首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献，搜索用时 656 毫秒
1.
The biologically active form of vitamine D(3) [1alpha,25(OH)(2)D(3)] has recently been described not only to influence bone metabolism but also to exert immunomodulating activities, which may have an impact on bone formation/resorption as well. In this study, we analysed the effects of 1alpha,25(OH)(2)D(3) on the cytokine pattern of porcine bone marrow-derived cells from piglets aged 1-3 weeks. After culture for 1 week, the number of osteoclasts was determined, with tartrate-resistant acid phosphatase (TRAP)-positive, multinucleated cells being considered osteoclasts. Cultured bone marrow cell-derived mRNA was subjected to semiquantitative RT-PCR specific for a panel of porcine cytokines (IL-1alpha, IL-6, IL-8, IL-10, and TNF-alpha). In addition, an immunofluorescence analysis using anti-porcine mAbs specific for IL-1beta, IL-2, IL-4, IL-6, IL-12, TNF-alpha, and IFN-gamma was performed. In order to prove the existence of a porcine homologue of the receptor activator of NF-kappaB ligand (RANKL) bone marrow cell- as well as porcine white blood cell-derived mRNA was investigated by RT-PCR using primer pairs specific for murine RANKL. Cell culture supernatant was analysed for soluble RANKL by means of an ELISA designed for quantification of human RANKL. By means of RT-PCR, expression of IL-1alpha, IL-6, IL-8, IL-10 and TNF-alpha mRNA could be found in cells cultured with and without 1alpha,25(OH)(2)D(3). Immunofluorescence analysis revealed that IL-1, IL-6, and TNF-alpha were produced by both stromal cells and osteoclasts. Besides its known osteoclastogenic effects, 1alpha,25(OH)(2)D(3) tended to downregulate the respective cytokines, but significantly upregulated RANKL expression. The homology between the porcine RANKL-specific sequence and the corresponding human RANKL sequence was 79%. The data found support the idea that porcine bone marrow cell cultures may provide a suitable alternative to murine systems in human osteological research.  相似文献   

2.
Using an established standardized exercise test on a high-speed treadmill, thirteen Thoroughbred racehorses were exercised to fatigue (failure); blood samples were obtained before exercise, at failure, and at 2, 6, 24, 48, and 72 h after exercise. The exercise test induced a systemic inflammatory response characterized by a mild transient endotoxemia, leukocytosis, increased leukocyte expression of mRNA for tumor necrosis factor-alpha (TNF-alpha), IL-1 beta, and IL-6, and increased circulating concentrations of TNF-alpha and prostaglandin F2 alpha (PGF 2 alpha), with the most pronounced changes being evident at failure and 2h after exercise. Expression of mRNA for IL-6, TNF-alpha, and IL-1 beta was increased by 120-fold, three-fold, and four-fold, respectively, when compared to pre-exercise values. Plasma concentrations of 6-keto-PGF1alpha and PGE2 did not change in response to the exercise test. Collectively, these findings indicate that brief, strenuous exercise induces endotoxemia and a systemic pro-inflammatory response in horses that persists for at least 2h.  相似文献   

3.
The distinct patterns of cytokine expression in CD4+ and CD8+ T cells are well understood in mice and humans. However, little information is available about cytokine expression in bovine CD4+ and CD8+ T cells. In this study, mRNA expression of 19 different cytokines was analyzed in CD4+ and CD8+ T cells of calves with or without Concanavalin A (Con A) stimulation. CD4+ and CD8+ T cell populations were enriched to 98% purity by positive selection using magnetic cell sorting (MACS). CD4+ T cells spontaneously expressed the mRNAs of interleukin-1alpha (IL-1alpha), IL-1beta, IL-2, IL-6, IL-7, IL-8, IL-10, IL-18, IFN-gamma, TNF-alpha, TNF-beta and TGF-beta, and augmented the mRNA expression of IL-10, IFN-gamma and TNF-beta after Con A stimulation. The mRNAs of IL-3, IL-4, IL-5, IL-13 and GM-CSF were newly expressed in Con A-stimulated CD4+ T cells. CD8+ T cells displayed spontaneous mRNA expression of IL-6, IL-18, TNF-alpha, TNF-beta and TGF-beta, and newly expressed the mRNA of IL-2, IL-7, interferon-gamma (IFN-gamma) and GM-CSF after Con A stimulation. It was found that CD4+ T cells expressed the mRNA of 17 cytokines except for IL-12 and IL-15, while CD8+ T cells expressed only the mRNA of 9 cytokines after Con A stimulation. The profile of cytokine mRNA expression was substantially different in the CD4+ and CD8+ T cells of calves, indicating that CD4+ T cells can be distinguished from CD8+ T cells by the cytokine gene expression of IL-1alpha, IL-1beta, IL-3, IL-4, IL-5, IL-8, IL-10 and IL-13. Differential cytokine expression between CD4+ and CD8+ T cells serve to interpret an individual function of T cell subsets in the immune system of calves.  相似文献   

