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D Chen

Publications and source records attributed to D Chen.

At least 379 records · Page 21Linked to original sources
PubMed1998

Molecular cloning and characterization of LR3, a novel LDL receptor family protein with mitogenic activity.

We report molecular cloning and initial functional characterization of a novel member of the low density lipoprotein receptor (LDLR) gene family. The cDNA was isolated from a human osteoblast cDNA library and encoded a 1,615 amino acids protein designated as LR3. It has, in the extracellular region, a cluster of three LDLR ligand binding repeats at a juxtamembrane position and four EGF precursor homology domains separated by YWTD spacer repeats. The entire ectodomain shares the same modular organization with the middle portion of the extracellular regions of two LDLR family members, LDLR-related protein (LRP), and gp330/megalin. LR3 mRNA was expressed in most of the adult and fetal tissues examined. The highest expression level was seen in aorta. In human osteosarcoma cells examined, LR3 mRNA was highly enriched in TE85 cells, moderately expressed in MG63 cells and primary human osteoblasts, and undetectable in SaOS-2 cells. NIH 3T3 cells transfected with either full length LR3 or its ectodomain showed significantly increased proliferation, whereas transfection of intracellular domain had no proliferative effect. We predict that LR3 is a multi-functional protein with potential mitogenic activity.

3T3 Cells↗
PubMed1998

Multiple forms of rat stomach histidine decarboxylase may reflect posttranslational activation of the enzyme.

Histidine decarboxylase (HDC) catalyzes the formation of histamine, which takes part in a variety of physiological processes including gastric acid secretion, neurotransmission and inflammation. While purified rat HDC is a homodimer of approximately 54 kDa subunits, molecular cloning of mammalian HDC has revealed that HDC mRNA encodes a 74 kDa protein. This discrepancy in molecular mass may be due to a posttranslational processing of the primary translated product of rat HDC mRNA. In the present study we demonstrate that full-length rat HDC expressed in Escherichia coli or in an in vitro transcription/translation system is enzymatically inactive, while expression of a C-terminus truncated HDC (reducing the molecular mass to 54 kDa) gave rise to a protein with high enzyme activity in the same expression systems. COS-7 cells expressing truncated HDC displayed high HDC activity, whereas COS-7 cells expressing full-length HDC displayed low activity. Western blot analysis of fetal rat liver and oxyntic mucosa of gastrin-stimulated rats revealed the presence of both full-length HDC (approximately 73 kDa) and a approximately 53 kDa subunit form in addition to an intermediate form of about 63 kDa. The results are in line with the view that rat HDC may be produced as an enzymatically inactive proenzyme which is processed to give rise to the active enzyme.

Animals↗
PubMed1998

Novel aspects of gastrin-induced activation of histidine decarboxylase in rat stomach ECL cells.

The ECL cells in the rat stomach respond to gastrin with secretion of histamine and activation of the histamine-forming enzyme histidine decarboxylase (HDC). In the present study, we have investigated factors that influence gastrin-induced activation of HDC. Gastrin-17 was given by continuous intravenous infusion to fasted and freely fed rats in various doses and for various periods of time. We found that: (1) ECL cells in fasted rats displayed one order of magnitude higher sensitivity to gastrin (3 h infusion) than did ECL cells in fed rats (ED50 0.4 versus 4.0 nmol kg(-1) h(-1)), while the maximum response to gastrin was two times greater in fed rats than in fasted rats; (2) HDC in both fasted and fed rats responded to a high gastrin dose (5 nmol kg(-1) h(-1)) in a biphasic manner with peak activity after 8 h in fasted rats and after 16 h in fed rats. In both groups, the activation was followed by a marked decline in the enzyme activity to almost prestimulation levels 24 h after start of the infusion. A low gastrin dose (0.4 nmol kg(-1) h(-1)) did not induce such a biphasic response. Maximum activation of HDC in fed rats occurred 6 days after starting the infusion of the low gastrin dose and was two times higher than the maximum activation observed after the high gastrin dose; (3) In fasted rats the HDC mRNA level rose in response to the high gastrin dose, peaked after 8 h (twofold increase) and then returned to the prestimulation level. In fed rats the increase was slower, reaching a plateau after 24 h that lasted for 6 days (twofold increase); (4) The translation inhibitor cycloheximide blocked the activation of HDC induced by gastrin (4 h infusion of 5 nmol kg(-1) h(-1)), while the transcription inhibitor actinomycin D, which suppressed the increase in HDC mRNA expression, did not.

