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Biomedical subjects

Ting Zhang

Publications and source records attributed to Ting Zhang.

At least 55 records · Page 3Linked to original sources
PubMed2005

[Preparation and identification of monoclonal antibodies against SARS-CoV putative protein X4].

OBJECTIVE: To obtain monoclonal antibodies against putative protein X4 of SARS-CoV for further study of the structure and function of protein X4. METHODS: Balb/c mice were immunized with recombinant Protein X4, hybridoma cell lines secreting monoclonal antibodies against protein X4 were screened by regular cell fusion and subcloning approach. The specificities of these monoclonal antibodies were determined by Western blotting and Immunofluorescence assay. RESULTS: Three hybridoma cell lines (8A5, 8H6 and 4A2) stable in secreting specific monoclonal antibodies were successfully obtained. They could bind specifically to protein X4 proved by Western blotting. All of them belonged to the IgG2a isotype proved by antigen mediated ELISA. Indirect Immunofluorescence assay indicated that they could specifically bind to protein X4 expressed in SARS-CoV infected Vero E6 cells. CONCLUSION: Monoclonal antibodies of high specificity against protein X4 have been successfully prepared, which laid the foundation for the further study of protein X4.

Animals↗
PubMed2005

CD8+ T cell depletion amplifies hepatitis C virus replication in peripheral blood mononuclear cells.

We investigated the ability of CD8+ T cells to inhibit hepatitis C virus (HCV) replication in peripheral blood mononuclear cells (PBMCs). PBMCs isolated from 11 of 20 HCV-infected subjects had no detectable HCV RNA. Removal of CD8+ T cells from these PBMCs resulted in detection of HCV RNA, and depletion of CD8+ T cells from PBMCs that had detectable HCV RNA amplified HCV replication. Reconstitution of CD8- PBMCs with autologous CD8+ T cells led to inhibition of HCV replication. Interferon-gamma produced by CD8+ T cells was partially responsible for CD8+ T cell-mediated noncytotoxic anti-HCV activity in PBMCs. This noncytotoxic anti-HCV activity was confirmed in HCV replicon cells. Supernatants from CD8+ T cell cultures inhibited HCV RNA expression in the replicon cells. These findings may have important implications for the immunopathogenesis of HCV in both immune and hepatic cells and are relevant to the development of host innate immunity-based anti-HCV interventions.

Antiviral Agents↗
PubMed2005

Morphine inhibits CD8+ T cell-mediated, noncytolytic, anti-HIV activity in latently infected immune cells.

Opiates have profound effects on the function of human immune cells and are a possible cofactor in the immunopathogenesis of human immunodeficiency virus (HIV) disease. We investigated the impact of morphine on CD8+ T cell-mediated, noncytotoxic, anti-HIV activity in latently infected human immune cells. Morphine inhibited the noncytotoxic, anti-HIV activity of CD8+ T cells in HIV latently infected cells (U1 and J1.1). Naltrexone abrogated the morphine-mediated, inhibitory effect on the noncytotoxic, anti-HIV activity of CD8+ T cells. Interferon-gamma (IFN-gamma), a potent antiviral cytokine produced by CD8+ T cells, was partially responsible for CD8+ T cell-mediated, noncytotoxic, anti-HIV activity. The anti-HIV activity of IFN-gamma was also compromised by morphine treatment. Further, morphine attenuated CD8+ T cell-mediated suppression of the HIV long-terminal repeat promoter activation. Morphine also inhibited CD8+ T cell-induced expression of the signal transducer and activator of transcription-1, an important transcriptional factor in the IFN signaling pathway. These data provide additional evidence to support the notion that opioids play a role in impairing the anti-HIV function of the immune system.

Anti-HIV Agents↗
PubMed2005

A tetracycline-regulatable adeno-associated virus vector for double-gene transfer.

