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Biomedical subjects

A Chen

Publications and source records attributed to A Chen.

At least 307 records · Page 17Linked to original sources
PubMed1994

Retinoic acid is required for and potentiates differentiation of acute promyelocytic leukemia cells by nonretinoid agents.

Patients with acute promyelocytic leukemia (APL) associated with the t(15;17) translocation and fusion of the promyelocytic leukemia (PML) and retinoic acid receptor-alpha (RAR-alpha) genes achieve complete remission but not cure with all-trans retinoic acid (RA), NB4, a cell line derived from a patient with t(15;17) APL that undergoes granulocytic differentiation when treated with pharmacologic doses of RA, was used as a model for differentiation therapy of APL. We found that NB4 cells are resistant to differentiation by nonretinoid inducers such as hexamethylene bisacetamide (HMBA), butyrates, vitamin D3, or hypoxanthine, all of which can induce differentiation in the commonly used HL60 leukemia cell line. Preexposure of NB4 cells to low concentrations of RA for a period as short as 30 minutes abolished resistance to nonretinoids and potentiated differentiation. Sequential RA and HMBA treatment yielded maximal differentiation by 3 days of drug exposure, whereas the effect of RA alone peaked after 6 days and yielded a smaller percentage of differentiated cells. RA also reversed NB4 cell resistance to butyrates and allowed for synergistic differentiation by these agents. Pretreatment with HMBA before exposure to RA failed to stimulate differentiation. Sequential RA/HMBA treatment also markedly increased the extent of differentiation of primary cultures of bone marrow and peripheral blood mononuclear cells from three APL patients. In one case RA/HMBA treatment overcame resistance to RA in vitro. Together, these results suggest that intermittent low doses of RA followed by either HMBA or butyrates may be a useful combination in the treatment of APL. This clinical strategy may help prevent or overcome RA resistance in APL.

Acetamides↗
PubMed1994

Human T cells express specific binding sites for C1q. Role in T cell activation and proliferation.

Although the receptor that binds to the collagen-like domain of human C1q (C1qR) is expressed on a wide variety of cell types, the presence or absence of this receptor on human T lymphocytes has been debatable. The current studies were undertaken to re-examine whether human T cells possess specific binding sites for C1q by using a combination of techniques, including radioligand binding studies, flow cytometric analysis, and epifluorescence imaging techniques. Radioligand binding studies indicate that both peripheral T cells and the cultured T cell line, MOLT4, bind 125I-labeled C1q in a specific and apparently saturable manner, reaching equilibrium within 30 min at 37 degrees C under conditions of subphysiologic (90 mM NaCl) ionic strength. Western blot analysis with anti-C1qR of membrane proteins derived from Raji and MOLT4 cells showed an apparent single band of approximately 60 kDa under nonreducing conditions. Furthermore, when peripheral blood T cells were stimulated with 12,-o-tetradecanoyl phorbol-13-ester acetate for 5 days at 37 degrees C and assessed by FACS for their ability to bind anti-C1qR, the mitogen-induced cells were found to bind 40 to 50% more than their unstimulated counterparts. In addition, both CD4+ and CD8+ T cells were found to bind anti-C1qR. When the cells were mitogen induced with either 12,-o-tetradecanoyl phorbol-13-ester acetate, Con A, or PWM for 48 h in the presence or absence of 50 micrograms/ml C1q then pulsed with 1 microCi [3H]thymidine for 16 h at 37 degrees C, proliferation was significantly inhibited (40 to 80%, n = 7) as assessed by reduced [3H]thymidine incorporation. Taken together, the data suggest that: 1) Human T cells express C1qR in which immunoblots reveal a 60-kDa single chain protein. 2) C1qR expression is up-regulated by mitogens that induce T cell proliferation. 3) The primary ligand, C1q, induces an antiproliferative signal, which suggests that the C1qR plays a role in T cell activation and proliferation. In addition, the data contribute to the characterization of C1qRs on cells in peripheral blood and indicate that all cells, with the exception of erythrocytes, bear functional C1q receptors.

Carrier Proteins↗
PubMed1994

VASE exon expression alters NCAM-mediated cell-cell interactions.

