Value through Innovation20 April 2014

Scientific Publications

Boehringer Ingelheim offers an overview of scientific publications on the following pages. This overview represents all publications of the last three years (YTD) where employees of Boehringer Ingelheim worldwide were involved.

25 publications regarding Inflammation
  • Author:
    Caesar M; Felk S; Zach S; Brønstad G; Aasly JO; Gasser T; Gillardon F
    Title:
    Changes in matrix metalloprotease activity and progranulin levels may contribute to the pathophysiological function of mutant leucine-rich repeat kinase 2
    Source:
    GLIA Article in Press (2014)
    Abstract:
    Increasing evidence suggests that Parkinson's disease (PD)-linked Leucine-rich repeat kinase 2 (LRRK2) has a role in peripheral and brain-resident immune cells. Furthermore, dysregulation of the anti-inflammatory, neurotrophic protein progranulin (PGRN) has been demonstrated in several chronic neurodegenerative diseases. Here we show that PGRN levels are significantly reduced in conditioned medium of LRRK2(R1441G) mutant mouse fibroblasts, leukocytes, and microglia, whereas levels of proinflammatory factors, like interleukin-1beta and keratinocyte-derived chemokine, were significantly increased. Decreased PGRN levels were also detected in supernatants of cultured human fibroblasts isolated from presymptomatic LRRK2(G2019S) mutation carriers, while mitochondrial function was unaffected. Furthermore, medium levels of matrix metalloprotease (MMP) 2 increased, whereas MMP 9 decreased in LRRK2(R1441G) mutant microglia. Increased proteolytic cleavage of the MMP substrates ICAM-5 and alpha-synuclein in synaptoneurosomes from LRRK2(R1441G) mutant mouse brain indicates increased net synaptic MMP activity. PGRN levels were decreased in the cerebrospinal fluid of presymptomatic LRRK2 mutant mice, whereas PGRN levels were increased in aged symptomatic mutant mice. Notably, PGRN levels were also increased in the cerebrospinal fluid of PD patients carrying LRRK2 mutations, but not in idiopathic PD patients and in healthy control donors. Our data suggest that proinflammatory activity of peripheral and brain-resident immune cells may particularly contribute to the early stages of Parkinson's disease caused by LRRK2 mutations. © 2014 Wiley Periodicals, Inc.
  • Author:
    Tauber J; Opatz T
    Title:
    Synthesis of Anti-inflammatory Macrolactones: (S)-Curvularin and Derivatives
    Source:
    Poster for FLOHET-15 in Gainesville/Florida and for Frühjahrssymposium des Jungchemikerforums der GDCh in Jena
  • Author:
    Profita M; Albano GD; Riccobono L; Di Sano C; Montalbano AM; Gagliardo R; Anzalone G; Bonanno A; Pieper MP; Gjomarkaj M
    Title:
    Increased levels of Th17 cells are associated with non-neuronal acetylcholine in COPD patients
    Source:
    Immunobiology Article in press (2014)
    Abstract:
    T-lymphocytes, including Th17-cells and T-cells expressing acetylcholine (ACh), are key components of systemic inflammation in chronic obstructive pulmonary disease (COPD). We investigated whether ACh promotes Th17 cells in COPD. ACh, IL-17A, IL-22, ROR.t, FOXP3 expression and AChIL-17A, AChIL-22, AChROR.t coexpression was evaluated in peripheral blood mononuclear cells (PBMC) from COPD patients (n = 16), healthy smokers (HS) (n = 12) and healthy control subjects (HC) (n = 13) (cultured for 48 h with PMA) by flow cytometry. Furthermore, we studied the effect of Tiotropium (Spiriva®) (100 nM) and Olodaterol (1 nM) alone or in combination, and of hemicholinium-3 (50 .M) on AChIL-17A, AChIL-22, AChROR.t, and FOXP3 expression in CD3+PBT-cells of PBMC from COPD patients (n = 6) cultured for 48 h with PMA. CD3+PBT-cells expressing ACh, IL-17A, IL-22 and ROR.t together with CD3+PBT-cells co-expressing AChIL-17A, AChIL-22 and AChROR.t were significantly increased in COPD patients compared to HC and HS subjects with higher levels in HS than in HC without a significant difference. CD3+FOXP3+PBT-cells were increased in HS than in HC and COPD. Tiotropium and Olodaterol reduced the percentage of CD3+PBT-cells co-expressing AChIL-17A, AChIL-22, and AChROR.t while increased the CD3+FOXP3+PBT-cells in PBMC from COPD patients, cultured in vitro for 48 h, with an additive effect when used in combination. Hemicholnium-3 reduced the percentage of ACh+IL-17A+, ACh+IL-22+, and ACh+ROR.t+ while it did not affect FOXP3+ expression in CD3+PBT-cells from cultured PBMC from COPD patients. We concluded that ACh might promote the increased levels of Th17-cells in systemic inflammation of COPD. Long-acting .2-agonists and anticholinergic drugs might contribute to control this event. © 2014.
