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X Cao

Publications and source records attributed to X Cao.

At least 37 records · Page 2Linked to original sources
PubMed2002

Lymphotactin cotransfection enhances the therapeutic efficacy of dendritic cells genetically modified with melanoma antigen gp100.

Lymphotactin (Lptn) is a C chemokine that attracts T cells and NK cells. Dendritic cells (DC) are highly efficient, specialized antigen-presenting cells and antigen-pulsed DC has been regarded as promising vaccines in cancer immunotherapy. The aim of our present study is to improve the therapeutic efficacy of DC-based tumor vaccine by increasing the preferential chemotaxis of DC to T cells. In this study, Lptn and/or melanoma-associated antigen gp100 were transfected into mouse bone marrow-derived DC, which were used as vaccines in B16 melanoma model. Immunization of C57BL/6 mice with DC adenovirally cotransfected with Lptn and gp100 (Lptn/gp100-DC) could enhance the cytotoxicities of CTL and NK cells, increase the production of IL-2 and interferon-gamma significantly, as compared with immunization with gp100-DC, Lptn-DC, LacZ-DC, DC or PBS counterparts. The Lptn/gp100-DC immunized mice exhibited resistance to tumor challenge most effectively. It was found that the tumor mass of mice vaccinated by Lptn/gp100-DC showed obvious necrosis and inflammatory cell infiltration. In vivo depletion analysis demonstrated that CD8(+) T cells are the predominant T cell subset responsible for the antitumor effect of Lptn/gp100-DC and CD4(+) T cells were necessary in the induction phase of tumor rejection, while NK cells were less important although they participated in the antitumor response either in the induction phase or in the effector phase. In the murine model with the pre-established subcutaneous B16 melanoma, immunization with Lptn/gp100-DC inhibited the tumor growth most significantly when compared with other counterparts. These findings provide a potential strategy to improve the efficacy of DC-based tumor vaccines.

Animals↗
PubMed2002

Pig amelogenin gene expresses a unique exon 4.

The pig amelogenin gene was isolated from a Lambda genomic library, and a 6.3 kb SalI/XbaI restriction fragment, inclusive of exons 3 through 7, was subcloned into a plasmid vector. DNA sequencing revealed two putative exon 4 sequences. The derived amino acid sequence of exon 4a, KSGRWGARLTAFVSSVQ, had previously been identified in a 190-amino-acid amelogenin isoform by protein sequencing. Exon 4b encoded the peptide DLYLEAIRIDRTAF, which is homologous to exon 4-encoded segments reported for human, mouse, and rat. Oligonucleotides from both of these exons were used to amplify cDNA generated from developing teeth. Amplification products were analyzed by agarose gel electrophoresis, cloned, and characterized by DNA sequencing. Exon 4a was found in transcripts encoding amelogenin isoforms having 190 and 73 amino acids. Exon 4b was found only in apparent splicing intermediates that retained intron 3, but was not detected in any final mRNA transcripts. Pig amelogenin having apparent molecular mass of 23 kD were isolated from the enamel matrix and characterized by mass spectrometry. Two mass values, 18,512.5, and 18,571.2 Da, were measured that match the values predicted for the 162-amino-acid cleavage product of the 173-amino-acid amelogenin, and the 165-amino-acid cleavage product of the 190-amino-acid amelogenin, which includes 17 amino acids encoded by exon 4a. We conclude that the pig amelogenin gene expresses a unique exon 4 that is not homologous to, or evolved from, the exon 4 segment expressed in humans and rodents.

Alternative Splicing↗
PubMed2002

Photoimmobilization of biomolecules within a 3-dimensional hydrogel matrix.

