Expression and Characterization of Catalytic Domain of T Cell Protein Tyrosine Phosphatase（△TC-PTP） —— Immunohistochemical Study of △TC-PTP Expression in Non-small Cell Lung Carcinomas

This study objective was to express and characterize the catalytic domain of the human T cell protein tyrosine phosphatase（△TC-PTP） and to study immunohistochemically the expression of △TC-PTP in human non-small cell lung cancers. △TC-PTP gene was PCR amplified with the cDNA of human TC-PTP as template, and cloned into the pT7 expression vector. The recombinant pT7-△TC-PTP was expressed in E. coli Rosetta （ DE3 ） host cells and puri- fied. The enzymatic characteristics of △TC-PTP including enzyme activity and kinetics assay were measured. The antiserum was prepared by immunizing rabbit with the purified recombinant △TC-PTP. Rabbit polyclonal antibody against △TC-PTP was purified by PVDF immobilized antigen affinity chromatography. Immunohistochemical staining of lung cancer tissues was performed with antibody against △TC-PTP protein. △TC-PTP gene was correctly cloned, expressed, and purified. The recombinant △TC-PTP had a highly catalytic activity of PTPase. Squamous cell lung carcinoma showed a significantly higher expression rate of △TC-PTP （76. 92%, 10/13 ） than adenocarcinoma （57.14%, 4/7） and normal lung tissue（20%, 1/5 ）. This study represents the first demonstration that △TC-PTP is highly expressed in human squamous cell lung carcinomas. In addition, this study provides an important basis for further studying the biological function of TC-PTP and its relationship with lung carcinomas and other diseases.     

Application of matrix solid-phase dispersion in the analysis of priority polycyclic aromatic hydrocarbons in fish samples

The performance of matrix solid-phase dispersion (MSPD) for the extraction of polycyclic aromatic hydrocarbons (PAHs) in fish tissue is described. The suitability of different solid supports was tested as well as the influence on the extraction efficiency of the natural fat content in samples. Under optimal conditions 0.6-0.8 g of tissue sample, are dispersed with 2 g of octadecylsiloxane (C18) and 0.5 g of anhydrous sodium sulphate and transferred to the top of a polyethylene solid-phase extraction cartridge which already contains 2 g of florisil and 1 g of C18. Cartridges were eluted with acetonitrile. The analysis of the extracts was carried out by high-performance liquid chromatography (HPLC) coupled with fluorescence detection. The proposed method provides detection limits between 0.04 and 0.32 ng/g for the different considered PAHs, below the maximum levels established by the some regulatory bodies for the six PAHs after recent oil spill episodes and European Union regulations. Recoveries over 80% were obtained for all compounds. Accuracy validation was carried out using the US National Institute of Standards and Technology (NIST) SRM 2977 reference material.     

Voltammetric determination of 5-hydroxydole-3-acetic acid in human gastric juice

A method for the determination of the major serotonin metabolite — 5-hydroxyindole-3-acetic acid (5-HIAA) in human gastric juice by cyclic voltammetry was described. The measurement conditions were investigated. The potential window was chosen from +0.1 to +0.9 V, the supporting electrolyte was 0.025 M PBS solution (pH 2.0). The method allowed determination in the concentration range from 2.0 ×10⁻⁷ to 2.0 ×10 ⁻⁵ M and a detection limit of 80 nM. When samples of gastric juice were analyzed with the method, we obtained the mean content of 5-HIAA in the gastric juice. Meanwhile, interference from other ions and substances were examined. The experimental results indicate that the method for the determination of gastric juice samples is successful.     

