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1.
Zhonghua Gan Zang Bing Za Zhi ; 26(12): 922-926, 2018 Dec 20.
Article in Chinese | MEDLINE | ID: mdl-30669785

ABSTRACT

Objective: To establish a new model of hepatic steatosis cells by optimizing the original ethanol or high fat, the present study proposed an in vitro hepatocyte steatosis model for the study of fatty liver. Methods: Oil red O staining was used to observe the effects of fetal bovine serum, oleic acid and ethanol on lipid accumulation in human liver cell line L02 in a concentration- and time-dependent manner. RT-PCR was used to detect the mRNA expression levels of PPAR-γ and AP-2, and the suitable conditions for the establishment of hepatocyte steatosis model were screened out. A t-test was used for comparison between the two groups, and one-way Analysis of Variance (ANOVA) was used in more than three groups. Results: Oil red O staining showed the number of reddish-orange lipid droplets in L02 cells gradually increased with the increase of fetal bovine serum, oleic acid and ethanol in a concentration - and time-dependent manner. Compared with 0.00% oleic acid and 2% ethanol, the count value of red particle was 100.00% ± 17.63% at the beginning and after 24 h, 0.003% oleic acid and 2% ethanol jointly acted in L02 cells. After incubation for 48 hours with 2% ethanol and serum-free DMEM medium, the accumulation of lipid droplets was the highest with a count value of 802.38%+71.06%(t = 42.36, P < 0.001). RT-PCR analysis showed the lipid accumulation induced by this method was positively correlated with the mRNA expression of PPAR-γ and AP-2. Conclusion: L02 cells were successfully exposed to high fat and ethanol, and the hepatocyte steatosis model was established and optimized, suggesting that the occurrence of hepatic cell steatosis was related to the up-regulation of PPAR-γ and AP-2.


Subject(s)
Fatty Liver , Hepatocytes , Cell Line , Humans , Lipid Metabolism , Oleic Acid
2.
Micron ; 91: 41-48, 2016 Dec.
Article in English | MEDLINE | ID: mdl-27721207

ABSTRACT

This paper studies the absolute thickness measurement of pyrolytic graphite spheroids (GSs) by using STEM-EELS mode with log-ratio method and Kramers-Kroning (K-K) method, taking the measured thickness from TEM image as reference that is the diameter of GSs ranging from 60 to 250nm. The effect of collection semi-angle (ß) on thickness measurement has been investigated. It is found that in general the thickness obtained by K-K analysis with surface effect corrected shows the best accuracy, followed by K-K sum rule and then log-ratio method for the three different collection semi-angles of 12.4, 17.3 and 21.1mrad applied. Of these angles, the smallest one gives an overestimated result and the largest one gives an underestimated result, whereas between the two, the angle of 17.3mrad that is about 2x convergence semi-angle (9.0mrad) is identified as more appropriate for K-K analysis. The surface-scattering correction, inelastic mean free path of GS and effect of refractive index n on thickness measurement for different ß angles are also investigated. Moreover, the optical property deduced from the data collected at the center of graphite spheroid, which is related to its microstructure, is characterized by K-K analysis.

3.
J Biol Regul Homeost Agents ; 29(1): 201-6, 2015.
Article in English | MEDLINE | ID: mdl-25864759

ABSTRACT

Joint detection of anti-dsDNA antibodies, anti-U1RNP, anti-SM antibodies, anti-SSA antibodies, anti-ribosomal P protein antibodies, anti-nucleosome antivodies (Anua), anti-histone antibodies (AHA) and antinuclear antibodies brings to the early diagnosis of systemic lupus erythematosus (SLE) and speculation of renal lesion degree of lupus nephritis patients in order to choose a specific therapeutic schedule. This paper analyzed the abnormal immunology features and connections of each pathological pattern of LN renal biopsy and probed into the essence in order to provide basis for diagnosis, treatment, pathological pattern speculation and forward assessment of LN. We chose 97 cases, treated them with renal biopsy and pathological pattern classification, analyzed pathological pattern distribution, different pathological patterns and the correlation of immunity index with anti-dsDNA antibodies, anti-U1RNP, anti-Sm antibodies, anti-SSA antibodies, anti-ribosomal P protein antibodies, Anua, AHA and ANA of the first renal biopsy were taken as the experiment index. The results showed that the morbidity of the male was distinctly lower than the female and the age of onset was much lower (P < 0.05); pattern I, pattern II, pattern III, pattern IV, pattern V, and pattern VI accounted for 1.0%, 3.1%, 12.4%, 47.4%,16.5%, 15.5%, 4.1%, 0%,respectively; among all the LN patients, there were respectively 59, 43, 28, 52, 51, 48, 36 and 93 cases in which anti-dsDNA antibody, anti-U1RNP antibody, anti-Sm antibody, anti-SSA antibody, anti-ribosomal P protein antibodies, Anua, AHA and ANA had increased and the positive rate was 60.8%, 44.3%, 28.9%, 53.6%, 52.6%, 49.5%, 37.1% and 95.9%, respectively. In conclusion, pattern IV is the most common of all pathological patterns of LN. Among the immunity index, anti- U1RNP antibodies and anti-SSA antibodies are positively correlated with anti-dsDNA antibodies; Anua is positively correlated with anti-dsDNA antibodies and AHA; anti-dsDNA antibodies, anti U1RNP antibodies, anti-Sm antibodies, anti SSA antibodies, AHA, anti-ribosomal P protein antibodies and ANA have no obvious correlation with LN renal lesions degree; Anua level of serum is positively correlated with LN renal lesions degree.


