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Biomedical subjects

R Sun

Publications and source records attributed to R Sun.

At least 37 records · Page 2Linked to original sources
PubMed2001

Microencapsulated bovine chromaffin cell xenografts into hemiparkinsonian rats: a drug-induced rotational behavior and histological changes analysis.

Bovine chromaffin cells were microencapsulated within alginate-polylysine-alginate (APA) membranes. Microencapsulated bovine chromaffin cells as well as unencapsulated cells and empty microcapsules were grafted into the brain of hemiparkinsonian rats with 6-hydroxydopamine (6-OHDA) lesions. Apomorphine-induced rotational behavior of the host animals and the survival of the grafted chromaffin cells were examined after transplantation. The animals receiving microencapsulated bovine chromaffin cells showed a significant decrease (17.6--35.6%) in apomorphine-induced rotation 1 week postimplantation that remained stable for the 10 month test period. Fluorescent histochemistry further revealed that microencapsulation increased the chromaffin cell survival with only a minimum host reaction for up to 10 months posttransplantation while the survival of free, unencapsulated chromaffin cells was only modest and was accompanied by a large inflammatory response. The reduction of apomorphine-induced rotations was correlated with the survival of bovine chromaffin cells in the host brain. The data indicate that encapsulation of bovine chromaffin cells in APA membranes reduces the host immune response to the xenograft and prolongs the viability of the grafted cells.

Animals↗
PubMed2001

Performance characteristics of the COBAS AmpliScreen HIV-1 test, version 1.5, an assay designed for screening plasma mini-pools.

BACKGROUND: The COBAS AmpliScreen HIV-1 test, version 1.5 (v1.5) (Roche Molecular Systems), is designed for screening pools composed of samples from 24 individual units of blood or plasma. A specimen-processing procedure (Multiprep) simultaneously concentrates and extracts HIV-1, HCV, and HBV particles from plasma and incorporates an HIV-1 internal control (IC) RNA. Processed samples are amplified by RT-PCR using HIV-1-specific primers and detected by hybridization of the amplified products to HIV-1- and IC-specific oligonucleotide probes. STUDY DESIGN AND METHODS: Plasma samples containing known quantities of HIV-1 were used to evaluate analytical sensitivity and precision and to validate a pool testing algorithm. Analytical specificity was evaluated by adding various viruses and bacteria to HIV-1-negative plasma. Seroconversion panels were tested to estimate the window-period reduction achieved by RNA testing. RESULTS: The analytical sensitivity of the test (concentration that yields > or = 95% positive results in a set of replicate tests) was 25 copies of HIV-1 RNA per mL of pooled plasma. Representative strains from all HIV-1 group M subtypes were reproducibly detected (> 95% positive results) at concentrations of 20 to 200 viral particles per mL. The test did not cross-react with a set of 31 viral and 5 bacterial isolates, and it yielded negative results on a panel of 500 blood samples from HIV-1-seronegative donors. Plasma samples containing abnormally high levels of Hb, albumin, triglycerides, or bilirubin did not interfere with the test. HIV-1 RNA was detected 2 to 14 days before HIV-1 antibody and 0 to 28 days before p24 antigen. The test specifically detected pools containing a single positive unit with 2400 HIV-1 RNA copies per mL and correctly identified the positive unit. CONCLUSION: The COBAS AmpliScreen HIV-1 test, v1.5, has sufficient sensitivity to detect a single infected unit containing 600 copies of HIV-1 per mL in a pool with 23 uninfected units and should reduce the window period between infection and seroconversion by at least 2 to 14 days.

Acquired Immunodeficiency Syndrome↗
PubMed2001

Function of Rta is essential for lytic replication of murine gammaherpesvirus 68.

