Participation of hepatic α/ß-adrenoceptors and AT1 receptors in glucose release and portal hypertensive response induced by adrenaline or angiotensin II.

It has been previously demonstrated that the hemodynamic effect induced by angiotensin II (AII) in the liver was completely abolished by losartan while glucose release was partially affected by losartan. Angiotensin II type 1 (AT1) and adrenergic (∝1- and ß-) receptors (AR) belong to the G-proteins superfamily, which signaling promote glycogen breakdown and glucose release. Interactive relationship between AR and AT1-R was shown after blockade of these receptors with specific antagonists. The isolated perfused rat liver was used to study hemodynamic and metabolic responses induced by AII and adrenaline (Adr) in the presence of AT1 (losartan) and ∝1-AR and ß-AR antagonists (prazosin and propranolol). All antagonists diminished the hemodynamic response induced by Adr. Losartan abolished hemodynamic response induced by AII, and AR antagonists had no effect when used alone. When combined, the antagonists caused a decrease in the hemodynamic response. The metabolic response induced by Adr was mainly mediated by ∝1-AR. A significant decrease in the hemodynamic response induced by Adr caused by losartan confirmed the participation of AT1-R. The metabolic response induced by AII was impaired by propranolol, indicating the participation of ß-AR. When both ARs were blocked, the hemodynamic and metabolic responses were impaired in a cumulative effect. These results suggested that both ARs might be responsible for AII effects. This possible cross-talk between ß-AR and AT1-R signaling in the hepatocytes has yet to be investigated and should be considered in the design of specific drugs.

Participation of hepatic α/β-adrenoceptors and AT1 receptors in glucose release and portal hypertensive response induced by adrenaline or angiotensin II

It has been previously demonstrated that the hemodynamic effect induced by angiotensin II (AII) in the liver was completely abolished by losartan while glucose release was partially affected by losartan. Angiotensin II type 1 (AT1) and adrenergic (∝1- and β-) receptors (AR) belong to the G-proteins superfamily, which signaling promote glycogen breakdown and glucose release. Interactive relationship between AR and AT1-R was shown after blockade of these receptors with specific antagonists. The isolated perfused rat liver was used to study hemodynamic and metabolic responses induced by AII and adrenaline (Adr) in the presence of AT1 (losartan) and ∝1-AR and β-AR antagonists (prazosin and propranolol). All antagonists diminished the hemodynamic response induced by Adr. Losartan abolished hemodynamic response induced by AII, and AR antagonists had no effect when used alone. When combined, the antagonists caused a decrease in the hemodynamic response. The metabolic response induced by Adr was mainly mediated by ∝1-AR. A significant decrease in the hemodynamic response induced by Adr caused by losartan confirmed the participation of AT1-R. The metabolic response induced by AII was impaired by propranolol, indicating the participation of β-AR. When both ARs were blocked, the hemodynamic and metabolic responses were impaired in a cumulative effect. These results suggested that both ARs might be responsible for AII effects. This possible cross-talk between β-AR and AT1-R signaling in the hepatocytes has yet to be investigated and should be considered in the design of specific drugs.

The hepatic clearance of recombinant tissue-type plasminogen activator decreases after an inflammatory stimulus.

We have shown that tissue-type plasminogen activator (tPA) and plasma kallikrein share a common pathway for liver clearance and that the hepatic clearance rate of plasma kallikrein increases during the acute-phase (AP) response. We now report the clearance of tPA from the circulation and by the isolated, exsanguinated and in situ perfused rat liver during the AP response (48-h ex-turpentine treatment). For the sake of comparison, the hepatic clearance of a tissue kallikrein and thrombin was also studied. We verified that, in vivo, the clearance of 125I-tPA from the circulation of turpentine-treated rats (2.2 +/- 0.2 ml/min, N = 7) decreases significantly (P = 0.016) when compared to normal rats (3.2 +/- 0.3 ml/min, N = 6). The AP response does not modify the tissue distribution of administered 125I-tPA and the liver accounts for most of the 125I-tPA (>80%) cleared from the circulation. The clearance rate of tPA by the isolated and perfused liver of turpentine-treated rats (15.5 +/- 1.3 microg/min, N = 4) was slower (P = 0.003) than the clearance rate by the liver of normal rats (22. 5 +/- 0.7 microg/min, N = 10). After the inflammatory stimulus and additional Kupffer cell ablation (GdCl3 treatment), tPA was cleared by the perfused liver at 16.2 +/- 2.4 microg/min (N = 5), suggesting that Kupffer cells have a minor influence on the hepatic tPA clearance during the AP response. In contrast, hepatic clearance rates of thrombin and pancreatic kallikrein were not altered during the AP response. These results contribute to explaining why the thrombolytic efficacy of tPA does not correlate with the dose administered.

The hepatic clearance of recombinant tissue-type plasminogen activator decreases after an inflammatory stimulus

We have shown that tissue-type plasminogen activator (tPA) and plasma kallikrein share a common pathway for liver clearance and that the hepatic clearance rate of plasma kallikrein increases during the acute-phase (AP) response. We now report the clearance of tPA from the circulation and by the isolated, exsanguinated and in situ perfused rat liver during the AP response (48-h ex-turpentine treatment). For the sake of comparison, the hepatic clearance of a tissue kallikrein and thrombin was also studied. We verified that, in vivo, the clearance of 125I-tPA from the circulation of turpentine-treated rats (2.2 + or - 0.2 ml/min, N = 7) decreases significantly (P = 0.016) when compared to normal rats (3.2 + or - 0.3 ml/min, N = 6). The AP response does not modify the tissue distribution of administered 125I-tPA and the liver accounts for most of the 125I-tPA (>80 percent) cleared from the circulation. The clearance rate of tPA by the isolated and perfused liver of turpentine-treated rats (15.5 + or - 1.3 µg/min, N = 4) was slower (P = 0.003) than the clearance rate by the liver of normal rats (22.5 + or - 0.7 µg/min, N = 10). After the inflammatory stimulus and additional Kupffer cell ablation (GdCl3 treatment), tPA was cleared by the perfused liver at 16.2 + or - 2.4 µg/min (N = 5), suggesting that Kupffer cells have a minor influence on the hepatic tPA clearance during the AP response. In contrast, hepatic clearance rates of thrombin and pancreatic kallikrein were not altered during the AP response. These results contribute to explaining why the thrombolytic efficacy of tPA does not correlate with the dose administered

Vena cava perfusion in situ: a tool for uptake studies.

