Responses of Plasma Acetate Metabolism to Hop (Humulus lupulus L.) in Sheep

An isotope dilution method using [1-13C]sodium (Na) acetate was conducted to determine the effect of feeding hop (Humulus lupulus L.) residues on plasma acetate metabolism in six adult crossbred sheep. The sheep were fed 63 g/kg BW0.75/d of either mixed hay (MH-diet) of orchardgrass (Dactylis glomerata L.) and reed canarygrass (Phalaris arundinacea L.) at a 60:40 ratio or MH-diet and hop-residues (Hop-diet) at 85:15 ratio with a crossover design for each of 3 week period. The isotope dilution method using single injection of [1-13C]Na acetate was performed thrice; before feeding (BF), 2 h after feeding (2F) and 4 h after feeding (4F), on the 21st day of each dietary treatment. Plasma acetate concentration tended to increase (P= 0.06) and turnover rate was numerically higher (P= 0.16) for MH-diet than Hop-diet. Plasma glucose, NEFA, VFA and lactic acid concentrations were similar between dietary treatments. In both the diets, although plasma concentration of acetate did not change, turnover rate increased significantly (P= 0.02) 2F than BF. Hop-residues did not show any negative impacts on acetate metabolism as well as physiology of animals in the present experimental conditions, hence thereby it could be used as an alternative to MH-diet for rearing sheep

Molecular targeting of hepatocyte growth factor by an antagonist, NK4, in the treatment of rheumatoid arthritis

INTRODUCTION: Hepatocyte growth factor (HGF) is a potent proangiogenic molecule that induces neovascularization. The HGF antagonist, NK4, competitively antagonizes HGF binding to its receptor. In the present study, we determined the inhibitory effect of NK4 in a rheumatoid arthritis (RA) model using SKG mice. METHODS: Arthritis was induced in SKG mice by a single intraperitoneal injection of β-glucan. Recombinant adenovirus containing NK4 cDNA (AdCMV.NK4) was also injected intravenously at the time of or 1 month after β-glucan injection. Ankle bone destruction was examined radiographically. The histopathologic features of joints were examined using hematoxylin and eosin and immunohistochemical staining. Enzyme-linked immunosorbent assays were used to determine the serum levels of HGF, interferon γ (IFN-γ, interleukin 4 (IL-4) and IL-17 production by CD4(+ )T cells stimulated with allogeneic spleen cells. RESULTS: The intravenous injection of AdCMV.NK4 into SKG mice suppressed the progression of β-glucan-induced arthritis. Bone destruction was also inhibited by NK4 treatment. The histopathologic findings of the ankles revealed that angiogenesis, inflammatory cytokines and RANKL expression in synovial tissues were significantly inhibited by NK4 treatment. Recombinant NK4 (rNK4) proteins inhibited IFN-γ, IL-4 and IL-17 production by CD4(+ )T cells stimulated with allogeneic spleen cells. CONCLUSIONS: These results indicate that NK4 inhibits arthritis by inhibition of angiogenesis and inflammatory cytokine production by CD4(+ )T cells. Therefore, molecular targeting of angiogenic inducers by NK4 can potentially be used as a novel therapeutic approach for the treatment of RA

Pharmacokinetic modeling of hepatocyte growth factor in experimental animals and humans

Hepatocyte growth factor (HGF) is under development for treatment of renal failure. This study was designed to clarify changes in HGF pharmacokinetics in renal failure and to establish a pharmacokinetic model applicable to single and repeated doses. The plasma concentration profile in mice with glycerol-induced acute renal failure was similar to that in normal mice, indicating a minimal contribution of kidney to systemic clearance of HGF. Nevertheless, accumulation of fluorescein-4-isocyanate-labeled HGF in renal tubules in both cases suggests the occurrence of efficient endocytosis of HGF in kidney. A pharmacokinetic model including plasma and liver compartments was constructed, incorporating both high- and low-affinity receptors for association and subsequent endocytosis of HGF because HGF is eliminated via specific receptor c-Met and heparin-like substance. The model well explained the plasma concentration profiles at all doses examined after bolus injection in animals and humans, and those during infusion in rodents. It includes externalization of receptors, which is negatively regulated by HGF, and can explain the gradual increase in trough concentration during repeated dosing in monkeys. Overall pharmacokinetic profiles of HGF are governed by at least two receptors and are well described by this pharmacokinetic model, which should assist in safe management of clinical trials. © 2012 Wiley Periodicals, Inc

Astrocytic dysfunction induced by ABCA1 deficiency causes optic neuropathy

Astrocyte abnormalities have received great attention for their association with various diseases in the brain but not so much in the eye. Recent independent genome-wide association studies of glaucoma, optic neuropathy characterized by retinal ganglion cell (RGC) degeneration, and vision loss found that single-nucleotide polymorphisms near the ABCA1 locus were common risk factors. Here, we show that Abca1 loss in retinal astrocytes causes glaucoma-like optic neuropathy in aged mice. ABCA1 was highly expressed in retinal astrocytes in mice. Thus, we generated macroglia-specific Abca1-deficient mice (Glia-KO) and found that aged Glia-KO mice had RGC degeneration and ocular dysfunction without affected intraocular pressure, a conventional risk factor for glaucoma. Single-cell RNA sequencing revealed that Abca1 deficiency in aged Glia-KO mice caused astrocyte-triggered inflammation and increased the susceptibility of certain RGC clusters to excitotoxicity. Together, astrocytes play a pivotal role in eye diseases, and loss of ABCA1 in astrocytes causes glaucoma-like neuropathy

