Introduction

Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease in Western countries, and it has been considered the liver expression of the metabolic syndrome¹. However, the cause and treatments of NAFLD are still controversial. Nitric oxide (NO) and its derivatives play important roles in the physiology and pathophysiology of the vascular system and liver². NO availability not only affects the vascular system but also induces changes in mitochondrial bioenergetics that result in increased accumulation of fatty acids in the liver. eNOS^-/- animals develop systemic alterations similar to type 2 diabetes and hypertension³. In addition, they also develop progressive NAFLD when fed with high-fat diet (HFD)⁴. In this work we investigated whether the total amount of intraperitoneal fat and liver fat-fraction in eNOS^-/- mice fed with HFD is ameliorated by pharmacological treatments for type 2 diabetes (Metformin) and hypertension (Losartan). Additionally, we studied if the administration of an inhibitor of endothelial NO synthesis (L-NAME)⁵ in wild type mice induces the same phenotype as observed in eNOS^-/- mice thereby establishing whether nitric oxide plays an important role in the development of NAFLD.

Methods

Study design is summarized in Fig.1. In vivo MRI: Whole body 3-point Dixon images were obtained using a 3T clinical MR scanner (Achieva, Philips Healthcare, Best, NL) and a 47mm single-loop surface coil. The acquisition parameters include TR/ TE1/ TE2/ TE3 = 15/ 2.3/ 3.5/ 4.7 ms; flip angle 10º; in plane resolution= 0.17 x 0.17 mm; slice thickness= 0.5 mm. Animal treatments: All treatments were administered in water as follow: L-NAME (0.5 mg/mL), Metformin (2 mg/mL), Losartan (0.3 mg/mL).

Results

eNOS^-/- mice fed with HFD have significantly more intraperitoneal fat and liver fat-fraction than WT and eNOS^-/- mice fed with normal chow (Fig. 2, Fig. 3A, Fig. 4A). WT mice that received L-NAME have similar phenotype as in eNOS^-/- mice (Fig. 3A and Fig. 3B). Lower amounts of intraperitoneal fat and liver fat-fraction were observed in eNOS^-/- mice fed with HFD and treated with Losartan and Metformin, with a slightly synergic effect when both treatments were administered together (Fig. 3B and Fig. 4B). Our results suggest that the treatment of hypertension and type 2 diabetes induced by the lack of eNOS, normalize the fat liver content.

Conclusion

Using in vivo MRI we demonstrated that liver fatty acid accumulation is accelerated in eNOS^-/- mice fed with HFD, which could be replicated in WT mice fed with L-NAME. Intraperitoneal and liver fat accumulation could be partially normalized after the administration of losartan and metformin. Our results show the relevance of noninvasive monitoring of liver fat fraction and body composition to evaluate the response and adherence to pharmacological treatments.

Acknowledgements

The British Heart Foundation (RG/12/1/29262). CONICYT-PIA, Anillo ACT1416. PuenteUC P1712/2017.

References

¹Cohen JC, Science 2011; ²Iwakiri Y, Trends Pharmacol Sci 2015; ³Takahashi T, J Diabetes Res 2014; ⁴Sheldon RD, J Appl Physiol 2014; ⁵Suda O, Circulation 2002