Endocytic delivery of vancomycin mediated by a synthetic cell surface receptor: rescue of bacterially infected Mammalian cells and tissue targeting in vivo.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
Avery August
Publications and source records attributed to Avery August.
Vitamin D receptor expression by the lung micro-environment is required for maximal induction of lung inflammation.
Mice lacking the vitamin D receptor (VDR) are resistant to airway inflammation. Pathogenic immune cells capable of transferring experimental airway inflammation to wildtype (WT) mice are present and primed in the VDR KO mice. Furthermore, the VDR KO immune cells homed to the WT lung in sufficient numbers to induce symptoms of asthma. Conversely, WT splenocytes, Th2 cells and hematopoetic cells induced some symptoms of experimental asthma when transferred to VDR KO mice, but the severity was less than that seen in the WT controls. Interestingly, experimentally induced vitamin D deficiency failed to mirror the VDR KO phenotype suggesting there might be a difference between absence of the ligand and VDR deficiency. Lipopolysaccharide (LPS) induced inflammation in the lungs of VDR KO mice was also less than in WT mice. Together the data suggest that vitamin D and the VDR are important regulators of inflammation in the lung and that in the absence of the VDR the lung environment, independent of immune cells, is less responsive to environmental challenges.
Animals↗
Tec kinase Itk forms membrane clusters specifically in the vicinity of recruiting receptors.
The Tec family of tyrosine kinases transduces signals from antigen and other receptors in cells of the hematopoietic system. In particular, interleukin-2 inducible T cell kinase (Itk) plays an important role in modulating T cell development and activation. Itk is activated by receptors via a phosphatidylinositol 3-kinase-mediated pathway, which results in recruitment of Itk to the plasma membrane via its pleckstrin homology domain. We show here that membrane localization of Itk results in the formation of clusters of at least two molecules within 80 A of each other, which is dependent on the integrity of its pleckstrin homology domain. By contrast, the proline-rich region within the Tec homology domain, SH3 or SH2 domains, or kinase activity were not required for this event. More importantly, these clusters of Itk molecules form in distinct regions of the plasma membrane as only receptors that recruit phosphatidylinositol 3-kinase reside in the same membrane vicinity as the recruited Itk. Our results indicate that Itk forms dimers in the membrane and that receptors that recruit Itk do so to specific membrane regions.
Cell Line↗
A kinase independent function for Tec kinase ITK in regulating antigen receptor induced serum response factor activation.
The Tec family kinases are critical downstream regulators of antigen receptor signals in lymphocytes. As kinases, they act on critical substrates to regulate signals such as calcium increase leading to activation of transcription factors such as NFAT, NFkappaB and SRF. We now show here that ITK, a member of the Tec family of tyrosine kinases, has a kinase independent function. Mutants of ITK that lack kinase activity or a kinase domain can rescue cells lacking Tec family kinases for antigen receptor induced SRF activation, but not for NFAT, AP-1 or NFkappaB activation. Furthermore, expression of these mutants in WT cells enhanced SRF activation. This kinase independent function required the SH2 domain since a mutant lacking both the kinase and SH2 domains was much less effective at rescuing SRF activation. This kinase-deleted mutant could partially rescue ERK activation, and interact with multiple tyrosine phosphorylated proteins during antigen receptor signaling, suggesting that ITK uses a scaffolding function that regulates signals leading to specific regulation of SRF activation.
Animals↗
Reduced airway hyperresponsiveness and tracheal responses during allergic asthma in mice lacking tyrosine kinase inducible T-cell kinase.