Rta, encoded primarily by open reading frame 50, is well conserved among gammaherpesviruses. It has been shown that the Rta proteins of Epstein Barr virus (EBV), Kaposi's sarcoma-associated herpesvirus (KSHV, or HHV-8), and murine gammaherpesvirus 68 (MHV-68; also referred to as gamma HV68) play an important role in viral reactivation from latency. However, the role of Rta during productive de novo infection has not been characterized in gammaherpesviruses. Since there are cell lines that can support efficient productive de novo infection by MHV-68 but not EBV or KSHV, we examined whether MHV-68 Rta plays a role in initiating viral lytic replication in productively infected cells. Rta, functioning as a transcriptional activator, can activate the viral promoter of early lytic genes. The amino acid sequence alignments of the Rta homologues suggest that the organizations of their functional domains are similar, with the DNA binding and dimerization domains at the N terminus and the trans-activation domain at the C terminus. We constructed two mutants of MHV-68 Rta, Rd1 and Rd2, with deletions of 112 and 243 amino acids from the C terminus, respectively. Rd1 and Rd2 could no longer trans-activate the promoter of MHV-68 gene 57, consistent with the deletions of their trans-activation domains at the C terminus. Furthermore, Rd1 and Rd2 were able to function as dominant-negative mutants, inhibiting trans-activation of wild-type Rta. To study whether Rd1 and Rd2 blocked viral lytic replication, purified virion DNA was cotransfected with Rd1 or Rd2 into fibroblasts. Expression of viral lytic proteins was greatly suppressed, and the yield of infectious viruses was reduced up to 10(4)-fold. Stable cell lines constitutively expressing Rd2 were established and infected with MHV-68. Transcription of the immediate-early gene, rta, and the early gene, tk, of the virus was reduced in these cell lines. The presence of Rd2 also led to attenuation of viral lytic protein expression and virion production. The ability of Rta dominant-negative mutants to inhibit productive infection suggests that the trans-activation function of Rta is essential for MHV-68 lytic replication. We propose that a single viral protein, Rta, governs the initiation of MHV-68 lytic replication during both reactivation and productive de novo infection.

Animals↗
PubMed2001

Transcription activation of polyadenylated nuclear rna by rta in human herpesvirus 8/Kaposi's sarcoma-associated herpesvirus.

Human herpesvirus 8 (HHV-8) (also known as Kaposi's sarcoma-associated herpesvirus) encodes a novel noncoding polyadenylated nuclear (PAN) RNA (also known as T1.1 or nut-1) during the early phase of lytic replication. PAN RNA is the most abundant transcript of HHV-8, comprising 80% of total poly(A)-selected transcripts in HHV-8-infected cells during lytic replication. We directly measured the abundance of PAN RNA by visualizing 1.1- to 1.2- kb PAN RNA in an ethidium bromide-stained gel from poly(A)-selected RNA. We further pursued the mechanisms by which PAN RNA expression is induced to such high levels. rta, an immediate-early gene of HHV-8, is a transactivator that is sufficient and necessary to activate lytic gene expression in latently infected cells. Ectopic expression of Rta was previously shown to induce PAN RNA expression from the endogenous viral genome and activate the PAN promoter in a reporter system. Here, we have identified the Rta-responsive element (RRE) in the PAN promoter. Deletion analysis revealed that the RRE is present in a region between nucleotides -69 and -38 of the PAN promoter. A promoter construct containing the 69 nucleotides upstream of the transcription start site of the PAN promoter was activated by Rta in the absence or presence of the HHV-8 genome. Rta activated the PAN promoter up to 7,000-fold in 293T cells and 2,000-fold in B cells. Electrophoretic mobility shift assays demonstrated that Rta formed a highly stable complex with the RRE of the PAN promoter. Our study suggests that Rta can induce PAN RNA expression by direct binding of Rta to the RRE of the PAN promoter. This study has highlighted an important mechanism controlling PAN RNA expression and also provides a model system for investigating how Rta transactivates gene expression during lytic replication.

Animals↗
PubMed2001

[Purification and biological activity of rh-leptin expressed in Escherichia coli].

The human leptin was successfully expressed with high level in E. coli under the control of PL promotor. The yield of recombinant protein was over 40% of total cellular protein and expressed as inclusion bodies. The recombinant human leptin (rh-leptin) was purified with gel filtration, anion-exchange and reverse chromatography. Refolding was achieved by gradually reducing denaturant using a diafiltration method. The refolded rh-leptin was characterized by SDS-PAGE, Western-blotting and its first 15 amino acid residues sequence of the N-terminal. The purified product was found to be biologically active, reducing the food intake and body weight gain upon testing in BALB/c mice.

