Effects of hypothermia on energy metabolism of metabolically loaded liver

The effects of hypothermia on adenine nucleotide level, mitochondrial metabolism and redox state of the remnant liver were studied in 25, 50, 70 and 93% hepatectomized rabbits. In 25% of the hepatectomized rabbits, energy charge levels and mitochondrial phosphorylative activities remained unchanged, while in 50 and 70% hepatectomized rabbits, the energy charge levels decreased maximally at 24 hours after hepatectomy with a concomitant rise of mitochondrial phosphorylative activity. In 93% hepatectomized rabbits, the energy charge levels decreased rapidly without enhancement in mitochondrial phosphorylative activity and these rabbits died within 12 hours after hepatectomy. Body cooling significantly increased the energy charge of the remnant liver with further enhancement in mitochondrial phosphorylative activity in 25, 50 and 70% hepatectomized rabbits, but did not increase the energy charge in 93% hepatectomized rabbits. On the other hand, during body rewarming, the energy charge levels of the remnant liver decreased. More than 60% of 70% hepatectomized rabbits died during body rewarming, while all of 50% or less hepatectomized rabbits remained alive. Thus, body cooling may provide remarkable protection for the remnant liver from an energy crisis under the condition that compensatory mitochondrial enhancements are present.     

Insulin requirements for hepatic regeneration following hepatectomy.

On the basis of changes in the adenine nucleotide the mitochondrial metabolism of the remnant liver, insulin requirements for hepatic regeneration were studied in diabetic rats treated with varying amounts of alloxan. Mildly diabetic rats with less than 30% inhibition in maximal portal insulin response to oral glucose load, showed a parabolic glucose tolerance pattern and could tolerate partial hepatectomy. Whereas, severely diabetic rats with more than 45% inhibition showed a linear glucose tolerance pattern and died within 24 hours after partial hepatectomy. In the former rats, the energy charge (ATP + 1/2ADP/ATP + ADP + AMP) levels of the remnant liver decrease slightly at an early period after partial hepatectomy but could be restored rapidly to normal levels with a concomitant rise of oxidative phosphorylation in remnant liver mitochondria. In contrast, the energy charge levels in the latter groups fell more markedly and could not be restored, because of insufficient enhancement of mitochondrial oxidative phosphorylation. It is suggested that an enhancement in mitochondrial phosphorylative activity of the remnant liver following partial hepatectomy is inhibited in proportion to the severity of impaired insulin secretion, resulting in a decrease of the potential functional capacity of liver.     

Mitochondrial phosphorylative activity and DNA synthesis in regenerating liver of diabetic rats

The changes in the incorporation rate of [¹⁴C]thymidine into hepatic cell DNA and the mitochondrial phosphorylative activity and hepatic energy charge following partial hepatectomy were studied in four groups of diabetic rats (Groups A, B, C, and D), in which diabetes was induced by intravenous injection of alloxan of 35, 49, 63, and 70 mg/kg of body wt. In normal rats, the mitochondrial phosphorylative activity was enhanced maximally during a transient decrease in the hepatic energy charge (ATP+1/2ADP/ATP+ADP+AMP) at 3 hr after partial hepatectomy. The incorporation rate of [¹⁴C]thymidine into hepatic cell DNA began about 18 hr later, and reached its peak between the 22nd and 24th hour. In diabetic rats, the enhancement of mitochondrial phosphorylative activity decreased in proportion to the severity of the decrease in the portal-fasting IRI level. The suppression by diabetes in the incorporation rate of [¹⁴C]thymidine into hepatic cell DNA was correlated positively with a decrease in the preceding enhancements in mitochondrial phosphorylative activity. In Groups A and B, the energy charge levels decreased but were restored rapidly, while those of Groups C and D decreased linearly without restoration. It is suggested that since biosynthetic enzymatic activities exhibit high activity at an energy charge above 0.85, the depression of DNA synthesis in the regenerating liver of diabetic rats is due, at least partially, to a decrease in the hepatic energy charge, which is based on suppression of an enhancement in mitochondrial phosphorylative activity following insulin deficiency.     

