ETA2022 Poster Presentations Graves’ Disease 2 and Orbitopathy (8 abstracts)
Macrophage-orbital fibroblasts interaction in context of hypoxic signaling for inflammatory processes during graves’ orbitopathy
Gina-Eva Görtz 1 , Anja Eckstein 2 , Christoph Jesenek 3 , Mareike Horstmann 4 , Svenja Philipp 5 , Kirsten Bruderek 6 , Michael Oeverhaus 7 , Anke Daser 6 , Nikolaos Bechrakis 8 , Sven Brandau 9 & Utta Berchner-Pfannschmidt 10
1University Hospital Essen, Molecular Ophthalmology, Essen, Germany; 2Univ.-Augenklinik, Department of Ophthalmology, University Eye Hospital Essen, Essen, Germany; 3University Hospital Essen, University of Duisburg-Essen, Molecular Ophthalmology, Department of Ophthalmology, Essen, Germany; 4Univerity Hospital Essen, Molecular Ophthalmology, Essen, Germany; 5Department of Ophthalmology, Molecular Ophthalmology, Essen, Germany; 6Department of Otorhinolaryngology, University of Duisburg-Essen, Essen, Germany; 7Universitätsklinikum Essen, Klinik für Augenheilkunde, Essen, Germany; 8Klinik für Augenheilkunde, Universitätsklinikum Essen, Department of Ophthalmology, Molecular Ophthalmology Group, University of Duisburg-Essen, Essen, Germany, Essen, Germany; 9Klinik für Hals-, Nasen-, Ohrenheilkunde, University Hospital Essen, Department of Otorhinolaryngology, Essen, Germany; 10Molekulare Ophthalmologie, Department of Ophthalmology, University Hospital Essen, Essen, Germany
Introduction: The inflammatory eye disease Graves’ orbitopathy (GO) is the main complication of Graves’ disease in patients. In previous studies we have shown that hypoxia and HIF-1 dependent pathways could play an important role in the pathogenic process of GO. Hypoxia is known to attract inflammatory cells and therefore maintains inflammation and recruitment of immune cells like macrophages (MQ). However, few is known about the specific contribution of MQ to the progression of orbitopathy. Therefore, we investigated the role and interaction of MQ and orbital fibroblasts (OF) in context of inflammation and hypoxia.
Methods & Results: We analyzed the expression levels of hypoxic marker HIF-1α, MQ marker CD68, proinflammatory cytokine TNFα and recruitment proteins CCL2, CCL5 and CCL20 in fat biopsies of control and GO patients by real-time PCR. We found that HIF-1α, CD68, TNFα, CCL2, CCL5 and CCL20 mRNA expression was increased in the fat tissue of GO patients. Next, we analyzed the cytokine profile of supernatants from fat biopsies with a multiplex ELISA. We could show an enhanced secretion of TNFα, CCL2 and CCL20 only under hypoxia while CCL5 was induced on protein level under normoxia as well as under hypoxia in GO tissue. An immunofluorescence stain of CD68 and TNFα was used to demonstrate the source of TNFα in the orbital tissue. The Immunofluorescence indicated that TNFα secretion occurs in conjunction with CD68 positive MQ. To further, investigate the inflammatory interaction of MQ and OF, we stimulated OF with TNFα or co-cultured them with M1-MQ from a THP-1 cell line under normoxic and hypoxic conditions. We found that OF expressed hypoxic marker HIF-1α, hypoxia target gene VEGF and immune marker ICAM-1 as well as chemokines CCL2, CCL5 and CCL20 most pronounced upon TNFα stimulation and hypoxia. M1-MQ enhanced the induction of HIF-1α and CCL2 in OF in addition to hypoxia alone, whereas proinflammatory inhibitors Etanercept and dexamethasone reduced this effect. Furthermore, we found that OF-macrophage co-culture enhanced adipogenic differentiation and adiponectin secretion under hypoxia. PX-478, a HIF-1α inhibitor, reduced the adipogenic differentiation of OF significantly.
Conclusion: In summary, our results show that hypoxia and macrophage-OF interactions have a cumulative effect. The findings indicate that the inflammatory milieu and hypoxic signaling in the orbit are characterized by TNFα positive macrophages, which interact with OF which results in constant inflammation and tissue remodeling. A combination of anti-inflammatory treatment and HIF-1α reduction could be an effective treatment option for GO.
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Endocrine Abstracts
ISSN 1470-3947 (print) | ISSN 1479-6848 (online)
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