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Biomedical subjects

D Yu

Publications and source records attributed to D Yu.

At least 19 recordsLinked to original sources
PubMed2006

Oncogenic BRAF regulates beta-Trcp expression and NF-kappaB activity in human melanoma cells.

Mutational activation of BRAF is a frequent event in human malignant melanomas suggesting that BRAF-dependent signaling is conducive to melanoma cell growth and survival. Previously published work reported that melanoma cells exhibit constitutive anti-apoptotic nuclear factor kappaB (NF-kappaB) transcription factor activation triggered by proteolysis of its inhibitor IkappaB. IkappaB degradation is dependent upon its phosphorylation by the IkappaB kinase (IKK) complex and subsequent ubiquitination facilitated by beta-Trcp E3 ubiquitin ligase. Here, we report that melanocytes expressing a conditionally oncogenic form of BRAF(V600E) exhibit enhanced beta-Trcp expression, increased IKK activity and a concomitant increase in the rate of IkappaBalpha degradation. Conversely, inhibition of BRAF signaling using either a broad-spectrum Raf inhibitor (BAY 43-9006) or by selective knock-down of BRAF(V600E) expression by RNA interference in human melanoma cells leads to decreased IKK activity and beta-Trcp expression, stabilization of IkappaB, inhibition of NF-kappaB transcriptional activity and sensitization of these cells to apoptosis. Taken together, these data support a model in which mutational activation of BRAF in human melanomas contributes to constitutive induction of NF-kappaB activity and to increased survival of melanoma cells.

Cell Line, Tumor↗
PubMed2006

Ectopic expression of glucagon-like peptide 1 for gene therapy of type II diabetes.

Glucagon-like peptide 1 (GLP-1) is a promising candidate for the treatment of type II diabetes. However, the short in vivo half-life of GLP-1 has made peptide-based treatments challenging. Gene therapy aimed at achieving continuous GLP-1 expression presents one way to circumvent the rapid turnover of GLP-1. We have created a GLP-1 minigene that can direct the secretion of active GLP-1 (amino acids 7-37). Plasmid and adenoviral expression vectors encoding the 31-amino-acid peptide linked to leader sequences required for secretion of GLP-1 yielded sustained levels of active GLP-1 that were significantly greater than endogenous levels. Systemic administration of expression vectors to animals using two diabetic rodent models, db/db mice and Zucker Diabetic Fatty (ZDF) rats, yielded elevated GLP-1 levels that lowered both the fasting and random-fed hyperglycemia present in these animals. Because the insulinotropic actions of GLP-1 are glucose dependent, no evidence of hypoglycemia was observed. Improved glucose homeostasis was demonstrated by improvements in %HbA1c (glycated hemoglobin) and in glucose tolerance tests. GLP-1-treated animals had higher circulating insulin levels and increased insulin immunostaining of pancreatic sections. GLP-1-treated ZDF rats showed diminished food intake and, in the first few weeks following vector administration, a diminished weight gain. These results demonstrate the feasibility of gene therapy for type II diabetes using GLP-1 expression vectors.

Adenoviridae↗
PubMed2006

Upregulation and activation of PKC alpha by ErbB2 through Src promotes breast cancer cell invasion that can be blocked by combined treatment with PKC alpha and Src inhibitors.

Although ErbB2 is known to enhance breast cancer metastasis, the signaling events responsible for this remain elusive. Alpha-isozyme of protein kinase C (PKCalpha), which is involved in cancer development and progression, has been suggested to be activated by ErbB2 without direct evidence. In addition, the roles of PKCalpha in ErbB2-mediated cancer cell malignancy have not been clearly identified. In this study, we investigated whether ErbB2 can activate PKCalpha and determined what role PKCalpha plays in ErbB2-mediated breast cancer cell invasion. We expressed wild-type and mutant ErbB2 with altered signaling capacities in MDA-MB-435 breast cancer cells and revealed that overexpression or activation of ErbB2 in MDA-MB-435 cells upregulated and activated PKCalpha and that downregulation of ErbB2 by small-interfering RNA decreased the expression and activity of PKCalpha in BT474 breast cancer cells. These in vitro results were supported by data from breast cancer patient samples. In 150 breast cancer tumor samples, ErbB2-overexpressing tumors showed significantly higher positive rates of PKCalpha membrane immunohistochemistry staining than that of ErbB2-low-expressing tumors. Mechanistically, we found that PKCalpha is co-immunoprecipitated with Src and PKCalpha expression and activity can be decreased by Src inhibitor PP2 and by the expression of a dominant-negative mutant of Src. Moreover, ErbB2-mediated upregulation of urokinase-type plasminogen activator receptor (uPAR) is reduced by either the PKCalpha inhibitor Go6976 or the Src inhibitor PP2, and the combination of Go6976 with PP2 is superior to either agent alone in suppressing uPAR expression and cell invasion. These results demonstrate that PKCalpha is critical for ErbB2-mediated cancer cell invasion and provide valuable insights for current and future PKCalpha and Src inhibitor clinical trials.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed2006

