Search PubMed⌕ Search

Biomedical subjects

H C Chang

Publications and source records attributed to H C Chang.

At least 199 records · Page 11Linked to original sources
PubMed1989

CD2-mediated adhesion facilitates T lymphocyte antigen recognition function.

The CD2 T lymphocyte-surface glycoprotein serves to mediate adhesion between T lymphocytes and their cognate cellular partners which express the specific ligand LFA-3. In addition, CD2 by itself or in conjunction with T-cell receptor stimulation, transduces signals resulting in T-lymphocyte activation. One or both of these functions seems to be physiologically important, given that certain anti-CD2 monoclonal antibodies block T-cell activation and that antigen-responsive memory T cells express a high level of CD2 relative to virgin T cells, which are largely antigen-unresponsive. Nevertheless, the contribution of the individual CD2 functions in T-cell responses has not been independently examined. To this end, human CD2 complementary DNAs encoding an intact LFA-3-binding adhesion domain, but lacking a functional cytoplasmic signal transduction element (CD2trans-), were introduced into an ovalbumin-specific, I-Ad restricted murine T-cell hybridoma. The antigen-specific response of T hybridoma cells expressing human CD2trans- protein was enhanced up to 400% when the human LFA-3 ligand was introduced into the I-Ad expressing murine antigen-presenting cells. In contrast, no augmentation was observed if human LFA-3 was absent or expressed on a third-party cell lacking the I-Ad restriction element. These results directly demonstrate the functional significance of adhesion events mediated between CD2 on the antigen-responsive T lymphocyte and LFA-3 on the presenting cell in optimizing antigen-specific T-cell activation.

Animals↗
PubMed1989

Perturbation of the idiotypic network. I. Induction with multiple alloantigen stimulation.

The effect of hyperimmunization on the immune network with allostimulated syngeneic lymphocytes responding to different haplotypes was analyzed. Ten different haplotypes were used to stimulate syngeneic donor mice. Control mice were multiply immunized with incomplete Freund's adjuvant alone or with syngeneic mixed lymphocyte culture-generated lymphocytes. BALB/c mice were immunized consecutively with alloreactive blasts or allogeneically stimulated spleen cells at 10-day intervals. After a rest period of 2 months, the ratio of T helpers to T suppressors was determined by immunofluorescent staining. The functional network was probed by immunizing the mice with phosphorylcholine (PC) coupled to hemocyanin. The sera were analyzed for anti-PC antibodies and TEPC15 (T15) idiotypic expression. The results demonstrated (i) a decrease in the level of anti-PC antibody titer and T15 idiotypic expression; (ii) a decrease in the number of T helper cells and an increase in the number of T suppressor cells; (iii) a loss of PC epitope specificity; (iv) an increase of IgM antibodies expressing T15 without anti-PC specificity; and (v) an elevated level of preimmune lymphocyte proliferation and Ig secretion. These results reveal a functional network linkage in the regulation of alloreactivity and antigen response and show how repeated exposure to alloantigens can induce a perturbation of the idiotypic network controlling the response of a non-alloantigen-related BALB/c strain dominant idiotype (T15).

Animals↗
PubMed1989

Perturbation of the idiotypic network. II. Transfer of suppression to neonatal mice.

The cellular mechanisms which are responsible for idiotype perturbation induced by repeated stimulation with either allogeneic mixed lymphocyte culture-generated syngeneic blasts or allogeneically stimulated syngeneic spleen cells were investigated as described in the preceding article. Using the splenic fragment culture system, the precursor frequencies of T and B cells for anti-phosphorylcholine (PC) antibodies and T15 idiotypic antibodies were determined in allogeneically challenged mice. Adoptive transfers of T cells to neonatal BALB/c mice induced suppression of the T15+ anti-PC responses. In addition, the effect of immunization with internal image-bearing anti-idiotopes on the level of anti-PC antibodies and T15 idiotype was determined. Results from this study demonstrated (i) a decrease in the precursor frequency in the PC-specific and idiotype-specific B cell repertoire; (ii) a decrease in the precursor frequency of T helper cells, which recognize idiotypes and anti-idiotypes; (iii) the possibility to transfer T15 suppression to neonatal mice; and (iv) the possibility to restore T15 dominance by anti-idiotypic antibody immunization. These data indicate that both the T and B compartments are involved in the maintenance of suppression induced by repeated exposure to alloantigen-sensitized syngeneic cells. Collectively these findings show that a nonspecific general suppression induced by allohyperimmunization can perturb the T15+ anti-PC response.

