Evidence for a role for genomic instability in radiation-induced mutagenesis.
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Animals↗
Biomedical subjects
Y Yu
Publications and source records attributed to Y Yu.
High-intensity focused ultrasound in the treatment of experimental liver tumour.
This project aimed to determine the adequacy and accuracy of high-intensity focused ultrasound (HIFU) for ablating experimental liver tumour, and to assess imaging methods for monitoring the therapeutic results. The rabbit liver pseudotumour model was established by injection of Freund's complete adjuvant into the liver; the animals then received HIFU therapy via laparotomy at the focal point of the beam (1.1 MHz, 500 W/cm2, 20 s). The rabbits were sacrificed at scheduled times after treatment and liver tumours were examined histologically. Sequential imaging of the liver tumour was performed before and after HIFU treatment. HIFU accurately destroyed the rabbit liver tumour and induced coagulation necrosis 24 h later. Sonographic imaging studies revealed that characteristic changes occurred. A hyperechoic mass turned to a hypoechoic lesion with no Doppler signal, and a high echogenic rim appeared 24 h after HIFU treatment, correlating well with the pathological changes of a sonoablated lesion. These results verify that HIFU has the power to ablate liver tumour quite adequately and accurately, and that sonography is useful for monitoring sonoablated liver tumour.
Animals↗
Involvement of MHC class I molecule and ICAM-1 in the enhancement of adhesion and cytotoxic susceptibility to immune effector cells of tumor cells transfected with the interleukin (IL)-2, IL-4 or IL-6 gene.
To investigate the molecular and cellular mechanisms involved in the reduced tumorigenicity and increased immunogenicity of interleukin-2 (IL-2)-, IL-4- or IL-6-gene-transfected B16 melanoma vaccine, we have analyzed the functional and phenotypic properties of these genetically engineered melanoma cells in the present study. The cytokine-gene-transfected B16 melanoma cells showed stronger adhesion to the lymphokine-activated killer (LAK) cells or cytotoxic T lymphocytes (CTL), and higher sensitivity to cytotoxicity of LAK cells or CTL. Using fluorescence-activated cell sorting analysis, we found that both MHC class I and ICAM-1 expression were increased after IL-2, IL-4 or IL-6 gene transfection. The increased level of MHC class I and ICAM-1 expression seems to be responsible for the high sensitivity of these gene-transfected B16 cells to LAK or CTL cytotoxicity because anti-(MHC class I) or anti-ICAM-1 mAb could inhibit the adhesion and cytotoxicity increment simultaneously. The CTL induction was partly inhibited by anti-ICAM-1 mAb and was completely blocked by anti-MHC class I mAb. These results suggested that the decreased tumorigenicity of IL-2-, IL- 4-, and IL-6-gene-transfected B16 melanoma cells may be partly due to the increased sensitivity to effector cell cytotoxicity mediated by increased expression of ICAM-1 or MHC class I molecules on the tumor cell surface after cytokine gene transfection.
Animals↗
Iodized oil enhances the thermal effect of high-intensity focused ultrasound on ablating experimental liver cancer.
The influence of the biological medium on high-intensity focused ultrasound (HIFU) therapy for ablating experimental liver cancer was studied. In study 1, the temperature rise in the focal zone in the presence of iodized oil or castor oil was observed in vitro. The results showed that HIFU with iodized oil produced a higher and faster temperature rise than did HIFU with castor oil, whether high-power (500 W/cm2) or relatively low-power (136 W/cm2) conditions were used (P = 0.0008 and P = 0.0004 respectively). With the excised liver samples, the temperature also rose higher and more rapidly after injection of iodized oil into the liver than when castor oil was injected (P = 0.0239), and the target liver tissue revealed more radically and extensive destruction with iodized oil than with castor oil. In study 2, 48 nude mice, bearing primary liver cancer LTNM4 implanted subcutaneously, were randomly divided into four groups. Group I (n = 12) were the controls, group II (n = 12) were injected with iodized oil alone, group III (n = 12) received HIFU treatment, and group IV (n = 12) were exposed to HIFU after iodized oil injection. Significant inhibition of tumor growth was seen in groups III and IV as compared with group I or group II (P < 0.05), the tumor growth inhibition rate on the 28th day after treatment being 87% and 93% respectively. Significantly improved survival was noted in groups III and IV compared with groups I and II (P < 0.05). Histologically, group IV showed more complete tumor necrosis than did group III. These data suggest that HIFU combined with iodized oil might have achieve of synergism, location and targeting in the treatment of liver cancer.
