Gelatin Polypeptides Research Articles

Flow distribution and cryoprotectant concentration in rabbit kidneys perfused with glycerol solutions

Rabbit kidneys were perfused at 10 °C with a solution containing gelatin polypeptides (Haemaccel), and glycerol was introduced, and then removed, using a technique that has previously been shown to result in viable kidneys. This involved increasing the concentration of glycerol in the perfusate from zero to a maximum of 3 m, holding it at this level for 30 min, and then decreasing it at the same rate to < 0.1 m. Measurements were made of the concentration of glycerol in cortex, cortico-medullary zone, and medulla at various stages of perfusion. During the experiments it was observed that vascular resistance increased dramatically toward the end of deglycerolization, and changes in regional perfusate flow were measured by the diffusable indicator method. It was found that renal tissue is effectively permeated by glycerol using this technique. The perfusate flow throughout all regions of the kidney was reduced during deglycerolization but the greatest effect was on cortico-medullary flow, which was found to be abnormally high during the initial stages of hypothermic perfusion, but was severely impaired when the glycerol was removed. The cryoprotectant was almost completely removed by the washout procedure adopted.

The influence of gelatin polypeptides, potassium, calcium and osmolality on the hypothermic perfusion of rabbit hearts

After a period of perfusion at 37 °C with a standard perfusate, rabbit hearts were cooled to +10 °C and perfused at this temperature for 5 hr with a variety of solutions. The hearts were then rewarmed to 37 °C and perfused again with the standard perfusate to assess heart function. The effects on subsequent normothermic function of including gelatin polypeptides (Haemaccel) and of increasing the osmolality and the concentrations of K + and Ca 2+ in the solutions used for hypothermic perfusion were studied. The best results were achieved with a noncolloidal electrolyte solution containing 20 m m K + and 4.8 m m Ca 2+ which gave an average maximum percentage recovery of function of 57.9 ± 7.1%. The addition of sufficient mannitol to raise the osmolality from 300 mOsm/Kg to 420 mOsm/Kg improved (but not significantly) the maximum percentage recovery of function to 61.2 ± 8.5%

Experimental studies on continuous hypothermic liver perfusion with a synthetic solution containing gelatin polypeptides (Haemaccel)

A rat liver model has been developed for studying preservation by continuous hypothermic perfusion. Perfusion for 5 and 24 hr with a defined solution containing Haemaccel as colloid at a temperature of 5–7 °C was found to result in good preservation of the metabolic activity of the livers as assessed by the ability of the tissue to synthesize urea and take up galactose. In common with other continuous perfusion methods, there was increasing evidence of cell damage as perfusion progressed, and there was an associated depletion of cellular K + and an increase in tissue water content. There was a considerable increase in vascular resistance, as indicated by a fall in flow rate at a constant pressure in the later stages of perfusion, similar to that which has been reported in other liver preservation experiments using plasma derivatives or Dextran. Further studies are envisaged to attempt to analyze, in a controlled fashion, the nature of the damage incurred by the liver during storage and the requirements for maintenance of a viable graft at hypothermia.

Renal preservation by hypothermic perfusion. IV. The use of gelatin polypeptides as the sole colloid.

Abstract Rabbit kidneys were perfused with a solution of extracellular electrolyte composition, rendered hypertonic with glucose and containing 1.75% Haemaccel (Hoechst) as the sole colloid. Perfusions were carried out at 10 °C for 24 and 48 hr using perfusion pressures of 20 or 40 mm Hg. Function, was tested by autografting with immediate contralateral nephrectomy. All the kidneys perfused at 20 mm Hg for 24 hr showed excellent life-sustaining function and 7 of 10 survived in the 40 mm Hg group; the difference was not statistically significant. However, all the kidneys perfused for 48 hr failed to sustain life, and histological examination revealed extensive breakdown of the microcirculation. The 24 hr results were similar to those previously obtained with an albumin-based perfusate, but the 48-hr results were inferior: However, the obvious advantages of a well-standardised, cheap, and easily stored perfusate are such as to justify further study of gelatin-derived colloids for organ preservation.

HYPOTHERMIC PERFUSION OF RABBIT KIDNEYS WITH SOLUTIONS CONTAINING GELATIN POLYPEPTIDES

Rabbit kidneys were perfused with a solution of extracellular electrolyte composition, made hypertonic with glucose and containing the gelatin polypeptide preparation Haemaccel (Hoechst) as the only colloid. Perfusions were carried out at 5 and 10 C for 19 hr, and function was tested by autografting. All of the kidneys perfused at the higher temperature showed immediate life-sustaining function after transplantation and contralateral nephrectomy, whereas only one graft of five perfused at the lower temperature showed any function. The suitability of the Haemaccel solution as a vehicle for introducing the cryoprotective agent glycerol was tested by perfusing kidneys for 4 hr with a solutiont containing 2% glycerol; the function of these organs was similar to that of kidneys transplanted without perfusion. Ultrastructural examination of kidneys perfused for 24 hr at 10 C showed excellent structural preservation, but measurements of water and ion contents and the penetration of marker molecules in nonmetabolizing kidneys showed 2.8% Haemaccel to be somewhat less effective than 6% bovine serum albumin in stabilizing these values. The Haemaccel perfusate is considered to be highly suitable for the introduction and removal of cryoprotective agents, and the results of hypothermic preservation by continuous perfusion are encouraging.

Hypothermic perfusion of rabbit kidneys with a perfusate containing gelatin polypeptides (haemaccel)

Hypothermic perfusion of rabbit kidneys with a perfusate containing gelatin polypeptides (haemaccel)

Gelatin: a poor substrate for a mammalian collegenase.

A rabbit tumor collagenase was purified more than 5000-fold. In this form it degrades native collagen in helical conformation at 37 degrees C, pH 7.6, into two fragments, but it had little capacity to cleave gelatin, an indication of the importance of higher-order structure of substrate for this enzyme in pure form. It is likely that, in vivo, enzymes other than collagenase degrade gelatin polypeptides produced by primary collagenolysis.

An endopeptidase from rheumatoid synovial tissue culture

1. 1.|An endopeptidase, capable of degrading gelatin to small fragments, has been found in the medium from cultures of rheumatoid synovium. This peptidase, which does not degrade undenatured collagen, has numerous characteristics in common with the rheumatoid synovial collagenase: It is inhibited by EDTA, by serum proteins and by sulfhydryl compounds such as cysteine; it has a broad pH optimum in the neutral range; its synthesis and release does not seem to be stimulated by addition of sucrose to the culture media. 2. 2.|The protease has a mol. wt. approximately twice that of the collagenase, and can hydrolyze a chromophore-peptide (Pbz- l-Pro-Leu-Gly-Pro- d-Arg) synthesized as a specific substrate for bacterial collagenase. Using a sensitive assay for neutral protease activity with 14C-labeled gelatin as substrate, the synovial collagenase has a limited capacity to degrade gelatin peptides to small fragments (< 5000 mol. wt.). 3. 3.|These data suggest that collagen may be degraded in vivo, extracellularly, by at least two enzymes—a collagenase to make an initial cleavage through the collagen molecule and a protease to degrade further the gelatin polypeptides resulting from collagenolysis.