Changes in hepatic energy metabolism in rats with acute necrotizing pancreatitis

In this study, changes of hepatic cellular ATP, ADP, and AMP, concentrations and mitochondrial oxidative phosphorylation were investigated in rats with experimental acute necrotizing pancreatitis (ANP). It was found that energy change (ATP + 1/2 ADP)/(ATP + ADP + AMP) of the liver decreased from 0.866 to 0.806 (P less than 0.05) 24 h after ANP, and to 0.769 (P less than 0.01) at 48 h. On the other hand, mitochondrial phosphorylative activity increased to 130% and 157% over the control at 12 h and 24 h respectively, and then rapidly dropped to 62% of normal value at 48 h. Blood ketone body ratio was positively correlated with hepatic energy charge level in ANP. The authors came to the following conclusions that: (1) In ANP, mitochondrial function damage resulted in decreased hepatic energy charge, which, in turn, led to hepatocellular impairment; (2) the measurement of blood ketone body ratio was a reliable indicator by which to assess the energy status of the liver in ANP.     

Changes in energy metabolism of allografts after liver transplantation

To evaluate the function of energy metabolism in allografts after liver transplantation, changes in hepatic energy charge levels, oxidative and phosphorylative activities of mitochondria and arterial blood ketone body ratio (acetoacetate/3-hydroxybutyrate; KBR) were studied in piglets. Hepatic energy charge levels decreased to 0.831 +/- 0.010 at 3 days and 0.836 +/- 0.009 at 3 weeks after operation compared to the preoperative value of 0.868 +/- 0.006 (p less than 0.01), and returned to 0.856 +/- 0.007 at 6 weeks. Mitochondrial oxidative and phosphorylative activities were moderately enhanced to 19.14 +/- 2.07 (10(-10) mol ATP/mg of mitochondrial protein/s) at 3 days and 20.89 +/- 1.72 at 3 weeks compared to the preoperative value of 16.74 +/- 2.36, and returned to 16.65 +/- 1.54 at 6 weeks. There was no significant difference in the concentrations of mitochondrial respiratory components, except in cytochrome c + c1. KBR decreased immediately at the beginning of the anhepatic phase and rapidly recovered to the preoperative level within 60 min after revascularization of allografts. There was no change in KBR during the postoperative course except in cases with clinical deterioration. From these results, it is suggested that the mitochondrial capacity for ATP synthesis was enhanced to compensate for the decreased energy charge level and that a decreased KBR is a sign of a critically deranged metabolic function in allografts.     

The Effects of Heterologous Liver Cross-Hemodialysis on Adenylate Energy Charge of the Remnant Liver after Major Hepatic Resection

The effect of liver support on the adenylate energy charge (ATP + 0.5ADP)/(ATP + ADP + AMP) of the remnant liver after major hepatic resection was studied in rabbits. The present study emphasized the principle of restoring the decreased energy charge level of the remnant liver after major hepatic resection by use of an ex vivo heterologous liver cross-he-modialysis with an interposed Cuprophan membrane. The energy charge level provides the cell with a very sensitive intracellular control mechanism. Regulatory enzymes from biosynthetic sequences exhibit very little activity at low levels of energy charge, and their activities increase sharply at high-energy charge levels. The energy charge level of the remnant liver maximally decreased from the control level of 0.860 to 0.767 at 24 hours after 70% hepatectomy. The energy charge level increased from 0.767 to 0.857 after two hours of cross-hemodialysis with an interposed Cuprophan membrane between the 24-hour, 70% hepatectomized rabbit and an ex vivo pig liver with high energy charge. The above results suggest that this ex vivo pig cross-hemodialysis may be effective for biosynthesis in the regenerative processes of the remnant liver.     

Mitochondrial changes in phospholipid molecular species during the increased oxidative phosphorylation after hepatectomy.

The changes in liver mitochondrial and microsomal phospholipid molecular species were analyzed during the period of remarkably increased oxidative phosphorylation following partial hepatectomy in rabbits. At 24 hours after hepatectomy, phosphorylative activity increased significantly from 69.7 +/- 5.5 to 118.5 +/- 5.7 nmol of ATP synthesized/min/mg protein, compared to the sham operated group. The ratio of phosphatidylethanolamine to phosphatidylcholine (PE/PC) in mitochondria increased significantly in the hepatectomy group compared with the sham operated group. Remarkable changes in molecular species were observed in mitochondrial phosphatidylethanolamine. 1-Stearoyl-2-arachidonoyl species decreased in the hepatectomy group. On the other hand, microsomal phospholipids hardly changed compared with mitochondrial ones. The change in content of 1-stearoyl-2-arachidonyl phosphatidylethanolamine in mitochondria tended to return to normal levels concomitant with the normalization of phosphorylative activity. The changes in content of mitochondrial phospholipids, especially phosphatidylethanolamine, might also be related to enhancement of phosphorylative activity.     

