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1.
Using RT-PCR and semi-quantitative PCR, mRNA expression for canine interferon (IFN)-gamma, interleukin (IL)-2, IL-4, IL-5, IL-10, tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta in peripheral blood mononuclear cells (PBMCs) was examined in dogs with or without demodicosis. mRNA expression for IFN-gamma as well as TNF-alpha in dogs with demodicosis (localized (LD) and generalized (GD)) was slightly lower than those in dogs without demodicosis (healthy controls). Expression of IL-5 mRNA in dogs with demodicosis was higher than that in control dogs, but there were no significant differences in IL-4 and IL-10 mRNA expression levels among the three groups. On the other hand, expression levels of TGF-beta mRNA in dogs with GD were higher than those in control dogs and dogs with LD. The expression levels of IL-5 and TGF-beta mRNA decreased in all three dogs with GD which showed resolution of the clinical signs. Taken together, these results suggest that the Th2-like response in PBMCs from dogs with demodicosis is up-regulated, and that subsequent increased expression of IL-5 and TGF-beta mRNA in dogs with GD is reversible after treatment. Therefore, these cytokines, particularly IL-5, might be a useful clinical index of the clinical course in demodicosis. Also, increased TGF-beta mRNA expression might be a key factor for revealing the difference in the mechanism of onset between LD and GD.  相似文献   

2.
The immune responses of hypothyroid dogs to canine thyroglobulin (cTg) were evaluated for the proliferative ability of peripheral blood mononuclear cells (PBMC). PBMC from three hypothyroid dogs with high titers of thyroglobulin autoantibody (TgAA) and 3 clinically normal dogs were cultured with 5, 10, or 20 microg/ml of cTg for 72 hr. The proliferative responses of the cells were determined by the level of incorporated BrdU. The numbers of cells expressing Thy-1, CD4, CD8 and IgG in the PBMC were counted by the immunofluorescence method. Proliferative responses to cTg were observed in the cells from hypothyroid dogs. The number of cells expressing IgG and CD8 in the hypothyroid dogs tended to be high compared with the clinically normal dogs. The CD4+ cells in cultures from hypothyroid dogs increased depending upon the amount of cTg. There was a significant (P<0.05) positive correlation between the number of CD4+ cells and the concentration of cTg in the cultures from hypothyroid dogs. These findings suggest a possible relationship between canine hypothyroidism and cellular immunity. Loss of self tolerance to thyroid antigens in CD4+ T cells may play an important role in the development of canine hypothyroidism.  相似文献   

3.
Human IL-13, like IL-4, is involved in the regulation of B-cell development, IgE synthesis and allergic responses. However, because IL-13 does not affect either murine Ig class switching or IgE production in vitro, the use of murine models to study the role of IL-13 in IgE-mediated diseases has been limited. In this communication, we report that recombinant protein of canine IL-13 (rcaIL-13) stimulates production of allergen-specific-IgE in vitro by peripheral blood mononuclear cells (PBMC) from flea allergen-sensitized dogs, and that this stimulation activity is specifically inhibited by recombinant protein of canine IL-13Ralpha2 and Fc fragment of canine IgG heavy chain (rcaIL-13Ralpha2-Fc). The data suggest that the regulatory effects of IL-13 on IgE production in canine PBMC are similar to those reported in humans. Thus, canine IL-13 may be a central mediator of allergic diseases in dogs, and allergic dogs may be excellent models for research on IgE-mediated diseases in humans.  相似文献   

4.
Dendritic cells (DCs) are the most potent antigen-presenting cells that are expected to be therapeutic agents for tumor immunotherapy. In this study, we generated DCs of sufficient number for DC-based immunotherapy from peripheral blood mononuclear cells (PBMC) in dogs. PBMC were cultured in the presence of phytohemagglutinin (PHA). On day 6, large adherent cells with dendrite-like projections were seen, and the number of these large cells with projections increased on day 8. These cells were positive for esterase staining. They expressed MHC class II, CD11b, CD8 and weakly CD4 on their surface. They tended to make contact with lymphocytes under culture conditions. We obtained about 2-5 x 10(6) of DCs from 10 ml of peripheral blood. These DCs phagocytosed HEK-293 cells by overnight co-culturing. These cells generated from PBMC are possible canine DCs and are applicable to clinical trials of DC-based whole tumor cell immunotherapy in dogs.  相似文献   

