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1.
Leukotoxin (Lkt) is the primary virulence factor secreted by Mannheimia haemolytica which causes pneumonia in ruminants. Previously, we have shown that CD18, the beta subunit of beta(2) integrins, mediates Lkt-induced cytolysis of ruminant leukocytes. CD18 associates with four distinct alpha subunits giving rise to four beta(2) integrins, CD11a/CD18 (LFA-1), CD11b/CD18 (Mac-1), CD11c/CD18 (CR4), and CD11d/CD18. It is not known whether all the beta(2) integrins serve as a receptor for Lkt. Since PMNs are the leukocyte subset that is most susceptible to Lkt, and Mac-1 expression on PMNs exceeds that of other beta(2) integrins, it is of interest to determine whether Mac-1 serves as a receptor for Lkt which necessitates the cloning of CD11b and CD18. In this study, we cloned and sequenced the cDNA encoding CD11b of Ovis canadensis (bighorn sheep) and Ovis aries (domestic sheep). CD11b cDNA is 3455 nucleotides long encoding a polypeptide of 1152 amino acids. CD11b polypeptides from these two species exhibit 99% identity with each other, and 92% with that of cattle, and 70-80% with that of the non-ruminants analyzed.  相似文献   

2.
Mannheimia haemolytica is an important etiological agent of pneumonia in domestic sheep (DS, Ovis aries). Leukotoxin (Lkt) produced by this organism is the principal virulence factor responsible for the acute inflammation and lung injury characteristic of M. haemolytica caused disease. Previously, we have shown that the leukocyte-specific integrins, beta(2) integrins, serve as the receptor for Lkt. Although it is certain that CD18, the beta subunit of beta(2) integrins, mediates Lkt-induced cytolysis of leukocytes, it is not clear whether CD18 of all three beta(2) integrins, LFA-1, Mac-1 and CR4, mediates Lkt-induced cytolysis of DS leukocytes. Since polymorphonuclear leukocytes, which express all three beta(2) integrins, are the leukocyte subset that is most susceptible to Lkt, we hypothesized that all three beta(2) integrins serve as the receptor for Lkt. The objective of this study was to determine whether DS LFA-1 serves as a receptor for M. haemolytica Lkt. We cloned the cDNA for DS CD11a, the alpha subunit of LFA-1, and co-transfected it along with the previously cloned cDNA for DS CD18, into a Lkt-non-suceptible cell line. Transfectants stably expressing DS LFA-1 were bound by Lkt. More importantly, Lkt lysed the DS LFA-1 transfectants in a concentration-dependent manner. Pre-incubation of Lkt with a Lkt-neutralizing monoclonal antibody (MAb), or pre-incubation of transfectants with MAbs specific for DS CD11a or CD18, inhibited Lkt-induced cytolysis of the transfectants. Exposure of LFA-1 transfectants to low concentrations of Lkt resulted in elevation of intracellular [Ca(2+)](i). Taken together, these results indicate that DS LFA-1 serves as a receptor for M. haemolytica Lkt.  相似文献   

3.
Pneumonia caused by Mannheimia haemolytica is an important disease of cattle (BO), domestic sheep (DS, Ovis aries) and bighorn sheep (BHS, Ovis canadensis). Leukotoxin (Lkt) produced by M. haemolytica is cytolytic to all leukocyte subsets of these three species. Although it is certain that CD18, the beta subunit of beta(2) integrins, mediates Lkt-induced cytolysis of leukocytes, whether CD18 of all three beta(2) integrins, LFA-1 (CD11a/CD18), Mac-1 (CD11b/CD18) and CR4 (CD11c/CD18), mediates Lkt-induced cytolysis of BO, DS and BHS leukocytes remains a controversy. Based on antibody inhibition experiments, earlier studies suggested that LFA-1, but not Mac-1 and CR-4, serves as a receptor for M. haemolytica Lkt. PMNs express all three beta(2) integrins, and they are the leukocyte subset that is most susceptible to Lkt. Therefore we hypothesized that all three beta(2) integrins serve as the receptor for Lkt. The objective of this study was to determine whether Mac-1 of BO, DS and BHS serves as a receptor for Lkt. cDNAs for CD11b of BO, DS and BHS were transfected into a Lkt-non-susceptible cell line along with cDNAs for CD18 of BO, DS and BHS, respectively. Transfectants stably expressing BO, DS or BHS Mac-1 specifically bound Lkt. These transfectants were lysed by Lkt in a concentration-dependent manner. Increase in intracellular [Ca(2+)](i) was observed in transfectants following exposure to low concentrations of Lkt indicating signal transduction through secondary messengers. Collectively, these results indicate that Mac-1 from these three species serves as a receptor for M. haemolytica Lkt.  相似文献   