鼻咽癌患者治疗过程中头发微量元素的变化

对比了３０名健康人与１００名初诊、三个月后复诊及六个月后复诊的鼻咽癌患者头发中的１１种微量元素的含量。发现病人的锌（Ｚｎ）、铜（Ｃｕ）、锰（Ｍｎ）及钴（Ｃｏ）的含量都比健康人低，而病人的钛（Ｔｉ）及砷（Ａｓ）含量则偏高。但经过三至六个月电疗及化疗之后，这些元素的含量普遍与健康人差异变小，这似乎是成功治疗的一个反映。     

Comparison of slurry sampling and microwave-assisted digestion for calcium, magnesium, iron, copper and zinc determination in fish tissue samples by flame atomic absorption spectrometry

The development of a slurry sampling method for the determination of calcium, copper, iron, magnesium and zinc in fish tissue samples by flame atomic absorption spectrometry is described. In comparison with microwave-assisted digestion, the proposed method is simple, requires short time and eliminates total sample dissolution before analysis. Suspension medium was optimized for each analyte to obtain quantitative recoveries from fish tissue samples without matrix interferences. Nevertheless, iron recoveries higher than 46% were not found. Treatment of samples slurried in nitric acid by microwave irradiation for 15-30 s at 75-285 W permitted to achieve efficient recoveries for calcium, iron, magnesium and zinc. Further improvement in the matrix effects for iron determination was accomplished by the use of an additional step of short microwave-assisted suspension treatment. However, standard addition method was required for calcium and copper determination, being necessary hydrochloric acid as suspension medium for the last one. Although copper could not be determined in the certified reference material using microwave-assisted digestion, the accuracy of the slurry sampling method was verified for all the investigated analytes. Detection limits were 22.8 ± 8.0, 0.884 ± 0.092, 5.07 ± 0.76, 35.5 ± 0.7 and 1.17 ± 0.04 μg g⁻¹ for calcium, copper, iron, magnesium and zinc, respectively. The standard deviations obtained using slurry sampling method and microwave-assisted digestion were not significantly different, and the mean relative standard deviation of the over-all method (n = 3) of the slurry sampling method for different concentration levels was below 12%.     

四川地区宫颈癌组织HPV18和HPV45 E6基因突变分析

采用聚合酶链反应技术对四川地区2003~2004年收集的60例宫颈癌患者的癌组织DNA进行人乳头瘤病毒(HPV) E6基因扩增, 获得HPV18和45型E6基因. 序列分析发现, 18型三例E6基因有同样的两处同义突变; 45型两例E6基因发生突变, 一例有两处碱基突变, 另一例发生六处碱基突变, 其中两处涉及氨基酸变化, 均位于E6抗原决定簇区. HPV45型E6基因中134位c→t, 157位c→t, 259位g→t和341位t→c的碱基点突变未见报道. 另外， 该地区HPV18和45型突变株之间存在碱基互变,  它们之间的最小差异比野生型HPV18和HPV45之间的差异小4.05%, 该数值比在非洲发现的突变株的要小很多, 该结果支持HPV18和HPV45可能起源于非洲的观点.     

GeneMCL in microarray analysis

Accurately and reliably identifying the actual number of clusters present with a dataset of gene expression profiles, when no additional information on cluster structure is available, is a problem addressed by few algorithms. GeneMCL transforms microarray analysis data into a graph consisting of nodes connected by edges, where the nodes represent genes, and the edges represent the similarity in expression of those genes, as given by a proximity measurement. This measurement is taken to be the Pearson correlation coefficient combined with a local non-linear rescaling step. The resulting graph is input to the Markov Cluster (MCL) algorithm, which is an elegant, deterministic, non-specific and scalable method, which models stochastic flow through the graph. The algorithm is inherently affected by any cluster structure present, and rapidly decomposes a graph into cohesive clusters. The potential of the GeneMCL algorithm is demonstrated with a 5,730 gene subset (IGS) of the Van't Veer breast cancer database, for which the clusterings are shown to reflect underlying biological mechanisms.     