Targeting and killing of prostate cancer cells using lentiviral constructs containing a sequence recognized by translation factor eIF4E and a prostate-specific promoter.

To develop a gene therapy that would selectively kill prostate cancer cells while sparing normal cells, we have constructed lentiviral vectors that contain a therapeutic gene with a short DNA sequence in the 5'-untranslated region (UTR) that is recognized by the translation initiation factor, eIF4E, which is often overexpressed in malignant cells. Infection of cancer (LNCaP, PC-3M, DU145, and MCF-7 cells) and noncancer cell lines (BPH-1, 267-B1, Plat-E, and Huvec-c cells) with lentivirus having a CMV-promoter and EGFP reporter resulted in high levels of EGFP expression in all cells, whereas, inclusion of the eIF4E UTR recognition sequence restricted high expression to cancer cells and Plat-E cells, which also express substantial levels of eIF4E. Infection of the cells with lentiviral vectors having this UTR in front of the HSV thymidine kinase suicide gene resulted in differential sensitivity to the killing effects of ganciclovir, with at least 100-fold more drug required to kill noncancer cells than cancer cells. Furthermore, in experiments where the CMV promoter was replaced by the prostate-specific ARR(2)PB promoter, the killing effects of ganciclovir were restricted to prostate cancer cells and not seen in nonprostate cancer cells. Our results indicate that combined translational regulation, by incorporation of an eIF4E-UTR recognition sequence into a therapeutic gene, together with transcriptional regulation with a prostate-specific promoter, may provide a means to selectively destroy prostate cancer cells while sparing normal prostate cells.

Cell Death↗
PubMed2006

Liver abscesses in children: a single center experience in the developed world.

OBJECTIVES: The aim of this study was to investigate the clinical and radiologic features, predisposing risk factors, and complications of children with pyogenic liver abscess (PLA) referred to a tertiary pediatric hepatology center. METHODS: We analyzed our database of all children referred to our unit over a 10 year period and performed a case note review of all patients with a radiologically proven PLA. RESULTS: PLA was diagnosed in 15 children (7 boys), 0.5% of all referrals. They presented at a median age of 10 years (range 2 months-15 years). In three children (2 boys), PLA was the first manifestation of chronic granulomatous disease. Among the others, five had radiologic evidence of other intra-abdominal pathology (1 with subsequently proven appendicitis), and four developed portal vein thrombosis with portal hypertension. The commonest isolated pathogen was Staphylococcus aureus. Combined treatment with guided aspiration and prolonged intravenous antibiotics was successful in all patients. CONCLUSION: PLA is a rare diagnosis in children in the developed world. It may be caused by primary neutrophil disorders even in the absence of a previous history of infection. Co-existent appendicitis, intra-abdominal sepsis, and ascending pylephlebitis must be sought because these children are at risk of developing portal vein obstruction and portal hypertension. Prolonged intravenous antibiotic treatment guided by microbiologic sensitivities is highly effective.

Adolescent↗
PubMed2005

Results of a search for cold flows of dark matter axions.

Theoretical arguments predict that the distribution of cold dark matter in spiral galaxies has peaks in velocity space associated with nonthermalized flows of dark matter particles. We searched for the corresponding peaks in the spectrum of microwave photons from axion to photon conversion in a cavity detector for dark matter axions. We found none and place limits on the density of any local flow of axions as a function of the flow velocity dispersion over the axion mass range 1.98 to 2.17 microeV.

