Effect of glucose, fructose, sorbitol and amino acid solutions employed in clinical medicine on the development of liver regeneration after partial hepatectomy

The authors studied the effect of glucose (G), fructose (F), sorbitol (So), 4% Nutramin S Spofa (NuS) and Nutramin C Spofa (NuC) on development of liver regeneration. NuS is used clinically to improve a negative nitrogen balance, while the amino acid composition of NuC has a beneficial effect on the ornithine cycle and thus helps to normalize the NH3 level in patients with liver failure. Glucose seems to be the best substance for an optimal onset of liver regeneration. Its onset in rats given NuC was slower than in animals given NuS, but 48 h after partial hepatectomy there was no difference between NuC and NuS as regards the total DNA content and 96 h after the operation there was no difference as regards either the number of mitoses or the number of liver cells. In addition to reducing the plasma ammonia level, therefore, NuC also has a favourable effect on the development on liver regeneration.     

Effect of the parenteral administration of amino acid solutions in different phases after partial hepatectomy on the initiation and development of liver regeneration in rats

Amino acid solutions enriched with branched-chain amino acids or pure branched-chain amino acid solutions were administered parenterally to female laboratory rats (pre-operative weight 230 +/- 30 g) which had been subjected to 65-70% partial hepatectomy (PH), and specific liver DNA activity, the hepatocyte mitotic index and other indicators of the initiation of liver regeneration were studied. Both solutions were infused in an hourly dose of 3.3 ml/kg body weight, during the following postoperative intervals: 1-6, 7-12, 1-12, 1-18 and 1-24 hours. The control rats continued to be fed on the standard laboratory diet after the operation. The results show that the infusion of an amino acid solution enriched with branched-chain amino acids had an inhibitory effect on the onset of DNA synthesis in the liver 18 hours after partial hepatectomy whatever the administration interval. The situation in the case of pure branched-chain amino acid solutions was the same. Twenty-four hours after PH, neither type of solution, irrespective of the infusion interval, was followed by an increase in DNA synthesis compared with the controls fed on the standard laboratory diet. Neither the hepatocyte mitotic index, nor the total liver DNA concentration, showed any changes indicative of stimulation of the initiation of liver regeneration. An infusion stress effect, evaluated from the decrease in the weight of the thymus, was found chiefly in the case of infusions lasting 12 h or longer.     

Acceleration of the onset of liver regeneration by carnitine in partially hepatectomized rats

Immediately and 6 h after removal of 70% of the liver tissue, rats were treated with L-carnitine (Carnitene, Sigma-Tau, Italy) and received an injection of 100, 200 or 1,000 mg/kg b.w. into their femoral vein. The control rats were given the same volume of saline solution. The rats were sacrificed 18, 21, 24 or 30 h after the operation. The development of liver regeneration was evaluated from the incorporation of 14C-thymidine into DNA and from the hepatocyte mitiotic activity. In rats given carnitine in a dose of 100 or 200 mg/kg b.w. significantly higher DNA specific activity values were found 18 and 21 h after partial hepatectomoy and higher hepatocyte mitotic activity values after 30 h. In rats given carnitine in a dose of 1,000 mg/kg b.w., DNA specific activity values 21 h after partial hepatectomy were lower than in the control group. We conclude that L-carnitine, in a dose of 100 or 200 mg/kg b.w. has an enhancing effect on the onset of liver regeneration after 70% hepatectomy.     

Effect of branched chain amino acids on liver regeneration after partial hepatectomy

The authors investigated the effect of the enrichment of commercial amino acid solutions with branched chain amino acids on the development of liver regeneration. Partial (65-70%) hepatectomy was performed on male Wistar rats (140-160 g body weight). Starting with the day of the operation, amino acid solutions normally used in clinical practice and the same solutions enriched with branched chain amino acids were administered by stomach tube; 24, 48 and 96 h after the operation the animals were decapitated. The onset of DNA synthesis was found to be more rapid in animals given the enriched solutions. Once regeneration had started, the stimulant effect of an increased supply of branched chain amino acid on liver regeneration was smaller. Nevertheless, even in the later phase after partial hepatectomy branched chain amino acids had a stronger stimulant effect on liver regeneration after partial hepatectomy than an energy supply in the form of sorbitol.     

