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John J Ryan

Publications and source records attributed to John J Ryan.

At least 19 recordsLinked to original sources
PubMed2006

Stat5 expression is required for IgE-mediated mast cell function.

The mast cell (MC) inflammatory response is now linked not only to atopy, but also to arthritis, multiple sclerosis, heart disease, and resistance to bacterial infection. In the current study, we demonstrate that the signal transducer and activator of transcription 5 (Stat5) is rapidly activated by IgE cross-linkage, and that its expression is critical to the MC response. Stat5-deficient (Stat5KO) MC demonstrated a significant decrease in IgE-mediated degranulation, leukotriene B4 production, cytokine secretion, and survival signals. The defect in cytokine production may be caused by decreased cytokine mRNA stability. Stat5KO MC-induced cytokine mRNAs normally following IgE cross-linkage, but these mRNAs were not sustained over time and were degraded at twice the rate observed in WT cells. Interestingly, the RNA destabilizing protein tristetraprolin was induced following IgE cross-linkage in Stat5KO but not wild-type cells. Moreover, reducing tristetraprolin expression via short hairpin RNA transfection significantly increased IL-13 production in Stat5KO MC. Our work demonstrates that Stat5 is a critical factor in IgE-induced MC activation, acting in part via posttranscriptional control of cytokine mRNA stability. These data have a direct impact on MC-associated inflammatory and autoimmune diseases.

Animals↗
PubMed2006

Interleukin-10 induces apoptosis in developing mast cells and macrophages.

Interleukin (IL)-10 is a potent immunoregulatory cytokine capable of inhibiting the inflammatory response. As mast cells and macrophages are central effectors of inflammation, we investigated the effects of IL-10 on mast cell and macrophage development from mouse bone marrow progenitors. Bone marrow cells were cultured in IL-3 + stem cell factor (SCF), giving rise to mixed populations of mast cells and macrophages. The addition of IL-10 greatly decreased the expansion of bone marrow progenitor cells through a mechanism requiring signal tranducer and activator of transcription-3 expression. The inhibitory effects were a result of the induction of apoptosis, which occurred with caspase-3 activation and reduced mitochondrial membrane potential. Supporting a role for the mitochondrion, bone marrow cells from p53-deficient or Bcl-2 transgenic mice were partly resistant to the effects of IL-10. Further, IL-10 decreased Kit receptor expression and inhibited survival signaling by SCF or IL-3. These data indicate that IL-10 induces an intrinsic, mitochondrial apoptosis cascade in developing mast cells and macrophages through mechanisms involving blockade of growth factor receptor function. The ability of IL-10 to inhibit survival could support immune homeostasis by dampening inflammatory responses and preventing chronic inflammation.

Animals↗
PubMed2006

PCB, PCDD and PCDF residues in fin and non-fin fish products from the Canadian retail market 2002.

Fish products (n=129) available on the Canadian retail market were collected and analyzed for levels of PCBs, PCDDs and PCDFs during the spring of 2002. The collection included samples from eight fish groups (Arctic char, crab, mussels, oysters, salmon, shrimp, tilapia, trout) from the wild and those raised on fish farms, as available. Sample collection included both domestic and imported fish products, however, no significant difference in residue levels was observed between these groups of fish products. Salmon samples were found to contain the highest concentration of sigmaPCBs (geometric mean 12.9 ng/g wet weight), while crab samples had greatest sigmaPCDD/F levels (geometric mean 0.002 ng/g wet weight). The geometric mean of the total toxic equivalents (WHO-TEQ) ranged from 0.06 pg WHO-TEQ/g whole weight in farmed shrimp to 1.1 pg WHO-TEQ/g whole weight in farmed salmon samples. PCB 153, 138, 118 and 101 were the dominant congeners observed in fish product samples studied, while 1,2,3,7,8-pentachlorodibenzodioxin and 2,3,7,8-tetrachlorodibenzofuran contributed the most to total PCDD and PCDF loadings. Lipid content was positively correlated to sigmaPCB levels; however, no relationship between lipid content and sigmaPCDD/F concentrations was established. SigmaPCB levels were below the Canadian guideline value for PCBs in fish and fish products (2000 ng/g). Similarly, 2,3,7,8-TCDD levels in all fish products were below the Canadian guideline value (0.020 ng/g).

