The effect of acute uraemia on gluconeogenesis in isolated perfused rat livers

Abstract. The effect of acute uraemia on glucose and urea formation by isolated perfused livers of fasting rats was investigated. The basal gluconeogenesis following nephrectomy was significantly increased as compared to normal and sham operated controls. Enhanced glucose formation was associated with an increase in both urea synthesis and output of the poorly metabolizable amino acids valine, leucine, and isoleucine. In the presence of a mixture of amino acids (serine, threonine, lysine, glutamic acid, ornithine, and citrulline) all added at near physiological concentrations, the stimulating effect of uraemia on gluconeogenesis was further enhanced. This was paralleled by an increased formation of urea and an increased uptake of the amino acids. It is concluded that acute uraemia may stimulate glucose synthesis by increased substrate supply. This seems to be achieved by at least two different mechanisms, namely increased protein degradation and accelerated amino acid utilization.     

Isozymes of Aspartate Aminotransferase in Rat Liver during Acute Uraemia

Abstract. Total activity of aspartate-aminotransferase (GOT; EC 2.6.1.1) and activities of the cytoplasmic (c-GOT) and mitochondrial (m-GOT) isozymes were measured in rat liver 24 and 48 h after bilateral nephrectomy.
24 h after nephrectomy no significant differences in enzyme activities could be detected between uraemic animals and sham-operated controls. However 48 h after nephthrectomy the total activity of GOT increased significantly. Fractional tissue extraction revealed an elevation of only the cytoplasmic isozyme (c-GOT), due to a selective increase of this enzyme fraction. No significant change could be noticed of the mitochondrial isozyme (m-GOT). These results are discussed with regard to an increased gluconeogenesis in rat liver 48 h after bilateral nephrectomy.     

Possible sites of interaction of acute renal failure with amino acid utilization for gluconeogenesis in isolated perfused rat liver

Experiments were performed to elucidate the mechanisms involved in the enhanced conversion of amino acids to glucose in acute uraemic rats. Increased gluconeogenesis from a mixture of serine, threonine, lysine, glutamate, ornithine and citrulline was confirmed using a non-recirculating perfusion system. Stimulation was concentration dependent, being most pronounced at physiological amino acid concentrations. Stimulation of glucose and urea formation could be mimicked by using serine alone whereas with lactate and pyruvate inhibition of gluconeogenesis was observed. Serine dehydratase activity was significantly elevated following nephrectomy. Further, the uptake of the non-metabolize amino acid alpha-aminoisobutyrate was considerably increased. It is concluded that serine may play a special role as substrate for the additional glucose formation in acute uraemic rats, probably mediated by an activation of serine dehydratase. Acceleration of amino acid transport seems to represent an additional component of stimulation of amino acid utilization in acute uraemia.     

Enzymatic and Metabolic Studies on Carbohydrate and Amino Acid Metabolism in Rat Liver during Acute Uraemia

Abstract. Enzyme activities of the glycolytic, gluconeogenic, and hexose monophosphate pathways were measured in the liver of starved rats 12 and 48 hours after bilateral nephrectomy. Control experiments (sham operated rats) revealed that alterations of enzyme activities were not due to uraemia but to starvation. Alanine-aminotransferase and aspartate- aminotransferase activities, however, were significantly elevated in rat liver 48 hours after nephrectomy when compared with sham operated controls. Concentrations of some of the gluconeogenic intermediates (3-phosphoglyceric acid, pyruvate, phosphoenolpyruvate and glucose-6-phosphate) were significantly higher in the liver of uraemic animals. Amino acid analysis showed an increase in only L-alanine concentration. It is suggested that the elevated content of pyruvate in the liver during acute uraemia is due to an inhibition of pyruvate degradation. Together with the elevated pyruvate concentration the increase in L-alanine could be explained as a consequence of the equilibrium of the alanine-aminotransferase reaction; K_app. of the reaction is not changed by uraemia. Increased activities of the transaminases and the elevated concentrations of the other metabolites measured might indicate that in the liver of nephrectomized rats there is enhanced gluconeogenesis from substrates other than pyruvate.     

