Antioxidant N-acetylcysteine attenuates the acute liver injury caused by X-ray in mice

The aim of this study was to evaluate the protective effects of different doses and administration modes of N-acetylcysteine (NAC) against X-ray-induced liver damage in mice. Kun-Ming mice were divided into four groups, each composed of six animals: two control groups and two NAC-treated groups. An acute study was carried out to determine alterations in lipid peroxidation (determined by measuring malondiadehyde (MDA) level), glutathione (GSH) content and superoxide dismutase (SOD) activity (assayed by colorimetric method), and DNA damage (characterized by DNA-single strand break using with comet assay) as well as cell apoptosis (measured by flow cytometry) at 12 h after irradiation. The results showed that there were dose-related decreases in MDA level, DNA damage and cell apoptosis, and dose-dependent increases in GSH content and SOD activity in all NAC-treated groups compared to control groups, indicating that pre-treatment or post-treatment with NAC significantly attenuates the acute liver damage caused by X-ray. In addition, significant positive correlations were observed between MDA level and DNA damage or cell apoptosis, implying that lipid peroxidation plays a major role in X-ray-induced liver injury. The data suggest that NAC exerts its radioprotective effect by counteracting accumulated reactive oxygen species in the liver through its properties as a direct antioxidant and a GSH precursor, when administered before or after X-ray irradiation.

Radiobiological response of human hepatoma and normal liver cells exposed to carbon ions generated by Heavy Ion Research Facility in Lanzhou

Human hepatoma and normal liver cells were irradiated with C-12(6+), ion beams (LET= 96.05 keV/mu m) and gamma-rays at Heavy Ion Research Facility in Lanzhou (HIRFL). The chromatid breaks and break types were detected using the premature chromosome condensation technique. Our experimental results showed that chromatid breaks seem to have a good relation with C-12(6+) absorbed dose and C-12(6+) are more effective to induce chromatid breaks as compared to they-rays. For C-12(6+) ion irradiation the major break was isochromatid break, while chromatid breaks were dominant for gamma-ray irradiation. We also observed that the Relative Biology Effectiveness (RBE) of C-12(6+) ion is about 2.5 times higher than that of gamma-rays.

两种保护剂对C离子诱导小鼠急性肝损伤的应答；Response of Two Protective Agents to Acute Injury Induced by ~(12)C~(6+) Ions in Mouse Liver

比较了N-乙酰半胱氨酸(NAC)及乙酰左旋肉毒碱(ALCAR)对12C6+离子照射小鼠的损伤效应,并探讨了其可能的作用机制。利用4Gy剂量的12C6+离子束对预先给予NAC(100mg/kg)和ALCAR(100mg/kg)保护的昆明小鼠进行单次全身照射。随后检测肝组织中总抗氧化能力(TAC)、DNA单链断裂和细胞凋亡率。结果显示,与照射对照组相比,提前给予NAC和ALCAR均极显著地增强了肝组织的抗氧化能力(P<0.001),减轻了12C6+离子导致的肝组织中DNA断裂(P<0.001)和细胞凋亡(P<0.001)。此外,还发现ALCAR组抗重离子辐照损伤的能力显著地高于NAC组(P<0.05)。实验结果提示了NAC和ALCAR可通过抵御组织内的氧化胁迫,阻止DNA链的断裂和细胞的凋亡,实现对C离子辐照损伤的保护效应。而且ALCAR比NAC可能更适合成为有潜力、有希望的抗C重离子辐射药物。

Phosphoproteome analysis of mouse liver using immobilized metal affinity purification and linear ion trap mass spectrometry

Since protein phosphorylation is a dominant mechanism of information transfer in cells, there is a great need for methods capable of accurately elucidating sites of phosphorylation. In recent years mass spectrometry has become an increasingly viable alternative to more traditional methods of phosphorylation analysis. The present study used immobilized metal affinity chromatography (IMAC coupled with a linear ion trap mass spectrometer to analyze phosphorylated proteins in mouse liver. A total of 26 peptide sequences defining 26 sites of phosphorylation were determined. Although this number of identified phosphoproteins is not large, the approach is still of interest because a series of conservative criteria were adopted in data analysis. We note that, although the binding of non-phosphorylated peptides to the IMAC column was apparent, the improvements in high-speed scanning and quality of MS/MS spectra provided by the linear ion trap contributed to the phosphoprotein identification. Further analysis demonstrated that MS/MS/MS analysis was necessary to exclude the false-positive matches resulting from the MS/MS experiments, especially for multiphosphorylated peptides. The use of the linear ion trap considerably enabled exploitation of nanoflow-HPLC/MS/MS, and in addition MS/MS/MS has great potential in phosphoproteome research of relatively complex samples. Copyright (C) 2004 John Wiley Sons, Ltd.

