[The protective effect of optimized target delineation in intensity modulated radiation therapy on swallowing function in patients with different TNM staging of nasopharyngeal carcinoma].

Objective:To investigate the protective effect of optimized target delineation in intensity modulated radiation therapy(IMRT) on swallowing function in patients with different TNM staging of nasopharyngeal carcinoma.Method:Fifty patients with nasopharyngeal carcinoma were enrolled in this study. They were randomly divided into 25 cases of experimental group and 25 cases of control group by random number table and received IMRT treatment. Patients in control group only received routine delineation of target areas, the patients in experimental group were given the delineation of the relevant parts of the swallowing on the basis of the control group.And then, the degree of dysphagia, xerostomia, weight loss, and quality of life were assessed in the two groups of patients during and after IMRT.Result:There was no significant change in the degree of dysphagia in stage Ⅱ patients during radiotherapy, but the degree of dysphagia in stage Ⅲ and Ⅳ patients increased with the increase of radiotherapy time. After the end of radiotherapy, there was no significant change in the degree of dysphagia in the control group of patients. Compared with the 0th week after the end of radiotherapy, the stage Ⅱ patients in experimental group showed significant improvement in week 12(P<0.05), while the stage Ⅲ and Ⅳ patients showed significant improvement in week 24 after radiotherapy(P<0.05). The degree of xerostomia of two groups of patients continued to increase with varying degrees during and after IMRT(P<0.05 or P<0.01). The weight of the two groups of patients during radiotherapy continued to decrease with the increase of radiotherapy time,and gradually recovered after the end of radiotherapy.And in the experimental group, the weight loss was significantly lower in week 12 and week 24 than in the control group (P<0.05). During radiotherapy, the quality of life scores of the two groups became lower and lower with the increase of radiotherapy time compared with the 0th week of radiotherapy. After the end of radiotherapy, the quality of life began to gradually improve, and in week 24 after the end of radiotherapy, the quality of life of the experimental group of patients was significantly higher than that of the control group (P<0.05).Conclusion:During radiotherapy of patients with nasopharyngeal carcinoma, the structural organs associated with swallowing function are given individualized target delineation can reduce the occurring of dysphagia due to radiotherapy and improve the quality of life of patients after radiotherapy.

Physical wounding and ethylene stimulated embryogenic stem cell proliferation and plantlet regeneration in protocorm-like bodies of Phalaenopsis orchids.

Phalaenopsis orchids have been regenerated by inducing protocorm-like bodies (PLBs) from etiolated leaf sections. However, the physiological and molecular mechanisms of secondary PLB development and subsequent proliferation have not been explored. Bisectionally cutting primary PLBs resulted in more secondary PLBs at 5 weeks, suggesting an embryogenic stem cell property imposed by wounding of primary PLB tissues. The ethylene precursors ethephon and 1-aminocyclopropanecarboxylic acid and the ethylene perception inhibitor silver nitrate increased PLB formation, while aminoethoxyvinylglycine decreased PLB formation. Ethylene content in wounded PLB explants increased over culture time in media containing ethylene precursors or inhibitors. mRNA levels of PhACS2, PhACS3, and PhACO were increased by ethephon and decreased by ethylene inhibitors. Expression of genes in the ethylene signaling pathway was enhanced following ethylene-precursor treatment and was mitigated by ethylene inhibitors during PLB proliferation. Transcription of PhETR and PhEIN3, as well as PhERS, PhCTR, and PhGTP, was significantly increased 12 h after ethylene treatment. Ethylene and physical wounding stimulated secondary PLB formation in Phalaenopsis, probably through ethylene biosynthesis and signal transduction.

Expression of ß-glucuronidase on the surface of bacteria enhances activation of glucuronide prodrugs.

