In vivo production of heat shock protein in mouse peritoneal macrophages by administration of lipopolysaccharide.

The in vivo production of heat shock protein was studied by administration of bacterial lipopolysaccharide (LPS) into mice. Heat shock protein 70 was detected in the extract of adherent peritoneal cells from mice injected intraperitoneally with LPS by using the immunoblotting method. The expression of heat shock protein 70 was found 2 days after injection of LPS and reached its peak 4 days after injection. The intraperitoneal injection of LPS induced the expression of heat shock protein 70, whereas its subcutaneous injection did not. The in vivo production of heat shock protein 70 was inhibited by administration of LPS together with quercetin, an inhibitor of accumulation of heat shock protein 70 mRNA. Tumor necrosis factor alpha enhanced LPS-induced heat shock protein production in vivo. There was a decrease of gamma delta T cells in the peritoneal cavity of mice injected intraperitoneally with LPS. It was suggested that bacterial LPS is a stressful agent which induces the in vivo heat shock protein response, and its administration leads to the production of heat shock protein 70 in peritoneal macrophages.     

Macrophage cytophilic antibody in mice: effect of bacterial lipopolysaccharide on the uptake of immunoglobulins by mouse peritoneal cells

Injection of lipopolysaccharide from Escherichia coli intravenously into mice affected the ability of their peritoneal macrophages to adsorb antisheep erythrocyte cytophilic antibodies in vitro. After a single intravenous injection of lipopolysaccharide, the alteration in antibody uptake took the form of a triphasic response. An early evanescent increase in adsorptive capacity took place, which reached a maximum 5 min after injection and was followed immediately by a phase of depressed uptake of antibody. Enhancement of antibody uptake then increased to reach a maximum about 24 hr after injection, before declining slowly. The second enhancement phase was inhibited by prior immunization with E. coli or after multiple injections of lipopolysaccharide. Exposure of macrophages to lipopolysaccharide in vitro did not alter their capacity to take up cytophilic antibody. Changes in the capacity of macrophages and polymorphonuclear neutrophil leukocytes to adsorb opsonized erythrocytes were induced by injection of lipopolysaccharide and were similar to those induced with respect to the uptake of cytophilic antibodies by macrophages.     

小鼠脾脏B淋巴细胞纯化培养方法的建立

背景：B淋巴细胞是机体免疫系统重要的参与者,目前的研究主要采用磁珠、补体等方法进行B淋巴细胞的分离纯化,但是这些方法费用高或者细胞损伤大、纯度低,体外分离、培养B淋巴细胞的方法还有待进一步改进。 目的：探讨从小鼠脾脏细胞中对B淋巴细胞同时进行分离、培养的方法。采用加入白细胞介素4、脂多糖或者CD3单克隆抗体及其组合,探讨体外分离、培养小鼠脾脏B淋巴细胞的适宜条件。 方法：体外分离培养小鼠脾脏细胞,随机分为7组,分别用白细胞介素4,CD3,脂多糖,白细胞介素4+CD3,白细胞介素4+脂多糖,CD3+脂多糖组进行干预、将未给予刺激的脾细胞作为对照组。用流式细胞仪检测在不同培养条件下小鼠脾脏细胞T、B淋巴细胞及其亚群的变化。 结果与结论：与对照组比较,白细胞介素4组淋巴细胞在培养后第3-5天数量达到高峰;脂多糖组在培养初期无明显作用,第3天开始淋巴细胞数量有明显的增加,第5天达到高峰;培养体系中加入CD3单克隆抗体,可导致T淋巴细胞消失,培养2 d后,可得到较为单一的B淋巴细胞,其细胞数量在第3天达到高峰。其中B220+IgD+成熟B淋巴细胞亚群数量显著增加。体外培养24 h后,各组B220+CD93+Transitional B淋巴细胞亚群均完全消失。结果说明,体外培养的小鼠脾脏细胞加入CD3单克隆抗体和白细胞介素4可以去除T淋巴细胞,并维持成熟B淋巴细胞的生存和增殖。  中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

Functional maturation of immature B cells accumulated in the periphery by an intraperitoneal administration of a traditional Chinese medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to)