Changes in blood glucose levels in relation to blood ketone body ratio following hypertonic glucose infusion in 70% hepatectomized rabbits

The effects of postoperative infusion of a hypertonic glucose solution on the blood glucose level, blood ketone body ratio (acetoacetate/beta-hydroxybutyrate), and plasma alanine and proline levels were studied in 70% hepatectomized rabbits (group A) and in rabbits 70% hepatectomized and, in addition, subjected to bile duct obstruction at 12 h after hepatectomy (group B). Glucose infusion was started at the end of hepatectomy and continued for 20 h. The blood glucose level in group A remained at approximately 300 mg/dl throughout the study; however, it reached 789 mg/dl in group B at 20 h. The blood ketone body ratio, which reflects hepatic mitochondrial redox potential, decreased from 0.90 +/- 0.09 in untreated rabbits to 0.38 +/- 0.05 in group A, and to 0.19 +/- 0.03 in group B at 20 h. As the blood ketone body ratio decreased, plasma proline and alanine levels increased rapidly (proline, r = -0.601, p less than 0.02; alanine, r = -0.640, p less than 0.001). In addition, the blood ketone body ratio was positively correlated with the hepatic energy charge level [(ATP + 0.5 ADP)/(ATP + ADP + AMP)] (r = 0.57, p less than 0.001). It is suggested that the entry of glucose and amino acids into the Krebs cycle is inhibited as the blood ketone body ratio decreases, and under such conditions the infused glucose tends to accumulate, resulting in severe hyperglycemia.     

Early metabolic disturbances in the liver following unilateral hepatic or common bile duct obstruction in rabbits.

The early metabolic disturbances in the liver following ligation of the left hepatic or the common duct were studied in the rabbit. In the rabbits subjected to ligation of the left hepatic bile duct, the phosphorylative activity of the liver mitochondria in the ligated lobe decreased to about 50% of controls 48 h after the ligation, while that in the nonligated lobe increased 2-fold. These rabbits continued to live. On the other hand, in the rabbits subjected to ligation of the common bile duct, the mitochondrial phosphorylative activity increased slightly at 3 h after the ligation but fell rapidly thereafter. The mitochondrial free NAD+ to NADH ratios and hepatic energy charge (ATP+1/2ADP/ATP+ADP+AMP) markedly decreased after the ligation. Most of these rabbits did not survive over 2 days. These results indicate that the metabolic disturbances in the jaundiced liver are due to an impairment of the oxidative phosphorylation. In addition, it is suggested that the maintenance of the energy balance by enhanced mitochondrial phosphorylative activity is readily impaired in the jaundiced liver.     

The role of enhanced mitochondrial phosphorylation in rat liver transplantation.

The effects of organ preservation on mitochondrial oxidative phosphorylation activity, adenylate hepatic energy charge, cytochrome content, and redox state of NAD+/NADH couple in rat liver transplantation were compared between a nonpreservation group and a preservation group with grafts preserved for 12 hr in Euro-Collin's solution. At 3 hr after transplantation, the energy charge in the preservation group decreased to 0.60 +/- 0.02 from the control value of 0.86 +/- 0.01, accompanied by a reduction of intramitochondrial redox state of NAD+/NADH couple. In contrast, in the nonpreservation group, the decrease in energy charge was minimally decreased to 0.79 +/- 0.04 due to the compensatory enhancement of mitochondrial oxidative phosphorylation activity. These results suggest that an enhanced mitochondrial ATP synthesis and a reduced intramitochondrial redox state are important factors affecting survival following rat liver transplantation.     