  • Author:
    Vallon V; Gerasimova M; Rose MA; Masuda T; Satriano J; Mayoux E; Koepsell H; Thomson SC; Rieg T
    Title:
    SGLT2 inhibitor empagliflozin reduces renal growth and albuminuria in proportion to hyperglycemia and prevents glomerular hyperfiltration in diabetic Akita mice
    Source:
    Am J Physiol 306 (2), F194-F204 (2014)
    Abstract:
    Our previous work has shown that gene knockout of the sodium-glucose cotransporter SGLT2 modestly lowered blood glucose in streptozotocin-diabetic mice (BG; from 470 to 300 mg/dl) and prevented glomerular hyperfiltration but did not attenuate albuminuria or renal growth and inflammation. Here we determined effects of the SGLT2 inhibitor empagliflozin (300 mg/kg of diet for 15 wk; corresponding to 60-80 mg.kg-1.day-1) in type 1 diabetic Akita mice that, opposite to streptozotocin-diabetes, upregulate renal SGLT2 expression. Akita diabetes, empagliflozin, and Akita + empagliflozin similarly increased renal membrane SGLT2 expression (by 38-56%) and reduced the expression of SGLT1 (by 33-37%) vs. vehicle-treated wild-type controls (WT). The diabetes-induced changes in SGLT2/ SGLT1 protein expression are expected to enhance the BG-lowering potential of SGLT2 inhibition, and empagliflozin strongly lowered BG in Akita (means of 187-237 vs. 517-535 mg/dl in vehicle group; 100-140 mg/dl in WT). Empagliflozin modestly reduced GFR in WT (250 vs. 306 (xl/min) and completely prevented the diabetes-induced increase in glomerular filtration rate (GFR) (255 vs. 397 (xl/min). Empagliflozin attenuated increases in kidney weight and urinary albumin/creatinine ratio in Akita in proportion to hyperglycemia. Empagliflozin did not increase urinary glucose/creatinine ratios in Akita, indicating the reduction in filtered glucose balanced the inhibition of glucose reabsorption. Empagliflozin attenuated/prevented the increase in systolic blood pressure, glomerular size, and molecular markers of kidney growth, inflammation, and gluconeogenesis in Akita. We propose that SGLT2 inhibition can lower GFR independent of reducing BG (consistent with the tubular hypothesis of diabetic glomerular hyperfiltration), while attenuation of albuminuria, kidney growth, and inflammation in the early diabetic kidney may mostly be secondary to lower BG.