We found that an intraperitoneal (ip) injection of a traditional Chinese herbal medicine, xiao-chai-hu-tang (Japanese name: shosaiko-to), induced accumulation of B lymphocytes (sIgM+) in the peritoneal cavity and spleen. 1) Cell surface marker analysis by a fluorescence-activated cell sorter (FACS) demonstrated that the accumulated B cells on day 4 or 7 after shosaiko-to administration (early phase) were composed mainly of sIgM+IgD- cells and suggested that these B cells maturated into sIgM+IgD+ cells on days 10 or 14 (late phase). Relative decrease of IgM+IgD+ cells at early phase was more profound in peritoneal cells (PC) than in spleen cells. 2) With respect to spleen lymphocytes, antibody responses to a thymus-independent (TI) antigen of type 2 (trinitrophenylated Ficoll) and a thymus-dependent (TD) antigen (sheep erythrocyte) were enhanced at late phase but not at early phase. In contrast, responses to trinitrophenylated lipopolysaccharide (TNP-LPS) as a TI-1 antigen and LPS as a B cell mitogen or a polyclonal B cell activator were enhanced markedly at early phase but declined at late phase. 3) With respect to peritoneal lymphocytes, responses to LPS were suppressed at early phase but recovered at late phase. Enhanced responses to TI and TD antigen at late phase in spleen lymphocytes and suppressed response to LPS at early phase in peritoneal lymphocytes may be explained by increases of IgM+IgD+ mature B cells and IgM+IgD- immature B cells, respectively, at those times. Enhanced responses to TI-1 or LPS in spleen lymphocytes at early phase may be explained by elevated sensitivity of IgM+IgD+ cells which reside in the spleen before shosaiko-to administration and receive the direct stimulation by shosaiko-to, or by acquired responsiveness of IgM+IgD- cells which migrate after stimulation with shosaiko-to.     

In vivo rapid reduction of alloantigen-activated CD8+ mature cytotoxic T cells by inhibitors of acidification of intracellular organelles, prodigiosin 25-C and concanamycin B

Prodigiosin (PrG) 25-C and concanamycin B (CMB) are immunosuppressants that specifically inhibit the induction of cytotoxic T cells (CTL) without affecting the function of B cells and helper T cells in vivo. Both compounds inhibit acidification of intracellular organelles and induce destruction of cytotoxic granules and degradation of perforin in vitro. Here we show that a single intraperitoneal (i.p.) injection of PrG 25-C, and of CMB, into mice eliminates cytotoxic activity 7 days after alloantigen stimulation (when mature CTL activity has been detected in control mice), with minimal effect on the alloantigen-specific antibody titre in serum. FK506 did not suppress the cytotoxic activity with this administration schedule. Suppression was accompanied by a decrease in the CD8+ population and in perforin expression of spleen cells induced by alloantigen stimulation. The suppression of CTL activity and decrease in CD8+ cell number was detected as early as 7 hr after the injection of compounds. These results suggest that inhibitors of acidification of intracellular organelles suppress CTL activity in vivo by reducing the number of mature CD8+ CTL.     

Effects of granulocyte/macrophage colony-stimulating factor on the development and differentiation of CD5-positive macrophages and their potential derivation from a CD5-positive B-cell lineage in mice.

In co-cultures of either the murine pre-B cell line J13, fetal liver cells, or adult peritoneal or bone marrow cells with ST2 mouse bone marrow stromal cells in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF), the development of CD5+ macrophages was demonstrated by immunohistochemical staining and flow cytometry. Although CD5+ macrophages were not present in the peritoneal cavities of normal mice, approximately 30% of the peritoneal macrophages in viable motheaten (mev/mev) mice, deficient in SHP-1 protein tyrosine phosphatase, expressed cell surface CD5 and B220, markers for B cells. In the mev/mev mice, GM-CSF level in peritoneal fluid was increased significantly. At 5 days after daily intravenous injection with GM-CSF, many CD5+ macrophages appeared in the peritoneal cavity and in omental milky spots of normal mice but fewer in osteopetrosis (op) mutant mice, deficient in macrophage (M)-CSF. These results indicate that GM-CSF, in combination with M-CSF, induces the development and differentiation of CD5+ macrophages in the peritoneal cavity, particularly in the omental milky spots of mice. In the peritoneal cavity of GM-CSF-treated mice, the percentages of hematopoietic progenitor cells doubly positive for CD5 and CD34 or c-kit and of macrophage precursor cells doubly positive for CD5 and ER-MP58 or ER-MP20 were increased significantly during the development of CD5+ macrophages and CD5 B cells, suggesting that CD5+ macrophages and B cells may share a bipotential progenitor in vivo.     

