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Biomedical subjects

H Lin

Publications and source records attributed to H Lin.

At least 325 records · Page 18Linked to original sources
PubMed1997

[Cloning and expression of heavy- and light-chain variable region genes of monoclonal antibody specific for human fibrin].

OBJECTIVE: To obtain the minimum molecule having activity to bind human fibrin. METHODS: The immunoglobulin heavy- and light-chain variable region (VH and Vkappa) genes were isolated from 8E5 hybridoma cells, which secreted a monoclonal antibody against human fibrin, by RT-PCR. An expression vector pOPE51-8E5 was constructed for the recombinant VH-Vkappa expression. The transformed E. coli JM 109 cells were propagated and induced by IPTG. RESULTS AND CONCLUSION: Expression product was found in the periplasmic space and inclusion bodies by SDS-PAGE and immunobloting. It was a 30000 single chain fragment (scFv) with antigen-binding specificity. The yield of the scFv was 28.9% of the total bacterial proteins.

Animals↗
PubMed1997

Synthesis and antinociceptive activity of [D-Met2, Pro5] enkephalin [N1,5-beta-D-2,3,4,6-O-tetraacetylglycosyl]--amide and [D-Met2, Pro5] enkephalinamide.

Tetra-O-acetylgalactopyranosylamine and tetra-O-acetylglucopyranosylamine of D-Met2, Pro5 enkephalin were designed and synthesized to enhance their membrane penetration, biological activity and resistance to proteolytic hydrolysis. Three approaches to the synthesis were attempted, which lead to a new synthetic scheme with a higher yield and enhanced ease of purification. The improved procedure involved attaching the tetra-O-acetylglycopyranosylamine to a t-Boc-Gly-Phe-Pro-OH peptide, removing the t-Boc, and condensing it with t-Boc-Tyr-D-Met-OH. Biological evaluation in vivo showed that these acetylglycopyranosylamine derivatives bind to mu and delta opioid receptors in homogenate binding assays and possess analgesic activity. The analgesic potency was less than that of the parent compound D-Met2, Pro5 enkephalin. These acetylglycopyranosylamine derivatives showed enhanced lipophilicity compared to their parent compound by a partition coefficient study and they also showed greater membrane permeability, using the rabbit cornea as a model system. These derivatives also are resistant to hydrolytic enzymes as compared to the endogenous met-enkephalin when evaluated in homogenized iris-ciliary body and aqueous humor from rabbit eyes.

Analgesics↗
PubMed1997

Inhibition of bufalin on pituitary and testicular function in rats.

The effects of bufalin on the secretion of testosterone and luteinizing hormone (LH) and the accumulation of testicular adenosine 3':5'-cyclic monophosphate (cAMP) were studied. Male rats were injected with bufalin, human chorionic gonadotropin (hCG), gonadotropin releasing hormone (GnRH), hCG plus bufalin or GnRH plus bufalin via a jugular catheter. Blood samples were collected at several intervals subsequent to the challenge. In the in vitro study, rat testis blocks were incubated with bufalin, hCG or both for 1 h. The anterior pituitary gland was incubated with bufalin, GnRH or both for 30 min. The media were analyzed for testosterone or LH. For studying cAMP accumulation, testicular blocks were incubated for 1 h with the medium containing isobutyl-1-methylxanthine. After incubation, tissues were extracted by ethanol before measuring cAMP concentration. A single intravenous injection of bufalin decreased the basal and hCG-stimulated levels of plasma testosterone. Administration of bufalin in vitro resulted in an inhibition of both basal and hCG-stimulated release of testosterone. Bufalin diminished cAMP accumulation in rat testes. However, the basal levels of plasma and medium LH were not altered by bufalin administration. Likewise, the LH response to GnRH was diminished by bufalin administration, both in vivo and in vitro. These results suggest that the inhibition of testosterone production by bufalin is partly caused by a decrease of testicular cAMP accumulation and LH response to GnRH in rats.

1-Methyl-3-isobutylxanthine↗
PubMed1997

The use of personal protective measures in control of malaria in a defined community.