New evidence for the role of TNF-alpha in liver ischaemic/reperfusion injury

Background Tumour necrosis factor‐alpha (TNF‐α) plays a key role in causing ischaemia/reperfusion (I/R) injury. I/R also causes activation of xanthine oxidase and dehydrogenase (XDH + XO) system that, via generated free radicals, causes organ damage. We investigated the effect of ischaemia, reperfusion and non‐ischaemic prolonged perfusion (NIP) on TNF‐α and XDH + XO production in an isolated perfused rat liver model. Materials and methods Rat livers underwent 150 min NIP (control group) or two hours of ischaemia followed by reperfusion (I/R group). TNF‐α (TNF‐α mRNA and protein level), XDH + XO production and bile secretion were determined in tissue and effluent at baseline, at 120 min of ischaemia, after 30 min of reperfusion (I/R group) and after 120 and 150 min of prolonged perfusion (control). Results Unexpectedly, neither ischaemia nor reperfusion had any effect on TNF‐α production. TNF‐α in effluent was 11 ± 4·8 pg mL^?1 at baseline, 7 ± 3·2 pg mL^?1 at the end of ischaemia, and 13 ± 5·3 pg mL^?1 after 30 min of reperfusion. NIP, however, caused a significant increase of TNF‐α synthesis and release. TNF‐α effluent level after 120 and 150 min of perfusion was 392 ± 78·7 pg mL^?1 and 408 ± 64·3 pg mL^?1, respectively. TNF‐α mRNA in tissue was also significantly elevated compared to baseline levels (1·31 ± 0·2 P < 0·001 and 1·38 P < 0·002, respectively). Decrease of liver function (expressed by bile secretion) during I/R and NIP was accompanied by significant XDH + XO elevation. Conclusion This is the first evidence that NIP, and not I/R, is the decisive trigger for TNF‐α production. This study leads to a better understanding of pathogenesis of liver I/R and perfusion damage.     

复方虫草精华对心脏缺血/再灌注损伤的保护作用

目的：观察复方虫草精华（Lungfit)对大鼠离体心脏缺血／再灌注损伤是否存在保护作用。方法：Lungfit分小、大两个剂量（160、325mg/kg.d^-1)给大鼠连续灌胃给药10天后，取心脏，用Langendorff装置逆行恒压灌流，左心室内置乳胶水囊和换能器相连，分别测定缺血前后各组心脏的心功能参数值。结果：缺血前，Lungfit灌胃大鼠离体心脏的各项心功能参数值与对照组相比无显著差异（P＞0．05）；缺血30min后的再灌注期间，Lungfit灌饲的大鼠心脏的心率、冠脉流量及心肌舒缩功能的改善明显优于对照组（P＜0．05或P＜0．01）。结论：Lungfit对心肌缺血／再灌注损伤具有保护作用。     

丹参对离体大鼠肝组织保护作用的实验研究

目的：探讨丹参对离体低温保存大鼠肝组织的保护作用及作用机理。方法：供肝切取采用模拟临床肝移植的标准取肝法，离体肝组织置于4℃保护液中。将72只SD大鼠随机分为3组：对照组（A组）32只（再根据不同保存时间分为1、2、3、6h4个亚组，每组各8只）、实验组（B组）32只（再根据不同保存时间分为1、2、3、6h4个亚组，每组各8只）和正常对照组8只（设为保存0h组）。肝脏保存1、2、3、6h后，测定肝细胞内三磷酸腺苷（ATP）等的含量，线粒体Ca^2＋含量及Ca^2＋-ATP酶活性。观察实验组、对照组肝细胞及线粒体形态学改变。结果：（1）实验组肝细胞ATP的含量、Ca^2＋-ATP酶活性、线粒体Ca^2＋含量与对照组比较，差异有显著性（P〈0．05）；（2）光电镜下见随保存时间延长，实验组细胞损伤较对照组轻微。结论：丹参能改善低温保存肝脏的能量代谢；减轻线粒体钙超载，减轻低温保存肝脏线粒体的损害，因此丹参能提高供肝保存质量，可用于供肝保存。     