Photokilling Squamous Carcinoma Cells SCCVII with Ultrafine Particles of Selected Metal Oxides

The ability of ultrafine particles of TiO₂, WO₃ and iron-doped TiO₂ to kill cancer cells in the presence of UV irradiation was investigated. The best photokilling effect on carcinoma cells SCVII cultured in vitro showed iron-doped TiO₂ ultrafine particles synthesized by the sol-gel procedure with starting chemicals Ti(IV)-isopropoxide and anhydrous Fe(II)-acetate. It was found that a small particle size and high dispersity influenced citotoxicity and photocatalytic efficiency. The remarkable photokiling effect of highly iron-doped TiO₂ ultrafine particles (the molar ratio Fe/Ti = 0.136) in the presence of UV irradiation was observed. The influence of ultrafine metal oxide particles on the inhibition of cancer cell proliferation was measured using a ³H-thymidine incorporation test. The possible mechanism involved in the photokilling of carcinoma cells with ultrafine particles of selected metal oxides was discussed.     

Synthesis, biochemical properties and molecular modelling studies of organometallic specific estrogen receptor modulators (SERMs), the ferrocifens and hydroxyferrocifens: evidence for an antiproliferative effect of hydroxyferrocifens on both hormone-dependent and hormone-independent breast cancer cell lines

A series of ferrocene derivatives based upon the structure of the antiestrogenic drug tamoxifen or of its active metabolite hydroxytamoxifen has been prepared and named by analogy ferrocifens and hydroxyferrocifens. This series includes 1-[4-(O(CH(2))(n)NMe(2))phenyl]-1-phenyl-2-ferrocenyl-but-1-ene and 1-[4-(-O(CH(2))(n)NMe(2))phenyl]-1-(4-hydroxyphenyl)-2-ferrocenyl-but-1-ene, with n=2, 3, 5 and 8, and 1-[4-(-O(CH(2))(2)NMe(2))phenyl]-1-(4-hydroxyphenyl)-2-ferrocenylethene. Most of these molecules have been synthesised by McMurry cross-coupling of the appropriate ketones, except for the ethene complexes, which were prepared by a four-step reaction sequence starting from the ferrocenylacetic acid. All these compounds were obtained as mixtures of Z and E isomers. The isomers were separated in the cases of the ferrocenyl derivatives of tamoxifen and hydroxytamoxifen (n=2). No isomerisation of the Z and E isomers occurred in DMSO after one day, while a 50:50 mixture of the isomers was obtained within one hour in chloroform. The X-ray structure of (E)-1-[4-(-O(CH(2))(2)NMe(2))phenyl]-1-(4-hydroxyphenyl)-2-ferrocenyl-but-1-ene has been determined. The relative binding affinity (RBA) values of the hydroxyferrocifens for the estrogen receptor alpha (ERalpha) was good to moderate, with values decreasing progressively with the length of the basic chain. The RBA values found for the estrogen receptor beta (ERbeta) are equal to or slightly less than those found for the alpha form. The lipophilicity of the hydroxyferrocifens are superior to the values found for estradiol and increase with lengthening of the chain. The antiproliferative effects of the four hydroxyferrocifens with n=2, 3, 5 and 8 were studied on four breast cancer cell lines (MCF7, MDA-MB231, RTx6 and TD5) possessing different levels of ERalpha. On MCF7 cells containing high levels of ERalpha, hydroxyferrocifens behave as antiestrogens. At a molarity of 1 microM the effect is close to that of hydroxytamoxifen (used for reference) when n=2 or 5, more marked when n=3, and weaker when n=8. Ferrocene alone has no effect. For the MDA-MB231 cells, classed as a hormone-independent breast cancer cell line, on the other hand, the hydroxyferrocifens show remarkable antiproliferative behaviour while the hydroxytamoxifen is completely inactive. Hydroxyferrocifens therefore show the unique property of being active both on hormone-dependent and on hormone-independent breast cancer cell lines. The molecular modelling study provides some clues for understanding of the antagonist effect of these molecules, while an additional cytotoxic effect due to the vectorised ferrocenyl unit is revealed in some occasions.