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1.
J M Sharma 《Veterinary immunology and immunopathology》1988,19(1):51-66
Spleen cells but not the thymus or the bursa cells of chicken embryos suppressed the in vitro mitogenesis of spleen cells of adult syngeneic or allogeneic chickens. The natural suppressor cell activity of embryo spleen was present at embryonation day 16, reached peak levels at embryonation day 18 and disappeared at hatch. The embryo spleen cells did not by themselves respond to phytohemagglutinin stimulation in vitro. The suppressive effect of embryonic spleen cells on adult spleen cells was present when the embryonic cells were added at the time of or after initiation of the adult spleen mitogenic cultures. When the embryonic cells were added to the cultures of adult spleen cells after the blastogenic response of the adult cells had peaked, the embryonic cells inhibited the incorporation of the label into adult spleen cell blasts. The suppressive activity of the embryonic spleen cells was mediated by soluble suppressor product(s) secreted by these cells, and direct cell-to-cell contact between embryonic and adult spleen cells was not necessary for suppression to occur. Infection of embryos with turkey herpesvirus and Marek's disease virus reduced the suppressor cell activity of embryonic spleen, although substantial residual suppressor cell activity remained in virus-infected embryos. Several pathogenic or non-pathogenic isolates of infectious bursal disease virus did not appreciably alter the suppressor cell activity of embryonic spleen cells. 相似文献
2.
We have previously shown that activation of primary cultures of chicken bone-marrow macrophages and embryo fibroblasts with supernatants of concanavaline A-stimulated or reticuloendotheliosis virus (REV)-transformed chicken spleen cells as source of IFN-gamma significantly decreases Eimeria tenella growth in vitro. In the present study, we used various chicken cell lines, HD11 macrophages and DU24 fibroblasts, both virally transformed, CHCC-OU2 fibroblasts and LMH hepatic epithelial cells, both chemically transformed, to replicate E. tenella in vitro. We confirmed the previous results by showing that HD11 macrophages pre-treated for 24h with recombinant chicken IFN-gamma (either produced in E. coli or by transfected COS cells), at doses ranging from 1000 to 10U/ml, drastically inhibited E. tenella replication as measured by [3H] uracil uptake after a further 70h of culture, as when treated with REV supernatant. Likewise the fibroblast and epithelial cell lines exhibited significant inhibitory activity on E. tenella replication after pre-treatment with recombinant chicken IFN-gamma, but were less sensitive (1000-100U/ml) than when treated with REV supernatant. Recombinant chicken IFN-alpha pre-treatment of all cell lines had no inhibitory effect on parasite development. 相似文献
3.
Continuous passage of MDCC-RP1, a highly tumorigenic Marek's disease (MD) lymphoblastoid cell line, in cell culture resulted in a gradual loss in ability of the cell line to cause progressive tumors in susceptible day-old chicks. Inoculation of day-old chicks with high-cell-culture-passaged (187th to 417th) nontumorigenic MDCC-RP1 cells gave excellent protection against challenge at 8 days with low-passaged tumorigenic MDCC-RP1 cells but failed to protect against primary tumors caused by inoculation with MD virus. Vaccination with the herpesvirus of turkeys, on the other hand, protected the chickens well against primary tumors caused by MD virus and against transplantable tumors caused by tumorigenic MDCC-RP1 cells, but it did not protect as well against another MD lymphoblastoid cell line, MDCC-RP4. It is unlikely, therefore, that vaccines prepared from passaged MDCC-RP1 cell lines will have value for protecting chickens against MD in the field. 相似文献
4.
A soluble substance that blocks B-cell response was extracted from spleen of deer mice (Peromyscus maniculatus) chronically infected with Trypanosoma brucei. Immunosuppressor activity of this substance was demonstrated by plaque-forming cell assays of spleen cells from Swiss/Webster mice inoculated with sheep red blood cells and simultaneously given the suppressor substance, and of spleen cell cultures treated with sheep red blood cells and suppressor substance. Studies by light and electron microscopy of spleen of immunosuppressed Swiss/Webster mice showed that the suppressor substance blocks germinal center formation and prevents plasma cell differentiation in the red pulp cords. 相似文献
5.
