ABSTRACT
Introduction: Fibrin tissue adhesive, which has applications in several areas of medicine, can be prepared by different methods. Aim: To compare fibrin tissue adhesives prepared by 3 different methods. Method: In this prospective experimental laboratory study, fibrin tissue adhesives prepared by the use of plasma fibrinogen (group 1), cryoprecipitation (group 2), and precipitation by ammonium sulfate (group 3) were tested on 15 rabbits and 10 fragments of dura mater. The quality of the clots was assessed in terms of the success of the healing process, local toxicity, graft adhesion capacity, and degree of adhesion of 2 fragments of dura mater produced. Results: All methods produced a clot with high adhesion and no toxicity, but tensile strength testing revealed that the glue produced from the ammonium sulfate-precipitated clot (group 3) was the strongest, requiring 39 g/cm ² to separate the fragments as opposed to 23 g/cm ² for group 2 and 13 g/cm ² for group 1. Conclusion: All methods produced good results as far as clot formation and non-toxicity, but ammonium sulfate precipitation produced the best tensile strength and was thus the most effective method of preparing fibrin tissue adhesive...
Subject(s)
Animals , Rabbits , Fibrin Tissue Adhesive/classification , Fibrin Tissue Adhesive/therapeutic use , Biological Therapy , Rabbits/surgery , Fibrinogen/isolation & purification , Plasma , Sutures , Tensile StrengthABSTRACT
Two fibrinogenolytic enzymes, Bothrops alternatus metalloprotease isoform (BaltMP)-I and II, were purified from Bothrops alternatus venom using Diethylaminoethyl (DEAE) Sephacel, Sephadex G-75 and Heparin-Agarose column chromatography. Purified BaltMP-I and II ran as single protein bands on analytical polyacrylamide gel electrophoresis and showed molecular weights of 29000 and 36000, respectively, under reducing conditions in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). BaltMP-II, but not BaltMP-I, displayed blood-clotting activity in bovine plasma, which was about 10-fold higher than that of the crude venom. Both enzymes were proteolytically active against bovine fibrinogen as substrate. When fibrinogen and each enzyme were incubated at 37°C, at a ratio of 1:100 (w/w), BaltMP-II cleaved preferentially the Aalpha -chain and more slowly the Bbeta -chain. The action of BaltMP-I was similar, but lower. None of the proteases degraded the gamma-chain of fibrinogen. The fibrinogenolytic activity of the enzymes was inhibited by 1,10-phenanthroline, suggesting they are metalloproteases. Since both enzymes were found to cause defibrinogenation when intraperitoneally (i.p.) administered to mice, they can be of medical interest as a therapeutic agent in the treatment and prevention of arterial thrombosis.(AU)
Subject(s)
Animals , Fibrinogen/isolation & purification , Bothrops , Crotalid Venoms/isolation & purification , Metalloproteases , Thrombosis , Protein IsoformsABSTRACT
The effect of sucrose (60 per cent w/w) and 1 M glycine as thermal stabilizers for fibrinogen in cryoprecipitate was studied. Sucrose (9.2 g) and glycine (0.9 g) were dissolved in 6 g of cryoprecipitate and the solution was pasteurized at 60 degrees C for 10 h. The preparation was then dialyzed for 20 h in phosphate buffered saline (PBS), lyophilized, stored for one week at -40 degrees C and resuspended in distilled water. The recovery of total proteins and fibrinogen in the final product averaged 66.4 +/- 4.1 per cent and 43.8 +/- 6.4 per cent of the initial contents, respectively (mean +/- SEM, N = 9). The pasteurization of cryoprecipitate in the presence of PBS (control experiments) produced extensive precipitation, which is characteristic of protein denaturation. Thus, this method partially protected fibrinogen and other proteins in cryoprecipitate from inactivation by prolonged exposure to heat during pasteurization