The neural cell adhesion molecule (NCAM) is found on cells as several related polypeptides formed by alternative splicing of the single NCAM gene. The alternatively spliced 30-bp VASE exon in the fourth immunoglobulin-like domain is the structural variation nearest those portions of the polypeptide proposed to mediate cell-cell adhesion. To test the ability of distinct forms of the NCAM molecules to mediate cell adhesion, L cells were transfected with expression vectors encoding rat 140 kD NCAM +/- the VASE exon. L cell lines which expressed these polypeptides were isolated and tested for self-aggregation in a low shear, rapid aggregation assay. Increased cellular aggregation of the transfectants was observed to be a function of the NCAM molecule expressed. These transfected cells showed segregation in a long term co-aggregation assay: cells expressing NCAM--VASE formed aggregates which tended to exclude cells expressing NCAM+VASE and vice versa. These results provide direct evidence that this small difference in NCAM structure is sufficient to allow segregation of cells.

Animals↗
PubMed1994

Use of model cell lines to study the biosynthesis and biological role of cancer-associated sialosyl-Tn antigen.

Sialosyl-Tn (STn) is a mucin-associated carbohydrate antigen that is not expressed by most normal epithelial cells but becomes expressed in several types of adenocarcinomas, where it is often associated with a poor prognosis. Little is known about the regulation of the STn phenotype in tumor cells and the immune response to STn antigen. In the present study, we established clonal cell lines in which virtually all of the cells were STn positive (designated LS-C) or STn negative (designated LS-B). These two cell lines, derived from a single parental cell line, LS174T, have the same total protein electrophoretic profiles but carry markedly different oligosaccharide structures on their mucin; the mucin from LS-C cells has only the Tn and STn structures, whereas LS-B cell mucin lacks these simple structures but carries more complex oligosaccharides. These results indicate that lack of STn expression by cells can be due to the lack of STn synthesis rather than inaccessibility of antibodies to bind to STn by steric hindrance. Both clones were similar in their growth rates, response to gamma-interferon, and sensitivity to lysis by lymphokine-activated killer cells. These cells may be important models for understanding the regulation of glycosylation at the cellular level and for further studies of tumor biology and immune response to STn antigen.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1994

Positive control mutations in the MyoD basic region fail to show cooperative DNA binding and transcriptional activation in vitro.

An in vitro transcription system from HeLa cells has been established in which MyoD and E47 proteins activate transcription both as homodimers and heterodimers. However, heterodimers activate transcription more efficiently than homodimers, and function synergistically from multiple binding sites. Positive control mutants in the basic region of MyoD that have previously been shown to be defective in initiating the myogenic program, can bind DNA but have lost their ability to function as transcriptional activators in vitro. Additionally, positive control mutants, unlike wild-type MyoD, fail to bind cooperatively to DNA. We propose that binding of MyoD complexes to high affinity MyoD binding sites induces conformational changes that facilitate cooperative binding to multiple sites and promote transcriptional activation.

Amino Acid Sequence↗
PubMed1994

Morphogenesis of catecholaminergic interneurons in the frog spinal cord.

During embryonic and larval development of the frog Xenopus laevis, a bilateral population of cerebrospinal fluid-contacting neurons matures in the ventral spinal cord. These cells are catecholaminergic and form a dispersed or nonrandom pattern of spacing within each of their spinal cord columns. In order to test the mechanisms underlying pattern formation of these neurons, it is first necessary to understand their normal morphogenesis. Morphogenetic changes were examined by using immunocytochemistry for tyrosine hydroxylase as a cell marker. Immunoreactivity in spinal cord cells was detected as early as 1.4 days (stage 28) of embryonic development. Subsequently, these cells underwent changes in shape, and rapid, regressive changes in cell size. The population emerges gradually during development, but the major characteristic of nonrandom spacing, their dispersion from other catecholaminergic cells, is apparent at early stages of differentiation. Increases in cell density occur over an extended period of time and can be divided into an initial phase of large, rapid changes and a subsequent plateau phase of gradual changes. The two longitudinal columns of catecholaminergic cells that are characteristic of older animals become apparent just before hatching, when density increases until cells on both sides of the midline are present in the same region. Although the dispersed pattern exists within each column, cross-correlation analysis shows that there is a random relationship between cells in opposite columns. During larval development, the catecholaminergic cell domain expands in both a rostral and caudal direction. The morphogenetic changes of the catecholaminergic cell population begin to show how the cells become partitioned within the floor plate region of the spinal cord.