Cytotoxicity of liver targeted drug-loaded alginate nanoparticles

In this study, novel liver targeted doxorubicin (DOX) loaded alginate (ALG) nanoparticles were prepared by CaCl2 crosslinking method. Glycyrrhetinic acid (GA, a liver targeted molecule) modified alginate (GA-ALG) was synthesized in a heterogeneous system, and the structure of GA-ALG and the substitution degree of GA were analyzed by H-1 NMR, FT-IR and elemental analysis. The drug release profile under the simulated physiological condition and cytotoxicity experiments of drug-loaded GA-ALG nanoparticles were carried out in vitro. Transmission electron micrographs (TEM) and dynamic light scattering (DLS) analysis showed that drug-loaded GA-ALG nanoparticles have spherical shape structure with the mean hydrodynamic diameter around 214 +/- 11 nm.

Biochemical effects of gadolinium chloride in rats liver and kidney studied by H-1 NMR metabolomics

The biochemical effects of gadolinium chloride were studied using high-resolution H-1 nuclear magnetic resonance (NMR) spectroscopy to investigate the biochemical composition of tissue (liver and kidney) aqueous extracts obtained from control and gadolinium chloride (GdCl3) (10 and 50 mg/kg body weight, intraperitoneal injection. i.p.) treated rats. Tissue samples were collected at 48, 96 and 168 h p.d. after exposure to GdCl3, and extracted using methanol/chloroform solvent system. H-1 NMR spectra of tissue extracts were analyzed by pattern recognition using principal components analysis. The liver damages caused by GdCl3 were characterized by increased succinate and decreased glycogen level and elevated lactate, alanine and betaine concentration in liver. Furthermore, the increase of creatine and lactate, and decrease of glutamate, alanine, phosphocholine, glycophosphocholine (GPC), betaine, myo-inositol and trimethylamine N-oxide (TMAO) levels in kidney illustrated kidney disturbance induced by GdCl3.

NMR and pattern recognition studies on the acute biochemical effects of Lu(NO3)(3)

Pattern recognition methods were applied to the analysis of 600 MHz H-1 NMR spectra of urine from rats dosed with compounds that induced organ-specific damage in the liver and kidney. Male Wistar rats were separated into groups (n=4) and each was treated with one of following compounds: HgCl2, CCl4, Lu(NO3)(3) and Changle (a kind of rare earth complex mixed with La, Ce, Pr and Nd). Urine samples from the rats dosed with HgCl2, CCl4 and Lu(NO3)(3) were collected over a 24 h time course and the samples from the rats administrated with Changle were gained after 3 months. These samples were measured by 600 MHz NMR spectroscopy. Each spectrum was data-processed to provide 223 intensity-related descriptors of spectra. Urine spectral data corresponding to the time intervals, 0-8 h (HgCl2 and CCl4), 4-8 (Lu(NO3)(3)) h and 90 d (Changle) were analyzed using principal component analysis (PCA). Successful classification of the toxicity and biochemical effects of Lu(NO3)(3) was achieved.

Arabinogalactan as a carrier for contrast agent in magnetic resonance imaging

Arabinogalactan-Gd-DTPA was synthesized by the reaction of diethylenetriaminepenta-acetic acid (DTPA) bisanhydride with polysaccharide in dry DMSO and characterized by FTIR, elemental analysis and ICP-AES. Its stability was investigated by competition with Ca2+, EDTA, DTPA. The t(1)-relaxivity is 8.06 mmol(-1) . L . s(-1) in D2O, 8.48 mmol(-1) . L . s(-1) in 0.725 mmol . L-1 BSA, respectively. t(1)-weighted MR imaging of rat kidney and liver showed a remarkable enhancement post injection of Arabinogalactan-Gd-DTPA. The results indicate that the arabinogalactan-Gd-DTPA is a potential contrast agent for MRI.