All T15 Id-positive antibodies (but not the majority of VHT15+ antibodies) are produced by peritoneal CD5+ B lymphocytes.

After adult irradiation and reconstitution with autologous bone marrow, BALB/c and C.B20 mice no longer utilize the T15 Id in response to phosphorylcholine. T15 Id expression can be restored by transfers of peritoneal B cells or by FACS-purified CD5+ IgM+ lymphocytes (but not by T lymphocytes) from syngeneic donors. Using bone marrow and peritoneal cell donors that are congeneic for heavy and light chain allotypes, the exclusive origin of the T15 Id in peritoneal B cells was ascertained. These conclusions have been essentially confirmed by immunization with either anti-T15 Id or anti-VHT15 antibodies conjugated to lipopolysaccharide. Thus, the production of VHT15-positive antibodies continues at control levels in bone marrow-reconstituted animals while no T15 Id production can be stimulated even in this protocol of direct B cell stimulation. These results constitute the first formal demonstration of the exclusive production of and Id by CD5+ B-cells.     

Surface immunoglobulin ligands and cytokines differentially affect proliferation and antibody production by human CD5+ and CD5- B lymphocytes.

Normal human peripheral blood B lymphocytes were separated into CD19+ CD5+ and CD19+ CD5- subsets by dual-color FACS sorting. In most experiments the cells were activated with Staphylococcus aureus Cowan I (SAC) and cultured in the absence or presence of recombinant human IL-1 alpha, IL-2, or IL-6, or combinations of these cytokines. Unstimulated CD5+ and CD5- B cells showed a comparable, low level of incorporation of [3H]thymidine into DNA. SAC stimulated proliferation of CD5+ and CD5- B cells, and this proliferation was augmented by IL-2 in the case of CD5- B cells. Anti-mu beads stimulated some proliferation of the CD5- subset and augmented SAC-induced proliferation of these cells. In contrast, anti-mu beads did not stimulate proliferation of the CD5+ subset and had no effect on SAC-induced proliferation of these cells. CD5+ B cells activated by anti-mu beads were stimulated to proliferate in the presence of IL-4, but not in the presence of IL-2. These observations support the interpretation that two signals are required for proliferation of CD5+ B cells. Using a two-step culture system, SAC activation itself did not induce Ig production by either subset of purified B cells. However, it primed the cells for antibody production in the presence of IL-2. IL-1 and IL-6 by themselves augmented antibody formation by these cells slightly, if at all. However, IL-6, and to a lesser extent IL-1, augmented antibody production in the presence of IL-2. Under the culture conditions used CD5- B cells produced IgM, IgG, and IgA whereas the CD5+ B cells produced almost exclusively IgM. The expression on B cells of surface activation markers was analyzed after culture for 2 days with SAC or anti-mu beads. In both subsets expression of Leu-23 and Leu-21 was increased, with some differences in intensity (Leu-23 greater in CD5+ cells, Leu-21 greater in CD5- cells). SAC increased IL-2R expression to a greater extent than anti-mu beads. In neither subset was expression of CD23 increased. These observations are discussed in the context of the possible role of the CD5+ subset of B lymphocytes as components of a system of natural immunity.     

In vitro IgG rheumatoid factor production by CD5-negative murine B cells in response to immune complexes of lipopolysaccharide.