It has been recognized that a three-dimensional cell invasive scaffold that provides both topographical and chemical cues is desirable in regenerative tissue engineering to encourage cell attachment, migration, regrowth and ultimately tissue repair. Carbohydrate hydrogels are attractive for such applications because they are generally biocompatible and able to match the mechanical properties of most soft tissues. Although carbohydrate hydrogels have been previously modified with cell adhesive peptides and proteins, complicated hydrogel matrix activation was required prior to biomolecule coupling and, perhaps more importantly, the overall immobilization yield was low at approximately 1%. In this study, we report the photo-immobilization of a model biomolecule, ovalbumin (OVA), to agarose gel. We describe two methods of modification where the photoactive moiety is coupled to either the protein (i.e. OVA) or the matrix (i.e. agarose) prior to immobilization. We found that the photo-immobilization yield depends on the location of the photoactive moiety. Using photoactive OVA, 1.8% of the OVA initially incorporated into the agarose gel is immobilized; using photoactive agarose, 9.3% of the OVA initially mixed with the agarose is immobilized. The latter is a significant improvement over previous yields and may be useful in attaining our goal of immobilizing a biomolecule gradient for guided tissue regeneration.

Benzophenones↗
PubMed2001

Inhibition of adenovirus cytotoxicity, replication, and E2a gene expression by adeno-associated virus.

Adeno-associated virus (AAV) and the other parvoviruses have long been known to inhibit proliferation of nonpermissive cells. The mechanism of this inhibition is not thoroughly understood. To learn how AAV interacts with host cells, we have begun an investigation into AAV's relationship with adenovirus (Ad), AAV's most efficient helper virus. AAV, but not UV-inactivated AAV, delayed Ad-induced cytotoxicity and inhibited Ad E2a gene expression. AAV, but not UV-inactivated AAV or a recombinant AAV vector, inhibited Ad DNA replication. To determine whether AAV or its replication (Rep) proteins alter Ad early gene expression, we measured steady state E2a mRNA levels in AAV and Ad coinfected cultures and in a cell line (Neo6) that inducibly expresses the Rep proteins. AAV, but not UV-AAV, and Rep expression resulted in diminution of E2a protein and mRNA levels. To determine whether the AAV Rep proteins directly affect the individual Ad early promoters, we constructed luciferase reporter plasmids containing each of the five early promoters. Cotransfection of Ad-luciferase and an AAV rep gene-expressing plasmid in HeLa cells demonstrated that Rep78 repressed the E1a, E2a, and E4 promoters but trans-activated the E1b and E3 promoters. In the presence of a cotransfected E1a-expressing plasmid, Rep78 repressed expression from all five promoters. These results indicate that Rep may have different effects on the Ad early promoters dependent upon the presence of the E1a trans-activating protein.

Adenoviridae↗
PubMed2001

Silicon surfaces as electron acceptors: dative bonding of amines with Si(001) and Si(111) surfaces.

The bonding of the trimethylamine (TMA) and dimethylamine (DMA) with crystalline silicon surfaces has been investigated using X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy, and density-functional computational methods. XPS spectra show that TMA forms stable dative-bonded adducts on both Si(001) and Si(111) surfaces that are characterized by very high N(1s) binding energies of 402.2 eV on Si(001) and 402.4 eV on Si(111). The highly ionic nature of these adducts is further evidenced by comparison with other charge-transfer complexes and through computational chemistry studies. The ability to form these highly ionic charge-transfer complexes between TMA and silicon surfaces stems from the ability to delocalize the donated electron density between different types of chemically distinct atoms within the surface unit cells. Corresponding studies of DMA on Si(001) show only dissociative adsorption via cleavage of the N-H bond. These results show that the unique geometric structures present on silicon surfaces permit silicon atoms to act as excellent electron acceptors.

Journal Article↗
PubMed2001

Intron distribution difference for 276 ancient and 131 modern genes suggests the existence of ancient introns.

o introns delineate elements of protein tertiary structure? This issue is crucial to the debate about the role and origin of introns. We present an analysis of the full set of proteins with known three-dimensional structures that have homologs with intron positions recorded in GenBank. A computer program was generated that maps on a reference sequence the positions of all introns in homologous genes. We have applied this program to a set of 665 nonredundant protein sequences with defined three-dimensional structures in the Protein Data Bank (PDB), which yielded 8,217 introns in 407 proteins. For the subset of proteins corresponding to ancient conserved regions (ACR), we find that there is a correlation of phase-zero introns with the boundary regions of modules and no correlation for the phase-one and phase-two positions. However, for a subset of proteins without prokaryotic counterparts (131 non-ACR proteins), a set of presumably modern proteins (or proteins that have diverged extremely far from any ancestral form), we do not find any correlation of phase-zero intron positions with three-dimensional structure. Furthermore, we find an anticorrelation of phase-one intron positions with module boundaries: they actually have a preference for the interior of modules. This finding is explicable as a preference for phase-one introns to lie in glycines, between G/G sequences, the preference for glycines being anticorrelated with the three-dimensional modules. We interpret this anticorrelation as a sign that a number of phase-one introns, and hence many modern introns, have been inserted into G/G "protosplice" sequences.