Animals↗
PubMed2001

[Study of the cytotoxity against human hepatocellular carcinoma cells induced by the MAGE-1 gene modified dendritic cells].

OBJECTIVE: To genetically modify dendritic cells (DC) with a tumor associated antigen gene MAGE-1 and to observe in vitro the cytotoxic effect induced by this genetically modified DC against human hepatocellular carcinoma (HCC) cell line SMMC7721, thus giving a primary verification of the possibility of this genetically modified DC to induce specific antitumor immunity against HCC and serving as a new type of vaccine. METHODS: The MAGE-1 gene was inserted into the retrovirus vector LXSN to construct the recombinant retrovirus LMSN. The monocyte-derived DCs were transfected by LMSN and control virus LXSN respectively as well as a third group was set up as non-transfected control. The MAGE-1 gene expression in LMSN transfected DC was identified by Western blot and the in vitro cytotoxities against SMMC7721 induced by three groups of DC were tested by MTT assay. RESULTS: A recombinant retrovirus LMSN containing the tumor rejection antigen gene MAGE-1 was successfully constructed and the MAGE-1 gene expression in LMSN transfected human monocyte-derived DC was induced. The LMSN transfected DC could induce very high lytic activity against SMMC7721 in vitro with the lytic activity of 78.9%+/-3.6%. Nevertheless, the LXSN transfected DC and non-transfected DC only induced relatively lower lytic activity as 34.7%+/-4.3% and 3.9%+/-2.0%, respectively. The difference of the lytic activities between those three groups as statistically significant (P<0.01). CONCLUSIONS: The MAGE-1 gene transfected DC can induce higher in vitro cytotoxicity against SMMC7721, suggesting the possibility of this genetically modified DC to induce specific antitumor activity and to serve as a new type of vaccine for HCC.

Animals↗
PubMed2001

[Effects of interferon-alpha on recurrence and metastasis of hepatocellular carcinoma after curative resection in nude mice].

OBJECTIVE: To study the effect of interferon-alpha (IFN-alpha) on the recurrence and metastasis of hepatocellular carcinoma (HCC) in nude mice, and to clarify if there is synergistic effect treated by combination of IFN-alpha and all-trans retinoic acid (ATRA). METHODS: The effect of IFN-alpha and/or ATRA on the proliferation of HCC cell lines was measured in vitro. The metastatic model of human HCC in nude mice LCI-D20 was used in present study. Curative resection was performed at 10th day after implantation in 44 nude mice. Drugs were given at the next day after resection. IFN-alpha was administered subcutaneously at doses of 3+/-10(5) U/day, 6+/-10(5) U/day, respectively. ATRA was administered p.o. at a dose of 20 mg/kg/day, and IFN-alpha 6+/-10(5)U/day combined with ATRA 20 mg/kg/day. The mice were sacrificed 35 days after treatment. The recurrent tumor size was measured and the presence of intrahepatic dissemination and lung metastases were recorded. RESULTS: The effect of IFN-alpha and/or ATRA on the proliferation of HCC cells SMMC7721, BEL-7402, BEL-7405, and MHCC97 was not obvious. The combination of IFN-alpha and ATRA had no synergistic effect in vitro. The lung metastatic rate, the liver recurrent rate, the size of main recurrent lesions, the number of intrahepatic disseminating nodules and the largest disseminating nodule of the controlled group was 100%(12/12), 100%(12/12), (1346.3+/-4.2 ) mm(3), 8.2+/-4.4, 864 mm(3), respectively; whereas it was 0, 87.5%(7/8), (8.7+/-2.9) mm(3), 2.3+/-0.6, and 7.8mm(3), respectively in the IFN-alpha 3+/-10(5) U/day treated group(P<0.05); 0, 12.5%(1/8), 0.5mm(3), 2, 0.5 mm(3) in the IFN-alpha 6+/-10(5)U/day treated group(P<0.05); 0, 12.5%(1/8), 1 mm(3), 2.5+/-0.7, 8 mm(3) in the IFN-alpha 3+/-10(5) U/day and ATRA treated group(P<0.05); 87.5%, 100%, (1472.6+/-5.6) mm(3), 7.3+/-3.8, 768.5 mm(3) in the ATRA treated group (P>0.05). CONCLUSIONS: IFN-alpha has an inhibitory effect on intrahepatic recurrence and lung metastasis of human HCC after curative resection in nude mice, and the effect is enhanced with increasing dose. IFN-alpha and ATRA have no synergistic effect according to in vivo and in vitro test. ATRA has no effect on recurrence and metastasis of HCC.

