Báo cáo khoa học: Muramyl-dipeptide-induced mitochondrial proton leak in macrophages is associated

Muramyl-dipeptide-induced mitochondrial proton leak in

macrophages is associated with upregulation of

uncoupling protein 2 and the production of reactive

oxygen and reactive nitrogen species

Takla G. El-Khoury, Georges M. Bahr and Karim S. Echtay

Faculty of Medicine and Medical Sciences and Faculty of Sciences, University of Balamand, Tripoli, Lebanon

Keywords

mitochondria; muramylpeptides; nitric oxide;

respiratory control ratio; superoxide anion;

UCP2

Correspondence

K. S. Echtay, Faculty of Medicine and

Medical Sciences, University of Balamand,

PO Box 100, Tripoli, Lebanon

Fax: +961 6 930279

Tel: +961 3 714125

E-mail: karim.echtay@balamand.edu.lb

(Received 5 May 2011, revised 13 June

2011, accepted 28 June 2011)

doi:10.1111/j.1742-4658.2011.08226.x

The synthetic immunomodulator muramyl dipeptide (MDP) has been

shown to induce, in vivo, mitochondrial proton leak. In the present work,

we extended these findings to the cellular level and confirmed the effects of

MDP in vitro on murine macrophages. The macrophage system was then

used to analyse the mechanism of the MDP-induced mitochondrial proton

leak. Our results demonstrate that the cellular levels of superoxide anion

and nitric oxide were significantly elevated in response to MDP. Moreover,

isolated mitochondria from cells treated with MDP presented a significant

decrease in respiratory control ratio, an effect that was absent following

treatment with a non-toxic analogue such as murabutide. Stimulation of

cells with MDP, but not with murabutide, rapidly upregulates the expres￾sion of the mitochondrial protein uncoupling protein 2 (UCP2), and pre￾treatment with vitamin E attenuates upregulation of UCP2. These findings

suggest that the MDP-induced reactive species upregulate UCP2 expression

in order to counteract the effects of MDP on mitochondrial respiratory

efficiency.

Introduction

Uncoupling proteins (UCPs) are members of the anion

carrier family molecules present in the inner mitochon￾drial membrane. Mammals express five UCP homo￾logues, UCP1–UCP5. UCP2 and UCP3 have 59% and

57% identity, respectively, with UCP1, and 73% iden￾tity with each other [1], whereas UCP4 and UCP5 (also

referred to as brain mitochondrial carrier protein 1,

BMCP1) have much lower sequence identity with

UCP1 [2,3]. UCP1 is the best characterized of these

proteins, mediating non-shivering thermogenesis in

brown adipose tissue by catalysing proton leak acti￾vated by long-chain fatty acids and inhibited by purine

nucleotides [4]. UCP2 is widely expressed in many tis￾sues with high levels detected in the spleen, thymus,

pancreatic b-cells, heart, lung, white and brown adi￾pose tissue, stomach, testis and macrophages, whereas

low levels have been reported in the brain, kidney,

liver and muscle [5]. UCP3 is expressed predominantly

in skeletal muscles and brown adipose tissues [6,7], at

hundred-fold lower concentration than UCP1 in brown

adipose tissue [8]. UCP4 and UCP5 are only present in

the brain [2,3]. Due to their homology to UCP1 and

their distribution in several mammalian tissues, it has

been initially postulated that these proteins can regu￾late mitochondrial oxidative phosphorylation through

uncoupling activity. However, the physiological function

of UCPs other than UCP1 has remained controversial.

Suggested functions include mild uncoupling, adaptive

Abbreviations

FCCP, fluorocarbonyl cyanide phenylhydrazone; LPS, lipopolysaccharide; MB, murabutide; MDP, muramyl dipeptide; PI, propidium iodide;

RCR, respiratory control ratio; ROS, reactive oxygen species; RNS, reactive nitrogen species; UCP, uncoupling protein.

3054 FEBS Journal 278 (2011) 3054–3064 ª 2011 The Authors Journal compilation ª 2011 FEBS