Animals↗
PubMed1998

Replication of R6K gamma origin in vitro: discrete start sites for DNA synthesis dependent on pi and its copy-up variants.

The regulation of the plasmid R6K gamma origin (gamma ori) is accomplished through the ability of the pi protein to act as an initiator and inhibitor of replication. Hyperactive variants of this protein, called copy-up pi, allow four to tenfold increases of gamma ori plasmid DNA in vivo. The higher activity of copy-up pi variants could be explained by an increase in the initiator function, a decrease in the inhibitor activity, or a derepression of a more efficient mechanism of replication that can be used by wt pi (pi35. 0) only under certain conditions. We have compared the replication activities of wt pi35.0 and copy-up pi mutants in vitro, and analyzed the replication products. It is shown that copy-up variants are several-fold more active than wt pi35.0 in replication. This appears to be due to enhanced specific replication activity of copy-up mutants rather than elevated fractions of protein proficient in DNA binding. Furthermore, biochemical complementation revealed that pi200 (copy-up) is dominant over wt pi35.0. The elevated activity of copy-up pi is not caused by an increased rate of replisome assembly as inferred from in vitro replication assays in which the lag periods observed were similar to that of wt pi35.0. Moreover, only one round of semiconservative, unidirectional replication occurred in all the samples analyzed indicating that copy-up pi proteins do not initiate multiple rounds of DNA synthesis. Rather, a larger fraction of DNA template replicates in the presence of copy-up pi as determined by electron microscopy. Two clusters of discrete DNA synthesis start sites are mapped by primer extension near the stability (stb) locus of the gamma ori. We show that the start sites are the same in the presence of wt pi35.0 or copy-up proteins. This comparative analysis suggests that wt pi35.0 and copy-up variants utilize fundamentally similar mechanism(s) of replication priming.

Amino Acid Substitution↗
PubMed1998

Phenylalanine hydroxylase from Chromobacterium violaceum. Uncoupled oxidation of tetrahydropterin and the role of iron in hyroxylation.

A gene encoding phenylalanine hydroxylase has been cloned from Chromobacterium violaceum and expressed in Escherichia coli. The purified phenylalanine hydroxylase contains copper, which does not support enzymatic activity. Upon removal of copper by dithiothreitol (DTT), the enzyme contains substoichiometric amounts of calcium and zinc but little or no redox-active metal ions. The copper-depleted hydroxylase catalyzes the phenylalanine-dependent oxidation of 6, 7-dimethyltetrahydropterin (DMPH4) by O2 in a reaction in which phenylalanine is not hydroxylated and does not appear to undergo a chemical change, and hydrogen peroxide is produced. Analogs of phenylalanine also activate the oxidation of DMPH4. Both the copper-phenylalanine hydroxylase and the copper-depleted hydroxylase catalyze the hydroxylation of phenylalanine in the presence of DTT and FeSO4 in a reaction in which hydrogen peroxide is not produced. The apparent values of Km for Fe2+ and DTT are 0.28 microM and 1.1 mM, respectively, at 1.0 mM phenylalanine, 120 microM DMPH4 and pH 7. 4 and 23 degreesC. The apparent value of kcat is 14.3 s-1 under these conditions. Glutathione, mercaptoethanol, and dihydrolipoate support the hydroxylation of phenylalanine essentially as well as DTT. Incubation of copper-depleted hydroxylase with FeSO4, phenylalanine, and DTT followed by gel permeation chromatography leads to an iron-hydroxylase containing approximately 1 molecule of iron per molecule of enzyme. The iron-hydroxylase displays an optical absorption band extending from 300 to 600 nm, and it catalyzes the hydroxylation of phenylalanine at the same maximum rate as the iron-activated hydroxylase but does not require added Fe2+. We conclude that iron participates in the hydroxylation of phenylalanine. Iron is not required for the oxidation of DMPH4, although it may exert a modest acceleration effect. A hypothetical mechanism is presented wherein the reaction of iron with the putative 4a-hydroperoxy-DMPH4 leads to 4a-hydroxy-DMPH4 and a high valent iron-oxy species. The iron-oxy species is postulated to react with phenylalanine in the hydroxylation process.

Amino Acid Sequence↗
PubMed1998

Exploring structure-activity relationships around the phosphomannose isomerase inhibitor AF14049 via combinatorial synthesis.

