Effect of hemodiabsorption and sorbent-based pheresis on amino acid levels in hepatic failure

Changes in plasma amino acid concentrations were measured in patients with hepatic failure during extracorporeal hemodiabsorption (using the Liver Dialysis Unit, "the Unit") or hemodiabsorption plus sorbent-based pheresis treatment (using the Liver Dialysis Plasmafilter Unit, "the PF-Unit") Systems. Eight patients with hepatic failure, grade 3 or 4 encephalopathy, elevated bilirubin and/or creatinine levels and respiratory or renal failure were treated for 1-3 days with the Unit alone. Three of these were also treated with the Unit containing 10 g of BCAA in the sorbent suspension. Four patients with hepatic failure treated with the PF Unit also had 10 g of branched chain amino acid (BCAA) added to the sorbents of the Unit portion of this device. Pre- and post-plasma samples were drawn and high performance liquid chromatography (HPLC) was used to separate and detect amino acids in the plasma. Both the Unit and the PF-Unit have the capability to selectively remove various amino acids, especially aromatic amino acids (AAA). The pre-treatment amino acid profiles of plasma were typical for hepatic failure, with abnormally high levels of phenylalanine, tyrosine, tryptophan, and methionine and decreased levels of valine, leucine and isbolucine. The average pre-treatment Fischer ratio (BCAA/AAA) for both Unit and PF-Unit patients was 1.43 (+/- 0.58). Treatments by both systems resulted in an increase of BCAA levels in blood and concomitant decrease of AAA levels, with an average Fischer ratio improvement of 30-38% for the Unit and PF-Unit without BCAA. The Fischer ratio improved by 90% (average) for the Unit with BCAA. Levels of many other amino acids (such as alanine, glycine, proline or lysine) increased during both Unit and PF-Unit treatments. The removal of strongly protein-bound toxin and amino acids such as tryptophan and sulphydryl amino acids was more effective by the PF-Unit. Both the Unit and the PF-Unit have the unique capability to remove toxic aromatic amino acids while increasing BCAA levels in patient. The increase in many amino acid levels may be related to the removal of toxins that interfere with normal amino acid metabolism. The addition of the PF module improves the removal of bilirubin and similarly protein-bound chemicals. Changes in amino acid profiles by the Unit and the PF-Unit contrast markedly with other extracorporeal devices.     

A controlled study of sorbent suspension dialysis in chronic liver disease and hepatic encephalopathy

To investigate the role of extracorporeal detoxification in cirrhotic patients with advanced hepatic encephalopathy not responding to medical treatment, 20 patients were randomized to receive six hours of additional sorbent dialysis or ongoing standardized medical treatment. Following treatment, the clinical stage of encephalopathy remained unchanged in both groups. Abnormal sensory evoked potentials improved following sorbent dialysis (N70 latency, 128 ms before versus 110 ms after treatment, P<0,05; cervico-cranial transmission, 7.7 ms versus 6.8 ms, P<0.01) indicating improvement in important aspects of cerebral function. In contrast, brain function remained unchanged following medical treatment (N70 latency, 114 ms versus 113 ms; cervico-cranial transmission, 7.7 ms versus 7.2 ms, P=NS, respectively). Serum benzodiazepine levels decreased significantly after sorbent dialysis but not after medical treatment. Biocompatibility of sorbent dialysis was limited and clinical complications occurred in a proportion of patients. In conclusion, a six-hour treatment with sorbent suspension dialysis did not ameliorate the clinical stage of HE but improved neurophysiologic function in cirrhotic patients who had not responded to conventional medical treatment.     