  • Author:
    Konstan MW; Döring G; Heltshe SL; Lands LC; Hilliard KA; Koker P; Bhattacharya S; Staab A; Hamilton A
    Title:
    A randomized double blind, placebo controlled phase 2 trial of BIIL 284 BS (an LTB4 receptor antagonist) for the treatment of lung disease in children and adults with cystic fibrosis
    Source:
    J Cystic Fibros Article in press (2014)
    Abstract:
    Background: Airway inflammation, mediated in part by LTB4, contributes to lung destruction in patients with cystic fibrosis (CF). LTB4-receptor inhibition may reduce airway inflammation. We report the results of a randomized, double-blind, placebo-controlled study of the efficacy and safety of the leukotriene B4 (LTB4)-receptor antagonist BIIL 284 BS in CF patients. Methods: CF patients aged . 6 years with mild to moderate lung disease were randomized to oral BIIL 284 BS or placebo once daily for 24 weeks. Co-primary endpoints were change in FEV1 and incidence of pulmonary exacerbation. Results: After 420 (155 children, 265 adults) of the planned 600 patients were randomized, the trial was terminated after a planned interim analysis revealed a significant increase in pulmonary related serious adverse events (SAEs) in adults receiving BIIL 284 BS. Final analysis revealed SAEs in 36.1% of adults receiving BIIL 284 BS vs. 21.2% receiving placebo (p = 0.007), and in 29.6% of children receiving BIIL 284 BS vs. 22.9% receiving placebo (p = 0.348). In adults, the incidence of protocol-defined pulmonary exacerbation was greater in those receiving BIIL 284 BS than in those receiving placebo (33.1% vs. 18.2% respectively; p = 0.005). In children, the incidence of protocol-defined pulmonary exacerbation was 19.8% in the BIIL 284 BS arm, and 25.7% in the placebo arm (p = 0.38). Conclusions: While the cause of increased SAEs and exacerbations due to BIIL 284 BS is unknown, the outcome of this trial provides a cautionary tale for the administration of potent anti-inflammatory compounds to individuals with chronic infections, as the potential to significantly suppress the inflammatory response may increase the risk of infection-related adverse events. © 2014 European Cystic Fibrosis Society.
  • Author:
    Kuzmich D; Bentzien J; Betageri R; DiSalvo D; Fadra-Khan T; Harcken C; Kukulka A; Nabozny G; Nelson R; Pack E; Souza D; Thomson D
    Title:
    Function-regulating pharmacophores in a sulfonamide class of glucocorticoid receptor agonists
    Source:
    Bioorg Med Chem Lett Article in press (2013)
    Abstract:
    A class of alpha-methyltryptamine sulfonamide glucocorticoid receptor (GR) modulators was optimized for agonist activity. The design of ligands was aided by molecular modeling, and key function-regulating pharmacophoric points were identified that are critical in achieving the desired agonist effect in cell based assays. Compound 27 was profiled in vitro and in vivo in models of inflammation. Analogs could be rapidly prepared in a parallel approach from aziridine building blocks. © 2013 Elsevier Ltd. All rights reserved.
  • Author:
    R. Lewis; L. Chachi; Y. Amrani; P. Bradding
    Title:
    A comparison of b2-adrenoceptor desensitisation induced by olodaterol and formoterol in human lung mast cells and airway smooth muscle cells
    Source:
    ERS, Barcelona, Spain, Sep 7-9, 2013
  • Author:
    Schülert N; Corradini L; Just S; Bernlöhr C; Külzer R; Doods H
    Title:
    The Bradykinin B1 receptor antagonist BI 113823 reverses hyperalgesia by reducing mechanosensitivity of peripheral afferents and nociceptive-specific dorsal horn neruons in a rat model of inflammatory pain
    Source:
    Deutscher Schmerzkongress
  • Author:
    Lacy BE; Wang F; Bhowal S; Schaefer E
    Title:
    On-demand hyoscine butylbromide for the treatment of self-reported functional cramping abdominal pain
    Source:
    Scand J Gastroenterol 48 (8), 926-935 (2013)
    Abstract:
    Objective. Hyoscine butylbromide (HBB) has been used for 60 years to treat cramping abdominal pain, but scientific evidence to support on-demand use is limited. The aim of this study was to identify a meaningful efficacy end point that differentiates between the effects of HBB (20-100 mg/day) and placebo when used on demand. Materials & methods. 175 patients were treated in a randomized, double-blind, placebo-controlled, two-arm parallel group study. After a 4-week run-in period, patients used HBB to treat 2 distinct episodes of abdominal pain associated with cramping (APC). Patients entered data into an electronic diary for up to 4 h/episode. Pain intensity was assessed using an 11-point numerical pain rating scale (NPRS). Patients were allowed to self-medicate with up to 4 additional doses of HBB every 30 min. Results. The adjusted mean difference for the change from baseline during the 4-h observation period for a reduction in abdominal pain was -0.7 (95% confidence interval (CI) -1.3, -0.1, p = 0.016) for episode 1 and -0.6 (95% CI -1.2, 0.0, p = 0.051) for episode 2. Patients in the HBB group recorded a clinically relevant reduction of at least 2 points on the NPRS (approximately 30% pain relief) earlier than patients in the placebo group (HBB: 45 min, placebo: 60 min). Reported adverse events were infrequent in both groups (10.2% and 10.3%). Conclusions. HBB is effective in the treatment of recurrent APC and is safe and well tolerated when used on demand. The change from baseline in the intensity of APC using the 11-point NPRS distinguished HBB from placebo. © 2013 Informa Healthcare.
