Plasma amino acid imbalance in alcoholic liver cirrhosis

Plasma amino acid concentrations and plasma glucagon and serum insulin levels were studied in male patients with compensated alcoholic and nonalcoholic liver cirrhosis. Age, nutritional status, and liver function tests were similar in both groups; none of the patients presented hepatic encephalopathy. Plasma valine and leucine concentrations were lower, and tyrosine, higher in alcoholic than nonalcoholic liver cirrhosis. As a result, the molar ratios of branched-chain amino acids (BCAA) to aromatic amino acids (AAA) were reduced markedly in this group. Although correlation coefficients comparing BCAA/AAA ratios and KICG in alcoholic and nonalcoholic liver cirrhosis were similar, a steeper regression line was observed in alcoholics. Plasma glucagon and proline levels were significantly higher in alcoholic than nonalcoholic liver cirrhosis, the former correlated with AAA concentrations only in alcoholic liver cirrhosis, but not with BCAA levels. These results indicated that alcoholic liver cirrhosis presented a more deranged plasma amino acid pattern than nonalcoholic, and the amino acid imbalances, except for depressed BCAA and elevated proline, were derived, in part, from the hyperglucagonemia.     

Plasma amino acids in four models of experimental liver injury in rats

Summary We studied the plasma amino acid profiles in four models of hepatic injury in rats. In partially hepatectomized rats (65% of liver was removed) we observed significant increase of aromatic amino acids (AAA; i.e. tyrosine and phenylalanine), taurine, aspartate, threonine, serine, asparagine, methionine, ornithine and histidine. Branched-chain amino acids (BCAA; i.e. valine, leucine and isoleucine) concentrations were unchanged. In ischemic and carbon tetrachloride acute liver damage we observed extreme elevation of most of amino acids (BCAA included) and very low concentration of arginine. In carbon tetrachloride induced liver cirrhosis we observed increased levels of AAA, aspartate, asparagine, methionine, ornithine and histidine and decrease of BCAA, threonine and cystine. BCAA/AAA ratio decreased significantly in partially hepatectomized and cirrhotic rats and was unchanged in ischemic and acute carbon tetrachloride liver damage. We conclude that a high increase of most of amino acids is characteristic of fulminant hepatic necrosis; decreased BCAA/AAA ratio is characteristic of liver cirrhosis; and decrease of BCAA/AAA ratio may not be used as an indicator of the severity of hepatic parenchymal damage.Abbreviations BCAA branched-chain amino acids (i.e. valine, leucine and isoleucine) - AAA aromatic amino acids (i.e. tyrosine and phenylalanine)     

某些内科系统疾病患者血浆游离氨基酸的变化及意义

为了解某些内科系统疾病的血氨基酸代谢特点。测定１７０例９种内科系统急慢性疾病患者血浆游离氨基酸浓度,并分别与５８例健康人血浆氨基酸浓度作对比分析。９种疾病的血浆胱氨基酸浓度不同程度升高,以尿毒症病人最为显著,炎症性病人血浆苯丙、蛋、赖氨酸和苯丙／酪比值显著升高,丙、组、精、脯氨酸和支／芳比值显著降低;糖尿病和甲亢病人血浆总游离氨基酸、谷、丙、酪、赖和支链氨酸显著升高;糠尿病病人支／芳比值显著升高,表明不同疾病的血氨基酸浓度有特征性变化,炎症是导致血氨基酸代谢紊乱的重要因素     

肝硬化与肝癌患者血浆游离氨基酸水平分析

采用高效液相色谱法分析了２５例肝硬化和１５例肝癌患者空腹血浆游离氨基酸水平的变化。结果表明,二者支链氨基酸（ＢＣＡＡ）如Ｖａｌ、Ｉｌｅ呈下降趋势,而芳香族氨基酸（ＡＡＡ）如Ｔｙｒ、Ｐｈｅ则呈上升趋势,ＢＣＡＡ／ＡＡＡ分子比值下降。丙氨酸（Ａｌａ）、蛋氨酸（Ｍｅｔ）、谷氨酰胺（Ｇｌｎ）及天门冬氨酸（Ａｓｐ）明显上升。其变化趋势二者存在差别。     

