IL-23 and IL-12, two structurally related heterodimeric cytokines sharing a common subunit, diver... more IL-23 and IL-12, two structurally related heterodimeric cytokines sharing a common subunit, divergently promote Th cell development and expansion. Both cytokines have been implicated in the pathogenesis of thyroid-associated ophthalmopathy (TAO), an autoimmune component of Graves disease. In TAO, CD34+ fibrocytes, putatively derived from bone marrow, can be identified in the orbit. There they masquerade as CD34+ orbital fibroblasts (OF) (CD34+ OF) and cohabitate with CD34− OF in a mixed fibroblast population (GD-OF). Slit2, a neural axon repellent, is expressed and released by CD34− OF and dampens the inflammatory phenotype of fibrocytes and CD34+ OF. In this study we report that thyrotropin (TSH) and the pathogenic, GD-specific monoclonal autoantibody, M22, robustly induce IL-23 in human fibrocytes; however, IL-12 expression is essentially undetectable in these cells under basal conditions or following TSH-stimulation. In contrast, IL-12 is considerably more inducible in GD-OF, cel...
Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2018
Human CD34 fibrocytes, circulating monocyte lineage progenitor cells, have recently been implicat... more Human CD34 fibrocytes, circulating monocyte lineage progenitor cells, have recently been implicated in thyroid-associated ophthalmopathy (TAO), the ocular manifestation of Graves' disease (GD). Fibrocytes express constitutive MHC class II (MHC-2) and, surprisingly, thyroglobulin (Tg) and functional thyrotropin (TSH) receptor (TSHR). Underlying expression of these thyroid proteins is the autoimmune regulator protein (AIRE). Fibrocytes respond robustly to TSH and thyroid-stimulating Igs by generating extremely high levels of inflammatory cytokines, such as IL-6. In TAO, they appear to infiltrate the orbit, where they transition to CD34 orbital fibroblasts (OF). There, they coexist with CD34 OF as a mixed fibroblast population (GD-OF). In contrast to fibrocytes, GD-OF express vanishingly low levels of MHC-2, Tg, TSHR, and AIRE. Further, the amplitude of IL-6 induction by TSH in GD-OF is substantially lower. The molecular basis for this divergence between fibrocytes and CD34 OF rema...
Orbital fibroblasts from patients with Graves' disease (GD-OF) express many different cytokin... more Orbital fibroblasts from patients with Graves' disease (GD-OF) express many different cytokines when treated with bovine thyrotropin (bTSH). The present study aimed to determine why TNF-α cannot be induced by bTSH in GD-OF. Fibrocytes and GD-OFs were cultivated from donors who were patients in a busy academic medical center practice. Real-time PCR, Western blot analysis, reporter gene assays, cell transfections, mRNA stability assays, ELISA, and flow cytometry were performed. We found that bTSH induces TNF-α dramatically in fibrocytes but is undetectable in GD-OF. The induction in fibrocytes is a consequence of increased TNF-α gene promoter activity and is independent of ongoing protein synthesis. It could be attenuated by dexamethasone and the IGF-1 receptor inhibiting antibody, teprotumumab. When separated into pure CD34+ OF and CD34- OF subsets, TNF-α mRNA became highly inducible by bTSH in CD34+ OF but remained undetectable in CD34- OF. Conditioned medium from CD34- OF inhib...
A relationship between the actions of TSH and IGF-1 was first recognized several decades ago. The... more A relationship between the actions of TSH and IGF-1 was first recognized several decades ago. The close physical and functional associations between their respective receptors (TSHR and IGF-1R) has been described more recently in thyroid epithelium and human orbital fibroblasts as has the noncanonical behavior of IGF-1R. Here we report studies conducted in lung fibroblasts from female wild-type C57/B6 (TSHR+/+) mice and their littermates in which TSHR has been knocked out (TSHR−/−). Flow cytometric analysis revealed that cell surface IGF-1R levels are substantially lower in TSHR−/− fibroblasts compared with TSHR+/+ fibroblasts. Confocal immunofluorescence microscopy revealed similar divergence with regard to both cytoplasmic and nuclear IGF-1R. Western blot analysis demonstrated both intact IGF-1R and receptor fragments in both cellular compartments. In contrast, IGF-1R mRNA levels were similar in fibroblasts from mice without and with intact TSHR expression. IGF-1 treatment of TSHR...
CD34(+) fibrocytes are bone marrow-derived monocyte progenitor cells that traffic to sites of tis... more 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 (TAO) 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 GD and healthy individuals. PTX-3 mRNA levels were determined by real-time PCR, protein by ELISA and Western blot, and PTX-3 gene promoter activity was assessed with reporter assays. P...