Effect of the parenteral administration of energy substrates in different postoperation phases on the initiation and development of liver regeneration in rats subjected to partial hepatectomy

DNA specific activity in the liver, the total DNA content of the liver and the mitotic index of the hepatocytes were studied after the infusion of glucose or lipid emulsions in female laboratory rats with a mean pre-operation weight of 250 +/- 30 g after partial (65-70%) hepatectomy (PH). The infusions were administered in the early prereplication phase (the 1st to 6th hour after the operation), in the late prereplication phase (the 7th to 12th hour after the operation), or continuously from the 1st to the 12th, or the 1st to the 24th, hour after partial hepatectomy. The effect of these parenterally administered energy substrates on the initiation of liver regeneration was evaluated 18 and 24 hours after partial hepatectomy. The results indicate that the infusion of glucose, in any interval after the operation, inhibited the initial phases of liver DNA synthesis (18 h after PH), but not its further development (24 h after PH). Neither the mitotic index of the hepatocytes, nor the total DNA content of the liver differed from the control groups in the case of rats given a glucose infusion. In the experimental groups given lipid emulsions, inhibition of liver DNA synthesis was recorded 18 h after PH only when the infusions were given from the 1st to the 12th or the 1st to the 18th hour after PH. The total DNA content of the liver 18 h after PH was raised in all the experimental groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of α-difluoromethylornithine on polyamine and DNA synthesis in regenerating rat liver: Reversal of inhibition of DNA synthesis by putrescine

The possibility that α-difluoromethylornithine, a specific, irreversible inhibitor of ornithine decarboxylase could be used to prevent the rise in hepatic putrescine and spermidine content following partial hepatectomy was tested. Administration of α-difluoromethylornithine at a dose of 400 mg/kg every 4 h reduced hepatic putrescine to <2 nmol/g, but had only a small effect on the rise in spermidine seen at 28 h after partial hepatectomy. Such treatment also reduced the rise in DNA synthesis produced by partial hepatectomy by up to 70%. The inhibitory effect towards DNA synthesis could be reversed by administration of putrescine which increased the hepatic putrescine content to about 30–40% of that in the regenerating control livers. These results suggest that accumulation of putrescine rather than spermidine is needed for DNA synthesis after partial hepatectomy. They also suggest that part, but not all of the rise in putrescine normally seen in the liver after partial hepatectomy is needed for the enhanced DNA synthesis associated with liver regeneration. Experiments with lower doses of α-difluoromethylornithine showed that a substantial part of the rise in hepatic ornithine decarboxylase activity could be abolished without affecting either the rise in spermidine content or the increase in DNA synthesis after partial hepatectomy.     

Effect of saline solutions administered parenterally in different postoperation phases on the regeneration of rat liver after partial hepatectomy

Two series of experiments were carried out on female laboratory rats with a mean pre-operation weight of 250 +/- 30 g, fed up to the time of the experiment on a standard laboratory diet with water ad libitum. In the first series the rats were subjected to 65-70% partial hepatectomy (PH) or to laparotomy (LAP) and their serum Na+, K+, Cl- and total calcium concentrations were determined 6, 12, 18 and 24 h after the operation. At given postoperation intervals the serum Na+, Cl- and total calcium concentrations in hepatectomized animals were lower than in the intact controls, while the K+ concentration was higher. In the second series of experiments, the rats were given infusions of physiological saline or Ringer solution at different intervals (1-6, 7-12, 1-12 and 1-24 h) after PH. Specific DNA activity in the liver, the hepatocyte mitotic index, the total DNA content of the liver and other indicators show that physiological saline infusions had an inhibitory, or at most a neutral effect on the initiation of liver regeneration, while the effect of the infusion of Ringer solution on the initiation of liver regeneration, in most of the given intervals, was indifferent. The regeneration response depends on the post-PH phase in which the solution is infused.     