Animals↗
Effects of cationic liposome-DNA complexes on pulmonary surfactant function in vitro and in vivo.
Cationic liposome-DNA complexes are being evaluated as potential gene therapy agents for the lung. Cations have strong effects on the biophysical functions of lung surfactant. Therefore, we assessed whether cationic liposomes [composed of N-(1-(2,3-dioleyloxy) propyl)-N,N,N-trimethyl-ammonium chloride and dioleylphosphatidylethanolamine] with or without DNA affect behavior of four types of surfactant in vitro. Experiments were carried out using a modified Wilhelmy surface balance. The ability of surfactants that contain protein and anionic lipids to lower surface tension was inhibited in the presence of cationic liposomes. Inactivation was less when DNA was preincubated with cationic liposomes. Surfactant that contained neither protein nor anionic lipids was not inactivated. Mechanical properties of the lung were studied to assess in vivo surfactant function after intratracheal instillation of a cationic liposome-DNA complex into adult rats. Pressure-volume deflation curves were shifted by 18% compared with those from normal (untreated) animals, but this effect was transient and not different from that observed in animals who received a similar volume of saline. These findings indicate that cationic liposomes alone may have deleterious effects on behavior of some surfactants possibly by disrupting charge interactions between negatively charged phospholipids and surfactant proteins. When DNA is added to liposomes before exposure to surfactants, the adverse charge interactions may be obviated by charge neutralization of liposomes by DNA.
Animals↗
Entamoeba histolytica stimulates interleukin 8 from human colonic epithelial cells without parasite-enterocyte contact.
BACKGROUND & AIMS: The mechanisms involved in the initiation of host mucosal inflammation in amebiasis are not fully understood. This study characterized the effect of Entamoeba histolytica components on interleukin 8 (IL-8) gene expression in human colonic cells. METHODS: Colonic cells were stimulated with amebic proteins, secretory components, or live trophozoites (separated with 0.45-microm pores), and the levels of IL-8 messenger RNA (mRNA) and protein were detected. RESULTS: Live amebae or their components enhanced IL-8 mRNA levels in the colonic cells (T84, LS174T, and Caco-2). In T84 cells, the accumulation of IL-8 mRNA induced by amebic components occurred in a dose- and time-dependent fashion. Increased secretion of IL-8 was noted after 12-hour stimulation; neutralizing antibodies against IL-15 or tumor necrosis factor alpha did not inhibit IL-8 production. Nuclear run-on assays showed that amebae-induced IL-8 gene occurred by a posttranscriptional mechanism. Cycloheximide treatment resulted in superinduction of IL-8 mRNA; however, dexamethasone inhibited E. histolytica-induced IL-8 gene expression. CONCLUSIONS: E. histolytica can directly stimulate the induction of IL-8 by colonic cells in the absence of cell-cell contact or injury.
Animals↗
Co-ordinated regulation of the plasma membrane calcium pump and the sarco(endo)plasmic reticular calcium pump gene expression by Ca2+.
The plasma membrane calcium pump (PMCA) and sarco(endo)plasmic reticular calcium pump (SERCA) are important components of the Ca2+ homeostasis system responsible for intracellular Ca2+ signaling, yet the factors which govern their expression remain unclear. Recently, we have found that overexpression of PMCA by a gene transfection approach caused a down-regulation of the SERCA pump [Liu B.F., Xu X., Fridman R., Muallem S., Kuo T.H. Consequences of functional expression of the plasma membrane calcium pump isoform 1a. J Biol Chem 1996; 271: 5536-5544]. The results suggest an interdependence between PMCA and SERCA gene expression which has prompted us to investigate further on the mechanisms that regulate the expression of these Ca2+ pump genes in various cultured cell lines. In the present study, we have analyzed the isoforms of the PMCA and SERCA in different cells and presented evidence in favor of a co-induction of the PMCA and SERCA gene expression by second messengers, such as protein kinase C, cAMP, and Ca2+. Ectopic expression of PMCA or SERCA led to downregulation of the endogenous forms of both pumps. Changes in the level of mRNAs were paralleled by corresponding altered pump protein contents. The Ca(2+)-mediated increase of gene expression is accompanied by increased transcription rates as indicated by nuclear run-on assay. The co-ordinated induction of the PMCA and SERCA gene expression by thapsigargin was not blocked by the cytosolic application of the Ca2+ chelator BAPTA. We conclude that genes encoding components of the major Ca2+ transport pathways, including pumps and IP3 receptor channels, are regulatorally linked and this link is provided by the Ca2+ load of the ER store. This study points to the importance of gene expression as an integral component in the regulation of cellular Ca2+ homeostasis.
