TSH-Mediated TNFα Production in Human Fibrocytes Is Inhibited by Teprotumumab, an IGF-1R Antagonist.

Hong Chen,Raymond S Douglas,Tünde Mester,Yi-Hsuan Wei,Shannon J C Shan,Neeraj Vij

doi:10.1371/journal.pone.0130322

Abstract

PurposeFibrocytes (FC) are bone marrow-derived progenitor cells that are more abundant and infiltrate the thyroid and orbit in Graves orbitopathy (GO). FCs express high levels of thyrotropin receptor (TSHR) and insulin-like growth factor-1 receptor (IGF-1R). These receptors are physically and functionally associated, but their role in GO pathogenesis is not fully delineated. Treatment of FCs with thyroid stimulating hormone (TSH) or M22 (activating antibody to TSHR) induces the production of numerous cytokines, including tumor necrosis factor α (TNFα). Teprotumumab (TMB) is a human monoclonal IGF-1R blocking antibody currently in clinical trial for GO and inhibits TSHR-mediated actions in FCs.AimTo characterize the molecular mechanisms underlying TSH-induced TNFα production by FCs, and the role of IGF-1R blockade by TMB.DesignFCs from healthy and GD patients were treated with combinations of TSH, M22, MG132 and AKTi (inhibitors of NF-κB and Akt, respectively), and TMB. TNFα protein production was measured by Luminex and flow cytometry. Messenger RNA expression was quantified by real time PCR.ResultsTreatment with TSH/M22 induced TNFα protein and mRNA production by FCs, both of which were reduced when FCs were pretreated with MG132 and AKTi (p<0.0001). TMB decreased TSH-induced TNFα protein production in circulating FCs from mean fluorescent index (MFI) value of 2.92 to 1.91, and mRNA expression in cultured FCs from 141- to 52-fold expression (p<0.0001). TMB also decreased M22-induced TNFα protein production from MFI of 1.67 to 1.12, and mRNA expression from 6- to 3-fold expression (p<0.0001).ConclusionTSH/M22 stimulates FC production of TNFα mRNA and protein. This process involves the transcription factor NF-κB and its regulator Akt. Blocking IGF-1R attenuates TSH/M22-induced TNFα production. This further delineates the interaction of TSHR and IGF1-R signaling pathways. By modulating the proinflammatory properties of FCs such as TNFα production, TMB may be a promising therapeutic agent for GO.

Highlights

Fibrocytes are bone marrow-derived progenitor cells of the monocyte lineage [1]
Treatment with thyroid stimulating hormone (TSH)/M22 induced tumor necrosis factor α (TNFα) protein and mRNA production by FCs, both of which were reduced when FCs were pretreated with MG132 and Akt inhibitor IV (AKTi) (p
Blocking insulin-like growth factor-1 receptor (IGF-1R) attenuates TSH/M22induced TNFα production. This further delineates the interaction of TSHR and IGF1-R signaling pathways

Summary

Introduction

Fibrocytes are bone marrow-derived progenitor cells of the monocyte lineage [1]. They normally constitute less than 1% of circulating leukocytes [1]. In conditions of inflammation and fibrosis, fibrocytes emerge from the bone marrow and can comprise up to 15% of circulating leukocytes [2,3,4]. Fibrocytes have a distinct phenotype as they express both leukocyte and fibroblast surface markers [5]. Fibrocytes have both the proinflammatory properties of leukocytes as well as tissue remodeling capabilities of fibroblasts, making them excellent mediators of inflammation. Accumulating evidence suggests that they play an important role in the pathogenesis of Graves disease (GD) and Graves orbitopathy (GO)

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