Extracellular activation of hydrophilic glucuronide prodrugs by ß-glucuronidase (ßG) was examined to increase the therapeutic efficacy of bacteria-directed enzyme prodrug therapy (BDEPT). ßG was expressed on the surface of Escherichia coli by fusion to either the bacterial autotransporter protein Adhesin (membrane ßG (mßG)/AIDA) or the lipoprotein (lpp) outermembrane protein A (mßG/lpp). Both mßG/AIDA and mßG/lpp were expressed on the bacterial surface, but only mßG/AIDA displayed enzymatic activity. The rate of substrate hydrolysis by mßG/AIDA-BL21cells was 2.6-fold greater than by pßG-BL21 cells, which express periplasmic ßG. Human colon cancer HCT116 cells that were incubated with mßG/AIDA-BL21 bacteria were sensitive to a glucuronide prodrug (p-hydroxy aniline mustard ß-D-glucuronide, HAMG) with an half maximal inhibitory concentration (IC50) value of 226.53±45.4 µM, similar to the IC50 value of the active drug (p-hydroxy aniline mustard, pHAM; 70.6±6.75 µM), indicating that mßG/AIDA on BL21 bacteria could rapidly and efficiently convert HAMG to an active anticancer agent. These results suggest that surface display of functional ßG on bacteria can enhance the hydrolysis of glucuronide prodrugs and may increase the effectiveness of BDEPT.

Infrared spectroscopy of the mass 31 cation: protonated formaldehyde vs methoxy.

Pulsed discharges containing methanol or ethanol produce ions having the nominal formula [C,H(3),O](+), i.e. m/z = 31. Similar ions resulting from electron impact ionization in mass spectrometers are long recognized to have either the CH(2)OH(+) protonated formaldehyde or CH(3)O(+) methoxy cation structures. The H(2)OCH(+) oxonio-methylene structure has also been suggested by computational chemistry. To investigate these structures, ions are expanded in a supersonic beam, mass-selected in a time-of-flight spectrometer, and studied with infrared laser photodissociation spectroscopy. Sharp bands in the O-H and C-H stretching and fingerprint regions are compared to computational predictions for the three isomeric structures and their vibrational spectra. Protonated formaldehyde is the most abundant isomer, but methoxy is also formed with significant abundance. The branching ratio of these two ion species varies with precursors and formation conditions.

Vibrational spectroscopy and theory of the protonated benzene dimer and trimer.

Protonated benzene cluster ions, H(C(6)H(6))(2)(+) and H(C(6)H(6))(3)(+), are produced in a pulsed electrical discharge source coupled to a supersonic expansion. Mass-selected complexes are investigated with infrared photodissociation spectroscopy in the 1000-3200 cm(-1) region using the method of argon tagging. The IR spectra of H(C(6)H(6))(2)(+)-Ar and H(C(6)H(6))(3)(+)-Ar contain broad bands in the high frequency region resulting from CH-π hydrogen bonds. Sharp peaks are observed in the fingerprint region arising from the ring modes of both the C(6)H(7)(+) and C(6)H(6) moieties. M06-2X calculations have been performed to investigate the structures and vibrational spectra of energetically low-lying configurations of these complexes. H(C(6)H(6))(2)(+) is predicted to have three nearly isoenergetic conformers: the parallel displaced (PD), T-shaped (TS), and canted (C) structures [Jaeger, H. M.; Schaefer, H. F.; Hohenstein, E. G.; Sherrill, C. D. Comput. Theor. Chem. 2011, 973, 47-52]. A comparison of the experimental dimer spectrum with those predicted for the three isomers suggests an average structure between the TS and PD conformers, which is consistent with the low energy barrier predicted to separate these two structures. No evidence is found for the C dimer even though it lies only 1.2 kcal/mol above the PD dimer. Although the trimer is also computed to have many low lying isomers, the IR spectrum limits the possible species present.

Establishment, characterization, virus susceptibility and transfection of cell lines from cobia, Rachycentron canadum (L.), brain and fin.

Establishment and characterization of two cobia, Rachycentron canadum, cell lines derived from cobia brain (CB) and cobia fin (CF) are described. Caudal fin and brain from juvenile cobia were dissociated for 30 and 10 min, respectively, in phosphate-buffered saline containing 0.25% trypsin at 25 degrees C. The optimal culture condition for both dissociated cells (primary cell culture) was at 28 degrees C in Leibovitz-15 medium containing 10% foetal bovine serum. The cells have been sub-cultured at a ratio of 1:2 for more than 160 passages over a period of 3 years. Origin of the cultured cells was verified by comparison of their sequences of mitochondrial cytochrome oxidase subunit I genes (cox I) with the cox 1 sequence from cobia muscle tissue. The cell lines showed polyploidy. No mycoplasma contamination was detected. Susceptibility to grouper iridovirus was observed for the CB cell line but not the CF cell line. Both cell lines expressed green fluorescent protein after being transfected with green fluorescent reporter gene driven by the cytomegalovirus promoter.