Animals↗
A role for p53 in DNA end rejoining by human cell extracts.
The tumor suppressor p53 is a major regulator in the response of human cells to DNA damage. In this study we assessed the role of p53 in the repair of DNA double-strand breaks in plasmid DNA using cell extracts from three human lymphoblastoid cell lines derived from the same donor. TK6, WI-L2-NS and TK6-E6-5e cells express wild-type, mutated and essentially no p53 protein, respectively. Total cellular extracts from TK6, WI-L2-NS and TK6-E6-5e cells were incubated with EcoRI linearized pUC19 DNA. Southern blot analysis of end-rejoined DNA indicated that the major products formed were linear multimers. There was approximately 2-fold greater end rejoining in WI-L2-NS and TK6-E6-5e extracts compared with TK6 extracts. Total DNA from end-rejoining reactions was purified and used to transform bacteria. Using the lacZ reporter gene as a measure of repair fidelity we found that misrepair, as indicated by white colonies, occurred at 4.1% to 6.5% of transformants, with no significant difference between the three cell lines. Gel analysis revealed that misrepair involved only deletions. Sequence analysis of 11 misrepaired products from each cell line showed 12 different deletions from 4 to 48 bp in length, but each cell line yielded similar product types. These results indicate that total cellular extracts from human lymphoblastoid cells lacking p53 or expressing mutated p53 have increased end-rejoining activity as compared with extracts from cells expressing wild-type p53. However, the p53 status does not influence the ratio of misrepair:correct repair, or the type of misrepair events.
Cell Extracts↗
Inhibition of murine chronic graft-versus-host disease by the chloroform extract of Tripterygium wilfordii Hook f.
The effects of chloroform extract of Tripterygium Wilfordii Hook f (TWH extract) on chronic graft-versus-host disease (GVHD) were examined in a murine experimental model. Chronic GVHD was induced by intravenous transfer of parental DBA/2 spleen cells into unirradiated (C57BL/6 x DBA/2)F1 recipient mice. The effects of TWH extract on GVHD were assessed by measuring both the degree of splenomegaly and the total serum IgE levels 3 weeks after the cell transfer. Subcutaneous administration of TWH extract once a day for 3 weeks suppressed chronic GVHD in a dose-dependent manner. Significant suppression of splenomegaly was first noted in mice treated with 7.5 micrograms/kg of the agent. The maximum inhibition was observed when mice were treated with more than 10.0 micrograms/kg (but not 5.0 micrograms/kg) caused complete suppression of serum IgE hyperproduction. The ability of donor T cells purified from recipient spleen cells to produce interleukin 4 in response to stimulation with anti-CD3 monoclonal antibody was significantly abrogated when recipient mice were treated with 10.0 micrograms/kg of the agent. These results strongly suggest that TWH extract will be an addition to the cohort of immunosuppressive therapies used in solid organ and bone marrow transplantation.
Animals↗
Molecular cloning of a functional murine arginine-specific mono-ADP-ribosyltransferase and its expression in lymphoid cells.
