Protecting Effects of Vasoactive Intestinal Polypeptide on Lymphocytes Against Metal Toxicity

Abstract

The hormonal neuropeptides calcitonin gene-related peptide (CGRP), cholecystokinin (CCK), neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP), were investigated for a potential protective effect on thymocytes after a toxic dose of nickel sulfate, giving an inhibition of DNA synthesis. There was a statistically significant increase in the synthesis of DNA from the level caused by nickel sulfate, with VIP, l0^-4-10^-5 mol/l, while the slightly stimulating effects obtained with CGRP, CCK and NPY, were statistically non-significant. This indicates that VIP, at least as pharmacological concentrations, might have protective effects on lymphocytes against metal toxicity.     

肠血管活性多肽或生长抑素抑制大鼠肠CD8~+淋巴细胞归巢

为观察肠血管活性多肽 (VIP )及生长抑素 (SST )对大鼠肠淋巴细胞在肠相关淋巴组织归巢的影响 ,本实验将 18只大鼠随机分为三组 ,每组 6只 ,分别从股静脉输入生理盐水、VIP组或SST ,从肠系膜淋巴管插管引流淋巴液。结果显示 ,大鼠经静滴VIP或SST后 ,5h内肠系膜淋巴管淋巴细胞总数降低 (P <0 0 5 ) ;肠淋巴液量和对照组比较无显著改变 (P >0 0 5 ) ;每毫升淋巴液中细胞数降低 (P <0 0 5 )。VIP组和SST组肠淋巴液中CD8+ 细胞比例降低 (P <0 0 5 )。两实验组回肠粘膜CD8+ 细胞数降低 (P <0 0 5 )。VIP或SST能减少肠粘膜CD8+ 淋巴细胞与其他器官及系统免疫的沟通 ,也抑制从其他器官及系统免疫中归巢至肠粘膜的CD8+ 细胞     

肠血管活性多肽对多器官功能障碍综合征大鼠肠淋巴细胞归巢的影响

为观察肠血管活性多肽 (VIP )对大鼠肠缺血再灌注致多器官功能障碍综合征 (MODS )时 ,肠淋巴细胞归巢至肠相关淋巴细胞 (GALT )的调节以及对MODS转归的影响。本研究用微型无创动脉夹夹住大鼠肠系膜动脉根部 4 5min ,松夹后再灌注6h ,制备MODS模型。将VIP分别从股静脉输入和从腹腔注入大鼠体内。收集肠淋巴液 ,计数淋巴细胞总数及T、B淋巴细胞比例 ,了解淋巴细胞进入血循环状况。体外用51Cr标记肠淋巴细胞 ,回输入大鼠体内 ,用γ计数器检测各器官组织内的51Cr 肠淋巴细胞 ,了解肠淋巴细胞归巢。检测血及肠淋巴TNF α、内毒素 ;测定血D 乳酸、谷丙转氨酶 (ALT )、肌肝 (Cr)、血氧分压 ;观察重要器官组织学改变 ,评价VIP对MODS时各脏器形态及功能改变。结果显示 ,VIP使MODS大鼠肠粘膜迁移至血循环肠淋巴细胞总数 [(0 4 2± 0 18)× 10 7/h]较未予处理的MODS组 [(0 2 8± 0 15 )× 10 7/h]显著增加 (P <0 0 5 ) ,以T细胞数上升明显。VIP减少了MODS大鼠归巢至肠粘膜的肠淋巴细胞 ,Peyer淋巴结及小肠分布的51Cr 细胞量分别占总51Cr量的 2 14 %± 1 4 9%、 1 5 8%± 0 4 2 % ,显著低于未予处理的MODS组 (5 0 4 %± 1 2 3%和 3 2 3%± 1 6 9% ,P <0 0 1及P <0 0 5 )。外源性给予MODS大鼠VIP后 ,肠淋巴     

The Effects of Serotonin on the Differentiation of Neurons Producing Vasoactive Intestinal Polypeptide in the Suprachiasmatic Nucleus of the Rat

The morphogenetic influences of serotonin on the differentiation of neurons synthesizing vasoactive intestinal polypeptide (VIP) in the suprachiasmatic nucleus were studied in rats. This was addressed by comparative morphofunctional analysis of VIP neurons in adult rats whose brains developed prenatally in conditions of normal and deficient serotonin metabolism. Serotonin deficiency was created in fetuses by treatment of their mothers with p-chlorophenylalanine (PCPA). Pregnant females in controls were treated with 0.9% NaCl. VIP neurons in experimental and control animals were found to show no differences in VIP mRNA concentrations and, probably, in the level of VIP synthesis. However, inhibition of serotonin synthesis led to an increase in the number of VIP-immunoreactive neurons and an increase in the VIP concentration within these cells. This was not associated with any change in neuron size, which was an indicator of the absence of functional hypertrophy accompanying activation of specific synthesis. Comparison of the data obtained here showed that during prenatal ontogenesis, serotonin has an imprinting influence on the differentiation of VIP neurons and is probably involved in the formation of the mechanism of VIP secretion.     

