. from 4 to 21 year (mean: 10.6 ± 6.0 years), and their weight The term “fatigue” indicates a clinical condition of athletic ranged from 407 to 600 Kg (mean: 495.2 ± 57.3 Kg). All horses causing the interruption of exercise or a reduction in animals, once included in the study, were stabled in boxes and its intensity (5). In case of high-intensity exercise, fatigue subjected to a standardized diet: hay and water ad libitum and is caused by the accumulation of catabolites in blood and integrated with complementary feed corresponding to about muscle which affects the contractile capacity of myocytes (6). 1% of their body weight divided into two daily rations. Clinical It is evident that as the threshold for “fatigue” increases, the assessment was performed daily to check for any symptoms improvement in the performance of the horse will increase that could cause withdrawing the animals from the study. as well. Studies in human athletes demonstrated that fatigue can be delayed and performance enhanced if blood glucose Study design levels are maintained by oral or intravenous supplementation Before starting the study, the animals were conditioned for of glucose (7). Given the crucial involvement of thiamine in 20 days to daily standardized training sessions lasting 35 glucose metabolism and the aerobic pathway, particularly in minutes and consisting of a warming-up phase (walk for the muscles, its importance is evident during exercise and in 10 minutes), followed by 5 minutes trot, 5 minutes gallop, 5 the onset of “fatigue” (1). minutes trot and 10 minutes walk. Thiamine deficiency can cause neurologic disorders and At the end of the 20 days, the horses underwent their thiamine supplementation can relieve symptoms or resolve first stress test (ST): 400 meters walk followed by 800 meters several neurologic diseases in cats (8, 9), dogs (10), cows (11, trot; the horses then performed an exercise at their maximum 12) and humans (13, 14). Reduction in serum lactate concen- potential (maximum speed) for 1,200 meters. Clinical and tration has also been observed in pigs after TPP administra- clinical pathology evaluations were performed before (T0) tion (15). Cocarboxylase was found to be beneficial in the and immediately after (T1) the ST. The post-stress evalua- ischemic canine myocardium due to its favorable systemic tions were made directly on the track immediately after each hemodynamic effects (16). horse had stopped. The effect of thiamine administration has been tested Starting from the first day, the administration of TPP was in human athletes with different results. In some studies commenced, at the dose of 1 mg/Kg b.w., I.V., twice a day thiamine was able to reduce serum lactate concentration and continued for seven days. At the end of the administra- and to improve resistance to fatigue (17, 18). On the other tion period, the horses underwent a second stress test (STTPP) hand, other surveys did not obtain similar results (19, 20). with the same protocol previous described and with clinical A previous study in athletic horses has shown that the oral and clinical pathology evaluation as previously performed (at administration of supplemental thiamine decreased the levels T0TPP and at T1TPP). of serum lactate during exercise (21). The horses were ridden by the same rider during both Thiamine pyrophosphate is the active ingredient of the ST and STTPP. veterinary drug Bicarbossilasi® (Teknofarma S.p.A., Torino, Italy). In the present study, the effect of thiamine pyro- Sampling and clinical pathology evaluations phosphate administration to athletic horses during intense Venous blood samples were collected from the jugular vein exercise has been tested in a clinical, cross over trial. into sterile tubes with and without ethylene diaminetetraacetic acid (EDTA) at T0, T1, T0TPP and T1TPP and maintained MATERIALS AND METHODS at 4°C until the delivery to the laboratory within 3 hours from collection. The following hematologic and biochemi- Animals cal parameters were measured within 12 hours from sample Ten pleasure horses (6 male and 4 female) normally used for collection: erythrocytes (red blood cells, RBC), packed cell recreational riding (e.g. trail riding) were included in the study. volume (PCV), hemoglobin (Hb), red cell distribution width 16 Laus, F. Israel Journal of Veterinary Medicine Vol. 72 (2) June 2017 Published in IVIS with the permission of the editor Close window to return to IVIS Research Articles (RDW), mean corpuscular volume (MCV), mean corpuscular Statistical analysis hemoglobin (MCH), mean corpuscular hemoglobin concen- All results are expressed as the mean value and standard devia- tration (MCHC), total leukocytes, neutrophils, lymphocytes, tion (SD) for each stage of the test. Data collected at T0 and monocytes, eosinophils, basophils and platelets along with T1 were compared with those obtained at T0TPP and T1TPP, PDWc (platelet distribution width) and MPV (mean platelet respectively, by the Student’s t-test. For serum lactate, the com- volume) (Cell Dyn 3500, Abbott, Chicago, IL, USA). parison was performed between each sampling stage before The biochemical profile included aspartate aminotrans- (T0, T1, T1+2h, T1+8h, T1+18h) and after (T0TPP, T1TPP, ferase (AST), gamma glutamyltransferase (GGT), creatine T1TPP+2h, T1TPP+8h, T1TPP+18h) thiamine administration. phosphokinase (CPK), total protein (TP), albumin (Alb), P value for statistical significance was set at <0.05 for urea, creatinine (Crea), glucose (Glu), alkaline phosphstase all tests. Statistical analyses were performed using the (ALP), lactate dehydrogenase (LDH), total bilirubin (TB), WINPEPI (PEPI-for-Windows) computer programs (22). direct bilirubin (DB), triglycerides (Tryg), cholesterol (Chol), calcium, phosphorus, magnesium, sodium, potassium, and chlorine (Targa 3000 plus, Biotecnica Instruments, Italy, RESULTS Rome). None of the horses showed clinical signs that could result in Blood lactate was analysed using a portable device withdrawal from the study.