4.
  相似文献   

5.
In situ hybridization (ISH) technique with a biotin-labeled probe was established for detecting feline interleukin 1 (IL-1) alpha mRNA in necropsied specimens. Homology between human IL-1 alpha cDNA used as a probe and feline IL-1 alpha mRNA was confirmed by means of dot blot hybridization using the biotin-labeled probe. Hence, we tried by this biotinylated probe to detect mRNA of IL-1 alpha in paraffin-embedded sections. The following results were obtained for the routine procedures: 1) coating slides with poly-L-lysine and/or heating at 60 degrees C at least for 6 hours gave an excellent result for the adhesion of the tissue sections, 2) 10 micrograms/ml solution of proteinase K treatment for 30 minutes or 50 to 100 micrograms/ml solution of proteinase K treatment for 10 to 30 minutes at 37 degrees C gave the good results in the detection of ISH signal, 3) suitable denaturation time of probes at 70 to 90 degrees C was 5 to 15 minutes, and 4) effective hybridization was obtained by incubation for 24 hours at 4 degrees C, for 18 to 24 hours at 25 degrees C or for 5 to 24 hours at 37 degrees C.  相似文献   

6.
We recently cloned the genes encoding chicken IL-15 and IL-15 receptor (R) alpha proteins. In this study, 12 monoclonal antibodies (mAbs) against recombinant chicken IL-15Ralpha were produced and characterized. By enzyme-linked immunosorbent assay (ELISA), all mAbs showed binding specificity for IL-15Ralpha, but not IL-2 or interferon-gamma, and identified a 25.0kDa protein by immunoblot analysis. Flow cytometric analysis revealed negligible expression of IL-15Ralpha on non-activated lymphocytes from the spleen, thymus or bursa, low but detectable expression on macrophages and high expression on concanavalin A-activated spleen lymphoblasts. Established chicken T cell (RP13) and macrophage (HD11) cell lines expressed substantially higher levels of IL-15Ralpha compared with a B cell line (RP9). Two mAbs inhibited IL-15 dependent proliferation of T cells suggesting that the tertiary structure of the protein domain of native IL-15Ralpha that binds to IL-15 is preserved in the recombinant receptor molecule. These mAbs will be useful reagents for further in vitro and in vivo studies of the biological functions of chicken IL-15 and its receptor.  相似文献   

7.
Mycobacterium avium subspecies paratuberculosis (MAP) is a facultative intracellular pathogen that resides in host macrophage cells. Presently, little is known about how MAP is able to subvert the normal bacteriocidal functions of infected macrophages. Previously, we reported that ileal tissues from MAP infected cattle contained high levels of interleukin-1 alpha (IL-1alpha) and tumor necrosis factor receptor-associated factor 1 (TRAF1), relative to ileal tissues from uninfected cattle. High-level expression of these two proteins could have profound effects on macrophage function, intracellular signaling, and apoptosis. We now demonstrate that high levels of TRAF1 protein are located primarily within macrophages infiltrating areas of MAP infection. We have also utilized cultured bovine monocyte-derived macrophage cells (MDM) either infected with live MAP or stimulated with recombinant IL-1alpha (rIL-1alpha) to determine if there is a relationship between IL-1alpha and TRAF1 expression. These studies have identified a dose dependent increase in TRAF1 protein levels in bovine MDM in response to infection with live MAP or following treatment with rIL-1alpha. Sustained TRAF1 protein expression was dependent upon interaction of rIL-1alpha with it's receptor and rIL-1beta was also able to enhance TRAF1 gene expression. Our results suggest that MAP may use the IL-1-TRAF1 system to enhance TRAF1 protein expression in infected bovine MDM. These novel results provide evidence for a new avenue of research on the effect of MAP and other intracellular pathogens on macrophage signaling and apoptosis.  相似文献   