暴发性肝损伤肝性脑病大鼠血浆及脑游离氨基酸变化的相关分析

本文测定了D-氨基半乳糖所致暴发性肝损伤肝性脑病大鼠血浆及脑勺浆游离氨基酸含量的变化,并作相应的相关分析。结果表明,正常大鼠血浆及脑内各种氨基酸含量间均无相关。肝性脑病大鼠血浆及脑勺浆多种氨基酸水平均有增高,脑内大多数氨基酸含量与血浆相应氨基酸的变化无相关关系,但一些与肝性脑病发生相关的氨基酸水平与其相应的血浆含量呈显著相关。提示肝性脑病的发生与血脑屏障氨基酸转运功能选择性改变有关。     

内毒素对实验性肝硬化大鼠血浆氨基酸系列的影响

实验性肝硬化大鼠血浆氨基酸系列改变表现为支／芳比值降低，但无肝性脑病表现，腹腔注射对正常大鼠无明显影响的微量内毒素，肝硬化大鼠即出现嗜睡与错迷症状，肝细胞损伤加重，氨基酸分析表明血浆谷氨酰胺、牛磺酸水平显著高于肝硬化组，酪氨酸水平与肝硬化组比较显著降低，提示内毒素能加剧肝硬化大鼠血浆氨基酸失衡，为肝性脑病重要的诱发因素。     

Plasma catecholamines in hepatic coma and liver cirrhosis: Role of octopamine

Summary Plasma levels of adrenaline, noradrenaline and octopamine were estimated by a radioenzymatic method in nine cirrhotic outpatients with encephalopathy and in ten patients with hepatic coma (coma grade III–IV). In the cirrhotic outpatients normal as well as elevated plasma levels of noradrenaline were found. Octopamine could not be detected in the plasma of these patients as well as of ten healthy volunteers. Elevated noradrenaline levels were present in all patients with hepatic coma. Plasma noradrenaline remained elevated or even further increased during the course of hepatic coma, whereas adrenaline was elevated less frequently. In eight of the ten patients with hepatic coma octopamine was again not detectable in plasma. Only in two patients high levels of octopamine up to 59.5 ng/ml could be found in addition to increased noradrenaline concentrations. The infusion of the branched chain amino acid L-valine had no influence on the plasma level of either noradrenaline or octopamine.The data indicate that the sympathetic nervous system is activated during the course of hepatic coma. An accumulation of octopamine is not a common finding in chronic liver disease and hepatic coma. Since in the two patients with elevated octopamine levels the rise in octopamine occured concomitantly with a rise in noradrenaline, a displacement of noradrenaline by the false neurotransmitter octopamine in the noradrenergic neuron of the peripheral sympathetic nervous system seems unlikely. The results indicate that the development of hypotension in the course of liver cirrhosis and hepatic coma cannot be related to a deficiency of noradrenaline.Deeply moved we have to inform the readers about the sudden death of our colleague and teacher Professor Dr. F. Wewalka     

Hepatic encephalopathy and cerebral blood flow improved by liver dialysis treatment

Eight acute liver failure patients, all in grade IV hepatic encephalopathy, were administered liver dialysis treatment with the Hemo Therapies Unit (Hemo Therapies Inc, San Diego, CA, USA). The patients were evaluated to determine whether the Glasgow Coma Scale score and cerebral blood flow improved with treatment. After the initial treatment, consciousness levels as measured by the Glasgow Coma Scale improved from a pre-treatment median of 5 (range 3 to 6) to a post-treatment median of 7 (range 5 to 9) (p=0.0005 by paired Wilcoxon test); mean blood flow velocity in the middle cerebral arteries as shown by transcranial Doppler sonography increased from a median of 37.85 cm/sec (range 20.3 to 114.0) to 57.90 (32.5 to 135.0) post-treatment (p=0.022); however, there was no significant change in the pulsatility index from a median of 1.18 (range 0.61 to 1.71) to 0.85 (range 0.70 to approximately 1.79) post-treatment (p=0.13). The 8 patients received 2 to 7 (median 5.5) times of daily 6-h liver dialysis treatments. Following the completion of all liver dialysis treatments, hepatic coma was fully resolved in 4 of 8 patients (50%) Three of 8 patients (37.5%) survived to hospital discharge, whereas 5 patients did not survive due to irreversible liver function and associated complications. In conclusion, liver dialysis treatment could improve hepatic encephalopathy, but the prognosis still depended on the underlying diseases.     