Malaria is one of the main health problems in the non-immune immigrant workers and army personnel of the malaria endemic areas in Myanmar. Due to changes in the vector bionomics and multiresistant strains of P. falciparum, chemoprophylaxis alone is not an effective means of control of malaria in them. So it is envisaged that the combined used of personal protective measures (deltamethrin impregnated bed-nets, scalves and hand-bands) and the chemoprophylaxis will be an effective means of control of malaria in the define group of people. The study also intended to find out the side effects of the deltamethrin and feasibility and acceptability of methods by the users. The study was conducted in Theini Township, Northern Shan State, from March to November 1993. The study population consisted of all ages of both sexes 554 and 440 persons in the test and control groups respectively. At the initial phase of the study, malaria infected persons from both the groups were treated. The experimental group received personal protective measures with impregnation of bed-nets using 25 mg ai/m2 of deltamethrin at 4 monthly intervals and the scarves and hand-bands at twice the concentration of the insecticides at monthly intervals. Chemoprophylaxis was given to both the groups at weekly intervals using age adjusted dosage of Pyrixine tablet (sulfadoxine-pyrimethamine). The parasitological, entomological, and epidemiological indices were collected at two month intervals in both the groups. The study clearly showed the impact of personal protective measures and chemoprophylaxis on malaria infection in the studied subjects. During the study period, the out patient malaria cases of the test group was 6% to 11.2% and that of the control group was 12% to 21.6% in Theini Hospital. The reinfection rate of the test group (0.9 to 4.7%) was also significantly lower than the control group (6.1 to 14.3%) from July to November. Acceptance of the treated bed-nets, scarves and hand-bands was high and good compliance was found in the follow up. The results of the study clearly showed that malaria can be controlled effectively in the defined group of persons for a malaria transmission season by using chemoprophylaxis and personal protective measures.

Adult↗
PubMed1997

Fusion expression of green fluorescent protein and HCV capsid antigene in Escherichia coli cells.

A chimeric gene of the green fluorescent protein (GFP) and hepatitis C virus (HCV) core antigene were constructed and expressed in E. coli cells. The expressed fusion protein was examined by Dot-ELISA and Western blot and the three antigenic determinants were detected. The GFP-Core fusion protein showed not only the striking green fluorescence under natural light but also the HCV antigenic activity. A new method of immunological diagnosis is greatly anticipated in the light of this fusion protein which can be seen as the HCV antigen tagged with the green fluorescent protein.

Animals↗
PubMed1996

A common mutant epidermal growth factor receptor confers enhanced tumorigenicity on human glioblastoma cells by increasing proliferation and reducing apoptosis.

Alterations of the EGFR gene occur frequently in human gliomas where the most common is an in-frame deletion of exons 2-7 from the extracellular domain, resulting in a truncated mutant receptor (deltaEGFR or de 2-7 EGFR). We previously demonstrated that introduction of deltaEGFR into human U87MG glioblastoma cells (U87MG.deltaEGFR) conferred remarkably enhanced tumorigenicity in vivo. Here, we show by cell-mixing experiments that the enhanced tumorigenicity conferred by deltaEGFR is attributable to a growth advantage intrinsic to cells expressing the mutant receptor. We analyzed the labeling index of the proliferation markers Ki-67 and bromodeoxyuridine and found that tumors derived from U87MG.deltaEGFR cells had significantly higher labeling indexes than those of tumors derived from U87MG cells that were either naive, expressed kinase-deficient mutants of deltaEGFR, or overexpressed exogenous wild-type EGFR. We also utilized terminal deoxynucleotidyl transferase-mediated nick end-labeling assays and showed that the apoptotic index of U87MG.deltaEGFR tumors was more than 4-fold lower than that of parental U87MG tumors. This decrease in cell death was inversely correlated with the expression level of Bcl-X(L), a negative regulator of apoptosis, which was more than 3-fold higher in U87MG.deltaEGFR-derived tumors than in those derived from parental cells. Similar observations were obtained in vitro in serum-free conditions. These results suggest that deltaEGFR exerts its pronounced enhancement of glioblastoma tumorigenicity by stimulating proliferation and inhibiting apoptosis and that the effects are directly attributable to its constitutively active signal.

Animals↗
PubMed1996

Enhanced tumorigenic behavior of glioblastoma cells expressing a truncated epidermal growth factor receptor is mediated through the Ras-Shc-Grb2 pathway.

A mutant epidermal growth factor receptor (DeltaEGFR) containing a deletion of 267 amino acids from the extracellular domain is common in human glioblastomas. We have previously shown that the mutant receptor fails to bind EGF, is constitutively phosphorylated, and confers upon U87MG glioblastoma cells expressing it (U87MG. DeltaEGFR), an increased ability to form tumors in mice. Here we demonstrate that the constitutively phosphorylated DeltaEGFR enhances growth of glioblastoma cells through increased activity of Ras: 1) there was an increase in the proportion of Ras present in the GTP-bound form, and 2) introduction of neutralizing anti-Ras 259 antibodies into U87MG and U87MG.DeltaEGFR cells by microinjection inhibited DNA synthesis to the same low level in both cell populations. We also show that the truncated EGF receptor constitutively associates with the adapter proteins Shc and Grb2 which are involved in the recruitment of Ras to activated receptors. Several derivatives of DeltaEGFR containing single, or multiple mutations at critical autophosphorylation sites were constructed and used to demonstrate that the major Shc binding site is Tyr-1148, and that Grb2 association occurs primarily through Tyr-1068. We conclude that the increased tumorigenic potential of glioblastoma cells expressing the truncated EGF receptor is due at least in part to Ras activation presumably involving the Shc and Grb2 adapter proteins.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Differential expression of lacZ in the liver and kidney of transgenic mice carrying chimeric lacZ-erythropoietin gene constructs with or without its 1.2 kb 3'-flanking sequence.