Subject(s)
Lupus Nephritis/immunology , Lupus Nephritis/pathology , Adolescent , Adult , Aged , Antibodies, Antinuclear/blood , Female , Histones/immunology , Humans , Lupus Erythematosus, Systemic/immunology , Lupus Erythematosus, Systemic/pathology , Male , Middle Aged , Nucleosomes/immunology , Ribonucleoprotein, U1 Small Nuclear/immunology , Ribosomal Proteins/immunology , Young Adult , snRNP Core Proteins/immunology
4.
Eur J Surg Oncol ; 40(3): 255-9, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24429027

ABSTRACT

OBJECTIVES: This research aims to specify the prognostic value of P-cadherin on recurrence and progression in non-muscle-invasive bladder cancers (NMIBC). METHODS: A total of 110 NMIBC cases were collected and P-cadherin protein was assessed by immunohistochemical test in these samples. Correlations between P-cadherin expression and clinicopathologic features were analyzed. For recurrence-free and progression-free survival, Kaplan-Meier log-rank test was used. Then Cox univariate and multivariate analyses were further performed. RESULTS: P-cadherin high expression correlated with tumor progression (P = 0.031). Kaplan-Meier results showed that patients with high P-cadherin expression had worse progression-free survival (P = 0.034) but not recurrence-free survival (P = 0.133) than low-expression patients. Cox regression results showed P-cadherin expression was an independent predictor for progression (P = 0.042) but not recurrence (P = 0.139) in NMIBC. CONCLUSIONS: Our results demonstrated that P-cadherin expression correlated with tumor progression and could be taken as an independent predictor for progression in NMIBC.


Subject(s)
Biomarkers, Tumor/analysis , Cadherins/analysis , Carcinoma, Transitional Cell/chemistry , Carcinoma, Transitional Cell/genetics , Neoplasm Recurrence, Local/genetics , Urinary Bladder Neoplasms/chemistry , Urinary Bladder Neoplasms/genetics , Adult , Aged , Analysis of Variance , Biomarkers, Tumor/genetics , Biopsy, Needle , Cadherins/genetics , Carcinoma, Transitional Cell/mortality , Carcinoma, Transitional Cell/pathology , Cohort Studies , Disease Progression , Disease-Free Survival , Female , Gene Expression Profiling/methods , Genetic Markers/genetics , Humans , Immunohistochemistry , Kaplan-Meier Estimate , Male , Middle Aged , Multivariate Analysis , Neoplasm Invasiveness/genetics , Neoplasm Invasiveness/pathology , Neoplasm Recurrence, Local/mortality , Neoplasm Staging , Predictive Value of Tests , Prognosis , Proportional Hazards Models , Retrospective Studies , Survival Analysis , Urinary Bladder Neoplasms/mortality , Urinary Bladder Neoplasms/pathology
5.
Oncogene ; 29(10): 1451-62, 2010 Mar 11.
Article in English | MEDLINE | ID: mdl-20101232

ABSTRACT

The ERalpha signaling pathway is one of the most important and most studied pathways in human breast cancer, yet numerous questions still exist such as how hormonally responsive cancers progress to a more aggressive and hormonally independent phenotype. We have noted that human breast cancers exhibit a strong direct correlation between ERalpha and E-cadherin expression by immunohistochemistry, suggesting that ERalpha signaling might regulate E-cadherin and implying that this regulation might influence epithelial-mesenchymal transition (EMT) and tumor progression. To investigate this hypothesis and the mechanisms behind it, we studied the effects of ERalpha signaling in ERalpha-transfected ERalpha-negative breast carcinoma cell lines, the MDA-MB-468 and the MDA-MB-231 and the effects of ERalpha knockdown in naturally expressing ERalpha-positive lines, MCF-7 and T47D. When ERalpha was overexpressed in the ERalpha-negative lines, 17beta-estradiol (E2) decreased slug and increased E-cadherin. Clones maximally exhibiting these changes grew more in clumps and became less invasive in Matrigel. When ERalpha was knocked down in the ERalpha-positive lines, slug increased, E-cadherin decreased, cells became spindly and exhibited increased Matrigel invasion. ERalpha signaling decreased slug expression by two different mechanisms: directly, by repression of slug transcription by the formation of a corepressor complex of ligand-activated ERalpha, HDAC inhibitor (HDAC1), and nuclear receptor corepressor (N-CoR) that bound the slug promoter in three half-site estrogen response elements (EREs); indirectly by phosphorylation and inactivation of GSK-3beta through phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt). The GSK-3beta inactivation, in turn, repressed slug expression and increased E-cadherin. In human breast cancer cases, there was a strong inverse correlation between slug and ERalpha and E-cadherin immunoreactivity. Our findings indicate that ERalpha signaling through slug regulates E-cadherin and EMT.