We studied the in vitro production of rheumatoid factor (RF) by spleen cells of normal adult mice. IgG RF cross-reactive with rabbit IgG was produced in response to immune complexes of TNP-lipopolysaccharide (LPS) with murine IgG anti-TNP antibody in an Fc-specific manner, but not to a mixture of IgG and LPS. Antibody-uncomplexed LPS induced little IgG RF production, but suppressed the subsequent IgG RF response to antibody-complexed LPS, whereas IgM RF was induced by either LPS or antibody-complexed LPS. The IgG RF production followed as rapid a time course as IgM RF production; the rate of IgG RF production reached its maximum soon after a lag period of 1 day and declined after 5 days. Treatment of splenic B cells from BALB/c mice with anti-Ly-1.2 antibody and rabbit complement resulted in a selective reduction of IgM RF production by 90%, with little effect of IgG RF production. These results suggest that IgG RF is derived primarily from CD5- memory B cells which have been developed in normal mice by an unknown mechanism. Unlike the CD5+ precursor cells for IgM RF, these memory cells are unresponsive to polyclonal stimulation by LPS but are activated by simultaneous stimulation by aggregated Fc epitopes and the mitogenic stimulus from LPS.     

Elimination of CD4(+)CD25(+) T cell accelerates the development of glomerulonephritis during the preactive phase in autoimmune-prone female NZB x NZW F mice

The role of CD4(+)CD25+ T cell in glomerulonephritis (GN) development during the preactive phase was investigated in autoimmune-prone female NZB x NZW F1 (B/WF1) mice. The administration of anti-mouse CD25+ T-cell monoclonal antibody (PC61.5) 3 days after birth induced the development of GN with an increase in IgG2a antinuclear antibody, productions of IL-6 and IFN-gamma, whereas TGF-beta1 production decreased, compared to untreated control mice. The present study results suggest that CD4(+)CD25+ T cells may, at least in part, downregulate the development of GN during the preactive phase in B/WF1 mice.     

Distinct surface phenotypes of B cells responsible for spontaneous production of IgM and IgG anti-DNA antibodies in autoimmune-prone NZB x NZW F1 mice

Autoimmune-prone NZB x NZW F1 (B/W F1) mice produce a high titer of anti-DNA antibodies, In vivo and in vitro studies showed that in the early life of these mice, the immunoglobulin isotype of these antibodies almost exclusively belongs to IgM class, however, IgG anti-DNA antibodies begin to develop when the mice are about 5-6 months old and the titer exceeds that of IgM antibodies from age 7 months on. We asked whether or not the B cell population responsible for IgM and IgG antibody production belongs to the same lineage. The surface phenotypes of B cell populations responsible for the spontaneous production of either IgM or IgG anti-DNA antibodies were examined using panning and sorting methods with several monoclonal antibodies to B cells, including CD5 (Ly-1) and Lp-3; the latter defines a unique B cell differentiation antigen. We obtained evidence that surface phenotypes of B cells secreting IgM anti-DNA antibodies belong to CD5+ Lp-3- and those of B cells secreting IgG anti-DNA antibodies which occur only in old B/W F1 mice belong to CD5- Lp-3+ subpopulations. The majority of peritoneal B cells were CD5+ Lp-3+ throughout the life span of the mice and anti-DNA antibody production was never evidenced. These findings were discussed in relation to age-associated changes of B cell populations in the spleen of this strain of mice.     

Human peripheral blood CD4+ and CD8+ T cells express Th1-like cytokine mRNA and proteins following in vitro stimulation with heat-inactivated Brucella abortus.

Defining the pattern of lymphokine production associated with Brucella abortus is critical for advancing the development of B. abortus as a vaccine carrier. In the present study we investigated the ability of heat-inactivated B. abortus or lipopolysaccharide from B. abortus to induce lymphokine production from purified human T cells in vitro. Gamma interferon (IFN-gamma), interleukin-2 (IL-2), IL-4, and IL-5 induction was assayed by mRNA-specific PCR and by enzyme-linked immunosorbent assay and bioassay for protein production. Following depletion of monocytes and B cells, B. abortus increased IFN-gamma and IL-2 mRNA expression in purified T cells compared with expression in unstimulated cells. In contrast, no IL-5 mRNA expression and only transient low-level IL-4 mRNA expression and no IL-4 protein secretion were detected. Phytohemagglutinin or phorbol myristate acetate plus ionomycin induced mRNA and protein for all these cytokines. Similar results were obtained with LPS purified from B. abortus. Removal of NK cells did not reduce lymphokine production, and enriched NK cells did not express IFN-gamma mRNA or secrete IFN-gamma protein in response to B. abortus, indicating that NK cells were not the responding population. Both CD4+ and CD8+ populations produced IFN-gamma and IL-2 in response to B. abortus. Preincubation of resting T cells with B. abortus or LPS from B. abortus for 7 days induced their differentiation into Th1-like cells as judged by their subsequent lymphokine response to phorbol myristate acetate plus ionomycin. These results suggest that B. abortus can induce differentiation of Th0 into Th1-type cells.     