Evolution, Molecular↗
PubMed2001

Dendritic cells support hematopoiesis of bone marrow cells.

BACKGROUND: We previously observed that vaccination of normal mice with bone marrow (BM) -derived dendritic cells (DCs) could increase the number of peripheral white blood cells (WBCs) and platelets. In the present study, we investigated the potential of DCs to support the hematopoiesis of BM cells in vitro and in vivo. METHODS: In the absence of exogenous cytokines, the expansion of CD34+ stem cells was observed when cultured with DC-derived supernatant or contact cocultured with DC. After culture in supernatant of DCs or contact coculture with DCs for 3 days, CD34+ progenitor cells were cultured in the semisolid media to test their ability to generate the clonogeneic cells. Then, BM cells combined with DCs or not were transferred into lethally irradiated syngeneic recipients to determine the effects of DCs on hematopoietic recovery. RESULTS: After culture in the supernatant of DCs, especially in the supernatant of OVA-DCs (OVA-stimulated DC), the proliferation of CD34+ stem cells and generation of clonogeneic cells were augmented in correspondence with the concentration of DCs. After contact coculture with DCs, the proliferation of CD34+ stem cells and generation of clonogeneic cells were more significant than that in noncontact cultures. Moreover, when cultured with DCs or supernatant of DCs, CD34+ progenitor cells were preferentially differentiated to megakaryocytes. After coculture with OVA-DCs, markedly greater generation of colony forming units-granulocyte/macrophages (CFU-GM): colony forming units-megakaryocytes (CFU-MK) was found than that in coculture with unstimulated DCs. Pretreatment of DC with antibodies to thrombopoietin (TPO), interleukin (IL) -6, IL-12, or anti-mouse intercellular adhesion molecule-1 (ICAM-1) could inhibit the ability of DCs to support the generation of CFU-GM, CFU-MK. After transplant with BM cells and DCs, the number of peripheral platelets of the recipients increased significantly and, to a lesser extent, peripheral WBC counts increased. The survival periods were significantly prolonged when the lethally irradiated mice were transplanted with BM cells combined with DCs or OVA-DCs. High levels of TPO, IL-6, and IL-12 could be detectable in the supernatant of DCs, and TPO expression by DCs was further confirmed by reverse transcription-polymerase chain reaction analysis and intracellular staining with anti-TPO antibody. CONCLUSIONS: We first demonstrated that DCs, especially antigen-stimulated DCs, can promote the expansion of hematopoietic progenitors and support hematopoiesis, preferentially support megakaryopoiesis of BM cells, by expressing soluble factors, including TPO, IL-6, IL-12, and by direct cell-to-cell interaction with stem cells in vitro and in vivo.

Animals↗
PubMed2001

DIgR1, a novel membrane receptor of the immunoglobulin gene superfamily, is preferentially expressed by antigen-presenting cells.

A novel membrane receptor of immunoglobulin gene superfamily (IgSF) has been identified from mouse dendritic cells (DC) and designated as DC-derived Ig-like receptor 1 (DIgR1). It encodes a 228-amino-acid (aa) residue polypeptide with a 21-aa signal peptide, a 20-aa transmembrane region, a 189-aa extracellular region, and a 19 aa intracellular region. Its extracellular region contains a single V domain of Ig. So it is a novel type I transmembrane glycoprotein of IgSF. DIgR1 shows significant homologies to human CMRF-35 antigens and polymeric immunoglobulin receptors (pIgR). The mRNA expression of DIgR1 was highly abundant in mouse spleen. The preferential expression of DIgR1 mRNA is observed in the known antigen-presenting cells (APC) including DC, monocytes/macrophages, and B lymphocytes. A 40 kDa of protein in NIH/3T3 cells transfected with the DIgR1 cDNA was detected by Western blot analysis using anti-DIgR1 polyclonal antibodies. The expression of DIgR1 protein on DC is not regulated by LPS stimulation. Further study should be conducted to investigate what were biological functions of DIgR1 in the immunobiology of APC.

