Ted T. Kung

New IL-17 Family Members Promote Th1 or Th2 Responses in the Lung: In Vivo Function of the Novel Cytokine IL-25

We have biologically characterized two new members of the IL-17 cytokine family: IL-17F and IL-25. In contrast to conventional in vitro screening approaches, we have characterized the activity of these new molecules by direct in vivo analysis and have compared their function to that of other IL-17 family members. Intranasal administration of adenovirus expressing IL-17, IL-17C, or IL-17F resulted in bronchoalveolar lavage neutrophilia and inflammatory gene expression in the lung. In contrast, intranasal administration of IL-25-expressing adenovirus or IL-25 protein resulted in the production of IL-4, IL-5, IL-13, and eotaxin mRNA in the lung and marked eosinophilia in the bronchoalveolar lavage and lung tissue. Mice given intranasal IL-25 also developed epithelial cell hyperplasia, increased mucus secretion, and airway hyperreactivity. IL-25 gene expression was detected following Aspergillus and Nippostrongylus infection in the lung and gut, respectively. IL-25-induced eosinophilia required IL-5 and IL-13, but not IL-4 or T cells. Following IL-25 administration, the IL-5+ staining cells were CD45R/B220+, Thy-1+/−, but were NK1.1-, Ly-6G(GR-1)-, CD4-, CD3-, and c-kit-negative. γ-common knockout mice did not develop eosinophilia in response to IL-25, nor were IL-5+ cells detected. These findings suggest the existence of a previously unrecognized cell population that may initiate Th2-like responses by responding to IL-25 in vivo. Further, these data demonstrate the heterogeneity of function within the IL-17 cytokine family and suggest that IL-25 may be an important mediator of allergic disease via production of IL-4, IL-5, IL-13, and eotaxin.

Characterization of a murine model of allergic pulmonary inflammation.

Pulmonary inflammation with eosinophil (EOs) infiltration is a prominent feature of allergic respiratory diseases such as asthma. In order to study the cellular response during the disease development, an animal model of IgE-mediated pulmonary inflammation with characteristic eosinophilia is needed. We developed a method for inducing severe pulmonary eosinophilia in the mouse and also studied the numbers of EOs in blood and bone marrow and the response to corticosteroid treatment. Animals were sensitized with alum-precipitated ovalbumin (OVA) and challenged with aerosolized OVA 12 days later when serum IgE levels were significantly elevated. Four to eight hours after challenge there were moderate increases in the number of EOs in the bone marrow and peripheral blood, but only a few EOs were observed in the lung tissue and in bronchoalveolar lavage (BAL) fluid. Twenty-four hours after challenge, there was a marked reduction of EOs in bone marrow, while the number of EOs peaked in the perivascular and peribronchial regions of the lung. Forty-eight hours after challenge, the highest number of EOs was found in the BAL fluid, making up > 80% of all cells in that compartment. The high levels of EOs in the lung tissue and BAL fluid lasted for 2-3 days and was followed by a more moderate but persistent eosinophilia for another 10 days. Nonsensitized animals showed no significant changes in the number of EOs in BAL fluid, lungs, blood or bone marrow. Histopathological evaluation also revealed epithelial damage, excessive mucus in the lumen and edema in the submucosa of the airways.(ABSTRACT TRUNCATED AT 250 WORDS)

ACTH accelerates recovery of neuromuscular function following crushing of peripheral nerve

Adrenocorticotropin (ACTH)-treated adrenalectomized rats subjected to crush denervation recover sensation and functional movement sooner than saline-treated rats. Axonal regeneration is accelerated, the number of large endplates and the frequency of preterminal branching are increased. ACTH has no effect on either intact or denervated muscles. The ameliorative action of ACTH during regeneration is apparently neurogenic and independent of corticoids.

A novel, orally active CXCR1/2 receptor antagonist, sch527123, inhibits neutrophil recruitment, mucus production, and goblet cell hyperplasia in animal models of pulmonary inflammation