Animals↗
PubMed1994

Role of the integrin alpha v beta 6 in cell attachment to fibronectin. Heterologous expression of intact and secreted forms of the receptor.

The integrin alpha v beta 6 has been shown to be a fibronectin-binding protein. To determine whether the cytoplasmic and transmembrane domains of alpha v beta 6 are necessary for binding to fibronectin, a truncated, secreted form of the integrin lacking these domains was engineered and expressed in Chinese hamster ovary cells. Fibronectin affinity chromatography demonstrated that the secreted integrin, like its full-length counterpart, was capable of binding fibronectin. Monoclonal antibodies were made to secreted alpha v beta 6 and to beta 6-transfected NIH 3T3 cells. In experiments designed to determine whether alpha v beta 6 can mediate cell attachment to fibronectin, full-length human beta 6 was expressed in Chinese hamster ovary cells and in the human colon carcinoma cell line SW480. beta 6-expressing cells were identified by alpha v beta 6-specific antibodies, and the beta 6-transfectants were used in cell-adhesion assays. In Chinese hamster ovary cells, human beta 6 associated with hamster alpha v but was incapable of mediating cell attachment to fibronectin. However, expression of beta 6 in these cells had the dominant negative effect of decreasing alpha v beta 5-dependent adhesion to vitronectin. In SW480 cells, beta 6 expression conferred the ability to bind to fibronectin even in the presence of inhibitory antibodies against beta 1 integrins. In such cells, fibronectin binding ability could be blocked by an antibody to alpha v integrins. These results constitute the first direct evidence that alpha v beta 6 mediates cell attachment to fibronectin.

3T3 Cells↗
PubMed1994

Myelodysplastic syndrome transforming to acute promyelocytic-like leukemia with trisomy and rearrangement of chromosome 11.

Variants of the t(15;17)(q22;q12-q21) chromosomal rearrangement associated with acute promyelocytic leukemia (APL) have been previously described and they frequently involve either chromosome 15 and/or 17. Previously we reported a rare variant t(11;17). We now describe two patients with myelodysplastic syndrome (MDS) that transformed to APL-like leukemia. Both had trisomy 11 at the diagnosis of APL-like leukemia. Following treatment for APL, patient 1 reverted to MDS and showed a normal karyotype. When leukemia recurred, his bone marrow karyotype was 47,XY,t(4;11), +11,der(22)t(1;22). Both patients were treated with all-trans retinoic acid (ATRA) for APL for 5 weeks, but failed to respond. The karyotype of patient 1 after ATRA treatment was 46,XY,t(4;11); the trisomy 11 had been lost and the bone marrow was replaced with immature myeloblasts without promyelocytes. In patient 2, the karyotype remained the same as at diagnosis, i.e., 47,X,-Y,dir ins(4;7),del(5), +6,del(7), +8, + 11,-18. Molecular analysis by reverse transcriptase PCR analysis showed the presence of wild type retinoic acid receptor alpha (RARA) and the absence of the PML-RARA chimeric gene associated with t(15;17). Additional analysis of PLZF, a new zinc finger gene associated with t(11;17), also showed the absence of this hybrid gene. These data support the concept that APL is a heterogeneous disorder and that variants with chromosome 11 rearrangement exist that do not respond to ATRA.

Adult↗
PubMed1994

Pathogenesis of IgA nephropathy: in vitro activation of human mesangial cells by IgA immune complex leads to cytokine secretion.