The Journal of Clinical Endocrinology & Metabolism, 2014
Context:Factors common to multiple autoimmune diseases have been sought vigorously. Graves' d... more Context:Factors common to multiple autoimmune diseases have been sought vigorously. Graves' disease (GD) and type 1 diabetes mellitus (T1DM) involve end-organ remodeling. Fibrocytes participate in inflammatory diseases and were recently shown to express thyroid-specific proteins such as the thyrotropin receptor and thyroglobulin.Objective:The objective of the study was to determine whether a broader repertoire of autoantigen expression, such as proteins associated with T1DM, can be ascribed to fibrocytes.Design, Setting, and Participants:Fibrocytes and fibroblasts were collected and analyzed from healthy individuals and those with autoimmune diseases in an academic clinical practice.Main Outcome Measures:Real-time PCR, Western blot analysis, gene promoter analysis, cell transfections, and flow cytometric cell sorting were performed.Results:Islet cell antigen ICA512 (IA-2) and islet cell autoantigen of 69 kDa (ICA69), two islet-specific proteins implicated in T1DM, are expressed ...
Ophthalmic plastic and reconstructive surgery, Jan 5, 2014
To investigate the phenotypic and functional characteristics of peripheral and tissue-infiltratin... more To investigate the phenotypic and functional characteristics of peripheral and tissue-infiltrating stem cells called fibrocytes in patients with idiopathic orbital inflammation (IOI). Seven patients with IOI were studied. In the 3 patients requiring orbital biopsy, fibrocytes were identified in orbital tissue from patients with IOI compared with healthy controls using immunohistochemistry. Fibrocytes from the peripheral blood of all 7 patients and controls were quantified and phenotyped by flow cytometry and immunofluorescence for expression of CD34, alpha smooth muscle actin, CD40, and collagen 1. Quantitation of CD40-mediated interleukin-6 (IL-6) production was measured using enzyme-linked immunosorbent assay. Orbital biopsy specimens from patients with IOI demonstrate tissue infiltration by fibrocytes (n = 3). Fibrocytes are present in the peripheral blood of IOI patients (n = 7) but are scarce in healthy donors (n = 19). Fibrocytes from IOI patients express substantial levels of...
Our goals are to construct a simple genetic clock that will stably oscillate in Escherichia coli ... more Our goals are to construct a simple genetic clock that will stably oscillate in Escherichia coli and to identify the design principles and parameters responsible for oscillations. We previously described a simple genetic circuit of linked activator and repressor operons that produced damped oscillations. Here, we altered the repression of the activator operon and identified an oscillator that produces improved oscillations over our initial system. We also explored mathematical models of the oscillator. Toy models were used to investigate the behaviors that may be obtained from our clock circuitry. Depending on parameters, the circuitry produced a wide array of oscillatory systems, including sinusoidal and relaxation oscillators. We also attempted to explicitly model all known interactions that affect the oscillator, producing a 32-dimensional ODE model. This model can produce results similar to those obtained in experiments, and we have begun attempts to fit experimental data to the model.
IL-23 and IL-12, two structurally related heterodimeric cytokines sharing a common subunit, diver... more IL-23 and IL-12, two structurally related heterodimeric cytokines sharing a common subunit, divergently promote Th cell development and expansion. Both cytokines have been implicated in the pathogenesis of thyroid-associated ophthalmopathy (TAO), an autoimmune component of Graves disease. In TAO, CD34+ fibrocytes, putatively derived from bone marrow, can be identified in the orbit. There they masquerade as CD34+ orbital fibroblasts (OF) (CD34+ OF) and cohabitate with CD34− OF in a mixed fibroblast population (GD-OF). Slit2, a neural axon repellent, is expressed and released by CD34− OF and dampens the inflammatory phenotype of fibrocytes and CD34+ OF. In this study we report that thyrotropin (TSH) and the pathogenic, GD-specific monoclonal autoantibody, M22, robustly induce IL-23 in human fibrocytes; however, IL-12 expression is essentially undetectable in these cells under basal conditions or following TSH-stimulation. In contrast, IL-12 is considerably more inducible in GD-OF, cel...
Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2018
Human CD34 fibrocytes, circulating monocyte lineage progenitor cells, have recently been implicat... more Human CD34 fibrocytes, circulating monocyte lineage progenitor cells, have recently been implicated in thyroid-associated ophthalmopathy (TAO), the ocular manifestation of Graves' disease (GD). Fibrocytes express constitutive MHC class II (MHC-2) and, surprisingly, thyroglobulin (Tg) and functional thyrotropin (TSH) receptor (TSHR). Underlying expression of these thyroid proteins is the autoimmune regulator protein (AIRE). Fibrocytes respond robustly to TSH and thyroid-stimulating Igs by generating extremely high levels of inflammatory cytokines, such as IL-6. In TAO, they appear to infiltrate the orbit, where they transition to CD34 orbital fibroblasts (OF). There, they coexist with CD34 OF as a mixed fibroblast population (GD-OF). In contrast to fibrocytes, GD-OF express vanishingly low levels of MHC-2, Tg, TSHR, and AIRE. Further, the amplitude of IL-6 induction by TSH in GD-OF is substantially lower. The molecular basis for this divergence between fibrocytes and CD34 OF rema...