4.
Leukotoxin (Lkt) and LPS are the major virulence determinants of Mannheimia haemolytica that contribute to the pathogenesis of bovine and ovine pneumonic pasteurellosis. We have previously identified bovine and ovine CD18 as the functional receptor for Lkt. LPS complexes with Lkt resulting in increased thermal stability and enhanced cytotoxic activity of Lkt. Cellular recognition of LPS involves several different molecules including CD14. We hypothesized that expression of ovine CD14 together with LFA-1 or Mac-1 would enhance Lkt-induced cytotoxicity. Ovine cDNA for CD14 was amplified by PCR and cloned into mammalian expression vectors. The 1122 bp cDNAs for bighorn sheep (BHS) and domestic sheep (DS) CD14 encode 373 amino acids which exhibit 99% identity with each other. Ovine CD14 plasmids were transfected either into HEK-293 cells, or previous HEK-293 transfectants stably expressing ovine LFA-1 or Mac-1. Flow cytometric analysis of transfectants confirmed the cell surface expression of CD14. The transfectants expressing LFA-1 or Mac-1 and the transfectants co-expressing CD14 with LFA-1 or Mac-1 did not show any significant difference in Lkt-induced cytotoxicity when incubated with LPS complexed Lkt. In contrast, incubation of the LFA-1 or Mac-1 and LFA-1/CD14 or Mac-1/CD14 transfectants with Lkt which lacks LPS, resulted in reduced cytotoxicity. None of the above transfectants showed any difference in [Ca2+](i) elevation when incubated with both types of Lkt preparations. Lkt did not induce any cytotoxicity or [Ca2+](i) elevation in ovine CD14 transfectants or parent HEK-293 cells. Based on these findings, we conclude that expression of CD14 together with LFA-1 or Mac-1 does not enhance Lkt-induced cytotoxicity, whereas LPS enhances cytotoxicity by complexing with Lkt.  相似文献   

5.
Previously, we have shown that CD18, the beta-subunit of beta(2)-integrins, serves as a receptor for leukotoxin (Lkt) secreted by Mannheimia (Pasteurella) haemolytica on bovine leukocytes. Anti-CD18 monoclonal antibodies (mAbs) inhibit Lkt-induced cytolysis of bighorn sheep (Ovis canadensis) leukocytes suggesting that CD18 may serve as a receptor for Lkt on the leukocytes of this species as well. Confirmation of bighorn sheep CD18 as a receptor for Lkt, and elucidation of the enhanced Lkt-susceptibility of bighorn sheep polymorphonuclear leukocytes (PMNs), necessitates the cloning and sequencing of cDNA encoding bighorn sheep CD18. Hence, in this study we cloned and sequenced the cDNA encoding CD18 of bighorn sheep, and compared with that of other animal species. The cDNA of bighorn sheep CD18 has an open reading frame (ORF) of 2310bp. CD18 sequences obtained individually from peripheral blood mononuclear cells (PBMCs) and PMNs were identical to each other. Comparison of the deduced 770-amino acid sequence of CD18 of bighorn sheep with that of domestic sheep, goats, cattle, humans and mice revealed 99, 98, 95, 82 and 80% identity, respectively. Availability of cloned bighorn sheep CD18 cDNA should allow the molecular characterization of M. haemolytica Lkt-receptor interactions in bighorn sheep and other ruminants that are susceptible to this disease.  相似文献   