Animals↗
PubMed1989

Structural and binding analysis of a two domain extracellular CD2 molecule.

The 50-kD CD2 (T11) surface glycoprotein on human T lymphocytes and thymocytes plays a critical role in T lineage cell activation and adhesion via its ligand LFA-3. To begin to define structure-function relationships in the extracellular segment of the transmembrane CD2 molecule, we have used a eukaryotic expression system and a CD2 cDNA to produce milligram amounts of recombinant soluble CD2 molecule that corresponds to the two extracellular segment exons. We show that this protein, termed T11ex2, behaves as a monomer in aqueous solution and includes a proteolytically resistant NH2-terminal fragment (domain I) encoded by the first extracellular segment exon. Circular dichroism analysis of T11ex2 demonstrates that its stabilized secondary structure is dependent on the intrachain disulfide bonds present in domain II. The T11ex2 monomer binds directly to the CD2 ligand LFA-3 with a dissociation constant of 0.4 microM. This relatively low affinity implies that cooperative binding resulting from an array of transmembrane CD2 molecules is important to facilitate physiologic T cell adhesion.

Antibodies, Monoclonal↗
PubMed1989

Deoxyribonuclease II purified from the isolated lysosomes of porcine spleen and from porcine liver homogenates. Comparison with deoxyribonuclease II purified from porcine spleen homogenates.

Porcine spleen DNase II (EC 3.1.22.1), one of the best-characterized DNases II, is subcellularly located in lysosomes because the enzyme is co-sedimented with two of the lysosomal marker enzymes, cathepsin D and acid phosphatase. The physicochemical properties, including the subunit structure, sensitivity to iodoacetate inactivation, native molecular weight and chromatographic behavior, of the DNase II purified from the isolated lysosomes of porcine spleen are indistinguishable from those of the same enzyme purified from the whole porcine spleen homogenate. DNase II can also be extracted from porcine liver with 0.05 M H2SO4 or 0.1 M NaCl and purified from either extract by a series of column chromatographies. The purified liver DNase II from either extract has the same subunit structure (alpha-chain, Mr 35,000 and beta-chain, Mr 10,000) as the purified DNase II of porcine spleen. The two liver extracts as well as the extracts of spleen and gastric mucosa contain DNase II with very similar properties on Sephadex G-100 gel filtration, on acid polyacrylamide gel electrophoresis under non-denaturing conditions, and on isoelectric focusing. The data strongly suggest that, for the same species of animal, the DNase II activities in various tissues are associated with protein molecules of identical structure.

Animals↗
PubMed1989

Isolation and characterization of a cDNA encoding a putative cytokine which is induced by stimulation via the CD2 structure on human T lymphocytes.

To define activation-specific sequences in human T lymphocytes, a cDNA library was constructed by subtractive hybridization using resting and stimulated peripheral blood lymphocyte pairs. Stimulation of peripheral blood lymphocytes was achieved by triggering with mitogenic anti-CD2 (T11) monoclonal antibodies. Differential library screening with cDNA probes derived from stimulated vs. resting peripheral blood lymphocytes led to identification of a novel sequence, termed HC21. The predicted primary and secondary structure of HC21 deduced from the translated nucleotide sequence suggest that the gene encodes a secreted protein of 92 amino acids including a 23-residue leader sequence. Northern blot analysis demonstrates that HC21 mRNA is induced in peripheral blood T lymphocytes and interleukin 2-dependent T cell clones within 10 min following stimulation via CD2, while steady-state RNA expression is maximal by 2 h. Although the kinetics of HC21 expression are similar after stimulation via the antigen/major histocompatibility complex receptor (CD3-Ti), CD3-Ti triggering leads to less accumulation of HC21 RNA than CD2 triggering. In contrast, recombinant interleukin 2 does not induce detectable HC21 RNA expression in T cell clones. Transient expression of the HC21 cDNA in COS cells results in a readily detectable protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of radiolabeled cell supernatants consistent with HC21 encoding a secreted product. Protein sequence analysis reveals a striking homology (up to 76% identity at amino acid level) with other members of a recently described lymphokine family whose functions are yet to be defined.

Amino Acid Sequence↗
PubMed1989

Cloning and expression of a gamma-interferon-inducible gene in monocytes: a new member of a cytokine gene family.