Specific inhibition of the synthesis of putrescine and spermidine by 1,3-diaminopropane in rat liver in vivo.

Chronic administration of 1,3-diaminopropane, a compound inhibiting mammalian ornithine decarboxylase (EC 4.1.1.17) in vivo, effectively prevented the large increases in the concentration of putrescine that normally occur during rat liver regeneration. Furthermore, repeated injections of diaminopropane depressed by more than 85% ornithine decarboxylase activity in rat kidney. Administration of diaminopropane 60 min before partial hepatectomy only marginally inhibited ornithine decarboxylase activity at 4 h after the operation. However, when the compound was given at the time of the operation (4 h before death), or any time thereafter, it virtually abolished the enhancement in ornithine decarboxylase activity in regenerating rat liver remnant. An injection of diaminopropane given 30 to 60 min after operation, but not earlier or later, depressed S-adenosyl-L-methionine decarboxylase activity (EC 4.1.1.50) 4 h after partial hepatectomy. Diaminopropane likewise inhibited ornithine decarboxylase activity during later periods of liver regeneration. In contrast to early regeneration, a total inhibition of the enzyme activity was only achieved when the injection was given not earlier than 2 to 3 h before the death of the animals. Diaminopropane also exerted an acute inhibitory effect on adenosylmethionine decarboxylase activity in 28-h regenerating liver whereas it invariably enhanced the activity of tyrosine aminotransferase (EC 2.6.1.5), used as a standard enzyme of short half-life. Treatment of the rats with diaminopropane entirely abolished the stimulation of spermidien synthesis in vivo from [14C]methionine 4 h after partial hepatectomy or after administration of porcine growth hormone. Both partial hepatectomy and the treatment with growth hormone produced a clear stimulation of hepatic RNA synthesis, the extent of which was not altered by injections of diaminopropane in doses sufficient to prevent any enhancement of ornithine decarboxylase activity and spermidine synthesis.     

Effects of carvacrol upon the liver of rats undergoing partial hepatectomy

The present study aims to investigate the possible effects of carvacrol obtained from origanum oil upon the regenerative feature of the liver subsequent to partial hepatectomy in rats.Male Wistar Albino rats, weighing 230±30 g, were divided into three experiment groups. Group I (n=8) were used as sham operation group. Group II (n=8) were applied saline solution and hepatectomy. Carvacrol and hepatectomy (73 mg/kg) were applied to Group III (n=8). One dose of test material was injected 1 h before 68% partial hepatectomy. At the end of the experiments, blood and organs were removed. The liver regeneration rate of the rats was calculated measuring the weights of their liver before and after the hepatectomy. Hematoxylin and eosin, interleukin-6 (IL-6) and proliferating cell nuclear antigen (PCNA) treatments were applied to liver sections. Aspartate transaminase (AST), alanine transaminase (ALT), tumor necrosis factor-alpha (TNF-α) and IL-6 levels were determined in serum samples.The liver regeneration, mitotic index and PCNA index increased significantly in rats of Group III over the Group II at the 72nd hour after partial hepatectomy. Histological evaluations were also similar with these results of PCNA and mitotic indexes. In AST, ALT, TNF-α and IL-6 levels, there was no statistically significant difference.According to these results, it is concluded that carvacrol increases the liver regeneration rate.     

