Search

Microvascular disorders are relevant in systemic sclerosis (SSc). Hyperviscosity, due to alterations of blood cells and plasma components, may play a role in the pathogenesis of microcirculatory disorders. An impaired availability of nitric oxide, related to polymorphisms in NOS3, the gene for endothelial cell nitric oxide synthase, might influence erythrocyte deformability. We undertook this study to investigate the hemorheologic profile in SSc and the role of NOS3 polymorphisms in modulating the hemorheologic status of SSc patients.We studied 113 consecutive SSc patients (75 with limited cutaneous SSc [lcSSc] and 38 with diffuse cutaneous SSc [dcSSc]) and 113 healthy controls. The hemorheologic profile was obtained by assessing whole blood viscosity (WBV; at shear rates of 0.512 and 94.5 seconds−1), plasma viscosity (PLV; at a shear rate of 94.5 seconds−1), and erythrocyte deformability index (DI). We determined NOS3 polymorphisms by molecular analysis.A marked alteration of hemorheologic parameters was found both in patients with lcSSc and in those with dcSSc compared with controls (P < 0.0001). In multivariate analysis, rheologic variables were significantly associated with the disease (for WBV at a shear rate of 94.5 seconds−1, odds ratio [OR] 5.4, 95% confidence interval [95% CI] 1.4–19.9, P = 0.01; for PLV, OR 2.8, 95% CI 1.2–6.5, P = 0.01; for DI, OR 3.9, 95% CI 1.4–10.8, P = 0.007), and NOS3 −786C and 894T alleles significantly affected the DI (for −786C allele, OR 2.3, 95% CI 1.01–5.4, P = 0.04; for 894T allele, OR 2.2, 95% CI 1.01–4.8, P = 0.04). The simultaneous presence of the −786C and 894T alleles represented a susceptibility factor for SSc (OR 2.8, 95% CI 1.4–5.7, P = 0.004).Our findings document an altered rheologic profile in SSc and demonstrate a relationship between this alteration and NOS3 polymorphisms, thus shedding light on a potential novel mechanism influencing the microcirculation in this disease. [ABSTRACT FROM AUTHOR]

Defective angiogenesis, resulting in tissue ischemia, is particularly prominent in the diffuse form of systemic sclerosis (SSc). The present study was undertaken to identify possible differences between normal and SSc microvascular endothelial cells (MVECs) in the expression of the cell‐associated urokinase‐type plasminogen activator (uPA)/uPA receptor (uPAR) system, which is critical in the angiogenic process. MVECs were isolated from the dermis of healthy individuals and from the dermis of patients with diffuse SSc. The uPA/uPAR system was examined at the protein and messenger RNA levels. Angiogenesis was assayed on Matrigel‐coated porous filters and plates to evaluate cell proliferation, invasion, and capillary morphogenesis. Cleavage of uPAR and the activity of matrix metalloproteinase 12 (MMP‐12) were evaluated by Western blotting. Compared with MVECs from healthy skin, MVECs from SSc patients showed higher expression of uPAR. However, in SSc MVECs, uPAR undergoes truncation between domain 1 and domain 2, as shown by flow cytometry, enzyme‐linked immunosorbent assay, and Western blotting, a cleavage that is known to impair uPAR functions. These properties of SSc MVECs were associated with poor spontaneous and uPA‐dependent invasion, proliferation, and capillary morphogenesis. The uPAR cleavage occurring in SSc MVECs was associated with overexpression of MMP‐12. SSc MVEC–conditioned medium impaired uPA‐dependent proliferation and invasion as well as capillary morphogenesis in normal MVECs in vitro. Both a general hydroxamate inhibitor of MMP activity and anti–MMP‐12 antibodies restored this SSc MVEC–induced impaired functioning. Overproduction of MMP‐12 by SSc MVECs accounts for the cleavage of uPAR and the impairment of angiogenesis in vitro and may contribute to reduced angiogenesis in SSc patients. [ABSTRACT FROM AUTHOR]

Postnatal angiogenesis relies on a proper response of endothelial cells to angiogenic stimuli. In systemic sclerosis (SSc), endothelial cells are unresponsive to angiogenic factors. Since circumstantial and experimental evidence points to tissue kallikreins as powerful effectors of the angiogenic response, we undertook this study to investigate the kallikrein pattern of normal and SSc endothelial cells in order to identify differences that can account for defective angiogenesis.Expression of 14 tissue kallikreins was studied by a microarray approach, by reverse transcription–polymerase chain reaction, and by Western blotting in endothelial cells isolated from the skin of clinically healthy subjects and SSc patients. Cell proliferation was quantified by direct cell counting. Invasion and capillary morphogenesis were evaluated in a Boyden chamber and in culture flasks layered with Matrigel. Cyclic nucleotide production was measured by enzyme immunoassay. MAP kinase and ERK activation were measured by Western blotting.Endothelial cells from SSc patients showed poor expression of kallikreins 9, 11, and 12 compared with endothelial cells from normal subjects. Antibodies against the relevant kallikreins on normal endothelial cells revealed that while kallikreins 9, 11, and 12 induced cell growth, only kallikrein 12 regulated invasion and capillary morphogenesis. Buffering of kallikrein 12 with antibodies resulted in the acquisition of an SSc‐like pattern by normal cells in in vitro angiogenesis. Reduction of cAMP and cGMP production and of ERK phosphorylation upon administration of antikallikrein antibodies revealed that the activity of kallikreins 9, 11, and 12 was mediated by kinins.Reduction of tissue kallikreins 9, 11, and 12 may be relevant to reduced angiogenesis in SSc patients. [ABSTRACT FROM AUTHOR]