Complement Regulation in Human Tenocytes under the Influence of Anaphylatoxin C5a.

Sandeep Silawal,Benjamin Kohl,Gundula Schulze-Tanzil,Jingjian Shi

doi:10.3390/ijms22063105

Abstract

A central part of the complement system, the anaphylatoxin C5a was investigated in this study to learn its effects on tenocytes in respect to understanding the potential expression of other crucial complement factors and pro-inflammatory mediators involved in tendinopathy. Human hamstring tendon-derived tenocytes were treated with recombinant C5a protein in concentrations of 25 ng/mL and 100 ng/mL for 0.5 h (early phase), 4 h (intermediate phase), and 24 h (late phase). Tenocytes survival was assessed after 24 h stimulation by live-dead assay. The gene expression of complement-related factors C5aR, the complement regulatory proteins (CRPs) CD46, CD55, CD59, and of the pro-inflammatory cytokines tumor necrosis factor (TNF)-α and interleukin (IL)-6 was monitored using qPCR. Tenocytes were immunolabeled for C5aR and CD55 proteins. TNFα production was monitored by ELISA. Tenocyte survival was not impaired through C5a stimulation. Interestingly, the gene expression of C5aR and that of the CRPs CD46 and CD59 was significantly reduced in the intermediate and late phase, and that of TNFα only in an early phase, compared to the control group. ELISA analysis indicated a concomitant not significant trend of impaired TNFα protein synthesis at 4 h. However, there was also an early significant induction of CD55 and CD59 mediated by 25 ng/mL anaphylatoxin C5a. Hence, exposure of tenocytes to C5a obviously evokes a time and concentration-dependent response in their expression of complement and pro-inflammatory factors. C5a, released in damaged tendons, might directly contribute to tenocyte activation and thereby be involved in tendon healing and tendinopathy.

Highlights

Tendinopathy has been defined as a chronic low grade inflammatory and degenerative musculoskeletal disease [1,2]
A low increase in pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α is associated with the pathological tendons and the affected tenocytes [4]
The aim of the study is to understand the effect of C5a, known as an inflammatory mediator, on human tenocytes with a particular focus on complement regulation and cytokines TNFα and IL-6

Summary

Introduction

Tendinopathy has been defined as a chronic low grade inflammatory and degenerative musculoskeletal disease [1,2]. The collagen tears resulting from so-called micro traumata after repetitive mechanical overloading have been a long known classical explanation for the pain and functional impairment in tendinopathy [3]. The inflammatory concept in the pathogenesis of tendinopathy has regained a wider focus [2]. A low increase in pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α is associated with the pathological tendons and the affected tenocytes [4]. An increased presence of inflammatory cells such as macrophages, mast cells, and T-cells in pathological tendons was emphasized in comparison to healthy tendons

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