Long-term growth of T cell cultures requires addition of Interleukin 2 (IL-2). In order to maintain bovine cultures, optimal conditions for bovine IL-2 production were defined using peripheral blood mononuclear cells (PBM). Irradiation and preculture enhanced IL-2 production possibly by reducing suppressor activity. IL-2 activity was also detected in Bovine Herpesvirus Type 1-stimulated cultures. Unlike mitogen-stimulated cultures, a wide variation in IL-2 activity was seen between supernatants produced by virus-stimulated cells from different animals indicating the clonal nature of antigen specific cells from individuals. Bovine IL-2-dependent cells used to quantitate IL-2 activity were characterized as: PNA, esterase negative, H4+ (anti Ia-like), B29+ (anti-pan T cell), and C5- (anti-monocyte). The observations that bovine IL-2 can maintain activated murine cells, CTLL-20 and HT-2, could lead to the replacement of rat IL-2 with bovine IL-2 in long-term murine cultures. Conditions described here result in large volumes of active medium. 相似文献
6.
To determine whether there is histamine-induced suppressor activity in macrophage-related functions other than in immunity, extracts and media from a macrophage cell line, RAW 264, were tested for suppressor effect on fibroplasia. The procedure consisted of priming confluent RAW 264 cells in culture with media or cellular extracts of washed mastocytes (P-815). The inoculum was removed from the RAW 264 cells by rinsing with fresh medium 24 hours later, and then with medium replacement and 3 more days of culture. The culture media or extracts of washed RAW 264 cells were tested for suppressor activity. The primed RAW 264 cells were lysed by 4 freeze-thaw cycles and cleared by centrifugation, and the resulting supernatant was tested on fibroblast (3T3) cell growth and wound healing in mice and for suppressor activity on T cells. Replication of 3T3 cells, as quantitated by uptake of [3H]thymidine, was reduced 75% when "suppressor" material from RAW 264 cells was added to 3T3 cultures and not when media or extracts of unprimed RAW 264 cells were added. Tensiometric measurements of wound breaking strength (full-thickness incised wounds) were reduced 31% by day 4 and 47% by postsurgical day 7 when "suppressor" RAW 264 extracts were instilled into wounds. Leukocyte cultures stimulated with phytohemagglutinin had a reduced uptake of [3H]thymidine (suppressed 90% to 95%) when exposed to primed RAW 264 extracts, whereas kidney cell culture lines were unaffected. The data obtained indicated that mastocyte (histamine)-induced suppressor factors are present for fibroblast activity as well as T-cell function. 相似文献
7.
J M Sharma 《Avian diseases》1981,25(4):882-893
Chickens of 2 genetic lines (lines P and N) were inoculated with a pathogenic strain of Marek's disease (MD) virus (MDV) and chronologically examined for disease response and natural killer (NK) cell expression. The NK cell reactivity was assayed in an in vitro cytotoxicity assay in which effector cells from the spleen of test chickens were reacted with 51Cr-labeled LSCC-RP9 target cells. Chickens of line P developed progressive debilitating disease and a high incidence of gross tumors and death. The NK cell reactivity of line-P chickens infected with MDV was significantly lower than that of uninfected control hatchmates. In contrast, NK cell levels were significantly elevated in MDV-inoculated line-N chickens that were resistant to MD and in chickens of lines P or N that had been inoculated with herpesvirus of turkeys (HVT). NK cell levels were also elevated in line P if chickens were vaccinated with HVT before infection with MDV. Inhibition of NK reactivity in susceptible chickens and elevation of reactivity in naturally resistant or vaccinated chickens may indicate a role for the NK cell system in regulating resistance to MD. 相似文献
8.
以纯化的酵母重组表达的犬细小病毒VP2单位免疫BALB／C小鼠,采用B淋巴细胞杂交瘤技术,通过ELISA方法筛选,获得4株能稳定分泌抗犬细小病毒CPV结构蛋白VP2的单克隆抗体杂交瘤细胞株。4株单克隆抗体中,2株属于IgG2b亚类,2株属于IgG1亚类,其腹水效价可达到1：51200和1：204800,细胞培养上清液效价可达1：256、1：512。ELISA分析表明,这些单抗仅与CPV及其VP2发生特异性反应,而与CDV和CAV-1及CAV-2没有交叉反应;荧光免疫染色病毒检测进一步表明单克隆抗体的特异性效果好。这些特异性单抗的制备为建立有效的检测犬细小病毒感染奠定了基础。 相似文献
9.