Phosphomannose Isomerase (PMI) has been shown by genetic methods to be an essential enzyme in fungal cell wall biosynthesis. The PMI inhibitor AF14049 was discovered as an unanticipated side product from high-throughput library screening against the enzyme from C, albicans. Solid-phase synthetic methods were developed and a series of libraries and discrete analogs synthesized to explore SAR around AF14049.

Amides↗
PubMed1998

Tie2 receptor ligands, angiopoietin-1 and angiopoietin-2, modulate VEGF-induced postnatal neovascularization.

Angiopoietin-1 (Ang1) has been recently identified as the major physiological ligand for the tyrosine kinase receptor Tie2 and assigned responsibility for recruiting and sustaining periendothelial support cells. Angiopoietin-2 (Ang2) was found to disrupt blood vessel formation in the developing embryo by antagonizing the effects of Ang1 and Tie2 and was thus considered to represent a natural Ang1/Tie2 inhibitor. In vivo effects of either angiopoietin on postnatal neovascularization, however, have not been previously described. Accordingly, we used the cornea micropocket assay of neovascularization to investigate the impact of angiopoietins on neovascularization in vivo. Neither Ang1 nor Ang2 alone promoted neovascularization. Pellets containing vascular endothelial growth factor (VEGF) alone induced corneal neovascularity extending from the limbus across the cornea. Addition of Ang 1 to VEGF (Ang1+VEGF) produced an increase in macroscopically evident perfusion of the corneal neovasculature without affecting macroscopic measurements of length (0.58+/-0.03 mm) or circumferential neovascularity (136+/-10 degrees). In contrast, pellets containing Ang2+VEGF promoted significantly longer (0.67+/-0.05 mm) and more circumferential (160+/-15degrees) neovascularity than VEGF alone or Ang1+VEGF (P<0.05). Excess soluble Tie2 receptor (sTie2-Fc) precluded modulation of VEGF-induced neovascularization by both Ang2 and Ang1. Fluorescent microscopic findings demonstrated enhanced capillary density (fluorescence intensity, 2.55+/-0.23 e+9 versus 1.23+/-0.17 e+9, P<0.01) and increased luminal diameter of the basal limbus artery (39.0+/-2.8 versus 27.9+/-1.3 microm, P<0.01) for Ang1+VEGF compared with VEGF alone. In contrast to Ang1+VEGF, Ang2+VEGF produced longer vessels and, at the tip of the developing capillaries, frequent isolated sprouting cells. In the case of Ang2+VEGF, however, luminal diameter of the basal limbus artery was not increased (26.7+/-1.9 versus 27.9+/-1.3, P=NS). These findings constitute what is to our knowledge the first direct demonstration of postnatal bioactivity associated with either angiopoietin. In particular, these results indicate that angiopoietins may potentiate the effects of other angiogenic cytokines. Moreover, these findings provide in vivo evidence that Ang1 promotes vascular network maturation, whereas Ang2 works to initiate neovascularization.

Angiopoietin-1↗
PubMed1998

A neuronal ryanodine receptor mediates light-induced phase delays of the circadian clock.

Circadian clocks are complex biochemical systems that cycle with a period of approximately 24 hours. They integrate temporal information regarding phasing of the solar cycle, and adjust their phase so as to synchronize an organism's internal state to the local environmental day and night. Nocturnal light is the dominant regulator of this entrainment. In mammals, information about nocturnal light is transmitted by glutamate released from retinal projections to the circadian clock in the suprachiasmatic nucleus of the hypothalamus. Clock resetting requires the activation of ionotropic glutamate receptors, which mediate Ca2+ influx. The response induced by such activation depends on the clock's temporal state: during early night it delays the clock phase, whereas in late night the clock phase is advanced. To investigate this differential response, we sought signalling elements that contribute solely to phase delay. We analysed intracellular calcium-channel ryanodine receptors, which mediate coupled Ca2+ signalling. Depletion of intracellular Ca2+ stores during early night blocked the effects of glutamate. Activators of ryanodine receptors induced phase resetting only in early night; inhibitors selectively blocked delays induced by light and glutamate. These findings implicate the release of intracellular Ca2+ through ryanodine receptors in the light-induced phase delay of the circadian clock restricted to the early night.

Animals↗
PubMed1998

Differential roles for bone morphogenetic protein (BMP) receptor type IB and IA in differentiation and specification of mesenchymal precursor cells to osteoblast and adipocyte lineages.