6.
Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia, produces Apx toxins that are recognized as major virulence factors. Recently, we showed that ApxIIIA-cytotoxic activity specifically targets Sus scrofa leukocytes. Since both LtxA from Aggregatibacter actinomycetemcomitans (aggressive periodontitis in humans) and LktA from Mannheimia haemolytica (pneumonia in ruminants) share this characteristic, respectively towards human and ruminant leukocytes, and because both use the CD18 subunit to interact with their respective LFA-1, we hypothesized that ApxIIIA was likely to bind porcine CD18 to exercise its deleterious effects on pig leukocytes. A β 2−integrin-deficient ApxIIIA-resistant human erythroleukemic cell line was transfected either with homologous or heterologous CD11a/CD18 heterodimers using a set of plasmids coding for human (ApxIIIA-resistant), bovine (-resistant) and porcine (-susceptible) CD11a and CD18 subunits. Cell preparations that switched from ApxIIIA-resistance to -susceptibility were then sought to identify the LFA-1 subunit involved. The results showed that the ApxIIIA-resistant recipient cell line was rendered susceptible only if the CD18 partner within the LFA-1 heterodimer was that of the pig. It is concluded that porcine CD18 is necessary to mediate A. pleuropneumoniae ApxIIIA toxin-induced leukolysis.  相似文献   

7.
Mannheimia (Pasteurella) haemolytica, the etiologic agent of bovine pneumonic mannheimiosis, produces an exotoxic leukotoxin. The leukotoxin (LktA) is a member of the RTX (repeats in toxin) family of bacterial cytolysins and is distinguished from other toxins by its unique target cell specificity to ruminant leukocytes occurring through binding to a specific receptor. We have demonstrated previously that the beta2 integrin LFA-1 is a receptor for LktA in bovine leukocytes and is involved in leukotoxin-induced biological effects. However the subunits within LFA-1 involved in binding to LktA, and post-binding signaling leading to cellular activation have not been well characterized. The purpose of our study was to pinpoint these precise subunits on bovine alveolar macrophages and to characterize their interaction with LktA. The results in this study indicate that although LktA can efficiently bind to the CD18 subunit of both LFA-1 and Mac-1, post-binding signaling events including elevation of intracellular calcium and CD18 tail phosphorylation are only observed through LFA-1. Furthermore, LktA also binds to the CD11a subunit of LFA-1. LktA binding to CD11a could be inhibited by a small molecule inhibitor of the I(inserted)-domain, the major ligand binding interface on CD11a. I-domain inhibition significantly blunts LktA-induced intracellular calcium elevation and tyrosine phosphorylation of the CD18 tail. Based on our results we suggest that LFA-1 serves as the functional leukotoxin receptor on bovine alveolar macrophages.  相似文献   

8.
Bovine herpesvirus-1 (BHV-1) has been reported to increase the susceptibility of cattle to respiratory disease caused by Mannheimia (Pasteurella) haemolytica A1. The principal virulence factor of M. haemolytica is a leukotoxin (LKT) that can specifically kill ruminant leukocytes following its binding to the beta2-integrin CD11a/CD18 (lymphocyte function-associated antigen 1 (LFA-1)). In this study, we investigated the effects of experimental infection of bovine peripheral blood mononuclear cells (MNCs) with BHV-1 in vitro, on the subsequent interaction of these cells with the M. haemolytica LKT. We found that BHV-1 infection increased LFA-1 expression (as assessed by flow cytometry), and subsequently enhanced LKT binding and cytotoxicity to bovine MNCs. We also found that BHV-1 infection increased CD18, IL-1beta, and IFN-gamma mRNA expression by MNCs. As previously reported for bovine polymorphonuclear neutrophils (PMNs), MNCs increased their expression of LFA-1, and their LKT binding and cytotoxicity, following exposure to IL-1beta, TNF-alpha, and IFN-gamma. These findings suggest that BHV-1 infection, and the resulting release of inflammatory cytokines, can stimulate expression of LFA-1 in bovine MNCs, thus enhancing the binding and biological effects of LKT. If such a mechanism occurs in vivo it might explain, in part, the increased susceptibility of BHV-1 infected cattle to bovine pasteurellosis.  相似文献   