Journal Article↗
PubMed2005

Prospective assessment of quality of life in ovarian cancer patients receiving whole abdomen hyperthermia and liposomal doxorubicin.

PURPOSE: Prospective assessment of quality of life (QoL) in patients with refractory, residual or recurrent ovarian cancer receiving whole abdomen hyperthermia and intravenous liposomal doxorubicin chemotherapy. METHODS: Treatment consisted of six cycles of intravenous liposomal doxorubicin at 40 mg m2 followed by whole abdomen hyperthermia with each cycle delivered every 4 weeks. QoL assessment was performed at baseline, prior to each cycle of chemotherapy and every 3 months during follow-up using self-administered questionnaires. Global QoL was rated on a seven-point scale and specific domains of QoL, disease related symptoms and treatment related toxicity were rated on a four-point scale. RESULTS: Thirty-two patients were enrolled on the study and 129 QoL questionnaires were completed. Average age was 57.9 (range 45-76); nine patients had persistent and 23 recurrent disease. Ten patients completed six cycles of therapy. Three patients returned follow-up surveys. Subjects rated their overall QoL and health at baseline as above average with mean scores 5.10 (95% CI=4.62-5.58) and 4.66 (95% CI=4.23-5.08), respectively. No significant change in overall QoL was found between baseline and cycles 4-6 of therapy. Mean ratings of overall health and subject reported differences in QoL between cycles were not significantly changed during therapy. Limited follow-up data were available, but scores suggest possible improvement in QoL for patients completing all therapy. Subjects rated the greatest negative impact on QoL in areas of role functioning and social functioning, where the mean (SD) over all cycles was 2.00 (0.67) and 1.98 (0.70), respectively. For physical symptoms, fatigue and sleep disturbance had the most negative impact on QoL with means (SD) of 2.26 (0.62) and 1.91 (0.70). The moderate treatment related toxicity seen in this study did not significantly impact patients reported QoL. CONCLUSIONS: Patients with unfavourable ovarian cancer responding to intravenous liposomal doxorubicin and whole abdomen hyperthermia maintained above average QoL during therapy. Limited data on patients completing protocol therapy demonstrated possible improvement in QoL.

Abdomen↗
PubMed2005

Phase I/II trial of intravenous Doxil and whole abdomen hyperthermia in patients with refractory ovarian cancer.

OBJECTIVE: A phase I/II study of Doxil combined with whole abdomen hyperthermia was conducted in patients with refractory ovarian cancer. Liposomal doxorubicin combined with hyperthermia has been shown to increase both liposomal delivery and drug extravasation into tumour xenografts resulting in enhanced cytotoxic effects. PATIENTS AND METHODS: Thirty patients with either recurrent or persistent epithelial ovarian cancer were enrolled. All patients had either measurable or assessable disease. Patients received intravenous (IV) Doxil at a dose of 40 mg m-2 as a 1-h infusion followed by whole abdomen hyperthermia. The phase I portion of the study was performed to determine the maximal tolerated dose (MTD) of hyperthermia. Quality of life (QoL) was performed at baseline, prior to each cycle and every 3 months. Plasma pharmacokinetic studies were performed with the first cycle. RESULTS: Ten patients participated in the phase I portion of the study which demonstrated that the MTD of hyperthermia was 60 min after either average vaginal and rectal temperatures of 40 degrees C had been achieved or after 30 min of power application, whichever was shorter. All 30 patients were either paclitaxel and/or platinum resistant initially or developed resistant disease. The median number of prior chemotherapeutic regimens was three (range 2-8) and six patients had been previously treated with Doxil. There were three partial responses for a response rate of 10% (95% CI: [2%, 27%]) and eight patients (27%; 95% CI: [12%, 46%]) had disease stabilization. The median time to progression or death was 3.4 months (95% CI: [2.6, 5.2]) and the median survival was 10.8 months (95% CI: [8.8, 17.4]). Twelve patients (40%) experienced palmar-plantar erythrodysesthesia (PPE), but only four (13%) experienced grade 3-4 PPE toxicity. Doxil systemic exposure was higher in those with grade 3-4 PPE compared to those with no PPE. None of the patients had grade 3-4 thermal toxicity due to hyperthermia. QoL was not decreased in patients responding to therapy. CONCLUSIONS: Therapy with intravenous Doxil and whole abdomen hyperthermia for patients with platinum/paclitaxel resistant ovarian cancer is feasible and does not negatively impact quality of life.