Studies on Urea Cycle Enzymes in Rat Liver during Acute Uraemia

Abstract. Activities of urea cycle enzymes were measured in the liver of starved rats 12 and 48 h after bilateral nephrectomy. Control experiments (sham-operated, starved rats) revealed that the activities of only two enzymes of the cycle are altered in the uraemic state: argininosuccinic acid synthetase (EC 6. 3. 4. 5.), which is considered to be rate limiting for urea production and carbamyl phosphate synthetase (EC 2. 7. 2. 5.). Alterations in ornithine concentration of the liver, a possible cause of an increased urea production rate, could not be detected previously (21). Our present results do not support the concept that a decrease of the activity of ornithine-6-amino transferase (EC 2. 6. 1. 13), leading to an increase in the ornithine content of the liver is responsible for the accelerated urea production rate in the liver of acute uraemic rats.     

The Mechanism of Action of Mercuric Chloride on the Isolated Perfused Rat Kidney

Abstract. Studies on 80 rat kidneys, perfused at constant flow, showed that mercuric chloride produced a marked increase in perfusion pressure within five minutes of administration. Adrenergic blocking agents (phentolamine and propranolol), angiotensin sensitivity depletion, bradykinin and low concentrations of mannitol had no effect on this increase in resistance. Perfusion of the kidney with 5 % mannitol solution, however, significantly reduced the increase of vascular resistance induced by mercuric chloride. Evidence is presented that mercuric chloride may evoke an increase in vascular resistance by inducing endothelial cell swelling, an action antagonised by the hypertonic effect of mannitol. Other possible contributory mechanisms are also discussed.     

Effect of Nicotinamide on Carbohydrate Metabolism in the Rat Liver during Starvation

Abstract. A single high dose of nicotinamide (500 mg/kg body weight) results in an eight-hour long inhibition of lipolysis in adipose tissue by transformation to nicotinic acid. In the present paper the effect of inhibited lipolysis on carbohydrate and amino acid metabolism after forty-eight hours starvation was examined in vivo. Within two hours after the injection of nicotinamide (Nam) the plasma level of FFA declines from 900 μval/l to 300 μval/l. The FFA level rises again between 8–12 h after Nam injection. The reduced supply of FFA to the liver leads to a reduced oxidation rate, characterized by a fall in the acetyl-CoA content of the liver and of ketone bodies in liver and blood. Simultaneously there is an increase in protein catabolism. This increase is characterized by increased urinary excretion of urea up to 10 h after Nam injection and by a higher urea production rate in liver slices of rats pretreated with Nam. In the liver the levels of the following amino acids rise very sharply SER (100%), CIT (400%), ORN (250%), ALA (300%), α-AB (470%), VAL (350%), LEU (250%), ILE (370%). The content of THR is reduced to 63%, and GLY to 50%. In comparison to the values of the control groups the contents of ASP, GLU, TYR, LYS, HIS and ARG are unchanged. The result of the increased precursor supply for gluconeogenesis is a higher rate of glucose production, characterized by an increase in blood glucose by 40 mg/100 ml, by an increase in liver glycogen content, by an increase in the concentrations of intermediates of the Embden-Meyerhof-pathway (mainly the concentrations of PEP (870%), 3PGA (480%), FDP (150%), G6P (250%) and glucose (220%). In liver slice experiments the glucose production rate from L-ALA rises significantly, compared to the control group, after pretreatment of the animals with Nam. The results show that a stimulation of gluconeogenesis is obtainable in vivo, although the oxidation rate of FFA is reduced. Thus, they support the assumption, that an increased fatty acid oxidation rate is not a primary condition of stimulated gluconeogenesis.     