9.
In this study we have used the expression of perforin to characterize subsets of porcine cytotoxic lymphocytes. Perforin positive lymphocytes expressed both CD2 and CD8, most were small dense lymphocytes (SDL) and up to 90% were CD3 negative. However, the numbers of perforin positive T-cells increased with the age of the animal and their populations increased after specific antigen stimulation in vitro. The remaining perforin positive lymphocytes were large and granular and contained more CD3+CD5+CD6+ T-cells (−40%) of which a substantial proportion also co-expressed CD4. Perforin was expressed in subpopulations of both CD8 and CD8β lymphocytes, but was not expressed in γδ T-cells or monocyte/macrophages. The perforin positive CD3− subset was phenotypically homogeneous and defined as CD5−CD6−CD8β−CD16+CD11b+. This population had NK activity and expressed mRNA for the NK receptor NKG2D, and adaptors DAP10 and DAP12. Perforin positive T-cells (CD3+) could be divided into at least three subsets. The first subset was CD4−CD5+CD6+CD11b−CD16− most were small dense lymphocytes with cytotoxic T-cell activity but not all expressed CD8β. The second subset was mainly observed in the large granular lymphocytes. Their phenotype was CD4+CD5+CD6+CD8β+CD16−CD11b− and also showed functional CTL activity. Thus not all of double positive T-cells are memory helper T-cells. The third subset did not express the T-cell co-receptor CD6, but up to half of them expressed another T-cell co-receptor CD5. The majority of this subset expressed CD11b and CD16, thus the third perforin positive T-cell subset was CD3+CD4−CD5+CD6−CD8β±CD11b+CD16+, and possessed MHC-unrestricted cytotoxicity and LAK activity.  相似文献   

10.
Mannheimia (Pasteurella) haemolytica A1 produces an extracellular leukotoxin (LKT) that is reported to bind the beta(2)-integrin CD11a/CD18 (LEA-1) on ruminant leukocytes. LKT binding induces activation, and subsequent cytolysis, of these cells. It is well known that active viral infection greatly increases the susceptibility of cattle to pasteurellosis. To better understand the mechanism by which this occurs, we investigated the effects of experimental in vivo infection of cattle with bovine herpes virus-1 (BHV-1) on the ex vivo interaction of bovine leukocytes with the M. haemolytica LKT. In this study, we demonstrated that active BHV-1 infection increased the expression of the beta(2)-integrin CD11a/CD18 (as defined by the mAb BAT75) on bovine peripheral blood neutrophils, enhanced the binding of LKT to bronchoalveolar lavage (BAL) leukocytes and peripheral blood neutrophils, and increased the killing of BAL leukocytes and peripheral blood leukocytes by LKT. In addition, BHV-1 greatly increased the number of BAL, resulting in many more LKT-responsive cells being present in the lungs. These findings might explain in part the increased susceptibility of BHV-1 infected cattle to pneumonic pasteurellosis.  相似文献   