To define activation-specific sequences in human peripheral blood lymphocytes (PBL), a cDNA library was constructed by subtractive hybridization using resting and stimulated PBL pairs. Stimulation of PBL was achieved by triggering with mitogenic anti-CD2 (T11) monoclonal antibodies. Differential library screening with cDNA probes derived from stimulated versus resting PBL led to identification of two novel sequences, termed HC11 and HC14. The predicted primary and secondary structure of HC11 deduced from the translated nucleotide sequence suggests that the gene encodes a secreted protein of 99 amino acids (aa), including a 23 aa residue leader sequence. Surprisingly, Northern blot analysis demonstrated that HC11 mRNA is induced predominantly in peripheral blood non-T cells. Subsequently, we observed that the HC11 mRNA is induced in macrophages and the monocytic line U-937 by gamma-IFN, raising the possibility that T cell-derived gamma-IFN induced upon anti-CD2 stimulation activated monocytes to express HC11 RNA. In support of this notion, neutralizing anti-gamma-IFN monoclonal antibody inhibits the induction of HC11 mRNA in PBL activated through anti-CD2 antibodies. These findings suggest that there is a molecular cascade involving T cell-produced lymphokines and monokines which serve as a means for intercellular communication. Transient expression of HC11 cDNA results in a readily detectable specific set of protein bands in SDS-PAGE analysis of supernatants from radio-labeled COS cells, consistent with HC11 encoding a secreted product(s). Protein sequence comparison reveals homology with other members of a recently described inducible cytokine family whose functions are yet to be defined.

Amino Acid Sequence↗
PubMed1989

The structural biology of CD2.

The CD2 molecule is a 50-55KD transmembrane glycoprotein expressed on the vast majority of thymocytes and virtually all peripheral T lymphocytes. Its functions are two-fold: adhesion and activation. CD2 serves to facilitate conjugate formation between the T-lineage cell and its cognate partner via intermolecular interaction of CD2 and LFA-3 on the former and latter cells, respectively. Perturbation of the CD2 extracellular segment by certain combinations of anti-CD2 MAbs or LFA-3 and a single anti-CD2 MAb activate T-lineage function. These CD2-mediated activation events also synergize with signals mediated through the TCR to augment T-cell response. Based on microchemical analysis of immunoaffinity-purified human CD2 and cDNA and genomic cloning of mouse and human molecules, considerable structural information is now available. The mature surface human CD2 molecule consists of 327 amino acids: a 185 aa extracellular segment; a 25 aa hydrophobic transmembrane segment; and a 117 aa cytoplasmic domain rich in prolines and basic residues. The CD2 gene is comprised of five exons which span approximately 12 Kb on chromosome 1. A similar protein structure and gene exon organization is found for the mouse CD2 homologue. The CD2 adhesion domain is approximately 103 aa in length and is encoded by a single exon (exon 2). This domain is resistant to proteolysis, even though it lacks any intrachain disulfides and, like the entire extracellular segment protein expressed in a baculovirus system, binds to its cellular ligand, LFA-3. The latter occurs with a micromolar Kd. This relatively low affinity suggests that multivalent interactions among CD2 monomers on the T cells and individual LFA-3 structures on the cognate partner are important in enhancing the avidity of the T-cell interaction with its target or stimulator cell. The affinity of the CD2 extracellular segment for LFA-3 is not affected by truncations in the CD2 cytoplasmic domain, implying that ligand binding is not regulated by intracellular mechanisms. Given that CD2 mRNA expression and surface CD2 copy number are increased by more than one order of magnitude post-TCR stimulation, it is more likely that adhesion via CD2 is modulated by alteration in surface copy number. Analysis of early transduction events occurring via CD3-Ti (TCR) and CD2 including single channel Ca2+ patch-clamp recordings on living human T lymphocytes indicate a virtual identity of signals.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1989

Effects of exercise and ticlopidine on platelet function and prostanoids in patients with old myocardial infarction.