BACKGROUND: Patients with allergic asthma have symptoms of a predominant T(H)2 response, including airway eosinophilic inflammation and increased mucous production in the lungs. This accompanies increased airways responsiveness, which can be life threatening. Because T(H)2 cells and cytokines have been implicated in contributing to these symptoms, pathways that control the development of these cells or that regulate their cytokine production represent good targets for controlling this disease. OBJECTIVE: We have previously shown that mice lacking the tyrosine kinase inducible T-cell kinase (ITK) have drastically reduced airway inflammation in a model of allergic asthma. However, it was not clear whether this translated into reduced airways hyperresponsiveness. We have analyzed tracheal responsiveness and airways hyperresponsiveness of wild-type (WT) and ITK null mice during induction of experimental allergic asthma. METHODS: Experimental allergic asthma was induced in WT and ITK knockout mice. Tracheal responses to carbachol, acetylcholine, and potassium chloride were analyzed. Airways hyperresponsiveness to methacholine challenge was also analyzed in allergen-challenged mice, along with lung and bronchoalveolar lavage fluid T(H)2 cytokine message and protein. RESULTS: ITK null mice have reduced tracheal responses to cholinergic challenge in vitro before as well as after allergen challenge. These mice also have reduced airways hyperresponsiveness in response to allergen challenge, which could be rescued by transferring WT splenocytes or purified WT CD4+ T cells. This reduced airways response was preferentially accompanied by reduced expression of T(H)2 cytokines in the lungs. CONCLUSION: Our results indicate that the tyrosine kinase ITK and its function in T cells represent an attractive target for antiasthmatic drugs. CLINICAL IMPLICATIONS: Modulating the expression or activity of ITK may be a novel strategy to block allergic airway inflammation.
Acetylcholine↗
Nutrients, nuclear receptors, inflammation, immunity lipids, PPAR, and allergic asthma.
The peroxisome proliferator-activated receptors (PPARs) belong to the larger superfamily of steroid/thyroid nuclear receptors. PPARgamma is expressed in a number of hematopoietic cells, including dendritic cells, eosinophils, macrophages, and T cells. A number of lipids and synthetic compounds interact with PPARgamma, that, depending on the cell type, results in the regulation of specific genes. There is now a large body of data indicating that allergic asthma is the result of a predominant type-2 helper T cell immune response including IL-4, -5 and -13, eosinophilic inflammation in the lungs, mucous production, and airway hyperresponsiveness (AHR). Targeting the production of these type-2 helper T cell mediated cytokines has been proposed as a way to regulate this disease. Because PPARgamma ligands can affect T cell cytokine production in vitro, we have examined whether these ligands affect symptoms of allergic asthma in a murine model of this disease. We discuss data showing that ciglitazone and GW1929, two agonistic ligands for PPARgamma, significantly inhibited airway inflammation during allergic asthma induction. Oral treatment with ciglitazone and GW1929 inhibited airway inflammation, with less of an effect on AHR. By contrast, intranasal exposure to GW1929 significantly reduced AHR following exposure to allergen, while GW9662, a PPARgamma antagonist, had no effect. In vitro, T cells from ciglitazone-treated mice secreted significantly less IL-4 and IFN-gamma in response to restimulation. These data suggest that PPARgamma agonists may be useful for the treatment of allergic asthma.
Animal Nutritional Physiological Phenomena↗
Differential regulation of cytokine production by CD1d-restricted NKT cells in response to superantigen staphylococcal enterotoxin B exposure.
NKT cells are a heterogeneous population characterized by the ability to rapidly produce cytokines, such as interleukin 2 (IL-2), IL-4, and gamma interferon (IFN-gamma) in response to infections by viruses, bacteria, and parasites. The bacterial superantigen staphylococcal enterotoxin B (SEB) interacts with T cells bearing the Vbeta3, -7, or -8 T-cell receptors, inducing their expansion and cytokine secretion, leading to death in some cases due to cytokine poisoning. The majority of NKT cells bear the Vbeta7 or -8 T-cell receptor, suggesting that they may play a role in regulating this response. Using mice lacking NKT cells (CD1d(-/-) and Jalpha18(-/-) mice), we set out to identify the role of these cells in T-cell expansion, cytokine secretion, and toxicity induced by exposure to SEB. We find that Vbeta8(+) CD4(+) T-cell populations similarly expand in wild-type (WT) and NKT cell-null mice and that NKT cells did not regulate the secretion of IL-2. By contrast, these cells positively regulated the secretion of IL-4 and IFN-gamma production and negatively regulated the secretion of tumor necrosis factor alpha (TNF-alpha). However, this negative regulation of TNF-alpha secretion by NKT cells provides only a minor protective effect on SEB-mediated shock in WT mice compared to mice lacking NKT cells. These data suggest that NKT cells may regulate the nature of the cytokine response to exposure to the superantigen SEB and may act as regulatory T cells during exposure to this superantigen.