Subject(s)
Breast Neoplasms/metabolism , Cadherins/metabolism , Estrogen Receptor alpha/metabolism , Transcription Factors/metabolism , Blotting, Western , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cadherins/genetics , Cell Line, Tumor , Epithelium/metabolism , Epithelium/pathology , Estradiol/pharmacology , Estrogen Receptor alpha/genetics , Female , Gene Expression Regulation, Neoplastic/drug effects , Glycogen Synthase Kinase 3/metabolism , Glycogen Synthase Kinase 3 beta , Histone Deacetylase Inhibitors/pharmacology , Humans , Immunohistochemistry , Mesoderm/metabolism , Mesoderm/pathology , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , RNA Interference , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/drug effects , Snail Family Transcription Factors , Transcription Factors/genetics
6.
Cell Death Differ ; 17(3): 522-33, 2010 Mar.
Article in English | MEDLINE | ID: mdl-19816511

ABSTRACT

The cellular FLICE inhibitory protein (c-FLIP) is an endogenous inhibitor of the caspase-8 proapoptotic signaling pathway downstream of death receptors. Recent evidence indicates that the long form of c-FLIP (c-FLIP(L)) is required for proliferation and effector T-cell development. However, the role of c-FLIP(L) in triggering autoimmunity has not been carefully analyzed. We now report that c-FLIP(L) transgenic (Tg) mice develop splenomegaly, lymphadenopathy, multiorgan infiltration, high titers of auto-antibodies, and proliferative glomerulonephritis with immune complex deposition in a strain-dependent manner. The development of autoimmunity requires CD4(+) T cells and may result from impaired thymic selection. At the molecular level, c-FLIP(L) overexpression inhibits the zeta chain-associated protein tyrosine kinase of 70 kDa (ZAP-70) activation, thus impairing the signaling pathway derived from ZAP-70 required for thymic selection. Therefore, we have identified c-FLIP(L) as a susceptibility factor under the influence of epistatic modifiers for the development of autoimmunity.


Subject(s)
CASP8 and FADD-Like Apoptosis Regulating Protein/metabolism , Lupus Erythematosus, Systemic/immunology , Mice, Inbred BALB C/immunology , Mice, Inbred C57BL/immunology , T-Lymphocyte Subsets/immunology , T-Lymphocytes/immunology , Thymus Gland/immunology , Animals , Apoptosis/physiology , Autoantibodies/metabolism , B-Lymphocytes/immunology , CASP8 and FADD-Like Apoptosis Regulating Protein/genetics , Cell Proliferation , Cytokines/metabolism , Dendritic Cells/immunology , Humans , Lupus Erythematosus, Systemic/pathology , Lupus Erythematosus, Systemic/physiopathology , Lymphocyte Activation , Mice , Mice, Transgenic , Phenotype , T-Lymphocyte Subsets/cytology , T-Lymphocytes/cytology , Thymus Gland/cytology , Transgenes , ZAP-70 Protein-Tyrosine Kinase/metabolism
7.
J Microsc ; 230(Pt 1): 143-59, 2008 Apr.
Article in English | MEDLINE | ID: mdl-18387049

ABSTRACT

In this paper, a probabilistic technique for compensation of intensity loss in confocal microscopy images is presented. For single-colour-labelled specimen, confocal microscopy images are modelled as a mixture of two Gaussian probability distribution functions, one representing the background and another corresponding to the foreground. Images are segmented into foreground and background by applying Expectation Maximization algorithm to the mixture. Final intensity compensation is carried out by scaling and shifting the original intensities with the help of parameters estimated for the foreground. Since foreground is separated to calculate the compensation parameters, the method is effective even when image structure changes from frame to frame. As intensity decay function is not used, complexity associated with estimation of the intensity decay function parameters is eliminated. In addition, images can be compensated out of order, as only information from the reference image is required for the compensation of any image. These properties make our method an ideal tool for intensity compensation of confocal microscopy images that suffer intensity loss due to absorption/scattering of light as well as photobleaching and the image can change structure from optical/temporal section-to-section due to changes in the depth of specimen or due to a live specimen. The proposed method was tested with a number of confocal microscopy image stacks and results are presented to demonstrate the effectiveness of the method.