Twenty-eight subjects, 16 with old myocardial infarction (OMI) and a control group of 12, received ticlopidine 250 mg/day for 4 weeks. All subjects underwent the standard Bruce protocol exercise test before and after therapy. Blood was collected from a peripheral vein to study platelet function at pre-exercise (rest), peak exercise (end point), and 6 min post-exercise (recovery). Platelet count, platelet aggregatory response, plasma thromboxane B2 (TXB2), and 6-keto-PGF 1 alpha were measured by Coulter counter, aggregometry, and radioimmunoassay, respectively. The platelet count, aggregatory response (induced by ADP, adrenalin, and collagen) and the plasma concentration of TXB2 and 6-keto-PGF 1 alpha were not significantly affected by exercise in the OMI and control groups. Overall, there were no significant changes in exercise duration, heart rate, and blood pressure in both groups before or after ticlopidine treatment, with some minor exceptions. There were no significant changes in the peripheral blood profiles, the bleeding and coagulation times, the blood chemistry data, and the platelet counts between the groups before or after treatment. There was a significant decrease in collagen- and ADP-induced platelet aggregation in both groups after ticlopidine therapy. Prior to treatment, a higher plasma TXB2 concentration was noted in the OMI group during and after exercise; the difference was no longer statistically significant after ticlopidine therapy when both groups were compared. There was a significant reduction in the TXB2 levels in the OMI group after ticlopidine treatment despite exercise. Before therapy, plasma 6-keto-PGF 1 alpha was significantly elevated in the OMI group before or after exercise but was not statistically significant after ticlopidine treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

6-Ketoprostaglandin F1 alpha↗
PubMed1988

Expression of a functional CD3-Ti antigen/MHC receptor in the absence of surface CD2. Analysis with clonal Jurkat cell mutants.

To investigate the requirement for CD2 expression in activation of T lymphocytes via the CD3-Ti antigen/MHC receptor complex, we produced and characterized a series of CD2- Jurkat variants. These mutants lack detectable surface CD2 as determined by indirect immunofluorescence, immunoprecipitation analysis, and specific radiolabeled antibody binding assay, but nevertheless, expressed normal numbers of CD3-Ti receptors. As expected, the combination of anti-CD2 antibodies, termed anti-T112 and anti-T113, which are mitogenic for resting T lymphocytes, failed to stimulate activation of these variants. In contrast, triggering of their CD3-Ti components resulted in the normal set of T lymphocyte-associated activation events, including phosphoinositide turnover, elevation in intracellular free calcium, early gene-induction events, and IL-2 production. Assuming that the Jurkat cell line is representative of normal cycling human T lymphocytes, we conclude that the presence of the CD2 molecule on the plasma membrane is not in itself a requirement for an operational CD3-Ti-alpha/beta receptor.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1988

A soluble CD4 protein selectively inhibits HIV replication and syncytium formation.

The CD4 (T4) molecule is expressed on a subset of T lymphocytes involved in class II MHC recognition, and is probably the physiological receptor for one or more monomorphic regions of class II MHC (refs 1-3). CD4 also functions as a receptor for the human immunodeficiency virus (HIV) exterior envelope glycoprotein (gp120) (refs 4-9), being essential for virus entry into the host cell and for membrane fusion, which contributes to cell-to-cell transmission of the virus and to its cytopathic effects. We have used a baculovirus expression system to generate mg quantities of a hydrophilic extracellular segment of CD4. Concentrations of soluble CD4 in the nanomolar range, like certain anti-CD4 monoclonal antibodies, inhibit syncytium formation and HIV infection by binding gp120-expressing cells. Perhaps more importantly, class II specific T-cell interactions are uninhibited by soluble CD4 protein, whereas they are virtually abrogated by equivalent amounts of anti-T4 antibody. This may reflect substantial differences in CD4 affinity for gp120 and class II MHC.

Animals↗
PubMed1988

The T 11 (CD 2) cDNA encodes a transmembrane protein which expresses T 11(1), T 11(2) and T 11(3) epitopes but which does not independently mediate calcium influx: analysis by gene transfer in a baculovirus system.

To determine whether a full-length human T 11 (CD 2) cDNA encodes a protein which expresses all three T 11 epitopes (T 11(1), T 11(2), T 11(3] and independently triggers activation in a cell other than a T lymphocyte, a baculovirus expression system was employed. Here we show that a recombinant T 11 cDNA-containing baculovirus can induce high-level expression of T 11(1), T 11(2) and T 11(3) epitopes on the surface of gut epithelial SF9 cells. However, in this environment the T 11 protein cannot be triggered to transduce a signal resulting in an elevation of the cytosolic free Ca2+ as is known to occur in T lymphocytes. These results support the notion that T 11 functions in a coordinate fashion with other intracellular lymphoid components.

Animals↗
PubMed1988

In vitro characteristics of human fetal cells obtained from chorionic villus sampling and amniocentesis.