Animals↗
PubMed2001

[Significance of the unbalanced expression of Th1/Th2 type cytokines in human glioma].

OBJECTIVE: To study the significance of the unbalanced expression of Th1/Th2 type cytokines in human glioma. METHODS: The gene expressions of Th1/Th2 type cytokines in 62 specimens of human glioma tissues, 4 glioma cell lines, peripheral blood mononuclear cell (PBMC) of 15 glioma patients, 5 specimens of normal adult brain tissue and 5 brain meningioma tissues were detected by semiquantitative reverse transcription polymerase chain reaction. IFN-gamma and IL-2 represent Th1 type cytokines. IL-4, IL-6, IL-10 and IL-13 represent Th2 type cytokines. RESULTS: There were obviously predominant expression of Th2 type cytokines in glioma cell lines (P < 0.01) and specimens of human glioma tissues (P < 0.01). The tendency of distinct expression of Th2 type cytokines in PBMC was also existent. There wasn't obvious discrepancy of the expression of two type cytokines in normal adult brain tissues and meningioma tissues. CONCLUSIONS: It is likely that the switching of Th1/Th2 type cytokines in gliomas as predominant expression of Th2 type cytokine genes is related to the origination of gliomas and the evasion of glioma cells from immune surveillance.

Adolescent↗
PubMed2001

[Influence of adjustment of balance of Th1/Th2 type cytokines on proliferation of glioma cells].

OBJECTIVE: To study the influence of adjustment of balance of Th1/Th2 by external cytokines on proliferation of glioma cells. METHODS: The gene expressions of Th1/Th2 type cytokines in C6, 9L, U251 and SHG44 glioma cells were detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). After the cells were induced with IFN-gamma + IL-4 McAb and IL-4 + IFN-gamma McAb respectively, we isolated the total RNA to proceed RT-PCR again. The evaluation of cell proliferation was proceeded by MTT assay method. RESULTS: There was obviously predominant expression of Th2 type cytokines in glioma cell lines (P < 0.01). The expression intensity of IFN-gamma was improved in IFN-gamma + IL-4 McAb groups and Th2 type cytokines were enhanced in IL-4 + IFN-gamma McAb groups. IFN-gamma and IL-4 McAb could cause the switch from Th2 to Th1, and could remarkably inhibit the proliferation of glioma cells in a dose-dependent way (P < 0.01). On the other hand, IL-4 and IFN-gamma McAb could strengthen the switch of Th2, and might stimulate the glioma cell growth, also in a dose-dependent way (P < 0.01). CONCLUSIONS: There is a Th2 preponderance in glioma cells. IFN-gamma and IL-4 McAb could regulate the switch from Th2 to Th0 or Th1, and inhibit the proliferation of glioma cells.

Antibodies, Monoclonal↗
PubMed2001

[Differentiation of natural killer cells into two functional subsets: NKh1 and NKh2].

OBJECTIVE: To verify the presence of functional subsets of natural killer cells based on the cytokine production. METHODS: NK cells were purified and cultured in complete RPMI1640 medium in the presence of either IFN gamma + anti-IL-4(classical Th1 polarization) or IL-4 + anti-IFN gamma (classical Th2 polarization) for three days, and then were collected and detected for type I/type II cytokines by RT-PCR method. RESULTS: NK cells were purified from 15 healthy donors, over 70% purity of NK cells were determined by flow cytometry. NK cells in peripheral blood expressed high level of type I cytokines, mainly IFN gamma, but low level of type II cytokines such as IL-10 and IL-13, IL-4 was not produced by NK cells. Cells cultured in IFN gamma + anti-IL-4 condition exhibited significantly increased level of IFN gamma, unchanged IL-2, and decreased type II cytokines. Cells grew in IL-4 + anti-IFN gamma condition exhibited increased IL-10 and IL-13, and decreased IFN gamma expressions. CONCLUSIONS: Based on the cytokine production, NK cells may be divided into two functional subsets in the same manner as that of T lymphocytes(e.g. Th1/Th2): NKh1 and NKh2. The biological characterization and phenotypic marker are under investigate.