8.
J Acoust Soc Am ; 117(5): 2742-51, 2005 May.
Article in English | MEDLINE | ID: mdl-15957745

ABSTRACT

The energy transmission in a mechanically linked double-wall structure into an acoustic enclosure is studied in this paper. Based on a fully coupled vibro-acoustic formulation, focus is put on investigating the effect of the air gap and mechanical links between the two panels on the energy transmission and noise insulation properties of such structures. An approximate formula reflecting the gap effect on the lower-order coupled frequencies of the system is proposed. A criterion, based on the ratio between the aerostatic stiffness of the gap cavity and the stiffness of the link, is proposed to predict the dominant transmitting path, with a view to provide guidelines for the design of appropriate control strategies. Numerical results reveal the existence of three distinct zones, within which energy transmission takes place following different mechanisms and transmitting paths. Corresponding effects on noise insulation properties of the double-wall structure are also investigated.


Subject(s)
Acoustics , Energy Transfer , Mechanics , Models, Theoretical , Vibration
9.
J Exp Med ; 194(9): 1339-48, 2001 Nov 05.
Article in English | MEDLINE | ID: mdl-11696598

ABSTRACT

B7H/B7RP (hereby called B7H) is a new member of the B7 family of costimulatory molecules and interacts with inducible costimulatory molecule (ICOS). Its function for CD8 T cells has not been reported. We report here that expression of B7H on the tumor cells reduced tumorigenicity and induced immunity to subsequent challenge with parental tumor cells. The immune protection correlates with an enhanced cytotoxic T lymphocyte (CTL) response against P1A, the major tumor antigen expressed in the J558 tumor. To understand the mechanism of immune protection, we adoptively transferred transgenic T cells specific for tumor antigen P1A into mice that bore P1A-expressing tumors. We found that while the transgenic T cells divided faster in mice bearing the B7H(+) tumors, optimal B7H-induced clonal expansion of P1CTL required costimulation by B7-1 and B7-2 on the endogenous host antigen-presenting cells (APCs). Interestingly, when B7H(+) and B7H(-) tumors were coinjected, P1CTL selectively eliminated the B7H(+) tumor cells. Moreover, B7H expressed on the tumor cells made them highly susceptible to destruction by CTL in vivo, even if the CTL was administrated into mice with large tumor burdens. Tumors that recurred in the P1CTL-treated mice lost transfected B7H and/or H-2L(d), the class I molecule that presents the P1A peptide. Taken together, our results reveal that B7H costimulates clonal expansion of, and cognate destruction by CD8(+) T lymphocytes in vivo.


Subject(s)
Proteins/immunology , T-Lymphocytes, Cytotoxic/immunology , Animals , Antigen-Presenting Cells/immunology , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/immunology , Clone Cells , Cytotoxicity, Immunologic/immunology , Female , Gene Expression , Inducible T-Cell Co-Stimulator Ligand , Ligands , Male , Mice , Mice, Inbred BALB C , Mice, Transgenic , Proteins/genetics , T-Lymphocytes, Cytotoxic/cytology , Tumor Cells, Cultured
10.
Cancer Res ; 61(18): 6860-7, 2001 Sep 15.
Article in English | MEDLINE | ID: mdl-11559562

ABSTRACT

Because of the low frequency of antigen-specific T cells, early events in the activation of tumor-specific T cells in vivo have not been well characterized. There is still no direct documentation on where the clonal expansion begins and how tumor antigens are presented to the host CD8 T cells to initiate it. Here we used transgenic T cells specific for a natural tumor antigen P1A to evaluate the kinetics, location, and modes of antigen presentation for initiating CTL response in vivo. Our results demonstrate that the initial activation of P1A-specific T cells takes place in the lymphoid organs. The activated T cells then migrate into tumors, where they undergo accelerated division and acquire distinct activation markers. The site of initiation cannot be altered by either local expression of costimulatory molecules or by intratumor injection of naïve T cells. Moreover, using genetic models that allow only one mode of antigen presentation, we show here that both cross-presentation of P1A by the host antigen-presenting cells, and direct antigen presentation and costimulation by the tumor cells are sufficient to initiate rapid T cell-clonal expansion in the lymphoid organ. These results provide direct evidence for two fundamental assumptions on the mechanisms of T-cell activation in vivo.


Subject(s)
Antigen Presentation/immunology , Antigens, Neoplasm/immunology , CD8-Positive T-Lymphocytes/immunology , Animals , B7-1 Antigen/immunology , Cross Reactions/immunology , Epitopes, T-Lymphocyte/immunology , Immunophenotyping , Immunotherapy, Adoptive , L-Selectin/immunology , Lymph Nodes/immunology , Lymphocyte Activation/immunology , Lymphocytes, Tumor-Infiltrating/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, SCID , Mice, Transgenic , Plasmacytoma/immunology , Spleen/immunology , Spleen/pathology
11.
Cell Immunol ; 202(1): 41-53, 2000 May 25.
Article in English | MEDLINE | ID: mdl-10873305

ABSTRACT

The identification of factors that regulate the proliferation and differentiation of double-positive (DP) into CD4(+) and CD8(+) single-positive (SP) thymocytes has proven difficult due to the inability of DP thymocytes to proliferate, expand, and differentiate into SP thymocytes in available cell culture media. Here we report on the ability of DP thymocytes to differentiate in a novel conditioned medium, termed XLCM, derived from the supernatant of mitogen activated human cord blood mononuclear cells. During a 5-day culture in XLCM in the absence of thymic stromal cells, DP thymocytes from normal mice and MHC double knockout mice (lack SP thymocytes) proliferate, expand, and differentiate into several (alphabetaTCR(+), NK1.1(+)alphabetaTCR(+), and gammadeltaTCR(+)) subsets of CD4(+) and predominantly CD8(+) SP thymocytes. These studies suggest that the use of XLCM may aid in the characterization of factors that regulate the differentiation of DP thymocytes into CD8(+) SP thymocytes.


