Immunohistochemical localization of androgen receptors in female mole rat (Spalax leucodon) tissues

Androgens exert their effects through androgen receptors (AR) in tissues. We investigated the distribution of AR in female mole rat tissues. Tissues were excised, fixed with 10% formalin and embedded in paraffin. Sections were stained after microwave antigen retrieval for immunohistochemistry. Immunostaining of AR immunostaining was detected in the nucleus or cytoplasm of the cells in the cerebral cortex, cerebellum, anterior pituitary, lung, liver, uterus and skin. Granulosa and some thecal cells in the ovary, cardiac muscle cells and adipose cells exhibited a nuclear reaction for AR. In the kidney, labeling of AR was restricted to the cytoplasm of tubule cells. We found that AR could be detected using immunohistochemistry in the nucleus or cytoplasm or both in the presence of androgens.     

豇豆花叶病毒衣壳蛋白编码区在中间组份的RNA核苷酸序列上的定位(英文)

对豇豆花叶病毒两个衣壳蛋白(VP37和VP23)的氨基端和羧基端氨基酸序列进行了分析,这些结果可以允许VP37和VP23编码区在病毒中间组份(M)RNA的核苷酸序列上进行基因定位。这两个编码区是相邻的,并表明,从M RNA的原始翻译产物中释出VP37和VP23的蛋白酶解部位,分别是谷氨酰胺-甲硫氨酸和谷氨酰胺-甘氨酸二肽序列。     

Solid-phase enzyme immunoassay of urokinase using monoclonal antibodies

Using two monoclonal antibodies directed against urokinase, we have developed a micro enzyme-linked immunosorbant assay (ELISA) to detect and measure urokinase in biological fluids. The system presents the following characteristics: simple and rapid procedure, reproducibility, sensitivity (urokinase levels down to 1 ng/ml) and evaluation of the enzyme in biological fluids such as urine, pleural effusions, and ascitic fluids without preliminary purification.     

Use of a bioelectrochemical cell for the synthesis of (bio)chemicals

A bioelectrochemical cell containing either d-glucose oxidase (β-d-glucose:oxygen 1-oxidoreductase, EC 1.1.3.4) or xanthine oxidase (xanthine:oxygen oxidoreductase, EC 1.2.3.2) plus dichlorophenol-indophenol as electron acceptor in one half-cell, and chloroperoxidase (chloride:hydrogen-peroxide oxidoreductase, EC 1.11.1.10) in the other half-cell is described. Due to a combination of chemical, biochemical and electrochemical reactions, electricity and specific (bio)chemicals can be produced in the cell simultaneously and in both compartments. Furthermore, the oxidases in a bioelectrochemical cell are not inactivated by H₂O₂ and as a result the operational lifetimes of the oxidases were increased about five-fold.     

Enzymatic bromination of barbituric acid and some of its derivatives

Barbituric acid, 1-methyl- and 1,3-dimethylbarbituric acid, some of its 5-phenyl derivatives, and 5-chlorobarbituric acid are presented as new substrates for the bromoperoxidase isolated from the brown alga Ascophyllum nodosum. This enzyme is able to convert these substrates into the corresponding 5-bromo or 5,5-dibromo derivatives in good yields. Kinetic measurements show that the structure of the examined substrates has little or no effect on the enzymatic rate of bromination. However, at low substrate concentration the reaction rate depends on both the concentration of the organic substrate and the concentration of hydrogen peroxide. A mechanism is proposed for the reactions of bromoperoxidase with its substrates. These reactions involve the formation of free hypobromous acid which can either brominate the organic halogen acceptor or produce singlet oxygen by a competing reaction with hydrogen peroxide.     