Animals↗
A role for the Tec family kinase ITK in regulating SEB-induced interleukin-2 production in vivo via c-jun phosphorylation.
BACKGROUND: Exposure to Staphylococcal Enterotoxin B (SEB), a bacterial superantigen secreted by the Gram-positive bacteria Staphyloccocus aureus, results in the expansion and eventual clonal deletion and anergy of Vbeta8+ T cells, as well as massive cytokine release, including Interleukin-2 (IL-2). This IL-2 is rapidly secreted following exposure to SEB and may contribute to the symptoms seen following exposure to this bacterial toxin. The Tec family kinase ITK has been shown to be important for the production of IL-2 by T cells stimulated in vitro and may represent a good target for blocking the production of this cytokine in vivo. In order to determine if ITK represents such a target, mice lacking ITK were analyzed for their response to SEB exposure. RESULTS: It was found that T cells from mice lacking ITK exhibited significantly reduced proliferative responses to SEB exposure in vitro, as well as in vivo. Examination of IL-2 production revealed that ITK null mice produced reduced levels of this cytokine in vitro, and more dramatically, in vivo. In vivo analysis of c-jun phosphorylation, previously shown to be critical for regulating IL-2 production, revealed that this pathway was specifically activated in SEB reactive Vbeta8+ (but not non-reactive Vbeta6+) T cells from WT mice, but not in Vbeta8+ T cells from ITK null mice. However, toxicity analysis indicated that both WT and ITK null animals were similarly affected by SEB exposure. CONCLUSION: These data show that ITK is required for IL-2 production induced by SEB in vivo, and may regulate signals leading IL-2 production, in part by regulating phosphorylation of c-jun. The data also suggest that perturbing T cell activation pathways leading to IL-2 does not necessarily lead to improved responses to SEB toxicity.
Animals↗
Actin depolymerization transduces the strength of B-cell receptor stimulation.
Polymerization of the actin cytoskeleton has been found to be essential for B-cell activation. We show here, however, that stimulation of BCR induces a rapid global actin depolymerization in a BCR signal strength-dependent manner, followed by polarized actin repolymerization. Depolymerization of actin enhances and blocking actin depolymerization inhibits BCR signaling, leading to altered BCR and lipid raft clustering, ERK activation, and transcription factor activation. Furthermore actin depolymerization by itself induces altered lipid raft clustering and ERK activation, suggesting that F-actin may play a role in separating lipid rafts and in setting the threshold for cellular activation.
Actins↗
Natural killer T cells: rapid responders controlling immunity and disease.
Natural killer T (NKT) cells are a subset of T cells that share properties of natural killer cells and conventional T cells. They are involved in immediate immune responses, tumor rejection, immune surveillance and control of autoimmune diseases. Most NKT cells express both an invariant T cell antigen receptor and the NK cell receptor NK1.1, and are referred to as invariant NKT cells. This invariant T cell receptor is restricted to interactions with glycolipids presented by the non-classical MHC, CD1d. These NKT cells rapidly produce high levels of interleukin (IL)-2, IFN-gamma, TNF-alpha, and IL-4 upon stimulation through their TCR. Most also have cytotoxic activity similar to NK cells. NKT cells are involved in a number of pathological conditions, and have been shown to regulate viral infections in vivo, and control tumor growth. They may also play both protective and harmful roles in the progression of certain autoimmune diseases, such as diabetes, lupus, atherosclerosis, and allergen-induced asthma.
Animals↗
Vitamin D receptor-deficient mice fail to develop experimental allergic asthma.
The active metabolite of vitamin D (1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3))) is known to modulate the immune response in Th1 cell-directed diseases. To investigate the role of vitamin D in Th2 cell-directed diseases, experimental allergic asthma was induced in vitamin D receptor (VDR) knockout and in wild-type (WT) mice. As expected, WT mice developed symptoms of airway inflammation with an influx of eosinophils, elevated Th2 cytokine levels, mucous production, and airway hyperresponsiveness. The administration of 1,25(OH)(2)D(3) had no effect on asthma severity. The only discernable effect of 1,25(OH)(2)D(3) on experimental allergic asthma in WT mice was an increased expression of two Th2-related genes (soluble CD23 and GATA-3) in lungs of BALB/c mice exposed to Ag through the nasal route only. By contrast, asthma-induced VDR knockout mice failed to develop airway inflammation, eosinophilia, or airway hyperresponsiveness, despite high IgE concentrations and elevated Th2 cytokines. The data suggest that although 1,25(OH)(2)D(3) induced these Th2-type genes, the treatment failed to have any affect on experimental asthma severity. However, VDR-deficient mice failed to develop experimental allergic asthma, suggesting an important role for the vitamin D endocrine system in the generation of Th2-driven inflammation in the lung.