8.
Immune responses to gastrointestinal helminth infections have received increasing attention due to similarities to allergen-induced responses. In fact, the whipworm parasite of swine, Trichuris suis, has been used in beginning clinical trials as an antidote to inflammatory bowel disease. This strategy was based on this similarity and the recognition that other worms have been documented to induce anti-inflammatory responses in the host. In an effort to understand the basis for this response, we hypothesized that the proteins and peptides secreted by T. suis stimulate local intestinal epithelial cells to produce anti-inflammatory cytokines. To test this hypothesis in a correlate system of the natural swine host, T. suis excretory secretory products (ESP) were used to treat both differentiated and undifferentiated intestinal pig epithelial cells (IPEC-1) in vitro as a model for the effect on villus tip and crypt epithelial cells in the vicinity of the worms. IPEC-1 were exposed to low-level doses (0.3mg/ml) of T. suis ESP, and IL-4, IL-6 and IL-10 cytokine responses were measured by an enzyme-linked immunosorbant assay (ELISA). IL-6 was the predominant cytokine produced, accompanied by moderate IL-10 secretion from both differentiated and undifferentiated cells. As expected, IL-4 was not produced by IPEC-1. Additionally, IL-6 and IL-10 cytokines were produced within 24h, suggesting that these two cytokines form part of the primary host response to T. suis infections. These data suggest that T. suis ESP could enhance host immune responses and modulation through the induction of enteric IL-6 and IL-10.  相似文献   

9.
  相似文献   

10.
OBJECTIVES: To evaluate the effects of equine recombinant interleukin-1alpha (rEqIL-1alpha) and recombinant interleukin-1beta (rEqIL-1beta) on proteoglycan metabolism and prostaglandin E2 (PGE2) synthesis by equine articular chondrocytes in explant culture. SAMPLE POPULATION: Near full-thickness articular cartilage explants (approx 50 mg) harvested from stifle joints of a 3-year-old and a 5-year-old horse. PROCEDURE: Expression constructs containing cDNA sequences encoding EqIL-1alpha and EqIL-1beta were generated, prokaryotically expressed, and the recombinant protein purified. Near full-thickness articular cartilage explants (approx 50 mg) harvested from stifle joints of a 3-year-old and a 5-year-old horse were separately randomized to receive rEqIL-1alpha or rEqIL-1beta treatments 10 to 500 ng/ml). Proteoglycan release was evaluated by 1,9-dimethylmethylene blue spectrophotometric analysis of explant media glycosaminoglycan (GAG) concentration and release of 35S-sulfate-labeled GAG to explant media. Proteoglycan synthesis was assessed by quantification of 35S-sulfate incorporation into proteoglycan. Explant media PGE2 concentrations were evaluated using a PGE2-specific enzyme-linked immunoassay. Data were collected at 48-hour intervals and normalized by DNA content. RESULTS: Proteoglycan release was induced by rEqIL-1alpha and rEqIL-1beta at concentrations > or =0.1 ng/ml, with 38 to 76% and 88 to 98% of total GAG released by 4 and 6 days, respectively. Inhibition of proteoglycan synthesis (42 to 64%) was observed at IL-1 concentrations > or = 0.1 ng/ml at 2 and 4 days. Increased PGE2 concentrations were observed at IL-1 concentrations > or = 0.1 ng/ml at 2 and 4 days. CONCLUSIONS AND CLINICAL RELEVANCE: The rEqIL-1 induced potent concentration-dependent derangement of equine chondrocyte metabolism in vitro. These findings suggest this model may be suitable for the in vitro study of the pathogenesis and treatment of joint disease in horses.  相似文献   