齐墩果酸预处理对大鼠肝脏缺血再灌注损伤中细胞凋亡的影响

目的：观察齐墩果酸（OA）预处理对肝脏缺血再灌注损伤中细胞凋亡的影响,探讨齐墩果酸预处理对肝脏保护作用的机制。方法雄性 SD 大鼠40只,随机分为假手术组（SH 组）、缺血再灌注组（IR 组）、0．5％羧甲基纤维素钠组（CM 组）和齐墩果酸预处理组（齐墩果酸组）,齐墩果酸组以100 mg/kg 的齐墩果酸混悬液,SH 和 IR 组以相同容积的水,CM 组以相同容积的0．5％CMC-Na 分别每日灌胃1次,连续7 d。第8天建立70％肝脏缺血再灌注模型,肝脏缺血60 min 后行再灌注3 h。检测大鼠血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）和LDH 水平;检测肝组织内丙二醛（MDA）、超氧化物歧化酶（SOD）、谷胱甘肽（GSH）水平变化;通过 HE 染色观察肝组织形态,利用 Tunel 染色检测肝组织凋亡状态;Western blot 检测肝脏组织 caspase3和 p53蛋白表达水平。结果与 IR 组相比齐墩果酸组血清肝酶水平及肝组织中 MDA 水平显著降低,而 SOD 和 GSH 活性则显著升高;与 IR组相比齐墩果酸组肝组织病理形态学损伤明显减轻;缺血再灌注后可见明显的肝实质细胞凋亡现象,齐墩果酸组凋亡细胞较 IR 组显著下降,且 caspase3和 p53蛋白表达亦减少。结论齐墩果酸预处理可减轻肝脏缺血再灌注损伤,可能与抑制 caspase3和 p53蛋白表达从而抑制细胞凋亡有关。     

Acute Renal Failure after Folate: NaKATPase in Isolated Rat Renal Tubule

Abstract. Using refinements of quantitative histochemistry, i.e. oil-well technique, enzymatic Pi analysis and NADP/ NADPH cycling, the activity of NaKATPase (E.C. 3.6.1.3), an enzyme involved in transmembrane ion transport, was measured in single dissected segments of the proximal and distal tubule of male rats 10 min., 30 min., 60 h and 14 days after folate administration (250 mg/kg body weight). 10 min. after injection the onset of acute renal failure was already apparent by an increase in blood urea of 45 per cent and cessation of urine flow. 60 h after folate the rats became polyuric. The kidney wet weight rose by 40 per cent in a few minutes after the injection due to universal tubular dilatation. During the first hours numerous folate casts were localized, mainly in the thick ascending iimb of Henle's loops. In addition precipitates were found intracellularly in the distal part of the proximal convoluted tubules. The straight portion of the proximal tubules was free of precipitates. 10 min. after the injection tubular segments containing folate material did not show Na K ATPase activity, whereas the folate free tubular segments revealed a residual activity of 30 per cent. Only 15 per cent of the tubules within one lyophilized section had collapsed proximal portions with control activity values. However, the less collapsed segments of the distal tubules revealed a loss of Na K ATPase activity of 55 per cent. Thus folate seems to affect the distal tubule first of all. 60 h after folate administration, when kidney growth reached its maximum, the whole nephron lost its Na K ATPase activity. No tubular cell degeneration or necrosis could be detected during the development stage or the sustained phase of renal failure. After addition of folate to renal homogenate, Na K ATPase exhibited activation (+ 35 per cent) and inhibition (–42 per cent and more) of activity plotted against folate concentration. A careful evaluation of the available evidence led to the conclusion that the acute renal failure induced by folate differs markedly from other models by a directly altered tubular cell metabolism in the development stage of oliguria. Acute tubular obstruction may initiate oliguria. However it seems unlikely that tubular obstruction will be the unique pathogenic factor for this syndrome, especially for the development of the tremendous kidney growth. The inhibition of tubular Na K ATPase activity after folate may indicate an impaired tubular active reabsorption capacity which would be involved in the acute renal failure. The most important finding is that glomerular filtration does not seem to be a primary factor in the development of oliguria after folate administration.     