Animals↗
PubMed2006

IL-3-mediated TNF production is necessary for mast cell development.

Mouse mast cell development and survival are largely controlled by the cytokines IL-3 and stem cell factor (SCF). We have found that IL-3 stimulation of bone marrow cells induces the production of TNF via a PI3K- and MAPK kinase/ERK-dependent pathway. Specifically, Mac-1-positive cells were responsible for TNF production, which peaked on days 7-10 of culture and decreased rapidly thereafter. The importance of IL-3-induced TNF secretion was demonstrated by the failure of TNF-deficient bone marrow cells to survive for >3 wk when cultured in IL-3 and SCF, a defect that was reversed by the addition of soluble TNF. The development of human mast cells from bone marrow progenitors was similarly hampered by the addition of TNF-blocking Abs. Cell death was due to apoptosis, which occurred with changes in mitochondrial membrane potential and caspase activation. Apoptosis appeared to be due to loss of IL-3 signaling, because TNF-deficient cells were less responsive than their wild-type counterparts to IL-3-mediated survival. In vitro cultured mast cells from TNF-deficient mice also demonstrated reduced expression of the high affinity IgE receptor, which was restored to normal levels by the addition of soluble TNF. Finally, TNF-deficient mice demonstrated a 50% reduction in peritoneal mast cell numbers, indicating that TNF is an important mast cell survival factor both in vitro and in vivo.

Animals↗
PubMed2006

Dioxins: an overview.

This review article summarizes what is known about human health following exposure to dioxins. It is meant primarily for health professionals but was also written with the general public in mind. The need for such an article became apparent to the authors following media inquiries at the time the then Ukraine presidential candidate Victor Yushchenko was deliberately poisoned with the most toxic dioxin, tetrachlorodibenzodioxin or TCDD.

Adipose Tissue↗
PubMed2006

TGFbeta1 induces mast cell apoptosis.

Mast cells are potent effectors of the inflammatory response, playing an important role in atopy, bacterial immunity, and animal models of arthritis, multiple sclerosis, and heart disease. Hence controlling mast cell numbers and responsiveness is essential for preventing inflammatory disease. We demonstrate that the cytokine transforming growth factor (TGF) beta1 is a potent inducer of mast cell apoptosis, a finding that was consistent in cultured mouse bone marrow-derived mast cells, peritoneal mast cells, and human mast cells. Cell death appeared to be caused by TGF-mediated repression of interleukin-3 (IL-3) receptor expression and function, leading to mitochondrial damage and activation of an apoptotic cascade acting via p53 and caspases. Although IL-3 receptor expression was reduced within 1 day of TGFbeta1 stimulation, apoptosis required at least 3 days to occur. This delay in onset is postulated to allow protective mast cell effector functions, protecting the host from infection while preventing the establishment of chronic inflammation. Our data support the theory that TGFbeta1 is an inhibitor of mast cell survival. The widespread expression of TGFbeta1 offers this cytokine as an ideal candidate for control of mast cell homeostasis.

Animals↗
PubMed2006

cis-Terpenones as an effective chemopreventive agent against aflatoxin B1-induced cytotoxicity and TCDD-induced P450 1A/B activity in HepG2 cells.

Aflatoxin B1 (AFB1) is a potent carcinogen, which can significantly increase the risk of hepatocellular carcinoma development through food contamination. In past decades, chemopreventive agents, such as oltipraz and chlorophyllins, have demonstrated that chemo-intervention is an effective approach to reduce hepatotoxicity by AFB1. However, because of the potential adverse effects of these agents, alternative novel mechanism-based chemopreventive agents are needed. We report here that novel cis-terpenones 1-3, which were synthesized as the precursors of natural product analogues in our laboratory, showed promising protective effects against AFB1-induced cytotoxicity in HepG2 cells. Chemo-protection was observed with increasing concentrations of cis-terpenones in the co-treatment of AFB1, and no cytotoxicity was observed with cis-terpenones alone. In addition, cis-terpenones 1-3 at 10 and microM effectively inhibited induced cytochrome P450 1A/1B activity by 50% in HepG2 cells, as indicated by an EROD assay. P450 1A/B is involved in the activation of many pre-carcinogens and is highly inducible in liver cells. These results suggested that novel terpenones 1-3 are candidates for the development of novel mechanism-based chemopreventive agents against AFB1 and other carcinogenic stimuli.