An increasing demand for polycistronic vectors that express multiple genes simultaneously has arisen in recent years to obtain an efficient gene therapy. Armed with the knowledge that the expression of transgene in mammalian cells often requires tight control, we constructed in this study a tetracycline-regulated double-gene adeno-associated virus (AAV) vector carrying green and red fluorescent protein genes and expressed it in PC12 cells. When detected by fluorescence microscope and fluorescence-activated cell sorting, gene expression was induced by 44-66-fold and could be reversibly controlled by doxycycline. This double-gene AAV vector may be useful for regulated expression of two genes or a marker to monitor transgene expression.

Adenoviridae↗
PubMed2005

Hepatitis C virus inhibits intracellular interferon alpha expression in human hepatic cell lines.

The chronicity of hepatitis C virus (HCV) infection raises the question of how HCV is able to persist in hepatic cells. We show that human primary hepatocytes and human hepatic cell lines (Huh7 and HepG2) spontaneously produce interferon (IFN)-alpha that is inhibited in the HCV replicon cells (Huh.8 and FCA-1). Silencing IFN-alpha gene expression by IFN-alpha small interfering RNA (siRNA) in the HCV replicon cells resulted in increased HCV replicon expression. The activation of IFN-alpha expression by interferon regulatory factor (IRF-7) led to the inhibition of HCV replicon expression, whereas the anti-IFN-alpha receptor antibody could partially block IRF-7-mediated HCV replicon inhibition. In addition, the blockade of IFN-alpha receptor by anti-IFN-alpha receptor antibody on the replicon cells increased HCV replicon expression. Among the HCV nonstructural (NS) proteins tested, NS5A is the most potent inhibitor of IFN-alpha expression by the hepatic cells. Investigation of the mechanism of HCV action on IFN-alpha showed that IRF-7-induced IFN-alpha promoter activation was inhibited in the HCV replicon cells. Furthermore, IRF-7 expression was restricted in the HCV replicon cells. In conclusion, we provide direct evidence that HCV undermines the intracellular innate immunity of the target cells, which may account for HCV persistence in hepatic cells.

Cells, Cultured↗
PubMed2005

Morphine withdrawal enhances hepatitis C virus replicon expression.

We previously demonstrated that morphine enhances hepatitis C virus (HCV) replication in human hepatic cells. Here we describe the impact of morphine withdrawal (MW), a recurrent event during the course of opioid abuse, on HCV replicon expression in human hepatic cells. MW enhanced both viral RNA and protein expression in HCV replicon cells. Blocking opioid receptors by treatment with naloxone after morphine cessation (precipitated withdrawal, PW) induced greater HCV replicon expression than MW. Investigation of the mechanism responsible for MW- or PW-mediated HCV enhancement showed that both MW and PW inhibited the expression of endogenous interferon-alpha (IFN-alpha) in the hepatic cells. This down-regulation of intracellular IFN-alpha expression was due to the negative impact of MW or PW on IFN-alpha promoter activation and on the expression of IFN regulatory factor 7 (IRF-7), a strong transactivator of the IFN-alpha promoter. In addition, both MW and PW inhibited the anti-HCV ability of recombinant IFN-alpha in the hepatic cells. These in vitro observations support the concept that opioid abuse favors HCV persistence in hepatic cells by suppressing IFN-alpha-mediated intracellular innate immunity and contributes to the development of chronic HCV infection.

Cell Line↗
PubMed2005

Preparation and characterization of a monoclonal antibody against CKLF1 using DNA immunization with in vivo electroporation.

