Endothelin mediation of insulin and glucose-induced changes in vascular contractility.

Although the prevalence of hypertension in diabetic patients is high and many factors participate, hyperinsulinemia cannot be discarded as a contributing factor. Insulin could act directly on smooth muscle altering intracellular calcium levels that mediate contraction and glucose transport or could induce the secretion of endothelin by the endothelial cells lining the vessels. The aim of the present report was to study the effect of different glucose and insulin concentrations on rat vascular smooth-muscle contractile characteristics and to determine whether insulin effects are mediated by endothelin. Femoral arteries obtained from Wistar rats were placed in an in vitro chamber and superfused with different glucose and/or insulin solutions. The contractile response to KCl 80 mmol/L, measured by the force generated, showed a significant decrease with high extracellular glucose concentrations (11 mmol/L). Insulin caused a dose-dependent increase in arterial contraction induced by KCl. This increase was significant when arteries were stimulated with 80 mmol/L KCl in the presence of 5.5 mmol/L glucose, but when 40 mmol/L KCl was used, an increase was observed with both 5.5 and 11 mmol/L glucose. The insulin-induced contraction was significantly reduced in the presence of hyperimmune anti-endothelin serum and in the presence of endothelin receptor ET(A) and ET(B) antagonists PD 151,242 and BQ-788, respectively. These results suggest that hyperinsulinemia and hyperglycemia may contribute to hypertension in diabetes and that responses to insulin are mediated partially by endothelin, thus explaining why non-insulin-dependent diabetes mellitus patients show an increase in arterial pressure before the onset of nephropathy.

Purification and characterization of a variant of human prothrombin: prothrombin Segovia.

A dysprothrombin designated prothrombin Segovia was isolated from the plasma of an individual with normal prothrombin antigen and prothrombin activity lesser than 25% of the control prothrombin activity. Activation by prothrombinase complex showed a lower amidolytic than clotting activity, which suggests a lesser generation of active intermediates than normal prothrombin. When prothrombin Segovia was activated by prothrombinase complex in the absence of factor Va, no thrombin formation was found by functional activities. SDS-PAGE analysis of the molecules derived by activation with prothrombinase complex, Taipan snake venom and Echis carinatus venom showed an accumulation of molecules not cleaved at bond Arg320-Ile321. This was more evident with Echis carinatus venom, which only acts on this bond. Our data suggest that the alteration of prothrombin Segovia impairs the scission of bond Arg320-Ile321.

The altered expression of inflammation-related molecules and secretion of IL-6 and IL-8 by HUVEC from newborns with maternal inactive systemic lupus erythematosus is modified by estrogens.

Systemic lupus erythematosus (SLE) predominantly affects women, especially those in reproductive age. Genetic contributions to disease susceptibility as well as immune dysregulation, particularly persistent inflammatory responses, are considered essential features. Our aim was to determine whether human umbilical vein endothelial cells (HUVEC) isolated from healthy newborns to women with inactive SLE show inflammation-related abnormalities that might lead to an early development of SLE in the offsprings. HUVEC isolated from six women with inactive SLE were stimulated with 2.5 ng/mL of TNF-alpha and/or physiological and pharmacological doses of 17-I(2) estradiol (E2). Then the expression of VCAM-1, ICAM-1, E-selectin, toll-like receptor-9 (TLR-9), heat shock protein 70 (HSP70) and HSP90 were measured. The concentrations of IL-6, IL-8, and IL-10 were also determined in maternal serum and in TNF-alpha stimulated and non-stimulated HUVEC culture supernatant. HUVEC from children with no family history of autoimmune disease served as controls. Our results showed that in HUVEC from SLE+ mothers, a constitutively low expression of adhesion molecules was enhanced by TNF-alpha treatment. The E2 (1 ng/mL) increased the expression of adhesion molecules but had no effect upon TNF-alpha-treated cells. IL-6 was constitutively higher in SLE+ HUVEC, whereas IL-8 was lower; E2 treatment diminished the latter. The E2 had no effect upon IL-6 and IL-8 secretions in TNF-alpha-treated cells. SLE+ HUVEC showed a disordered cytoskeleton and overexpressed HSP70, HSP90, and TLR-9. Our results indicate that endothelial cells of newborns to SLE+ mothers are in a proinflammatory condition which can be upregulated by estrogens.

