Abstract-Interactions between integrins and growth factor receptors play a critical role in the development and healing of the vasculature. This study mapped two binding domains on fibronectin (FN) that modulate the activity of the angiogenic factor, vascular endothelial growth factor (VEGF T he growth, repair, and regeneration of blood vessels are complex processes that involve coordinated regulation of endothelial cell proliferation, migration, and differentiation. 1 One of the most important vascular morphogens is vascular endothelial growth factor (VEGF). VEGF has been shown to play a major role in vasculogenesis and angiogenesis by gene deletion studies. 2,3 Targeted disruption of the VEGF receptor Flk-1 (VEGFR-2) in mice resulted in failure of blood-island formation and endothelial differentiation. 4 Flk-1 is also the first endothelial receptor tyrosine kinase to be expressed in the hemangioblast. 5 We and others recently demonstrated that the hematopoietic progenitor cell CD34 ϩ can differentiate into endothelial cells, and that VEGF was one of the critical factors promoting this differentiation. 6,7 Interactions between cells and their extracellular matrix (ECM) play an integral role in blood vessel development. The earliest ECM protein expressed in the embryo during vasculogenesis is fibronectin (FN). 8 Gene deletion studies have demonstrated that both FN and its major integrin receptor, ␣ 5 ␤ 1 , are critical for vasculogenesis and angiogenesis in the developing embryo. 9 -11 Collectively, these observations suggest important roles for FN and its integrin receptor, ␣ 5 ␤ 1 , in vasculogenesis and angiogenesis.In this study, we show that novel VEGF binding domains of FN are required for promoting the specific association of the FN receptor integrin ␣ 5 ␤ 1 with the VEGF receptor, Flk-1. This association between VEGF and FN is required for the full effects of VEGF-induced endothelial cell migration and proliferation. This study demonstrates that FN can profoundly affect VEGF biological activity and consequently the behavior of endothelial cells through their coordinated effects on Flk-1 and ␣ 5 ␤ 1 . Materials and Methods Solid-Phase VEGF Binding AssayECM proteins and FN peptides were purchased from Sigma and Gibco and were purified further by gel filtration and ion exchange chromatography. Microtiter plates were coated with the appropriate ECM proteins (50 L; 10 g/mL) in 100 mmol/L bicarbonate buffer (pH 9) overnight at 4°C.

We retrospectively analyzed the results of 75 living-related pediatric renal transplants performed at our center between January 1986 and December 1999. The major causes of end-stage renal disease (ESRD) were glomerulonephritis (26%) and nephrolithiasis (16%), while the etiology was unknown in 50%. The mean age of the recipients was 12 yr (range 6-17 yr) and that of the donors was 39 yr (range 20-65 yr). The majority (73%) of donors were parents. Eighty five per cent of donors were one-haplotype matched and the rest identical. Immunosuppression was based on a triple drug regimen. Thirty per cent of recipients were rapid metabolizers of cyclosporin A (CsA) (area under the curve [AUC]: < 6,000 ng/mL/h), while 16% were slow metabolizers (AUC: > 8,000 ng/mL/h). Forty three (57%) children encountered 59 rejection episodes, the majority of which (59%) were recorded in the first month post-transplant. Seventy-four per cent of the rejection episodes were steroid sensitive and the rest, except two, were resolved by therapy with antithymocyte globulin (ATG) or orthoclone thymocyte 3 (OKT3). After a mean follow-up of 37 months, 17 (22%) grafts had chronic rejection and 76% of these recipients had previously experienced acute rejection episodes. The overall infection rate was high, necessitating two hospital admissions/patient/year. The majority (53%) of the infections were bacterial. Urinary tract infections (UTIs) were seen in 17 (23%) recipients. Twelve of these had ESRD as a result of stone disease and eight grafts were lost because of UTIs. Eight per cent of recipients developed tuberculosis (TB), and extra-pulmonary lesions were seen in 50%. Surgical complications were encountered in eight patients. Free medication to all recipients and parental support ensured a compliance rate of 93%. Baseline growth deficit was seen in children of the two groups studied (the 6-12 yr and 13-17 yr age-groups), with Z-scores of - 2.39 and - 2.12, respectively. No growth catch-up was observed at 12 and 24 months in either group. Post-donation complications were seen most commonly in donors > 50 yr of age and included: proteinuria (> 300 mg/24 h, four patients), hypertension (three patients), and diabetes (one patient). Twenty-four grafts were lost, 54% as a result of immunological and the rest as a result of non-immunological causes, and 17 recipients died during the follow-up period. Infections were the main cause of patient and graft loss. Overall 1- and 5-yr graft and patient survival rates were 88% and 65%, and 90% and 75%, respectively.

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