Aflatoxin B1↗
PubMed2005

Impaired FcepsilonRI-dependent gene expression and defective eicosanoid and cytokine production as a consequence of Fyn deficiency in mast cells.

Fyn kinase is a key contributor in coupling FcepsilonRI to mast cell degranulation. A limited macroarray analysis of FcepsilonRI-induced gene expression suggested potential defects in lipid metabolism, eicosanoid and glutathione metabolism, and cytokine production. Biochemical analysis of these responses revealed that Fyn-deficient mast cells failed to secrete the inflammatory eicosanoid products leukotrienes B4 and C4, the cytokines IL-6 and TNF, and chemokines CCL2 (MCP-1) and CCL4 (MIP-1beta). FcepsilonRI-induced generation of arachidonic acid and normal induction of cytokine mRNA were defective. Defects in JNK and p38 MAPK activation were observed, whereas ERK1/2 and cytosolic phospholipase A2 (S505) phosphorylation was normal. Pharmacological studies revealed that JNK activity was associated with generation of arachidonic acid. FcepsilonRI-mediated activation of IkappaB kinase beta and IkappaBalpha phosphorylation and degradation was defective resulting in a marked decrease of the nuclear NF-kappaB DNA binding activity that drives IL-6 and TNF production in mast cells. However, not all cytokine were affected, as IL-13 production and secretion was enhanced. These studies reveal a major positive role for Fyn kinase in multiple mast cell inflammatory responses and demonstrate a selective negative regulatory role for certain cytokines.

Animals↗
PubMed2005

Biological consequences of trinuclear platinum complexes: comparison of [[trans-PtCl(NH3)2]2mu-(trans-Pt(NH3)2(H2N(CH2)6-NH2)2)]4+ (BBR 3464) with its noncovalent congeners.

[[trans-PtCl(NH(3))(2)](2)mu-(trans-Pt(NH(3))(2)(H(2)N(CH(2))(6)NH(2))(2))](4+) (BBR 3464) is a 4+ cationic trinuclear platinum drug that has undergone phase II clinical trials in the treatment of ovarian and lung cancers. The chemical structure of BBR 3464 is distinct from that of clinically used agents such as cisplatin and oxaliplatin. As a consequence, the modes of DNA binding and the structures of BBR 3464 DNA adducts are also structurally distinct from those formed by cisplatin and oxaliplatin. Previous chemical and spectroscopic measurements on BBR 3464 had elucidated a significant noncovalent contribution to DNA binding. To examine this effect further, the biological activity of two BBR 3464 analogs that bind DNA only through noncovalent interactions was investigated in this study, and their cellular effects were compared with those caused by the "parent" drug. The compounds were [[trans-PtL(NH(3))(2)](2)mu-(trans-Pt(NH(3))(2)(H(2)N(CH(2))(6)NH(2))(2))](n+), with L = NH(3), n = 6 for compound I, and L = H(2)N(CH(2))(6)NH(3), n = 8 for compound II. All compounds induce caspase-dependent apoptosis in both primary mast cells and transformed mastocytomas, although with a smaller IC(50) value in the transformed cells. In cells deficient in either the tumor suppressor proteins p53 or Bax, apoptosis was least affected in the case of II, but in all cases the effect of p53 deficiency was greater than that of Bax. Surprisingly, cellular uptake was actually enhanced for the more highly charged compounds, resulting in significant (micromolar) cyotoxicity for II. Cellular accumulation was enhanced in mastocytomas over primary mast cells, suggesting a mechanism for enhancement of tumor cell selectivity.

Animals↗
PubMed2005

IFN-gamma induces apoptosis in developing mast cells.