Vasoactive Intestinal Polypeptide Suppressed Experimental Autoimmune Encephalomyelitis by Inhibiting T Helper 1 Responses

Vasoactive intestinal peptide (VIP) has been found to act as a potent anti-inflammatory factor through regulating the production of both anti- and pro-inflammatory mediators and promoting Th2-type responses. In this study, we used myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis (EAE) model in C57BL/6 mice to investigate the potential effects of VIP on multiple sclerosis. Our results showed that in vivo treatment of EAE-induced mice with VIP had great protective benefit at both clinical and histological levels. Disease suppression was associated with the inhibition of T cells proliferation, shifting of the immune response toward a Th2-type response and influencing the expression of pro-inflammatory cytokines including IFN-γ, IL-6 and IL-2 as well as chemotactic factors such as RANTES. In conclusion, the study provides evidence that VIP had great protective effect on EAE through its inhibition actions on pathogenic T cells and through a specific effect on the Th1 response.Haiyan Li and Yunhua Mei contributed equally to this work     

血管活性肠肽对树突状细胞免疫调节作用研究进展

神经系统与免疫系统通过神经肽、神经递质和细胞因子相互作用。血管活性肠肽(VIP)作为一种重要的神经肽参与炎症反应和免疫应答的调控。近年研究表明VIP能通过与树突状细胞(DC)表面的受体结合调节DC的表型和功能成熟,在DC体内迁移和功能成熟中发挥重要作用。VIP主要通过调节DC膜表面黏附分子和共刺激分子的表达、趋化因子及其受体的表达和趋化活性,诱导Th细胞分化以及抗原提呈功能发挥作用。     

Differentiation and Function of Intestinal Intraepithelial Lymphocytes

Intestinal intraepithelial lymphocytes (i-IEL) are phenotypically diverse and consist of both thymically derived and extrathymically derived cells. Extrathymically derived i-IEL are clearly different from thymically derived peripheral T cells in their phenotype and repertoire selection. The major locus of differentiation of extrathymically derived i-IEL appears to be the intestinal epithelium because recombination activating gene (RAG)-1 is expressed in CD3^? i-IEL. Extrathymic differentiation however does not imply independence from the thymus as athymic mice have increased numbers of CD3^?CD8^? and CD3^?CD8αα⁺ i-IEL but decreased numbers of CD3⁺CD8αα⁺ i-IEL when compared to euthymic mice. We speculate from these results that thymus-derived cytokine(s)/factor(s) may support differentiation from CD3^?CD8αα⁺ to CD3⁺CD8αα⁺ i-IEL in the intestinal epithelium.

I-IEL seem to have some role in immune surveillance because they reside at a site which may represent a first line of defense against pathogenic organisms. This idea is supported by the reports showing in vivo activation of i-IEL under conditions of intestinal infection or tumor-bearing state. In vitro analyses showed cytotoxicity and cytokine production of i-IEL but their true function(s) in vivo is(are) not well known. Clearly more analysis on the in vivo function(s) of i-IEL are needed in order to clarify the true role(s) of i-IEL.     

Generation of Cytotoxic T Lymphocytes Against Ly Alloantigen

Cytotoxic T lymphocytes specific for immune alloantigens controlled by alleles of the Ly system have been induced in vivo. These results were obtained either in a secondary type of response or by treating mice before immunization with a single dose of cyclophosphamide (80 mg/kg).     

Induction of Cytotoxic Lymphocytes Directed Against Public Alloantigens

The response of H-2k lymphocytes to in vitro stimulation by H-2b and HI2d spleen cells has been monitored for the production of cytotoxic lymphocytes (CL). Using cells from H-2b strains CL which were capable of lysing both H-2b and H-2d target cells (TC) were generated. These results, together with competitive inhibitor studies of the cytotoxic activity against H-2d TC, demonstrated that T lymphocytes can respond to both public and private alloantigens of the H-2 complex. When H-2d cells were used for stimulation two of the three strains tested induced CL with cytotoxic activity towards both H-2d and H-2b TC. These observations indicate that the T cell response to alloantigen, leading to the production of cytotoxic lymphocytes, can be multicomponent like the established B cell response and can be expected to be subject to as much phenotypic variation. The full complexity may only be revealed by examing a number of combinations of stimulating strains, methods of stimulation and different kinds of target cells.     