Cells, Cultured↗
PubMed2001

[The Raman spectra study on crystal-octadecyl hydrogen succinate crystal].

Raman spectra of crystal-octadecyl hydrogen succinate (OHS), whose geometric disposition of the experiment has y(zx)y, y(zz)y, and y(xz)y,y(xx)y respectively, have been recorded and analyzed with the purpose of understanding the structure of OHS crystal. It shows that some of the Raman-active modes of alkyl chain appear polarization. The analysis for crystal structure indicates that the molecular of OHS arrange in layers along [100] and [001] by the mode of dimers, and the molecular layers of chain pile up along [010]. The other groups of OHS corresponding Raman vibration peaks have been analyzed as well.

English Abstract↗
PubMed2001

[ATR-FTIR studies on the packing film].

FTIR and ATR-FTIR of the packing film have been studied. The dimension of thickness is about 0.034 m/m, the right and wrong sides of this packing film is composed of polypropylene and polyethylene glycol terephthalate, respectively.

English Abstract↗
PubMed2000

Dihydrofolate reductase from Kaposi's sarcoma-associated herpesvirus.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the first human virus known to encode dihydrofolate reductase (DHFR), an enzyme required for nucleotide and methionine biosynthesis. We have studied the purified KSHV-DHFR enzyme in vitro and analyzed its expression in cultured B-cell lines derived from primary effusion lymphoma (PEL), an AIDS-associated malignancy. The amino acid sequence of KSHV-DHFR is most similar to human DHFR (hDHFR), but the viral enzyme contains an additional 23 amino acids at the carboxyl-terminus. The viral DHFR, overexpressed and purified from E. coli, was catalytically active in vitro. The K(m) of KSHV-DHFR for dihydrofolate (FH(2)) was 2.4 microM, which is significantly higher than the K(m) of recombinant hDHFR (rhDHFR) for FH(2) (390 nM). K(m) values for NADPH were similar for the two enzymes, about 1 microM. KSHV-DHFR was inhibited by folate antagonists such as methotrexate (K(i): 200 pM), aminopterin (K(i): 610 pM), pyrimethamine (K(i): 29 nM), trimethoprim (K(i): 2.3 microM), and piritrexim (K(i): 3.9 nM). In all cases, K(i) values for these folate antagonists were higher for KSHV-DHFR than for rhDHFR. The viral enzyme was expressed at levels two- to tenfold higher than hDHFR in PEL cell lines as an early lytic cycle gene. KSHV-DHFR mRNA and protein appeared from 6 to 24 h after chemical induction of the KSHV lytic cycle. Epitope-tagged KSHV-DHFR and rhDHFR both localized to the nucleus of transfected cells, while other KSHV nucleotide metabolism genes localized to the cytoplasm. DHFR activity was not essential for viral replication in cultured PEL cells. Since hDHFR was not detectable in peripheral blood mononuclear cells (PBMCs), KSHV-DHFR may function to provide increased DHFR activity in vivo in infected cells that have little or none of their own enzyme.

Amino Acid Sequence↗
PubMed2000

Balance of mass, momentum, and energy in splintering central collisions for 40Ar up to 115 MeV /Nucleon

For central collisions of (17-115)A MeV 40Ar+Cu, Ag, Au, an overall balance is determined for the average mass, energy, and longitudinal momentum. Light charged particles and fragments are separated into forward-focused and isotropic components in the frame of the heaviest fragment. Energy removal by the isotropic component reaches 1-2 GeV. For such high deposition energies, statistical multifragmentation models predict much more extensive nuclear disassembly than is observed.