Animals↗
Regulation of CXC chemokine receptor 4-mediated migration by the Tec family tyrosine kinase ITK.
Chemokines are critical in controlling lymphocyte traffic and migration. The CXC chemokine CXCL12/SDF-1alpha interacts with its receptor CXCR4 to induce the migration of a number of different cell types. Although an understanding of the physiological functions of this chemokine is emerging, the mechanism by which it regulates T cell migration is still unclear. We show here that the Tec family kinase ITK is activated rapidly following CXCL12/SDF-1alpha stimulation, and this requires Src and phosphatidylinositol 3-kinase activities. ITK regulates the ability of CXCL12/SDF-1alpha to induce T cell migration as overexpression of wild-type ITK-enhanced migration, and T cells lacking ITK exhibit reduced migration as well as adhesion in response to CXCL12/SDF-1alpha. Further analysis suggests that ITK may regulate CXCR4-mediated migration and adhesion by altering the actin cytoskeleton, as ITK null T cells were significantly defective in CXCL12/SDF-1a-mediated actin polymerization. Our data suggest that ITK may regulate the ability of CXCR4 to induce T cell migration.
Animals↗
Modulation of tracheal smooth muscle responses in inducible T-cell kinase knockout mice.
It has been shown that the Tec family nonreceptor tyrosine kinase inducible T cell kinase (ITK) plays a role in the activation of naive T cells and in the differentiation of T helper (TH2)-type cells producing cytokines in a model of allergic inflammation, thereby possibly indirectly mediating hyperresponsivenes of airway smooth muscle tone. Using excised tracheae from wild type (WT) mice and those lacking ITK, we conducted a series of in vitro experiments in which isometric smooth muscle tones were assessed in response to several agonists to determine whether the absence of ITK would affect the responsiveness of tracheal smooth muscle cells. The resulting change in contractile responses was evaluated by measuring agonist cumulative concentration-response curves (CCRC). Our results indicate that the cholinergic agonist acetylcholine (ACh) and its analog carbachol (CCh) exhibited comparable CCRC profiles in contracting isolated tracheae from both WT and ITK-/- mice, with no alteration in their efficacies. However, the EC50 values for the two agonists were found to be significantly higher in ITK-/- tracheae than in those from WT mice, suggesting an alteration of the potencies of these cholinergic agonists in the trachea of ITK-/- mice. Moreover, we found that the depolarizing agent potassium chloride (KCl) had a significantly lower efficacy in contracting ITK-/- tracheae compared to those from WT mice. This difference in KCl efficacy was abolished in the presence of a calcium (Ca2+) voltage-dependent channel (VDC) agonist, Bay K8644, suggesting a modulation of the KCl induced permeability of VDC Ca2+ channels in the trachea of ITK-/- mice. Taken together, these results suggest that the presence of ITK may play a modulating role in the pharmacomechanical as well as in the electromechanical coupling of airway smooth muscle contraction.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Peroxisome proliferator-activated receptor gamma ligands attenuate immunological symptoms of experimental allergic asthma.
Asthma is characterized by a predominant T(H)2 type immune response to airborne allergens. Controlling T(H)2 cell function has been proposed as therapy for this disease. We show here that ligands for the nuclear receptor peroxisome proliferator activated receptor (PPAR)gamma significantly reduced the immunological symptoms of allergic asthma in a murine model of this disease. A PPARgamma ligand, 15-deoxy-delta(12,14)-prostaglandin J(2), significantly inhibited production of the T(H)2 type cytokine IL-5 from T cells activated in vitro. More importantly, in a murine model of allergic asthma, mice treated orally with ciglitazone, a potent synthetic PPARgamma ligand, had significantly reduced lung inflammation and mucous production following induction of allergic asthma. T cells from these ciglitazone treated mice also produced less IFNgamma, IL-4, and IL-2 upon rechallenge in vitro with the model allergen. Our results suggest that ligands for PPARgamma may be effective treatments for asthmatic patients.