自发性高血压鼠脑底动脉血管活性肠多肽能神经纤维的起源

应用免疫组织化学 ABC法和神经节切除术 ,观察了 2 0只自发性高血压鼠 (分手术 、 、 组和对照组 )脑底动脉血管活性肠多肽能神经纤维的起源。结果显示 ,对照组脑底血管的大脑前动脉、大脑中动脉、大脑后动脉和基底动脉壁上均可见棕褐争的免疫反应阳性纤维 ,纤维似细曲线状 ,呈网状走行。手术 组作一侧蝶腭神经节切除术 ,双侧脑底动脉主要分支的阳性纤维明显减少 ;手术 组作一侧耳神经节切除术 ,双侧大脑后动脉和基底动脉壁上的阳性纤维密度减少 ;手术 组作一侧颈上神经节切除术 ,基底动脉的阳性纤维减少。结果表明 :自发性高血压鼠一侧脑底动脉主要分支的血管活性肠多肽能神经纤维起源于双侧蝶腭神经节、耳神经节和颈上神经节。提示血管活性肠多肽能神经可能在高血压鼠的脑血管运动和高血压的发生和发展方面起着重要的作用     

Expression of Activation and Costimulatory Elements by Human Intestinal Intraepithelial Lymphocytes

It is unclear whether human intestinal intraepithelial T lymphocytes (iIEL) are resting or activated cells. To address this question, an improved isolation procedure was developed for small bowel iIEL, which were analysed by two-colour flow cytometry and compared with resting and mitogen-activated peripheral blood lymphocytes. iIEL expression of CD44 isoforms, Bcl-2 and Ki67 antigen was also determined in tissue sections. iIEL expressed CD69 at levels comparable with 48-72 h phytohaemagglutinin blasts, but did not express CD25 or CD95. iIEL were Bcl-2+ but not Ki67+. alphaEbeta7 and alpha4B7 expression was relatively high, whereas alphaLbeta2, CD5 and CD28 were expressed at low density. Isolated iIEL expressed CD44 (core epitopes) at lower levels than peripheral blood lymphocytes, although almost all CD44 contained splice variant 6 (CD44v6). Peripheral blood lymphocytes expressed CD44 at very high density, but little CD44v6, even after activation. However, in tissue sections, iIEL showed differential labelling with CD44 core epitope antibodies and no detectable CD44v6, implying CD44 receptor occupancy or epitope masking in situ. Thus, normal iIEL express a quasi-activated phenotype with unusual patterns of adhesion receptors, which may act as costimulatory elements. These may permit iIEL to assume effector functions, with absence of CD25 preventing entry into the cell cycle, thereby maintaining an apoptosis-resistant phenotype.     

Enhanced Cytotoxicity of Human Lymphocytes Against Rabies-Infected Cells by Rabies-Specific Antibodies

Human anti-rabies immune sera enhanced the in vitro cytotoxicity of human lymphocytes against rabies virus-infected green-monkey kidney cells. The immune sera were collected from patients immunized with rabies vaccine produced either in human diploid cells or in nervous tissue. Significant cytotoxicity was observed even with high serum dilutions, indicating that the K-cell assay might be a sensitive tool for detection of anti-rabies antibodies.     

Origin and Immunoregulation of Autoantibodies Against Intestinal Alkaline Phosphatase

In the sera of patients with acute bactetial infections specific autoantibodies (sIAPa) of the imtnuno-globulin class G (IgG) were found which bind to intestinal alkahne phosphatase (IAP) through the Fab portion. This was demonstrated using immunoaffinity (IA) isolation of sIAPa from patients' sera (particularly bacterial meningitis and ventriculitis) digestion with pepsin, purification of F(ab')₂ fragments on protein A and subsequently binding on IAP coupled to CNBr (cyanogen bromide)-activated Sepharose. Immunoblots using specific anti-Fc and anti-Fab antibodies showed that the bulk of F(ab')₂ fragments had bound. Additionally, binding of native IAP to the F(ab')₂ fragments was observed after separation of F(ab')₂ fragments using isoelectric focusing (IEF), blotting onto nitro cellulose and incubation with IAP. Moreover, we have demonstrated the occurrence of natural anti-IAP autoantibodies (nIAPa) which were isolated from sera of healthy individuals using IA chromatography. Investigation of isotype distribution revealed that IgG but not IgM or IgA were predominant even among nIAPa. The nIAPa fraction exhibited lower binding efficiencies on IEF blots than the sIAPa fraction, however, in contrast to sIAPa, cross-reactions with other autoantigens were observed for nIAPa. NIAPa and sIAPa did not show subclass restriction. As revealed by IEF the spectrotypes of sIAPa were found to be patient-specific, poly- to oligoclonal and stable during longer periods.     