Sch527123 [2-hydroxy-N,N-dimethyl-3-[[2-[[1(R)-(5-methyl-2-furanyl)propyl]amino]-3,4-dioxo-1-cyclobuten-1-yl]amino]ben-zamide] is a potent, selective antagonist of the human CXCR1 and CXCR2 receptors (Gonsiorek et al., 2007). Here we describe its pharmacologic properties at rodent CXCR2 and at the CXCR1 and CXCR2 receptors in the cynomolgus monkey, as well as its in vivo activity in models demonstrating prominent pulmonary neutrophilia, goblet cell hyperplasia, and mucus production. Sch527123 bound with high affinity to the CXCR2 receptors of mouse (Kd = 0.20 nM), rat (Kd = 0.20 nM), and cynomolgus monkey (Kd = 0.08 nM) and was a potent antagonist of CXCR2-mediated chemotaxis (IC50 ∼3–6 nM). In contrast, Sch527123 bound to cynomolgus CXCR1 with lesser affinity (Kd = 41 nM) and weakly inhibited cynomolgus CXCR1-mediated chemotaxis (IC50 ∼1000 nM). Oral treatment with Sch527123 blocked pulmonary neutrophilia (ED50 = 1.2 mg/kg) and goblet cell hyperplasia (32–38% inhibition at 1–3 mg/kg) in mice following the intranasal lipopolysaccharide (LPS) administration. In rats, Sch527123 suppressed the pulmonary neutrophilia (ED50 = 1.8 mg/kg) and increase in bronchoalveolar lavage (BAL) mucin content (ED50 =<0.1 mg/kg) induced by intratracheal (i.t.) LPS. Sch527123 also suppressed the pulmonary neutrophilia (ED50 = 1.3 mg/kg), goblet cell hyperplasia (ED50 = 0.7 mg/kg), and increase in BAL mucin content (ED50 = <1 mg/kg) in rats after i.t. administration of vanadium pentoxide. In cynomolgus monkeys, Sch527123 reduced the pulmonary neutrophilia induced by repeat bronchoscopy and lavage (ED50 = 0.3 mg/kg). Therefore, Sch527123 may offer benefit for the treatment of inflammatory lung disorders in which pulmonary neutrophilia and mucus hypersecretion are important components of the underlying disease pathology.

Inhibition of Pulmonary Eosinophilia and Hyperreactivity by Antibodies to lnterleukin-5

Eosinophils infiltrate into the lungs during asthma and may cause the damage associated with pulmonary inflammation. In allergic animal models, antibodies to interleukin (IL)-5 inhibit pulmonary eosinophilia, tissue damage and hyperreactivity. Sch 55700, a humanized antibody against human IL-5, inhibits eosinophilia in these models with an extended biological duration. On the basis of this dosing regimen and the humanized nature of Sch 55700, it is anticipated that the host response leading to tolerance would be minimized.

Induction of lysosomal and plasma membrane-bound sialidases in human T-cells via T-cell receptor

Among the three isoenzymes of neuraminidase (Neu) or sialidase, Neu-1 has been suggested to be induced by cell activation and to be involved in IL (interleukin)-4 biosynthesis in murine T-cells. In the present study, we found that antigen-induced airway eosinophilia, a typical response dependent on Th2 (T-helper cell type 2) cytokines, as well as mRNA expression of Th2 cytokines, including IL-4, are suppressed in Neu-1-deficient mice, thereby demonstrating the in vivo role of murine Neu-1 in regulation of Th2 cytokines. To elucidate the roles of various sialidases in human T-cell activation, we investigated their tissue distribution, gene induction and function. Neu-1 is the predominant isoenzyme at the mRNA level in most tissues and cells in both mice and humans, including T-cells. T-cells also have significant levels of Neu-3 mRNAs, albeit much lower than those of Neu-1, whereas the levels of Neu-2 mRNAs are minimal. In human T-cells, both Neu-1 and Neu-3 mRNAs are significantly induced by T-cell-receptor stimulation, as is sialidase activity against 4-methylumbelliferyl- N -acetylneuramic acid (a substrate for both Neu-1 and Neu-3) and the ganglioside G(D1a) [NeuAcalpha2-3Galbeta1-3GalNAcbeta1-4(NeuAcalpha2-3)Galbeta1-4Glcbeta1-cer] (a substrate for Neu-3, but not for Neu-1). The expression of the two sialidase genes may be under differential regulation. Western blot analysis and enzymic comparison with recombinant sialidases have revealed that Neu-3 is induced as a major isoform in activated cells. The induction of Neu-1 and Neu-3 in T-cells is unique. In human monocytes and neutrophils stimulated with various agents, the only observation of sialidase induction has been by IL-1 in neutrophils. Functionally, a major difference has been observed in Jurkat T-cell lines over-expressing Neu-1- and Neu-3. Upon T-cell receptor stimulation, IL-2, interferon-gamma, IL-4 and IL-13 are induced in the Neu-1 line, whereas in the Neu-3 line the same cytokines are induced, with the exception of IL-4. Taken together, these results suggest an important immunoregulatory role for both Neu-1 and Neu-3 in humans.

Inhibition of experimental acute pulmonary inflammation by pirfenidone.

