Patients with Hemophilia Have Increased Numbers and Pericyte Coverage of Synovial Vessels.
Authors: Zetterberg Eva , Morfini Massimo , Wallensten Richard , Palmblad Jan , Margareta Holmström 
Published: October 2019

Abstract
Abstract 3053
Poster Board II-1029
In patients with hemophilia, repeated joint bleedings leads to synovitis and bleeding arthropathy is the major cause of morbidity in these patients.
The synovitis is characterized by a highly vascular synovial membrane with prominent proliferation of synovial fibroblasts and infiltration by inflammatory cells. This chronic inflammation, as well as a direct toxic effect of blood on chondrocytes ultimately leads to cartilage and bone destruction and a crippling arthropathy. It has recently been shown that. Inflammatory cells isolated from hemophilic joints synthesize pro angiogenic factors (matrix metalloproteas-9, basic fibroblast growth factor and cyclooxygenas-2). Angiogensis in the adult is a complex process involving breakdown of connective tissue, proliferation and migration of endothelial cells but also recruitment of supporting cells, pericytes, important for vessel maturity. The aim of this study was to determine whether repeated joint bleedings in hemophilia patients induce a pro angiogenic reaction [increased micro vascular density (MVD) and expression of vascular endothelial growth factor (VEGF)] in the hemophilic joint and to study if also pericytes are involved in the process. After informed consent, synovial biopsies were collected from patients with severe hemophilia (n=5) when undergoing knee surgery on clinical grounds. As control, synovial biopsies from one patient undergoing diagnostic arthroscopy were used. Biopsies were snap frozen in liquid nitrogen and sectioned by cryotome. After fixation, sections were double stained for CD34 (for detection of endothelial cells) and SMA-a (for detection of pericytes) by immuno fluorescence.
Sections were also stained for VEGF by immuno histochemistry. As shown in Table 1 patients with hemophilia had a very high MVD and pericyte coverage as compared to control samples. We also measured levels of VEGF by ELISA in plasma from 23 patients and 4 controls, but in all samples the concentration was below detection level. However, in synovial biopsies from patients with hemophilia, VEGF was clearly expressed by the endothelial cells as well as mononuclear cells present in the section.
Table 1 Angiogenesis parameters in patients with hemophilia Synovial biopsies plasma MVD (vessels/HPF) Pericyte coverage (%) VEGF+ (cells/HPF) VEGF (pg/mL Hemophilia patients 17±8 91±0.1 22±19 <9 Control 2 40 0 0
Based on these preliminary data we suggest that hemophilic arthropaty is characterized by increased angiogenesis and mobilization of pericytes. Although no increase of VEGF was detected systemically it was shown to be expressed locally, within the inflamed joint. Anti-angiogenic drugs are currently successfully used in different types of cancers and a specificVEGF inhibitor for topical administration has successfully been developed for proliferative diabetic retinopathy. The results presented in this study could be part of a rationale to design studies using these drugs also in hemophilic arthropaty, a condition currently lacking specific treatment.

Disclosures
No relevant conflicts of interest to declare.