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Endogenous pancreatic protease activity and methods for impeding their function.

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Summary

Pefabloc and Trasylol inhibit proteases during islet isolation. However, Pefabloc reduced insulin secretion, making it unsuitable, while Trasylol interfered with collagenase.

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Area of Science:

  • Biochemistry
  • Cell Biology
  • Transplantation Science

Background:

  • Serine protease inhibitors like Pefabloc and Trasylol are used in islet isolation to enhance yields by inhibiting endogenous proteases.
  • Protector Solution (PS) has been reported to improve Pefabloc's efficiency.

Purpose of the Study:

  • To evaluate the in vitro effects of Pefabloc and Trasylol on protease activities (trypsin, chymotrypsin, elastase, liberase, thermolysin).
  • To assess the impact of Pefabloc on islet function and insulin release.
  • To determine the efficacy of Pefabloc with or without Protector Solution (PS).

Main Methods:

  • EnzCheck protease assay was employed to quantify protease activities.
  • Protease inhibition was measured for Pefabloc (0.4 mmol/L, with/without PS) and Trasylol (0.43 micromol/L).
  • Islet insulin release was assessed after incubation or perifusion with Pefabloc.

Main Results:

  • Pefabloc significantly inhibited trypsin, chymotrypsin, elastase, and a switch sample, but not liberase or thermolysin.
  • Trasylol inhibited all tested enzymes except elastase and the switch sample.
  • Pefabloc's inhibitory potency was lost when initially diluted in PS.
  • Islet insulin release was significantly reduced when exposed to Pefabloc.

Conclusions:

  • Pefabloc effectively inhibits in vitro protease activity when used correctly but negatively impacts islet insulin secretion, rendering it unsuitable for islet isolation.
  • Trasylol is not compatible with collagenase due to its inhibition of liberase and thermolysin.