各型肝病患者红细胞内氨基酸检测的临床研究

本文对３０例正常人和１０８例各型肝病患者红细胞内氨基酸进行检测,结果显示,各型肝炎与正常人比较红细胞内氨基酸均表现不同程度的增高和降低。如正常人红细胞内１７种氨基酸的总量为３５５４．４７μｍｏｌ／Ｌ,支／芳比值３．０６±０．４０;急性肝炎总量３４２３．２５μｍｏｌ／Ｌ,支／芳比值２．４８±０．３０;慢性肝炎（轻度）总量３３２９．３３μｍｏｌ／Ｌ,支／芳比值２．３９±０．２２;慢性肝炎（中度）总量３２１９．３２μｍｏｌ／Ｌ,支／芳比值１．８３±０．４０;肝硬化总量３７６２．３３μｍｏｌ／Ｌ,支／芳比值１．３６±０．４１。     

Application of cross-linked beta-cyclodextrin polymer for adsorption of aromatic amino acids

Beta-cyclodextrin (beta-CyD) was cross-linked by hexamethylene diisocyanate and the polymer was investigated for adsorption of aromatic amino acids (AAA) from phosphate buffer. High adsorption rates were observed at the beginning and the adsorption equilibrium was then gradually achieved in about 45 min. The adsorption of AAA decreased with the increase of initial concentration and also temperature. Under the same conditions, the adsorption efficiencies of AAA were in the order of L-tryptophan (L-Trp) > L-phenylalanine (L-Phe) > L-tyrosine (L-Tyr). Much higher adsorption values, up to 52.4 and 43.0 mg/g for L-Trp and L-Phe, respectively, at 50 mmol/L and 3.2 mg/g for L-Tyr at 2 mmol/L, were obtained with the beta-CyD polymer at 37 degrees C. It was shown that the adsorption of AAA on the beta-CyD polymer was consistent with the Freundlich isotherm equation. The adsorption of mixed aromatic amino acids and branched-chain amino acids (BCAA) showed that AAA were preferentially adsorbed with adsorption efficiencies 10-24%, while those of BCAA were lower than 2%. It seems that the structure and hydrophobicity of amino acid molecules are responsible for the difference in adsorption, by influencing the strength of interactions between amino acid molecule and the polymer.     

研究荧光标记BCAA及AAA在大鼠体内代谢

使用DNS-Cl标记BCAA及AAA等六种氨基酸,观察这些氨基酸在大鼠小肠的吸收,血液的清除以及肝脏、肌肉、肾脏、脑等器官的摄取及排空情况。结果:小肠在30分钟开始吸收,4小时部分排空;血液在4小时开始清除,16小时清除完毕;肝脏、肌肉、肾脏、脑组织等,都在30分钟全部或部分摄取,但各器官对不同氨基酸,排空的时间不尽相同。     

Encapsulation of rat hepatocyte spheroids for the development of artificial liver

Hepatocyte spheroids and hepatocyte were immobilized in chitosan/alginate capsules formed by the electrostatic interactions between chitosan and alginate. After encapsulation, there was a 10% decrease in the viability of spheroids due to the exposure of the cells to a pH 6 during the encapsulation process. However, the encapsulated hepatocyte spheroids maintained over 50% viability and liver specific functions for 2 weeks while the encapsulated hepatocytes, free hepatocytes and free hepatocyte spheroids showed low viability and liver specific functions. Therefore, encapsulated hepatocyte spheroid might be applied to the development of a bioartificial liver.     

Adipose Tissue Dysfunction and Altered Systemic Amino Acid Metabolism Are Associated with Non-Alcoholic Fatty Liver Disease

Background

Fatty liver is a major cause of obesity-related morbidity and mortality. The aim of this study was to identify early metabolic alterations associated with liver fat accumulation in 50- to 55-year-old men (n = 49) and women (n = 52) with and without NAFLD.

Methods

Hepatic fat content was measured using proton magnetic resonance spectroscopy (¹H MRS). Serum samples were analyzed using a nuclear magnetic resonance (NMR) metabolomics platform. Global gene expression profiles of adipose tissues and skeletal muscle were analyzed using Affymetrix microarrays and quantitative PCR. Muscle protein expression was analyzed by Western blot.

