Exploring the Effects of a High-Fat Diet and Gut Microbiomes on RIP140 Knockdown Mice Resistant to Diet-Induced Type II Diabetes

Jacob Jensen, Yi-Wei Lin, Li-Na Wei Department of Pharmacology, University of Minnesota Medical School, University of Minnesota

Introduction and Background

Figure 1:

Metabolic diseases, such as obesity and diet-induced type II diabetes, are a prevalent concern in Western society. These diseases are primarily linked to a high fat diet, which is composed of trans and saturated fats, high fructose corn syrup, and carbohydrates1. Unfortunately, because of the high glucose conditions associated with diet-induced type II diabetes, the inflammation of the arterial walls is accelerated2. Typically, the recruitment and activation of adipose tissue macrophages (ATM) play a crucial role in obesity-induced inflammation and insulin resistance. In addition, gut microbiota may play a significant role in metabolic disorders. Changing the gut microbiota could lead to the elimination of metabolic syndrome3. This study explored the effects of feeding macrophage-specific RIP140 knockdown mice (MφRIP140KD) a high-fat diet and different types of mice feces. Typically, wild type mice are more prone to diet-induced diabetes while MφRIP140KD show less severe symptoms4.

Figure 1: (A) Oil red O staining of aortic sinus (left panel) and quantitative relative lesion size (middle panel) after 16 weeks of western diet feeding. Average lesion size of ApoE null mice was set as 1. Relative macrophagic RIP140 mRNA expression level is shown on the right panel. (B) en face oil red O staining of aorta and quantification.

Hypothesis 1.  A high-fat diet will increase the expression of RIP140, ATM, inflammation, and insulin

resistance. 2.  When wild type mice are fed the feces of knockdown mice, their gut microbiota’s will alter,

making them less susceptible to diet induced diabetes. 3.  When knockdown mice are fed the feces of wild type mice, their gut microbiota’s will alter,

making them more susceptible to diet induced diabetes4.

Materials and Methods

Results 1 Results 3

Conclusion

•  C57/BL6 mice were used as wild type and MφRIP140KD transgenic mice were generated •  Experimentations commenced on 8 week old mice •  Mice from hypothesis one were fed diets for 12 weeks. Mice from hypotheses two and three were

fed diets for 7 weeks. Afterwards, they were fed their own feces for 7 weeks •  Normal diet contained 18% calories from diet and no detectable amounts of cholesterol. High fat

diet contained 60% calories from fat and 345 mg cholesterol/mg •  Glucose tolerance test (GTT) and insulin tolerance test (ITT) were performed every three weeks

after over night fasting •  Oxygen consumption (vO2) was measured via indirect calorimetry for 2 days •  Fluorescein isothiocyanate labeled dextran was used to measure the intestine’s permeability •  Mice were euthanized to collect adipose tissue. All mice were euthanized by exposure to a legal

dosage of CO2 •  Adipose tissue was fixed in frozen liquid nitrogen for protein and mRNA analysis. Macrophage

harvested from collected tissues was plated at 1x107 cells/plate in the reagent DMEM for 8 days •  Following 8 days, a western blot of the cultured adipose tissue was performed

Figure 2: •  Microbiome profiles of Wild Type (WT) and MφRIP140KD

animals under Normal Diet and High-Fat Diet conditions •  Diet and RIP140 expression in mice change gut microbiome

profiles

Results 2

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Figure 3: •  (A) Glucose tolerance test of WT and KD mice after microbiome

transportation and 8 weeks of high-fat diet feeding •  (B) Insulin tolerance test of WT and KD mice after microbiome

transportation and 8 weeks of high-fat diet feeding •  Microbiome transplantation from MφRIP140KD mice improves

diet-induced insulin resistance

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•  Intestinal permeability analysis of mice after microbiome transplantation after 8 weeks of high-fat diet feeding. WT/WT: WT as donors and WT as recipients, WT/KD: WT as donors and KD as recipients, KD/WT: KD as donors and WT as recipients, KD/KD: KD as donors and KD as recipients.

•  Microbiome transplantations can possibly create affordable clinical treatments for obesity and diet induced type II diabetes

•  RIP140 expression alters gut microbiome in mice fed with a normal or high-fat diet •  Microbiome transplantation from MφRIP140KD mice to WT mice prevents diet-

induced metabolic disorders •  Microbiome transplantations can lead to creating affordable clinical treatments for

obesity and diet-induced type II diabetes

References 1)  Hu, F. B. Globalization of Diabetes The role of diet, lifestyle, and genes. Diabetes

Care 34, 1249-1257 (2011). 2)  Chait, A. & Bornfeldt, K. E. Diabetes and atherosclerosis: is there a role for

hyperglycemia? J. Lipid Res. 50, S335-S339 (2009). 3)  Liu, P.-S. et al. Reducing RIP140 expression in macophage alters ATM infiltration,

facilitates white adipose tissue browning, and prevents high-fat diet-induced insulin resistance. Diabetes 63, 4021-4031 (2014).

4)  Ho, P.-C., Tsui, Y.-C., Feng, X., Greaves, D. R. & Wei, L.-N. NF-κB-mediated degradation of the coactivator RIP140 regulates inflammatory responses and contributes to endotoxin tolerance. Nat. Immunol. 13, 379–386 (2012)

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Funding •  Sponsored by the Undergraduate Research Opportunities Project

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