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Interaction of mouse immunoglobulins with a mast cell tumor HC was studied by the rosette technique. All mast cells formed rosettes, and this could be inhibited by IgG1 myeloma proteins, less efficiently by IgG2 , and not by IgA or IgM. Antigen antibody complexes formed under appropriate con ditions also inhibit, with at least 100-fold greater efficiency than free im munoglobulin. Antisera containing reaginic activity also inhibit rosettes in the presence of antigen, but on heating the sera, a concomitant loss of reaginic activity and rosette inhibition occurs. It is proposed that there is a specific receptor on the mast cell surface for IgG1 and reaginic immunoglobulins, and that binding to this site in the form of a complex of correct antigen ratio, will initiate the anaphylactic process. 相似文献
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Feng Huang Xiaoyun Tong Huaming Deng Liangqing Fu Ronghua Zhang 《Cellular & molecular immunology》2008,5(6):433-438
Gab2 plays an important role in FcεRI mediated signal events which lead to degranulation from mast cells. The present study was designed to investigate the effect of the synthetic Gab2 (scaffolding adapter Grb2-associated binder 2) siRNA on the antigen-induced activation of RBL-2H3 cells. A double stranded siRNA against Gab2- mRNA was synthesized and transfected into RBL-2H3 cells. After 6 h, cells were then sensitized with dinitrophenyl (DNP)-specific IgE overnight and challenged with dinitrophenyl-human serum albumin (DNP-HSA) to induce mast cell degranulation before supernatants were collected. Effects of Gab2 siRNA on antigen-induced release of β-hexosaminidase and histamine, cytokine production and regulation of the proteins in the pathway were measured by enzymatic assay, EIA, ELISA and Western blotting. Treatment with Gab2 siRNA significantly decreased Gab2 expression, inhibited the FcεRI-mediated mast cell release of β-hexosaminidase and histamine, reduced the production of IL-4 and TNF-α and inhibited the phosphorylation of Akt, PKC8 and p38 mitogen-activated protein kinase (MAPK). Data showed that Gab2 siRNA could suppress the antigen-induced activation of RBL-2H3 cells and suggested a possible mechanism through inhibition of signaling molecules downstream of Gab2 in the 2+ FcεRI-mediated Ca -independent pathway. Furthermore, potential usefulness of Gab2 knock-down as a method for inhibition of mast cell-mediated allergic reactions was demonstrated. Cellular & Molecular Immunology. 2008; 5(6):433-438. 相似文献
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Synaptotagmin (Syt) constitutes a family of membrane-trafficking proteins, so far nearly 20 Syts have been discovered. Extensive work showed that synatotagmins were a potential Ca^2+ sensor for regulated exocytosis. This study was to investigate the expression and location of synaptotagmin II (Syt2) in RBL-2H3 (RBL) and its role in regulating exocytosis of RBL. The expression of Syt2 in RBL was confirmed by Western blot. The recombinant expression vector pEGFP-N1-Syt2 was constructed and transfected into RBL by electroporation, the stable transfectant RBL-Syt2-S expressing fusion protein Syt2-EGFP were obtained and Syt2 was highly concentrated at plasma membrane with little detected in cytoplasm. To analyze the role of Syt2 during exocytosis of RBL, the release of cathepsin D was assayed by immunoblotting. Compared with control, the release of cathepsin D by RBL-Syt2-S was markedly decreased. The results indicated that Syt2 played a negative regulation in exocytosis of lysosomes in RBL. Cellular & Molecular Immunology. 2005;2(3):205-209. 相似文献
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The CD45 protein is a transmembrane tyrosine phosphatase that is required for normal T and B cell receptor-mediated signaling. In order to study the function of this phosphatase in mast cells, we have isolated a CD45-deficient variant from the rat basophilic leukemia cell line (RBL-2H3), a tumor analog of mucosal mast cells. The secretory response as well as the inositol 1,4,5-triphosphate (InsP3) formation to FcRI and ionophore stimuli were similar in the RBL-2H3 cell line and its derived CD45-deficient sub-population. However, pretreatment with the phorbol ester TPA, which directly activates protein kinase C (PKC), caused a marked increase in mediator release and InsP3 production in the CD45-deficient variant compared to the parental RBL-2H3 cells. These findings suggest that CD45 might directly or indirectly modify the activity of PKC or the InsP3-dephosphorylating phosphatase. 相似文献