  • Author:
    McHugh Kevin P; Crotti Tania N; Li Jun; Hill Jon; Nabozny Gerald H; Goldring Steven R
    Title:
    Novel targets for blocking osteoclast-mediated resorption in inflammatory disorders.
    Abstract:
    -
  • Author:
    Thome Diane; Souza Donald; Behera Aruna; Zheng Jie; Swantek Jennifer; Nabozny Gerald H
    Title:
    Development of a quantitative model of endotoxin-induced calvarial bone erosion utilizing micro-computed tomography and PAX-it imaging analysis software.
    Source:
    Arthritis Rheum 64 (10), 512 (2012)
    Abstract:
    Background/Purpose: To facilitate an understanding of the biological factors involved in inflammation- mediated bone loss, in vitro systems and murine models of bone loss, such as endotoxin-induced calvarial erosion have been utilized. However, challenges exist with in vivo model robustness, throughput and quantitation of bone erosions. Assessment of erosion is typically determined via micro-computed tomography (MicroCT) but quantitation has been hampered by mouse to mouse variability in bone morphology and lack of consistent robust erosion obtained by endotoxin alone. We set out to develop a robust, quantitative in vivo endotoxin-induced calvarial erosion model that would be amenable for in vivo testing of potential therapeutic agents and would allow for gaining insight into processes involved in inflammatory bone loss. Methods: Lipopolysaccharide (LPS) was injected with or without receptor activator of NF-kB ligand (RANKL), over the calvaria of mice. On day 5 or 8, mice were euthanized and processed for MicroCT analysis. Skulls were scanned at medium resolution, 70 kVp, 114 .mu.A. Scans were rendered into 3-D. Optimal viewing angles were chosen for creation of TIFF images, which were then examined using Pax-IT imaging analysis software (MIS, Inc), using a 4-pass analysis to generate 4 areas of erosion representing each of the 4 calvarial plates. The values obtained in the 4 areas of erosion were summed to provide a measure of total area of calvarial erosion. For further validation aimed at demonstration of the quantitative nature of the model, in vivo testing of therapeutic agents known to play a role in osteoclastogenesis was implemented. Results: Five-day dose-response studies using RANKL + LPS in B10.RIII mice resulted in severe erosive events, but dose-dependent relationships were not consistent. We compared 5-day and 8-day models. At Day 8, mice exhibited consistent and strong dose-dependent calvarial erosion with co-administration of 25 .mu.g LPS and 10ug RANKL per mouse. We determined inclusion of RANKL was necessary for optimal erosion. Summating four 8-day studies using B10.RIII mice, co-administration of 25 .mu.g LPS and 10 .mu.g RANKL led to an average area of erosion of 4080 + 259 mm2, compared to the naive (untreated) average area of erosion of 506 + 40 mm2. The microCT/Pax-IT analysis method facilitated reproducible, quantitative results. Importantly, therapeutic agents known to play a role in osteoclastogenesis demonstrated statistically significant reductions in bone erosion further emphasizing the quantitative nature of this model. Conclusion: We developed a robust, quantitative endotoxin plus RANKL-induced calvarial bone erosion model through use of MicroCT and Pax-IT analysis. This model and quantitation methodology allows for testing therapeutic agents to facilitate understanding of the biological factors and processes involved in bone erosion. The ultimate goal is to impact identification of better therapeutic options for patients.
