Biomedical subjects

Yuan-Yuan Chen

Publications and source records attributed to Yuan-Yuan Chen.

13 recordsLinked to original sources

PubMed2006

Cell-free bioassay for measurement of dioxins based on fluorescence enhancement of fluorescein isothiocyanate-labeled DNA probe.

This study aims to develop a rapid and sensitive cell-free bioassay of dioxins. It is known that dioxin ligand can bind heterodimeric aryl hydrocarbon receptor (AhR) and triggers the formation of the complex of dioxin-AhR, AhR nuclear translocator (ARNT), and dioxin-responsive element (DRE) region of the DNA. The hypothesis of the proposed method is that if FITC were labeled at the DRE sequence, its fluorescence intensity would be enhanced when the complex forms because the interaction interface of the binding components (AhR, ARNT, and DRE) creates a rather hydrophobic condition that is in favor of FITC emission. Effects of modification site of FITC on the DNA probes on binding efficiency between the complex components and fluorescence emission enhancement were evaluated by surface plasmon resonance and fluorescence analysis, respectively. Results showed that the labeling site at the second base at the 5' end apart from the core region (5'-TNGCGTG-3') of DRE did not obviously interfere with the binding between the DNA probe and dioxin-AhR/ARNT hybrid but presented significant fluorescence emission enhancement. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) was used as the typical toxin in this study. The method had a linear range of 1-100 pM, with detection limit of 0.1 pM (0.64 fg/assay) and coefficient of variation of 5.6% (n = 10, 50 pM TCDD in transformed cytosol). The whole detection cycle was approximately 4 h. The method was also used to estimate the toxic equivalents (TEQ) of 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD) and 1,2,3,4,7,8-hexachlorodibenzo-p-dioxin (HxCDD). Measurement of TEQs of the mixture of TCDD, PeCDD, and HxCDD were highly consistent with the predicted data. The average recovery using fly ash extract was approximately 93%.

Biological Assay↗

PubMed2006

The key DNA-binding residues in the C-terminal domain of Mycobacterium tuberculosis DNA gyrase A subunit (GyrA).

As only the type II topoisomerase is capable of introducing negative supercoiling, DNA gyrase is involved in crucial cellular processes. Although the other domains of DNA gyrase are better understood, the mechanism of DNA binding by the C-terminal domain of the DNA gyrase A subunit (GyrA-CTD) is less clear. Here, we investigated the DNA-binding sites in the GyrA-CTD of Mycobacterium tuberculosis gyrase through site-directed mutagenesis. The results show that Y577, R691 and R745 are among the key DNA-binding residues in M.tuberculosis GyrA-CTD, and that the third blade of the GyrA-CTD is the main DNA-binding region in M.tuberculosis DNA gyrase. The substitutions of Y577A, D669A, R691A, R745A and G729W led to the loss of supercoiling and relaxation activities, although they had a little effect on the drug-dependent DNA cleavage and decatenation activities, and had no effect on the ATPase activity. Taken together, these results showed that the GyrA-CTD is essential to DNA gyrase of M.tuberculosis, and promote the idea that the M.tuberculosis GyrA-CTD is a new potential target for drug design. It is the first time that the DNA-binding sites in GyrA-CTD have been identified.

Adenosine Triphosphatases↗

PubMed2006

Reversible immobilization of proteins with streptavidin affinity tags on a surface plasmon resonance biosensor chip.

Dissociation of biotin from streptavidin is very difficult due to their high binding affinity. The re-use of streptavidin-modified surfaces is therefore almost impossible, making devices containing them (e.g. surface plasmon resonance (SPR) sensor chips) expensive. This paper describes a new protocol for reversible and site-directed immobilization of proteins with streptavidin affinity tags on the streptavidin-coated SPR biosensor chip (SA chip). Two streptavidin affinity tags, nano-tag and streptavidin-binding peptide (SBP tag), were applied. They both can specifically interact with streptavidin but have weaker binding force compared to the biotin-streptavidin system, thus allowing association and dissociation under controlled conditions. The SA chip surface could be regenerated repeatedly without loss of activity by injection of 50 mM NaOH solution. The fusion construct of a SBP tag and a single-chain antibody to mature bovine prion protein (scFv-Z186-SBP) interacts with the SA chip, resulting in a single-chain-antibody-modified surface. The chip showed kinetic response to the prion antigen with equilibrium dissociation constant K (D) approximately equal to 4.01 x 10(-7). All results indicated that the capture activity of the SA chip has no irreversible loss after repeated immobilization and regeneration cycles. The method should be of great benefit to various biosensors, biochips and immunoassay applications based on the streptavidin capture surface.