Effects of partial deprivation of portal blood on arterial blood ketone body ratio in rabbits

To examine the effects of portal blood deprivation on energy metabolism of the liver, we studied: (1) the ketone body ratio (acetoacetate/beta-hydroxybutyrate) in liver tissue, which is in equilibrium with the free NAD+/NADH ratio in liver mitochondria, in the ligated lobe (LL) and nonligated lobe (NLL), (2) the hepatic energy charge [EC = (ATP + 1/2 ADP)/(ATP + ADP + AMP)] in both LL and NLL, and (3) the arterial blood ketone body ratio (BKBR) after left portal vein branch ligation (PBL) in rabbits. As found in LL after PBL, portal blood deprivation decreased the tissue ketone body ratio. The EC in LL significantly decreased after PBL, but recovered 7 days after PBL since the LL became atrophic. The BKBR remained within the normal range, even when 60% of the total liver was deprived of portal blood.     

Role of mitochondrial enhancement in maintaining hepatic energy charge level in endotoxin shock

The purpose of this study is to clarify the nature of apparent abnormalities in fuel substrate utilization which occur during progressive endotoxin shock and to relate these findings with its implications for circulatory behavior. After the administration of endotoxin, marked differences appeared in the time courses of hepatic energy charge and mitochondrial oxidative and phosphorylative activities between the early stage with normal blood pressure and the late stage with low blood pressure. In the early stage, the hepatic energy charge was maintained at near normal levels with a concomitant enhancement in mitochondrial oxidative and phosphorylative activities. In the late stage, this enhancement of mitochondrial oxidative and phosphorylative activities was depressed concomitant with a fall in hepatic energy charge. The mitochondrial enhancement was further associated with a fall in mitochondrial redox state, a rise in ketone body formation, and normoglycemia, indicating an acceleration of free fatty acid β-oxidation and gluconeogenesis at the early stage. In comparison, mitochondrial inhibition was accompanied by a further fall in mitochondrial redox state and hypoglycemia, indicating an inhibition of gluconeogenesis at the late stage. It is suggested that an enhancement in mitochondrial oxidative and phosphorylative activities is a protective mechanism which compensates for the fall in hepatic energy charge, and thus plays an essential role in the survival and recovery.     

Significance of mitochondrial enhancement in hepatic energy metabolism in relation to alterations in hemodynamics in septic pigs with severe peritonitis

Liver mitochondrial phosphorylative activity, hepatic adenine nucleotides, and hemodynamic parameters were studied in 23 pigs with peritonitis induced by cecal ligation and perforation. Between 2 and 7 days after treatment, the onset of an apparent hyperdynamic state, characterized by an increased cardiac index and decreased total peripheral resistance index, occurred. In this period, the energy charge level was barely maintained and the occurrence of liver mitochondrial enhancement was observed. However, 10-14 days after treatment, the onset of a hypodynamic state with its inverse patterns in hemodynamics took place. In this period, the energy charge level fell to 0.68, concomitant with a marked decrease in mitochondrial function. Further, a high positive correlation between the cardiac index and liver mitochondrial phosphorylative activity was noted (r = 0.85, p less than 0.01). These results suggest that liver mitochondria play a major role in the metabolic and hemodynamic adaptations occurring during sepsis.     

Effects of glucagon and insulin on poly(ADP-ribose) polymerase activity and blood ketone body ratio after partial hepatectomy in rats

The effects of glucagon and insulin on liver nuclear poly(ADP-ribose) polymerase activity and blood ketone body ratio after rat partial (68%) hepatectomy were examined. Liver weight regeneration rate was enhanced by glucagon and insulin after 5th posthepatectomy day. The maximal value of poly(ADP-ribose) polymerase activity without glucagon and insulin was revealed as 368 +/- 64 pmole/mg/min on 5 days after the hepatectomy. In contrast, the enzyme activity with glucagon and insulin reached to the peak value as 253 +/- 42 pmole/mg/min on 2 days after the hepatectomy. The amounts of DNA per nuclear protein showed similar changes with the changes of poly (ADP-ribose) polymerase activity after the hepatectomy. Blood ketone body ratio showed almost similar changes in both groups, except transitional decrease in the group without glucagon and insulin on 5th postoperative day. It is suggested that, to promote remnant liver regeneration, the combined therapy of glucagon and insulin may act directly to nucleic acid metabolism through the changes of poly(ADP-ribose) polymerase activity and preserve energy charge level by the suppression of NAD consumption by massive poly(ADP-ribose) formation.     