5.
In this study, we summarized the major phenotypic/functional aspects of circulating leukocytes following canine immunization with Leishvaccine and Leishmune?. Our findings showed that Leishvaccine triggered early changes in the innate immunity (neutrophils and eosinophils) with late alterations on monocytes. Conversely, Leishmune(?) induced early phenotypic changes in both, neutrophils and monocytes. Moreover, Leishvaccine triggered mixed activation-related phenotypic changes on T-cells (CD4+ and CD8+ and B-lymphocytes, whereas Leishmune(?) promoted a selective response, mainly associated with CD8+ T-cell activation. Mixed cytokine profile (IFN-γ/IL-4) was observed in Leishvaccine immunized dogs whereas a selective pro-inflammatory pattern (IFN-γ/NO) was induced by Leishmune? vaccination. The distinct immunological profile triggered by Leishvaccine and Leishmune? may be a direct consequence of the distinct biochemical composition of these immunobiological, i.e. complex versus purified Leishmania antigen along with Bacillus Calmette-Guérin (BCG) versus saponin adjuvant. Both immunobiologicals are able to activate phagocytes and CD8+ T-cells and therefore could be considered as a putative vaccines against canine visceral leishmaniasis (CVL).  相似文献   

6.
Interleukin-12 (IL-12) plays a pivotal role in regulating cellular immune responses involving autoimmunity, infectious disease, and cancer. Human recombinant (hr) IL-12 is being evaluated for therapy of human cancer. We investigated the potential of hrIL-12 to activate canine peripheral blood mononuclear cells (PBMC) using proliferation and cytotoxicity as readouts. Human rIL-12 caused increased proliferation of PBMC, and enhanced lysis of allogeneic canine tumor targets mediated by PBMC from normal dogs in vitro. In addition, antibody-dependent cellular cytotoxicity (ADCC) mediated by canine PBMC was enhanced by hrIL-12. These results indicate that hrIL-12 is recognized by canine immune cells, triggering a number of immune responses in canine PBMC, that may be important for immunotherapy of canine cancer. Information from this investigation provides impetus for evaluation of the effects of hrIL-12 on PBMC from tumor-bearing dogs and should be helpful in the development of hrIL-12 as an immune cell activator in vivo in the dog.  相似文献   

7.
REASONS FOR PERFORMING STUDY: An assay has been developed that measures EHV-1 specific interferon gamma synthesis (IFNgamma), a cytokine produced following the activation of memory T lymphocytes and therefore a measure of cell mediated immunity. The method requires validation in the field. OBJECTIVES: To measure the frequency of EHV-1 specific, IFNgamma synthesising peripheral blood mononuclear cells (PBMC) in a population of Thoroughbred horses, and examine its relationship with age, gender, premises and history of vaccination or field infection with EHV-1. METHODS: Lymphocytes from 200 Thoroughbred horses were stimulated with EHV-1 in vitro, and IFNgamma detected using a monoclonal antibody and indirect immunofluorescence. Percent positive cells were enumerated by flow cytometric analysis and the results described and compared statistically between groups. RESULTS: The frequency of IFNgamma+ PBMC was significantly higher in animals age >5 years compared with 2-4 years, in females vs. males, on stud farms vs. training yards and following vaccination of 2-year-olds with inactivated virus compared with nonvaccinates. Age strongly confounded all these associations and care must therefore be taken interpreting these results. Mares exposed to a field infection with EHV-1 also had higher frequencies of IFNgamma+ PBMC than other vaccinated horses. CONCLUSIONS: The frequency of EHV-1 specific, IFNgama+ PBMC among the sample Thoroughbred population was diverse but lowest in young, unvaccinated horses-in-training. POTENTIAL RELEVANCE: The frequency of EHV-1 specific lymphocytes synthesising IFNgamma in this population may be associated with its susceptibility to infection with this virus. This easy technique may be applied to monitor the antigenicity of vaccines and their effectiveness at stimulating cellular immunity.  相似文献   

8.
The effects of salivary gland extract (SGE) from R. sanguineus were examined on the production of IgG1 and IgG2 and the mRNA expression of IFN-gamma, IL-2, IL-4, IL-5 and IL-10 in the mononuclear cells from canine peripheral blood, treated with concanavalin A (ConA) in vitro. SGE suppressed the ConA-induced production of IgG2. It also inhibited the expression of IFN-gamma, IL-2 and IL-5 mRNA in a dose-dependent manner. No dose-dependent suppression was observed of IL-10 mRNA expression although a significant effect was observed at a SGE protein concentration of 25 microg/ml. SGE had no effect on the mRNA expression of IL-4. These results suggest that the suppression of IgG2 production by SGE from R. sanguineus was caused by the suppression of IFN-gamma production.  相似文献   

9.
10.
Serum Cystatin C (sCys-C) is one of the most important serum markers of renal function assessment in dogs. The purpose of this study was to determine the sCys-C concentration in dogs with visceral leishmaniasis (VL). In the study, 16 dogs with VL and 10 clinical healty dogs (control) were used. Mean sCys-C concentration of the infected dogs was significantly higher than that of the control group (p < 0.05). Mean serum creatinine concentration was lower and mean blood urea nitrogen, albumin and globulin concentrations were higher in dogs with VL; however, these changes were not statistically significant. Mean total protein and phosphorus concentrations were found to be higher in dogs with VL than healthy dogs (p < 0.05). No significant correlation had been determined between sCys-C and other variables. Visceral leishmaniasis in dogs has increased sCys-C concentration indicating a possible renal impairment; however, further studies are needed to be performed together with renal biopsies in the investigation sCys-C in dogs with VL.  相似文献   