A protein mono-ADP-ribosyltransferase (ADPRT), anchored in the cell membrane as a glycosylphosphatidylinositol (GPI)-anchored cell-surface enzyme, was recently described on murine cytotoxic T cells (CTL). Expression of this enzyme was shown to exert regulatory functions on CTL proliferation and cytotoxic activity, presumably by modulating activity of the protein tyrosine kinase p56(lck), which is associated with the CTL co-receptor CD8. Here we report on the molecular cloning and expression of this important regulatory enzyme. The ADPRT coding sequence was derived by making use of ADPRT sequence homologies from different vertebrate species. A cDNA fragment of the enzyme coding sequence was generated by reverse transcription polymerase chain reaction (RT-PCR) from murine T-cell lymphoma SL12, which expresses the cell-surface ADPRT. The cDNA fragment was found to share extensive homology with the corresponding sequences of human and rabbit muscle ADPRT. In Northern blot hybridization, this cDNA fragment generates a strong hybridization signal with RNA from murine heart and skeletal muscle. Weak signals are seen with SL12, thymus, and spleen. Therefore, a murine skeletal muscle cDNA library was used to identify and obtain the coding sequence of the ADPRT gene. It is shown that the nucleic acid open reading frame sequence of the murine skeletal muscle gene shares 80.3% and 76.3% homology with the sequences of the human and rabbit muscle genes, respectively. Semiquantitative RT-PCR with intron-spanning primers shows that the ADPRT mRNA is present in lymphoid organs, cytotoxic T cells, and T-cell lines. Transfection of the ADPRT coding sequence into EL4 cells results in expression of the enzyme as a functional GPI-anchored cell-surface protein, able to ADP-ribosylate the arginine analog agmatine as well as cell-surface molecules.
ADP Ribose Transferases↗
Influence of selective decontamination of the digestive tract on cell-mediated immune function and bacteria/endotoxin translocation in thermally injured rats.
OBJECTIVE: To determine the influence of pretreatment with selective decontamination of the digestive tract (SDD) on systemic immunosuppression, and the relationship between bacteria/endotoxin translocation and abnormalities of immune function in thermally injured rats. DESIGN, MATERIALS, AND METHODS: Animals were subjected to a 40% full-thickness scald injury, and divided into SDD-treated and control groups. The treatment group received SDD (polymyxin E, tobramycin, and 5-flucytosine) by gavage twice daily for 3 days before the experiment and continued for 5 days after thermal injury. The control group was given the same amount of water. The parameters reflecting cell-mediated immunity, including splenocyte proliferation in response to mitogens, interleukin 2 (IL-2) production, and lymphocyte subpopulation, were measured before injury and 1 and 5 days after burn, respectively. MEASUREMENTS AND MAIN RESULTS: Thermal injury resulted in marked reduction in splenocyte proliferative response to T-cell mitogens, IL-2 production, and T-helper/suppressor cells (CD4/CD8) ratio. Prophylactic treatment with SDD significantly decreased the incidences of bacterial translocation and endotoxemia, prevented suppressive mitogenic response and inadequate IL-2 production (p < 0.05-0.01) but did not affect the abnormal ratio of CD4 to CD8 T lymphocytes in blood (p > 0.05). CONCLUSIONS: These results suggest that bacteria/endotoxin translocation from the gut appears to be involved in cell-mediated immune dysfunction as a consequence of thermal injury. Pretreatment with SDD might attenuate postburn immunosuppression by preventing translocation events.
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Risk factors for bladder cancer: a case-control study in northeast China.
A case-control study of risk factors for bladder cancer was carried out in Heilongjing Province, China. Between May 1989 and May 1990, 217 histologically confirmed cases of bladder cancer and 254 controls with non-neoplastic and non-urine system disease were recruited. Individuals were interviewed in the wards of six major hospitals. Controls were matched by sex, age and area of residence. Information was collected concerning economic status, occupation, histories of smoking and consumption of alcohol, use of tea, the taking of analgesics, dietary histories and previous diseases. Odds ratios (ORs) were calculated from stratified analysis and conditional logistic regression models. Increased risk was observed with increasing times per year and number of years of saccharine use. Compared with non-users, the use of saccharine for more than 19 times per year, and for more than 15 years, the adjusted ORs were 3.9 (95% CI = 1.8-8.67) and 5.1 (95% CI = 2.3-11.6), respectively. Statistically significant associations were also found for diseases related to the urinary system (OR = 2.8; 95% CI = 1.1-7.6). Increased consumption of fruit and vegetable may reduce the risk of bladder cancer. Cigarette smoking had no effect on the risk of bladder cancer in both genders. There was no association between the consumption of alcohol or tea, or types of water supply, with bladder cancer risk.
Adult↗
Decision theoretic steering and genetic algorithm optimization: application to stereotactic radiosurgery treatment planning.
