Biomedical subjects

Babette Weksler

Publications and source records attributed to Babette Weksler.

3 recordsLinked to original sources

PubMed2006

Classical pathway complement activation on human endothelial cells.

Endothelial cells regulate vascular integrity and express complement binding proteins including gC1qR/p33 (gC1qR), which recognize C1q, a subunit of the first component of the classical complement pathway. Experiments were performed to investigate classical complement pathway activation on resting endothelial cells and endothelial cells exposed to shear stress. C1q deposition and C4 activation (C4d) were demonstrated by solid phase ELISA and flow cytometry on human microvascular and umbilical vein endothelial cells after exposure to serum or plasma. C4d deposition was accompanied by downstream complement activation including C3b and C5b-9 deposition. C4 activation failed to occur in C1q depleted serum, but was not affected by Factor B depleted serum, confirming classical complement pathway activation. Moreover, C4 activation occurred following exposure of endothelial cells to purified C1 and C4, in the absence of other plasma proteins, and in the absence of detectable cell surface IgG and IgM. Shear stress (18 dynes/cm2) increased C1q (n=9, p<0.05) and C4d (n=9, p<0.05) deposition approximately two-fold, and enhanced endothelial cell gC1qR expression (n=7, p<0.05). Treatment of endothelial cells with anti gC1qR monoclonal antibody F(ab')2 fragments reduced C4d deposition by approximately 20% (n=5, p<0.05). These data demonstrate direct classical complement pathway activation on endothelial cells. gC1qR appears to play a minor but definable role, whereas cell surface IgG or IgM are not required.

Animals↗

PubMed2004

Ex vivo evaluation of erythrocytosis-enhanced platelet thrombus formation using the cone and plate(let) analyzer: effect of platelet antagonists.

Red blood cells (RBC) contribute significantly to haemostasis and thrombosis under oscillatory flow conditions, and erythrocytosis has been associated with increased thrombotic risk. To measure the dynamic influences of RBC on platelets, we used a recently described cone and plate(let) analyzer (CPA), evaluating the effect of haematocrit (Hct) on platelet function in whole blood under arterial flow conditions (1800/s, 2 min, 25 degrees C). Anticoagulated blood, reconstituted to varying haematocrits with autologous RBC, demonstrated a significant increase in adherent platelet aggregate formation at Hct levels >45%. This increase was not affected by pretreatment of blood with 0.05 mmol/l aspirin, but was prevented by antagonists of P2Y1, P2Y12, or P2X1, ADP and ATP receptors, and by converting exogenous ADP to ATP with creatine phosphate/creatine phosphokinase. As negligible platelet granule secretion was measured during CPA analysis, but metabolic inhibition of RBC with sodium azide or glutaraldehyde fixation inhibited erythrocytosis-enhanced increases in platelet aggregate size, adenine nucleotides contributing to shear-induced platelet aggregate formation appear to be derived from erythrocytes. These findings support the use of CPA for ex vivo evaluation of the contribution of RBC to platelet function and its inhibition under physiological shear conditions.

Anticoagulants↗

PubMed2002

Anti-platelet factor 4/heparin antibodies from patients with heparin-induced thrombocytopenia provoke direct activation of microvascular endothelial cells.

Heparin-induced thrombocytopenia (HIT) is a serious complication that occurs in approximately 1-5% of patients treated with heparin and may be associated with severe thrombotic events. HIT is mediated by antibodies directed mostly to epitope(s) formed by complexes between heparin or other anionic mucopolysaccharides and platelet factor 4 (PF4). Anti-PF4/heparin IgG antibodies from six patients with HIT were affinity purified and assessed for interaction with human microvascular and macrovascular endothelial cells (EC). The antibodies directly activated primary cultures of human bone marrow microvascular EC (HBMEC) and SV40 immortalized HBMEC (TrHBMEC) only in the presence of PF4, but did not activate macrovascular human umbilical vein EC (HUVEC) under the same conditions. These antibodies were found to bind to TrHBMEC through the F(ab)(2) portion of the anti-PF4/heparin IgG. TrHBMEC activation was characterized by an augmented release of IL-6, von Willebrand factor, soluble thrombomodulin, and by an elevated expression of the adhesion molecules P-selectin, E-selectin and vascular cellular endothelial molecule-I to different degrees. Enhanced monocyte adhesion to PF4/heparin antibody-treated TrHBMEC (33-72% adhesion) was also observed. None of these effects occurred with unstimulated HUVEC. However, pre-treatment of HUVEC with tumor necrosis factor-alpha resulted in the same changes observed with microvascular EC exposed to the HIT antibodies. Our findings indicate that anti-PF4/heparin antibodies directly activate microvascular EC while interaction with macrovascular EC requires pre-activation. These results may explain some of the specific clinical manifestations in HIT.

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