Results

Increased branched-chain amino acid (BCAA), aromatic amino acid (AAA) and orosomucoid were associated with liver fat accumulation already in its early stage, independent of sex, obesity or insulin resistance (p<0.05 for all). Significant down-regulation of BCAA catabolism and fatty acid and energy metabolism was observed in the adipose tissue of the NAFLD group (p<0.001for all), whereas no aberrant gene expression in the skeletal muscle was found. Reduced BCAA catabolic activity was inversely associated with serum BCAA and liver fat content (p<0.05 for all).

Conclusions

Liver fat accumulation, already in its early stage, is associated with increased serum branched-chain and aromatic amino acids. The observed associations of decreased BCAA catabolism activity, mitochondrial energy metabolism and serum BCAA concentration with liver fat content suggest that adipose tissue dysfunction may have a key role in the systemic nature of NAFLD pathogenesis.     

Perfusion of 3D encapsulated hepatocytes—A synergistic effect enhancing long‐term functionality in bioreactors

Long‐term primary cultures of hepatocytes are essential for bioartificial liver (BAL) devices and to reduce and replace animal tests in lead candidate optimization in drug discovery and toxicology tests. The aim of this work was to improve bioreactor cultures of hepatocyte spheroids by adding a more physiological perfusion feeding regime to these bioreactor systems. A continuous perfusion feeding was compared with 50% medium replacement (routinely used for in vitro tests) at the same dilution rate, 0.125 day⁻¹, for three operative weeks. Perfusion feeding led to a 10‐fold improvement in albumin synthesis in bioreactors containing non‐encapsulated hepatocyte spheroids; no significant improvement was observed in phase I drug metabolizing activity. When ultra high viscous alginate encapsulated spheroids were cultured in perfusion, urea synthesis, phase I drug metabolizing activity and oxygen consumption had a threefold improvement over the 50% medium replacement regime; albumin production was the same for both feeding regimes. The effective diffusion of albumin in the alginate capsules was 7.75.10⁻⁹ cm² s⁻¹ and no diffusion limitation for this protein was observed using these alginate capsules under our operational conditions. In conclusion, perfusion feeding coupled with alginate encapsulation of hepatocyte spheroids showed a synergistic effect with a threefold improvement in three independent liver‐specific functions of long‐term hepatocyte spheroid cultures. Biotechnol. Bioeng. 2011; 108:41–49. © 2010 Wiley Periodicals, Inc.     

Bioartificial liver system based on choanoid fluidized bed bioreactor improve the survival time of fulminant hepatic failure pigs

Bioartificial liver (BAL) support system has been proposed as potential treatment method for end-stage liver diseases. We described an improved BAL system based on a choanoid fluidized bed bioreactor containing alginate-chitosan encapsulated primary porcine hepatocytes. The feasibility, safety, and efficiency of this device were estimated using an allogeneic fulminant hepatic failure (FHF) model. FHF was induced with intravenous administration of D-galactosamine. Thirty FHF pigs were divided into three groups: (1) an FHF group which was only given intensive care; (2) a sham BAL group which was treated with the BAL system with empty encapsulation, and (3) a BAL group which was treated with the BAL system containing encapsulated freshly isolated primary porcine hepatocytes. The survival times and biochemical parameters of these animals were measured, and properties of the encapsulations and hepatocytes before and after perfusion were also evaluated. Compared to the two control groups, the BAL-treated group had prolonged the survival time and decreased the blood lactate levels, blood glucose, and amino acids remained stable. No obvious ruptured beads or statistical decline in viability or function of encapsulated hepatocytes were observed. This new fluidized bed BAL system is safe and efficient. It may represent a feasible alternative in the treatment of liver failure.     