Insulin suppresses PDK-4 expression in skeletal muscle independently of plasma FFA

Starvation and experimental diabetes induce a stable increase in pyruvate dehydrogenase kinase (PDK) activity in skeletal muscle, which is largely due to a selective upregulation of PDK-4 expression. Increased free fatty acid (FFA) level has been suggested to be responsible for the upregulation. Because these metabolic states are also characterized by insulin deficiency, the present study was designed to examine whether insulin has a significant effect to regulate PDK mRNA expression in rat skeletal muscle. In study 1, overnight-fasted rats received an infusion of saline or insulin for 5 h (n = 6 each). During the insulin infusion, plasma glucose was clamped at basal levels (euglycemic hyperinsulinemic clamp). A third group (n = 6) received Intralipid infusion during the clamp to prevent a fall in plasma FFA. PDK-2 mRNA level in gastrocnemius muscle was not altered by insulin or FFA (i.e., Intralipid infusion). In contrast, PDK-4 mRNA level was decreased 72% by insulin (P < 0.05), and Intralipid infusion prevented only 20% of the decrease. PDK-4 protein level was decreased approximately 20% by insulin (P < 0.05), but this effect was not altered by Intralipid infusion. In study 2, overnight-fasted rats were refed or received an infusion of saline or nicotinic acid (NA, 30 micromol/h) for 5 h (n = 5 each). During the refeeding and NA infusion, plasma FFA levels were similarly (i.e., 60-70% vs. saline control) lowered. Muscle PDK-4 mRNA level decreased 77% after the refeeding (P < 0.05) but not after the NA infusion. In conclusion, the present data indicate that insulin had a profound effect to suppress PDK-4 expression in skeletal muscle and that, contrary to previous suggestions, circulating FFA had little impact on PDK-4 mRNA expression, at least within 5 h.     

Hepatic pyruvate metabolism during liver regeneration after partial hepatectomy in the rat

Hepatic pyruvate kinase (PK) and pyruvate dehydrogenase (PDHa) specific activities were decreased after partial hepatectomy or sham operation. The decreases were more marked and sustained after partial hepatectomy. These activity changes ensure that hepatic carbon flux after partial hepatectomy is predominantly in the direction of gluconeogenesis. The decrease in PK specific activity observed after partial hepatectomy was associated with a decreased PK activation ratio (activity measured at 0.15 mM PEP: activity measured at 5.0 mM PEP), and hepatic concentrations of PEP were increased. The low hepatic PDHa activity observed at the first day after partial hepatectomy occurred concomitantly with an increased fatty acid concentration. PDHa activity increased after inhibition of lipolysis. The results indicate that carbohydrate utilization is unimportant for hepatic energy supply during liver regeneration. There was no evidence that the control of PK or PDH in the regenerative liver after partial hepatectomy differed from that observed in the liver of the unoperated rat.     

Specific inhibition of the synthesis of putrescine and spermidine by 1,3-diaminopropane in rat liver in vivo

Chronic administration of 1,3-diaminopropane, a compound inhibiting mammalian ornithine decarboxylase (EC 4.1.1.17) in vivo, effectively prevented the large increases in the concentration of putrescine that normally occur during rat liver regeneration. Furthermore, repeated injections of diaminopropane depressed by more than 85% ornithine decarboxylase activtivity in rat kidney.Adminsitration of diaminopropane 60 min before partial hepatectomy only marginally inhibited orthine decarboxylase activity at 4 h after the operation. However, when the compound was given at the time of the operation (4 h before death), or any time thereafter, it virtually abolished the enhancement in ornithine decarboxylase activity in regenerating rat liver remnant.An injection of diaminopropane given 30 to 60 min after operation, but not earlier or later, depressed $\text{S-}\text{adenosyl}\text{-}\text{l}\text{-}\text{methionine}$ decarboxylase activity (EC 4.1.1.50) 4 h after partial hepatectomy.Diaminopropane likewise inhibited ornithine decarboxylase activity during later periods of liver regeneration. In contrast to early regeneration, a total inhibition of the enzyme activity was only achieved when the injection was given not earlier than 2 to 3 h before the death of the animals.Diaminopropane also exerted an acute inhibitory effect on adenosylmethionine decarboxylase activity in 28-h regenerating liver whereas it invariably enhanced the activity of tyrosine aminotransferase (EC 2.6.1.5), used as a standard enzyme of short half-life.Treatment of the rats with diaminopropane entirely abolished the stimulation of spermidien synthesis in vivo from [¹⁴C] methionine 4 h after hepatectomy or after administration of porcine growth hormone.Both partial hepatectomy and the treatment with growth hormone produced a clear stimulation of hepatic RNA synthesis, the extent of which was not altered by injections of diaminopropane in doses sufficient to prevent any enhancement of ornitine decarboxylase activity and spemedicine synthesis.     