为获得分泌抗大豆凝集素(SBA)单克隆抗体的杂交瘤细胞,以纯化的SBA为抗原,免疫BALB/c小鼠,加强免疫3 d后取小鼠脾细胞与骨髓瘤细胞(SP2/0)融合,应用有限稀释法和间接ELISA 方法克隆筛选出2株分泌抗SBA单克隆抗体的杂交瘤细胞系A7、F12。细胞上清抗体效价均在1∶2×103以上,腹水抗体效价均为1∶1×106,单抗亚型鉴定结果均为IgG2b型,分子质量为189.6 ku,亲和常数为7.1×107 mol/L,Western blotting结果表明,2株单抗具有较高的特异性。该McAb的制备为建立SBA定量检测方法奠定了基础。 相似文献
10.
M A Qureshi L Miller H S Lillehoj M D Ficken 《Veterinary immunology and immunopathology》1990,26(3):237-250
A new chicken mononuclear cell line (MQ-NCSU) has been established. The starting material used to initiate this cell line was a transformed spleen from a female Dekalb XL chicken which had been experimentally challenged with the JM/102W strain of the Marek's disease virus. After homogenization, a single cell suspension of splenic cells was cultured using L.M. Hahn medium supplemented with 10 microM 2-mercaptoethanol. Under these culture conditions, a rapidly proliferating cell was observed and then expanded after performing limiting dilution cultures. These cells were moderately adherent and phagocytic for sheep red blood cells and Salmonella typhimurium. When tested against a panel of monoclonal antibodies (mAb) using the flow cytometry, MQ-NCSU cells stained readily with anti-chicken monocyte specific (K-1) mAb but did not stain with mAb detecting T-helper, T-cytotoxic/suppressor, and NK cells. MQ-NCSU cells expressed very high levels of Ia antigens and transferrin receptors. In addition, cell-free supernatant obtained from MQ-NCSU culture contained a factor which exhibited cytolytic activity against tumor cell targets. Based on their cultural, morphological, and functional characteristics and mAb reactivity profile, we conclude that MQ-NCSU cell line represents a malignantly-transformed cell which shares features characteristic of cells of the mononuclear phagocyte lineage. 相似文献
11.
猪瘟病毒E2基因在真核细胞中表达及抗E2蛋白单克隆抗体的制备 总被引：4，自引：1，他引：4
利用DNA重组技术将猪瘟病毒（CSFV）石门株囊膜蛋白E2基因插入逆转录病毒载体pBABE-puro中构建重组逆转录病毒载体pBABE-puro-E2,该重组逆转录病毒载体与pVSVg质粒经磷酸钙共转染法转入293T细胞中包装逆转录病毒假病毒.用包装的假病毒感染SP2/0细胞,经嘌呤霉素筛选阳性细胞后进行流式细胞术（FACS）分析,结果表明CSFV E2基因在SP2/0细胞膜上成功表达.将表达E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠,成功诱导小鼠产生了抗E2蛋白的抗体.取免疫小鼠脾细胞与SP2/0骨髓瘤细胞融合,经克隆和筛选获得了4株稳定分泌抗猪瘟病毒E2蛋白单克隆抗体的杂交瘤细胞株,所分泌的单抗可与CSFV产生特异性反应并具有中和活性. 相似文献
12.
Cells with the histological and ultrastructural characteristics of large granular lymphocytes (LGL) have been obtained in culture from both cattle and red deer (Cervus elaphus) reacting with 'sheep-associated' malignant catarrhal fever (MCF). Such cells have been derived from thymus, lymph node and spleen suspensions as well as from cerebrospinal fluid cells and cultured cornea. On most occasions their presence was observed only transitorily but by providing the cells with feeder monolayers and, or, interleukin-2, several lines were maintained indefinitely, and some became independent of these factors after prolonged culture. A similar cell line was also derived from a Père David's deer affected with MCF at Whipsnade zoological park. Functionally, cultured LGL were cytotoxic to both primary cell cultures and cell lines and their cytotoxicity was not restricted to histocompatible target cells. These findings suggest that the cultured cells have natural killer cell-like activity and that they are important targets for the agent of MCF in cattle and deer. One cell line derived from a red deer transmitted the disease but none of the cells generated from cattle did. 相似文献
13.