Mast cells are critical effectors of allergic disease, and are now implicated in immune responses observed in arthritis, multiple sclerosis, and heart disease. Because of their role in inflammation, understanding how mast cells develop is of clinical importance. In this study we determined the effects of IFN-gamma on mast cell survival. Using in vitro culture of bone marrow cells in IL-3 plus stem cell factor, we found that the addition of IFN-gamma induced apoptosis, as exhibited by the presence of subdiploid DNA and caspase activation. IFN-gamma-mediated apoptosis was Stat1-dependent, and was accompanied by loss of mitochondrial membrane potential. Apoptosis was reduced in cultures of bone marrow cells derived from p53- or Bax-deficient mice, as well as H2K-Bcl-2 transgenic mice. IFN-gamma hyperresponsiveness has been shown to result in inflammatory disease and death in mice lacking the regulatory protein suppressor of cytokine signaling (SOCS)-1. Bone marrow cells from SOCS-1 knockout (KO) mice failed to give rise to viable mast cells after culture in IL-3 plus stem cell factor, with profound apoptosis occurring as the cultures matured. However, bone marrow cells lacking both SOCS-1 and IFN-gamma survived normally. This in vitro defect in mast cell development was recapitulated in vivo. SOCS-1 KO mice demonstrated a 67% decrease in peritoneal mast cell numbers relative to wild-type mice, a deficiency that was reversed in SOCS-1/IFN-gamma KO mice. These data demonstrate the potent regulatory effects of IFN-gamma on mast cell survival and show that this cytokine can elicit mast cell death in vitro and in vivo.

Animals↗
PubMed2005

Kidney transplantation and gender disparity.

Gender inequity in access to hemodialysis and kidney transplantation has created a public health crisis in the US. Women have a lower chance of receiving hemodialysis and kidney transplant than men, but they constitute the majority of living kidney donors. Research has shown that economic factors such as greater income of men may encourage females to be donors; while gender-bias on part of physicians or institutions, lack of social support networks and differences in health-seeking behaviors compared to men are cited as reasons for this imbalance. We suggest various strategies to improve participation of women in the transplant process by education; raising awareness by publishing gender-specific data for dialysis and transplant centers; education and workshops to eliminate gender-bias within institutions and health-care providers and establishment of gender-specific support groups. Transplant teams that are more sensitive to the social complexities of women's lives may lead to increased understanding of the effects of renal disease and indicate measures that need to be in place in order to address this gender disparity in the treatment of renal failure. Research needs to be done to elucidate the underlying medical, societal or psychological processes that lead to gender bias in the field of kidney transplantation.

Female↗
PubMed2005

TGF-beta 1 inhibits mast cell Fc epsilon RI expression.

Mast cell activation through the high affinity IgE receptor (FcepsilonRI) is a critical component of atopic inflammation. The cytokine TGF-beta1 has been shown to inhibit IgE-dependent mast cell activation, possibly serving to dampen mast cell-mediated inflammatory responses. We present proof that TGF-beta1 inhibits mast cell FcepsilonRI expression through a reversible pathway that diminishes protein, but not mRNA, expression of the FcepsilonRI subunit proteins alpha, beta, and gamma. The stability of the expressed proteins and the assembled cell surface complex was unaltered by TGF-beta1 treatment. However, TGF-beta1 decreased the rate of FcepsilonRI beta-chain synthesis, arguing that this inhibitory cytokine exerts its effects at the level of mRNA translation. TGF-beta1 consistently diminished FcepsilonRI expression on cultured human or mouse mast cells as well as freshly isolated peritoneal mast cells. The related cytokines, TGF-beta2 and TGF-beta3, had similar effects. We propose that TGF-beta1 acts as a negative regulator of mast cell function, in part by decreasing FcepsilonRI expression.

Animals↗
PubMed2005

TGF-beta1 inhibits late-stage mast cell maturation.

OBJECTIVE: The objective of this study was to determine the effects of transforming growth factor (TGF)-beta1 on mast cell development. MATERIALS AND METHODS: Mast cells were cultured from mouse bone marrow in interleukin (IL)-3 + stem cell factor, in the presence or absence of TGF-beta1. We assessed mast cell development by measuring the expression of kit, T1/ST2, FcvarepsilonRI, and Fcgamma receptors. Cell morphology was determined by histochemical staining. Alterations in FcvarepsilonRI subunit expression were measured by Western blot analysis. Adoptive transfer of cultured mast cells into mast cell-deficient W/W(v) mice was used to determine if the in vivo environment could reverse the inhibitory effects of TGF-beta1. RESULTS: TGF-beta1 decreased FcvarepsilonRI, c-kit, T1/ST2, and FcgammaR expression, and inhibited granule formation in developing mast cells. Accessory cells were not required for this inhibition. Smad3 deficiency did not alter the response of bone marrow cells to TGF-beta1. TGF-beta1 inhibited expression of the FcvarepsilonRI alpha subunit protein, without decreasing beta or gamma proteins. Mast cells derived in the presence of TGF-beta1 were functionally impaired, as IgE-mediated cytokine secretion was greatly reduced. The changes in granule formation and surface antigen expression were long-standing, as they were not reversed by transfer to W/W(v) mice. CONCLUSIONS: TGF-beta1 may contribute to mast cell homeostasis by inhibiting maturation from bone marrow precursors. The effects of TGF-beta1 result in greatly diminished expression of cell surface markers, reduced granulation, and lack of responsiveness to IgE-mediated activation. Thus TGF-beta1 can serve as a potent and multifunctional regulator of mast cell maturation.