Ability of the Met kinase inhibitor crizotinib and new generation EGFR inhibitors to overcome resistance to EGFR inhibitors

Purpose: Although EGF receptor tyrosine kinase inhibitors (EGFR-TKI) have shown dramatic effects against EGFR mutant lung cancer, patients ultimately develop resistance by multiple mechanisms. We therefore assessed the ability of combined treatment with the Met inhibitor crizotinib and new generation EGFR-TKIs to overcome resistance to first-generation EGFR-TKIs. Experimental Design: Lung cancer cell lines made resistant to EGFR-TKIs by the gatekeeper EGFR-T790M mutation, Met amplification, and HGF overexpression and mice with tumors induced by these cells were treated with crizotinib and a new generation EGFR-TKI. Results: The new generation EGFR-TKI inhibited the growth of lung cancer cells containing the gatekeeper EGFRT790M mutation, but did not inhibit the growth of cells with Met amplification or HGF overexpression. In contrast, combined therapy with crizotinib plus afatinib or WZ4002 was effective against all three types of cells, inhibiting EGFR and Met phosphorylation and their downstream molecules. Crizotinib combined with afatinib or WZ4002 potently inhibited the growth of mouse tumors induced by these lung cancer cell lines. However, the combination of high dose crizotinib and afatinib, but not WZ4002, triggered severe adverse events. Conclusions: Our results suggest that the dual blockade of mutant EGFR and Met by crizotinib and a new generation EGFR-TKI may be promising for overcoming resistance to reversible EGFR-TKIs but careful assessment is warranted clinically. © 2013 Nanjo et al

CpG Oligodeoxynucleotides as a Future Vaccine for Allergic Diseases

An astounding feature of the DNA sequences termed CpG motifs is the induction of immune and inflammatory responses in a senseless manner. CpG motifs exist abundantly in microbes and evoke innate immunity that constitutes the first line of defense against microbial infections in vertebrates. CpG motifs that essentially work in an antigen-nonspecific fashion, however, turn into novel immunomodulators that can manipulate acquired immunity in an antigen-specific manner if oligodeoxynucleotides containing CpG motifs (CpG ODNs) are directly conjugated to the antigen. CpG ODNs with potent polyclonal Th1-inducing ability show promise for application in immunotherapy whereby neutralization of dominant allergy-prone Th2 cells is achieved by inducing allergen-specific Th1 cells. The underlying mechanisms include an unexpected enhancement of dendritic cell function as a linker between innate and acquired immunity. In the foreseeable future the mainstream therapeutic role of corticosteroids in anti-inflammatory therapy for allergic diseases could possibly be replaced by immunotherapy using CpG ODN-conjugated antigens

Prevention of Tracheal High-Dose Tolerance Induction by Granulocyte-Macrophage Colony Stimulating Factor- Dependent Restoration of Antigen-Presenting Cell Function

The intrusion of airborne allergens into airways elicits eosinophilic inflammation, as represented by bronchial asthma. It has been shown that excessive amounts of allergen in murine trachea lead to an unexpected evasion of deleterious eosinophilic inflammation by inducing T cell tolerance. In the present study, the mechanisms of tracheal high-dose tolerance are examined with regard to accessory cell functions and the effects of pro-inflammatory cytokines on tolerance. Antigen-induced tracheal eosinophilia was suppressed on instillation of high doses of antigen into the trachea, while concurrent instillation of granulocyte-macrophage colony stimulating factor (GM-CSF) with the antigen restored the diminished responses. The restoration of eosinophilic infiltration by GM-CSF occurred in parallel with an increase in interleukin (IL)-4 production by CD4+ T cells from the mediastinal lymph nodes. This was found to reflect the empowerment of antigen-presenting cells by GM-CSF, because the impaired ability of Ia+ cells from the tolerant mice to stimulate IL-4-producing T cells is restored by GM-CSF administration. The prevention of tolerance by up-regulating accessory cell functions is a feature unique to GM-CSF, because another pro-inflammatory cytokine, IL-iβ, failed to empower antigen-presenting cells. Thus, besides the induction of transforming growth factor-β-secreting CD4+ T cells, high-dose tolerance in the trachea includes an impairment of the accessory cell functions that support IL-4 production from T cells, which was reversed by GM-CSF. This report is the first demonstration that GM-CSF breaks the T cell tolerance of IL-4-producing T helper cells

CD44 on blood eosinophils as a novel marker of bronchial asthma management

Bronchial asthma (BA) is characterized by infiltration of the respiratory tracts by eosinophils. A wide variety of adhesion molecules is involved in the binding of eosinophils to the vascular endothelium and their subsequent transmigration from the circulation to the airways, while little is known about CD44 expression on eosinophils. In the present study we introduce a novel staining combination with which surface markers on eosinophils can be analyzed without purification prior to staining and examined whether the expression of CD44 on peripheral blood eosinophils could be monitored as a marker of the control of BA. Staining of eosinophils with anti-CD16 and anti-VLA-4 monoclonal antibodies enabled us to delineate eosinophils as VLA-4highCDl6- cells from other leukocyte populations in the whole blood. CD44 was constitutively expressed on resting eosinophils and expression increased following cytokine-mediated activation. In all BA patients examined, CD44high eosinophils were enriched in sputum relative to peripheral blood, indicating that eosinophils in sputum were more activated than those in the blood. By comparing the extent of CD44 expression on blood eosinophils from poorly controlled and well-controlled asthma patients, we unexpectedly found that the density of CD44 expression is lower on blood eosinophils from the poorly controlled group. Thus, the extent of CD44 expression on blood eosinophils defined as VLA- 4highCD16- cells is a novel marker indicative of the management of BA