Aged↗
PubMed2004

Use of advances in technology for maritime risk assessment.

The maritime industry is moving toward a "goal-setting" risk-based regime. This opens the way to safety engineers to explore and exploit flexible and advanced risk modeling and decision-making approaches in the design and operation processes. In this article, following a brief review of the current status of maritime risk assessment, a design/operation selection framework and a design/operation optimization framework are outlined. A general discussion of control engineering techniques and their application to risk modeling and decision making is given. Four novel risk modeling and decision-making approaches are then outlined with illustrative examples to demonstrate their use. Such approaches may be used as alternatives to facilitate risk modeling and decision making in situations where conventional techniques cannot be appropriately applied. Finally, recommendations on further exploitation of advances in general engineering and technology are suggested with respect to risk modeling and decision making.

Algorithms↗
PubMed2004

Activation of the phosphatidylinositol-3' kinase pathway and DNA synthesis by a mutant insulin-like growth factor I receptor lacking the NPXY motif.

We have investigated the role of the NPXY motif in the insulin-like growth factor I receptor (IGF-IR) by focusing on the activation of the phosphatidylinositol-3' kinase (PI3-K) pathway and DNA synthesis following IGF-I stimulation. For this purpose, we established stable R-cell lines, which are deficient in endogenous IGF-IR, and express human IGF-IR lacking the whole NPEY(950) sequence (DeltaNPEY). The DeltaNPEY cells showed an apparent autophosphorylation of IGF-IR, albeit with reduced sensitivity to stimulation compared with cells expressing similar levels of wild-type IGF-IR. Activation of insulin receptor substrate (IRS)-1 and IRS-2 was severely impaired in DeltaNPEY cells even at high concentrations of IGF-I. However, recruitment of p85, a regulatory subunit of PI3-K, to activated IRS-2 was similar between the cell lines, but recruitment of p85 to IRS-1 was reduced in DeltaNPEY cells. Essentially similar levels of p85- or phosphotyrosine-associated PI3-K and Akt activities were observed between the cell lines, although the sensitivity to stimulation was reduced in DeltaNPEY cells. Activation of extracellular signal-regulated kinase and DNA synthesis were virtually unaffected by the mutation, in terms of both sensitivity to stimulation and responsiveness. DNA synthesis was completely inhibited by the PI3-K inhibitor, LY294002. These results indicate that the IGF-IR is able to activate the PI3-K pathway and induce DNA synthesis in a normal fashion without the NPXY motif when the receptor is fully activated.

Amino Acid Motifs↗
PubMed2003

Regulation of the expression of the prostate-specific antigen by claudin-7.

Claudins are a family of proteins involved in forming tight junctions between cells. Here we describe two forms of claudin-7 (CLDN-7), a full-length form of CLDN-7 with 211 amino-acid residues and a C-terminal truncated form with 158 amino-acid residues. These two forms of CLDN-7 are able to regulate the expression of a tissue-specific protein, the prostate-specific antigen (PSA), in the LNCaP prostate cancer cell line. We also found that the expression of CLDN-7 is responsive to androgen stimulation in the LNCaP cell line, suggesting that this protein is involved in the regulatory mechanism of androgen. Both forms of claudin-7 are expressed in human prostate, kidney and lung samples, and in most samples, the full-length form of claudin-7 was predominant. However, in some prostate samples from healthy individuals, the truncated form of claudin-7 is predominantly expressed. Our results demonstrated that unlike other claudins, CLDN-7 has both structural and regulatory functions, and the two forms of CLDN-7 may be related to cell differentiation in organ development.

Adenocarcinoma↗
PubMed2003

Effect of PDE5 inhibition combined with free oxygen radical scavenger therapy on erectile function in a diabetic animal model.