Amino Acid Sequence↗
PubMed2001

Molecular cloning and characterization of a novel calcium/calmodulin-dependent protein kinase II inhibitor from human dendritic cells.

Calcium/calmodulin-dependent protein kinase II (CaMKII) is a widely distributed protein kinase that regulates numerous physiological functions. Inhibitors of CaMKII are useful tools for investigating the CaMKII functions. Here we identified a novel CaMKII inhibitor protein (CaM-KIIN) from the human dendritic cell cDNA library by large-scale random sequencing. Human CaM-KIIN contains 79 amino acids, which shares 98% identity and 98% positives with rat CaMKII inhibitor protein beta and 65% identity and 78% positives with rat CaMKII inhibitor alpha. Human CaM-KIIN mRNA expression was detectable in various tissues and cell lines by Northern blot and RT-PCR. To investigate its biological functions, full-length human CaM-KIIN was overexpressed in colon adenocarcinoma LoVo cells. When expressed in LoVo cells, it could inhibit cell proliferation, block cell growth, and decrease the viable cell number. These results characterize a potential cellular inhibitor protein of CaMKII that plays an important role in the regulation of cell growth.

Adenocarcinoma↗
PubMed2001

[Treatment of spontaneous metastatic lung cancer with tumor antigen-pulsed, interleukin-18 gene-modified dendritic cells].

OBJECTIVE: To investigate the effect of tumor antigen-pulsed, interleukin-18 (IL-18) gene-modified dendritic cells in treatment of spontaneous metastatic lung cancer. METHODS: 3LL Lewis lung cancer cells were injected into the footpads of C57BL/6 mice to establish a spontaneous metastatic lung cancer model. Ninety-six mice with lung cancer were divided into 8 groups, 12 in each. treated differently. One group was treated by subcutaneous vaccination for two times of tumor antigen peptide Mut1-pulsed, IL-18 gene-modified dendritic cells (DC-IL-18/Mut1) that were derived from normal bone marrow. The other groups were treated with other measures. After treatment, the lung weight, number of metastatic nodes on the lung surface, survival time, and NK and CTL activities were examined. RESULTS: Compared with the mice treated with Mut1-pulsed control LacZ gene-modified DC and those treated with untreated DC, the tumor-bearing mice treated with DC-IL-18/Mut1 had the lightest lung weight (215 mg +/- 20 mg Vs 398 mg +/- 23 mg and 987 mg +/- 45 mg, t = 14.7 and 38.4, P < 0.01), the least lung metastatic nodes (0 Vs 7.8 +/- 2.7 and 49, P < 0.01), the longest survival time (chi(2) = 6.78 and 10.49 respectively, P < 0.01), the strongest cytotoxic T cell activity (53.4 +/- 3.1 Vs 41.3 +/- 2.6 and 9.8 +/- 2.1, t = 13.4 and 15, 7 respectively, P < 0.01), and increased proportions of CD4 + Tcells, CD8 + Tcells, and NK cells. CONCLUSION: Tumor antigen-pulsed, IL-18 gene-modified dendritic cells have a significant therapeutic effect on spontaneous netastatic lung cancer through induction of anti-tumor immunological responses.

Animals↗
PubMed2001

A transcriptionally [correction of transcriptively] active complex of APP with Fe65 and histone acetyltransferase Tip60.

Amyloid-beta precursor protein (APP), a widely expressed cell-surface protein, is cleaved in the transmembrane region by gamma-secretase. gamma-Cleavage of APP produces the extracellular amyloid beta-peptide of Alzheimer's disease and releases an intracellular tail fragment of unknown physiological function. We now demonstrate that the cytoplasmic tail of APP forms a multimeric complex with the nuclear adaptor protein Fe65 and the histone acetyltransferase Tip60. This complex potently stimulates transcription via heterologous Gal4- or LexA-DNA binding domains, suggesting that release of the cytoplasmic tail of APP by gamma-cleavage may function in gene expression.

Acetyltransferases↗
PubMed2001

Effect of contact time and inhibitor concentration on the affinity mediated adsorption of cells to surfaces.