Bacteremic shock: aspects of high-energy metabolism of rat liver following living Escherichia coli injection

Sublethal and lethal doses of living Escherichia coli (E. coli) (serotype: O-18) were injected intravenously into rats. The amounts of live E. coli injected were 0.5 ml of 0.8 – 1.0 × 10⁹ or 2.5 – 3.0 × 10⁹ organisms/ml saline per 100 g body weight for the sublethal (SL) or lethal (L) groups, respectively. The adenylate energy charge (ATP + 1/2ADP/ATP + ADP + AMP) and free NAD⁺/NADH ratios of the mitochondrial and cytoplasmic compartments in rat livers were measured at 3, 6, 12, and 24 hr following E. coli injection. At 6 hr in both groups, the hepatic energy charge levels were decreased from 0.856 in controls to 0.823 (SL) and 0.812 (L), associated with elevated mitochondrial NAD⁺/NADH ratios (P < 0.05). At 12 hr the sublethal group maintained the above state. In the lethal group, however, the hepatic energy charge was markedly decreased (P < 0.001), associated with remarkably decreased NAD⁺/NADH ratios in both mitochondria and cytoplasm (P < 0.01 and P < 0.001). In both groups the total ketone body (acetoacetate + β-hydroxybutyrate) contents were decreased with an elevation of mitochondrial NAD⁺/NADH. From these results, it is concluded that the difference between the sublethal and lethal groups is characterized by the progressive deterioration of the high-energy level in the lethal group, possibly due to restricted mitochondrial respiration. In addition, it is suggested that the elevated mitochondrial NAD⁺/NADH ratio may play a role in decreased ketone body contents in the liver following E. coli injection.     

Influence of colloidal carbon phagocytosis as a metabolic load on the hepatic energy metabolism in rats

The influence of phagocytosis on hepatic energy metabolism was investigated in rats injected with colloidal carbon. Following injection of 6 mg colloidal carbon per 100 g body wt (BW) the hepatic energy charge potential (ECP) remained unchanged. However, oxidative phosphorylation of the isolated mitochondria (PR) increased significantly to 120 and 124% of the control at 1 and 3 hr, respectively, and returned to the normal range at 6 hr. The pyruvate level rose to 195 and 150% at 3 and 6 hr. The lactate level was elevated to 195 and 143% at 3 and 6 hr. Following injection of 20 mg colloidal carbon per 100 g BW, the ECP decreased from 0.865 to 0.783 at 1 hr, but recovered to the normal range thereafter. The PR increased more markedly to 135% at 1 hr, concomitant with an elevation of the ketone body ratio. At 3 hr it remained elevated at 118%, but returned to the normal level at 6 hr. The pyruvate and lactate levels increased to more than 250% at 1 and 3 hr. At 6 hr, the lactate level returned to the control level, but the pyruvate level showed still higher level than the control. These results indicate that phagocytosis exerts an acute metabolic load on the total liver. It was tentatively proposed that parenchymal and nonparenchymal cells in the liver have interrelation in the energy metabolism, and that an enhancement of hepatocellular mitochondrial phosphorylative activity occurs to assist the accelerated metabolism due to phagocytosis.     

Clinical Experience of Postoperative Hepatic Failure Treatment with Pig or Baboon Liver Cross-Hemodialysis with an Interposed Membrane

The present study emphasizes the principle of using liver support to restore the blood ketone body ratio (acetoacetate/β-hydroxybutyrate), which reflects the redox potential of liver mitochondria and correlates with hepatic energy charge (ATP + 0.5ADP/ATP + ADP + AMP). Eleven surgical patients with grade IV hepatic coma were treated by an ex vivo pig or baboon liver cross-hemodialysis with an interposed Cuprophan membrane when their blood ketone body ratios had decreased to below 0.4 compared with the normal of above 0.7. Three patients were treated by cross-hemodialysis using a standard Cuprophan membrane dialyzer without increase of blood ketone body ratio and without marked beneficial effect. However, five of eight patients who had bood ketone body ratios of above 0.25 became fully alert after treatment by cross-hemodialysis using the larger pore size and greater surface area Cuprophan membrane, concurrent with a rise in the decreased blood ketone body ratio, and three of them were later discharged. By contrast, in the three patients with blood ketone body ratios below 0.25, there was no restoration of consciousness and no improvement in their blood ketone body ratios by this liver support. It is suggested that, as long as the blood ketone body ratio remained over 0.25, this metabolic liver support is effective in restoring grade IV hepatic coma.     