Animals↗

PubMed2006

[Fusion tags technology and their applications].

Fusion tags are originally developed to facilitate the purification of recombinant protein from crude extracts. In recent years, the discovery of different tags and the development of fusion strategy make the function of fusion tags diversified. However, there was no a cure-all fusion tag for different applications. We here give an overview of fusion tag technology and the different applications of fusion tags, including the purification, detection and oriented immobilization of recombinant protein, the visualization of bioevent in vivo, the enhancement of the yield of protein, the improvement of the solubility and stability of the expressed protein.

Recombinant Proteins↗

PubMed2005

Effects of long-term treatment with candesartan on hemodynamics and organ damage in spontaneously hypertensive rats.

UNLABELLED: This study was designed to investigate the effects of candesartan on blood pressure (BP) and blood pressure variability (BPV) reductions, baroreflex sensitivity (BRS) amelioration, and organ protection in spontaneously hypertensive rats (SHR). METHODS: Studies were performed in two groups of SHR (n = 13 for control rats; and n = 20 for candesartan-treated rats) and one group of WKY rats (n = 13). Candesartan (3 mg /kg/d) was given in rat chow for 4 months. BP was then continuously recorded for 24 hours in conscious state. After the determination of BRS, rats were killed for organ-damage evaluation. RESULTS: Long-term treatment with candesartan significantly reduced BP and BPV expressed by both standard deviation and variation coefficient of BP, enhanced BRS and produced obvious organ protection. Compared with BP level, BPV and BRS values showed a closer or similar relationship with organ-damage parameters in SHR. Multiple regression analysis showed that the decrease in left ventricular hypertrophy was most closely associated with the increase in BRS, whereas the decrease in aortic hypertrophy was most closely associated with the decrease in 24-hour systolic BPV, and the amelioration in renal lesions, with the increase in BRS and the decrease in 24-hour systolic BPV. CONCLUSION: long-term treatment with candesartan results in organ protection in SHR. Besides BP reduction, the decrease in BPV and the restoration of BRS are significantly related to this organ protection.

Administration, Oral↗

PubMed2005

Expression and mutational analysis of tyrosine kinase receptors c-kit, PDGFRalpha, and PDGFRbeta in ovarian cancers.

Most women with epithelial ovarian cancer are diagnosed with advanced disease. Despite surgery and initial tumor reduction by standard chemotherapy, the tumors frequently recur and the patients eventually die of their disease. New drugs that inhibit tyrosine kinase receptors (TKRs) are being investigated for treatment and this study was undertaken to determine the expression and mutational state for 3 TKRs (c-kit, platelet-derived growth factor receptor [PDGFR] alpha, and PDGFR beta) in ovarian cancer. Tissue arrays containing 84 epithelial ovarian tumors were studied by immunohistochemistry with antibodies specific for c-kit, PDGFR alpha, and PDGFR beta. Immunoreactivity was detected in 78% of the tumor to at least one TKR. PDGFR alpha was expressed in the largest percentage of ovarian tumors (58%) whereas 29% expressed PDGFR beta. Two commercial antibodies against c-kit were studied and 33% of the tumors stained with one but only 6% were interpreted as positive with the second antibody. Activation of TKRs may occur through mutations but, by sequence analysis, no mutations were detected in 6 ovarian tumors with elevated immunoreactivity for each of the TKRs (c-kit, PDGFR alpha, and PDGFR beta). Tyrosine kinase receptors could also be activated through autocrine or paracrine stimulation of receptor by its ligand. Of 43 (35%) tumors tested for both c-kit receptor and its ligand (stem cell factor), 15 expressed both proteins indicating the possibility that this autocrine stimulation feedback loop is a factor in the growth of some ovarian cancers. This study demonstrates that PDGFR alpha, PDGFR beta, and c-kit are expressed in a high percentage of epithelial ovarian cancers suggesting that tyrosine kinase inhibitors may be useful in the treatment of these tumors.