11.
This communication reports the cloning of cDNAs encoding two canine IL-13 receptor alpha chains (caIL-13Ralpha1 and caIL-13Ralpha2). As described for the members of type-I cytokine receptors, both caIL-13Ralpha1 and caIL-13Ralpha2 were found to contain the highly conserved motifs, such as cysteine and tryptophan residues in their N-terminal portion and the WSXWS at C-terminus. The isolated caIL-13Ralpha1 cDNA contains 1547 nucleotides with an open reading frame that encodes 405 amino acid residues. Canine IL-13Ralpha1 is 82.0 and 69.3% identical to human and mouse IL-13Ralpha1s, respectively, at the amino acid level. Canine IL-13Ralpha1 has an almost identical cytoplasmic domain to its human and mouse counterparts. The isolated caIL-13Ralpha2 cDNA contains 1454 nucleotides and encodes an open reading frame of 386 amino acid residues. Canine IL-13Ralpha2 is 62.6 and 47.5% identical to its human and mouse counterparts, respectively, at the amino acid level. Using RT-PCR with caIL-13Ralpha1 and caIL-13Ralpha2 specific primers, mRNAs of caIL-13Ralpha1 and caIL-13Ralpha2 were detected in most dog tissues. In addition, RT-PCR detected caIL-13Ralpha1 mRNA in one of two canine mastocytoma (C2 but not Br) cell lines and in a canine macrophage-derived cell line (DH82). CaIL-13Ralpha2 mRNA was detected in all three canine cell lines.  相似文献   

12.
Inflammatory cell activation by interleukin-1 (IL-1) has been the subject of studies in laboratory animals and man, and was the aim of the present study. At physiological concentrations of 0.01-100 units/ml, human recombinant IL-1 alpha was shown to be a chemokinetic and chemotactic agent for canine neutrophils in vitro and to cause dose-dependent and selective neutrophil infiltration following intradermal injection of 1 or 250 units. The findings support a putative role for IL-1, and IL-1-induced products such as IL-8, as mediator of immunoinflammatory reactions in which neutrophil infiltration is prominent.  相似文献   

13.
The chemotactic activity of zymosan-activated serum (ZAS) and of two concentrations of recombinant human IL-8 (IL-8(25), 25 ng/ml; IL-8(50), 50 ng/ml) for ovine polymorphonuclear granulocytes (PMNs) was tested in a modified Boyden chamber. Thick cellulose acetate filters and the leading front method were used to quantify the movements of the cells. Both ZAS and IL-8(25) exerted a chemotactic effect on ovine PMNs (P < 0.01): IL-8(50) induced a more homogeneous response (P < 0.001). To verify the characteristics of the responsiveness to the chemokines after short-term (st) or long-term (lt) repeated samplings, chemotaxis was investigated 1 (T1st), 2 (T2st), 24 (T3st) and 48 h (T4st) after the basal sampling (T0st) and 15 days (T1lt) after the basal sampling (T0lt). No differences in chemotaxis were found in long-term repeated samplings. In contrast an increase in the responsiveness to IL-8(25) and to IL-8(50) (P < 0.05) was detected at T2st in comparison with T0st. Furthermore, the significance of the distance run by activated PMNs compared with the controls, increased from T0st to T2st, as a sign of a more homogeneous response to the chemokines. In the absence of evident changes in circulating leucocyte numbers and in serum cortisol concentrations, these findings could be interpreted as a consequence of a different expression of chemoattractant receptors on the membrane of PMNs collected at different times.  相似文献   

14.
Attachment and migration of bovine chondrocytes cultured in vitro were significantly suppressed by the addition of interleukin (IL)-1alpha at the concentration of 1 ng/ml or more (p<0.05). The application of hyaluronic acid (HA) at the concentration of 10 micro g/m l or more significantly recovered the attachment of chondrocytes (p<0.05) and the application of HA at 100 micro g/ml concentration recovered the migration of chondrocytes suppressed by IL-1alpha. These results suggest that the application of HA for inflammatory arthropathies or chondrocyte transplantation might be helpful to preserve the properties of chondrocytes and its extracellular matrix against inflammatory conditions.  相似文献   