P Quere 《Veterinary immunology and immunopathology》1992,32(1-2):149-164
Marek's disease virus (MDV)-transformed lymphoblastoid cells (MDCC-MSB1, -PA9 and -RP1) added to chicken splenic lymphocytes after treatment with mitomycin, suppress the lymphoproliferative response to T-cell mitogens (concanavalin A or phytohemagglutinin) by 40-70%. This suppressive activity was observed in syngeneic as well as in allogeneic combinations of cell lines and responder lymphocytes. The suppressive effect disappeared when the addition of MD-transformed cell lines to the responder cultures was delayed for 24 h. Treatment with glutaraldehyde, instead of mitomycin, greatly weakened the suppressive activity of the MD lymphoblastoid cells. A reduction of interleukin 2 (IL-2)-like activity produced by responder lymphocytes was observed after mixing with mitomycin-treated lymphoblastoid cells, but also, although slightly less, with the same glutaraldehyde-treated cells. Nevertheless no membrane fluorescence was observed, using INN-CH16 monoclonal antibody on MDV-induced lymphoblastoid cell lines to check up on the presence of IL-2 receptor-like structure. All the three lines exhibited a CD4+, CD8- phenotype. 相似文献
14.
应用提纯的猪流行性腹泻(PED)病毒抗原,建立了1D_8、5D_7、4B_1、2B_8、1A_4 5株抗PED病毒的单克隆抗体杂交瘤细胞林。用ELISA对此5株单抗的培养上清及诱生的腹水测定,培养上清效价为1D_8 1:1280、5D_7 1:640、4B_1 1:320、2B_8 1:320、1A_4 1:640;腹水效价为1D_8 10~(-7)、5D_7 10(-7)、4B_1 10(-7)、2B_8 10(-5)、1A_4 10(-4)_8应用兔抗鼠IgG和IgM标准抗血清做免疫双扩散试验,5株单抗中有1株属IgG类4株属IgM类。将单抗提纯后,应用夹心间接法ELISA及间接免疫荧光试验(IFA)进行特异性鉴定,初步证明,5株单抗除与PED病毒反应外,不与猪传染性胃肠炎病毒(TGEV)、禽传染性支气管炎病毒(IBV)、犬冠状病毒(CCV)、人肠道冠状病毒(HCV)和熊冠状病毒发生交叉反应。取效价高而稳定的1D_8、5D_7、4B_1细胞株分泌的McAb,以间接免疫荧光试验及直接ELISA检测PED病毒,初步证明MCAb在PED的诊断上具有敏感性高、特异性强的优点,可以作为PED病毒检测的一种新试剂。 相似文献
15.
Chicken thymus, spleen, and bursa lymphocytes were isolated by different methods and incubated under differing conditions in order to obtain and characterize avian lymphokines. The biological activity of lymphokine-containing cell culture supernatants was measured by their antiviral activity (interferon(IFN)-units) and by their capacity to induce cytostatic effects in bone-marrow-derived macrophages (50% cytostasis-inducing dose, CID). Lymphokine production by thymus lymphocytes required concanavalin A (ConA)-stimulation, while spleen cells, when cultured at high density, released CID and IFN activities into the culture medium even without mitogen-stimulation. By way of comparison, the highest lymphokine content was found in the supernatant of lymphocyte cultures, which were incubated for 72 hours at 41 degrees C after stimulation with an optimal ConA dose. For stimulation of thymus lymphocytes 30 micrograms ConA/ml were found to be optimal, independent of serum content and cell density in the cultures. In contrast, the optimal ConA dose for spleen lymphocytes not only depended on the serum content but also on the cell density in the cultures and varied within a range of 2.5 micrograms and 45 micrograms ConA/ml. 相似文献
16.
Kabeya H Yamasaki A Ikariya M Negishi R Chomel BB Maruyama S 《Veterinary microbiology》2007,119(2-4):290-296
This study was conducted to analyze cytokine production mechanisms in mice after Bartonella henselae stimulation. BALB/c mice were inoculated intraperitoneally with 3 x 10(6) colony forming units of B. henselae (Houston-1 strain) twice at 10-day interval. Spleen cells were harvested from the mice and stimulated with the organisms. Following the stimulation, interferon-gamma (IFN-gamma) and interleukin-4 (IL-4), IL-10, IL-12 and tumor necrosis factor-alpha (TNF-alpha) were measured in the culture supernatants of the spleen cells by ELISA. The spleen cells specifically secreted IFN-gamma, but not IL-4, indicating that T helper 1 (Th1) cells were activated following B. henselae stimulation. In addition, IL-10 and TNF-alpha productions were also detected in the culture supernatants of spleen cells. Neutralization of IL-10 in the culture supernatants significantly enhanced the production of IFN-gamma from the spleen cells stimulated with B. henselae. These results indicate that B. henselae predominantly stimulated Th1 cells and resulted in secreting IFN-gamma, however the production was partially inhibited by IL-10, which was produced simultaneously. 相似文献
17.