Adenocarcinoma↗

PubMed2005

Lack of association of cytomegalovirus with human brain tumors.

Cytomegalovirus (CMV) is thought to possess oncogenic properties and has been linked with a number of human malignancies. CMV infection was recently described in association with malignant gliomas. The intent of the present study was to further investigate the reported association between CMV and malignant gliomas. Tissue from 22 brain tumors of various histologic types and grades, four normal brains, six breast carcinomas, six colon carcinomas, six lung carcinomas, and six sarcomas were evaluated for the presence of CMV by polymerase chain reaction (PCR), in situ hybridization, and immunohistochemical methods. None of the brain tumors or normal brain tissue tested demonstrated evidence of CMV pp65 or early nuclear proteins by immunohistochemistry. In addition, no CMV RNA or DNA was detected in these cases by in situ hybridization and PCR. None of the carcinomas or sarcomas evaluated were positive for CMV by immunohistochemistry, in situ hybridization, or PCR. The findings of the present study suggest that CMV is not significantly associated with brain tumors in humans.

Brain Neoplasms↗

PubMed2005

Neoadjuvant chemotherapy followed by late-course accelerated hyperfractionated radiation therapy for locally advanced non-small-cell lung cancer: long-term results of a phase I/II clinical trial.

Toxicity, response, and long-term results of a definitive chemotherapy/radiation therapy (RT) protocol in patients with unresectable stage III non-small-cell lung cancer (NSCLC) were evaluated. Two cycles of cisplatin-based chemotherapy were delivered before RT, and another 2 cycles were added for patients who responded to the first 2 cycles of chemotherapy. The first course of radiation covered the primary lesion and elective nodal regions, given in 2 Gy per fraction, 5 days a week for a dose of 40 Gy. Late-course hyperfractionated accelerated RT was delivered to the gross tumor twice a day for an additional 27 Gy within 2 weeks, using 1.5 Gy per fraction. Fifty-three patients with unresectable stage IIIA (N2) and IIIB NSCLC were eligible for analysis. Twelve patients developed grade 3 neutropenia, and 3 patients developed grade 4 neutropenia. Grade 2 or 3 esophagitis was observed in 14 and 2 patients, respectively, and grade 2 or 3 pneumonitis was observed in 9 and 1 patient, respectively. Six patients developed grade 2 and 1 patient developed grade 3 late lung toxicity. The median survival time was 15.5 months. Twenty-six of 53 patients (49%) have died of locoregional progression inside the thorax. The distant metastasis rate was 59.5% (22 of 37 patients) for those who did not respond to chemotherapy and 18.8% (3 of 16 patients) for those who responded to chemotherapy (P = 0.006). Late-course hyperfractionated accelerated RT combined with induction chemotherapy was well tolerated and yielded long-term results that compare favorably with those of studies using 2 cycles of induction chemotherapy and conventional fractionated RT. However, local control was still discouraging.

Adult↗

PubMed2004

Epstein-Barr virus plays no role in the tumorigenesis of small-cell carcinoma of the lung.

Epstein-Barr virus infection has been associated with lymphoepithelioma-like carcinoma of the lung in Asian patients. Recently, Epstein-Barr virus proteins or genomic DNAs were detected in pulmonary squamous-cell carcinoma, adenocarcinoma, and undifferentiated small-cell carcinoma in American patients. We studied 23 cases of small-cell carcinoma of the lung for evidence of Epstein-Barr virus infection by in situ hybridization, immunohistochemistry, and polymerase chain reaction methods. Of the 23 cases, 13 cases were primary small-cell carcinoma of the lung and 10 cases were metastatic small-cell carcinoma of the lung to the brain (one case), liver (two cases), and lymph nodes (seven cases). None of the 23 cases was positive for Epstein-Barr virus-encoded small nonpolyadenylated RNA (EBER)-1 by in situ hybridization. By immunohistochemistry, eight cases showed focal positivity for Epstein-Barr virus nuclear antigen-1. The positive immunostaining was focal and was observed in tumor cells, vascular endothelial cells, and lymphocytes, suggesting nonspecific staining. None of the 23 cases was positive for the transactivating immediate-early BZLF1 (ZEBRA) and latent membrane protein (LMP-1). Only one case was positive for the BamHI W region and LMP-1 gene by polymerase chain reaction assay. Some tumor cells in the BamHI W region positive case were also positive for Epstein-Barr virus nuclear antigen-1. Our study indicates that rare cases of American small-cell carcinoma of the lung may contain Epstein-Barr virus-infected cells, but it is unlikely that Epstein-Barr virus plays a role in the tumorigenesis of small-cell carcinoma of the lung.

