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Biomedical subjects

L B Jaques

Publications and source records attributed to L B Jaques.

At least 19 recordsLinked to original sources

Determination of absolute amounts of heparin and of dextran sulfate in plasma in microgram quantities.

Heparin and dextran sulfates 8000 are separated from citrated plasma by absorption on epichlorohydrin triethanolamine cellulose columns followed by elution with 1.1 and 1.4 mol/L NaCl in 0.05 mol/L glycine-HCl buffer. The eluate is desalted with Sephadex G25-40, dried, and dissolved in water. A 1 microliters sample is applied to an agarose gel slide. After electrophoresis, the slide is fixed and stained with toluidine blue. The sulfated polysaccharide band(s) is identified by relative electrophoretic migration. The total amount of drug is estimated by matching its optical density with that of a band on one of a set of slides with graded amounts of heparin or dextran sulfate. The reaction with toluidine blue measures the total polyelectrolyte, not just the small proportion of the drug with anticoagulant activity. Pooled normal plasma showed a trace of chondroitin and no heparin. Recovery of heparin and hydrogenated dextran sulfate that was added to pooled normal plasma was complete (lowest concentration tested was 10 micrograms/ml); however, recovery for unhydrogenated dextran sulfate declined consistently by 9 micrograms/ml for concentrations below 50 micrograms/ml, setting a limit for its recovery. Plasma samples taken from patients for coagulation tests were examined by this procedure, and in so doing, steps were ascertained to improve the procedure for routine use. Results were compared with values for prothrombin time and activated partial thromboplastin times obtained on the same samples by the clinical laboratory. Because the procedure provides an independent parameter for measurement in patients who have received heparin therapy, insight into different patient responses to the drug is therefore possible. With minor modifications, the procedure can be used for heparans, dermatans, and chondroitins, because it allows identification and microscale quantitation on the basis of charge, molecular weight, and carbohydrate structure.

Chromatography

Anticoagulant activity and operative blood loss after intrapulmonary heparin.

The safety and efficacy of heparin given by the intrapulmonary route are further assessed in this study. A single dose of heparin (2000 units/kg) was given by intratracheal instillation in dogs and measurements of plasma heparin concentration, whole blood clotting time and partial thromboplastin time made at intervals for 48 h. These values rose progressively and in parallel for 7 h and remained elevated for 48 h. A series of operations was performed on dogs within 3 h of a single dose of intrapulmonary heparin (1500 units/kg). Operations involving minimal dissection (small bowel resection) and extensive dissection (resection of muscle) were performed in two separate groups. Within each group the animals were randomly given heparin or saline. In the limited dissection group there were no differences in operative blood loss, wound healing, or sequential haemoglobin and haematocrit measurements. In the group subjected to muscle resection there was increased postoperative wound drainage and a slightly greater fall in haemoglobin and haematocrit in those given heparin. It is concluded that heparin is absorbed from the lung causing significant changes in coagulation parameters. Even with the relatively high dose of 1500 units/kg, operations were performed with minimal hazard. Intrapulmonary heparin may have important clinical applications after further investigation.

Animals

Cellular control of heparin in blood.

Many investigators have observed the uptake of exogenous heparin by cells of the reticuloendothelial system (R.E.S.). When heparin is administered by the intravenous, intramuscular, subcutaneous, intraperitoneal and intratracheal routes the anticoagulant response observed is of varying magnitude. This has led us to examine the literature for evidence of a distribution of heparin between the cellular and blood compartments. A re-evaluation of such evidence has provided a new perspective on the pharmacokinetics of heparin. This is presented here in the cellular pool concept which is based on the premise that there exists in the body a pool of cells which takes up a portion of the administered heparin, stores it and later releases it to the circulation. This concept provides a rational explanation for the different types of anticoagulant response obtained with different modes of administration.

Biopharmaceutics

A novel method to separate the layers of the intestine.

By applying cellulose acetate paper to the lumenal and serosal surface of the intestine of rats, we have divided the intestine into three layers. Examining the layers histologically, we have shown that most of the villi are removed from the mucosal surface, ther serosa alone is removed from the serosal surface, and muscle layers. This appears to provide a rapid procedure for separation of the layers of the intestine for histological and biochemical studies.

Animals

Heparin via the lung.

Only recently has it been realized that heparin is absorbed when administered into the lung. Im mice, rats, dogs and humans a large dose of intrapulmonary heparin has been shown to cause a moderate degree of hypocoagulability lasting from 48 hours to 14 days depending on the species. This study is part of an ongoing program investigating the effects of the intrapulmonary administration of heparin. Heparin, 1500 units/kg body weight was instilled into the lung in a group of 12 dogs and an equal volume of saline was given in a control group of 10 dogs. Various hemodynamic and metabolic measurements were made at intervals. As expected there was a prolonged moderate increase in clotting time, but no significant effects from intrapulmonary administration of heparin were demonstrated by any of the other measurements. Much work has yet to be done before intrapulmonary heparin can be used clinically but it has potential importance in the management of thromboembolic disease.

Animals

The mast cell/heparin paradox.

Purified heparin extracted from tissues rich in mast cells remains the ideal rapid anticoagulant in clinical practice. Nevertheless, there are grounds for doubting that an injection of commercial heparin corresponds to the release of heparin-containing granules from the mast cells. The metachromatic granule contains much more than heparin--chondroitins, heparitins, histamine (in some species 5-hydroxytryptamine also), and a variety of enzymes. Shed granules, released by trauma of any kind, are ingested by connective-tissue phagocytes and are digested. Commercial heparin, on the other hand, is taken up by cells of the reticuloendothelial system and is stored there. This apparent paradox can be resolved by conceding that the mast cell is primarily concerned with the connective tissue, as Ehrlich saw it a century ago, and that, within these broad limits, it can express itself in a variety of ways.

Animals

Heparin and sulfated mucopolysaccharides--a micro system for quantitative differentiation and determination.

Heparin (Hep), hyaluronic acid, chondroitins (sulfate) A, B, and C, and heparins (sulfate) A, B, C, and D were subjected to microelectrophoresis in barbital-agarose gel, fixed with cetylpyridinium chloride and stained with toluidine blue. The optical densities of the resulting bands were compared with optical densities obtained upon reaction with azure A in aqueous solution and with the carbazole reagent. A linear relation was obtained between optical density and concentration of purified sulfated mucopolysaccharide (SMP). Less than 1 microgram of Hep and 2 microgram of other SMPs are required for measurement by electrophoresis, while about 30 microgram of each is required with the carbazole reagent. The optical density of a mixture of SMPs was equal to the sum of the densities for the individual SMPs upon microelectrophoresis. It was demonstrated that the individual SMPs in mixtures were distinguishabed by reaction with specific enzymes and by changes in migration in agarose with barbital, phthalate, ethylenediamine, or propanediamine buffers, permitting ready demonstration and quantitation of various SMP species. Examples are shown of the application of the procedure to measure the total SMPs and individual SMPs in tissue extracts. The method is sensitive, reproducible, flexible, and measures quantities 1/30th of those measured colorimetrically, yet is relatively unaffected by protein, carbohydrate, or inorganic electrolytes.

Buffers