Flavobion and thioctacid lessen irradiation-induced latent injury of the liver]

The effect of hepatoprotective drugs (flavobion and thioctacid) and a single whole-body irradiation (5.7 Gy of gamma radiation) on the regeneration of rat liver was examined. Liver regeneration was estimated on the basis of chosen morphological parameters on hour 30 after partial hepatectomy. Radiation-induced latent injury to intact rat liver 30 min before partial hepatectomy manifested in remaining regenerating liver by slowing-down of the increase in liver weight, cellularity and inhibition of the mitotic activity and in more frequent chromosome aberrations. Both hepatoprotective agents, especially thioctacid, used i.p. 60 min before irradiation, i.e. 90 min before partial hepatectomy, alleviate the manifestations of latent injury in this low proliferating organ as indicated by an increase in cellularity and mitotic index as compared with unprotected animals. Furthermore, the preparations tested decreased the frequency of radiation-induced chromosome aberrations.     

Effect of autonomic denervation on DNA synthesis during liver regeneration after partial hepatectomy

The regulatory role of autonomic nerves in liver regeneration after partial hepatectomy was studied in rats by bilateral subdiaphragmatic vagotomy or splanchnicectomy. 1. In control rats the wet weight of the regenerating liver was restored to approximately 80% of the preoperative weight 72 h after partial hepatectomy. Restoration of the liver weight was significantly impaired in vagotomy rats, but not in splanchnicectomy. Increases in the DNA and protein contents of the regenerating liver were also suppressed by vagotomy. 2. Hepatic DNA synthesis, measured as the incorporation of [methyl-3H]thymidine into DNA at various times after partial hepatectomy, was significantly less in vagotomized rats, and slightly more in splanchnicectomized rats than in control rats. The onset of DNA synthesis triggered by partial hepatectomy was also delayed by vagotomy. 3. The increases in activities of hepatic aspartate transcarbamoylase and thymidine kinase, the key enzymes in synthesis of pyrimidine nucleotides via the de novo and salvage pathways respectively, during liver regeneration, were significantly suppressed and retarded in vagotomized rats. Conversely, splanchnicectomy tended to stimulate these enzyme inductions after partial hepatectomy. 4. During starvation the plasma insulin level decreased after partial hapatectomy in control and vagotomized rats, as in sham-operated rats, but showed a transient increase 6 h after partial hepatectomy in splanchnicectomized rats. It is concluded that vagotomy inhibits and delays DNA synthesis and proliferation of liver cells after partial hepatectomy, whereas splanchnicectomy tends to stimulate these processes. The data also suggest that parasympathetic innervation of the liver may play an important regulatory role in liver regeneration.     