11.
High Mobility Group Box 1-Protein (HMGB1) is a nuclear chromosomal protein occurring ubiquitary in mammalian tissues. HMGB1 demonstrates cytokine function and induces inflammation when actively released by haematopoietic cells or passively released during cell necrosis. This study aimed at the determination of HMGB1 expression in different cell types and at the evaluation of the role of HMGB1 in PBMC proliferation. Therefore we investigated the HMGB1 mRNA expression level in different canine haematopoietic cell types and the influence of exogenous rhHMGB1 on canine PBMC proliferation. Differentiated haematopoietic blood cells showed lower relative HMGB1 expression levels compared to CD34+ haematopoietic stem cells. Relative HMGB1 expression seemed also to decrease during differentiation of CD34+ stem cells into dendritic cells. Furthermore, peripheral blood CD14+ monocytes and granulocytes showed a lower relative HMGB1 expression in comparison to CD3+ T-lymphocytes. When exogenous rhHMGB1 at low concentrations was added to single PBMC cultures an increase of proliferation was obvious. However, in higher concentrations HMGB1 lost its stimulative effect. In conclusion, HMGB1 is broadly expressed in canine haematopoietic cells with highest levels in haematopoietic stem cells. HMGB1 induced directly PBMC proliferation.  相似文献   

12.
13.
Genetic variation in concanavalin A (Con A)-induced proliferation and interleukin 2 (IL-2) production by peripheral blood mononuclear cells (PBMC) was studied in blood collected from 96 piglets, aged 7 weeks. The piglets were the offspring of seven sires and 24 dams. Pronounced differences between litters from various dams were observed in the immune parameters measured. Also, large individual differences in the magnitudes of Con A-induced proliferation and IL-2 production were seen for PBMC collected from individual pigs within each litter. Both the time course and magnitude of IL-2 activity showed genetic variation, as results from the offspring of the seven sires differed significantly. However, only the time course, not the magnitude, of proliferation differed among the offspring groups. It was possible to establish a rank order for the sires based on the IL-2 production of PBMC by their offspring. As IL-2 has a key role in regulating the immune response, mitogen-induced IL-2 activity seems to be a good candidate as a general marker for cell-mediated immunity in pigs.  相似文献   

14.
Interleukin 4 (IL-4) is expected to play a dominant role in the development of T helper (Th) 2 cells. Th2 immune responses with expression of relatively large amounts of interleukin 4 (IL-4) but little interferon gamma (IFN-gamma) are characteristic for chronic helminth infections. But no information is available about IL4 expression during early Fasciola hepatica (F. hepatica) infections in cattle. Therefore, we investigated F. hepatica specific IL-4 and IFN-gamma mRNA expression in peripheral blood mononuclear cells (PBMCs) from calves experimentally infected with F. hepatica. Cells were collected prior to infection and on post-inoculation days (PIDs) 10, 28 and 70. Interestingly, PBMCs responded to stimulation with F. hepatica secretory-excretory products (FhSEP) already on PID 10 and expressed high amounts of IL-4 but not of IFN-gamma mRNA suggesting that F. hepatica induced a Th2 biased early immune response which was not restricted to the site of infection. Later in infection IL-4 mRNA expression decreased whereas IFN-gamma mRNA expression increased slightly. Isolated lymph node cells (LNCs) stimulated with FhSEP and, even more importantly, non-stimulated LN tissue samples indicated highly polarized Th2 type immune responses in the draining (hepatic) lymph node, but not in the retropharyngeal lymph node. During preliminary experiments, two splice variants of bovine IL-4 mRNA, boIL-4delta2 and boIL-4delta3, were detected. Since a human IL-4delta2 was assumed to act as competitive inhibitor of IL-4, it was important to know whether expression of these splice variants of bovine IL-4 have a regulatory function during an immune response to infection with F. hepatica. Indeed, IL-4 splice variants could be detected in a number of samples, but quantitative analysis did not yield any clue to their function. Therefore, the significance of bovine IL-4 splice variants remains to be determined.  相似文献   