Phosphodiesterase (PDE) inhibitors represent an important advance in the treatment of erectile dysfunction (ED). In spite of widespread use and generally good efficacy, as a class they remain ineffective in 15-57% of men. Specific cohorts of patients with severe vascular or neurogenic basis to their ED, such as diabetic men or those who have undergone radical pelvic surgery, demonstrate lower response rates with PDE inhibition treatment. We believe that circulating levels of nitric oxide (NO) may be enhanced through delivery of adequate concentrations of free oxygen radical scavenger molecules such as vitamin E. Higher levels of NO, theoretically, should produce increased penile blood flow with the potential for a synergistic effect when combined with a PDE5 inhibitor. With this hypothesis in mind, 20 adult male Sprague-Dawley streptozotocin-induced (60 mg/kg i.p.) diabetic rats were divided into four therapeutic groups (n=5). Group I--control animals received peanut oil, group II--vitamin E 20 IU/day, group III--sildenafil 5 mg/kg/day and group IV--vitamin E 20 IU/day plus sildenafil 5 mg/kg/day, by oral gavage daily for 3 weeks. Erectile function was assessed as a rise in intracavernous pressure following cavernous nerve electrostimulation. Penile tissue was harvested to determine the changes in tissue morphology including neuronal nitric oxide synthase, smooth muscle alpha-actin and endothelial cell integrity. PDE5 protein content and activity were measured. Significant increases in intracavernous pressure were measured in the animals receiving combined vitamin E plus sildenafil treatment. Immunohistochemical staining showed increases of neuronal nitric oxide synthase, endothelial cell and smooth muscle cell staining. Western blot analysis did not show significant differences of PDE5 protein between the groups. However, higher PDE5 activity was measured in the sildenafil group and lower activity of PDE5 was recorded in the cohort receiving vitamin E with sildenafil. Vitamin E enhanced the therapeutic effect of the PDE5 inhibitor in a meaningful way in this animal model of diabetes. This study indicates a potential means of salvaging erectile function among patients who are refractory to sildenafil.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed2003

Toll-like receptor 9: modulation of recognition and cytokine induction by novel synthetic CpG DNAs.

Bacterial and synthetic DNA containing unmethylated 2'-deoxyribo(cytidine-phosphate-guanosine) (CpG) dinucleotides in specific sequence contexts activate the vertebrate innate immune system. A molecular pattern recognition receptor, Toll-like receptor 9 (TLR9), recognizes CpG DNA and initiates the signalling cascade, although a direct interaction between CpG DNA and TLR9 has not been demonstrated yet. TLR9 in different species exhibits sequence specificity. Our extensive structure-immunostimulatory activity relationship studies showed that a number of synthetic pyrimidine (Y) and purine (R) nucleotides are recognized by the receptor as substitutes for the natural nucleotides deoxycytidine and deoxyguanosine in a CpG dinucleotide. These studies permitted development of synthetic YpG, CpR and YpR immunostimulatory motifs, and showed divergent nucleotide motif recognition pattern of the receptor. Surprisingly, we found that synthetic immunostimulatory motifs produce different cytokine induction profiles compared with natural CpG motifs. Importantly, we also found that some of these synthetic immunostimulatory motifs show optimal activity in both mouse and human systems without the need to change sequences, suggesting an overriding of the species-dependent specificity of the receptor by the use of synthetic motifs. In the present paper, we review current understanding of structural recognition and functional modulation of TLR9 receptor by second-generation synthetic CpG DNAs and their potential application as wide-spectrum therapeutic agents.

Animals↗
PubMed2002

Short-term assay for the identification of neurotoxic compounds and their liver derived stable metabolites.

The release of stable neurotoxic metabolites from liver after metabolic activation of xenobiotics can be investigated in cultures of primary rat hepatocytes as metabolic activation system and embryonic chicken brain cell cultures as target. It was shown that adult rat hepatocyte cell cultures are a reliable bioactivating system for xenobiotics such as cyclophosphamide and isophenphos resulting in the release of stable metabolites into the supernatant. The cultured embryonic chicken brain cells were able to discriminate between the toxicity of parent drugs and its metabolite(s) and between metabolites with an unspecific cytotoxic activity (cyclophosphamide) and metabolites with a high potential to damage specific nerve cell population(s) (isophenphos). The observed neurotoxicity of the isophenphos metabolites is not an acute effect but induced only after a prolonged exposure period. The present study provides evidence that the subsequent use of hepatocytes and brain cell cultures has the potential to be used as an in vitro screening system for the identification of specific and non-specific neurotoxic compounds. Solely stable metabolites are taken into account, since in the in vivo situation only stable metabolites have a change to reach the nervous system. Our data suggest that the present approach is able to discriminate (a) between cell-type and organ specific toxicity, (b) between neurotoxicity derived after long-term or acute exposure, and (c) between the neurotoxicity induced by the native test compound or stable metabolites.