Pirfenidone, a putative tumor necrosis factor-alpha (TNF-alpha) inhibitor, has recently gained recognition for its therapeutic use in the treatment of idiopathic pulmonary fibrosis. As pulmonary fibrosis may be the result of lung inflammatory processes, we examined the anti-inflammatory potential of pirfenidone in several models of acute pulmonary inflammation. In antigen-induced allergic paradigms, 24 h after antigen challenge, sensitized mice or guinea pigs develop a prominent pulmonary inflammation, reflected by a significant increase in the number of recoverable bronchoalveolar lavage (BAL) total cells and eosinophils. In both species, the pretreatment of animals with pirfenidone (10 and 30 mg/kg) resulted in a dose-dependent inhibition of the antigen-induced pulmonary inflammation, which was reflected by a significant decrease in the BAL eosinophils and total cells by the 30 mg/kg dose. In a non-allergic model of pulmonary inflammation, rats challenged with intratracheal LPS develop a significant increase in BAL neutrophils and total cells, along with significant increases in TNF-alpha and IL-6. Pretreatment with pirfenidone (3 and 30 mg/kg) showed a dose-dependent inhibition of the LPS-induced pulmonary inflammation, reflected by a significant decrease in the number of BAL total and neutrophilic cells at both the 3 and 30 mg/kg dose. However, pirfenidone had no effect on the peak BAL levels of TNF-alpha. In contrast, pirfenidone significantly inhibited BAL levels of IL-6. In summary, we have shown that pirfenidone can inhibit allergic and non-allergic inflammatory cell recruitment and that its pulmonary anti-inflammatory activity is independent of TNF-alpha inhibition.

Inhibition of pulmonary eosinophilia and airway hyperresponsiveness in allergic mice by rolipram: involvement of endogenously released corticosterone and catecholamines

This study investigates the role of adrenal‐derived catecholamines and corticosterone on the inhibition by rolipram, a phosphodiesterase (PDE)‐4 inhibitor, of pulmonary eosinophilia and airway hyperresponsiveness (AHR) in allergic mice. The following experimental groups were studied in mice sensitized and challenged with ovalbumin (OVA): normal, adrenalectomized, propranolol (β‐adrenoceptor antagonist) and metyrapone (corticosterone synthesis inhibitor) treated. These interventions were studied both in the absence and in the presence of rolipram. Eosinophil numbers in the bronchoalveolar lavage (BAL) and AHR to methacholine were measured 24 h after OVA challenge. Treatment of sensitized mice with rolipram (0.3–10 mg kg−1, p.o.), inhibited pulmonary eosinophilia and the AHR to methacholine in OVA‐challenged mice. Adrenalectomy increased the number of eosinophils in the BAL of OVA‐challenged mice but had no effect on AHR to methacholine. Adrenalectomy attenuated both the rolipram‐induced inhibition of BAL eosinophilia and AHR to methacholine in OVA challenged mice. Propranolol (10 mg kg−1, p.o.) had no effect on the inhibition of eosinophilia by rolipram but attenuated the inhibition of AHR to methacholine in OVA challenged mice. On the other hand, metyrapone (10 mg kg−1, p.o.) attenuated the inhibition of eosinophilia by rolipram but had no effect on the inhibition of AHR to methacholine in OVA challenged mice. Metyrapone‐treatment alone increased the number of eosinophils in the BAL of OVA‐challenged mice. These results identify an important role for adrenal‐derived catecholamines and corticosterone on the inhibition of pulmonary eosinophilia and AHR by rolipram in allergic mice.

IN VITRO AND IN VIVO IMMUNOPHARMACOLOGICAL PROFILE OF SCH 40120

Sch 40120 (10-(3-chlorophenyl) - 6,8,9,10- tetrahydrobenzo [b] [1,8] naphthyridin-5 (7H)-one) is a leukotriene inhibitor that is also a potent inhibitor of acute inflammatory responses in rodent systems. In the present study, we have evaluated the effects of this drug on immune function as well as its activity in models of immune mediated chronic inflammatory disease. Sch 40120 was particularly effective in suppressing T cell proliferative responses in vitro. Antigen-specific and poly-clonally-induced in vitro antibody responses were also inhibited by the drug. However, the in vivo potency of Sch 40120 in suppressing immune responses and in inhibiting the pathological changes seen in rodent models of autoimmune disease (EAE and adjuvant arthritis) was somewhat less than that previously observed in models of acute inflammation. Nevertheless, the spectrum of activities exhibited by Sch 40120 suggests that it will be particularly useful in the treatment of psoriasis where T lymphocytes have been implicated in the development of disease and leukotrienes appear to have a role in the persistence of psoriatic plaques.