Administration, Oral↗
Differential regulation of NFAT and SRF by the B cell receptor via a PLCgamma-Ca(2+)-dependent pathway.
NFAT and SRF are important in the regulation of proliferation and cytokine production in lymphocytes. NFAT activation by the B cell receptor (BCR) occurs via the PLCgamma-Ca(2+)-calcineurin pathway, however how the BCR activates SRF is unclear. We show here that like NFAT, BCR regulation of SRF occurs via an Src-Syk-Tec-PLCgamma-Ca(2+) (Lyn-Syk-Btk-PLCgamma-Ca(2+)) pathway. However, SRF responds to lower Ca(2+) and is less dependent on IP(3)R expression than NFAT. Ca(2+)-regulated calcineurin plays a partial role in SRF activation, in combination with diacylglycerol (DAG), while is fully required for NFAT activation. Signals from the DAG effectors protein kinase C, Ras and Rap1, and the downstream MEK-ERK pathway are required for both SRF and NFAT; however, NFAT but not SRF is dependent on JNK signals. Both SRF and NFAT were also dependent on Rac, Rho, CDC42 and actin. Finally, we show that Ca(2+) is not required for ERK activation, but instead for its association with nuclear areas of the cell. These data suggest that combinatorial assembly of signaling pathways emanating from the BCR differentially regulate NFAT and SRF, to activate gene expression.
Animals↗
CD28-dependent HIV-1 transcription is associated with Vav, Rac, and NF-kappa B activation.
Activation of HIV-1-infected T cells through the T cell receptor and costimulatory molecule CD28 induces proviral transcription; however, the mechanism behind this enhanced virus expression is unknown. Jurkat T cells and primary CD4+ T cells expressing a CD8 alpha/CD28 chimeric receptor containing a mutation at tyrosine 200 in the cytoplasmic tail were unable to fully induce HIV-1 proviral transcription in response to CD8 alpha/28 receptor cross-linking in comparison to CD28 costimulation. The loss of transactivation seen with the mutant chimeric receptor correlated with a decrease in Vav tyrosine phosphorylation. CD28-dependent activation of HIV-1 transcription also required the GTPase activity of Rac1, which was not activated during costimulation with the mutated receptor. Furthermore, the mutated receptor was unable to induce NF-kappa B DNA binding or transactivation, as demonstrated by electromobility shift assays and HIV-1 long terminal repeat and NF-kappa B-dependent reporter constructs. These studies show that signaling events initiated by tyrosine 200 of CD28 are required for efficient expression of HIV-1 transcription in activated T cells.
Antigens, CD↗
Attenuation of immunological symptoms of allergic asthma in mice lacking the tyrosine kinase ITK.
Allergic asthma patients manifest airway inflammation and some show increases in eosinophils, T(H)2 cells, and cytokines, increased mucous production in the lung, and elevated serum IgE. This T(H)2-type response suggests a prominent role for T(H)2 cells and their cytokines in the pathology of this disease. The Tec family nonreceptor tyrosine kinase inducible T cell kinase (ITK) has been shown to play a role in the differentiation and/or function of T(H)2-type cells, suggesting that ITK may represent a good target for the control of asthma. Using a murine model of allergic asthma, we show here that ITK is involved in the development of immunological symptoms seen in this model. We show that mice lacking ITK have drastically reduced lung inflammation, eosinophil infiltration, and mucous production following induction of allergic asthma. Notably, T cell influx into the lung was reduced in mice lacking ITK. T cells from ITK(-/-) mice also exhibited reduced proliferation and cytokine secretion, in particular IL-5 and IL-13, in response to challenge with the allergen OVA, despite elevated levels of total IgE and increased OVA-specific IgE responses. Our results suggest that the tyrosine kinase ITK preferentially regulates the secretion of the T(H)2 cytokines IL-5 and IL-13 and may be an attractive target for antiasthmatic drugs.
Allergens↗
hCMV IE Promoter is responsive to B cell receptor signaling in a Bruton's tyrosine kinase dependent manner: caution in their use as internal controls.
Explore the source record for details and available documents.
Agammaglobulinaemia Tyrosine Kinase↗