Ncb5or Deficiency Increases Fatty Acid Catabolism and Oxidative Stress

Ming Xu,Wenfang Wang,Jennifer R Frontera,Melanie C Neely,Jianghua Lu,Daniel Aires,Fong-Fu Hsu,John Turk,Russell H Swerdlow,Susan E Carlson,Hao Zhu

doi:10.1074/jbc.m110.196543

Abstract

The endoplasmic reticulum-associated NADH cytochrome b(5) oxidoreductase (Ncb5or) is widely distributed in animal tissues. Ncb5or(-/-) mice develop diabetes at age 7 weeks and have increased susceptibility to the diabetogenic oxidant streptozotocin. Ncb5or deficiency also results in lipoatrophy and increased hepatocyte sensitivity to cytotoxic effects of saturated fatty acids. Here we investigate the mechanisms of these phenomena in prediabetic Ncb5or(-/-) mice and find that, despite increased rates of fatty acid uptake and synthesis and higher stearoyl-CoA desaturase (SCD) expression, Ncb5or(-/-) liver accumulates less triacylglycerol (TAG) than wild type (WT). Increased fatty acid catabolism and oxidative stress are evident in Ncb5or(-/-) hepatocytes and reflect increased mitochondrial content, peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) expression, fatty acid oxidation rates, oxidative stress response gene expression, and oxidized glutathione content. Ncb5or(-/-) hepatocytes readily incorporate exogenous fatty acids into TAG but accumulate more free fatty acids (FFA) and have greater palmitate-induced oxidative stress responses and cell death than WT, all of which are alleviated by co-incubation with oleate via TAG channeling. A high fat diet rich in palmitate and oleate stimulates both lipogenesis and fatty acid catabolism in Ncb5or(-/-) liver, resulting in TAG levels similar to WT but increased intracellular FFA accumulation. Hepatic SCD-specific activity is lower in Ncb5or(-/-) than in WT mice, although Ncb5or(-/-) liver has a greater increase in Scd1 mRNA and protein levels. Together, these findings suggest that increased FFA accumulation and catabolism and oxidative stress are major consequences of Ncb5or deficiency in liver.

Highlights

In vitro reconstitution experiments show that the NADH cytochrome b5 oxidoreductase (Ncb5or) homolog microsomal cytochrome b5 (Cyb5A) and its cognate reductase Cyb5R3 provide electrons for the stearoyl-CoA desaturase (SCD) reaction [8]
Defects in early neonatal development lead to increased mitochondrial biogenesis, fatty acid catabolism, and oxidative stress, and these phenotypic abnormalities are exacerbated by exposure to excess saturated fatty acid (SFA) and attenuated by monounsaturated fatty acid (MUFA)
It is notable that hepatic TAG stores in Ncb5orϪ/Ϫ mice exhibited fatty acid desaturation indices that were significantly lower than those of wild type (WT) mice as early as age 3 weeks when fed with chow

Summary

Introduction

In vitro reconstitution experiments show that the Ncb5or homolog microsomal cytochrome b5 (Cyb5A) and its cognate reductase Cyb5R3 provide electrons for the SCD reaction [8]. The other two (D371Y and L424M) occur at conserved residues in the b5R domain and are found as somatic mutations in 7% of human breast cancers [15] It is not yet known how these Ncb5or mutations affect its structure and function or contribute to disease pathogenesis. Ncb5orϪ/Ϫ hepatocytes incorporate fatty acids readily into TAG but accumulate more FFA and exhibit reduced content of MUFA relative to SFA at a given level of SCD expression, suggesting impaired SCD function. These defects in Ncb5orϪ/Ϫ cells appear to be responsible for their enhanced susceptibility to the cytotoxic effects of oxidants and palmitic acid

Methods

Results

Conclusion