川芎嗪促进大鼠部分肝切除后修复性再生机制的研究

目的:探讨大鼠部分肝切除后肝功能的变化及川芎嗪对肝修复性再生能力的影响。方法:在相同月龄的动物中按体重均衡的原则随机分组。实验动物共分为5组:设正常对照组(对照组)、青年假手术对照组(假术组)、青年肝切除组(青切组)、中年肝切除组(中切组)和中年肝切除治疗组(切治组),每组动物10只,常规饲养,自由饮水。参照Higgins and Aderson给大鼠施行肝脏70%切除手术,中切组大鼠术前以川芎嗪(200 mg/kg/d)腹腔注射7d,其余组注射生理盐水。假术组大鼠以同样的手术程序打开腹腔但不施行肝部分切除术。各组施行手术动物在切除术后24 h沿腹中线切开动物腹腔,于腹主动脉两髂分支处取血分离血清,切取所有肝脏,待测。采用试剂盒法分别测定各组血清中丙氨转氨酶(ALT)、谷草转氨酶(AST)含量;肝脏匀浆后,采用八木国夫法检测肝组织中丙二醛(MDA)含量,采用western blot法测定肝组织中核增殖抗原(PCNA)和铜锌超氧化物歧化酶(SOD1)、锰超氧化物歧化酶(SOD2)的蛋白表达。结果:与中切组相比手术动物相比,切治组组大鼠血清中ALT、AST水平显著降低(P<0.05),肝细胞中PCNA表达显著升高(P<0.05),肝组织中MDA含量显著降低(P<0.05);肝组织中SOD1、SOD2表达显著增加。结论:肝脏切除70%后,肝功受损,氧化应激增加,但核增殖能力增强。川芎嗪可以抑制肝切手术导致的氧化应激损伤,促进SOD的表达,抑制MDA的升高,降低ALT、AST水平,提高PCNA的表达。提示中年大鼠肝切除后肝功能受损与氧化应激相关,给予抗氧化药物能够促进肝再生修复能力,青年肝切除手术大鼠肝的修复能力强于中年动物。     

Effect of parenteral administration of carnitine on liver regeneration in partially hepatectomized rats

Male Wistar rats were subjected to 65-70% hepatectomy and either immediately or 18 h after surgery were given a 6-hour infusion containing 3 ml of either Ringer solution or aqua pro injectione alone or with L-carnitine in doses 8 mg (12.4 mumol), 40 mg (62 mumol) and 200 mg (310.2 mumol)/kg b.w. The rats were killed 6, 18, 24 and 30 h after surgery. The changes in the DNA specific activity and in the mitotic activity demonstrate that L-carnitine has a stimulating, dose-dependent effect on liver regeneration. This effect acts both during early post-hepatectomy, the prereplicative period and in the subsequent replicative period.     

Poly(ADP-ribose) polymerase activity in regenerating liver of hypothyroid rats

The role of PARP, a nuclear enzyme involved in DNA synthesis, repair and cell transformation, was studies during liver regeneration in hypothyroid animals. Hypothyroidism was induced by in vivo administration of propylthiouracil. In regenerating euthyroid animals PARP activity is stimulated showing an early and significant increase at 1.5 h with a maximum at 6 h after partial hepatectomy. Such an increase returns to control values within 18 h preceding the onset of DNA synthesis. A markedly different behavior, with respect to euthyroids, has been evidenced in hypothyroid rats. At first, liver PARP level was about 2-fold higher in non regenerating hypothyroid rats with respect to control euthyroids. During regeneration, PTU-treated animals show a net decrease in PARP activity, with a minimum at 6-9 h after partial hepatectomy. The activity returns to control levels within 24 days. The minimum in PARP activity anticipates, also in this case, the onset of DNA synthesis, which exhibits a maximum at 15-18 h. During liver regeneration PARP activity shows modifications related to the beginning of de novo DNA synthesis. Furthermore, these variations in turn undergo the effects of hypothyroidism.     

Activity of glucose-6-phosphatase during liver regeneration]

During hepatic regeneration a drop in the liver glycogen content along with a lower blood glucose level have been observed. These data are difficult to correlate with the rise of blood glucagon and the drop of insulin shown at the same times after partial hepatectomy. Therefore, liver glucose-6-phosphatase activity has been studied at 1.5, 4, 15 and 24 h, since that enzyme is involved in the release of glucose from the cell. 4 and 15 h after partial hepatectomy a remarkable decrease in glucose-6-phosphatase activity is observed. These results are discussed in view of the higher metabolic demand of regenerating liver.