Journal Article↗
PubMed2000

Immunological effects of BCG as an adjuvant in autologous tumor vaccines.

The role of Bacillus Colmette-Guérin (BCG) as an adjuvant in autologous tumor vaccines was examined. In nine patients with renal cell cancer, irradiated tumor cells alone (wild-type, WT) or with BCG were inoculated intradermally into contralateral thighs. Seven to 10 days later, the draining vaccine-primed lymph nodes (WT-VPLN and BCG-VPLN) were excised. BCG increased the number of harvested VPLN cells by 10-fold (mean +/- SE = 61.8 +/- 20.6/x10(-7)/patient). BCG-VPLN had significantly greater percentages of CD3(+) and CD4(+) T cells compared to WT-VPLN. Both groups of VPLN cells were activated in vitro with anti-CD3 or anti-CD3/CD28 mAbs followed by expansion in IL-2. Anti-CD3/CD28 activation resulted in greater expansion of CD4(+) T cells compared to anti-CD3. After activation, VPLN cells were stimulated with irradiated autologous tumor targets and cytokines (IFN-gamma, GM-CSF, IL-10) released into the supernatants were measured 24 h later. Anti-CD3/CD28-activated BCG-VPLN cells were found to have a greater release of IFN-gamma compared with that of WT-VPLN cells, which was not observed significantly with IL-10 or GM-CSF. BCG resulted in increased VPLN cell yield as well as enhanced type 1 (IFN-gamma release) immune responses of VPLN cells to autologous tumor without upregulating type 2 (IL-10 release) responses. Anti-CD3/CD28 was superior to anti-CD3 activation in this cellular response.

Adjuvants, Immunologic↗
PubMed2000

Gene gun application in the generation of effector T cells for adoptive immunotherapy.

We utilized the gene gun to transfect subcutaneous D5 melanoma and MT-901 mammary carcinoma tumors in situ with a granulocyte/macrophage-colony-stimulating factor (GM-CSF) plasmid complexed to gold particles. There was diminished tumor growth following bombardment with GM-CSF plasmid, which was apparent only during the period of administration. Transgenic GM-CSF was produced by the skin overlying the tumors and not by the tumors themselves. GM-CSF plasmid bombardment resulted in increased cell yields within tumor-draining lymph nodes (TDLN) with at least a 12-fold increase in the percentage of dendritic cells (8.9%) compared to controls (0. 7%). Secondarily activated TDLN cells from animals transfected with GM-CSF demonstrated enhanced cytokine release (interferon gamma, GM-CSF and interleukin-10) in response to tumor stimulator cells compared to controls, and had an increased capacity to mediate tumor regression in adoptive immunotherapy. There was a small, but detectable, non-specific immune adjuvant effect observed with gold particle bombardment alone, which was less than with GM-CSF plasmid. The adjuvant effect of GM-CSF plasmid required peri-tumoral transgene expression since gene bombardment away from the tumor was ineffective.

Animals↗
PubMed2000

Application of membrane-based dendrimer/DNA complexes for solid phase transfection in vitro and in vivo.

In this study a general description of the use of solid support membranes as the device for DNA delivery mediated by PAMAM dendrimers is presented. In contrast to the other DNA carriers, dendrimer/DNA complexes retain the ability to transfect after drying, which enabled coating or incorporation of complexes into poly(DL-lactide-co-glycolide) or collagen-based bioerodable membranes. These studies provide support for the use of this technology for in vitro and in vivo transfection of skin cells. Expression of luciferase or green fluorescent protein from pCF1-Luc and pEGFP1 plasmids indicated that dendrimer/DNA complexes can mediate transfection after dissociation from the solid support and/or when retained on the surface of the membranes. Modification of the membranes by incorporation of an anionic lipid, phosphatidyl glycerol (PG) at 1-5% concentrations, resulted in more efficient in situ transfection, particularly with dendrimer/DNA complexes formed at the low charge ratios (1-5). We also report data supporting the feasibility of membrane-based dendrimer/DNA complexes, particularly formed at lower than neutralizing conditions, for topical in vivo delivery of DNA to hairless mouse skin.

Administration, Topical↗