15.
Oligodeoxynucleotides containing the cytosine-phosphate-guanine (CpG) motif (CpG-ODNs) have been shown to induce T(H)1 immune responses in animals. Since the sequences of CpG-ODNs that induce T(H)1 responses are considered to vary among animal species, it is necessary to identify effective CpG-ODNs in each animal. In order to identify the sequences of CpG-ODNs that induce T(H)1 responses in dogs, mRNA expression and protein production of IFN-gamma were examined in peripheral blood mononuclear cells (PBMCs) from healthy dogs treated with 11 kinds of synthetic CpG-ODNs. One of the 11 CpG-ODNs (No. 2 CpG-ODN, 5'-GGTGCATCGATGCAGGGGGG-3') was shown to significantly increase mRNA expression and protein production of IFN-gamma in canine PBMCs in a manner dependent on the sequence of the CpG motif. This CpG-ODN also enhanced the expression of IL-12 p40 mRNA in canine PBMCs, whereas expression of IL-12 p35, IL-18, and IL-4 mRNAs was not induced by this CpG-ODN. These results indicate that this CpG-ODN was able to produce IFN-gamma by induction of T(H)1-skewed immune response in dogs. CpG-ODNs may be useful for inducing prophylactic and therapeutic immunity against allergic diseases, viral infection, and tumors in dogs.  相似文献   

16.
The incidence of canine visceral leishmaniasis (CVL) is increasing in the Mediterranean region. Many drugs have been tested for treatment of CVL, but little is known regarding their effect on test immune responses. In our study, three dogs naturally infected with Leishmania infantum and five dogs experimentally infected with the same strain, were treated with dimethasulfonate pentamidine (Lomidine) and the immune response evaluated before, and after, treatment. After the last injection, animals began to gain weight and the major clinical signs disappeared. Antibody titers gradually decreased to low levels, six months after treatment. At the same time, antigen specific lymphoproliferation reappeared in the sampled animals. This study shows that, after treatment, immune cellular responses to leishmanial antigens, involved in protection against Leishmania infection, were established.  相似文献   

17.
18.
The effects of chlorpromazine (CPZ), pentoxifylline (PTX) and dexamethasone (DEX) on mRNA expression of lipopolysaccharide (LPS)-induced proinflammatory cytokines were examined in bovine peripheral blood mononuclear cells (PBMCs) in vitro. The expression of inflammatory cytokine mRNAs was analyzed by RT-PCR and Southern blot hybridization in bovine PBMCs. CPZ and DEX decreased the expression of cytokine mRNA (such as interleukin-1 beta and tumor necrosis factor-alpha) after stimulation with LPS in a dose-dependent manner. However, pretreatment with PTX had no inhibitory effect on the mRNA expression of proinflammatory cytokines. These results indicated that pretreatment with CPZ and DEX might be effective to reduce the production of LPS-induced inflammatory cytokines in bovine PBMCs in vitro.  相似文献   

19.
Programmed cell death (PCD), or apoptosis, is initiated in response to various stimuli, including virus infection. A number of studies have shown that deregulation of apoptosis is an important feature of virus-induced immunosuppression for various viral diseases. In the present study, CapHV-1 was found to cause apoptosis in mitogen-stimulated as well as nonstimulated caprine peripheral blood mononuclear cells (PBMC). Apoptotic index, as quantified by fluorescent dyes, revealed a significant increase in the percentage of apoptotic cells at 24 and 48 h postinfection as compared to their respective noninfected controls. Apoptosis specific internucleosomal laddering in DNA from CapHV-1 infected PBMC was seen in agarose gel electrophoresis. No DNA fragmentation was observed in control noninfected PBMC. Virus-induced apoptosis was reduced by Z-VAD-FMK, an aspecific caspase inhibitor, by AC-DEVD-CHO (caspase-3-specific) and AC-VEID-CHO (caspase-6-specific) treatment. PCD in CapHV-1 infected peripheral blood mononuclear cells occurs at the G0/G1 phase of the cell cycle. However, penetration of virus particles and infection was not required for PCD, as UV-inactivated CapHV-1 induced apoptosis of mitogen-stimulated bovine peripheral blood mononuclear cells in vitro.  相似文献   

20.
为了解牛病毒性腹泻病毒(BVDV)感染对干扰素(IFN)mRNA转录时相的影响,探讨宿主-病毒之间的相互关系,用非致细胞病变(noncytopathic,NCP)和致细胞病变(cytopathic,CP)型BVDV感染临床健康BVDV检测阴性的荷斯坦奶牛外周血单核细胞(PBMC),利用实时荧光定量PCR技术对感染后IFN-α、β、γmRNA转录水平的变化进行定量分析。结果表明,CP型和NCP型BVDV感染PBMC后,Ⅰ型IFN(IFN-α、β)均呈现出不同程度的转录水平上调,且差异极显著(P〈0.01);只有IFN-α在CP型BVDV感染后4,12h(P〈0.5)出现转录下调。IFN-γ在整个感染过程中均呈现出不同程度的转录水平上调,且差异显著(P〈0.05)。这表明2种生物型BVDV感染可引起PBMC中IFN mRNA转录水平升高。  相似文献   

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