牛磺酸对大鼠心肌梗塞及血管活性物质的影响

本实验结扎在鼠左冠状动脉前降支建立心肌梗塞模型，观察牛磺酸对心肌梗塞的治疗作用及对血和活性物质的影响。结果显示外源性注射牛磺酸可减少大鼠心肌梗塞坏死面积和缩小梗塞范围：血清ＣＰＫ、ＣＫ ＭＢ活性下降，ＭＤＡ含量降低；牛磺酸还可使血浆内皮素、甘丙素、Ｐ物质、ｃＡＭＰ和ｃＧＭＰ水下下降，提示牛磺酸对急性心肌缺血坯死有明显的保护作用。     

Shared Idiotypic Determinants on B and T Lymphocytes Reactive Against the Same Antigenic Determinants

Normal rat lymphocyte populations house a high percentage of lymphocytes with idiotypic, antigen-binding receptors for the major histocompatibility complex antigens of the rat. These receptors can be isolated from normal serum or lymphocyte supernatants. Idiotypic, antigen-binding molecules released from normal Lewis lymphocytes were thus isolated using anti-(Lewis anti-DA) immuno adsorbents. Analysis by SDS polyacrylamide electrophoresis using molecules labeled by external or internal means (125I or 3H) demonstrated that B lymphocytes produce molecules of 'conventional' 7S-8S IgM type. T-lymphocyte-derived molecules had a molecular weight of around 150,000 and consisted of two chains of similar size. Such single chains would succumb to proteolysis by normal serum factors to yield fragments in the size range of 30,000-40,000 daltons. All three groups of T-cell-derived molecules expressed both antigen-binding and idiotypic markers. No evidence was obtained that any light chains are linked to the T-receptor molecules. Serological analysis of the T-cell molecules failed to prove the existence of any determinants of constant immunoglobulin type, nor did these molecules express antigenic markers of major histocompatibility complex types.     

Shared Idiotypic Determinants on B and T Lymphocytes Reactive Against the Same Antigenic Determinants

In the rat the major histocompatibility locus antigens are determined by the Ag-B locus. In the present article evidence is presented that the sera and urine ui normal adult rats contain naturally occurring antibody-like molecules with reactivity to allogeneic Ag-B antigens. Such molecules can be shown to contain both antigen-binding capacity for the relevant antigens and the idiotypic markers signifying such specific reactivity. The molecules could be shown to be composed of two groups of molecules, one around 7S IgG in size and the other around 35,000 in molecular weight. Only the smaller molecules were found in the urine. From other data we know that the 7S molecules are produced by B lymphocytes and the 35,000-molecular-wfight molecules by T cells. Purified natural anti-Ag-B factors, when inoculated into rabbits or chickens, lead to the production of specific anti-idiotypic antibodies that will selectively inactivate rat T lymphocytes with the capacity to react against the relevant Ag-B antigens while leaving other reactivity intact. We thus conclude that the present system allows the purification of naturally occurring idiotypic B- and T-cell products with antigen-binding specificity for further biochemical and functional analysis.     

Effects of Lentinan on Cytotoxic Functions of Human Lymphocytes

The in vitro effects of lentinan on natural killer (NK), antibody-dependent cell-mediated cytotoxicity (ADCC), lectin-dependent cell-mediated cytotoxicity (LDCC) and mitogen-induced blast transformation were studied in patients with solid tumors and chroyic lymphocytic leukemia (CLL). NK activity was measured against Cr-labelled K-562 targets, ADCC against antibody-coated chicken red cells. LDCC and natural cell-mediated cytotoxicity (NCMC) was assessed using ³H-thymidine prelabelled HEp-2 targets. Mitogen (PHA-and Con A-) induced blast transformation was measured by thymidine incorporation.

Blastogenesis and LDCC was not influenced by lentinan. 1 μg/ml lentinan increased NCMC of tumor-bearing subjects. The most prominent enhancement of NK and ADCC activity was seen in CLL patients, where a dose-related increase was seen (from 0.01 to 1 μg/ml).