Cell detachment by shear stress under conditions of laminar flow was used to investigate the effect of incubation time and soluble binding competitors on affinity mediated cell/surface interactions. Fractional attachment between yeast and a Concanavalin A (Con A) coated surface was studied as a function of adhesion time prior to exposure to shear in a parallel plate flow chamber. Two, four and sixteen hours adhesion times gave rise to significantly different fractional attachment profiles, with four hours giving greater cell retention.The effect of dextran as a competitive displacer of pre-attached cells was also examined using a number of exposure regimes. While the presence of dextran in the displacement buffer led to higher fractional displacement of pre-attached cells, this effect was magnified if an equilibration period between dextran solution and pre-attached cells was allowed before detachment was attempted. The decline in fractional attachment increased with incubation time up to 30 min, with longer periods resulting in a smaller effect. Pre-incubation of the Con A surface with dextran prior to the introduction of cells led to a 60% reduction in attachment.Attempts to determine critical shear values were complicated by the presence of a tightly bound cell fraction of approximately 15% that was not removed at the highest shear values used.

Journal Article↗
PubMed2001

Protein kinase C epsilon-Src modules direct signal transduction in nitric oxide-induced cardioprotection: complex formation as a means for cardioprotective signaling.

An essential role for protein kinase C epsilon (PKCepsilon) has been shown in multiple forms of cardioprotection; however, there is a distinct paucity of information concerning the signaling architecture that is responsible for the manifestation of a protective phenotype. We and others have recently shown that signal transduction may proceed via the formation of signaling complexes (Circ Res. 2001;88:59-62). In order to understand if the assembly of multiprotein complexes is the manner by which signaling is conducted in cardioprotection, we designed a series of experiments to characterize the associations of Src tyrosine kinase with PKCepsilon in a conscious rabbit model of nitric oxide (NO)-induced late preconditioning. Our data demonstrate that PKCepsilon and Src can form functional signaling modules in vitro: PKCepsilon interacts with Src; the association with PKCepsilon activates Src; and adult cardiac cells receiving recombinant adenoviruses encoding PKCepsilon exhibit increased Src activity. Furthermore, our results show that NO-induced late preconditioning involved PKCepsilon-Src module formation and enhanced the enzymatic activity of PKCepsilon-associated Src. Inhibition of PKC blocked cardioprotection, module formation, and PKCepsilon-associated Src activity, providing direct evidence for a functional role of the PKCepsilon-Src module in the orchestration of NO-induced cardioprotection in conscious rabbits.

Animals↗
PubMed2001

Cloning and characterization of Siglec-10, a novel sialic acid binding member of the Ig superfamily, from human dendritic cells.

The Siglecs (sialic acid-binding Ig-like lectins) are a subfamily of I-type lectins, which specifically recognize sialic acids. Nine members of the family have been identified thus far. We have obtained a novel cDNA clone from a human dendritic cell cDNA library encoding a protein with sequence and structural features of the Siglec family, hence designated as Siglec-10. The full-length Siglec-10 cDNA encodes a type 1 transmembrane protein containing four extracellular immunoglobulin-like domains, a transmembrane region, and a cytoplasmic tail with two classical immunoreceptor tyrosine-based inhibitory motifs. The N-terminal V-set Ig domain has most of the amino acid residues typical of the Siglecs. Siglec-10 shows the closest homology to Siglec-5 and Siglec-3/CD33. Various cells and cell lines including monocytes and dendritic cells express Siglec-10. High levels of mRNA expression were seen in peripheral blood leukocytes, spleen, and liver. When expressed on COS-7 cells, Siglec-10 was able to bind human red blood cells and soluble sialoglycoconjugates in a sialic acid-dependent manner. The identification of Siglec-10 as a new Siglec family member and its expression profile, together with its sialic acid-dependent binding capacity, suggest that it may be involved in cell-cell recognition by interacting with sialylated ligands expressed on specific cell populations.

Amino Acid Motifs↗
PubMed2001

Requirement of CHROMOMETHYLASE3 for maintenance of CpXpG methylation.