Mouse peritoneal cells: their ability to elicit or produce antibody after exposure to antigen

The functional activities of peritoneal cells in exudates of normal mice were studied.

Antigen-containing peritoneal cells were tested for their ability to induce antibody by transfer into normal syngeneic recipients. Collection of peritoneal cells at varying time intervals following intraperitoneal injection of formalin-killed pneumococci or capsular polysaccharide, resulted in a decrease with time in the ability of these cells to induce antibody in the recipient mice. This finding could be ascribed to changes in the cell population in the peritoneum as a result of antigen administration rather than to loss of immunogenic material from these cells. Evidence for this comes from experiments in which peritoneal cells collected ½-hour after antigen administration were maintained in vitro for up to 24 hours. The immunogenicity of carbohydrate antigen in these cells persists for at least 24 hours. Furthermore our morphological observations showed a drop in macrophages following antigen administration.

Our findings indicate that peritoneal exudates also contain immunocompetent cells. For this study we used recipient mice irradiated with 650–800 r, which completely suppressed their antibody response. Peritoneal cells collected 2 hours after injection of formalin-killed pneumococci led to serum antibody in the irradiated recipients 7 days later. By partially fractionating peritoneal cells on glass surfaces, it could be shown that the cells not adhering to glass (lymphocytes and medium sized cells) were mainly responsible for this antibody response. Cells adhering to glass which contained most of the antigen-containing large macrophages and also medium sized cells, were much less effective in inducing antibody. In addition, peritoneal cells induced with thioglycollate medium consisting mainly of large macrophages and very few lymphocytes, actively induced antibody in normal recipients but failed to restore antibody production in the highly irradiated mice. The transfer of antibody formation to irradiated mice thus is due to immunocompetent cells in normal peritoneal exudates.

     

IgG1 production by sIgD+ splenic B cells and peritoneal B-1 cells in response to IL-5 and CD38 ligation.

CD38 ligation on mouse B cells by CS/2, an anti-mouse CD38 mAb, induces proliferation, IL-5 receptor alpha chain expression and tyrosine phosphorylation of Bruton's tyrosine kinase. Furthermore, stimulation of splenic B cells with IL-5 together with CS/2 induces Blimp-1 expression and differentiation into Ig-producing cells. Here we examined the role of IL-5 in IgG1 and IgA production by B cells isolated from the spleen and peritoneal cavity. CD38 recognized by CS/2 was expressed in the follicular mantle B cells surrounding the germinal center, sIgD+ splenic B cells and peritoneal B cells. IL-5 induced IgG1 production in splenic sIgD+ B cells stimulated with CS/2, while it was ineffective to induce IgA production. Among the various cytokines tested, only IL-5 had a synergistic effect on IgG1 production with CS/2. IL-5 could induce the generation of S micro-Sgamma1 reciprocal recombination DNA products in CS/2-stimulated B cells. IL-4 was ineffective to induce either micro-gamma1 switch recombination or IgG1 secretion with CS/2, demonstrating that IL-5 promotes both micro-gamma1 switch recombination and IgG1 secretion in an IL-4-independent manner. The peritoneal B-2 cells exhibited both IgG1 and IgA production in response to IL-5 plus CS/2, while B-1 cells produced IgG1. These results imply that the pattern of differentiation to Ig-producing cells seen with peritoneal B cells is not identical to the pattern seen with splenic B cells and that peritoneal B-2 cells contain precursors of IgA-producing cells responding to IL-5 plus CS/2.     