Amino acid disturbances in experimental hepatic coma rats

In our previous paper (1), we discussed the in-vitro adsorption spectrum of plasma amino acids by coated charcoal hemoperfusion. We now describe the general amino acid disturbances in plasma, CSF and cerebrum in the galactosamine induced hepatic coma model in rats. All aromatic amino acids (AAA) increased significantly in the three compartments studied. Branched chain amino acids (BCAA) were either elevated to a much lesser extent or remained unchanged. The molar ratio of BCAA:AAA was consequently reduced. This is similar to findings in fulminant hepatic failure in human and other animal models. Most of the 20 amino acids analyzed were increased significantly in the plasma. Increases of amino acids were minimal in the brain tissue studied, i.e. the cerebrum.     

Hepatic encephalopathy. Pathogenesis and therapy

Hepatic encephalopathy is a frequent complication of cirrhosis. Portal-systemic shunts and depression of hepatic function are the primary underlying abnormalities. Arterial blood ammonia levels are frequently elevated during hepatic encephalopathy and are lower when a clinical improvement is established. Glutamine synthesis is part of the metabolic pathway for ammonia cerebral detoxification that induces ATP and glutamate (excitatory neurotransmitter) depletion. Plasma levels of branched chain amino acids are reduced in patients with cirrhosis, this event allows aromatic amino acids to cross the hemato-liquoral barrier through exchange with glutamine. Cerebral excess of aromatic amino acids promotes the synthesis of octopamine and feniletiletanolamine, weak neurotransmitters. Benzodiazepine-like substances may affect GABA-ergic transmission by interacting with their receptors on the GABA-benzodiazepine complex. Therapy is aimed at controlling the events that may precipitate the acute encephalopathy, at reducing the ammonia levels, and correcting the neurotransmission abnormalities.     

Transition of branched-chain amino acids and tyrosine ratio (BTR) in the blood of acute hepatitis patients

The molar ratio of branched-chain amino acids to tyrosine (BTR) correlates well with the Fischer ratio, and can be measured in a short period of time. It is regarded as the method of analysis that will eventually replace the Fischer ratio. But clinical significance of BTR in terms of acute liver disorders has not been examined thoroughly as of yet. In this study, we measured BTR of 34 patients with acute hepatitis, and examined the transition of the acute period of acute hepatitis and its recovery process. Thirty-four patients diagnosed with acute viral hepatitis became subjects of examination (16 patients of A type, 15 patients of B type, 1 patient of C type, 2 patients of non-A, non-B, non-C type). Out of the 34 patients, 11 were in serious stages (HPT under 40%), including 3 in fulminant condition. By using preserved serum obtained during the acute period (within 1 week of the highest transaminase value), recovery period (within 4 weeks), and treatment period (3 months and later), measurements were conducted with Diacolor:BTR (enzymatic analysis, ONO Pharmaceutical Co., Ltd.), and the results were compared with those of 50 healthy subjects (25 men, 25 women). BTR correlated well with the Fischer ratio for chronic hepatic patients, and with albumin (Alb), PT, and ICGR15 as well, proving that it is useful as an indicator of hepatic reserve ability. But BTR has not been thoroughly examined as it relates to acute liver disorders. In this study, BTR fell in the acute period, correlating with the serious period, proving that it is a useful indicator. For acute liver damage, BTR supports conventional indicators (Alb, Ch-E, HGF, etc.) for assessing serious damage. Also, it has been suggested that measuring the passage of BTR could be the indicator of true recovery, including amino acid metabolism for liver disorders.     

Effect of charcoal hemoperfusion on amino acid disturbance in experimental hepatic coma

Using the galactosamine induced hepatic coma rat model, we studied the effects of coated charcoal hemoperfusion on amino acids in plasma and CSF in grade III hepatic coma. We found that 1 hour of hemoperfusion significantly reduced AAAs and BCAAs in plasma and increased the molar ratio of BCAA:AAA. When rats in grade III coma were treated with 2 consecutive 1-hour hemoperfusions, the substantial reduction of plasma AAAs resulted in a significant decrease of AAA levels in CSF.     