Nuclear magnetic resonance studies on the effects of Rare Earth in rats urine

Male Wistar rats were administrated orally with La(NO3)(3) at doses of 0. 05, 0. 2, 2. 0, 10 and 20 mg/kg body weight. Urine was collected over a 24 h period after dosing. Resonances for a large number of low molecular weight metabolites were assigned in a high resolution H-1 NMR spectra of rat urine. The variation of some low molecular weight metabolites in urine provided a sensitive measurement of Rare Earth induced renal and liver lesions, in which DMA, DMG, urea, Kg, TMAO, succinate, citrate and amino acids have been suggested as NMR markers for renal damage and ethanol, lactate, taurine as the markers for liver damage. The method could be applicable to study of the toxicological effects of other compounds and drugs.

Effects of La3+ and Gd3+ on Ca2+ influx in rat hepatoma H-35 cells

Effects of La3+ and Gd3+ on Ca2+ influx were investigated in rat hepatoma H-35 cells by measuring the initial rate of Ca-45(2+) uptake. It was found that the maximum initial rate of Ca2+ uptake was increased six- to ten-fold at low concentrations of La3+ and Gd3+. Kinetic analyses by measuring the initial rate of Ca2+ influx at different external Ca2+ concentrations indicated the existence of two intracellular exchangeable components in the basal Ca2+ system, with low and high affinities for Ca2+, and only one class of Ca2+ binding sites was observed in the La3+- or Gd3+-treated cells. For high affinity, La3+ and Gc(3+) increased both kinetic parameters K-m and V-max of basal Ca2+ influx. La3+ and Gd3+ compete directly with Ca2+ for Ca2+ binding site for low affinity. The kinetics is competitive.

Molecular characterization of beef liver catalase by scanning tunneling microscopy

Beef liver catalase molecules can stick tenaciously to the highly oriented pyrolytic graphite (HOPG) surface which has been activated by electrochemical anodization. The immobilized sample is stable enough for high resolution scanning tunneling microscope (STM) imaging. When the anodized conditions are controlled properly, the HOPG surface will be covered with a very thin oxide layer which can bind the protein molecules. Individual molecules of native beef liver catalase are directly observed in detail by STM, which shows an oval-shape structure with a waist. The dimensions of one catalase molecule in this study are estimated as 9.0 x 6.0x 2.0 nm(3), which are in good agreement with the known data obtained from X-ray analysis, except the height can not be exactly determined from STM. Electrochemical results confirm that the freshly adsorbed catalase molecules maintain their native structures with biological activities. However, the partly unfolding structure of catalase molecules is observed after the sample is stored for 15 days, this may be caused by the long-term interaction between catalase molecules and the anodized HOPG surface.

Biochemical characterization of selenium-containing catalytic antibody as a cytosolic glutathione peroxidase mimic

A selenium-containing catalytic antibody (Se-4A4), prepared by converting reactive serine residues of a monoclonal antibody (4A4) raised against a GSH derivative into selenocysteines, acts as a mimic of cytosolic glutathione peroxidase (cGPX). To clarify the mechanism of action of this catalytic antibody, detailed studies on kinetic behaviour and biological activity were carried out. A rate of acceleration (k(cat)/K-m/k(uncat)) 10(7)-fold that of the uncatalytic reaction is observed. Under similar conditions, the turnover number (k(cat)) of Se-4A4 is 42% of that of the natural rabbit liver cGPX. The Se-4A4 reaction involves a Ping Pong mechanism, which is the same as that of the natural cGPX. The selenocysteine residue is located in the binding site of the antibody and is shown to be crucial for this activity. Of the thiol compounds tested, only GSH is able to serve as substrate for Se-4A4. It was demonstrated, using the free-radical-damage system (hypoxanthine/xanthine oxidase) of cardiac mitochondria, that Se-4A4 can protect mitochondria from free-radical damage at least 10(4)-fold more effectively than the natural cGPX.