Development of a bioartificial liver employing xenogeneic hepatocytes

Liver failure is a major cause of mortality. A bioartificial liver (BAL) employing isolated hepatocytes can potentially provide temporary support for liver failure patients. We have developed a bioartificial liver by entrapping hepatocytes in collagen loaded in the luminal side of a hollow fiber bioreactor. In the first phase of development, liver-specific metabolic activities of biosynthesis, biotransformation and conjugation were demonstrated. Subsequently anhepatic rabbits were used to show that rat hepatocytes continued to function after the BAL was linked to the test animal. For scale-up studies, a canine liver failure model was developed using D-galactosamine overdose. In order to secure a sufficient number of hepatocytes for large animal treatment, a collagenase perfusion protocol was established for harvesting porcine hepatocytes at high yield and viability. An instrumented bioreactor system, which included dissolved oxygen measurement, pH control, flow rate control, an oxygenator and two hollow fiber bioreactors in series, was used for these studies. An improved survival of dogs treated with the BAL was shown over the controls. In anticipated clinical applications, it is desirable to have the liver-specific activities in the BAL as high as possible. To that end, the possibility of employing hepatocyte spheroids was explored. These self-assembled spheroids formed from monolayer culture exhibited higher liver-specific functions and remained viable longer than hepatocytes in a monolayer. To ease the surface requirement for large-scale preparation of hepatocyte spheroids, we succeeded in inducing spheroid formation in stirred tank bioreactors for both rat and porcine hepatocytes. These spheroids formed in stirred tanks were shown to be morphologically and functionally indistinguishable from those formed from a monolayer. Collagen entrapment of these spheroids resulted in sustaining their liver-specific functions at higher levels even longer than those of spheroids maintained in suspension. For use in the BAL, a mixture of spheroids and dispersed hepatocytes was used to ensure a proper degree of collagen gel contraction. This mixture of spheroids and dispersed cells entrapped in the BAL was shown to sustain the high level of liver-specific functions. The possibility of employing such a BAL for improved clinical performance warrants further investigations.     

常用口服营养液中游离氨基酸的测定与分析

我科实验室于９５年对全国市售的８种富含氨基酸成份的口服液进行了游离氨基酸测定,结果显示：１号液总量为８８９０．９８μｍｏｌ／Ｌ,支／芳比值３．６６;２号液总量为４２９５．９６μｍｏｌ／Ｌ,支／芳比值２．９９;３号液总量为１０４４５．８８μｍｏｌ／Ｌ,支／芳比值３．１０;４号液总量为２５８２０．５μｍｏｌ／Ｌ,支／芳比值２．８１;５号液总量为５５３．５μｍｏｌ／Ｌ,支／芳比值３．７０;６号液总量为１１６８．５６μｍｏｌ／Ｌ,支／芳比值１．３８;７号液总量为３３９１．７１μｍｏｌ／Ｌ,支／芳比值２．４９;８号液总量为２８５１．１４μｍｏｌ／Ｌ,支／芳比值０．２２。正常人血清中游离氨基酸总量为２９６２．８μｍｏｌ／Ｌ,支／芳比值３．２７±０．５８。     

Blood amino acids in chronic HBsAg positive liver disease

Typical changes in blood aminoacid concentrations have been described in patients with severe liver disease. In this study we measured the serum amino acid levels, by Beckman Aminoacid Analyzer, in 11 healthy subjects and 24 HBsAg-positive patients with biopsy-proven liver disease (4 CPH, 10 CAH, 10 cirrhosis). A significant decrease in total aminoacids was observed in CAH and cirrhosis groups (-24% and -22% respectively). The three branched chain aminoacids (BCAA = val + leu + isoleu) were reduced by 24% (P less than 0.002) and 37% (P less than 0.001) in the CAH and cirrhosis groups respectively. Tyrosine was the only of the aromatic aminoacids (AAA) to increase in cirrhotics (+ 34%, P less than 0.02). The molar ratio BCAA/AAA was 3.6 in controls, 3.8 in CPH, 3.1 in CAH (P less than 0.025) and 1.9 in cirrhosis (P less than 0.001). A linear correlation was found between molar ratio BCAA/AAA and serum albumin in all patients (P less than 0.001). These results document the presence of specific quantitative changes in serum aminoacids of HBsAg positive patients, which appear related to severity of liver disease and comparable to the alterations described in non viral chronic liver disease.     