18.
Leukocytes isolated from intraepithelium, lamina propria, and aggregated lymphatic follicles of the small intestine of healthy adult cattle were tested for their ability to produce interleukin 2 (IL-2) by in vitro stimulation of cells with mitogens. Supernatants from interepithelial leukocytes, lamina propria leukocytes, and cultures stimulated with concanavalin A (conA), phytohemagglutinin, and pokeweed mitogen contained growth factors with the capacity to maintain proliferation of a bovine IL-2-dependent lymphoblastoid cell line. Interleukin-2 activity was demonstrated in supernatants of all 3 conA-stimulated leukocyte populations as early as 20 hours after initiation of culture, reached peak values at 30 to 50 hours, and decreased by 72 hours. Although quantitative variations of IL-2 production were observed between various cell types and among cattle, conA was the most potent in inducing IL-2 activity in all 3 leukocyte populations. Supplementation of culture medium with 2-mercaptoethanol or phorbol myristrate acetate neither induced IL-2 production nor enhanced mitogen-induced IL-2 production. Addition of indomethacin to conA-stimulated cultures enhanced IL-2 production. Although depletion of adherent cells did not affect IL-2 production, total elimination of Ia-positive accessory cells inhibited its production by all 3 cell populations. Lymphocytes responsible for IL-2 production in aggregated lymphatic follicle population were presumptive T cells because they were nylon wool-nonadherent, B26A positive (monoclonal antibody directed against pan T cells), pIg45A negative (antibody directed against pan B cells), and considered peanut agglutination-positive. 相似文献
19.
The purpose of this study was to determine whether avian pathogenic Escherichia coli produced cytotoxic activity. Culture supernatants of 20 E. coli strains isolated from cellulitis lesions in chickens, five E. coli strains from avian septicemia, five from swollen head syndrome, and five from the feces of healthy chickens were incubated with primary chicken embryo fibroblast (CEF) cells, primary chicken kidney (PCK) cells, a quail fibroblast cell line (QT-35), and four mammalian cell lines (human epithelioid cervical carcinoma, African green monkey kidney, Chinese hamster ovary, and human larynx epidermoid carcinoma). Cytotoxicity was observed with supernatants from the 30 avian pathogenic strains on the two primary chicken cells (CEF and PCK). The highest dilution of culture supenatant that induced cytotoxic changes in 50% of the cells was 1/64. Supernatants from the five strains from normal feces were noncytotoxic, and none of the supernatants was cytotoxic for the QT-35 or the four mammalian cell lines. The cytotoxic effect, which was observed as early as 2 hr after exposure of the cells, was maximal at 6 hr and was evident as vacuolation, morphologically indistinguishable from that previously reported for culture supernatants of Helicobacter pylori. Like the activity in H. pylori, the cytotoxicity of the avian pathogenic strains was destroyed by heating at 70 C for 30 min and by exposure to proteolytic enzymes and was retained by filtration with a 100,000 molecular weight cut-off ultrafilter. Supernatants of two vacuolating cytotoxin-positive cultures of H. pylori failed to induce vacuolation of the CEF and PCK cells but caused the characteristic vacuolation in HeLa and Vero cells. The observations suggest that avian pathogenic E. coli produce a cytotoxin that is similar to the cytotoxin of H. pylori but may be specific for avian cells. 相似文献
20.
Chicken primary enterocytes: inhibition of Eimeria tenella replication after activation with crude interferon-gamma supernatants 总被引：1，自引：0，他引：1
A reproducible and original method for the preparation of chicken intestine epithelial cells from 18-day-old embryos for long-term culture was obtained by using a mechanical isolation procedure, as opposed to previous isolation methods using relatively high concentrations of trypsin, collagenase, or EDTA. Chicken intestine epithelial cells typically expressed keratin and chicken E-cadherin, in contrast to chicken embryo fibroblasts, and they increased cell surface MHC II after activation with crude IFN-gamma containing supernatants, obtained from chicken spleen cells stimulated with concanavalin A or transformed by reticuloendotheliosis virus. Eimeria tenella was shown to be able to develop until the schizont stage after 46 hr of culture in these chicken intestinal epithelial cells, but it was not able to develop further. However, activation with IFN-gamma containing supernatants resulted in strong inhibition of parasite replication, as shown by incorporation of [3H]uracil. Thus, chicken enterocytes, which are the specific target of Eimeria development in vivo, could be considered as potential local effector cells involved in the protective response against this parasite. 相似文献