[Thrombin-antithrombin complexes and F1+2 prothrombin fragment in coronary revascularization surgery. Relationship with graft occlusion]. / Complejos trombina-antitrombina y fragmento F1+2 de protrombina en cirugía de revascularización coronaria. Relación con la oclusión del injerto.

PURPOSE: To evaluate the possible role played by activation of the coagulation system in the early occlusion of grafts in patients subjected to aortocoronary bypass. PATIENTS AND METHODS: This study includes 30 patients subjected to coronary revascularization surgery. The plasma rates of thrombin-anti-thrombin complexes (TAT) and 1+2 prothrombin fragments (F1+2) were measured before and after surgery. The studies were performed by enzyme immunoassay techniques. Selective angiography was performed in every patient on the 10th day after surgery. Descriptive statistics, Student's t test and Mann-Whitney U test were used for comparison of means ans statistical analysis. RESULTS: Occlusive lesion was shown by angiography in 15 of the 30 patients. The TAT and F1+2 levels showed a statistically significant increase (p less than 0.001) along the postoperative period, without any correlation regarding the presence or absence of occlusion. The preoperative concentration of TAT was significantly higher in patients with thrombotic occlusion (p less than 0.05). CONCLUSIONS: 1) Marked activation of the coagulation system is present in patients subjected to coronary revascularization surgery. 2) The pre-operative determination of TAT may be of value for predicting early graft occlusion.

Thrombin-antithrombin complexes and prothrombin fragment 1+2 in aorto-coronary bypass surgery: relation to graft occlusion.

Graft thrombotic occlusion is a common complication in patients undergoing aorto-coronary bypass surgery. Clotting activation seems to contribute to the thrombotic event. We have determined the plasma concentrations of two hemostatic markers, thrombin-antithrombin (TAT) complexes and prothrombin fragment 1+2 (F 1+2) in 100 patients undergoing revascularization procedures of whom 81 underwent shunt angiography. Angiographically proven graft occlusion was present in 19 patients (23.5%). A significant increase of both parameters was observed immediately after surgery and on postoperative days 1 and 5 (p < 0.001), although a relationship to graft occlusion could not be demonstrated. However, the preoperative TAT concentration was higher in patients developing graft occlusion (p < 0.01). We conclude that there is a marked clotting activation in patients undergoing aorto-coronary bypass surgery, as demonstrated by elevated TAT and F 1+2 concentrations. Preoperative TAT values can be good markers of early graft occlusion.

Characterization of von Willebrand factor in primary pulmonary hypertension.

The aim of this study was to determine the value of von Willebrand factor (vWF), a well-characterized endothelial cell protein secretion, as a marker for prognosis in patients with primary pulmonary hypertension (PPH). Venous and arterial blood samples were obtained from 18 clinically diagnosed PPH patients and 12 case controls matched for age and sex. Plasma vWF antigen was determined by enzyme-linked immunosorbent assay (ELISA). The patients' multimeric vWF pattern was analyzed by sodium dodecylsulfate (SDS)-agarose-acrylamide electrophoresis, Western blot, and densitometric analysis. vWF sialic acid content was determined by a lectin-based ELISA. The PPH patients showed a higher content of vWF antigen in venous (P = 0.0026) and arterial (P = 0.0094) blood samples than controls. The mean vWF sialic acid content of the PPH patients corresponded to 37.7% of the mean value for the control group. On the basis of the hemodynamic response to vasodilator trial, the PPH patients were grouped as responders or nonresponders. The latter group showed a significantly higher plasma vWF antigen antecubital vein/radial artery ratio, an increased number of unusually large vWF multimers, and a diminished content of vWF sialic acid in comparison with the first group. We believe that our results establish the nature of vWF alterations that are related to endothelial cell damage in patients with primary pulmonary hypertension and that this could be of value when establishing the prognosis in this group of patients.