Involvement of tissue kallikrein but not plasma kallikrein in the development of symptoms mediated by endogenous kinins in acute pancreatitis in rats

Abstract

In order to investigate the mechanism of kinin release leading to vascular symptoms in acute interstitial-oedematous pancreatitis, the novel, selective inhibitors of tissue kallikrein, (2S,2'R)-2-(2'-amino-3'-(4'-chlorophenyl)propanoylamino)-N-(3-guanidinopropyl)-3-(1-naphthyl)propanoamide (FE999024, CH-2856), and of plasma kallikrein, (2'S,2"R)-4-(2'-(2"(carboxymethylamino)-3"-cyclohexyl-propanoylamino)-3'-phenyl-propanoylamino)piperidine-1-carboxamidin (FE999026, CH-4215), were used in experimental caerulein-induced pancreatitis in rats. Oedema formation and plasma protein extravasation during the 2 h infusion of caerulein were inhibited in a dose-dependent manner by i.p. pretreatment with FE999024 (7-60 micromol kg(-1)) while FE999026 had no effect at the same doses. Haemoconcentration and hypovolaemia associated with the pancreatic oedema formation during pancreatitis were significantly attenuated by FE999024 at a dose of 20 micro mol kg(-1). The reduction in circulating plasma volume was not affected by FE999026. Accumulation of amylase and lipase in the pancreas was dose-dependently reduced by FE999024 while enzyme activities in the blood serum were increased by FE999024 at 60 micromol kg(-1) indicating improved enzyme removal from the tissue. Enzyme activities in the tissue and in the blood remained unaffected by FE999026. FE999024 (20 micromol kg(-1)) largely inhibited increased tissue kallikrein-like activity in the pancreas during acute pancreatitis and also strongly attenuated influx of plasma kallikrein into the tissue. FE999026 (20 micromol kg(-1)) significantly inhibited plasma kallikrein-like activity in the pancreas but had no effect on tissue kallikrein-like activity. In conclusion, vascular kinin-mediated symptoms observed during oedematous pancreatitis in the rat are caused by the action of tissue kallikrein in the pancreas whereas an involvement of plasma kallikrein seems to be unlikely.

Figure 1

Chemical structure of (a) the tissue kallikrein inhibitor, FE999024 ((2S,2′R)-2-(2′-amino-3′-(4′′chlorophenyl)propanoylamino)-N-(3-guanidinopropyl)-3-(1-naphthyl)propanoamide; C₂₆H₃₁CIN₆O₂, mol. wt. 495.03), and (b) the plasma kallikrein inhibitor, FE999026 ((2′S,2′′R)-4-(2′-(2′′)-carboxymethylamino)-3′′-cyclohexyl-propanoylamino)-3′-phenyl-propanoylamino)piperidine-1-carboxamidine; C₂₆H₄₀N₆O₄, mol. wt. 500.65).

Figure 2

Effect of (a,b) the tissue kallikrein inhibitor, FE999024 (7–60 μmol kg⁻¹), and (c,d) the plasma kallikrein inhibitor, FE999026 (7–60 μmol kg⁻¹), on pancreatic oedema formation (a,c) and plasma protein extravasation (b,d) during acute pancreatitis induced by caeruelin (CRL; 4 nmol kg⁻¹ h⁻¹ i.v. for 2 h).The inhibitors were given i.p. 30 min prior to the infusion of caerulein or its vehicle, phosphate-buffered saline (4 ml kg⁻¹ h⁻¹). Control animals not receiving an inhibitor were injected i.p. with 154 mM NaCl solution (1 ml kig⁻¹). Significance of difference of caerulein vs saline infusion. ††P<0.01, significance of difference from caerulein without kallikrein inhibitor: ^*P<0.05; means+s.e.mean; n=5–14.

Figure 3

Effect of (a) the tissue kallikrein inhibitor, FE999024, and (b) the plasma kallikrein inhibitor, FE999026, on hypovolaemia during caerulein-induced pancreatitis. The inhibitors (7–60 μmol kg⁻¹, i.p.) or their solvent (154 mM NaCl, 1 ml kg⁻¹), were administered 30 min prior to the start of the infusion of caerulein (CRL; 4 nmol kg⁻¹ h⁻¹ for 2 h) or its vehicle, phosphate-buffered saline (4 ml kg⁻¹ h⁻¹). Peak changes in circulating plasma volume (in per cent) were calculated from repeated measurements of haematocrit in arterial blood. Significance of difference of caerulein vs saline infusion: ††P<0.01, significance of difference from caerulein without kallikrein inhibitor: ^*P<0.05; means+s.e.mean; n=5–10.

Figure 4

Effect of (a,b) the tissue kallikrein inhibitor, FE999024, and (c,d) the plasma kallikrein inhibitor, FE999026, on amylase activity in the pancreatic tissue (a,c) and in the blood serum (b,d). Acute pancreatitis was induced by i.v. infusion of caerulein (CRL; 4 nmol kg⁻¹ h⁻¹ for 2 h) while control animals were infused with phosphate-buffered saline (4 ml kg⁻¹ h⁻¹). The kallikrein inhibitors (7–60 μmol kg⁻¹) or their vehicle (154 mM NaCl solution, 1 ml kg⁻¹) were injected i.p. 30 min prior to the start of the infusion. Significance of difference of caerulein vs saline infusion: ††P<0.01, significance of difference from caerulein without kallikrein inhibitor: ^*P<0.05; means+s.e.mean; n=5–13.

Figure 5

Effect of the tissue kallikrein (t-KK) inhibitor, FE999024, and the plasma kallikrein inhibitor, FE999026, on (a) t-KK-like activity and (b) p-KK-like activity in the pancreatic tissue, determined using the synthetic substrates, S-2266 and S2302, respectively. Acute pancreatitis was induced by i.v. infusion of caerulein (CRL; 4 nmol kg⁻¹ h⁻¹ for 2 h) while control animals were infused with phosphate-buffered saline (4 ml kg⁻¹ h⁻¹). The kallikrein inhibitors (20 μmol kg⁻¹) or their vehicle (154 mM NaCl solution, 1 ml kg⁻¹) were injected i.p. 30 min prior to the start of the infusion. Significance of difference of caerulein vs saline infusion: ††P<0.01; significance of difference from caerulein without kallikrein inhibitor: ^*P<0.05; means+s.e.mean; n=4–8.