Incidence rates of oral cancer and oral pre-cancerous lesions in a 6-year follow-up study of a Taiwanese aboriginal community.

BACKGROUND: In a previous article, we reported the prevalence rates of oral mucosal lesions in an aboriginal community from an epidemiological survey of oral pre-cancerous lesions. METHODS: Since 1997, the authors have started regular follow-up of the study population originally investigated. Thus, it has been possible to obtain incidence rates for the various oral pre-cancerous lesions and conditions. RESULTS: There were 194 persons without any oral lesion in the 1997 screening. During the clinical follow-up investigation and during the analysis of biopsies from pre-cancerous lesions, we discovered six new lesions (including cancer and pre-cancerous lesions) from five participants. All of the five persons were areca/betel quid chewers, and only one mixed areca/betel quid chewing with cigarette smoking habit. The age-standardized incidence rates for quid lesion, oral submucous fibrosis (OSF) and squamous cell carcinoma (SCC) were 267.0, 374.1 and 146.2 per 100,000 person-years, respectively, for areca/betel quid chewers. CONCLUSIONS: As compared with the rates from India and the general Taiwanese population, the study community encountered a serious problem of oral lesions.

The effects of chewing areca/betel quid with and without cigarette smoking on oral submucous fibrosis and oral mucosal lesions.

OBJECTIVES: The purpose of this study was to investigate the risk of areca/betel quid chewing with or without cigarette smoking on oral submucous fibrosis (OSF) and other oral mucosal lesions. METHODS: A stratified case-control study was designed. There were in total 102 patients with oral mucosal lesions or OSF (confirmed pathologically) in the case group. OSF (n = 62) and oral mucosal lesions (n = 62) in 102 subjects were separately analyzed for men and women investigating their risks. RESULTS: For OSF, people with both smoking and chewing habits had a statistically significant odds ratio (OR) 8.68 (95% CI = 1.87, 40.23). For the group of people with chewing habit only and without any lifetime cigarette smoking habit, the OR was 4.51 (95% CI = 1.20, 16.94). For other oral mucosal lesions, people with mixed habits and chewing only had also significant risks (OR = 8.37 and 3.95, respectively). For both OSF and other oral lesions, the ORs of mixed habits and chewing only were both higher in women than in men. CONCLUSIONS: The areca/betel quid used in Taiwan does not contain any tobacco product. The only way of areca/betel quid could synergize with any tobacco product is through cigarette smoking. A statistically significant association with oral mucosal lesions and OSF was still found in the group of areca/betel quid chewing only.

Malignant mesothelioma of the tunica vaginalis testis.

Malignant mesothelioma of the tunica vaginalis is rare, and is usually not diagnosed until surgery is undertaken. Reports on the ultrasound features of this tumour are limited. We present an unusual case with ultrasound features mimicking an adenomatoid tumour.

TCL1 oncogene expression in B cell subsets from lymphoid hyperplasia and distinct classes of B cell lymphoma.