Orbital fibroblasts from patients with Graves' disease (GD-OF) express many different cytokin... more Orbital fibroblasts from patients with Graves' disease (GD-OF) express many different cytokines when treated with bovine thyrotropin (bTSH). The present study aimed to determine why TNF-α cannot be induced by bTSH in GD-OF. Fibrocytes and GD-OFs were cultivated from donors who were patients in a busy academic medical center practice. Real-time PCR, Western blot analysis, reporter gene assays, cell transfections, mRNA stability assays, ELISA, and flow cytometry were performed. We found that bTSH induces TNF-α dramatically in fibrocytes but is undetectable in GD-OF. The induction in fibrocytes is a consequence of increased TNF-α gene promoter activity and is independent of ongoing protein synthesis. It could be attenuated by dexamethasone and the IGF-1 receptor inhibiting antibody, teprotumumab. When separated into pure CD34+ OF and CD34- OF subsets, TNF-α mRNA became highly inducible by bTSH in CD34+ OF but remained undetectable in CD34- OF. Conditioned medium from CD34- OF inhib...
A relationship between the actions of TSH and IGF-1 was first recognized several decades ago. The... more A relationship between the actions of TSH and IGF-1 was first recognized several decades ago. The close physical and functional associations between their respective receptors (TSHR and IGF-1R) has been described more recently in thyroid epithelium and human orbital fibroblasts as has the noncanonical behavior of IGF-1R. Here we report studies conducted in lung fibroblasts from female wild-type C57/B6 (TSHR+/+) mice and their littermates in which TSHR has been knocked out (TSHR−/−). Flow cytometric analysis revealed that cell surface IGF-1R levels are substantially lower in TSHR−/− fibroblasts compared with TSHR+/+ fibroblasts. Confocal immunofluorescence microscopy revealed similar divergence with regard to both cytoplasmic and nuclear IGF-1R. Western blot analysis demonstrated both intact IGF-1R and receptor fragments in both cellular compartments. In contrast, IGF-1R mRNA levels were similar in fibroblasts from mice without and with intact TSHR expression. IGF-1 treatment of TSHR...
CD34(+) fibrocytes are bone marrow-derived monocyte progenitor cells that traffic to sites of tis... more 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 (TAO) 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 GD and healthy individuals. PTX-3 mRNA levels were determined by real-time PCR, protein by ELISA and Western blot, and PTX-3 gene promoter activity was assessed with reporter assays. P...
The Journal of Clinical Endocrinology & Metabolism, 2014
Context:Factors common to multiple autoimmune diseases have been sought vigorously. Graves' d... more Context:Factors common to multiple autoimmune diseases have been sought vigorously. Graves' disease (GD) and type 1 diabetes mellitus (T1DM) involve end-organ remodeling. Fibrocytes participate in inflammatory diseases and were recently shown to express thyroid-specific proteins such as the thyrotropin receptor and thyroglobulin.Objective:The objective of the study was to determine whether a broader repertoire of autoantigen expression, such as proteins associated with T1DM, can be ascribed to fibrocytes.Design, Setting, and Participants:Fibrocytes and fibroblasts were collected and analyzed from healthy individuals and those with autoimmune diseases in an academic clinical practice.Main Outcome Measures:Real-time PCR, Western blot analysis, gene promoter analysis, cell transfections, and flow cytometric cell sorting were performed.Results:Islet cell antigen ICA512 (IA-2) and islet cell autoantigen of 69 kDa (ICA69), two islet-specific proteins implicated in T1DM, are expressed ...
Ophthalmic plastic and reconstructive surgery, Jan 5, 2014
To investigate the phenotypic and functional characteristics of peripheral and tissue-infiltratin... more To investigate the phenotypic and functional characteristics of peripheral and tissue-infiltrating stem cells called fibrocytes in patients with idiopathic orbital inflammation (IOI). Seven patients with IOI were studied. In the 3 patients requiring orbital biopsy, fibrocytes were identified in orbital tissue from patients with IOI compared with healthy controls using immunohistochemistry. Fibrocytes from the peripheral blood of all 7 patients and controls were quantified and phenotyped by flow cytometry and immunofluorescence for expression of CD34, alpha smooth muscle actin, CD40, and collagen 1. Quantitation of CD40-mediated interleukin-6 (IL-6) production was measured using enzyme-linked immunosorbent assay. Orbital biopsy specimens from patients with IOI demonstrate tissue infiltration by fibrocytes (n = 3). Fibrocytes are present in the peripheral blood of IOI patients (n = 7) but are scarce in healthy donors (n = 19). Fibrocytes from IOI patients express substantial levels of...
Our goals are to construct a simple genetic clock that will stably oscillate in Escherichia coli ... more Our goals are to construct a simple genetic clock that will stably oscillate in Escherichia coli and to identify the design principles and parameters responsible for oscillations. We previously described a simple genetic circuit of linked activator and repressor operons that produced damped oscillations. Here, we altered the repression of the activator operon and identified an oscillator that produces improved oscillations over our initial system. We also explored mathematical models of the oscillator. Toy models were used to investigate the behaviors that may be obtained from our clock circuitry. Depending on parameters, the circuitry produced a wide array of oscillatory systems, including sinusoidal and relaxation oscillators. We also attempted to explicitly model all known interactions that affect the oscillator, producing a 32-dimensional ODE model. This model can produce results similar to those obtained in experiments, and we have begun attempts to fit experimental data to the model.
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Papers by Stephen Atkins