Cumulative evidence indicates that osteoblasts and adipocytes share a common mesenchymal precursor and that bone morphogenetic proteins (BMPs) can induce both osteoblast and adipocyte differentiation of this precursor. In the present study, we investigated the roles of BMP receptors in differentiation along these separate lineages using a well-characterized clonal cell line, 2T3, derived from the mouse calvariae. BMP-2 induced 2T3 cells to differentiate into mature osteoblasts or adipocytes depending upon culture conditions. To test the specific roles of the type IA and IB BMP receptor components, truncated and constitutively active type IA and IB BMP receptor cDNAs were stably expressed in these cells. Overexpression of truncated type IB BMP receptor (trBMPR-IB) in 2T3 cells completely blocked BMP-2-induced osteoblast differentiation and mineralized bone matrix formation. Expression of trBMPR-IB also blocked mRNA expression of the osteoblast specific transcription factor, Osf2/ Cbfa1, and the osteoblast differentiation-related genes, alkaline phosphatase (ALP) and osteocalcin (OC). BMP-2-induced ALP activity could be rescued by transfection of wild-type (wt) BMPR-IB into 2T3 clones containing trBMPR-IB. Expression of a constitutively active BMPR-IB (caBMPR-IB) induced formation of mineralized bone matrix by 2T3 cells without addition of BMP-2. In contrast, overexpression of trBMPR-IA blocked adipocyte differentiation and expression of caBMPR-IA induced adipocyte formation in 2T3 cells. Expression of the adipocyte differentiation-related genes, adipsin and PPARgamma, correlated with the distinct phenotypic changes found after overexpression of the appropriate mutant receptors. These results demonstrate that type IB and IA BMP receptors transmit different signals to bone-derived mesenchymal progenitors and play critical roles in both the specification and differentiation of osteoblasts and adipocytes.

Adipocytes↗
PubMed1998

SNAP-23 requirement for transferrin recycling in Streptolysin-O-permeabilized Madin-Darby canine kidney cells.

Fusion of recycling and transcytotic vesicles with the apical and basolateral plasma membrane domains of Madin-Darby canine kidney (MDCK) cells requires the N-ethylmaleimide-sensitive factor and is sensitive to botulinum neurotoxin serotype E (BoNT/E). BoNT/E is thought to selectively proteolyze the 25,000-dalton synaptosomal associated protein (SNAP-25), a protein found in neurons or cells of neuroendocrine origin. However, SNAP-25 is not found in MDCK cells. One possible target for BoNT/E in MDCK cells is SNAP-23, a newly described SNAP-25 homolog that is found in several organs including kidney. Currently, the function of SNAP-23 is unknown. We have reconstituted transferrin recycling in permeabilized MDCK cells to assess the role of SNAP-23 in the endocytic traffic of this protein. We find that: (i) SNAP-23 is expressed in MDCK cells and is found both at the basolateral plasma membrane and associated with apical and basolateral vesicles, (ii) canine SNAP-23 is cleaved by BoNT/E, (iii) transferrin recycling is N-ethylmaleimide-sensitive factor-dependent and BoNT/E-sensitive, and (iv) addition of either exogenous SNAP-23 or anti-SNAP-23 antibodies inhibits ligand recycling. Our observations suggest that SNAP-23 may be required for fusion of recycling vesicles with the basolateral membrane of polarized MDCK cells.

Animals↗
PubMed1998

Transcription elongation factor P-TEFb mediates Tat activation of HIV-1 transcription at multiple stages.

Tat stimulates human immunodeficiency virus type 1 (HIV-1) transcription elongation through recognition of the transactivation response (TAR) RNA stem-loop structure at the 5' end of nascent viral transcripts. Recently, a human transcription elongation factor P-TEFb, consisting of CDK9 kinase, cyclin T and other associated factors, has been shown to interact with Tat to restore Tat activation in HeLa nuclear extract depleted of P-TEFb. Here, we report the purification of a P-TEFb complex fraction containing epitope-tagged wild-type CDK9 or kinase-inactive CDK9 and five tightly associated polypeptides. Only wild-type P-TEFb complex with an active CDK9 kinase was able to hyperphosphorylate the C-terminal domain of RNA polymerase II and mediate Tat transactivation in P-TEFb-depleted HeLa nuclear extract. Tat also stimulated transcription elongation by recruitment of the P-TEFb complex to the HIV-1 promoter through a Tat-TAR interaction. A possible mechanism for P-TEFb to become associated with polymerase elongation complexes and function as a general elongation factor was demonstrated by an interaction of P-TEFb with double-stranded RNA molecules through an 87 kDa subunit. Finally, P-TEFb was found to interact with and phosphorylate Tat-SF1, a Tat cofactor required for Tat transactivation. Our data indicate that the various subunits of the human P-TEFb complex may play distinct roles at multiple stages to mediate Tat activation of HIV-1 transcription elongation.

