Background: VEGF-A is a key regulator of rheumatoid (RA) and osteoarthritis (OA). During articular inflammation in OA and RA there is increased synovial angiogenesis and upregulation of angiogenic growth factors such as VEGF-A. VEGF-A comprises two splice variant families, VEGF-Axxxa and VEGF-Axxxb (xxx represents the number of amino acids, from 121 to 206), resulting from alternative splice site selection in exon 8. This site selection is controlled by Serine/Arginine Rich Splicing Factor Kinase 1 (SRPK1), which phophorylates Serine/Arginine Rich Splicing Factor 1 (SRSF1), inducing it to translocate to the nucleus. In most normal tissues, VEGF-Axxxb isoforms predominate, with anti-nociceptive and anti-angiogenic functions. I contrast, in pathological conditions such as inflammation and solid tumours, VEGF-Axxxa isoforms predominate, with pro-nociceptive and pro-angiogenic functions. VEGF-A has been proposed as a therapeutic target in RA and OA. To date, there are no published data on the functionally distinct VEGF-A splice variants in either RA or OA.
Objectives: To determine the patterns of, and relationships between, VEGF-A, SRPK1, and SRSF1 expression and activation and synovial inflammation in human RA and OA.
Methods: The study was approved by the Nottingham Research Ethics Committee 1 (05/Q2403/24) and Derby Research Ethics Committee 1 (11/H0405/2). Tissues were selected from age- and sex-matched cases in the University of Nottingham joint tissue repository. Post-mortem (PM) samples of healthy knee synovium (n=14, no history of arthritis or knee pain in the 12 months prior to death, no significant articular or synovial pathology) and arthroplasty-derived synovium samples from OA (n=35) or RA (n=14) patients were compared. OA samples were selected to represent the variety of inflammation levels, from low to high grade (0-3, Haywood et al., 2003). 8um thick sections were stained for SRSF1, SRPK1, total VEGF-A and VEGFxxxb via immunohistochemistry. Expression was estimated as fractional area, relative staining intensity (VEGF-Axxxb), and SRSF1 activation quantified by the degree of nuclear localisation. Statistical analyses were performed using Kruskal-Wallis followed by Dunn’s tests and Spearman’s rank correlations.
Results: SRPK1 expression was similar across all conditions. SRSF1 showed significantly higher expression in the OA tissue compared to PM (H(2)= 11.29, p=0.002; OA median=0.2, IQR(0.15, 0.28); PM median=0.09, IQR(0.07, 0.16)), and significantly higher nuclear localisation (indicating activation) in RA vs. OA, and in both RA and OA vs PM (H(2)=37.65, p<0.0001 RA cf. PM; p=0,007 OA cf. PM; RA median=89, IQR(83, 93); OA median=36.1, IQR(29, 42); PM median=19.8, IQR(14,21)). Nuclear SRSF1 was significantly correlated with inflammation score (r= 0.52, p<0.05). Total VEGF-A expression was significantly increased in RA compared to PM and OA (H(2)=23.3, p<0.001 RA cf. PM; RA median=0.4, IQR(0.37,0.59); PM median=0.18, IQR(0.15,0.2)) and was also correlated with the severity of inflammation (r=0.47 p<0.05). VEGF-Axxxb showed no changed in expression in OA or RA, although VEGF-Axxxb staining intensity was significant higher in RA samples, compared to controls (H(2)=7.2 p=0.02; RA median=2.3(1, 4); PM median=0.9 (0.7, 1.4)).
Conclusion: Increased levels of SRSF1 activation, and the association of total VEGF-A expression with inflammation score, support the hypothesis that there is activation of alternative splicing in inflamed synovium in RA and OA. Targeting this pathway could be a novel therapeutic strategy in OA & RA.
References: HAYWOOD L., MCWILLIAMS D. F., PEARSON C. I., GILL S. E., GANESAN A., WILSON D. & WALSH D. A. 2003. Inflammation and angiogenesis in osteoarthritis. Arthritis Rheum, 48, 2173-7.
Disclosure of Interests: Dimitrios Amanitis: None declared, Seyed Shahtaheri: None declared, Daniel Daniel McWilliams: None declared, Nicholas Beazley-Long: None declared, David Walsh Grant/research support from: 2016: Investigator-led grant from Pfizer Ltd (ICRP) on Pain Phenotypes in RA; non-personal financial disclosure (payment to University)., Consultant of: DAW has undertaken paid consultancy to Pfizer Ltd, Eli Lilly and Company and GSK Consumer Healthcare., Paid instructor for: 2019: Consultancy to Love Productions; consultancy on programme design, contribution to programme content on self-management of chronic pain (payments to University)
2019: Consultancy to AbbVie Ltd; 13.06.19; presentation on RA pain at EULAR, Madrid, and webinar (payments to University).
2019: Consultancy to Eli Lilly and Company Ltd. 06.06.19 Centre for Collaborative Neuroscience, Windlesham, Surrey, UK (payment to University).
2019: Consultancy to Pfizer (payment to University).
2018: Consultancy to Pfizer. 07.12.18. USA. 1 day. Tanezumab (payment to University).
2018: Consultancy to Pfizer. 23.11.18. Manchester UK. 1 day. Tanezumab (payment to University).
2018: Consultancy to Pfizer. 1.11.18. Skype. 4h. Tanezumab (payment to University).
2018: Consultancy to GlaxoSmithKline Plc. 1 day. Pain in RA and anti-GM-CSF (payment to University).
2018: Consultancy to Pfizer Ltd; Presentation at OARSI; non-personal financial disclosure (payment to University)
2018: Consultancy to Pfizer Ltd; Patient preference study; non-personal financial disclosure (payment to University)
2017: Consultancy to Pfizer Ltd; personal financial disclosure
2017: Consultancy to Pfizer Ltd through Nottingham University; non-personal financial disclosure (payment to University).
2015: Consultancy to GSK Consumer Healthcare; personal financial disclosure., Speakers bureau: 2019: Irish Society of Rheumatology: speaker fees (personal pecuniary), Lucy Donaldson Shareholder of: LFD is a co-inventor on patents protecting alternative RNA splicing control and VEGF-A splice variants for therapeutic application in a number of different conditions. LFD is also a founder equity holder in, and consultant to, Exonate Ltd, and Emenda Therapeutics Ltd. Both companies have with a focus on development of alternative RNA splicing control for therapeutic application in a number of different conditions, including ophthalmology (www.exonate.com), analgesia and arthritis (www.emendatherapeutics.com)., Consultant of:
LFD is a co-inventor on patents protecting alternative RNA splicing control and VEGF-A splice variants for therapeutic application in a number of different conditions. LFD is also a founder equity holder in, and consultant to, Exonate Ltd, and Emenda Therapeutics Ltd. Both companies have with a focus on development of alternative RNA splicing control for therapeutic application in a number of different conditions, including ophthalmology (www.exonate.com), analgesia and arthritis (www.emendatherapeutics.com).
Statistics from Altmetric.com
If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.