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Synaptotagmin(Syt)constitutes a family of membrane-trafficking proteins,so far nearly 20 Syts have beendiscovered.Extensive work showed that synatotagmins were a potential Ca~(2+) sensor for regulated exocytosis.Thisstudy was to investigate the expression and location of synaptotagmin Ⅱ(Syt2)in RBL-2H3(RBL)and its role inregulating exocytosis of RBL.The expression of Syt2 in RBL was confirmed by Western blot.The recombinantexpression vector pEGFP-N1-Syt2 was constructed and transfected into RBL by electroporation,the stabletransfectant RBL-Syt2-S expressing fusion protein Syt2-EGFP were obtained and Syt2 was highly concentrated atplasma membrane with little detected in cytoplasm.To analyze the role of Syt2 during exocytosis of RBL,therelease of cathepsin D was assayed by immunoblotting.Compared with control,the release of cathepsin D byRBL-Syt2-S was markedly decreased.The results indicated that Syt2 played a negative regulation in exocytosis oflysosomes in RBL.Cellular & Molecular Immunology.2005;2(3):205-209. 相似文献
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Etsuko Hirose Miyoko Matsushima Kenzo Takagi Yui Ota Keiko Ishigami Tatsuya Hirayama Yuta Hayashi Toshinobu Nakamura Naozumi Hashimoto Kazuyoshi Imaizumi Kenji Baba Yoshinori Hasegawa Tsutomu Kawabe 《Inflammation》2009,32(2):99-108
Kaempferol is one of the most commonly found dietary flavonoids. The exposure to kaempferol is known to inhibit degranulation
from mast cells, but the inhibitory mechanism of degranulation has not been clarified yet. In this study, we investigated
the involvement of heme oxygenase (HO)-1 in the anti-allergic action of kaempferol against degranulation in rat basophilic
leukemia (RBL-2H3) cells. Our results demonstrate upregulation of HO enzymatic activity after short (15 min) exposure to kaempferol,
followed by the induction of HO-1 expression in protein. The involvement of HO-1 in the kaempferol-induced inhibition of degranulation
was confirmed using tin protoporphyrin IX (SnPP), a HO-1 inhibitor. These findings strongly suggest that kaempferol exerts
anti-allergic actions via activation of the HO-1.
Etsuko Hirose and Miyoko Matsushima have contributed equally to this work. 相似文献
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《Immunological investigations》2013,42(8):503-515
The CD45 protein is a transmembrane tyrosine phosphatase that is required for normal T and B cell receptor-mediated signaling. In order to study the function of this phosphatase in mast cells, we have isolated a CD45-deficient variant from the rat basophilic leukemia cell line (RBL-2H3), a tumor analog of mucosal mast cells. The secretory response as well as the inositol 1,4,5-triphosphate (InsP3) formation to FcRI and ionophore stimuli were similar in the RBL-2H3 cell line and its derived CD45-deficient sub-population. However, pretreatment with the phorbol ester TPA, which directly activates protein kinase C (PKC), caused a marked increase in mediator release and InsP3 production in the CD45-deficient variant compared to the parental RBL-2H3 cells. These findings suggest that CD45 might directly or indirectly modify the activity of PKC or the InsP3-dephosphorylating phosphatase. 相似文献
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目的:研究SynaptotagminⅡ(简称syt2)在RBL-2H3(简称RBL)中的表达及其在胞吐中的作用。方法:通过Westernblot检测Syt2在RBL中的表达情况。用高保真酶扩增Syt2,与pEGFP-N1构建全长反义基因表达载体,电穿孔转染RBL,G418筛选获得稳定转染细胞。通过钙离子载体和抗原刺激,应用Westernblot检测稳定转染细胞和对照细胞分泌的组织蛋白酶D,分析Syl2对胞吐的影响。结果:在RBL中检测到Syt2表达。构建了pEGFP-N1-Syt2-AS质粒,插入片段测序结果与GenBank登录号NM012665(ratsyt2)序列完全一致。经转染和G418筛选,获得了稳定转染细胞RBL-Syt2-AS,Westernblot结果显示,两株RBL-Syt2-AS表达的syt2均明显减少,分别只有对照的8%和10%。经刺激后,RBL-Syt2.AS分泌的组织蛋白酶D较对照明显增加。结论:Syt在RBL中表达,其对RBL溶酶体胞吐起负调控作用。 相似文献
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先天免疫是机体抗细菌、抗病毒和抗肿瘤等的第一防线.Toll样受体(Toll-like recep-tors)是先天免疫的关键受体.它可以识别致病微生物的分子模式(pathogen associated moleculal pat-terns)而激活先天免疫细胞和进一步调整后天免疫系统.近来发现的Toll样受体在癌症的发生和发展中也扮演着重要角色.Toll样受体的激活因子在抗癌免疫疗法中可用作免疫调节剂(immunoadju-vants)或细胞毒素的药物.在这里,我们将简述当前关于Toll样受体的研究进展和在癌症的发生发展和癌症免疫疗法的作用. 相似文献
