Study of In Vivo Glucose Metabolism in High-fat Diet-fed Mice Using Oral Glucose Tolerance Test (OGTT) and Insulin Tolerance Test (ITT)

摘要

The current article describes the generation and metabolic characterization of high-fat diet-fed mice as a model of diet-induced insulin resistance and obesity. It further features detailed protocols to perform the oral glucose tolerance test and the insulin tolerance test, monitoring whole-body alterations of glucose metabolism in vivo.

摘要

Obesity represents the most important single risk factor in the pathogenesis of type 2 diabetes, a disease which is characterized by a resistance to insulin-stimulated glucose uptake and a gross decompensation of systemic glucose metabolism. Despite considerable progress in the understanding of glucose metabolism, the molecular mechanisms of its regulation in health and disease remain under-investigated, while novel approaches to prevent and treat diabetes are urgently needed. Diet derived glucose stimulates the pancreatic secretion of insulin, which serves as the principal regulator of cellular anabolic processes during the fed-state and thus balances blood glucose levels to maintain systemic energy status. Chronic overfeeding triggers meta-inflammation, which leads to alterations in peripheral insulin receptor-associated signaling and thus reduces the sensitivity to insulin-mediated glucose disposal. These events ultimately result in elevated fasting glucose and insulin levels as well as a reduction in glucose tolerance, which in turn serve as important indicators of insulin resistance. Here, we present a protocol for the generation and metabolic characterization of high-fat diet (HFD)-fed mice as a frequently used model of diet-induced insulin resistance. We illustrate in detail the oral glucose tolerance test (OGTT), which monitors the peripheral disposal of an orally administered glucose load and insulin secretion over time. Additionally, we present a protocol for the insulin tolerance test (ITT) to monitor whole-body insulin action. Together, these methods and their downstream applications represent powerful tools to characterize the general metabolic phenotype of mice as well as to specifically assess alterations in glucose metabolism. They may be especially useful in the broad research field of insulin resistance, diabetes and obesity to provide a better understanding of pathogenesis as well as to test the effects of therapeutic interventions.

引言

In the developed world, obesity and diabetes reached epidemic dimensions due to physical inactivity and the excess consumption of processed food, effects which are driven by rapid urbanization, industrialization as well as globalization. Although research on insulin resistance and it's co-morbidities, such as hyperlipidemia and atherosclerosis, has gained prominence during the last decades, the complex biological mechanisms which regulate metabolism in health and disease remain incompletely understood and there is still an urgent need for new treatment modalities to prevent and treat these diseases¹.

Insulin, and....

研究方案

All methods described here have been approved by the Animal Care and Use Committee of the Medical University of Vienna and conducted according to the Federation of European Laboratory Animal Science Associations (FELASA). Please note that all procedures described in this protocol should only be performed after institutional and governmental approval as well as by staff that are technically proficient.

1. HFD-fed mice

NOTE: Maintain all C57BL/6J mice on a 12-h light/dark cycle with free access to food and water.

1. At 6 weeks of age, place mice for 8-12 weeks on an HFD (40-60% fat calories) to induce....

结果

Figure 1 illustrates a schematic time table for metabolic phenotyping of mice on diets. At an age of approximately 6 weeks, mice should be placed on an HFD, while an LFD-group may serve as the control group. Importantly, body weight should be determined weekly to observe if there is an expected increase in body weight. Any kind of stress (e.g., noise or aggressive male behavior) can interfere with body weight gain and should be eliminated immediately.......

讨论

With the high prevalence of diabetes and associated diseases in the world's population, there is a strong requirement for research addressing the molecular mechanism, prevention, and treatment of disease¹⁹. The presented protocol describes well-established methods for the generation of HFD mice, a robust animal model used for metabolic research, as well as the conduction of the OGTT and ITT, which are potent tools for the assessment of whole-body metabolic alterations such as insulin resistanc.......

材料

Name	Company	Catalog Number	Comments
Mouse strain: C57BL/6J	The Jackson Laboratory	664	LFD/HFD
Accu Chek Performa - Glucometer	Roche	6870228	OGTT/ITT
Accu Chek Performa - Strips	Roche	6454038	OGTT/ITT
D-(+)-Glucose solution	Sigma-Aldrich	G8769	OGTT
Actrapid - Insulin	Novo Nordisk	417642	ITT
Reusable Feeding Needles	Fine Science Tools	#18061-22	OGTT; 22 gauge (-24 gauge for young mice)
Omnifix-Fine dosing syringes	Braun	9161406V	OGTT/ITT
Sterican Insulin needle (30G x 1/3"; ø 0.30 x 13 mm)	Braun	304000	ITT; lean mice
Sterican (G 27 x 3/4"; ø 0.40 x 20 mm)	Braun	4657705	ITT; mice on HFD
96 Well PCR Plates, non-skirted, flexible	Braintree Scientific, Inc.	SP0016	OGTT
Ultrasensitive Mouse Insulin ELISA kit	Crystam Chem	90080	OGTT
Rodent Diet with 60% kcal% fat	Research Diets Inc	D12492	mice on HFD
Rodent Diet with 10% kcal% fat.	Research Diets Inc	D12450B	mice on LFD
BRAND micro haematocrit capillary	Sigma-Aldrich	BR749321	OGTT/ITT
Vaseline - creme	Riviera	P1768677	OGTT/ITT