急性肝损伤大鼠模型的血浆氨基酸代谢变化模式

以Ｄ－氨基半乳糖（Ｄ－Ｇａｌａｃｔｏｓａｍｉｎｅ,Ｄ－ＧａｌＮ）造成急性肝损伤（急性肝炎、急性肝坏死）大鼠模型后、对照观察了急性肝损伤大鼠血浆氨基酸的变化,建立了大鼠急性肝损伤时血浆氨基酸的变化模式并对其发生机理进行了探讨。大鼠血浆氨基酸的测定采用聚酰薄层荧光分析技术,其测定结果是：急性肝炎组,酪氨酸（Ｔｙｒ）、天冬氨酸（Ａｓｐ）、谷氨酰胺（Ｇｌｎ）和鸟氨酸（Ｏｒｎ）升高,精氨酸（Ａｒｇ）下降,其余氨基酸无显著变化。急性肝坏死组,除Ａｒｇ显著下降外,其余所有氨基酸都显著升高,而两组支链氨基酸（ＢＣＡＡ）／芳香族氨基酸（ＡＡＡ）克分子比值均显著下降。     

Changes in plasma phenylalanine, isoleucine, leucine, and valine are associated with significant changes in intracranial pressure and jugular venous oxygen saturation in patients with severe traumatic brain injury

Changes in plasma aromatic amino acids (AAA?=?phenylalanine, tryptophan, tyrosine) and branched chain amino acids (BCAA?=?isoleucine, leucine, valine) levels possibly influencing intracranial pressure (ICP) and cerebral oxygen consumption (SjvO(2)) were investigated in 19 sedated patients up to 14?days following severe traumatic brain injury (TBI). Compared to 44 healthy volunteers, jugular venous plasma BCAA were significantly decreased by 35% (p?相似文献   

用高效液相色谱定量分析分支链氨基酸

目的：周2,4-二硝基氟苯（DNFB）对分支链氨基酸衍生后,采用优化的高效液相色谱（HPLC）对其进行定量分析。方法：色谱柱为AgilentZORBAXEclipsAAA（4．6mm×150mm,5-Micron）,流动相为乙酸盐缓冲液（pH6．4）-乙腈,流速1．0mL／min,检测波长360nm。结果：用HPLC法测定分支链氨基酸的浓度为20-200mg／L时线性关系良好,3种分支链氨基酸的R。均在0．9997以上,平均回收率高,RSD≤0．56％（n=6）。结论：此方法快速、准确、重现性好,适合于对发酵液中分支链氨基酸的定量分析。     

HPLC法分析乳清蛋白对糖尿病模型小鼠血浆氨基酸组分的影响

为寻求有效的氨基酸分离方法并探讨乳清蛋白对2型糖尿病防治的作用机制,采用HPLC法分析乳清蛋白中氨基酸组分及含量;分别以0％、10％、20％和40％的乳清蛋白(WP)灌胃1型、2型糖尿病模型组、正常组小鼠,4周后观察各组血浆氨基酸的变化.乳清蛋白中亮氨酸、异亮氨酸、缬氨酸分别占氨基酸总量的14.40％、5.93％和5....     

Phenotype of Hepatocyte Spheroids Behavior within Thermo-Sensitive Poly(NiPAAm-co-PEG-g-GRGDS) Hydrogel as a Cell Delivery Vehicle

Aggregates (spheroids) of specific cells are often regarded as a better form in artificial organs and mammalian cell bioreactors for improved cell-specific functions. Freshly harvested primary rat hepatocytes, cultivated as spheroids and entrapped in an adhesion molecules of Arg–Gly–Asp (RGD)-conjugated extracellular matrix, have been examined for differentiated morphology and enhanced liver-specific functions. A copolymer of RGD conjugated p(NiPAAm-co-PEG) hydrogel was used to entrap hepatocytes in the forms of spheroids. Over 28 days, entrapped the spheroids had a higher viability and produced albumin and urea at constant rates, while there was slight increase in cell numbers and reduction of albumin secretion in single cell culture in the hydrogel. Hepatocytes cultured in this way are a potentially useful three-dimensional cell system for application in a bioartificial liver device and bioreactor.The first two authors (Keun-Hong Park and Kun Na) are equally contributed to this work.