Hinge-thiol coupling of monoclonal antibody to silanized iron oxide particles and evaluation of magnetic cell depletion

Iron oxide particles of average size 0.5-1.5 microns, covered by a silane coat carrying amino groups (Bio-Mag, Advanced Magnetics, Boston), were derivatized by reaction with N-[(gamma-maleimidobutyryl)oxy]-succinimide (GMBS), N-hydroxysuccinimidyl iodoacetate (NHIA), 2-iminothiolane (2-It), or N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP). The derivatized particles were suitable for the reaction with sulfhydryl groups and subsequently coated with monoclonal antibodies (MoAbs) of different classes and isotypes (IgM, IgG1, IgG2a, IgG2b, IgG3) as well as polyclonal rabbit anti-mouse IgG (RAM). The antibodies were reduced by dithiothreitol (DTT) and covalently conjugated to the BioMag derivatives via liberated sulfhydryls of the hinge region. The observed conjugation ratios, expressed as protein/iron (micrograms/mg), could be reproducibly varied for optimization. These ratios were dependent on the type and amount of antibody offered for coupling to the derivatized particles, decreasing as follows: polyclonal = IgM greater than IgG2b greater than IgG2a = IgG3 greater IgG1. The conjugation ratios were also dependent on the type and amount of the spacer used to derivatize the BioMag particles, decreasing as follows: GMBS greater than NHIA greater than 2-It greater than SPDP. The magnetically responsive magnetite-antibody conjugates ("magneto-beads"), carrying MoAb BMA 081 (anti-CD8; IgG2a), MoAb BB10 (anti-CD10/CALLA; IgG2b), MoAb VIL-A1 (anti-CD10; IgM), and polyclonal RAM, coupled similarly via 3.6 mumol of GMBS spacer per mg of Fe, were further investigated with respect to a depletion effect on specific cell subsets. The rates of cell depletion were found to be strongly dependent on the individual characteristics of the antibody used.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence That the 32,000-Dalton Protein Encoded by Bottom-Component RNA of Cowpea Mosaic Virus is a Proteolytic Processing Enzyme

Translation of middle-component RNA of cowpea mosaic virus in vitro produced two polypeptides of 95 and 105 kilodaltons (95K and 105K, respectively) with overlapping amino acid sequences, which were specifically cleaved by a protease encoded by the bottom-component RNA. The proteolytic cleavage was studied by the addition of antibodies raised against various bottom-component RNA-encoded proteins to extracts prepared from bottom-component RNA-inoculated cowpea protoplasts. Since antiserum to the 32K polypeptide efficiently inhibited the proteolytic activity of such extracts, although antiserum to VPg or to the 170K polypeptide did not, evidence was obtained which indicates that the 32K polypeptide represents the protease involved. Fractionation of proteolytically active extract by glycerol gradient centrifugation demonstrated that 32K polypeptides do not exist as free proteins but are aggregated to the bottom-component RNA-encoded 170K, 84K, 60K, or 58K polypeptides. Maximal proteolytic activity was observed for 32K polypeptides associated with 170K polypeptides, suggesting that the activity was unstable and confined to newly synthesized molecules.     

Management of women at increased risk for breast cancer: preliminary results from a new program

OBJECTIVE: To examine the characteristics of malignant tumours that develop in women undergoing surveillance for increased risk for breast cancer and to identify presentation patterns in order to determine the respective roles of mammography, clinical breast examination (CBE) and breast self-examination (BSE). SETTING: Breast Diagnostic Clinic and Familial Breast Cancer Clinic at Toronto-Sunnybrook Regional Cancer Centre. PARTICIPANTS: A total of 1044 women evaluated for breast cancer risk from Oct. 1, 1990, to Dec. 31, 1996, of whom 381 were categorized as being at high risk, 204 as being at moderate risk, 401 as being at slightly increased risk and 58 as being at no appreciably increased risk. PROGRAM COMPONENTS: Comprehensive review and discussion of risk factors, clinical assessment, surveillance recommendations that include mammography, CBE and BSE, genetics consultation (Familial Breast Cancer Clinic) and psychosocial support. Data are captured prospectively, updated at each visit and audited every 3 to 6 months. PROGRAM OUTCOMES: During the study period breast cancer was diagnosed in 24 patients, 12 in the high-risk group, 4 in the moderate-risk group and 8 in the group at slightly increased risk. The mean age at diagnosis was 47 (range 32 to 82) years. Ten cases of cancer were diagnosed during surveillance (incident cancer), 5 in women under age 50. The mean length of time from initial assessment to diagnosis was 28.6 (range 12 to 51) months. Of the 24 women, 17 reported a family history of breast cancer. The mean age at diagnosis in this cohort was 45.5 years, and the diagnosis was made under age 50 in 10 patients (59%). The mean earliest age at which breast cancer was diagnosed in a family member was 42.5 years. CONCLUSIONS: These preliminary results suggest that surveillance of women at increased risk for breast cancer may be useful in detecting disease at an early stage. The regular performance of mammography, CBE and BSE appears necessary to achieve these results.     

