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Pentraxin-3 Is a TSH-Inducible Protein in Human Fibrocytes and Orbital Fibroblasts.

ABSTRACT: CD34(+) fibrocytes are bone marrow-derived monocyte progenitor cells that traffic to sites of tissue injury and repair. They putatively infiltrate the orbit in thyroid-associated ophthalmopathy where they appear to transition into CD34(+) orbital fibroblasts (OFs) that interact with residential CD34(-) fibroblasts. A unique phenotypic attribute of fibrocytes and CD34(+) OFs is their expression of the functional thyrotropin receptor (TSHR) and other "thyroid-specific" proteins. When activated through TSHR, fibrocytes express a number of cytokines and other inflammatory genes. Here we sought to determine whether pentraxin-3 (PTX-3), an acute-phase protein involved in inflammation and autoimmunity, might be induced by TSH in fibrocytes and OFs. These cells were collected from patients with Graves disease and healthy individuals. PTX-3 mRNA levels were determined by real-time PCR, protein was determined by ELISA and Western blot, and PTX-3 gene promoter activity was assessed with reporter assays. PTX-3 expression was induced by TSH in both cell types, regardless of the health status of the donor and was a consequence of increased steady-state PTX-3 mRNA levels. M22, a TSHR-activating monoclonal antibody, also induced PTX-3. The induction could be attenuated by dexamethasone and by IGF-I receptor-blocking antibodies, teprotumumab and 1H7. TSH effects were mediated through phosphatidylinositol 3-kinase/AKT, mammalian target of rapamycin/p70(s6k), Janus tyrosine kinase 2 pathways, and enhanced PTX-3 mRNA stability. These findings indicate that PTX-3 is a TSH target gene, the expression of which can be induced in fibrocytes and OFs. They suggest that PTX-3 might represent a previously unidentified nexus between the thyroid axis and the mechanisms involved in tissue remodeling.

SUBMITTER: Wang H

PROVIDER: S-EPMC4606754 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

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Pentraxin-3 Is a TSH-Inducible Protein in Human Fibrocytes and Orbital Fibroblasts.

Wang Hao H Atkins Stephen J SJ Fernando Roshini R Wei Rui-Li RL Smith Terry J TJ

Endocrinology 20150819 11

CD34(+) fibrocytes are bone marrow-derived monocyte progenitor cells that traffic to sites of tissue injury and repair. They putatively infiltrate the orbit in thyroid-associated ophthalmopathy where they appear to transition into CD34(+) orbital fibroblasts (OFs) that interact with residential CD34(-) fibroblasts. A unique phenotypic attribute of fibrocytes and CD34(+) OFs is their expression of the functional thyrotropin receptor (TSHR) and other "thyroid-specific" proteins. When activated thr ...[more]

PMID: 26287404

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Project description: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 ? (TNF?). 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 TNF? 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-?B and Akt, respectively), and TMB. TNF? protein production was measured by Luminex and flow cytometry. Messenger RNA expression was quantified by real time PCR.ResultsTreatment with TSH/M22 induced TNF? 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 TNF? 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 TNF? 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 TNF? mRNA and protein. This process involves the transcription factor NF-?B and its regulator Akt. Blocking IGF-1R attenuates TSH/M22-induced TNF? production. This further delineates the interaction of TSHR and IGF1-R signaling pathways. By modulating the proinflammatory properties of FCs such as TNF? production, TMB may be a promising therapeutic agent for GO.

Project description:ContextThe molecular basis for anatomically dispersed clinical manifestations in Graves' disease (GD) eludes our understanding. Bone marrow-derived, pluripotent fibrocytes represent a subset of peripheral blood mononuclear cells and infiltrate the orbital and thyroid tissues in GD. These cells may be involved in the pathogenesis of thyroid-associated ophthalmopathy (TAO).ObjectiveThe objective of the study was to quantify fibrocyte display of functional cell surface TSH receptor (TSHR), identify the profile of chemokines they express after TSHR activation, and determine whether circulating TSHR(+) peripheral blood fibrocytes are more frequent in situ in patients with TAO.Design/setting/participantsUsing a newly developed technique, fibrocytes were directly identified in peripheral blood from 31 patients with TAO and 19 healthy subjects receiving care at a multidisciplinary academic center.Main outcome measuresThe frequency in situ of fibrocytes (collagen 1(+), CD45(+), CD34(+), CD14(+), CD86(+) peripheral blood mononuclear cells) was assessed by multiparameter flow cytometry and correlated to clinical disease activity and smoking status. Levels of TSHR-displaying fibrocytes and their response to TSH and TSHR-activating antibody, M22, were measured by flow cytometry, Luminex, and real-time PCR.ResultsThe levels of TSHR expression by fibrocytes are substantially higher than those found in orbital fibroblasts. Moreover, the frequency of TSHR(+) fibrocytes in patients with TAO was greater than that in healthy subjects in situ. Their abundance is not influenced by disease activity or smoking history. These cells produce high levels of several cytokines and chemokines including IL-8, regulated upon activation, normal T cell expressed and secreted, and monocyte chemoattractant protein-1 when treated with TSH or M22. TSH induces IL-8 production at the pretranslational level. This induced cytokine can be detected in intact fibrocytes ex vivo.ConclusionsFrequency of circulating TSHR(+) fibrocytes is markedly increased in patients with TAO, and they express proinflammatory chemokines in response to TSH. Because they infiltrate both orbit and thyroid in GD, they may represent the link between systemic immunoreactivity and organ-specific autoimmunity.

Dataset Information

Pentraxin-3 Is a TSH-Inducible Protein in Human Fibrocytes and Orbital Fibroblasts.

Publications

Pentraxin-3 Is a TSH-Inducible Protein in Human Fibrocytes and Orbital Fibroblasts.

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