IgA immune complex (IC) plays a crucial role in the pathogenesis of IgA nephropathy (IgAN). As IgA-IC is not itself cytotoxic, other mediators may be involved in the pathogenesis. In order to elucidate the mechanisms by which IgA-IC mediates renal injury in IgAN, the ability of IgA-IC to 'activate' cultured human mesangial cells (HMC) was studied. HMC were incubated with nephritogenic IgA-IC, containing a MOPC-315 plasmacytoma-derived IgA anti-dinitrophenyl (DNP) and DNP-conjugated bovine serum albumin. The cells showed morphological changes, an accelerated rate of proliferation, and increased production of interleukin-1 (IL-1), interleukin-6 (IL-6), platelet activating factor (PAF) and generation of superoxide anion. The enhancement of IL-1 and IL-6 mRNA expression in HMC incubated with IgA-IC was identified by dot blot analysis. Northern blot hybridization also demonstrated an augmented IL-6 mRNA expression in HMC treated with IgA-IC. These results suggest that nephritogenic IgA-IC may amplify the proliferation of HMC and the production of immune/chemical mediators and superoxide anion thereby resulting in the renal lesions of IgAN.

Adolescent↗
PubMed1994

Isoprenyl diphosphate synthases: protein sequence comparisons, a phylogenetic tree, and predictions of secondary structure.

Isoprenyl diphosphate synthases are ubiquitous enzymes that catalyze the basic chain-elongation reaction in the isoprene biosynthetic pathway. Pairwise sequence comparisons were made for 6 farnesyl diphosphate synthases, 6 geranylgeranyl diphosphate synthases, and a hexaprenyl diphosphate synthase. Five regions with highly conserved residues, two of which contain aspartate-rich DDXX(XX)D motifs found in many prenyltransferases, were identified. A consensus secondary structure for the group, consisting mostly of alpha-helices, was predicted for the multiply aligned sequences from amino acid compositions, computer assignments of local structure, and hydropathy indices. Progressive sequence alignments suggest that the 13 isoprenyl diphosphate synthases evolved from a common ancestor into 3 distinct clusters. The most distant separation is between yeast hexaprenyl diphosphate synthetase and the other enzymes. Except for the chromoplastic geranylgeranyl diphosphate synthase from Capsicum annuum, the remaining farnesyl and geranylgeranyl diphosphate synthases segregate into prokaryotic/archaebacterial and eukaryotic families.

Alkyl and Aryl Transferases↗
PubMed1994

Prostate cancer screening: role of the digital rectal examination and prostate-specific antigen.

BACKGROUND: This study was designed to determine the efficacy of digital rectal examination (DRE) and serum prostate-specific antigen (PSA) for early detection of prostate cancer in men > or = 50 years of age. METHODS: A prospective single-center clinical trial was conducted to screen 644 asymptomatic men, who were elicited by newspaper and radio advertisements, with DRE and PSA. Quadrant biopsy examinations of the prostate were performed if PSA > 4 ng/ml or if DRE was suspicious. RESULTS: Thirty-seven percent of the men (n = 241) had an abnormality of DRE or elevated PSA. Of the 163 patients who underwent transrectal ultrasound and quadrant biopsies of the prostate, 77% had normal biopsies, 14 (8%) had prostatic intraepithelial neoplasia, and 24 (15%) had carcinoma of the prostate. PSAs ranged from 0.3 to 65.5 ng/ml, with a mean of 2.35 and a median of 1.6. Ninety-five patients had a PSA > 4 ng/ml, of whom 17 had a PSA > 10 ng/ml. Sensitivity of PSA was 75% and specificity 87%; for DRE the sensitivity was 75% and the specificity 69%. Clinical stage of patients who underwent radical prostatectomy was B1 in 15 and B2 in five. Fifteen of 20 patients (75%) had organ-confined disease; the other five had specimen-confined disease. No patient was found to have nodal involvement. CONCLUSION: The combination of PSA and DRE seems to improve the stage of diagnosis of patients with prostate cancer. Larger, randomized studies will be necessary to evaluate the effect of screening on overall survival.

Aged↗
PubMed1994

The alpha v beta 6 integrin promotes proliferation of colon carcinoma cells through a unique region of the beta 6 cytoplasmic domain.