Biomarkers, Tumor↗

PubMed2004

Radiation therapy concurrent with weekly paclitaxel for locoregionally advanced nasopharyngeal carcinoma: outcomes of a phase I trial.

PURPOSE: The purpose of this study was to define the maximum tolerated dose (MTD) by describing the dose-limiting toxicity (DLT) of paclitaxel given as a 3-hour intravenous infusion concurrently with conventional radiotherapy in patients with locoregionally advanced nasopharyngeal carcinoma (NPC). MATERIALS AND METHODS: Patients with locoregionally advanced NPC were enrolled into a dose-escalating study. Toxicity was graded according to Common Toxicity Criteria 2.0. MTD was defined when 2 of 6 patients developed DLT. The starting dose of paclitaxel was 20 mg/m2 once weekly, with a subsequent dose escalation of 10 mg/m2 in cohorts of 3 new patients. Radiation therapy was administered in a conventional technique over 7 weeks in 2.0-Gy/daily fractions for 5 days/wk up to total doses of 68-70 Gy. RESULTS: From November 2000 to June 2001, 16 patients completed chemoradiotherapy. On the first-dose level (20 mg/m2), no patient experienced DLT. On the next dose level with 30 mg/m2, 1 patient experienced DLT with grade 3 mucositis, which lasted for 5 weeks, and among the additional 3 patients, no one developed DLT. On the third dose level with 40 mg/m2, 1 patient developed grade 3 mucositis and another grade 3 dermatitis, and both of them lasted more than 3 weeks. To make the trial more credible, another 4 patients were added to the 30-mg/m2 level, and no DLT occurred. Thus, the accrual of patients stopped. CONCLUSION: Combined modality with paclitaxel given weekly, as a 3-hour infusion concomitant to conventional radiotherapy, is feasible for locoregionally advanced NPC. The dose recommended for a phase II trial is 30 mg/m2 with mucositis and dermatitis as DLT, and other toxicity is mild.

Adult↗

PubMed2003

Epstein-Barr virus infection is not associated with fibroadenomas of the breast in immunosuppressed patients after organ transplantation.

Epstein-Barr virus has been linked to an increasing number of nonhematolymphoid conditions. Epstein-Barr virus was recently described in association with fibroadenomas of the breast occurring in immunosuppressed patients. To further investigate the potential association of Epstein-Barr virus with fibroadenoma in the context of immune dysfunction, 11 cases of fibroadenoma of the breast in immunosuppressed organ transplant recipients were examined. Cases were evaluated for the presence of Epstein-Barr virus by polymerase chain reaction, in situ hybridization, and immunohistochemical methods. The presence of Epstein-Barr virus genomic DNA was studied by polymerase chain reaction amplification using primers flanking the BamHI-W fragment of the Epstein-Barr virus genome, as well as the Epstein-Barr virus nuclear antigen-4 and latent membrane protein-1 genes. Cases were also evaluated for the presence of defective heterogeneous Epstein-Barr virus DNA. In addition, morphologic analysis by in situ hybridization for Epstein-Barr virus-encoded RNA-1 and immunohistochemistry for latent membrane protein-1 were performed. Epstein-Barr virus DNA was detected in 4 of 11 (36%) cases with BamHI-W polymerase chain reaction. Polymerase chain reaction studies for Epstein-Barr virus nuclear antigen-4 and latent membrane protein-1 genes were positive in two and four cases, respectively. No defective Epstein-Barr virus genomes were identified in any of the cases. Quantitative polymerase chain reaction demonstrated low levels of Epstein-Barr virus in the fibroadenomas studied. Despite the detection of Epstein-Barr virus genomes in a subset of the cases examined, the constituent epithelial and stromal components of all fibroadenomas demonstrated no evidence of Epstein-Barr virus-encoded RNA-1 by in situ hybridization or latent membrane protein-1 expression by immunohistochemistry. Rare Epstein-Barr virus-encoded RNA-1-positive lymphocytes were observed in some cases, which may account for the positive polymerase chain reaction results. The findings of the present study argue against a significant relationship between Epstein-Barr virus and fibroadenomas of the breast in the setting of transplant-associated immunosuppression.