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OBJECTIVE: Nicorandil is a potassium channel opener and nitric oxide (NO) donor, and the hypothesis was tested that these modes of action may inhibit cellular degranulation and release of tumour necrosis factor-alpha (TNF-alpha). MATERIALS AND METHODS: TNF-alpha and beta-hexosaminidase secretion were measured from rat basophilic leukemia cells (RBL-2H3) activated via the high affinity IgE receptor with dinitrophenyl-albumin (DNP-A) challenge in the presence of nicorandil. Inhibitors of K+ openers and NO were pre-incubated with the RBL-2H3 cells to determine the principal mode of action. RESULTS: Nicorandil significantly inhibited the release of TNF-alpha in a dose-dependent manner (p < 0.001, ANOVA) reaching a maximum inhibition of DNP-A 74.1% at 10(-3) M, (p < 0.001). Similarly it inhibited beta-hexosaminidase release (p < 0.001, ANOVA) with maximal inhibition at 10(-3) M (p < 0.001). Other K+ openers did not show this effect. Neither the potassium channel blocker glibenclamide nor the guanylyl cyclase inhibitor, ODQ, could reverse this inhibition, but when added in combination reduced the effect by 47%. CONCLUSIONS: Nicorandil is able to inhibit degranulation and TNF-alpha release of RBL cells stimulated through the IgE receptor, and requires both the K+ opening and nitric oxide donor activity, which may represent a novel method for inhibiting cytokine release. 相似文献
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《Immunological investigations》2013,42(6):470-480
The present study was designed to investigate the anti-allergic effects of ethanolic extract of Alternanthera sessilis (AS-1) in rat basophilic leukemia (RBL-2H3) cells. It significantly reduced the β-hexosaminidase release from anti-DNP-IgE sensitized RBL-2H3 cells. AS-1also inhibited the IgE antibody-induced increase in Interleukin-6 (IL-6), TNF-α, IL-13 and IL-4 production in these cells. The inhibitory effect of AS-1 on these cytokine was found to be nuclear factor-KB (NF-kB) dependent, as it attenuated the degradation of IKBa and nuclear translocation of NFkB. In addition, AS-1 significantly attenuated the DNP HAS-induced intracellular Ca2+ release from these cells, which makes us speculate strongly that the decreased intracellular Ca2+ is involved in the inhibitory effect of AS-1 on β-hexoaminidase release. Taken together, anti-allergic effects of AS-1 suggest possible therapeutic application of this extract in allergic diseases. 相似文献
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Toll样受体是广泛表达在哺乳动物细胞表面的跨膜信号传导受体,其通过识别多种类型的病原体相关分子模式及一些内源性配体,激活天然免疫系统,同时通过诱导树突状细胞分化成熟,调控获得性免疫反应的建立。大多数TLRs的配体可诱导机体产生Th1型免疫应答,然而在某些条件下也可导致Th2型免疫应答的发生。弄清TLRs参与调节Th0分化的机制,可为今后在感染免疫、自身免疫、超敏反应等方面进行深入研究及相关疾病的治疗提供新的切入点。 相似文献
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Bhattacharyya SP 《Immunology letters》2001,75(2):131-136
Receptor-mediated signal transduction pathways of cells involved in allergy and inflammations are extremely significant. Lyn is a member of the Src family of non-receptor protein tyrosine kinases and is associated with a number of cell surface receptors, including the B-cell antigen receptor and immunoglobulin E receptor (FcepsilonRI). Lyn is necessary for FcepsilonRI-mediated mast cell activation. To investigate how the level of Lyn is maintained in mast cell activation, it was studied whether Lyn binds to ubiquitin and is ubiquitinated for proteasomal degradation in cells. In the yeast two hybrid system, Lyn specifically interacted with ubiquitin in vivo. Furthermore, Lyn bound to ubiquitin-conjugated Sepharose beads in vitro and was efficiently competed by soluble ubiquitin. Pulse-chase experiments indicated intracellular degradation of Lyn was associated with the generation of a high molecular weight complex in the presence of proteasome-specific inhibitor, lactacystin. This high molecular weight complex cross-reacted with anti-Lyn and anti-ubiquitin demonstrating the ubiquitination Lyn. Overexpression of Lyn and ubiquitin in COS 7.2 cells also resulted in the ubiquitination of Lyn in the presence of lactacystin, supporting the ubiquitination of Lyn by a proteasome specific pathway. 相似文献
