Inhibition of Inducible Nitric Oxide Synthase Prevents IL-1β-Induced Mitochondrial Dysfunction in Human Chondrocytes.

Annett Eitner,Gunther O Hofmann,Arne Wilharm,Rebecca Raab,Hans-Georg Schaible,Sylvia Müller,Thomas Kamradt,Christian König

doi:10.3390/ijms22052477

Abstract

Interleukin (IL)-1β is an important pro-inflammatory cytokine in the progression of osteoarthritis (OA), which impairs mitochondrial function and induces the production of nitric oxide (NO) in chondrocytes. The aim was to investigate if blockade of NO production prevents IL-1β-induced mitochondrial dysfunction in chondrocytes and whether cAMP and AMP-activated protein kinase (AMPK) affects NO production and mitochondrial function. Isolated human OA chondrocytes were stimulated with IL-1β in combination with/without forskolin, L-NIL, AMPK activator or inhibitor. The release of NO, IL-6, PGE2, MMP3, and the expression of iNOS were measured by ELISA or Western blot. Parameters of mitochondrial respiration were measured using a seahorse analyzer. IL-1β significantly induced NO release and mitochondrial dysfunction. Inhibition of iNOS by L-NIL prevented IL-1β-induced NO release and mitochondrial dysfunction but not IL-1β-induced release of IL-6, PGE2, and MMP3. Enhancement of cAMP by forskolin reduced IL-1β-induced NO release and prevented IL-1β-induced mitochondrial impairment. Activation of AMPK increased IL-1β-induced NO production and the negative impact of IL-1β on mitochondrial respiration, whereas inhibition of AMPK had the opposite effects. NO is critically involved in the IL-1β-induced impairment of mitochondrial respiration in human OA chondrocytes. Increased intracellular cAMP or inhibition of AMPK prevented both IL-1β-induced NO release and mitochondrial dysfunction.

Highlights

Introduction published maps and institutional affilPro-inflammatory cytokines contribute significantly to the initiation and progression of osteoarthritis (OA) via up-regulation of catabolic processes [1,2]
The current study aimed to evaluate whether increased nitric oxide (NO) production is responsible for IL-1β-induced mitochondrial dysfunction in human OA chondrocytes obtained from knee joints during arthroplasty
The IL-1β concentration 0.1 ng/mL evoked a high release of NO, which only slightly increased at higher IL-β concentrations (Figure 1a)

Summary

Introduction

Pro-inflammatory cytokines contribute significantly to the initiation and progression of osteoarthritis (OA) via up-regulation of catabolic processes [1,2]. The impairment of the mitochondrial function of chondrocytes is thought to be an important factor in the pathophysiology of OA [3,4,5]. IL-1β induces upregulation of inducible NO synthase (iNOS) and the production of nitric oxide (NO). This mediator regulates numerous putative pathogenic processes in the cartilage (see below), and may alter mitochondrial respiration and ATP production in chondrocytes [3]. Whether IL-1β-induced NO production is causally responsible for mitochondrial impairment and whether inhibition of iNOS can prevent IL-1β-induced mitochondrial dysfunction has not yet been reported

Methods

Results

Discussion

Conclusion