In vitro and in vivo conditional sensitization of hepatocellular carcinoma cells to TNF-induced apoptosis by Taxol

High mortality among hepatocellular carcinoma (HCC) patients reflects both late diagnosis and low curability, due to pharmacoresistance. Taxol (TAX) is toxic for many human HCC-derived cell lines, yet its clinical efficacy on HCCs is poor. Combining TAX with other drugs appears a promising possibility to overcome such refractoriness. We analyzed whether combining tumor necrosis factor (TNF) with TAX would improve their toxicity. Human HCC-derived cell lines were treated with TAX or TNF, alone or combined. Apoptosis was assessed by morphology and flow-cytometry. Several pro- and anti-apoptotic molecules were evaluated by western blotting and/or enzymatic assay. After a 24 hour treatment, TNF was ineffective and TAX modestly cytotoxic, whereas HCC cells were conditionally sensitized to TNF by TAX. Indeed some relevant parameters were shifted to a prodeath setting: TNF-receptor 1 was increased, SOCS3, c-FLIP and pSTAT3 were markedly downregulated. These observations provide a significant clue to critically improve the drug susceptibility of HCC cells by combining 2 agents, TAX and TNF. The sequential application of TAX at a low dosage followed by TNF for only a short time triggered a strong apoptotic response. Of interest, prior TAX administration could also sensitize to TNF-induced apoptosis in the Yoshida AH-130 hepatoma transplanted in mice. Therefore, scrutinizing the possibility to develop similar combination drug regimens in suitable preclinical models seems highly advisable.     

Patterns of change over time in darter (Teleostei: Percidae) assemblages of the Arkansas River basin,northeastern Oklahoma,USA

Rivers and streams are among the most threatened ecosystems worldwide, and their fish assemblages have been modified by anthropogenic habitat alteration and introductions of non‐native species. Consequently, two frequently observed patterns of assemblage change over time are species loss and biotic homogenization. In the present study, we compared contemporary (2006–2007) and historical (1948–1955) assemblages of darters, a group of small benthic fishes of the family Percidae, in the Arkansas River drainage of northeastern Oklahoma, USA. Results showed species loss between the two sampling periods, with historical estimates of overall species diversity across the study area exceeding contemporary estimates by five to eight species. Assemblages showed a low degree of darter similarity based on species presence and absence, with pairwise site comparisons (Jaccard's similarity index) between historical and contemporary samples averaging < 0.35. No significant homogenization or differentiation of assemblages occurred. Range expansion of widespread species, one of the primary mechanisms of biotic homogenization, was not observed; rather, all species occurred at a smaller proportion of sites in contemporary samples. Our results highlight the threat posed by anthropogenic habitat alteration to taxonomic groups such as darters, most of which are habitat specialists. However, our results suggest that biotic homogenization is unlikely to occur in the absence of immigration, especially if assemblages are subjected to ‘novel disturbances’ such as dam construction and watershed‐scale habitat degradation which negatively affect all components of the assemblage.