Description and evaluation of a diabetic dyslipidemia model in Gottingen minipigs.

D Celdrán, F.M Sánchez-Margallo, B Moreno-Lobato, M.F. Martín-Cancho. Minimally Invasive Surgery Centre Jesus Uson, Pharmacy unit. 10071, Spain

Introduction: Diabetes affects more than 340 million people all over the world. Expectations from the World Health Organization suggest that the number of deaths will double by 20301. Over 60 million people live with diabetes in Europe. The cost of diabetes complications accounts for 5-10% of total healthcare spending in several European countries. Despite recent advances in diagnosis and treatment of diabetes, its complications still represent a major public health challenge; In the field of diabetes, the pig has gained considerable interest as a preclinical model because of their metabolic, biochemical and physiological responses closely resemble those observed in humans 2,3,

Objective: The aim of this study is to provide data from a detailed protocol for the induction of diabetes and dyslipidemia in Gottingen minipigs, to contribute in the development of a safe and stable diabetes model mimicking as faithfully as possible the real clinical situation.

Material and methods: The study protocol was approved by the Institutional Animal Care and Use Committee. Twentythree ten months old males Gottingen minipigs with an average weight of 17.44 ± 1.75 kg were used. Six animals were used as non treated controls while in the other seventeen the diabetes induction was carried out by Streptozotocin intravenous administration (150 mg/kg). During the first three days after administration an anti-inflammatory / analgesic treatment was established (buprenorphine patch [35 µg/h/72h] and flunixin meglumine [2 mg/kg/IM/12h]). After induction, if the fasting glucose exceeded 400 mg/dl, animals received a subcutaneous combination of short-acting and retard insulin (3IU). For establishing a diabetic dyslipidemia (hypertriglyceridemia and hypercholesterolemia) the maintenance feed was supplemented with animal fat, raising the saturated fat and cholesterol level of the diet, reaching a content of 15% and 1% of the daily ration respectively. To assess the establishment of the dyslipidemic diabetes status weekly monitoring of glucose, total cholesterol, triglycerides, HDL-cholesterol and LDL-cholesterol was conducted during 35 weeks. At the end of the study tissue samples from the kidney, liver and coronary artery were taken and immediately fixed with 4% formaldehyde for histological examination. The thoracic aorta were also removed for the later Sudan IV staining for fatty steaks. A Kolmogorov Smirnov test was used to assure that data were normally distributed. Analysis of variance (ANOVA) for repeated measures was used to study changes with time, followed by a Tukey´s test to examine deviation from control values in each group. Intergroup differences were analysed by 2-way ANOVA. For all analyses, values of P < 0.05 were considered significant.

Results: The diabetic induction protocol described produces an intense and irreversible diabetic state, 48h after STZ administration mean glycaemia showed a significant (p≤0.05) increase from fasting baseline, these levels remained then significantly elevated until the end of the study. An increase in cholesterol and triglycerides levels was obtained between the diabetic and control group. Conclusion: This protocol and results contribute to the refinement of animals use by providing specific useful information that may allow diabetes research groups to reduce initial studies to set up a reliable diabetic dyslipidemia model in Gottingen minipigs.

1.- W.H.O. Factsheet nº312, August 2011; Consulted 2-09-2011. Available on line at: http://www.who.int/mediacentre/factsheets/fs312/en/

2.- Larsen MO, Rolin B. Use of the Göttingen minipig as a model of diabetes, with special focus on type 1 diabetes research. ILAR J 2004; 45(3):303-13. Review.

3.- Larsen MO, Wilken M, Gotfredsen CF, Carr RD, Svendsen O, Rolin B. Mild streptozotocin diabetes in the Göttingen minipig. A novel model of moderate insulin deficiency and diabetes. Am J Physiol Endocrinol Metab 2002; 282(6):E1342-51