Treatment planning for stereotactic radiosurgery and fractionated radiotherapy is currently a labor intensive, operator-dependent process. Many degrees of freedom exist to make rigorous optimization intractable except by computationally intelligent techniques. The quality of a given plan is determined by an aggregate of clinical objectives, most of which are subject to competing tradeoffs. In this work, we present an autonomous scheme that couples decision theoretic guidance with a genetic algorithm for optimization. Ordinal ranking among a population of viable treatment plans is based on a generalized distance metric, which promotes a decreasing hyperfrontier of the efficient solution set. The solution set is driven toward efficiency by the genetic algorithm, which uses the tournament selection mechanism based on the ordinal ranking. Goals and satisficing conditions can be defined to signal the ultimate and the minimum achievement levels in a given objective. A conventionally challenging case in radiosurgery was used to demonstrate the practical utility and the problem-solving power of the decision theoretic genetic algorithm. Treatment plans with one isocenter and four isocenters were derived under the autonomous scheme and compared to the actual treatment plan manually optimized by the expert planner. Quality assessment based on dose-volume histograms and normal tissue complication probabilities suggested that computational optimization could be driven to offer varying degrees of dosimetric improvement over a human-guided optimization effort. Furthermore, it was possible to achieve a high degree of isodose conformity to the target volume in computational optimization by increasing the degree of freedom in the treatment parameters. The time taken to derive an efficient planning solution was comparable and usually shorter than in the manual planning process, and can be scaled down almost linearly with the number of processors. Overall, the autonomous genetic algorithm scheme was found to be powerful and versatile as a computationally intelligent counterpart to human-guided strategies in treatment optimization for stereotactic radiosurgery and radiotherapy.
Algorithms↗
Multiobjective decision theory for computational optimization in radiation therapy.
Machine-guided iterative optimization in radiation oncology requires ordinal or cardinal ranking of competing treatment plans. When the clinical objectives are multifaceted and incommensurable, the ranking formalism must take into account the decision maker's tradeoff strategies in a multidimensional decision space. To capture the decision processes in treatment planning, a multiobjective decision-theoretic scheme is formulated. Ranking among a group of candidate plans is based on a generalized distance metric. A dynamic metric weighting function is defined based on the state energy of the decision system, which is assumed to undergo thermodynamic cooling with iteration time. The decision maker is required to specify a baseline ranking of the objectives, which is taken to be the ground state of the decision system. This decision-theoretic formalism was applied to idealized cases in stereotactic radiosurgery and prostatic implantation, using the genetic algorithm as the optimization engine. The optimization pathways and the outcome at limited horizons indicated that the combined scheme of decision-theoretic steering and iterative optimization was robust and produced treatment plans consistent with the user's expectation. The effect of treatment uncertainties was simulated using imperfect objectives; however, certain recurring plans could be identified as optimized baseline solutions. Overall, the present formalism provides a realistic alternative to complete utility assessment or human-guided exploration of the efficient solution set.
Algorithms↗
Long-range interactions at the HO promoter.
The SWI5 gene encodes a zinc finger DNA-binding protein required for the transcriptional activation of the yeast HO gene. There are two Swi5p binding sites in the HO promoter, site A at -1800 and site B at -1300. Swi5p binding at site B has been investigated in some detail, and we have shown that Swi5p binds site B in a mutually cooperative fashion with Pho2p, a homeodomain protein. In this report, we demonstrate that Swi5p and Pho2p bind cooperatively to both sites A and B but that there are differences in binding to these two promoter sites. It has been shown previously that point mutations in either Swi5p binding site only modestly reduce HO expression in a PHO2 strain. We show that these mutant promoters are completely inactive in a pho2 mutant. We have created stronger point mutations at the two Swi5p binding sites within the HO promoter, and we show that the two binding sites, separated by 500 bp, are both absolutely required for HO expression, independent of PHO2. These results create an apparent dilemma, as the strong mutations at the Swi5p binding sites show that both binding sites are required for HO expression, but the earlier binding site mutations allow Swi5p to activate HO, but only in the presence of Pho2p. To explain these results, a model is proposed in which physical interaction between Swi5p proteins bound to these two sites separated by 500 bp is required for activation of the HO promoter. Experimental evidence is presented that supports the model. In addition, through deletion analysis we have identified a region near the amino terminus of Swi5p that is required for PHO2-independent activation of HO, suggesting that this region mediates the long-range interactions between Swi5p molecules bound at the distant sites.