Administration of Silica Sensitizes Lipopolysaccharide Responsiveness of Murine Macrophages but Inhibits T and B Cell Priming by Inhibition of Antigen Presenting Function

Macrophages play a key role in natural host defense against infection by a variety of pathogens. In addition, macrophages initiate the development of acquired immunity via antigen processing and presentation. The role of macrophages in resistance to pathogens, the development of autoimmune diseases and the induction of acquired immunity has been studied by treatment of rodents with reagents which are cytotoxic. We have studied the effects of one such reagent, silica, on the function of spleen macrophages and peritoneal exudate cells (PEC). Intraperitoneal administration of silica caused the accumulation of spleen macrophages and neutrophils, reduction in the number of B cells and had a modest effect on T cell abundance. The percentage of CD11b⁺ PEC was not affected by silica treatment but total PEC recovery was diminished 5-8 fold. Silica treatment did not cause release of TNF-alpha or IL-1-beta but, when stimulated with lipopolysaccharide (LPS) in vitro after silica treatment, PEC or spleen macrophages produced elevated levels of both cytokines compared to controls. In contrast, release of IL-12 from non-LPS treated PEC was stimulated 4-5 fold by silica treatment. In addition, sensitivity to LPS toxicity in vivo was significantly enhanced by silica. The ability of macrophages to present antigen to a T cell clone in vitro was found to be dramatically inhibited by silica treatment, as was the ability to prime antigen-specific T cells and B cells by antigen injection. Collectively these data demonstrate that silica treatment enhances macrophage sensitivity to LPS exposure but inhibits antigen processing and presentation.     

Recombinant Dirofilaria immitis polyprotein that stimulates murine B cells to produce nonspecific polyclonal immunoglobulin E antibody

Nonspecific immunoglobulin E (IgE) production is an event characteristically observed in parasitic helminth infections, but its mechanisms are still unclear. To define these mechanisms, we prepared a recombinant Dirofilaria immitis protein (rDiAg) and assessed its effect on nonspecific IgE production. rDiAg preferentially induced nonspecific IgE production, without eliciting specific IgE production, as well as a Th2-type cytokine profile (high interleukin-4 [IL-4] and IL-10 production but low gamma interferon production) in BALB/c mice. rDiAg significantly elicited the proliferative response of naive B cells. This response was not abolished by polymyxin B, an inhibitor of lipopolysaccharide (LPS), and rDiAg normally expanded splenic B cells from LPS nonresponder C3H/HeJ mice. Thus, the mitogenic effect of rDiAg was not due to LPS contamination. rDiAg also enhanced levels of CD23 expression on splenic B cells. Splenic B cells produced marked levels of IgE when cultured with the combination of rDiAg and IL-4 (rDiAg-IL-4), whereas peritoneal B cells produced negligible levels of IgE. rDiAg-IL-4-induced IgE production by splenic B cells was synergistically increased by coculture with peritoneal B cells. rDiAg-driven IL-10 secretion was higher in peritoneal B cells than in splenic B cells. IgE production by splenic B cells cocultured with peritoneal B cells was decreased to a level comparable to that by splenic B cells in the presence of a neutralizing anti-IL-10 monoclonal antibody. Collectively, these results suggest that rDiAg-induced polyclonal expansion and IgE class switching of splenic B cells contribute to nonspecific IgE production and that these responses are enhanced by peritoneal B-cell-derived IL-10.     

乙肝疫苗增强细胞因子诱导的杀伤细胞对于转乙型肝炎病毒基因小鼠的治疗作用

目的 观察乙肝疫苗是否增强细胞因子诱导的杀伤细胞(cytokine-induced killer cell,CIK)对于乙型肝炎病毒转基因小鼠(hepatitis B virus transgenic mice,HBV-Tg)的抗病毒作用.方法 给予HBV-Tg腹腔注射CIK细胞,皮下注射乙肝疫苗,用荧光定量PCR检测外周血中HBV DNA水平变化,流式细胞仪检测外周血中T淋巴细胞亚群,并用HE染色观察肝脏的组织病理改变.结果 CIK细胞降低了HBV-Tg鼠外周血中病毒载量,血中CD3~+、CD4~+及CD8~+细胞增多,乙肝疫苗和CIK细胞联合应用后,这种作用增强.结论 乙肝疫苗增强了CIK对于HBV-Tg小鼠的治疗作用,这种增强作用是否通过增加体内CD3~+、CD4~+及CD8~+细胞,尤其是CD8~+细胞而实现,有待进一步研究.     