11.
Thyroxine-binding prealbumin (TBPA) and thyroxine-binding globulin (TBG) were isolated from canine serum and partially characterized. TBPA was isolated by retinol-binding protein (RBP) affinity chromatography and further purified by preparative agarose gel electrophoresis or FPLC ion exchange chromatography. TBG was purified by thyroxine (T4)-Sepharose chromatography followed by gel filtration on Sephacryl S-300 and preparative electrofocusing in a granulated dextran gel. Molecular weights were estimated by SDS-polyacrylamide gradient gel electrophoresis. Canine TBPA had a tetramer molecular weight of 56,000, an extinction coefficient of 12.8 cm2cg−1, an isoelectric point of 5.26–5.70 and a microheterogeneity pattern similar to that of human TBPA. Partial immunochemical identity with human TBPA was also found. Plasma concentrations of TBPA were measured by rocket immunoelectrophoresis in 43 normal and 35 hypothyroid dogs. Reference levels for TBPA ranged between 205 and 474 mg/l. Hypothyroid dogs had a mean TBPA level of 315.0 mg/l (SD: 91.1 mg/l). TBG had a molecular weight of 75,000 and an isoelectric point of 5.0. No immunochemical identity with human TBG was found. Gel filtration of serum on Sephacryl S-200, identification of T4-binding proteins with 125I-T4, and protein- and lipoprotein staining of fractions was performed. Thyroxine-binding was found to TBG in the β-globulin region, TBPA in the 2-region, albumin, and to the high density lipoprotein (HDL2) in the 1-region and the very low density lipoprotein (VLDL) in the pre-β region. A corresponding band to the latter protein in serum was masked by TBG and TBPA, and T4-binding in the 1-region was not always seen in serum. Many similarities were found between man and dog regarding TBPA, but not TBG. The differences in structure of TBG may in part be responsible for the low serum T4 levels and rapid T4 metabolism seen in dogs.  相似文献   

12.
Molt induced by infusion of a gonadotropin-releasing hormone agonist (GnRH-A, ([D-leu6, Pro9]-GnRH N-ethylamide]) or feed withdrawal (FW) has been used as a model to study interactions between ovarian activity and thymosin β4 during molting in domestic hens. Thirty-three laying hens were divided into three groups: 1, controls, 2, GnRH-A infusion induced molt (GnRH-A), or 3, FW induced molt. All groups had reduced daylength. Blood was sampled weekly and assayed for concentrations of thymosin β4 and progesterone (P4). Plasma P4 concentrations were significantly depressed in both treatment groups compared to controls, indicating ovarian regression. Plasma P4 concentrations had returned to control values in the GnRH-A group by 28 d after the start of treatment, while P4 was still depressed in the FW group at day 42 when the experiment ended. Plasma concentrations of thymosinβ4 were elevated relative to controls from day 7 through day 14 in the GnRH-A group and from day 7 until day 28 in the FW group. It is concluded that plasma concentrations of thymosin β4 are elevated during molting in domestic hens, but the elevation is not attributable to depressed P4 concentrations.  相似文献   

13.
Following activation of granulocytes, L-selectin (CD62L) is generally shed from the cellular surface, whereas Mac-1 (CD11b/CD18) expression is well known to increase. However, a number of studies in bovines and humans show that the expression of L-selectin may increase as well. This urged us to examine the possible existence of both L-selectin and Mac-1 reservoirs in bovine neutrophil and eosinophil populations through the use of flow cytometry in combination with an optimized method for cell membrane permeabilization. Augmented L-selectin and Mac-1 expression was detected in both granulocyte populations upon saponin treatment. Confocal microscopic studies indicated that both molecules exhibit a different pattern of subcellular localization. Incubation with sialidase revealed the existence of hidden L-selectin epitopes at the cell surface, while no additional Mac-1 epitopes were exposed. Platelet-activating factor stimulation decreased surface and total expression of L-selectin to the same extent in both populations, but solely affected Mac-1 surface expression on eosinophils. Moreover, cytoskeletal actin filaments and microtubules were found to be involved in the regulation of Mac-1 surface expression on bovine neutrophils and eosinophils. In marked contrast, expression of L-selectin was minimally affected by cytoskeleton perturbing agents. The present study indicates that L-selectin and Mac-1 adhesion molecules reside in distinctly located reservoirs in bovine granulocytes and can be selectively mobilized upon in vitro stimulation.  相似文献   

