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Previous investigations have demonstrated specific receptors and associated mitogenic actions for insulin and insulinlike growth factors I and II (IGF-I and II) in postnatal bovine aortic smooth muscle. Using fetal tissue we have observed different patterns of binding and action for these peptides. Smooth muscle cells isolated from near-term fetal bovine aortae were studied in early passage. Specific receptors for both IGF-I and IGF-II were identified. Specific binding averaged 5.7%/2.5 X 10(5) cells for IGF-I, and 16.2% for IGF-II, and 0.3% for insulin. High affinity Kd for both IGF receptors were nanomolar. IGF-II was fivefold less potent than IGF-I in displacing IGF-I binding. IGF-I showed no affinity for the IGF-II receptor. Insulin, at physiologic concentrations, was incapable of displacing either IGF-I or IGF-II binding. Cellular incorporation of [methyl-3H]thymidine was stimulated at the lowest dose of IGF-I tested, 0.5 ng/ml. IGF-II showed no effect up to 100 ng/ml, after which a sharp increase in incorporation was noted. Insulin had a similar effect only at concentrations greater than 0.5 micrograms/ml, with a maximal response noted at 5 to 10 micrograms/ml. Our results indicate that fetal bovine aortic smooth muscle cells have an abundance of IGF receptors but lack specific insulin receptors. In addition, IGF-II binding levels are three times higher than for IGF-I. These results are consistent with observations in other species, in which a predominance of IGF over insulin receptors has been demonstrated in fetal tissue, and provide further evidence for a role for the IGFs in embryonic cellular metabolism.

Septicemic/endotoxic-induced adult respiratory distress syndrome (ARDS) remains a major clinical problem. The present study was to determine in the E. coli endotoxemic sheep ARDS model the efficacy of combination prophylaxis with antithrombin-III (AT-III) and alpha 1-proteinase inhibitor (alpha 1-PI). We reasoned that 1) AT-III supplementation would ameliorate the endotoxin-induced coagulopathy, 2) alpha 1-PI supplementation would attenuate pulmonary damage caused by neutrophil elastase and inactivation of AT-III by neutrophil elastase, and 3) the therapeutic effects of this combination would be additive or synergistic. The typical increases in lung lymph flow microvascular permeability to protein, transvascular protein flow and transvascular protein clearance, and decrease in systemic arterial PO2 were prevented or significantly attenuated during 5 hours of endotoxemia by the AT-III/alpha 1-PI combination pretreatment. Limited efficacy was observed with AT-III pretreatment, and none was seen with alpha 1-PI alone. Results of this study demonstrate that combining AT-III and alpha 1-PI prophylaxis prevents or attenuates indices of ARDS during gram-negative endotoxemia and that this efficacy is due to a statistically significant synergism between AT-III and alpha 1-PI.

We have examined the antithrombin effects of various phosphate-containing polyanions (including linear polyphosphates, polynucleotides and the phosphoserine glycoprotein, phosvitin) on the glycosaminoglycan-binding plasma proteinase inhibitors, antithrombin III (ATIII) and heparin cofactor II (HCII). These phosphate-containing polyanions accelerate the HCII-thrombin reaction, as much as 1600-fold in the case of phosvitin. The HCII-thrombin reaction with both phosvitin and polynucleotides appears to follow the ternary complex mechanism. The HCII-thrombin complex is rapidly formed in the presence of these phosphate polyanions (each at 10 micrograms/ml) when 125I-labeled thrombin is incubated with human plasma (ex vivo). None of these phosphate polyanions accelerate the ATIII-thrombin reaction. Our results suggest that the antithrombotic effect of these phosphate-containing polyanions is mediated by HCII activation and not by ATIII.

A new method of determining the oligosaccharide composition of commercial glycosaminoglycan heparin is described in which heparin was first depolymerized using heparin lyase (EC 4.2.2.7), and then analysed by a single h.p.l.c. step. All 20 of the porcine and bovine heparins examined were found to contain a small number of major oligosaccharide components, which on average comprised 86% of their mass. The five most abundant oligosaccharides have defined chemical structures. Although the relative abundance of oligosaccharides varied, the heparins examined were surprisingly similar. Porcine, bovine, low-Mr, and high and low antithrombin III (ATIII)-affinity heparins, however, each had distinctly different proportions of these major oligosaccharide components. The concentrations of one of these five oligosaccharides, containing a portion of the ATIII binding site, correlated with the anticoagulant activity of the ATIII-affinity-fractionated porcine-mucosal heparins from which it was derived. An additional oligosaccharide of undetermined structure was found in significant quantities in both bovine heparin and high ATIII-affinity porcine-mucosal heparin. The correlation between oligosaccharide concentration and anticoagulant activity suggests that the oligosaccharide is derived from a structural variant of the ATIII-binding site. Finally, for the heparins examined chondroitin/dermatan sulphate formed 0.6-7.4% of their mass.

Breast cancer tissue from 190 patients was studied for immunocytochemically reactive estrogen and progesterone receptors (ER, PR). Parallel cytosol ER and PR assays were performed on 159 of these patients using the dextran-coated charcoal (DCC) method. For the immunocytochemical determination, monoclonal antibodies to ER (ER-ICA kit) and PR were used in an immunoperoxidase procedure. Agreement between the two techniques in postmenopausal patients was better than in the premenopausal group (ER, kappa = 0.597 vs. 0.398; PR, kappa = 0.460 vs. 0.329). The median ER cytosol concentration in receptor-positive postmenopausal patients was significantly higher than in receptor-positive premenopausal patients (87 vs. 31 fmol/mg cytosol protein, p less than 0.001). A similar trend was also found in the immunocytochemical ER assay (270 vs. 207 histoscore units, p greater than 0.05). Significantly higher cytosol ER contents were found in patients with low serum estradiol concentration. The proportion of ER-negative tumors was slightly higher in the premenopausal patients by both methods. In the PR assays (biochemical or immunocytochemical) there were no significant differences between the two patient groups in the proportion of PR-negative tumors or in the median PR content in PR-positive tumors.

To examine alterations of the kallikrein-kinin system and of the complement due to the bolus injection of newer non-ionic contrast agents, venous blood samples were taken before and 3 min after angiography. There were no adverse contrast reactions clinically evident. Prekallikrein, kallikrein inhibition, beta-factor XIIa inhibition, C1-esterase inhibitor, C1q, C3, ATIII, HMW-kininogen, fibrinogen and factor XII were determined. Bolus injection of the contrast medium caused an activation of the kallikrein-kinin system (p less than 0.05) with reduction of prekallikrein, kallikrein-inhibition, beta-factor XIIa inhibition and C1-esterase inhibitor. The levels of C1q and C3 were also decreased (p less than 0.05) indicating an activation of the complement. Our results demonstrate, that angiography causes a significant activation of the kallikrein-kinin as well as of the complement system in spite of the use of newer non-ionic contrast agents.