Chemokine-like factor 1 (CKLF1) is a newly cloned human cytokine from PHA-stimulated U937 cells in our laboratory, which belongs to a novel gene family and has at least three alternative RNA splicing forms: CKLF2, CKLF3, and CKLF4. Former studies demonstrated that CKLF1 has a chemotaxis effect on different leukocytes both in vitro and in vivo. It can also stimulate the proliferation of mouse skeletal muscle cells and bone marrow cells. This study was designed to generate CKLF1-specific monoclonal antibody (MAb) for further exploration of its structure and function. Mice were immunized intramuscularly with naked plasmid DNA encoding CKLF1 gene with in vivo electroporation. Hybridomas were generated by the fusion of the spleenocytes to Sp2/0 myeloma cells. One hybridoma cell line designated as M4, which is stable in secreting anti- CKLF1 MAb, was generated. It belongs to the IgG1 isotype and is specific for the unique C-terminal domain of CKLFs protein. The specificity of this antibody has been assessed by enzyme-linked immunosorbent assay (ELISA) and Western blot. Our results demonstrated that intramuscular injection of naked DNA encoding CKLF1 gene combined with in vivo electroporation is an effective and simple method to raise MAbs that can be used for basic and clinical research of CKLF1 protein.

Animals↗
PubMed2005

Alcohol suppresses IL-2-induced CC chemokine production by natural killer cells.

BACKGROUND: Natural killer (NK) cells are a critical component of the host innate immune system. We investigated whether alcohol impairs NK cell function, particularly production of CC chemokines induced by interleukin (IL)-2, the natural ligands for CCR5 receptor. METHODS: Primary NK cells and NK cell line (YTS) were cultured with or without alcohol (10 to 80 mM) for three hours. The culture supernatants were then harvested and used to treat human peripheral blood monocyte-derived macrophages and a HeLa cell line, which expresses CD4, CCR5, and CXCR4 receptors (MAGI cells). CC chemokine expression by YTS and primary NK cells treated with or without alcohol was analyzed with the real-time RT-PCR and ELISA. [Ca(2)(+)]i and Western blot assays were used to determine calcium-mediated intracellular signaling pathway and NF-kappaB p65 expression. HIV strains (Bal and UG024) were used to infect macrophages and MAGI cells. In addition, ADA (macrophage-tropic strain) and murine leukemia virus (MLV) envelope-pseudotyped HIV infection was carried out in macrophages. HIV infectivity was determined by HIV reverse transcriptase (RT) and beta-galactosidase activity assays. RESULTS: Alcohol inhibited IL-2-induced CC chemokine (CCL3 and CCL4) expression by NK cells. Functional tests demonstrated that this reduced expression of CC chemokines was associated with diminished anti-HIV ability of NK cells. Alcohol also reduced the ability of NK cells to response to CCL3-mediated chemotaxis. Alcohol inhibited IL-2-induced NF-kappaB p65 protein expression and calcium mobilization by NK cells. CONCLUSIONS: Alcohol, through the inhibition of IL-2-induced NF-kappaB p65 protein expression and intracellular calcium mobilization, suppressed NK cell production of CC chemokines. This suppression of CC chemokine production was associated with diminished anti-HIV activity of NK cells. Thus, by inhibiting NK cell-mediated innate immunity against HIV, alcohol consumption may have a cofactor role in the immunopathogenesis of HIV disease.

Adult↗
PubMed2005

[Mutations in exon 7 of the phenylalanine hydroxylase (PAH) gene in chinese patients with phenylketonuria].

To study mutation in exon 7 of the gene for the phenylalanine hydroxylase(PAH), the mutations in exon 7 and flanking sequence of PAH gene were detected by means of SSCP analysis and DNA sequencing, in 147 unrelated Chinese children with phynelketonuria and their parents. Thirteen different mutations, including 11 missense, 1 deletion and 1 splice mutation, were revealed in 90/294 mutant alleles (30.61%). The prevalent mutations were R243Q (22.8%) and Ivs7nt2t->a (2.38%). Seven novel mutations were identified: G239D, R241fsdelG, G247S, E280G, L255S, R261Q, P281L. These new mutations have not been described in Chinese PKU population and the first 4 mutants have not been reported and thus been submitted to www.pahdb,mcgill.ca. The missense was the most common type. The deletion and frameshift mutations were detected for the first time in Chinese PKU population. This study showed the mutation characteristics and their distribution in exon 7 of PAH gene and proved that the exon 7 was the hot region of PAH gene mutation in Chinese PKU population .