阿魏酸钠与卡托普利对离体大鼠心脏的药理性预适应作用及机制实验研究

目的:研究阿魏酸钠(sodiumferulate,SF)和卡托普利(captopril,CP)诱导药理性预适应(pharmacologicalprecondition)对离体大鼠心脏的保护作用及其机制。方法:将大鼠离体心脏行Langdorff灌流(灌流液为任-乐氏液),并将大鼠随机分为6组:对照组,损伤(I/R)组,缺血预适应(IP)组,SF组,CP组,SF CP组。然后行心功能检查、ATP酶活性测定以及钙含量测定,并行统计分析。结果:SF和CP均能增加冠脉流出量,改善心功能;降低心肌组织钙超载;减少心律失常发生率及严重程度;提高心肌Na -K -ATP酶、Ca2 -ATP酶、Mg2 -ATP酶活性。而二者联合应用后,上述心肌保护作用未得到明显改善。结论:(1)SF和CP均能对离体大鼠心脏起药理性预适应保护作用;(2)SF预适应保护作用机理与卡托普利预适应保护作用机理可能相同;(3)两种药物若应用于临床心肌保护中,不宜联合应用。     

CO2超声造影对鼠肝影响的观察

通过对１２只Ｓｐｒａｇｕｅ－Ｄａｗｌｅｙ大鼠进行经股动脉肝脏ＣＯ２超声造影，并 与对照组比较，结果显示ＣＯ２微泡进入肝脏后。血清谷丙转氨酶升与对照组相比无明显差异；病理切片显示ＣＯ２组中仅２个肝细胞灶性能脂肪变性；电镜仅显示ＣＯ２组肝细胞有脂滴存在，但无明显的细胞器改变。     

STANDING检查在急性孤立性眩晕患者中的应用

目的:调查STANDING检查在急性孤立性眩晕患者中的特异性、敏感性和实用性。方法:收集急性孤立性眩晕患者,完成神经内科常规检查后,采用STANDING方法检查。入院后第2天完善MRI、DWI、眼震电图、冷热水试验,并随访2周,以明确诊断。结果:61例患者完成研究。STANDING检查的敏感性为97.0%,特异性为96.3%,准确性为95.1%,中枢性眩晕预测值为96.7%,周围性眩晕预测值为92.9%。平均检查时间为(10±2)min。结论:STANDING检查有助于提高眩晕的诊疗水平。     

The Role of Cyclic Adenosine Monophosphate in Adrenergic Effects on Ventricular Vulnerability to Fibrillation in the Isolated Perfused Rat Heart

The relation between myocardial tissue cyclic AMP (cAMP) and the vulnerability to ventricular fibrillation was assessed in the isolated perfused rat heart by measurement of ventricular fibrillation threshold (VFT) and vulnerable period duration (VP). Exogenous dibutyryl cyclic AMP (DBcAMP) reduced VFT and increased VP by a concentration-related action whereas exogenous cAMP did not. Theophylline (1.0 mmol/liter) increased the tissue content of cAMP by 58% (P < 0.001) and caused a leftward shift in the concentration-response curve to DBcAMP. An effect of cAMP on VFT and VP could be shown in the presence of phosphodiesterase inhibition by theophylline. beta-1-Adrenergic receptor blockade with atenolol did not alter the concentration-response curve for VFT when DBcAMP was administered. Epinephrine (100 nmol/liter to 1 mumol/liter) also increased vulnerability to VF; this effect was accompanied by a concentration-related increase in tissue cAMP, but inconsistent changes in tissue ATP, phosphocreatine and potassium. The concentration-response curve of VFT to epinephrine was shifted leftward by theophylline and rightward by atenolol.The increases in vulnerability to fibrillation in the isolated perfused rat heart, in response to DBcAMP, theophylline or epinephrine, could be related more closely to changes of tissue cAMP than to effects on tissue high energy phosphates or potassium. The effect of epinephrine and theophylline on vulnerability to ventricular fibrillation is mediated via alterations in the intracellular level of cAMP in the isolated perfused rat heart.     