Cell-matrix interactions are assumed to be important in regulating differentiation and tumor cell growth; however, the precise roles of individual matrix receptors in producing cellular responses are still unclear. We have previously described the alpha v beta 6 integrin, an epithelial cell fibronectin receptor expressed in many carcinoma cell lines. Here we show that heterologous expression of alpha v beta 6 in a human colon carcinoma cell line (SW480) enhances the proliferative capacity of these cells, both in vitro and in vivo in nude mice. This property of alpha v beta 6 correlates with the presence of an 11-amino acid region at the COOH terminus of the beta 6 cytoplasmic domain. This 11-amino acid sequence is required for the growth stimulatory effect, but not for other functions of the beta 6 cytoplasmic domain, such as promoting cell adhesion and focal contact localization.

Amino Acid Sequence↗
PubMed1994

Enhanced T cell migration to sites of microscopic CNS disease: complementary treatments evaluated by 2- and 3-D image analysis.

Novel therapies are being developed to attack tumour or other abnormal cells within the brain. A general problem is the need for delivery to sites of microscopic disease. Leukocytes offer an attractive solution; they are able to both move through tissue and recognize abnormal targets. Leukocytes may act as effectors, or as vehicles for drugs, retroviral vectors or other agents. Here, we illustrate complementary ways of enhancing leukocyte migration to sites of microscopic central nervous system (CNS) disease. Enhanced T cell migration to sites of disseminated tumour is used as the example. Computer-assisted image analysis is used to evaluate migration patterns in 2 and 3 dimensions. Shared regulatory features in the migration of tumour and responding cells, and the opportunities and questions they imply, are discussed.

Brain Diseases↗
PubMed1994

Evidence that L-(+)-lactate dehydrogenase deficiency is lethal in Streptococcus mutans.

In order to construct an effector strain for the replacement therapy of dental caries, we wished to combine the properties of low-level acid production and high-level colonization potential in a strain of Streptococcus mutans. To this end, we made a deletion in the lactate dehydrogenase (LDH) gene cloned from the bacteriocin-producing S. mutans strain JH1000. However, we were unable to substitute the mutant for the wild-type allele by transformation with linear DNA fragments. The mutated gene, carried on a suicide vector, was shown by Southern analysis to integrate into the JH1000 chromosome to yield transformants carrying both the wild-type gene and mutated LDH gene. Three spontaneous self-recombinants of one heterodiploid strain were isolated by screening 1,500 colonies for a loss of the tetracycline resistance encoded by the gene used to mark the LDH deletion. In all three cases, Southern analysis showed that a loss of tetracycline resistance was accompanied by a loss of the mutated LDH gene, resulting in restoration of the wild-type genotype. However, screening the same number of colonies for self-recombinants that did not make lactic acid during anaerobic growth in Todd-Hewitt broth failed to identify clones in which the wild-type allele was lost. A second, simpler screening of more than 80,000 colonies grown aerobically on glucose tetrazolium medium to identify low-level-acid-producing colonies was also unsuccessful. These results are interpreted as indicating that LDH deficiency is lethal in S. mutans under the cultivation conditions used in these experiments. The physiological bases for this hypothesis are described.

Blotting, Southern↗
PubMed1994

L-(+)-lactate dehydrogenase deficiency is lethal in Streptococcus mutans.

The previously cloned gene for L-(+)-lactate dehydrogenase (LDH) from Streptococcus mutans was mutagenized in vitro. An Escherichia coli transformant which expressed a thermolabile LDH activity was identified. The ldh(Ts) gene was introduced into S. mutans on a suicide vector to create a heterodiploid expressing both wild-type and thermolabile LDH activities. Self-recombinants which had only one ldh gene were isolated. One of these clones expressed only the thermolabile LDH activity. This isolate grew well at 30 degrees C but did not grow at 42 degrees C under a variety of cultivation conditions, thereby proving that LDH deficiency is lethal in S. mutans in the absence of compensatory mutations.

Blotting, Southern↗
PubMed1994

Phase I and pharmacokinetic study of recombinant human granulocyte-macrophage colony-stimulating factor given in combination with fluorouracil plus calcium leucovorin in metastatic gastrointestinal adenocarcinoma.