Adult↗
PubMed2005

[NEMO Delta 4-10 deletion of NEMO gene in Chinese incontinentia pigmenti cases].

OBJECTIVE: Incontinentia pigmenti (IP) is a rare X-linked dominant disorder that affects ectodermal tissues. In IP, mutations in NEMO lead to the complete loss of NF-kB activation creating a susceptibility to cellular apoptosis in response to TNF-alpha. Recently, a second nonfunctional copy of the gene, Delta NEMO was identified, opposite in direction to NEMO. Almost 90% of IP whose gene mutation type had been recognized have a recurrent genomic deletion of exons 4-10 of the NEMO (IKK gamma) gene, called NEMO Delta 4-10, which is necessary to activate the NF-kB pathway. Therefore, PCR-based detection of the NEMO deletion is a diagnostic measurement for IP. This study sought to analyze the NEMO Delta 4-10 deletion in NEMO gene of Chinese IP cases. METHODS: Seven IP cases and part of their families totally 15 persons were enrolled in this study. The 7 IP cases were aged 41 days to 8 years. Among them 1 was male and 6 were female. Four cases had family history of IP, the other 3 were sporadic cases. Fifty healthy children without any congenital diseases were taken as normal control group. According to the gene characteristics of IP, by PCR measurement NEMO Delta 4-10 deletion in NEMO gene was tested with specific primers In2/JF3R, and NEMO Delta 4-10 deletion in pseudogene Delta NEMO was checked out by primers Rev-2/JF3R. RESULTS: Five out of the 7 tested cases (case 1, 2, 3, 4, and 6) showed NEMO Delta 4-10 deletion in NEMO gene. The mothers of case 1 and case 6, 1a and 6a, also suffered from this disease, and their results were just the same as their daughters. For pseudogene Delta NEMO only case 2 and case 4 were proved having NEMO Delta 4-10 deletion, while other cases and families had negative results. For normal control group, NEMO Delta 4-10 deletion was not found either in NEMO gene or in their pseudogene Delta NEMO. CONCLUSION: Incontinentia pigmenti in most cases were caused by NEMO Delta 4-10 deletion in NEMO gene.

Child↗
PubMed2005

[Molecular identification of medicinal plants: Dendrobium chrysanthum, Dendrobium fimbriatum and their morphologically allied species by PCR-RFLP analyses].

AIM: To establish a simple method for molecular identification of original plants of D. chrysanthum and D. fimbriatum using molecular marker rDNA ITS region. METHODS: Restriction patterns of ITS fragments were obtained using PCR-RFLP method. The PCR products of D. chrysanthum and its morphologically allied species were digested at 37 degrees C by Cla I and Apa LI, those of D. fimbriatum and its morphologically allied species were digested by Sph I. RESULTS: D. chrysanthum, D. fimbriatum and their morphologically allied species could be identified by predicted restriction profiles of PCR-RFLP. The botanical origin of twenty-five fresh samples of "Shihu" collected in markets was identified by this method. CONCLUSION: The results showed that PCR-RFLP analysis of the rDNA ITS region is a feasible, simple and inexpensive method for determining the botanical origin of the traditional Chinese medicine "Shihu".

DNA, Plant↗
PubMed2004

Chrysin and its phosphate ester inhibit cell proliferation and induce apoptosis in Hela cells.