Effect of alternative pathway therapy on branched chain amino acid metabolism in urea cycle disorder patients

Urea cycle disorders (UCDs) are a group of inborn errors of hepatic metabolism caused by the loss of enzymatic activities that mediate the transfer of nitrogen from ammonia to urea. These disorders often result in life-threatening hyperammonemia and hyperglutaminemia. A combination of sodium phenylbutyrate and sodium phenylacetate/benzoate is used in the clinical management of children with urea cycle defects as a glutamine trap, diverting nitrogen from urea synthesis to alternatives routes of excretion. We have observed that patients treated with these compounds have selective branched chain amino acid (BCAA) deficiency despite adequate dietary protein intake. However, the direct effect of alternative therapy on the steady state levels of plasma branched chain amino acids has not been well characterized. We have measured steady state plasma branched chain and other essential non-branched chain amino acids in control subjects, untreated ornithine transcarbamylase deficiency females and treated null activity urea cycle disorder patients in the fed steady state during the course of stable isotope studies. Steady-state leucine levels were noted to be significantly lower in treated urea cycle disorder patients when compared to either untreated ornithine transcarbamylase deficiency females or control subjects (P<0.0001). This effect was reproduced in control subjects who had depressed leucine levels when treated with sodium phenylacetate/benzoate (P<0.0001). Our studies suggest that this therapeutic modality has a substantial impact on the metabolism of branched chain amino acids in urea cycle disorder patients. These findings suggest that better titration of protein restriction could be achieved with branched chain amino acid supplementation in patients with UCDs who are on alternative route therapy.     

肝性脑病患者血清中IL-6和IL-18的表达及意义

目的 检测肝性脑病患者血清中IL-6和IL-18的表达水平并探讨其与天冬氨酸转移酶（AST）、丙氨酸转氨酶（ALT）、总胆红素（TBil）、白蛋白（ALB）、肌酐（Cr）及血浆氨的相关性.方法 40例肝硬化合并肝性脑病的住院患者,定为A组,其中临床分期为Ⅲ～Ⅳ期的18例患者定为A1组,Ⅰ～Ⅱ期的22例患者定为A2组;20例肝硬化患者定为B组;20名健康人作为正常对照,定为C组.用酶联免疫吸附法（ELISA）检测所有研究对象血清中IL-6和IL-18的浓度,并分析其与AST、ALT、TBil、ALB、Cr及血氨的相关性.结果 A1组和A2组IL-6和IL-18浓度均明显高于B组和C组,差异有统计学意义（P＜0.05）;血清IL-6和IL-18水平均与血氨浓度呈正相关（P＜0.05）,与AST、ALT、TBil、ALB及Cr均无明显相关性（P＞0.05）.结论 肝性脑病患者血清中IL-6和IL-18均明显增高而且与血氨存在相关性,IL-6和IL-18可能与血氨存在协同作用,共同参与了肝性脑病的发病.     

The clinical usefulness of the molar ratio of branched-chain amino acids to tyrosine (BTR) in discriminating stage of chronic liver diseases]

We determined the molar ratio of branched-chain amino acids to tyrosine (BTR) in plasma and in serum by enzymatic method and compared it with Fischer ratio (the molar ratio of branched-chain amino acids to tyrosine and phenylalanine) in plasma obtained by conventional HPLC method. BTR in plasma and in serum was well correlated with plasma Fischer ratio. The normal range (mean +/- 2SD) of BTR was determined to be 4.41-10.05 in 210 normal subjects. In addition, we investigated the distribution of BTR values in patients with various liver diseases. BTR value decreased according to the severity of liver disease. We evaluated the clinical usefulness of BTR in patients with chronic liver diseases by cumulative distribution analysis (CDA) graph and receiver operating characteristic curve (ROC) analysis. The area under the curve for BTR analyzed by ROC for CH versus LC.HCC group was the highest (86.3%) of any for various concurrently-measured liver function tests, and was significantly higher than AST/ALT, ALT, AST, gamma-GT (each, p less than 0.001) and ALB (p less than 0.05). These diagnostic results showed that BTR is a superior indicator in discriminating between liver cirrhosis and chronic hepatitis.     