Subject(s)
CD8-Positive T-Lymphocytes/cytology , Cell Culture Techniques/methods , Culture Media, Conditioned , T-Lymphocyte Subsets/cytology , Thymus Gland/cytology , Animals , CD8-Positive T-Lymphocytes/immunology , Cell Differentiation/drug effects , Cell Lineage , Chemokines/analysis , Culture Media, Conditioned/chemistry , Cytokines/analysis , Cytokines/pharmacology , Female , Fetal Blood/chemistry , Fetal Blood/immunology , Humans , Interleukin-4/pharmacology , Killer Cells, Natural , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Receptors, Antigen, T-Cell, gamma-delta , T-Lymphocyte Subsets/immunology , Thymus Gland/immunology
12.
Chirality ; 12(5-6): 383-8, 2000 Jun.
Article in English | MEDLINE | ID: mdl-10824157

ABSTRACT

The condensation of o-(diphenylphosphino)benzaldehyde and various chiral diamine gives a series of diimino-diphosphine tetradentate ligands, which are reduced with excess NaBH4 in refluxing ethanol to afford the corresponding diaminodiphosphine ligands in good yield. The reactivity of these ligands toward trans-RuCl2(DMSO)4 and [Rh(COD)Cl]2 had been investigated and a number of chiral Ru(II) and Rh(I) complexes with the PNNP-type ligands were synthesized and characterized by microanalysis and IR, NMR spectroscopic methods. The chiral Ru(II) and Rh(I) complexes have proved to be excellent catalyst precursors for the asymmetric transfer hydrogenation of aromatic ketones, leading to optically active alcohols in up to 97% ee.


Subject(s)
Ketones/metabolism , Ruthenium Compounds/chemical synthesis , Ruthenium/chemistry , Chemistry/methods , Hydrogen/chemistry , Ligands , Magnetic Resonance Spectroscopy , Models, Chemical , Ruthenium Compounds/chemistry , Spectrophotometry, Infrared , Stereoisomerism
13.
J Exp Med ; 191(6): 1017-30, 2000 Mar 20.
Article in English | MEDLINE | ID: mdl-10727463

ABSTRACT

Activation-induced cell death (AICD) is a mechanism of peripheral T cell tolerance that depends upon an interaction between Fas and Fas ligand (FasL). Although c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) may be involved in apoptosis in various cell types, the mode of regulation of FasL expression during AICD in T cells by these two MAPKs is incompletely understood. To investigate the regulatory roles of these two MAPKs, we analyzed the kinetics of TCR-induced p38 MAPK and JNK activity and their regulation of FasL expression and AICD. We report that both JNK and p38 MAPK regulate AICD in T cells. Our data suggest a novel model of T cell AICD in which p38 MAPK acts early to initiate FasL expression and the Fas-mediated activation of caspases. Subsequently, caspases stimulate JNK to further upregulate FasL expression. Thus, p38 MAPK and downstream JNK converge to regulate FasL expression at different times after T cell receptor stimulation to elicit maximum AICD.


Subject(s)
Apoptosis/immunology , Calcium-Calmodulin-Dependent Protein Kinases/physiology , Lymphocyte Activation/immunology , Membrane Glycoproteins/biosynthesis , Mitogen-Activated Protein Kinases/physiology , T-Lymphocytes/enzymology , T-Lymphocytes/immunology , fas Receptor/metabolism , Animals , Apoptosis/drug effects , Apoptosis/genetics , Caspase Inhibitors , Caspases/metabolism , Cell Line , Enzyme Activation/drug effects , Enzyme Inhibitors/pharmacology , Fas Ligand Protein , Hybridomas , Imidazoles/pharmacology , JNK Mitogen-Activated Protein Kinases , Ligands , Lymphocyte Activation/drug effects , Lymphocyte Activation/genetics , Membrane Glycoproteins/deficiency , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Mice , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Mutant Strains , Mitogen-Activated Protein Kinases/metabolism , Pyridines/pharmacology , T-Lymphocytes/cytology , p38 Mitogen-Activated Protein Kinases
14.
Immunology ; 98(2): 159-70, 1999 Oct.
Article in English | MEDLINE | ID: mdl-10540214

