Differential effects of Th1 versus Th2 cytokines in combination with hypoxia on HIFs and angiogenesis in RAReport as inadecuate

Differential effects of Th1 versus Th2 cytokines in combination with hypoxia on HIFs and angiogenesis in RA - Download this document for free, or read online. Document in PDF available to download.

Arthritis Research and Therapy

, 14:R180

First Online: 06 August 2012Received: 08 March 2012Revised: 27 June 2012Accepted: 06 August 2012


IntroductionHypoxia and T-helper cell 1 Th1 cytokine-driven inflammation are key features of rheumatoid arthritis RA and contribute to disease pathogenesis by promoting angiogenesis. The objective of our study was to characterise the angiogenic gene signature of RA fibroblast-like synoviocytes FLS in response to hypoxia, as well as Th1 and T-helper cell 2 Th2 cytokines, and in particular to dissect out effects of combined hypoxia and cytokines on hypoxia inducible transcription factors HIFs and angiogenesis.

MethodsHuman angiogenesis PCR arrays were used to screen cDNA from RA FLS exposed to hypoxia 1% oxygen or dimethyloxalylglycine, which stabilises HIFs. The involvement of HIF isoforms in generating the angiogenic signature of RA FLS stimulated with hypoxia and-or cytokines was investigated using a DNA-binding assay and RNA interference. The angiogenic potential of conditioned media from hypoxia-treated and-or cytokine-treated RA FLS was measured using an in vitro endothelial-based assay.

ResultsExpression of 12 angiogenic genes was significantly altered in RA FLS exposed to hypoxia, and seven of these were changed by dimethyloxalylglycine, including ephrin A3 EFNA3, vascular endothelial growth factor VEGF, adipokines angiopoietin-like ANGPTL-4 and leptin. These four proangiogenic genes were dependent on HIF-1 in hypoxia to various degrees: EFNA3 >ANGPTL-4 >VEGF >leptin. The Th1 cytokines TNFα and IL-1β induced HIF-1 but not HIF-2 transcription as well as activity, and this effect was additive with hypoxia. In contrast, Th2 cytokines had no effect on HIFs. IL-1β synergised with hypoxia to upregulate EFNA3 and VEGF in a HIF-1-dependent fashion but, despite strongly inducing HIF-1, TNFα suppressed adipokine expression and had minimal effect on EFNA3. Supernatants from RA FLS subjected to hypoxia and TNFα induced fewer endothelial tubules than those from FLS subjected to TNFα or hypoxia alone, despite high VEGF protein levels. The Th2 cytokine IL-4 strongly induced ANGPTL-4 and angiogenesis by normoxic FLS and synergised with hypoxia to induce further proangiogenic activity.

ConclusionThe present work demonstrates that Th1 cytokines in combination with hypoxia are not sufficient to induce angiogenic activity by RA FLS despite HIF-1 activation and VEGF production. In contrast, Th2 cytokines induce angiogenic activity in normoxia and hypoxia, despite their inability to activate HIFs, highlighting the complex relationships between hypoxia, angiogenesis and inflammation in RA.


CIAcollagen-induced arthritis

DMEMDulbecco-s modified Eagle-s medium


DMSOdimethyl sulfoxide

EFNA3ephrin A3

ELISAenzyme-linked immunosorbent assay

FLSfibroblast-like synoviocytes

HIFhypoxia-inducible factor

HMEChuman microvascular endothelial cell

HREhypoxia response element

HSFhuman skin fibroblast



mAbmonoclonal antibody

PBSphosphate-buffered saline

PCRpolymerase chain reaction

RArheumatoid arthritis

siRNAsmall interfering RNA

ThT-helper cell

TNFtumour necrosis factor

UTRuntranslated region

VEGFvascular endothelial growth factor.

Electronic supplementary materialThe online version of this article doi:10.1186-ar3934 contains supplementary material, which is available to authorized users.

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Author: Helene Larsen - Barbara Muz - Tak L Khong - Marc Feldmann - Ewa M Paleolog

Source: https://link.springer.com/

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