Activation of the TCL1 oncogene has been implicated in T cell leukemias/lymphomas and recently was associated with AIDS diffuse large B cell lymphomas (AIDS-DLBCL). Also, in nonmalignant lymphoid tissues, antibody staining has shown that mantle zone B cells expressed abundant Tcl1 protein, whereas germinal center (GC; centrocytes and centroblasts) B cells showed markedly reduced expression. Here, we analyze isolated B cell subsets from hyperplastic tonsil to determine a more precise pattern of Tcl1 expression with development. We also examine multiple B cell lines and B lymphoma patient samples to determine whether different tumor classes retain or alter the developmental pattern of expression. We show that TCL1 expression is not affected by Epstein-Barr virus (EBV) infection and is high in naïve B cells, reduced in GC B cells, and absent in memory B cells and plasma cells. Human herpesvirus-8 infected primary effusion lymphomas (PEL) and multiple myelomas are uniformly TCL1 negative, whereas all other transformed B cell lines tested express moderate to abundant TCL1. This observation supports the hypothesis that PEL, like myeloma, usually arise from post-GC stages of B cell development. Tcl1 protein is also detected in most naïve/GC-derived B lymphoma patient samples (23 of 27 [85%] positive), whereas most post-GC-derived B lymphomas lack expression (10 of 41 [24%] positive). These data indicate that the pattern of Tcl1 expression is distinct between naïve/GC and post-GC-derived B lymphomas (P < 0.001) and that the developmental pattern of expression is largely retained. However, post-GC-derived AIDS-DLBCL express TCL1 at a frequency equivalent to naïve/GC-derived B lymphomas in immune-competent individuals (7 of 9 [78%] positive), suggesting that TCL1 down-regulation is adversely affected by severe immune system dysfunction. These findings demonstrate that TCL1 expression in B cell lymphoma usually reflects the stage of B cell development from which they derive, except in AIDS-related lymphomas.

Inhibition of protein kinase C and proto-oncogene expression by crocetin in NIH/3T3 cells.

Crocetin, a carotenoid isolated from the seeds of Gardenia jasminoides, was found to be a potent inhibitor of tumor promotion induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in mouse skin. When mouse fibroblast NIH/3T3 cells were treated with TPA alone, protein kinase C (PKC) translocated from the cytosolic fraction to the particulate fraction. Pretreatment with 60 and 120 microM crocetin for 15 min inhibited the TPA-induced PKC activity in the particulate fraction by 50% and 66%, respectively, but did not affect the level of PKC protein. Crocetin also reduced the level of TPA-stimulated phosphorylation of cellular proteins. Cells pretreated with crocetin (120 microM) had 55% less PKC [3H]phorbol dibutyrate-binding capacity. Suppression of TPA (100 ng/mL)-induced c-jun and c-fos gene expression was also observed in the mouse fibroblast cells pretreated with crocetin (30, 60, and 120 microM). Our results provided a basis for understanding the inhibitory effect of crocetin on TPA-mediated tumor promotion.

Clinical evaluation of the bone marrow imaging agent 99Tcm-phytate in the detection of bone metastases.

The aims of this study were to assess whether 99Tcm-phytate can detect metastatic skeletal lesions, and to compare it with 99Tcm-methylene diphosphonate (99Tcm-MDP) and 99Tcm-labelled human serum albumin nanocolloids (99Tcm-NC). Twenty-four patients with multiple bony metastases, investigated by 99Tcm-MDP whole-body scintigraphy, underwent 99Tcm-phytate bone marrow imaging. A separate bone marrow scintigram with 99Tcm-NC was performed in 20 of the patients. All of the metastatic lesions detected on the 99Tcm-phytate scintigrams exhibited photon-abundant foci only. Most of the 99Tcm-phytate scintigrams detected fewer metastatic lesions than the corresponding bone scintigrams. Visual comparison of the 99Tcm-NC images showed that 13 of 20 99Tcm-NC images were superior to the 99Tcm-phytate images in the detection of metastatic involvement of the skeleton. Thus 99Tcm-phytate should not be used as a bone marrow imaging agent for the detection of skeletal metastases.

Use of adverse conditions to stimulate a cellular stress response by equine articular chondrocytes.

OBJECTIVE: To determine the response of equine articular cartilage cells to heat and calcium stresses. DESIGN: Analysis of newly synthesized, [35S]methionine-labeled proteins after treatment of isolated primary equine chondrocytes. PROCEDURE: Primary cultures of equine articular chondrocytes were incubated at temperatures ranging from 37 to 42 C for heat stress experiments or incubated in the presence or absence of the intracellular calcium pump inhibitor, thapsigargin, for calcium stress experiments. Patterns of new protein synthesis were determined by incubating with [35S]methionine followed by separation of proteins by use of one- or two-dimensional gel electrophoresis and visualization of labeled proteins by use of fluorography. RESULTS: Equine chondrocytes cultured at temperature of 42 C had increased synthesis of specific proteins, compared with the profile of protein synthesis in control chondrocytes cultured at 37 C. These changes were characteristic of the heat shock stress response described in a number of other mammalian cell-types. Equine chondrocytes cultured in the presence of thapsigargin also had increased synthesis of specific proteins. Two-dimensional gel electrophoresis of these newly synthesized proteins revealed the changes to be consistent with the induction of the glucose-regulated protein family of stress proteins. CONCLUSIONS: Changes in the pattern of new protein synthesis can be induced in differentiated equine articular chondrocytes by heat shock or calcium stress. These responses are characteristic of a widely described mammalian stress response that has been postulated to be involved in cellular protective mechanisms. The ability of equine chondrocytes to mount a robust stress response may be important in the processes of tissue damage and recovery in articular joints of horses.