ABSTRACT

The induction of an immune response or tolerance is mediated by corresponding subsets of dendritic cells (DC). However, the property of tolerogenic DC is not clear. Recently, we have characterized a population of CD11c+ splenic DC derived from long-term mixed leucocyte culture (LT-MLC), which are able to proliferate upon stimulation and have a strong primary mixed leucocyte reaction (MLR)-stimulating activity in conventional MLR. In this study, we show that, in contrast to the irradiated ones, non-irradiated LT-MLC-derived DC induce polyclonal antigen-specific T-cell hyporesponsiveness when cocultured with allogeneic splenocytes for 3-11 days. The degree of the hyporesponsiveness increased with the length of coculture. Although these DC expressed major histocompatibility complex class II and B7 costimulatory molecules, which are down-regulated during coculture, they expressed very low or undetectable CD40 before and after coculture, respectively. The CD40-deficient DC spontaneously produce interleukin-10 (IL-10), but not IL-12. The skewed balance between IL-10 and IL-12 is associated with their capability to induce T-cell hyporesponsiveness, because a neutralizing antibody to IL-10, exogenous recombinant IL-12 or lipopolysaccharide (LPS) significantly blocked the hyporesponsiveness. Accordingly, infusion of a small number of non-irradiated LT-MLC-derived DC (5x105) significantly prolonged the survival of a vascularized heterotopic murine heart transplant, whereas irradiated DC accelerated graft rejection. These data suggest that CD40-deficient DC producing IL-10, but not IL-12 can induce T-cell hyporesponsiveness in vitro and in vivo.


Subject(s)
Adoptive Transfer , CD40 Antigens/immunology , Dendritic Cells/immunology , Immune Tolerance , Interleukin-10/immunology , T-Lymphocytes/immunology , Animals , Cells, Cultured , Flow Cytometry , Graft Rejection/immunology , Interleukin-12/immunology , Lipopolysaccharides/pharmacology , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, Inbred C57BL , Transplantation, Homologous
15.
J Immunol ; 162(7): 3819-29, 1999 Apr 01.
Article in English | MEDLINE | ID: mdl-10201899

ABSTRACT

Optimal T cell activation requires two signals, one generated by TCR and another by the CD28 costimulatory receptor. In this study, we investigated the regulation of costimulation-induced mitogen-activated protein kinase (MAPK) activation in primary mouse T cells. In contrast to that reported for human Jurkat T cells, we found that p38 MAPK, but not Jun NH2-terminal kinase (JNK), is weakly activated upon stimulation with either anti-CD3 or anti-CD28 in murine thymocytes and splenic T cells. However, p38 MAPK is activated strongly and synergistically by either CD3/CD28 coligation or PMA/Ca2+ ionophore stimulation, which mimics TCR-CD3/CD28-mediated signaling. Activation of p38 MAPK correlates closely with the stimulation of T cell proliferation. In contrast, PMA-induced JNK activation is inhibited by Ca2+ ionophore. T cell proliferation and production of IL-2, IL-4, and IFN-gamma induced by both CD3 and CD3/CD28 ligation and the nuclear expression of the c-Jun and ATF-2 proteins are each blocked by the p38 MAPK inhibitor SB203580. Our findings demonstrate that p38 MAPK 1) plays an important role in signal integration during costimulation of primary mouse T cells, 2) may be involved in the induction of c-Jun activation and augmentation of AP-1 transcriptional activity, and 3) regulates whether T cells enter a state of functional unresponsiveness.


Subject(s)
CD28 Antigens/immunology , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Lymphocyte Activation , Mitogen-Activated Protein Kinases , Receptors, Antigen, T-Cell/immunology , Signal Transduction/immunology , T-Lymphocytes/enzymology , T-Lymphocytes/immunology , Activating Transcription Factor 2 , Animals , CD3 Complex/immunology , CD3 Complex/metabolism , Calcium/physiology , Cell Nucleus/drug effects , Cell Nucleus/enzymology , Cell Nucleus/metabolism , Cells, Cultured , Cyclic AMP Response Element-Binding Protein/antagonists & inhibitors , Cyclic AMP Response Element-Binding Protein/biosynthesis , Cytokines/antagonists & inhibitors , Cytokines/metabolism , Drug Synergism , Enzyme Activation , Humans , Imidazoles/pharmacology , JNK Mitogen-Activated Protein Kinases , Jurkat Cells , Lymphocyte Activation/drug effects , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Proto-Oncogene Proteins c-jun/antagonists & inhibitors , Proto-Oncogene Proteins c-jun/biosynthesis , Pyridines/pharmacology , Signal Transduction/drug effects , T-Lymphocytes/drug effects , T-Lymphocytes/metabolism , Transcription Factors/antagonists & inhibitors , Transcription Factors/biosynthesis , p38 Mitogen-Activated Protein Kinases
16.
J Immunol ; 161(6): 2930-7, 1998 Sep 15.
Article in English | MEDLINE | ID: mdl-9743355