Tryptophan and tyrosine ratios to neutral amino acids in endogenous depression. Relation to antidepressant response to amitriptyline and lithium + L-tryptophan

The plasma ratios of tryptophan and tyrosine to those amino acids that compete with them during transport across the blood-brain barrier have been determined in depressed patients before and after treatment for four weeks with amitriptyline or lithium + L-tryptophan. There was no relation between the absolute plasma concentrations of free or total tryptophan or tyrosine and the clinical response to amitriptyline. There was also no relation between pre-treatment ratio of plasma tyrosine to competing amino acids and response to amitriptyline, but depressives with subnormal tryptophan ratio improved significantly more than patients with supernormal tryptophan ratio with comparable serum drug levels. The therapeutic response to lithium + L-tryptophan was predicted neither by the absolute plasma concentrations of free or total tryptophan or tyrosine nor by the tyrosine ratio, but there was also a trend towards greater improvement in patients with subnormal compared with supernormal tryptophan ratio. The results suggest that the pre-treatment plasma ratio of tryptophan to competing amino acids is a useful predictor of clinical response to amitriptyline. The possible mode of action of amitriptyline and lithium + L-tryptophan is briefly discussed.     

Plasma tyrosine/neutral amino acid ratio correlated with clinical response to nortriptyline in endogenously depressed patients

Ratios in plasma of tryptophan (Trp) and tyrosine (Tyr) to other large neutral amino acids were determined in 26 endogenous depressives before and after treatment with nortriptyline in doses adequate to achieve a steady-state serum level between 70 and 130 ng/ml, i.e., within the recommended therapeutic range. Pretreatment plasma Trp ratio and Tyr ratio were normal and did not change significantly during treatment. The plasma Trp and Tyr concentrations and the plasma Trp ratio showed no significant association with the therapeutic response. However, the pretreatment plasma Tyr ratio correlated significantly and directly with the final Hamilton rating score, and inversely with the per cent reduction of Hamilton rating score. Moreover, depressives with plasma Tyr ratio below the normal mean showed significantly greater clinical improvement than patients with higher plasma Tyr ratio with comparable serum nortriptyline levels. Evidence has been presented that biochemical variables in depressed patients are important determinants of clinical improvement following pharmacotherapeutic treatment. Moreover, the results suggest that the plasma Tyr ratio may be a guideline for antidepressant response to nortriptyline.     

Glutamine depletion in patients with Crimean-Congo hemorrhagic fever

Crimean-Congo hemorrhagic fever (CCHF) is a viral disease. There is not enough knowledge about plasma amino acid levels in CCHF. Therefore, we investigated plasma amino acid levels in patients with CCHF and the association between the levels of these amino acids and disease severity. The plasma amino acid levels (including glutamate [Glu], aspartate [Asp], glutamine [Gln], asparagine [Asn] and gamma-aminobutyric acid [GABA]) in CCHF patients and controls were measured by using liquid chromatography-mass spectrometry. Plasma levels of Gln were lower while Asp, Glu, and GABA levels were higher in patients. In fatal CCHF patients, we found the plasma level of Asn was increased whereas the plasma level of GABA was decreased. This study is the first in the literature to evaluate the plasma Gln, Glu, Asn, Asp, and GABA levels in CCHF patients. We found that the plasma Gln levels were significantly lower in CCHF patients while Asp, Glu, and GABA levels were elevated. Considering that these amino acids are important for immune cells, the plasma amino acid levels of CCHF patients may contribute to the understanding of the pathophysiology of disease and it can be important for supportive treatment of CCHF.     