Base Sequence↗
PubMed1998

Negative regulation of the heat shock transcriptional response by HSBP1.

In response to stress, heat shock factor 1 (HSF1) acquires rapid DNA binding and transient transcriptional activity while undergoing conformational transition from an inert non-DNA-binding monomer to active functional trimers. Attenuation of the inducible transcriptional response occurs during heat shock or upon recovery at non-stress conditions and involves dissociation of the HSF1 trimer and loss of activity. We have used the hydrophobic repeats of the HSF1 trimerization domain in the yeast two-hybrid protein interaction assay to identify heat shock factor binding protein 1 (HSBP1), a novel, conserved, 76-amino-acid protein that contains two extended arrays of hydrophobic repeats that interact with the HSF1 heptad repeats. HSBP1 is nuclear-localized and interacts in vivo with the active trimeric state of HSF1 that appears during heat shock. During attenuation of HSF1 to the inert monomer, HSBP1 associates with Hsp70. HSBP1 negatively affects HSF1 DNA-binding activity, and overexpression of HSBP1 in mammalian cells represses the transactivation activity of HSF1. To establish a biological role for HSBP1, the homologous Caenorhabditis elegans protein was overexpressed in body wall muscle cells and was shown to block activation of the heat shock response from a heat shock promoter-reporter construct. Alteration in the level of HSBP1 expression in C. elegans has severe effects on survival of the animals after thermal and chemical stress, consistent with a role for HSBP1 as a negative regulator of the heat shock response.

3T3 Cells↗
PubMed1998

Acetylsalicylate administered during simulated ischemia reduces the recovery of neuronal function in the in vitro rabbit retina.

Aspirin is widely used as an analgesic, in the secondary prevention of stroke, and has recently been suggested to be a putative neuroprotective agent, yet whether it acts directly on the central nervous system (CNS) is not yet clarified. We therefore examined the effect of lysine acetylsalicylate (L-ASA, 4-2000 microM) on neuronal function under normal conditions and following 1 h of ischemia using the in vitro rabbit retina preparation. L-ASA inhibited the light-evoked compound action potentials, but not the electroretinogram, in a concentration-dependent manner. In addition, L-ASA (2000 microM, but not 4, 40 or 200 microM) administered during ischemia, reduced the recovery of neuronal function compared to control (untreated) retinas. L-ASA therefore inhibits CNS neurotransmission, but not phototransduction, in a concentration-dependent manner. In addition, high concentration L-ASA impairs the recovery of neuronal function following an ischemic episode.

Action Potentials↗
PubMed1998

Cadherin-6 mediates the heterotypic interactions between the hemopoietic osteoclast cell lineage and stromal cells in a murine model of osteoclast differentiation.

Osteoclasts are multinucleated cells of hemopoietic origin that are responsible for bone resorption during physiological bone remodeling and in a variety of bone diseases. Osteoclast development requires direct heterotypic cell-cell interactions of the hemopoietic osteoclast precursors with the neighboring osteoblast/stromal cells. However, the molecular mechanisms underlying these heterotypic interactions are poorly understood. We isolated cadherin-6 isoform, denoted cadherin-6/2 from a cDNA library of human osteoclast-like cells. The isolated cadherin-6/2 is 3,423 bp in size consisting of an open reading frame of 2,115 bp, which encodes 705 amino acids. This isoform lacks 85 amino acids between positions 333 and 418 and contains 9 different amino acids in the extracellular domain compared with the previously described cadherin-6. The human osteoclast-like cells also expressed another isoform denoted cadherin-6/1 together with the cadherin-6. Introduction of cadherin-6/2 into L-cells that showed no cell-cell contact caused evident morphological changes accompanied with tight cell-cell association, indicating the cadherin-6/2 we isolated here is functional. Moreover, expression of dominant-negative or antisense cadherin-6/2 construct in bone marrow-derived mouse stromal ST2 cells, which express only cadherin-6/2, markedly impaired their ability to support osteoclast formation in a mouse coculture model of osteoclastogenesis. Our results suggest that cadherin-6 may be a contributory molecule to the heterotypic interactions between the hemopoietic osteoclast cell lineage and osteoblast/bone marrow stromal cells required for the osteoclast differentiation. Since both osteoclasts and osteoblasts/bone marrow stromal cells are the primary cells controlling physiological bone remodeling, expression of cadherin-6 isoforms in these two cell types of different origin suggests a critical role of these molecules in the relationship of osteoclast precursors and cells of osteoblastic lineage within the bone microenvironment.