Investigation of lipid peroxidation in regenerating rat liver]

The purpose of this study was to estimate the effects of lipid peroxidation in regenerating rat liver. Partial 70% hepatectomy was performed in rat according to Higgings and Anderson. EPC (alpha Toc: Ascorbic acid = 6:4, radical scavenger) was injected intravenously (5mg/kg weight) one hour before operation. Lipid peroxidation in regenerating liver reached a peak at 24 hours after operation. The administration of EPC markedly suppressed the increase of lipid peroxide in the plasma and remnant liver and that of GPT after hepatectomy, with subsequent good liver regeneration ratio. Moreover, the pretreatment with EPC remarkably elevated the activity of thymidine kinase (index of DAN synthesis). The EPC administration had not notable effects on the level of plasma ketone body ratio in animal but pathologically caused early advent of glycogen granule in the remnant liver tissue after partial hepatectomy, which reflected restoration of mitochondrial energy level. The results of the present study suggest that scavenger may be useful not only for impairment of liver dysfunction but also for recovery of mitochondrial energy level and DNA synthesis after liver resection.     

Effects of partial ischemia and reflow on mitochondrial metabolism in rat liver

To clarify the effects of partial ischemia and reflow on the mitochondrial metabolism of the rat liver, the afferent vessels supplying the left lateral and left half of medial lobes were occluded and then reperfused after given time periods of ischemia (30, 60, 90 and 120 min, groups A, B, C and D, respectively). Samplings were taken at 0, 10 and 60 min after reperfusion. The energy charge levels of ischemic lobes decreased rapidly from 0.85 +/- 0.01 in the sham group to 0.38 +/- 0.11, 0.35 +/- 0.07 and 0.34 +/- 0.06 in groups B, C and D, respectively. The phosphorylative activities of mitochondria isolated from ischemic lobes decreased gradually along with the time of ischemia. The reversal of mitochondrial function and energy charge levels following reperfusion was noted in groups A and B. In nonischemic lobes, the phosphorylation rate (nmol ATP/mg/min) increased from 90 +/- 6 in sham group to 125 +/- 12 and 130 +/- 9; 131 +/- 5 and 130 +/- 6; 123 +/- 6 and 122 +/- 17, and 138 +/- 6 and 138 +/- 13 at 10 and 60 min after reflow in groups A, B, C and D, respectively (p less than 0.05). The energy charge level of nonischemic lobes decreased from 0.85 +/- 0.01 of sham group to 0.80 +/- 0.03 in group D (p less than 0.05). From these results, it is concluded that the transitional zone for the reversal of mitochondria function and energy metabolism following prolonged liver ischemia appears at around 60 min.(ABSTRACT TRUNCATED AT 250 WORDS)     

Impaired hepatic energy metabolism in rat acute pancreatitis: protective effects of prostaglandin E2 and synthetic protease inhibitor ONO 3307.

To evaluate the effects of acute pancreatitis on the energy metabolism of the liver and on the fragility of hepatic cells and subcellular organelles, we studies (1) the arterial blood ketone body ratio (BKBR) (aceto acetate/beta-hydroxy butyrate), which is in equilibrium with the free NAD+/NADH ratio in liver mitochondria; (2) the hepatic energy charge (EC) = (ATP + 1/2 ADP)/(ATP + ADP + AMP); (3) the cathepsin B leakage from hepatic lysosomes and the malate dehydrogenase leakage from hepatic mitochondria in vitro; and (4) the protective effects of prostaglandin E2 (PGE2) and a new synthetic protease inhibitor ONO 3307 on hepatic injury in acute pancreatitis induced in rats by a supramaximal dose of caerulein. Decreased BKBR and hepatic EC as well as increased hepatic lysosomal and mitochondrial fragility were observed in rats with this type of acute pancreatitis, and both PGE2 and ONO 3307 had a significant protective effect against hepatic injury in these rats, especially ONO 3307. These results suggest that impaired hepatic energy metabolism is closely related to increased hepatic lysosomal and mitochondrial fragility and that some proteases, which are derived from pancreatitis and are susceptible to inhibition by ONO 3307, seem to play an important pathological role in this liver injury induced by pancreatitis. Therefore, it is important to take care of the liver in patients with acute pancreatitis.