The cytotoxicity of plasma from patients with acute hepatic failure to isolated rabbit hepatocytes.

The cytotoxicity of plasma from patients with various types of liver disease to rabbit hepatocytes maintained in vitro has been investigated using a microcytotoxicity assay system. Plasma from patients with fulminant hepatic failure and uncomplicated viral hepatitis showed significant cytotoxicity compared to controls. The cytotoxic effect of plasma from patients with fulminant hepatic failure was reduced by charcoal haemoperfusion, or heating and dialysis. The bile acids chenodeoxycholic acid and lithocholic acid were cytotoxic when added to control human plasma. Such toxic factors may be responsible for the delay in liver regeneration often seen in patients with fulminant hepatic failure.     

Phenylbutyrate exerts adverse effects on liver regeneration and amino acid concentrations in partially hepatectomized rats

Phenylbutyrate is recommended in urea cycle disorders and liver injury to enhance nitrogen disposal by the urine. However, hypothetically there may be adverse responses to the use of phenylbutyrate in the treatment of liver disease because of its role as a histone deacetylase inhibitor and its stimulatory effect on branched‐chain alpha‐keto acid dehydrogenase, the rate‐limiting enzyme in the catabolism of branched‐chain amino acids (BCAA; valine, leucine and isoleucine). We report the effects of phenylbutyrate on liver regeneration and amino acid levels in plasma of partially hepatectomized (PH) rats. Phenylbutyrate or saline was administered at 12‐h intervals to PH or laparotomized rats. Phenylbutyrate delayed the onset of liver regeneration compared to the saline‐treated controls, as indicated by lower hepatic DNA specific activities 18 and 24 h post‐PH, decreased hepatic fractional protein synthesis rates 24 h post‐PH and lowered the increases in liver weights and hepatic protein and DNA contents 48 h after PH. Hepatic DNA fragmentation (a hallmark of apoptosis) was higher in the phenylbutyrate‐treated animals than in controls. Phenylbutyrate decreased the glutamine and BCAA concentrations and the ratio of the BCAA to aromatic amino acids (phenylalanine and tyrosine) in the blood plasma in both hepatectomized and laparotomized animals. In conclusion, the delayed onset of liver regeneration and the decrease in BCAA/AAA ratio in blood suggest that phenylbutyrate administration may be disastrous in subjects with acute hepatic injury and BCAA supplementation is needed when phenylbutyrate is used therapeutically.     

Phenylbutyrate therapy for maple syrup urine disease

Therapy with sodium phenylacetate/benzoate or sodium phenylbutyrate in urea cycle disorder patients has been associated with a selective reduction in branched-chain amino acids (BCAA) in spite of adequate dietary protein intake. Based on this clinical observation, we investigated the potential of phenylbutyrate treatment to lower BCAA and their corresponding α-keto acids (BCKA) in patients with classic and variant late-onset forms of maple syrup urine disease (MSUD). We also performed in vitro and in vivo experiments to elucidate the mechanism for this effect. We found that BCAA and BCKA are both significantly reduced following phenylbutyrate therapy in control subjects and in patients with late-onset, intermediate MSUD. In vitro treatment with phenylbutyrate of control fibroblasts and lymphoblasts resulted in an increase in the residual enzyme activity, while treatment of MSUD cells resulted in the variable response which did not simply predict the biochemical response in the patients. In vivo phenylbutyrate increases the proportion of active hepatic enzyme and unphosphorylated form over the inactive phosphorylated form of the E1α subunit of the branched-chain α-keto acid dehydrogenase complex (BCKDC). Using recombinant enzymes, we show that phenylbutyrate prevents phosphorylation of E1α by inhibition of the BCKDC kinase to activate BCKDC overall activity, providing a molecular explanation for the effect of phenylbutyrate in a subset of MSUD patients. Phenylbutyrate treatment may be a valuable treatment for reducing the plasma levels of neurotoxic BCAA and their corresponding BCKA in a subset of MSUD patients and studies of its long-term efficacy are indicated.