In vitro characteristics of human fetal cells have been investigated after chorionic villus sampling at the first trimester and amniocentesis at the second trimester of pregnancy. Light microscopy revealed heterogeneous morphology of cell types in both the chorionic villus culture and the amniotic fluid cultures. Based on the experiments performed, chorionic villus cells are more sensitive to pronase, trypsin, and versene during subculture and have a higher DNA content per single cell and release more [125I]-Beta-human chorionic gonadotropin into culture medium than those found in amniotic fluid cells. The practical applications of this study are discussed.

Amniocentesis↗
PubMed1988

Adhesion domain of human T11 (CD2) is encoded by a single exon.

The 50-kDa T11 (CD2) T-lymphocyte surface glycoprotein facilitates physical adhesion between T-lineage cells and their cognate cellular counterparts (cytotoxic T-lymphocytes-target cells, helper T lymphocytes-antigen-presenting cells, or thymocytes-thymic epithelium) as well as signaling through the antigen-specific T3-Ti receptor complex. To examine the relationship between the structure and function of the T11 molecule, we have utilized a baculoviral expression system to produce milligram quantities of the hydrophilic extracellular T11 segment. Enzyme cleavage, microsequencing, and HPLC analyses of the expressed protein in conjunction with genomic cloning information show that the domain involved in cellular adhesion is encoded by a single 321-base-pair exon.

Amino Acid Sequence↗
PubMed1987

Human T lymphocyte activation.

The T lymphocyte receptor for antigen and histocompatibility molecules is a molecular complex comprised of five polypeptide chains. Both the 49KD alpha and 43KD beta chains are immunoglobulin-like and thus contain variable domains responsible for ligand binding. In contrast, the 20-25KD T3 gamma, delta and epsilon chains are monomorphic structures presumably involved in transmembrane signalling. The alpha and beta subunits are disulfide bonded to each other and held in noncovalent association with the T3 chains. T3-Ti receptor crosslinking leads to conformational modification of a second T lineage specific molecule, termed the 50KD T11 structure and in turn, protein kinase C activation, elevation in intracytoplasmic free calcium and Na+/H+ antiport stimulation.

Cell Differentiation↗
PubMed1987

Molecular cloning and expression of T11 cDNAs reveal a receptor-like structure on human T lymphocytes.

The T11 (CD2) sheep-erythrocyte-binding protein is a T-cell surface molecule involved in activation of T lymphocytes and thymocytes, including those lacking the T3-Ti antigen-receptor complex. The primary structure of T11 was deduced from protein microsequencing and cDNA cloning. The mature human protein appears to be divided into three domains: a hydrophilic 185 amino acid external domain bearing only limited homology to the T-cell surface protein T4 and the immunoglobulin kappa light chain variable region, a 25 amino acid hydrophobic transmembrane segment, and a 126 amino acid cytoplasmic domain rich in prolines and basic residues. Transfection of cDNAs encoding either the 1.7- or the 1.3-kilobase T11 mRNA into COS-1 cells resulted in expression of surface T11 epitopes as well as sheep-erythrocyte-binding capacity. The predicted structure is consistent with the possibility that T11 functions in signal transduction.

Amino Acid Sequence↗
PubMed1987

Quantitation of urinary methylmalonic acid by gas chromatography mass spectrometry and its clinical applications.

Urine methylmalonic acid (MMA) concentrations were detected in 79 Chinese patients by gas chromatography mass spectrometry (GC/MS), using a selected ion monitoring program. 10 of the 79 patients were found to have cobalamin deficiency. Their urine MMA amounts were all elevated with a mean value 1376 ng/microliter (11.66 mmol/l), ranging from 40.46 to 3900 ng/microliter (0.34-33.05 mmol/l). The remaining 69 cases were found to be unrelated to cobalamin deficiency. Their mean urine MMA was 3.62 ng/microliter (30.0 mumol/l), ranging from 0-17.47 ng/microliter (0-148.0 mumol/l). In this study, we found that urine MMA detected by GC/MS was a simple, rapid, convenient, specific and sensitive method for the diagnosis of cobalamin deficiency. The urine MMA concentrations in cases not due to cobalamin deficiency would not exceed 20 ng/microliter (169.5 mumol/l), whereas in cobalamin deficiency the urine MMA levels always exceeded 20 ng/microliter, or were even much higher. No overlapping of the results of urine MMA between these 2 groups of patients could be seen in our study. Detection of urine MMA is useful in the demonstration or exclusion of cobalamin deficiency in any suspect patients.

Aged↗