To improve the biological activities of chrysin (CR), we synthesize Diethyl Chysin-7-yl phosphate (CPE: C(19)H(19)O(7)P) and tetraethyl bis-phosphoric ester of chrysin (CP: C(23)H(28)O(10)P(2)) through a simplified Atheron-Todd reaction. The interactions of the CR and CPE with lysozyme were explored by electrospray ionization mass spectrometry (ESI) and fluorescence spectrometry method. Experimental results indicate that CPE could form the noncovalent compound with lysozyme, while the interaction of the CR with lysozyme was not detected. In addition, whether and how the compounds CPE and CP affect proliferation and apoptosis in human cervical cancer Hela cells were investigated. Moreover, the effects of CPE and CP in Hela cells were compared with that of the nonmodified CR compound. The Hela cells were co-cultured with CR, CP, and CPE as experimental groups, respectively, and corresponding control groups treated without CR, CP, and CPE. The proliferation and apoptosis were detected using MTT assay, HCl denatured-methyl green-pyronin staining, PCNA immunohistochemistry and TUNEL techniques. The cell growth IC(50), relative absorbance (RA), proliferating index (PI), PCNA-IR (immunoreactivity IR) integration value (IV), and apoptosis index (AI) were calculated and their correlation was analyzed in each group. The results show that all CR, CP, and CPE could inhibit proliferation and induce apoptosis in Hela cells. Moreover, the effects of CP and CPE were more potent than that of CR. The CP and CPE were proved to be a kind of stronger apoptosis inducers than nonphosphated CR. There was a negative correlation between proliferation and apoptosis. In conclusion, the CR, CP, and CPE could effectively inhibit growth by down-regulated expression of PCNA, and induce apoptosis in Hela cells. The efficiency of the modified CP and CPE preceded nonmodified CR compounds. The CP and CPE may be a new potential anti-cancer drug for therapy of human cervical carcinoma.

Antineoplastic Agents↗
PubMed2004

Natural killer cells inhibit hepatitis C virus expression.

Natural killer (NK) cells are critical in host innate defense against certain viruses. The role of NK cells in controlling hepatitis C virus (HCV) remains obscure. We examined whether NK cells are capable of inhibiting HCV expression in human hepatic cells. When NK cells are cultured with the HCV replicon-containing hepatic cells, they have no direct cytolytic effect but release soluble factor(s) suppressing HCV RNA expression. Media conditioned by NK cell lines (NK-92 and YTS) or primary NK cells isolated from healthy donors contain interferon gamma (IFN-gamma) and potently inhibit HCV RNA expression. Ligation of CD81 on NK cells inhibits IFN-gamma production and results in decreased anti-HCV activity. In addition, the antibodies to IFN-gamma or IFN-gamma receptors abolish the anti-HCV activity of NK cell-conditioned media. The role of IFN-gamma in NK cell-mediated, anti-HCV activity is supported by the observation that NK cell-conditioned media enhanced expression of signal transducer and activator of transcription-1, a nuclear factor that is essential in IFN-gamma-mediated antiviral pathways. NK cell-conditioned media have the ability to stimulate intracellular IFN-alpha expression in the hepatic cells, suggesting a mechanism responsible for NK cell-mediated, anti-HCV activity. Thus, NK cells hold the potential to play a vital role in controlling HCV replication in hepatic cells using an IFN-gamma-dependent mechanism.

Antigens, CD↗
PubMed2004

Exploratory spatial data analysis for the identification of risk factors to birth defects.

BACKGROUND: Birth defects, which are the major cause of infant mortality and a leading cause of disability, refer to "Any anomaly, functional or structural, that presents in infancy or later in life and is caused by events preceding birth, whether inherited, or acquired (ICBDMS)". However, the risk factors associated with heredity and/or environment are very difficult to filter out accurately. This study selected an area with the highest ratio of neural-tube birth defect (NTBD) occurrences worldwide to identify the scale of environmental risk factors for birth defects using exploratory spatial data analysis methods. METHODS: By birth defect registers based on hospital records and investigation in villages, the number of birth defects cases within a four-year period was acquired and classified by organ system. The neural-tube birth defect ratio was calculated according to the number of births planned for each village in the study area, as the family planning policy is strictly adhered to in China. The Bayesian modeling method was used to estimate the ratio in order to remove the dependence of variance caused by different populations in each village. A recently developed statistical spatial method for detecting hotspots, Getis's 7, was used to detect the high-risk regions for neural-tube birth defects in the study area. RESULTS: After the Bayesian modeling method was used to calculate the ratio of neural-tube birth defects occurrences, Getis's statistics method was used in different distance scales. Two typical clustering phenomena were present in the study area. One was related to socioeconomic activities, and the other was related to soil type distributions. CONCLUSION: The fact that there were two typical hotspot clustering phenomena provides evidence that the risk for neural-tube birth defect exists on two different scales (a socioeconomic scale at 6.84 km and a soil type scale at 22.8 km) for the area studied. Although our study has limited spatial exploratory data for the analysis of the neural-tube birth defect occurrence ratio and for finding clues to risk factors, this result provides effective clues for further physical, chemical and even more molecular laboratory testing according to these two spatial scales.