ABSTRACT

The TCR is a multisubunit complex composed of the clonotypic alpha/beta disulfide-linked heterodimer and noncovalently linked invariant CD3 gamma epsilon and CD3 delta epsilon and TCR zeta chains. Recent studies demonstrate that the surface expression of CD3 components can occur independently of the clonotypic TCR complexes in both thymocytes and splenic T cells. In this study, we report that free noncovalently associated TCR alpha beta heterodimers that exist independently of CD3 and TCR zeta chains are expressed on the cell surface of immature thymocytes and peripheral T cells, but not of T cell lines and T cell hybridomas. This suggests that the regulation of surface expression of TCR alpha beta heterodimers differs between primary T cells and T cell lines or T cell hybridomas. The isolation and biochemical characterization of surface clonotype-independent CD3 complexes and free membrane-associated TCR alpha beta complexes may provide a structural basis for the quantitative difference in amount of T cell proliferation stimulated by anti-CD3 epsilon and anti-TCR beta.


Subject(s)
CD3 Complex/metabolism , Membrane Proteins/metabolism , Receptors, Antigen, T-Cell, alpha-beta/metabolism , Receptors, Antigen, T-Cell/metabolism , T-Lymphocyte Subsets/metabolism , Animals , CD3 Complex/immunology , Calcium-Binding Proteins/metabolism , Calnexin , Cell Differentiation/immunology , Cell Membrane/immunology , Cell Membrane/metabolism , Humans , Hybridomas/immunology , Hybridomas/metabolism , Immune Sera/pharmacology , Jurkat Cells/immunology , Jurkat Cells/metabolism , Lymphocyte Activation/immunology , Macromolecular Substances , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Receptors, Antigen, T-Cell, alpha-beta/biosynthesis , Receptors, Antigen, T-Cell, alpha-beta/immunology , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/ultrastructure
17.
J Biol Chem ; 273(38): 24867-73, 1998 Sep 18.
Article in English | MEDLINE | ID: mdl-9733792

ABSTRACT

Aryl hydrocarbon receptor (AhR) nuclear translocator (Arnt) gene has been isolated and characterized from a mouse genomic DNA library. The gene is about 60 kilobases long and split into 22 exons. An unusual exon/intron junctional sequence was found in the 11th intron of the gene that begins with GC at its 5'-end. The exon/intron arrangement of mArnt gene differs greatly from those of the other members of the same basic-helix-loop-helix/PAS family. The gene is TATA-less and has several transcription start sites. The promoter region of the mArnt gene is GC-rich and contains a number of putative regulatory DNA sequences such as two GC-boxes, a cAMP-responsive element, E-box, AP-1 site, and CAAT-box. Deletion experiments revealed that all these DNA elements made substantial contributions to a high level of expression of the gene, except for the cAMP-responsive element. Of all, two GC-boxes displayed the most dominant enhancing effects. It was demonstrated that there exist specific factors binding to these DNA elements in the nuclear extracts of HeLa cells. Among them, Sp1 and Sp3, and CAAT-box binding factor-A were identified to bind the GC-boxes and CAAT-box, respectively. Expression of MyoD in HeLa cells stimulated the Arnt promoter activity by binding to the E-box.


Subject(s)
Gene Expression Regulation , Promoter Regions, Genetic , Transcription Factors/biosynthesis , Transcription Factors/genetics , Amino Acid Sequence , Animals , Aryl Hydrocarbon Receptor Nuclear Translocator , Base Composition , Base Sequence , Cell Nucleus/metabolism , DNA Primers , DNA-Binding Proteins/metabolism , Exons , Genomic Library , HeLa Cells , Helix-Loop-Helix Motifs , Humans , Introns , Mice , Molecular Sequence Data , MyoD Protein/biosynthesis , MyoD Protein/metabolism , Oligonucleotide Probes , Receptors, Aryl Hydrocarbon/biosynthesis , Receptors, Aryl Hydrocarbon/chemistry , Receptors, Aryl Hydrocarbon/genetics , Recombinant Fusion Proteins/biosynthesis , Regulatory Sequences, Nucleic Acid , Sequence Alignment , Sequence Homology, Amino Acid , Transcription Factors/chemistry
18.
Shanghai Kou Qiang Yi Xue ; 6(2): 68-70, 1997 Jun.
Article in Chinese | MEDLINE | ID: mdl-15159931

ABSTRACT

It is the first step that the data about relative teeth's shape are catched for CAD/CAM system in dental rehabilitation.That is how to measure the teeth's shape.The digtal speckle correlation method are introduced to measure the teeth's shape in the article.For one posterior tooth,we measured the five surfaces that the occlusal surface and the four axial surfaces.According to some special marks,we changed the five group's local data into one coordinate and reconstructed the three dimensional shape of the tooth.At last we drew the graph with the computer.In this paper the author also discussed some factors about the measure system's sensitive and precise and the extent scale.etc.