Animals↗
PubMed2002

Expression of the mel gene from Pseudomonas maltophilia in Bacillus thuringiensis.

AIMS: The objective of this work was to express a novel mel gene, responsible for melanin formation, in Bacillus thuringiensis. METHODS AND RESULTS: A novel mel gene from Pseudomonas maltophilia was sub-cloned into B. thuringiensis using a shuttle vector plasmid and electroporation. Results revealed that the mel gene was expressed under the control of the CryIIIA promoter in B. thuringiensis and conferred u.v. protection on the recipient strain. CONCLUSIONS: The novel mel gene from Ps. maltophilia expressed in B. thuringiensis conferred u.v. protection on the recipient strain. SIGNIFICANCE AND IMPACT OF THE STUDY: Products containing B. thuringiensis for pest control are sensitive to u.v. degradation. As melanin has the ability to act as a u.v. absorber, a recombinant B. thuringiensis strain producing melanin provides a new stability for B. thuringiensis preparations.

Bacillus thuringiensis↗
PubMed2001

Multiple signaling pathways involved in activation of matrix metalloproteinase-9 (MMP-9) by heregulin-beta1 in human breast cancer cells.

Matrix metalloproteinase-9 (MMP-9) plays important roles in tumor invasion and angiogenesis. Secretion of MMP-9 has been reported in various cancer types including lung cancer, colon cancer, and breast cancer. In our investigation of MMP-9 regulation by growth factors, MMP-9 was activated by heregulin-beta1 as shown by zymography in both SKBr3 and MCF-7 breast cancer cell lines. Increase in MMP-9 activity was due to increased MMP-9 protein and mRNA levels, which mainly results from transcriptional upregulation of MMP-9 by heregulin-beta1. Heregulin-beta1 activates multiple signaling pathways in breast cancer cells, including Erk, p38 kinase, PKC, and PI3-K pathways. We examined the pathways involved in heregulin-beta1-mediated MMP-9 activation using chemical inhibitors that specifically inhibit each of these heregulin-beta1-activated pathways. The PKC inhibitor RO318220 and p38 kinase inhibitor SB203580 completely blocked heregulin-beta1-mediated activation of MMP-9. MEK-1 inhibitor PD098059 partially blocked MMP-9 activation, whereas PI3-K inhibitor wortmannin had no effect on heregulin-beta1-mediated MMP-9 activation. Therefore, at least three signaling pathways are involved in the activation of MMP-9 by heregulin-beta1. Since MMP-9 is tightly associated with invasion/metastasis and angiogenesis, our studies suggest that blocking heregulin-beta1-mediated activation of MMP-9 by inhibiting the related signaling pathways may provide new strategies for inhibition of cancer metastasis and angiogenesis.

Breast Neoplasms↗
PubMed2001

Intratumoral delivery of an interferon gamma retrovirus-producing cells inhibits growth of a murine melanoma by a non-immune mechanism.

Interferon gamma (IFNgamma) is a potent inhibitor of cell growth effective against a wide range of tumor-derived cell lines. We cloned murine IFNgamma cDNA into a retroviral vector and created a packaging cell line (Am-gamma) producing this IFNgamma-encoding retrovirus. In a pilot experiment, admixing and co-injection of equal numbers of retrovirus-producing and target B16 melanoma cells led to high rates of infection and strong suppression of neoplastic growth. This effect was observed in the absence of measurable systemic production of IFNgamma and could be reproduced in animals lacking cytotoxic immune responses. Tumor angiogenesis was unaffected and no increase in apoptosis was apparent; however, mitotic indices were greatly reduced in Am-gamma-containing abortive tumors. We thus concluded that IFNgamma directly affects proliferation of B16 cells. Indeed, exposure of B16 cells to IFNgamma in vitro inhibits cell division, as measured by a thymidine incorporation assay. Most importantly, repeated intratumoral injections of Am-gamma stunted growth of established B16 melanomas in 75% of treated animals. Thus, this approach can serve as a prototype for new anti-cancer modalities.

Adenoviridae↗