Bayes Theorem↗
PubMed2004

The nature of phosphorylated chrysin-protein interactions involved in noncovalent complex formation by electrospray ionization mass spectroscopy.

In the work described in this paper, chrysin was phosphorylated by a modified Atheron-Todd reaction. The structure of phosphorylated chrysin was determined by elemental analysis, NMR, ESI-MS/MS, and X-ray data. Electrospray ionization results showed that the phosphorylated flavonoids could form noncovalent complexes with many proteins, such as lysozyme, myoglobin, bovine insulin, and cytochrome c, while noncovalent complexes were not detected in the mixed solution of the chrysin and proteins. The research shows that the phosphorylated flavonoids possess relatively stronger affinities and form noncovalent complexes with the proteins more easily than the non-phosphorylated compounds.

Animals↗
PubMed2004

EDR2 film dosimetry for IMRT verification using low-energy photon filters.

Recently the EDR2 (extended dose range) film has been introduced commercially for applications in radiation therapy dosimetry. In addition to characterizing the wide dynamic range, several authors have reported a reduced energy dependence of this film compared to that of X-Omatic Verification (XV) films for megavoltage photon beams. However, those investigations were performed under limited geometrical conditions. We have investigated the dosimetric performance of EDR2 film for the verification of IMRT fields at more clinically relevant conditions by comparing the film doses with the doses measured with an ion chamber and XV films. The effects of using a low energy scattered photon filter on EDR2 film dosimetry was also studied. In contrast to previous reports our results show that EDR2 film still exhibits considerable energy dependence (a maximum discrepancy of 9%, compared with an ion chamber) at clinically relevant conditions (10 cm depth for IMRT fields). However, by using the low-energy filters the discrepancy is reduced to within 3%. Therefore, EDR2 film, in combination with the filters, is found to be a promising two-dimensional dosimeter for verification of IMRT treatment fields.

Equipment Failure Analysis↗
PubMed2004

Compare two methods of measuring DNA damage induced by photogenotoxicity of fluoroquinolones.

AIM: To compare two methods of measuring DNA damage induced by photogenotoxicity of fluoroquinolones (FQ). METHODS: Lomefloxacin (LFLX), sparfloxacin (SPFX), ciprofloxacin (CPFX), and levofloxacin (LELX) were tested by comet assay and photodynamic DNA strand breaking activity under the different conditions of UVA irradiation. RESULTS: In comet assay, photogenotoxicity was evident at SPFX 1 mg/L, LFLX 5 mg/L, and CPFX 5 mg/L, and LELX 10 mg/L. In photodynamic DNA strand-breaking activity, SPFX and LFLX induced the conversion of the supercoiled form into the nicked relaxed form at 10-50 micromol/L, while CPFX at 25 micromol/L and LELX at 50 micromol/L. CONCLUSION: There were good correlations between the two methods to detect DNA damage induced by phototoxicity of fluoroquinolones. Photodynamic DNA strand breaking activity was a good method to detect DNA damage induced by photogenotoxicity of fluoroquinolones as well as comet assay.

Animals↗