Epigenetic silenced alleles of the Arabidopsis SUPERMAN locus (the clark kent alleles) are associated with dense hypermethylation at noncanonical cytosines (CpXpG and asymmetric sites, where X = A, T, C, or G). A genetic screen for suppressors of a hypermethylated clark kent mutant identified nine loss-of-function alleles of CHROMOMETHYLASE3 (CMT3), a novel cytosine methyltransferase homolog. These cmt3 mutants display a wild-type morphology but exhibit decreased CpXpG methylation of the SUP gene and of other sequences throughout the genome. They also show reactivated expression of endogenous retrotransposon sequences. These results show that a non-CpG DNA methyltransferase is responsible for maintaining epigenetic gene silencing.

Alleles↗
PubMed2001

Enhancement of antigen-presenting ability of B lymphoma cells by immunostimulatory CpG-oligonucleotides and anti-CD40 antibody.

Immunostimulatory oligodeoxynucleotides containing the CpG motifs (CpG-ODN) can activate antigen-presenting cells including dendritic cells, macrophages, B cells, and enhance production of Thl cytokines. So, CpG-ODN has been regarded as a promising immune adjuvant. Using the A20 B lymphoma cell model, we investigated the effect of CpG-ODN on the immunogenicity of B lymphoma cells and whether CpG-ODN could enhance the antigen-presenting ability of B lymphoma cells. After incubation with CpG-ODN, proliferation of A20 cells remained unchanged. But CpG-ODN stimulation up-regulated the expression of MHC-I, MHC-II, CD40, ICAM-1 molecules in A20 cells, enhanced the antigen uptake ability of A20 cells, and promoted A20 cell production of IgM and IgG. More importantly, A20 cells activated by CpG-ODN could stimulate allogeneic T cells in MLR and antigen-primed T cells to proliferate more efficiently, suggesting the antigen-presenting ability of A20 B lymphoma cells could be enhanced by CpG-ODN stimulation and CpG-ODN-activated B lymphoma cells might be used as a potent cellular vaccine. Although anti-CD40 mAb was as effective as CpG-ODN at activating A20 cells and A20 cells expressed more CD40 molecules after CpG-ODN stimulation, a combination of CpG-ODN and anti-CD40 mAb had no synergistic effect on A20 cell activation. Our data expanded the potential application of CpG-ODN as an immunotherapeutic agent in cancer treatment.

Animals↗
PubMed2001

The involvement of TNF-alpha-related apoptosis-inducing ligand in the enhanced cytotoxicity of IFN-beta-stimulated human dendritic cells to tumor cells.

TNF-alpha-related apoptosis-inducing ligand (TRAIL) is characterized by its preferential induction of apoptosis of tumor cells but not normal cells. Dendritic cells (DCs), besides their role as APCs, now have been demonstrated to exert cytotoxicity or cytostasis on some tumor cells. Here, we report that both human CD34(+) stem cell-derived DCs (CD34DCs) and human CD14(+) monocyte-derived DCs (MoDCs) express TRAIL and exhibit cytotoxicity to some types of tumor cells partially through TRAIL. Moderate expression of TRAIL appeared on CD34DCs from the 8th day of culture and was also seen on freshly isolated monocytes. The level of TRAIL expression remained constant until DC maturation. TRAIL expression on immature CD34DCs or MoDCs was greatly up-regulated after IFN-beta stimulation. Moreover, IFN-beta could strikingly enhance the ability of CD34DCs or MoDCs to kill TRAIL-sensitive tumor cells, but LPS did not have such an effect. The up-regulation of TRAIL on IFN-beta-stimulated DCs partially contributed to the increased cytotoxicity of DCS: Pretreatment of TRAIL-sensitive tumor cells with caspase-3 inhibitor could significantly increase their resistance to the cytotoxicity of IFN-beta-stimulated DCS: In contrast, NF-kappaB inhibitor could significantly increase the sensitivity of tumor cells to the killing by nonstimulated or LPS-stimulated DCS: Our studies demonstrate that IFN-beta-stimulated DCs are functionally cytotoxic. Thus, an innate mechanism of DC-mediated antitumor immunity might exist in vivo in which DCs act as effectors to directly kill tumor cells partially via TRAIL. Subsequently, DCs act as APCs involved in the uptake, processing, and presentation of apoptotic tumor Ags to cross-prime CD8(+) CTL cells.

Adjuvants, Immunologic↗