The Effect of Agmatine on Ischemic and Nonischemic Isolated Rat Heart

OBJECTIVE: the natural polyamines play a protective role during ischemic injury. We studied the effects of agmatine on ischemic and nonischemic isolated rat hearts. METHODS: Thirty-one rats were randomly assigned to one of four experimental groups. Sixteen rats were injected with saline (group 1, n = 9; group 3, n = 7), and 15 rats were injected with 100 mg/kg of agmatine (group 2, n = 8; group 4, n = 7). Injections were given twice: 24 hours and 1 hour before the experiment. Using the modified Langendorf model, rat hearts were perfused with Krebs-Henseleit solution for 105 minutes during phase 1 of the experiment (groups 1 and 2). During phase 2, hearts were exposed to 45 minutes of global ischemia (groups 3 and 4). RESULTS: During phase 1, no statistically significant differences were observed between the agmatine and the control groups. During phase 2, agmatine caused a significant increase in left ventricular pressure (P <.003). At the end of reperfusion, P(max) was 111% +/- 10% from the baseline levels versus only 82% +/- 5% in the control group. After 20 minutes of reperfusion, dP/dt (first-time derivative of the ventricular pressure) in the agmatine group reached full recovery of 106% +/- 12% versus only 64% +/- 14% in the saline group (P =.059). Agmatine also caused a significant increase in coronary flow rate (P <.004) throughout the reperfusion period. Quantitative immunohistochemical staining disclosed reduced cell damage in the agmatine-treated hearts (P <.02) versus the control group. CONCLUSION: Agmatine injection given before induced ischemia improves hemodynamic recovery by mechanisms that may be attributed to its vasodilatory properties.     

Effect of Peritubular Protein Concentration on Reabsorption of Sodium and Water in Isolated Perfused Proximal Tubules

Micropuncture studies have indicated that variation in peritubular oncotic pressure influences net transport of fluid out of the proximal tubule. The present in vitro studies on isolated perfused rabbit proximal convoluted tubules were designed to examine whether protein concentration gradient must act across the peritubular capillary membrane to influence reabsorption, or whether it can exert a direct effect across the tubular basement membrane 71 proximal tubules were perfused with ultrafiltrate made isosmolal to bathing fluids, the latter having identical electrolyte composition as the perfusing ultrafiltrate, but adjusted to three oncotic pressures: hypooncotic, protein 0.0 g/100 ml; control isooncotic serum, protein 6.4 g/100 ml; and hyperoncotic, protein 12.5 g/100 ml. Net volume flux (nl/mm per min), net Na flux (nEq/mm per min), unidirectional Na flux from bath to lumen (nEq/mm per min), and passive permeability coefficient (x 10(-5) cm/sec) for Na (P(Na)), urea (P(urea)), and sucrose (P(sucrose)) were determined using isotopic techniques. When the bath was hypooncotic, there was (as compared with isooncotic serum) a significant decrease in net volume (38%) and net sodium (40%) flux, but no change in P(Na), P(urea), or transtubular potential; however, P(sucrose) increased significantly (78%). In experiments in which hyperoncotic bath was used, there was (compared with isooncotic serum) an increase in net volume (28%) and net sodium (30%) flux, but transtubular potential difference did not change significantly.These data demonstrated that changes in the ambient protein concentration gradient exert direct effects upon proximal tubular reabsorption. Because penetration of sucrose (an index of intercellular movement) but not urea (an index of transcellular movement) varied with changes in tubular reabsorption, it is suggested that oncotic pressure acts by altering the rate of back-leak of reabsorbate through extracellular pathways between tubular cells. It is concluded that an effect of protein concentration on reabsorption can be exerted directly across the basement membrane, without necessary interposition of the capillary bed.     

The Effect of Protein Deficient Diet on Rat Kidneys

In 31 sera from patients with hepatocellular or cholestatic jaundice unconjugated bilirubin was practically normal. In 176 newborn infants with physiologic or haemolytic jaundice and in one patient with constitutional hyperbilirubinaemia unconjugated bilirubin was elevated. Total bilirubin was increased in both groups and was approximately equal to unconjugated bilirubin in the latter group. The method is diagnostically superior to the indirect van den Bergh reaction and to chromatographic analysis in distinguishing unconjugated and conjugated hyperbilirubinaemia.