PURPOSE: To determine the toxicities and potential for dose escalation of intravenous (IV) bolus fluorouracil (5-FU) given with 500 mg/m2/d leucovorin (LCV) and granulocyte-macrophage colony-stimulating factor (GM-CSF). PATIENTS AND METHODS: Thirty-seven patients received escalating doses of 5-FU/LCV on days 1 to 5 with subcutaneous GM-CSF either 5 or 10 micrograms/kg/d starting on day 6 or 3 micrograms/kg/d starting on day 1. 5-FU was escalated from 370 mg/m2/d by 15% increments between patient cohorts and within patients according to tolerance. RESULTS: With GM-CSF starting on day 6, dose-limiting toxicity occurred during cycle no. 1 in all three patients entered at 5-FU 490 mg/m2/d. However, individual patients tolerated 5-FU doses up to 644 mg/m2/d. When all cycles were analyzed, grade 3 to 4 mucositis and grade 4 granulocytopenia complicated < or = 15% and < or = 6% of cycles with 5-FU doses < or = 560 mg/m2/d (115 cycles). With GM-CSF starting on day 1, dose-limiting granulocytopenia occurred during cycle no. 1 in five of 10 patients entered at 5-FU 490 mg/m2/d. Although the granulocyte nadirs were significantly lower at each 5-FU dose level with the concurrent GM-CSF schedule (eg, 490 mg/m2/d: median, 879/microL v3,286/microL; two-tailed P [P2] < .001), dose-limiting granulocytopenia complicated < or = 16% of cycles with 5-FU < or = 560 mg/m2/d (99 cycles); > or = grade 3 mucositis occurred in < or = 20% of cycles. Grade 3 to 4 diarrhea was unusual with either GM-CSF schedule. Most patients treated with GM-CSF > or = 5 micrograms/kg/d required dose reductions for constitutional toxicity; 3.0 to 3.8 micrograms/kg/d was better tolerated. Venous thrombosis occurred in 17% of patients (concurrent v sequential GM-CSF, 29% v 5%; P2 = .08). The median delivered 5-FU dose-intensity for GM-CSF starting either on day 6 or on day 1 was 615 and 647 mg/m2/wk (P2 = .41), respectively. Pharmacologic exposure to 5-FU increased with higher doses of 5-FU, and concurrent GM-CSF administration did not affect 5-FU clearance. CONCLUSION: A starting dose of 425 mg/m2/d of 5-FU with LCV on days 1 to 5 could be safely combined with GM-CSF starting either on day 1 or day 6, with further 5-FU dose escalation according to individual tolerance.

Adenocarcinoma↗
PubMed1994

Effects of beta subunit cytoplasmic domain deletions on the recruitment of the integrin alpha v beta 6 to focal contacts.

The integrin alpha v beta 6 is expressed primarily in epithelial cells and mediates attachment to the extracellular matrix protein fibronectin. We recently observed that alpha v beta 6 localizes to focal contacts when cells that express the receptor are plated on fibronectin. To determine which regions of the beta 6 cytoplasmic domain are required for recruitment to focal contacts, the wild type human beta 6 cytoplasmic domain are required for recruitment to focal contacts, the wild type human beta 6 subunit and mutants containing selective deletions within the beta 6 cytoplasmic domain were stably expressed in Chinese Hamster Ovary cells. The beta 6 cytoplasmic domain is composed of a 41 amino acid region that is highly conserved among integrin beta subunits, and a unique 11 amino acid carboxy terminal extension. The 11 amino acid extension was not required for localization to focal contacts; in fact, its removal appeared to increase receptor localization. However, removal of any of the 3 conserved regions previously identified as important for the localization of beta 1 integrins to focal contacts (Reszka, et al., 1992, J. Cell Biol. 117:1321-30) eliminated recruitment of beta 6 to focal contacts. These data suggest that, as with the integrin beta 1 subunit, multiple regions within the first 41 amino acids of the beta 6 cytoplasmic domain are necessary for recruitment of alpha v beta 6 to focal contacts. However, the unique 11 amino acid carboxy terminus of beta 6 is not required for this recruitment, and may actually play a negative regulatory role.

Actins↗