Amino Acid Sequence↗
PubMed1998

[A molecular variant of angiotensinogen gene is associated with myocardial infarction in Chinese].

OBJECTIVE: A molecular variant of the angiotensinogen(AGT) gene with threonine instead of methionine at position 235(i.e.,with M235T polymor- phism) has been shown to be associated with essential hypertension, preeclampsia, coronary atherosclerosis, coronary heart disease(CHD), myocardial infarction(MI) in Caucasians and in Japanese. The purpose of the present study was to assess whether the M235T polymorphism was associated with MI in a Chinese population. METHODS: M235T polymorphism in exon 2 of AGT gene was determined by polymerase chain reaction(PCR) and restriction endonuclease analysis in a study of 57 patients with MI and 76 non-CHD individuals as control. RESULTS: The frequencies of T235 allele(0.82) and 235TT genotype (0.70) in the MI group were higher than those in the control subjects(0.63 and 0.42 respectively, P=0.013 and P<0.025). AGT gene 235TT genotype was at significantly increased risk of MI(odds ratio 3.65, P=0.016) in analysis adjusted for several main CHD risk factors. CONCLUSION: There is a significant association between AGT gene 235TT genotype and MI, this genotype might be an independent risk for MI in Chinese population.

Aged↗
PubMed1998

Nitric oxide synthase modulates angiogenesis in response to tissue ischemia.

We tested the hypothesis that endothelial nitric oxide synthase (eNOS) modulates angiogenesis in two animal models in which therapeutic angiogenesis has been characterized as a compensatory response to tissue ischemia. We first administered L-arginine, previously shown to augment endogenous production of NO, to normal rabbits with operatively induced hindlimb ischemia. Angiogenesis in the ischemic hindlimb was significantly improved by dietary supplementation with L-arginine, compared to placebo-treated controls; angiographically evident vascularity in the ischemic limb, hemodynamic indices of limb perfusion, capillary density, and vasomotor reactivity in the collateral vessel-dependent ischemic limb were all improved by oral L-arginine supplementation. A murine model of operatively induced hindlimb ischemia was used to investigate the impact of targeted disruption of the gene encoding for ENOS on angiogenesis. Angiogenesis in the ischemic hindlimb was significantly impaired in eNOS-/- mice versus wild-type controls evaluated by either laser Doppler flow analysis or capillary density measurement. Impaired angiogenesis in eNOS-/- mice was not improved by administration of vascular endothelial growth factor (VEGF), suggesting that eNOS acts downstream from VEGF. Thus, (a) eNOS is a downstream mediator for in vivo angiogenesis, and (b) promoting eNOS activity by L-arginine supplementation accelerates in vivo angiogenesis. These findings suggest that defective endothelial NO synthesis may limit angiogenesis in patients with endothelial dysfunction related to atherosclerosis, and that oral L-arginine supplementation constitutes a potential therapeutic strategy for accelerating angiogenesis in patients with advanced vascular obstruction.

Animals↗
PubMed1998

Reversal of tolerance to human MUC1 antigen in MUC1 transgenic mice immunized with fusions of dendritic and carcinoma cells.

Immunological unresponsiveness established by the elimination or anergy of self-reactive lymphocyte clones is of importance to immunization against tumor-associated antigens. In this study, we have investigated induction of immunity against the human MUC1 carcinoma-associated antigen in MUC1 transgenic mice unresponsive to MUC1 antigen. Immunization of adult MUC1 transgenic mice with irradiated MUC1-positive tumor cells was unsuccessful in reversing unresponsiveness to MUC1. By contrast, fusions of dendritic cells with MUC1-positive tumor cells induced cellular and humoral immunity against MUC1. Immunization with the dendritic cell fusions that express MUC1 resulted in the rejection of established metastases and no apparent autoimmunity against normal tissues. These findings demonstrate that unresponsiveness to the MUC1 tumor-associated antigen is reversible by immunization with heterokaryons of dendritic cells and MUC1-positive carcinoma cells.

Adult↗