Erythropoietin (EPO) plays a key role in erythropoiesis and is expressed predominantly in the fetal liver and in the adult kidney. The EPO gene is up-regulated at the transcriptional level under hypoxic/anemic conditions. We studied the role of the 5'- and 3'-flanking sequences of the mouse EPO gene in its tissue-specific and hypoxia-induced expression by developing transgenic mouse lines carrying chimeric EPO-lacZ gene constructs. Transgenic mice carrying a 6.5 kb segment of the 5'-sequence and most of the EPO gene in which lacZ was substituted for exon 1 (5'-lacZ-EPO) demonstrated induction of lacZ expression following hypoxia/ anemia induction in both the liver and kidney of adult mice. However, transgenic mice carrying the above construct along with the 1.2 kb 3'-flanking sequence (5'-lacZ-EPO-3') showed a high level of lacZ expression following hypoxia/anemia induction in adult kidney but not in adult liver. With the aim of further understanding the role of the 3'-flanking sequence in tissue-specific expression of the EPO gene, we studied the interactions of protein factors with this 1.2 kb 3' region and demonstrated that multiple sets of protein factors interact tissue specifically with a 10 bp sequence, TCAAAGATGG, located downstream of the previously characterized 3' hypoxia-responsive enhancer element.

Animals↗
PubMed1996

A combinatorial library strategy for the rapid humanization of anticarcinoma BR96 Fab.

We have used a combinatorial mutagenesis strategy to humanize BR96, a monoclonal antibody that binds to the Lewis Y class of tumor antigens. This approach allows simultaneous assessment of hundreds of humanized variable regions to identify the molecules that best preserve affinity, thus overcoming the major drawback of current humanization procedures, the requirement to construct and analyze each humanized antibody separately. Murine residues of BR96 were mutated to human if they were solvent-exposed residues that did not participate in the formation of the antigen binding site and were not at the interface of the light and heavy chain. At positions that might be involved in binding to antigen, the choice between the murine and human residue was more difficult. Murine and human alternatives were incorporated into a combinatorial library at positions representing buried residues that might affect the structural integrity of the antigen binding site. By encoding this library of humanized BR96 Fabs in an M13 phage vector, we rapidly identified several candidates with nearly identical antigen binding, within 2-fold, of the chimeric Fab. Additional mutagenesis directed at sites suggested in the literature as potentially important for antigen binding in a similar anti-Lewis Y antibody yielded no further improvements.

Amino Acid Sequence↗
PubMed1996

Earth-Based Observations of the Galileo Probe Entry Site

Earth-based observations of Jupiter indicate that the Galileo probe probably entered Jupiter's atmosphere just inside a region that has less cloud cover and drier conditions than more than 99 percent of the rest of the planet. The visual appearance of the clouds at the site was generally dark at longer wavelengths. The tropospheric and stratospheric temperature fields have a strong longitudinal wave structure that is expected to manifest itself in the vertical temperature profile.

Journal Article↗
PubMed1996

Eradication of established intracranial rat gliomas by transforming growth factor beta antisense gene therapy.

Like human gliomas, the rat 9L gliosarcoma secretes the immunosuppressive transforming growth factor beta (TGF-beta). Using the 9L model, we tested our hypothesis that genetic modification of glioma cells to block TGF-beta expression may enhance their immunogenicity and make them more suitable for active tumor immunotherapy. Subcutaneous immunizations of tumor-bearing animals with 9L cells genetically modified to inhibit TGF-beta expression with an antisense plasmid vector resulted in a significantly higher number of animals surviving for 12 weeks (11/11, 100%) compared to immunizations with control vector-modified 9L cells (2/15, 13%) or 9L cells transduced with an interleukin 2 retroviral vector (3/10, 30%) (P < 0.001 for both comparisons). Histologic evaluation of implantation sites 12 weeks after treatment revealed no evidence of residual tumor. In vitro tumor cytotoxicity assays with lymph node effector cells revealed a 3- to 4-fold increase in lytic activity for the animals immunized with TGF-beta antisense-modified tumor cells compared to immunizations with control vector or interleukin 2 gene-modified tumor cells. These results indicate that inhibition of TGF-beta expression significantly enhances tumor-cell immunogenicity and supports future clinical evaluation of TGF-beta antisense gene therapy for TGF-beta-expressing tumors.