While studying the uptake of trypsin and thrombin by the perfused rat liver, we verified that these proteins are internalized neither by hepatocytes nor Kupffer cells. These results raised the possibility that the enzymes might be binding to endothelial cells, either hepatic or vascular. In order to find out if the binding of enzymes to endothelial surface is a liver cell-specific phenomenon, we devised a system to perfuse the rat inferior cava vein in situ. After exsanguination, the vein was perfused with the recirculation of 30 mL of Krebs/BSA solution propellered by a pulsatile flow pump (10 mL/min). The liver was not exsanguinated, but to assure that the organ was indeed excluded from the circuit during the experiment at the end of the perfusion time we added China ink in the perfusion fluid. We verified that trypsin is extracted from the perfusion fluid by the vena cava as efficiently as by the liver, suggesting that the most of the infused trypsin is removed mainly by vascular endothelial cells when the liver perfusion model is used. On the other hand, thrombin is removed mainly by the liver cells since the uptake by the vena cava was insignificant.

Clearance of the alpha 2 macroglobulin-trypsin complex and uptake of trypsin by perfused liver.

The uptake and degradation of the alpha 2 macroglobulin-trypsin (alpha 2 m-trypsin) complex have been studied using isolated liver cells but not in the liver as a whole. We report the clearance of the complex by the isolated and exsanguinated liver of Wistar male rats, weighing 150- 280 g, and compare it with that of the free enzyme. The hepatic clearance of the alpha 2m-trypsin complex follows a pattern with a distribution phase followed by an elimination phase, which contrasts with that of trypsin where only the distribution phase is observed. The extraction of trypsin from the perfusate is Ca(2+)-independent (156 +/- 14 pmol/g liver in the presence of 2.5 mM Ca2+, N = 9, versus 140 +/- 8 pmol/g liver in its absence, N = 7) and is not affected by 100 mM NH4Cl (152 +/- 7 pmol/g liver, N = 6), 100 U/ml heparin (164 +/- 14 pmol/g liver, N = 5), 30 microliters/ml carbon particle suspension (150 +/- 13 pmol/g liver, N = 7) or an acute-phase situation induced by turpentine (125 +/- 10 pmol/g liver, N = 6) (P > 0.05, ANOVA). The hepatic clearance of the alpha 2m-trypsin complex is Ca(2+)-dependent (1.8 +/- 0.2 ml/min in the presence of Ca2+, N = 8, versus 0.6 +/- 0.03 ml/min in its absence, N = 4), affected by NH4Cl (< 0.1 ml/min, N = 7), heparin (1.1 +/- 0.2 ml/min, N = 6) and the acute-phase (0.6 +/- 0.1 ml/min, N = 6) but not by the carbon particle suspension (1.8 +/- 0.2 ml/min, N = 7). These results show that trypsin is not internalized by hepatocytes (no NH4Cl effect) or Kupffer cells (no carbon particle effect) and that the alpha 2m-trypsin complex is internalized in a Ca(2+)-dependent process by hepatocytes, but not by Kupffer cells, and is affected by an acute-phase reaction.

Clearance of the alpha2 macroglobulin-tripsin complex and uptake of trypsin by perfused liver

The uptake and degradation of the alpha2macroglobulin-trypsin (alpha2m-trypsin) complex have been studied using isolated liver cells but not in the liver as a whole. We report the clearance of the complex by the isolated and exsanguinated liver of Wistar male rats, weighing 150-280 g, and compare it with that of the free enzyme. The hepatic clearance of the alpha2m-trypsin complex follows a pattern with a distribution phase followed by an elimination phase, which contrasts with that of trypsin where only the distribution phase is observed. The extraction of trypsin from the perfusate is Ca2+ -independent (156 + 14 pmol/g liver in the presence of 2.5 mM Ca2+, N = 9, versus 140 + 8 pmol/g liver in its absence, N = 7) and is not affected by 100 mM NH4Cl (152 + 7 pmol/g liver, N = 6), 100 U/ml heparin (164 + 14 p/mol/g liver, N = 5), 30 mul/ml carbon particle suspension (150 + 13 pmol/g liver, N = 7) or an acute-phase situation induced by turpentine (125 + 10 pmol/g liver, N = 6) (P>0.05, ANOVA). The hepatic clearance of the alpha2m-trypsin complex is Ca2+ -dependent (1.8 + 0.2 ml/min in the presence of Ca2+, N = 8, versus 0.6 + 0.03 ml/min in its absence, N = 4), affected by NH4Cl (<0.1 ml/min, N = 7), heparin (1.1 + 0.2 ml/min, N = 6) and the acute-phase (0.6 + 0.1 ml/min, N = 6) but bot by the carbon particle suspension (1.8 + 0.2 ml/min, N = 7). These results show that trypsin is not internalized by hepatocytes (no NH4Cl effect) or Kupffer cells (no carbon particle effect) and that the alpha2m-trypsin complex is internalized in a Ca2+ -dependent process by hepatocytes, but not by Kupffer cells, and is affected by an acute-phase reaction.