A comparative study of PCR product detection and quantitation by electro-chemiluminescence and fluorescence.

Amplification and detection of target DNA sequences are made possible in a polymerase chain reaction (PCR) by using a mixture of biotinylated and ruthenium(II) trisbipyridal (Ru(bpy)3(2+))-end-labelled primers. In this way, biotin for capture and Ru(bpy)3(2+) for detection are directly incorporated into the PCR product obviating subsequent probe hybridization. PCR of a bacterial DNA template from Alteromonas species strain JD6.5 using a cocktail of biotin- and Ru(bpy)3(2+)-labelled primers amplified a 1 kilobase region. Serial dilution of PCR product followed by magnetic separation with Streptavidin (SA)-coated magnetic beads and an electrochemiluminescence (ECL) assay using the semi-automated QPCR System 5000 demonstrated sensitive (pg range) DNA detection. ECL assay of probe hybridization to a human immunodeficiency virus (HIV) sequence also produced pg level sensitivity. Quantitative DNA determination by ECL assay correlated well with visual detection of DNA in electrophoretic gels. However, DNA detection by ECL assay was 10 to 100 times more sensitive than conventional ethidium bromide staining. The combination of DNA-based magnetic separation with ECL assay provides a very sensitive and rapid method of quantitating DNA which, owing to its rapid and facile nature, may have many applications in the research, environmental monitoring, industrial and clinical fields.

The intracellular Ca(2+)-pump inhibitors thapsigargin and cyclopiazonic acid induce stress proteins in mammalian chondrocytes.

Primary cultures of mammalian articular chondrocytes respond to treatment with the intracellular Ca(2+)-pump inhibitors thapsigargin (TG) and cyclopiazonic acid by specific changes in protein synthesis consistent with a stress response. Two-dimensional gel electrophoresis of newly synthesized proteins confirmed that the response was consistent with the induction of glucose-regulated proteins. The effects of low-dose TG (10 nM), measured by changes in [35S]methionine labelling of newly synthesized proteins, can first be observed by 10 h and are maximal by 24 h. The pattern of changes induced by TG is shared with cyclopiazonic acid, but effects of both perturbants differ significantly from changes induced by heat shock. Upon removal of TG, normal protein synthesis is restored by 48 h. Immunoblots showed increased concentrations of the stress proteins HSP90, HSP72/73 and HSP60 in chondrocytes treated with TG, but induction of newly synthesized heat-shock proteins by TG was not apparent on [35S]methionine-labelled gels. The alterations in protein synthesis induced by Ca(2+)-pump inhibitors were unaffected by BAPTA-AM loading, which clamped cytosolic Ca2+ at resting levels. We conclude that inhibition of intracellular Ca(2+)-pump activity can elicit a stress response, which has important implications for the interpretation of chronic use of Ca(2+)-pump inhibitors. In particular, the activation of the cellular shock response should be considered in interpreting the regulation of protein synthesis and cell survival by Ca(2+)-pump inhibitors such as TG.

Purification and Properties of a Highly Active Organophosphorus Acid Anhydrolase from Alteromonas undina.

A highly active organophosphorus acid anhydrolase from Alteromonas undina was purified to homogeneity and found to be composed of a single polypeptide chain with a molecular weight of 53,000. With diisopropylfluorophosphate as a substrate, the purified enzyme has a specific activity of approximately 575 mumol/min/mg of protein. The enzyme has optimum activity at pH 8.0 and 55 degrees C and is stimulated by sulfhydryl reducing agents and manganese. It is capable of rapidly hydrolyzing a wide range of nerve agents and several chromogenic phosphinates.