Animals↗
PubMed1996

Adenovirus-mediated transfer of CCAAT/enhancer-binding protein-alpha identifies a dominant antiproliferative role for this isoform in hepatocytes.

CCAAT/enhancer-binding protein (C/EBP) isoforms are thought to be important regulators of the hepatocyte phenotype. However, the specific physiological roles of different isoforms are poorly understood because hepatocytes express multiple C/EBPs, and various isoforms have overlapping functions. To identify the functions of C/EBPalpha in mature hepatocytes, replication-defective adenovirus vectors were used to efficiently and homogeneously overexpress the mouse C/EBPalpha gene in a SV40 virus-conditionally transformed rat hepatocyte line that can be induced to express C/EBPbeta and C/EBPdelta but that has little endogenous C/EBPalpha expression. Hepatocytes were infected with a recombinant adenovirus vector carrying the cDNA for C/EBPalpha driven by Rous sarcoma virus promoter elements (AdCEBPalpha) or a similar vector carrying the Escherichia coli lacZ gene (Adbetagal). Staining for beta-galactosidase demonstrated an infection efficiency of 100% at a multiplicity of infection of 25 plaque-forming units/cell and persistence of foreign gene expression for at least 9 days. Cultures infected with AdCEBPalpha had 50-fold higher levels of C/EBPalpha mRNA and protein than those infected with Ad-beta-gal, but similar expression of C/EBP-beta. Infection with AdCEBPalpha inhibited proliferation in cells expressing little C/EBPbeta, even when proliferation was driven by the SV40 transforming antigen, and also blunted mitogenic induction of the c-myc proto-oncogene in nontransformed cells with high levels of C/EBPbeta. Although overexpression of C/EBPalpha consistently increased C/EBPalpha DNA binding activity, it was not sufficient for albumin expression. Infection with AdCEBPalpha only increased albumin mRNA levels in nontransformed cells that also expressed relatively high levels of C/EBPbeta. Thus, in hepatocytes, C/EBPalpha has a dominant antiproliferative function, but must interact with other factors to regulate hepatocyte-specific gene expression.

Adenoviridae↗
PubMed1996

The time course of CTLAIg effect on cardiac allograft rejection.

T cell response to alloantigen is dependent not only on T cell receptor activation, but also upon co-stimulation through the CD28 receptor, as T cell receptor activation alone is insufficient for an optimal immune response. The CD28 receptor on helper T cells interacts with its ligand B7 on activated B cells/macrophages as the co-stimulus to support T cell activity. The natural ligand for CD28, B7 is expressed in both constituitive and inducible forms. Expression of the inducible form, B7.1 is the most co-stimulatory and may peak at 48 hr following antigen presentation. CTLA4Ig (a soluble CD28 receptor analog) binds both B7's and inhibits CD28 activation. This experiment was undertaken to examine the optimal time course of CTLA4Ig effect at or following antigen presentation. In vivo studies used a rat MHC mismatch heterotopic cardiac allograft transplant model (Brown-Norway to Lewis). Controls received no immunotherapy. Experimental recipients received CTLA4Ig (0.05 mg i.p.) the day of transplant only or had CTLA4Ig x 7-21 days begun at Day 0, 3, or 5 following transplant. Rejection was defined as a lack of allograft contraction. Allograft survival was best when CTLA4Ig was present on Day 2. These findings demonstrate that CTLA4Ig was most effective 48 hr following antigen presentation, perhaps reflecting induction of tolerance at a time of maximal CTLA4Ig/B7.1 blockade. CTLA4Ig given later, at a time when the recognition/rejection process has already begun was not as effective indicating the lack of immunosuppressive function of CTLA4Ig itself and confirms CTLA4Ig's mechanism of co-stimulation blockade.

Abatacept↗
PubMed1996

Study on the relationship between cytogenetics and phenotypic effect in Turner's syndrome.

The cytogenetics and clinical stigmata in 5 cases of Turner's syndrome were studied. Three of them were non-mosaic 1(Xq) and two with partial monosome of a X chromosome short are (Xp21), whose DNA replication patterns of inactive X chromosome were analyzed by RBG technique. Results showed that differences between the replication patterns in cases of X chromosome deletion (Xp21) and normal females existed; that the behavior of abnormal X expressed nonrandom inactivation. It was suggested that the phenotype may be closely related with both X chromosome replication pattern and its inactivation behavior, which might be useful in genetic counselling.

Adolescent↗