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We studied the properties of the ectokinase activity on the outer cell surfaces of RBL-2H3 cells and examined the phosphorylation of exogenous substrates to clarify the substrate specificity of the ectokinases on RBL-2H3 cells. Among the several protein substrates tested, casein was the most strongly phosphorylated with [gamma-32P]ATP, and the net incorporation of 32P into casein was 0.65 pmol P/50 microg/10(6) cells. Casein kinase II peptide was also phosphorylated with [gamma-32P]ATP. The phosphorylation of casein and casein kinase II peptide was almost completely inhibited by the addition of 3 microg/ml of cell-impermeable K252b. Phosphorylation of casein and casein kinase II peptide was also observed by [gamma-32P]GTP. Western blot analysis using anti-casein kinase II antibody revealed a 44-kDa casein kinase band in the membrane fraction and Fc epsilonRI complexes. The immunofluorescence microscopic analysis using anti-casein kinase II antibody showed the existence of casein kinase II on the surface of the cells. This is the first report about the existence of ectokinase on mast cells. 相似文献
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Fabrizio Conti Francesca Romana Spinelli Cristiano Alessandri Guido Valesini 《Clinical reviews in allergy & immunology》2011,40(3):192-198
A variety of immune mechanisms, both humoral and cellular, are involved in the onset and amplification of the inflammatory
response in lupus nephritis (LN). Accumulating evidence substantiates the view that innate immunity pathways may also amplify
inflammatory reactions within the kidneys. Toll-like receptors (TLRs) are essential modulators of the innate immune response
thanks to their ability to rocognize conserved molecular patterns that are microbe specific and other danger signals. Their
recognition of endogenous molecules released from injured cells may also contribute to renal inflammation. Studies conducted
in vivo and in vitro provide experimental evidence for the functional role of TLRs in LN. Intriguingly, these data suggest
that pharmacological TLR signal suppression could be a useful approach to the treatment of systemic lupus erythematosus. 相似文献
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Kim GC Lee SG Park BS Kim JY Song YS Kim JM Yoo KS Huh GY Jeong MH Lim YJ Kim HM Yoo YH 《International archives of allergy and immunology》2003,131(2):101-110
BACKGROUND: Magnoliae flores (MF), the buds of Magnolia denudata Desrousseaux, have been successfully used for the management of allergic diseases in Korea. The purpose of the present study was to determine their causal role in inducing apoptosis of mast cells and to verify the underlying mechanism. METHODS: The viability of mast cells was assessed by the trypan blue exclusion test. Induction of apoptosis was confirmed by DNA fragmentation, nuclear staining and DNA hypoploidy. Western blotting and immunofluorescent staining were performed to study the alterations in expression level and translocation of apoptosis-related proteins. Mitochondrial membrane potential (MMP) change and cytochrome C release were assayed. RESULTS: We present several lines of evidence indicating that MF induce apoptosis. Changes in cell morphology, generation of DNA fragmentation, cell cycle arrest, activation of caspase-3, and PARP and DFF degradations were demonstrated. The reduction of MMP and the release of cytochrome C to cytosol were also shown. Either PTP blockers, bongkrekic acid and cyclosporin A, or pancaspase inhibitors, Boc.D-fmk and zVAD-fmk, did not prevent the release of cytochrome C. Bax protein content was increased, and Bax was translocated from cytosol into mitochondria at early time points after MF treatment. CONCLUSIONS: We have demonstrated that MF induce mitochondria- and caspase-dependent mast cell apoptosis. Our observations contribute new insights to the role of MF and support the view that the clinical effect of MF may depend on their pharmacological efficacy in regulating mast cell apoptosis. 相似文献
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目的研究羧胺三唑(CAI)对RBL-2H3肥大细胞增殖、凋亡及活化脱颗粒的影响,探索CAI的抗感染作用机制。方法以C48/80诱导RBL-2H3细胞活化脱颗粒模型,中性红染色法观察细胞脱颗粒的形态学,分别用ELISA法和底物显色法检测细胞培养上清中组胺和β-氨基己糖苷酶的释放水平,CCK-8法测定细胞活力,Hoechst 33342荧光染色法检测细胞凋亡。结果与对照组相比,10、20和40μmol/L CAI能够不同程度抑制C48/80诱导的RBL-2H3细胞脱颗粒反应,20和40μmol/L CAI能够降低C48/80诱导的组胺释放(P0.01),40μmol/L CAI能够降低β-氨基己糖苷酶的释放(P0.01)。另外,所用各浓度的CAI对细胞增殖和凋亡均无明显影响。结论 CAI能有效抑制RBL-2H3肥大细胞的活化脱颗粒,此作用并不是通过细胞毒发挥作用的。CAI可能部分通过下调肥大细胞的功能活化,发挥其抗感染作用。 相似文献
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