14.
Molecular cloning and expression analysis of pig CD81   总被引：1，自引：0，他引：1  
CD81, also known as TAPA-1 (target of antiproliferative antibody 1), is a member of the tetraspanin family of proteins and a component of the B cell co-receptor complex. Several studies have shown that CD81 plays significant roles in a variety of immune responses, including activation of B cells and T cells. In this study, we cloned pig Cd81 cDNA using RT-PCR coupled with rapid amplification of cDNA ends (RACE)-PCR and determined the complete cDNA sequence of pig Cd81. Pig Cd81 cDNA contains an open reading frame (711 bp) encoding 236 amino acids. The identity of pig CD81 with those of human, cattle, rat, and mouse are 90.30%, 92.26%, 86.22%, and 86.22%, respectively. Alignment of the CD81 amino acid sequence with those of mammalian species showed that the large extracellular loop (LEL) is the most divergent, whereas other domains are largely conserved. Pig Cd81 mRNA was detected by RT-PCR in a broad range of tissues, including lymphoid tissues as well as nonlymphoid tissues, indicated variety of cellular functions of CD81 in most pig tissues. Flow cytometry analyses demonstrated that human CD81 antibody recognizes a pig CD81 on the cell surface. Further, immunohistochemistry analysis using human CD81 antibody on pig spleen was revealed that CD81 expression is widely diffused in spleen tissue. Future study will be focused on defining the functional role of CD81 during the course of pig infectious diseases.  相似文献   

15.
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16.
An immunoradiometric assay and serum extraction procedure were developed to measure dimeric inhibin in porcine serum with minimal interference by putative inhibin-binding proteins. Assay sensitivity was 50 pg/tube, and it incorporated antibodies against the N-terminal region of inhibin's -subunit, -(1–25)-Ab, and against the C-terminal region of inhibin's βA-subunit. To determine whether inhibin-binding proteins were present in porcine serum, serum was incubated with [125I]-recombinant human (rh)-inhibin and then chromatographed by gel filtration. Radioiodinated rh-inhibin was associated with protein(s) >600 kDa. Radioiodinated rh-inhibin also was incubated with 2-macroglobulin, an inhibin-binding protein in human and rat serum. Elution profiles were similar for serum and 2-macroglobulin. Serum- and 2-macroglobulin-[125I]rh-inhibin complexes dissociated upon exposure to 8 M urea. Porcine serum was treated with urea, after which inhibin was isolated and concentrated. The recovery of rh-inhibin added to starting serum was 28%. Concentrations of endogenous dimeric inhibin were <28 pg/ml in serum collected from sows at random stages of the estrous cycle and were <21 pg/ml in serum collected from sows 2 d postweaning. Results demonstrate that 1) concentrations of dimeric inhibin are low in porcine serum, and 2) an inhibin-binding protein(s), consistent with 2-macroglobulin, is present in porcine serum.  相似文献   

17.
The effect of endotoxin on follicular growth and on secretion of LH, estradiol-17β, progesterone and cortisol during the proestrous phase in cattle was investigated. Holstein heifers were treated with PGF2 at 11–13 d after ovulation to induce luteolysis. At 42 hr after PGF2 treatment, heifers were administered either lipopolysaccharide (LPS; Escherichia coli, O111:B4, 5 μg/kg, n = 6) or saline (control; n = 6) by i.v. bolus injection. Ovarian structures were monitored daily by transrectal ultrasonography, and blood samples were collected at various times for hormonal analysis. The duration from PGF2 treatment to ovulation was significantly longer in the LPS group (8.0 ± 1.3 d) than in the control group (4.2 ± 0.2 d). LPS significantly reduced the pulse frequency of LH for 6 hr after the administration, and increased the mean concentration and pulse amplitude of LH from 3 to 6 hr after the administration. The plasma concentrations of progesterone and cortisol were transiently increased after LPS administration. The plasma concentration of estradiol-17β was significantly decreased at 24 hr after LPS administration compared to that in the controls. Five of six LPS-treated heifers exhibited no preovulatory LH surge until 120 hr after PGF2 treatment and the remaining heifer exhibited the surge at 108 hr after PGF2 treatment, while the LH surge was observed at 54–78 hr after PGF2 treatment in control heifers. These results suggest that endotoxin disrupts progression of the proestrous phase of cattle, interrupting the preovulatory estradiol rise and thus delaying the LH surge and the subsequent ovulation.  相似文献   