Base Sequence↗
Increased expression of EGFR in gastric mucosa of aged rats.
Although in Fischer-344 rats aging is found to be associated with increased gastric mucosal proliferative activity, little is known about the intracellular events that regulate this process. The present investigation examines the age-related changes in gastric mucosal tyrosine kinase activity and expression of epidermal growth factor receptor(EGFR) and its structural and functional analog p185c-erbB-2, the protein product of c-erbB-2/c-neu protooncogene. We observed a significantly higher intrinsic tyrosine kinase activity and tyrosine phosphorylation of EGFR and p185c-erbB-2 in the gastric mucosa of 24-mo-old (aged) rats than in that of their 4- or 12-mo-old counterparts. This was associated with increased levels of EGFR protein and steady-state mRNA levels of EGFR and p185c-erbB-2. In addition, we also observed threefold higher steady-state mRNA levels of transforming growth factor-alpha (TGF-alpha; one of the primary ligands of EGFR) in the gastric mucosa of aged rats than in that of 4-mo-old (young) animals. This was accompanied by a fivefold increase in the relative concentration of the 18-kDa precursor form of TGF-alpha in gastric mucosal membranes but not in the cytosol. In conclusion, our data demonstrate that aging is associated with increased tyrosine kinase activity of EGFR and p185c-erbB-2 in the gastric mucosa. Moreover, the observation that aging results in increased accumulation of TGF-alpha in gastric mucosal membranes raises the possibility that the membrane-bound TGF-alpha could partly be responsible for the constitutively active EGFR-induced signaling pathway in the gastric mucosa of aged rats and, in turn, for stimulation of mucosal proliferative activity.
Aging↗
Elevated right atrial pressure does not reduce collateral blood flow to ischemic myocardium.
Right atrial pressure (RAP) may become substantially elevated during heart failure and has been reported to reduce collateral flow to the ischemic myocardium of isolated hearts. The effect of elevated RAP on blood flow to collateral-dependent and normal myocardium of in situ hearts was studied in 20 open-chest anesthetized dogs with acute occlusion of the left anterior descending coronary artery. Regional myocardial blood flow was measured with radioactive microspheres while RAP was elevated by restricting right ventricular (RV) outflow with constant aortic pressure. Increasing RAP from 3.8 +/- 0.5 to 21.5 +/- 0.8 and then to 34.3 +/- 0.9 mmHg did not reduce blood flow to any transmural region of LV normal or collateral-dependent myocardium. Further elevation of RAP to 49.3 +/- 1.1 mmHg reduced subepicardial but not subendocardial collateral flow. Blood flow to normal RV increased. Retrograde flow and peripheral coronary pressure increased as RAP was elevated. Previously injected 11-micron microspheres were present in the retrograde flow when RAP was elevated; thus retrograde capillary flow contributed to the retrograde flow. The results explain discrepancies among previous reports, and they are consistent with a waterfall phenomenon in the coronary collateral circulation.
Animals↗
Ultrastructural study of glomerular capillary loops at different perfusion pressures as revealed by quick-freezing, freeze-substitution and conventional fixation methods.
Wistar rat kidneys were perfused with some fixatives at different pressures of 100, 150, 200 or 250 cm H2O via the aorta and were studied by the quick-freezing and freeze-substitution (QF-FS) or deep-etching (QF-DE) methods, in addition to the conventional immersion or perfusion fixation method. Different parts of glomerular capillary walls were selected for morphometric analyses. It was demonstrated that the widths of glomerular slit diaphragms and the spaces between foot processes were more widely dilated at higher perfusion pressures (200 and 250 cm H2O) than those seen at both normal perfusion pressure (150 cm H2O) and lower perfusion pressure (100 cm H2O). On the other hand, the glomerular basement membranes were thinner at higher perfusion pressures. By the QF-FS and QF-DE methods, the foot processes showed different shapes from those revealed by the conventional preparation methods, even at the same perfusion pressure. It is concluded that the widths of glomerular slit diaphragms and glomerular basement membranes and the spaces between foot processes may be significantly changed in vivo, depending on the hemodynamics in the glomerular capillary.
Animals↗