19.
Immunology ; 91(1): 135-44, 1997 May.
Article in English | MEDLINE | ID: mdl-9203977

ABSTRACT

The mechanisms contributing to the proliferation and differentiation of antigen-presenting cell (APC) precursors upon antigen stimulation or tissue injury are poorly understood. Herein, we report the induction of a population of dendritic cell-like cells (DLC) with potent antigen-presentation function from unfractionated spleen cells by means of repetitive allostimulation in long-term mixed leucocyte cultures (LT-MLC). Initially, only a few adherent DLC were observed. By 4-6 weeks, however, there were large numbers of DLC which survived persistently. Features of these DLC are closely related to dendritic cells (DC), including (1) dendritic, veiled or spiny-processed morphology; (2) expression of a wide array of leucocyte surface markers including DC-associated or restricted antigens: 33D1, NLDC-145, CD11c (N418), heat-stable antigen (HSA), CD44, B7-1 and B7-2; (3) ability to migrate to draining lymph nodes and white pulp area of spleen; (4) expression of high level of major histocompatability complex (MHC) class II molecules and (5) more potent mixed leucocyte reaction (MLR)-stimulating capacity than peritoneal macrophages and APC-enriched spleen cells. DLC-stimulated MLR was inhibited by monoclonal antibodies (mAbs) to B7-1, B7-2, intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), leucocyte-function associated antigen-1 (LFA-1) or very-late activation antigen-4 (VLA-4) by 30-55%. When maintained for more than 2 months, the DLC did not lose their MLR-stimulating activity, but many surface markers were down-regulated except for Mac-2 and VCAM-1, which remained stable or were up-regulated, respectively. In short-term culture, the addition of granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin (IL)-2 enhanced proliferation of DLC, while tumour necrosis factor-alpha (TNF-alpha) and IL-4 did not. IL-4 suppressed not only 'spontaneous', but also GM-CSF-enhanced proliferation, suggesting that cytokines play a differential role in DLC proliferation. These results confirm that professional APC can proliferate in response to repetitive antigen stimulation, and their proliferation is differentially regulated by cytokines. A comparison study of DLC with typical DC is being carried out in our laboratory.


Subject(s)
Antigen Presentation/immunology , Dendritic Cells/immunology , Animals , Cell Division/immunology , Cell Movement/immunology , Cell Separation , Cytokines/immunology , Dendritic Cells/cytology , Dendritic Cells/ultrastructure , Flow Cytometry , Immunophenotyping , Isoantigens/immunology , Lymphocyte Culture Test, Mixed , Lymphoid Tissue/immunology , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, Inbred C57BL
20.
Immunology ; 90(3): 448-54, 1997 Mar.
Article in English | MEDLINE | ID: mdl-9155654

ABSTRACT

Polymorphonuclear leucocyte (PMNL) accumulation in extravascular tissues and inflammatory exudates is dependent on their migration through blood vessel endothelium and then through connective tissue. Previously we utilized a barrier of human synovial and dermal fibroblasts (HSF or HDF) grown on microporous filters, as a model of PMNL migration through connective tissue. Those studies showed that beta 2 (CD18) and the beta 1 integrins, very late activation antigen-5 (VLA-5) and VLA-6, in part mediate this PMNL migration. Here we report that VLA-4, which can also be expressed at low levels on activated PMNL, is also involved in PMNL migration induced by C5a through fibroblast (HSF and HDF) barriers, because monoclonal antibody (mAb) to VLA-4 significantly inhibited (by 20-30%) PMNL migration. Blocking the function of CD18, VLA-5 or VLA-6 was not required for detection of the VLA-4-mediated migration. Combination treatment with mAb to VLA-4 and with mAb to VLA-5 or to VLA-6 further inhibited PMNL migration, irrespective of whether CD11/CD18 mechanisms were blocked with anti-CD18 mAb or not. Treatment of PMNL with a peptide based on the VLA-4-binding domain in the CS-1 fragment of fibronectin, but not a control peptide, inhibited PMNL migration to a comparable extent to treatment with mAb to VLA-4. A low level of VLA-4 was expressed on C5a-activated PMNL, detected by immunofluorescence flow cytometry. These results suggest that VLA-4 can be mobilized by human peripheral blood PMNL and can, in addition to VLA-5, VLA-6 and CD11/CD18 integrins, mediate PMNL migration through connective tissue. This is in marked contrast to PMNL transendothelial migration, where beta 1 integrins appear to play no significant role.


Subject(s)
Connective Tissue/immunology , Integrins/immunology , Neutrophils/immunology , Receptors, Lymphocyte Homing/immunology , Receptors, Very Late Antigen/immunology , CD18 Antigens/immunology , Carrier Proteins/pharmacology , Cell Culture Techniques , Cell Movement/drug effects , Cell Movement/immunology , Fibroblasts/immunology , Humans , Integrin alpha4beta1 , Oligopeptides/pharmacology , Receptors, Very Late Antigen/metabolism , Skin/immunology , Synovial Membrane/immunology
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