18.
Scientific literature on the use of antibiotic growth promoters (AGP) in beef cattle consuming forage-diets was reviewed. Database summarizes 136 comparisons between untreated and AGP-treated cattle from 48 bibliographic references. Performance data of cattle receiving AGP either alone or in combination with 17 β-estradiol implants was statistically analyzed. Forage quality, in terms of average daily gain (ADG), differentially influenced (P = 0.1) the effect of AGP on beef cattle performance. As the quality of forage increased, the estimated net ADG response to monensin decreased and that to lasalocid increased. ADG increased quadratically (P < 0.01) with increasing doses of monensin (R2 = 0.71) or lasalocid (R2 = 0.63). Ionophore-dosage level quadratically improved (P = 0.01; R2 = 0.52) feed conversion (FCONV) of cattle without affecting their dry matter intake (DMI; P > 0.1). A linear relationship (P < 0.01) between ADG and dose of tetronasin (R2 = 0.64) or lysocellin (R2 = 0.52) was also observed. The combination of monensin and 17β-estradiol implants resulted in an additive effect on ADG of grazing cattle. The experimental results reviewed show that, in beef cattle consuming forage-based diets, ionophores improve ADG and FCONV in a dose-dependent manner, with little or no effect on DMI. In addition, results suggest that forage quality influences the direction of the ADG response to AGP supplementation in cattle. Summarized data from beef cattle implanted with 17β-estradiol and/or supplemented with monensin indicate that the combined response of these two compounds on cattle grazing high-quality pastures has no effect on ADG over that obtained by 17β-estradiol alone.  相似文献   

19.
The chemoattractant activity of a new chemotactic factor, 'Gasserokine' produced by Lactobacillus gasseri JCM1131T, has been proposed as a novel immunological function of probiotic lactic acid bacteria. The focus of the present study was to understand the mechanism of the chemotaxis induced by Gasserokine, using activation of an adhesion molecule, Mac-1 (CD11b/CD18) on macrophages. The macrophage chemotaxis to Gasserokine was abolished by preincubation of macrophages with the anti-Mac-1 mAb. Gasserokine induced rapid serine phosphorylation of CD18 molecules within 1 min of stimulation, but the effect was short-lived. Substantial tyrosine phosphorylation was observed in CD18-associated protein of macrophages stimulated by Gasserokine. The tyrosine phosphorylation was confirmed in macrophages stimulated with Gasserokine and also serine/threonine phosphorylation was detected on CD18 molecules by laser microscopy using a double immunostaining method. These results suggest that selective activation of intracellular signaling cascades, such as the mitogen-activated protein kinase pathway, are related to the macrophage chemotaxis induced by Gasserokine.  相似文献   

20.
It is well known that the immune system changes with age during development and maturation in Holstein cattle. But age-related changes in leukocytes and T cell subsets in peripheral blood of Japanese Black cattle still remain unclear. The aim of the present study was to investigate comparative changes of leukocytes (granulocytes, monocytes, B cells and T cells) and T cell subsets (CD4+, CD8+, γδ, CD8+γδ and WC1+γδ T cells) in Japanese Black cattle aged 0.5, 1, 2, 6, 18 and 36–41 (adult) months on flow cytometry using specific monoclonal antibodies for the cell surface markers. T cell proportion was approximately 40% in 2-month-old cattle and decreased to 20.6% in adults. In contrast, B cell proportion significantly increased from 7.4% to 28.2% with age. In T cell subsets the percentage of CD4+ T cells significantly increased from 40.5% to 60%, but that of WC1+γδ T cell subset significantly decreased with age. The percentages of CD8+ and CD8+γδ T cells did not change. The present study details the proportional changes in leukocyte and T cell subsets with age in the peripheral blood of Japanese Black cattle and these findings are similar to those described for Holstein cattle.  相似文献   

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