15.
The canine cytokine IL-31 induces pruritus in dogs and can be detected in dogs with atopic dermatitis; however very little is understood around its interactions with specific canine cells. We hypothesize that IL-31 is involved in the progression of allergic skin disease by coordinating the interaction between the immune system with skin and neuronal systems. The goal of the following work was to identify cells that produce IL-31 as well as cells that may respond to this cytokine. Peripheral blood mononuclear cells (PBMCs) were collected from naïve and house dust mite (HDM) allergen-sensitized beagle dogs and used for ex vivo characterization of cytokine production assessed using ELISpot and quantitative immunoassay. Sensitization to HDM allergen induced a T-helper type 2 (Th2) cell phenotype characterized by an increase in the production of IL-4 protein. Interestingly, repeated allergen challenge over time also resulted in an increase in IFN-γ. Further evaluation showed that co-stimulation of Th2 polarized cells with antigen and the bacterial component Staphylococcus enterotoxin B (SEB) produced higher levels of IL-31 compared to either stimulant alone. Production of IL-31 when PBMCs were stimulated by T cell mitogens suggests T cells as a source of IL-31. Quantitative real-time PCR was utilized to determine expression of the IL-31 receptor alpha chain in canine cell lines and tissue. Canine monocytic cells, keratinocytes, and dorsal root ganglia were shown to express the IL-31 receptor alpha chain mRNA. In a multifaceted disease such as canine atopic dermatitis, the combination of Th2 polarization and microbial presence may lead to IL-31 mediated effects driving inflammation and pruritus by immune cells, keratinocytes, and direct neuronal stimulation.  相似文献   

16.
  相似文献   

17.
By in situ hybridization with biotinylated human interleukin 1 alpha (IL-1 alpha) cDNA probe, distribution of feline IL-1 alpha mRNA-expressing cells was examined in the tissues from 49 cases diagnosed as feline infectious peritonitis (FIP) by pathological examination. IL-1 alpha mRNA-expressing cells were found in visceral peritoneum, lymphoid organs, liver, kidney, pancreas, digestive tract, lung, pleura, brain, palpebral conjunctiva, and bone marrow. Hybridization signals for IL-1 alpha mRNA were mostly located in the local inflammatory lesions on the serosal surface of various organs and omentum, which were frequently involved in the lesions of FIP (27.8 +/- 5.1 cells/mm2). Morphological examination suggested that they were infiltrated macrophages. However, few IL-1 alpha mRNA-expressing macrophages were in the lesions of other organs. These data suggested that IL-1 alpha produced from macrophages in the local inflammatory sites might participate in the initiation and development of the lesions on the visceral peritoneum in FIP.  相似文献   

18.
  相似文献   

19.
This paper describes the cloning and sequence analysis of the cDNAs encoding the T helper (Th) 2 cytokines of llama including interleukin-4 (IL-4), IL-10 and IL-13. The cDNAs encoding for IL-4, IL-10 and IL-13 were amplified using specific primers designed from reported sequences of bovine cytokine genes. The cDNAs for llama IL-4, IL-10 and IL-13 were found to be 402, 537 and 411 bp in length, with open reading frames encoding 133, 178 or 136 amino acids, respectively. Homology analyses of nucleotide and deduced amino acid sequences of llama IL-4, IL-10 and IL-13 and phylogenetic analysis based on their nucleotide sequences indicated the close relationship in these cytokine genes between llama and eutherian mammalian order Artiodactyla (pig, cattle) and Perissodactyla (horse).  相似文献   

20.
In order to apply for reducing graft versus host disease in allogeneic stem cell transplantation, the study concerning the induction of specific T cell anergy was designed. Normal allogeneic lymphocytes, which were co-cultured with IL-10-treated immature dendritic cells in the first mixed leukocyte culture (MLC), were cultured with mature dendritic cells of the same origin as IL-10-treated immature dendritic cells in the second MLC. By co-culturing with IL-10-treated immature dendritic cells, the response of normal lymphocytes to mature dendritic cells cultured from the same individual as that of IL-10-treated dendritic cells was markedly reduced, compared with the lymphocytes cultured with non-treated dendritic cells or IL-10-treated dendritic cells from a third party individual. The present study demonstrated that antigen specific T cell anergy was generated by priming allogeneic lymphocytes with IL-10-treated immature dendritic cells. These data suggested the applicability of IL-10-treated recipient dendritic cells for the induction of recipient cell-specific donor T cell anergy in donor graft.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号