Background: Hemophilic joint disease secondary to recurrent hemarthroses is one of the most disabling and costly complications of hemophilia. Prior to widespread use of prophylactic factor concentrates, children in the United States with severe hemophilia A and B (X-linked recessive disorders with \<1% factor VIII/IX (FVIII/FIX) activity) experienced an average of 30-35 hemarthroses per year reported for FVIII deficiency (3,4 ). Clinical and sub clinical hemarthroses during childhood results in synovitis (hypertrophied synovium characterized by villous formation, markedly increased vascularity and chronic inflammatory cells (5) eventually leading to pannus formation and destructive arthritis (6, 7). At this time, synovial bleeding may be related not only to clotting factor deficiency but also to pre-existing vascular damage and inflammation, which is difficult to control clinically. Use of factor concentrate prophylaxis results in hemophilia patients experiencing fewer joint bleeds, less rapid deterioration of joint function and fewer days lost from school or work. However, it may be complicated by the unpredictable development of an inhibitor (a high-affinity, polyclonal, function-neutralizing antibody directed against FVIII/FIX), with an incidence of about 25% and 6% respectively (8) . These individuals have limited treatment options or are treated with products that are less efficacious in treating the joint bleed along with potential deleterious side effects such as thrombogenecity( 9) thus favoring joint disease development.

Primary prophylaxis (infusion of FVIII concentrates (25 - 40 u/kg thrice weekly or FIX concentrates - 80-100u/kg twice a week starting at age 1-2 yrs) before the onset of joint bleeds is used in Sweden since the 1960s to keep the trough level of factor VIII/FIX \> 1%, converting a severe hemophilia patient (FVIII/FIX activity \<1%) into a milder form (FVIII/FIX \>5%) (10 ) . This strategy is expensive (\~ $77,760/year for a 20 kg child based on the use of 3000 to 6,000 u/kg/yr with recombinant factor VIII), and may require the use of venous access devices in young children, which is complicated by severe infections, bleeding and thrombosis (11) . Secondary prophylaxis on the other hand, which involves the use of FVIII/FIX concentrates after "target joints" (at least four bleeds occurring into a single joint in the previous six months) have been identified may limit bleeding and subsequent joint damage. However, progression of existing joint disease continues and it is unclear whether secondary prophylaxis can actually prevent joint deterioration (12) . Furthermore, studies comparing primary and secondary prophylaxis in relation to cost-effectiveness and long-term joint morbidity suggest that primary prophylaxis improved long-term joint outcome but was twice as expensive (13 ) . For these reasons, the optimum age, subject population, and timing of prophylaxis is highly debated. Finally, therapeutic options for individuals who fail or cannot use prophylaxis (inhibitor patients) or refuse prophylaxis include isotopic (IS) and surgical synovectomy. Isotopic synovectomy involves intraarticular injection of 32P- colloid with the intent of scarring off the synovium leading to a subsequent reduction in hemarthroses (14) , both procedures being recommended for patients with chronic synovitis and ongoing hemarthroses. Again, the timing of these strategies in relation to the onset of synovitis remain unclear. Hence, if prophylaxis is not started early (before the occurrence of joint bleeds) and subject population is not optimized, a strategy to detect and monitor synovitis and joint arthropathy is urgently needed so that prophylaxis and synovectomy can be timed based on evidence to reap optimum benefits. Furthermore, in hemophilic children who complain of joint pains, clinical examination sometimes, may not clearly define whether the symptoms are related to a joint bleed, synovitis or surrounding soft tissue bleeds. Studies in animals suggest that cartilage damage can occur concurrently with synovial damage (15) contributing to joint arthropathy. Therefore, it seems that determination of both synovial and cartilage changes would be imperative and may help to guide prophylaxis.

Traditionally, hemophilic arthropathy has been diagnosed by clinical examination and plain radiographs of joints, which together tend to underestimate the extent of joint destruction (16) . Magnetic Resonance Imaging (MRI) can estimate the degree of bony damage associated with hemophilic joint disease (14 , 17 -19). The investigators have previously shown the utility of ultrasound-power Doppler sonography( USG-PDS) in detecting synovitis associated with hemophilic joint disease when compared to MRI ( 1) . The need for sedation in children and high costs ($ 2500 for MRI with sedation versus $ 600 for USG- PDS - no sedation at this institution) override the utility of this tool when repeated studies may be required for closer surveillance of joint disease progression. Visualization of cartilage is clinically relevant because benefits of both prophylaxis and synovectomy are realized only if there is minimal damage to cartilage. Furthermore, there is scattered evidence to suggest that isotopic synovectomy in a joint affected by bony arthropathy can lead to progression of the arthropathy leading to crippling arthritis.

The pathogenesis of HJD is not well defined. Neoangiogenesis is a critical factor in processes, such as tumor growth and inflammatory arthritis (20). Increased vascularity and neoangiogenesis have been implicated in the progression of musculoskeletal disorders and tumor growth. Vascular endothelial growth factor (VEGF), the principal signaling molecule in angiogenesis, can be induced by hypoxia and certain cytokines through interaction with its receptors, VEGFR1 and VEGFR2 (21 -23). The synovitic pannus in other joint diseases that share histologic similarities with hemophilic joint disease (HJD) have enhanced oxygen demand and show evidence of de novo blood vessel formation, including endothelialization of the synovium( 24) . Further, VEGF expression in the serum has been correlated with disease activity in rheumatoid arthritis ( 25) . Endothelialization may occur as a result of mature endothelial cell migration or through the recruitment of bone marrow (BM)-derived endothelial progenitor cells (EPCs) and hematopoietic progenitor cells (HPCs) from the peripheral circulation (26) . Importantly, proliferating synovium can secrete chemocytokines, such as VEGF, that might promote recruitment of endothelial cells (ECs) to sites of active angiogenesis ( 25) . Co-localization of hypoxia-inducible factor- 1 (HIF-1 α) which is a transcription factor involved in the induction of VEGF and produced in response to hypoxia within the joint and VEGF emphasizes the role of hypoxia in the up-regulation of angiogenesis in rheumatoid joint diseases ( 27).

The investigators have previously observed a 4-fold elevation in proangiogenic factors (vascular endothelial growth factor-A \[VEGF-A\], stromal cell-derived factor-1, and matrix metalloprotease-9) and proangiogenic macrophage/monocyte cells (VEGF+/CD68+ and VEGFR1+/CD11b +) in the synovium and peripheral blood of hemophilic joint disease (HJD) subjects along with significantly increased numbers of VEGFR2+/AC133 + endothelial progenitor cells and CD34+/ VEGFR1+ hematopoietic progenitor cells. Sera from HJD subjects induced an angiogenic response in endothelial cells that was abrogated by blocking VEGF, whereas peripheral blood mononuclear cells from HJD subjects stimulated synovial cell proliferation, which was blocked by a humanized anti-VEGF antibody (bevacizumab). Human synovial cells, when incubated with HJD sera, could elicit up-regulation of HIF-1α mRNA with HIF-1α expression in the synovium of HJD subjects, implicating hypoxia in the neoangiogenesis process. The investigators results provided evidence of local and systemic angiogenic response in hemophilic subjects with recurrent hemarthroses suggesting a potential to develop surrogate biologic markers to identify the onset and progression of hemophilic synovitis( 2). Therefore, evidence of increased synovial vascularity on USG-PDS and elevated angiogenic markers suggestive of increased vascularity in hemophilic joint disease subjects provides a compelling opportunity to develop surrogate biological markers for hemophilic joint disease. This would also further aid in tailoring strategies such as prophylaxis and synovectomy in an individual patient.