Induction of interleukin-12 production in mouse macrophages by berberine,a benzodioxoloquinolizine alkaloid,deviates CD4+ T cells from a Th2 to a Th1 response

In this study we investigated whether berberine-mediated induction of interleukin-12 (IL-12) production in antigen-presenting cells could regulate a cytokine profile of antigen-primed CD4+ T helper (Th) cells. Pretreatment with berberine induced IL-12 production in both macrophages and dendritic cells, and significantly increased the levels of IL-12 production in lipopolysaccharide-stimulated macrophages and in CD40 ligand-stimulated dendritic cells. Importantly, berberine pretreatment of macrophages increased their ability to induce interferon-gamma (IFN-gamma) and reduced their ability to induce IL-4 in antigen-primed CD4+ T cells. Berberine did not influence the macrophage cell surface expression of the class II major histocompatibility complex molecule, the co-stimulatory molecules CD80 and CD86, and intracellular adhesion molecule-1. Addition of neutralizing anti-IL-12p40 monoclonal antibody to cultures of berberine-pretreated macrophages and CD4+ T cells restored IL-4 production in antigen-primed CD4+ T cells. The in vivo administration of berberine resulted in the enhanced induction of IL-12 production by macrophages when stimulated in vitro with lipopolysaccharide or heat-killed Listeria monocytogenes, leading to the inhibition of the Th type 2 cytokine profile (decreased IL-4 and increased IFN-gamma production) in antigen-primed CD4+ T cells. These findings may point to a possible therapeutic use of berberine or medicinal plants containing berberine in the Th type 2 cell-mediated immune diseases such as allergic diseases.     

外源性IL-23对病毒性心肌炎小鼠Th17细胞增殖的影响

目的:观察小鼠重组IL-23(rIL-23)对病毒性心肌炎(VMC)小鼠Th17细胞增殖的影响。方法:BALB/c小鼠腹腔注射柯萨奇病毒B3(CVB3)建立VMC小鼠模型,1周后磁珠分离脾CD4+T淋巴细胞后进行体外培养,分别加入植物血凝素(PHA)与重组小鼠IL-23(rIL-23)或PHA单独刺激进行体外培养5天。流式细胞术测定培养后Th17细胞占CD4细胞的百分比、RT-PCR测定培养细胞IL-17 mRNA的表达、ELISA法测定培养物上清液中IL-17的水平。结果:与单纯PHA刺激比较,rIL-23刺激后Th17细胞明显增加(1.82%±0.17%比4.70%±1.29%),且培养细胞IL-17 mRNA及上清液中IL-17浓度也增加,两组比较有统计学差异(P均<0.05)。结论:外源性rIL-23可以增加并维持Th17细胞的增殖。     

Studies on the Role of CD43 in Human B-Cell Activation and Differentiation

The monoclonal antibody (MoAb) B1B6 to human leucocyte sialoglycoprotein, CD43, induces aggregation of T cells and delivers progression signals early during activation of both T and B cells in the presence of primary activators of protein kinase C. In this report we further studied the role of CD43 in human B-cell activation and differentiation. About 5-10% of resting tonsillar B cells are CD43+. In the presence of TPA or antibodies to CDw40, the proportions of CD43+ cells drastically increased. The expression was optimal on day 3 of culture, when up to 80% and 50%, respectively, were CD43+. Whereas MoAb B1B6 together with TPA induced a three- to fivefold higher proliferative response as compared to TPA alone, antibodies to CDw40 did not synergize with MoAb B1B6 in B-cell proliferation. Tonsillar populations depleted of CD43+ B cells responded with lower proliferation to TPA alone or to TPA and B1B6 or anti-CDw40 antibodies. MoAb B1B6 did not affect the production of IgM or IgG as induced by pokeweed mitogen in the presence of autologous T cells, from either peripheral blood or tonsillar B cells. Neither did it affect the IgG production from the CD43+ BSF-2 sensitive Epstein-Barr virus-transformed lymphoblastoid cell line CESS. The results show that CD43 is upregulated on B cells during activation. Furthermore, CD43+ B cells are included in the population which responds to signals delivered by TPA, anti-CD43 or anti-CDw40 antibodies, and the proliferation of this population is not merely due to an expansion of the small population of CD43+ cells present among these cells. Moreover, the epitopes recognized by MoAb B1B6 are not involved in the differentiation of and ultimate Ig-secretion from activated B cells.