Adolescent↗

PubMed2002

Recurrent B-cell neoplasms after Rituximab therapy: an immunophenotypic and genotypic study.

Rituximab has been widely used to treat relapsing or advanced stage B-cell neoplasms with an efficacy of about 50%. However, approximately 40-50% of Rituximab treated patients will recur. It is not clear whether these recurrent diseases have the same immunophenotype as that of the original tumors. At the City of Hope, we treated 91 cases of CD20-positive B-cell neoplasms with Rituximab in combination with chemotherapy and hematopoietic stem cell transplantation between August 1999 and December 2000. Thirty-five of 91 patients (38%) experienced recurrence during the time period within one year from treatment. Tumor cells from all of the recurrent patients expressed one or more B cell antigens (CD19, CD20, CD22, CD45RA, or CD79a). However, thirteen of 35 recurrent cases showed aberrant loss of CD20 expression (37%) by immunohistochemical or flow cytometric studies. Pre- and post-Rituximab treated tumor cell DNA was successfully extracted from archival paraffin sections, hematoxylin and eosin (H and E) stained slides, smears, or frozen cells in 10 of 13 CD20 negative recurrent cases. PCR studies for immunoglobulin (Ig) heavy chain gene rearrangements were performed in all these cases. Five cases showed identical Ig heavy chain gene rearrangements in the paired specimens. PCR assay for Ig kappa (kappa) gene rearrangement was performed in the five paired specimens lacking detectable Ig heavy chain gene rearrangements; 2 of them showed identical Igkappa gene rearrangements. Three pairs showed unmatched Ig heavy chain and Igkappa gene rearrangements, probably due to poor quality of recovered DNA. Aberrant loss of CD20 antigens may be a mechanism of treatment resistance and should be considered in the immunophenotyping of recurrent Rituximab-treated B-cell neoplasms; therefore, a panel of B cell markers is recommended for the immunologic diagnosis of recurrent B cell malignancies after Rituximab therapy. Seven of ten pairs of recurrent CD20-negative cases showed identical Ig heavy chain and Igkappa gene rearrangements by PCR assay, strongly suggesting that the pre- and post-Rituxan treated B cell neoplasms are clonally-related.

Antibodies, Monoclonal↗

PubMed2002

Epstein-Barr virus nuclear antigen (EBNA)-1 carboxy-terminal and EBNA-4 sequence polymorphisms in nasal natural killer/T-cell lymphoma in the United States.

Epstein-Barr virus (EBV) polymorphisms were examined in 12 cases of nasal natural killer (NK)/T-cell lymphoma diagnosed in the United States (U.S.-NL) with respect to the EBV-associated nuclear antigen (EBNA)-1 carboxy (C)-terminal region and the EBNA-4 region. A single dominant EBV strain was found in all cases. EBNA-1 sequences were remarkably homogeneous, showing either a P-ala (2/12) or P-ala variant (9/12) sequence. Other EBNA-1 subtypes known to be common in U.S.-reactive samples, such as P-thr or V-leu, were not identified. The final case had a base deletion with frame shift and premature stop codon. EBNA-1 C-terminal amino acid substitutions were common at codons 499 (10/12 cases), 502 (7/12), 524 (9/12), and 528 (6/12), all previously reported "hot spots." However, unlike previous reports of other EBV-associated neoplastic and reactive tissues, mutations were absent at residues 487 and 492. Mutations within HLA-A11-restricted immunogenic EBNA-4 epitopes 399-408 and 416-424 occurred in 3 of 12 cases but were not associated with HLA-A11 status. In summary, the exclusive finding of P-ala variant or P-ala EBNA-1 sequences in U.S.-NL cases differs from that reported in U.S.-reactive and non-U.S.-NL cases. Although the significance of this difference is not known for certain, it may be related to geographic and/or site-specific variations, rather than oncogenicity per se.

Amino Acid Sequence↗