Adoptive Transfer↗
PubMed2005

The role of Src family kinases in mast cell effector function.

Src family protein tyrosine kinases (SrcPTK) play a central role in immunoglobulin E (IgE)-mediated activation of mast cells. Functional coupling of the high-affinity IgE receptor (FcepsilonRI) is initiated by the SrcPTK family member, Lyn, through an antigen aggregation-dependent transphosphorylation. Because Lyn is the 'initiating' kinase, an essential role in mast cell effector function was conferred. Recent studies challenge this view. Evidence demonstrating that Lyn kinase is dispensable for mast cell degranulation is now available. In contrast, another SrcPTK family member, Fyn, is required for degranulation and cytokine production. New studies, on mast cells expressing FcepsilonRIbeta ITAM mutants, show that the loss of Lyn interaction with FcepsilonRI has only a modest inhibitory effect on mast cell degranulation and an enhancing effect on lymphokine production, although many of the biochemical signals (including FcepsilonRI phosphorylation) were significantly impaired. In vivo studies on Lyn-null mice also demonstrated that this kinase is a negative regulator of IgE production and anaphylaxis, whereas Fyn kinase is required for anaphylaxis but not for IgE production. Collectively, these studies argue that sustained Lyn kinase activity negatively regulates mast cell responses. This suggests the possible existence of Lyn polymorphisms that may contribute in allergic disease.

Animals↗
PubMed2004

Autocrine actions of macrophage-derived catecholamines on interleukin-1 beta.

Previous studies indicate that norepinephrine and epinephrine modulate production of interleukin-1(beta) (IL-1(beta)) by activated macrophages, but it is not known if macrophage-derived catecholamines affect IL-1(beta). In this study, recruited peritoneal macrophages from CBA/J female mice were activated with lipopolysaccharide (LPS) and treated with vehicle or adrenergic receptor antagonists for 24 h. Extracellular and intracellular levels of IL-1(beta) were measured with ELISA. Treatment with the beta-adrenergic receptor antagonists propranolol or ICI 118,551 increased LPS-induced production of IL-1(beta), whereas treatment with the alpha-adrenergic antagonists phentolamine or yohimbine decreased IL-1(beta). These findings demonstrate that adrenergic receptor antagonists unmask autocrine actions of macrophage-derived catecholamines on IL-1(beta) that may influence the inflammatory response.

Adjuvants, Immunologic↗
PubMed2004

Negative regulation of immunoglobulin E-dependent allergic responses by Lyn kinase.

A role for Lyn kinase as a positive regulator of immunoglobulin (Ig)E-dependent allergy has long been accepted. Contrary to this belief, Lyn kinase was found to have an important role as a negative regulator of the allergic response. This became apparent from the hyperresponsive degranulation of lyn-/- bone marrow-derived mast cells, which is driven by hyperactivation of Fyn kinase that occurs, in part, through the loss of negative regulation by COOH-terminal Src kinase (Csk) and the adaptor, Csk-binding protein. This phenotype is recapitulated in vivo as young lyn-/- mice showed an enhanced anaphylactic response. In vivo studies also demonstrated that as lyn-/- mice aged, their serum IgE increased as well as occupancy of the high affinity IgE receptor (FcepsilonRI). This was mirrored by increased circulating histamine, increased mast cell numbers, increased cell surface expression of the high affinity IgE receptor (FcepsilonRI), and eosinophilia. The increased IgE production was not a consequence of increased Fyn kinase activity in lyn-/- mice because both lyn-/- and lyn-/- fyn-/- mice showed high IgE levels. Thus, lyn-/- mice and mast cells thereof show multiple allergy-associated traits, causing reconsideration of the possible efficacy in therapeutic targeting of Lyn in allergic disease.

Animals↗