  • Author:
    McHugh Kevin P; Crotti Tania N; LiJun; Hill Jon; Nabozny Gerald H; Goldring Steven R; Purdue P Edward
    Title:
    Novel targets for blocking osteoclast-mediated resorption in inflammatory disorders.
    Source:
    Arthritis Rheum 64 (10), 12-13 (2012)
    Abstract:
    Background/Purpose: Osteoclasts are specialized myeloid lineage cells that mediate both pathologic and physiologic bone remodeling Fully differentiated and functional osteoclasts are found exclusively associated with the bone surface, indicating that interaction with the bone substrate plays a pivotal role in the regulation of osteoclast biology. Most in vitro studies on the formation and activation of osteoclasts have been performed using cells adhered to tissue culture plastic, and there is little information regarding the specific molecular mechanisms by which adherence to the bone surface regulates the terminal stages of osteoclast differentiation. To address this discrepancy, we have compared the transcriptional profiles of osteoclasts generated on a variety of substrates, including authentic bone and have identified the unique molecular signatures that are dependent and independent of integrin beta 3, a prototypical osteoclast regulator of osteoclast function. Methods: Bone marrow derived macrophages from wild-type and integrin beta 3 deficient mice were cultured in the presence or absence of RANKL on plastic, hydroxyapatite, or calvarial bone discs. RNA was isolated at different stages of osteoclast generation and microarray analysis and pathway mapping were utilized to identify osteoclast signaling pathways regulated by the cytokine RANKL, bone substrate, and integrin beta 3. Pathway analysis was performed using GSEA and Ingenuity pathway analysis. Results: Microarray analysis revealed RANKL-induced molecular profiles that were uniquely and specifically regulated in osteoclasts differentiated on the authentic bone substrate compared to the other substrates. Pathway analysis revealed coordinated downregulation by bone of a cluster of genes associated with cell cycle progression. The expression level of integrin beta 3, which is induced by RANKL during osteoclast differentiation, is further modulated by culture on the bone substrate. Interestingly, bone regulated genes could be divided into those that were dependent and independent of integrin beta 3. Conclusion: In addition to cytokines and growth factors, interaction of osteoclasts and their precursor with the bone substrate regulates pathways that are involved in osteoclast formation and activation. Although integrin beta 3 has been regarded as the essential mediator of osteoclast attachment and activation, molecular pathways independent of this integrin also modulate the genetic program during osteoclastogenesis. Analysis of these genes and their pathways provides novel targets and approaches for therapeutic targeting of osteoclast-mediated bone loss in inflammatory and related bone disorders.
  • Author:
    Berg Karen L; Hanidu Adedayo; Hill Jon; JiangXiaoyu; Freeman Tom; Swantek Jennifer; Nabozny Gerald H
    Title:
    Analysis of gene expression patterns in rheumatoid arthritis (RA) synovial macrophages from patients undergoing disease flare.
    Source:
    Arthritis Rheum 64 (10), 512 (2012)
    Abstract:
    Background/Purpose: Synovial macrophages play a key role in RA pathogenesis. Their numbers are greatly increased in RA synovium, their phenotype is consistent with a pro-inflammatory function, and clinical data indicate all efficacious RA therapies lead to a significant reduction of this cell type in the diseased joint. Studies probing global gene expression in RA synovial macrophages have been limited in scope. We set out to further our understanding of synovial macrophage biology.both in RA and additional arthropathies.through gene expression profiling, gene mining and pathway analysis in synovial macrophages from patients undergoing disease flare. Methods: Synovial fluids were collected from 10 RA patients in disease flare (treatments included oral DMARDs and/or biologics), and CD14+ cells isolated by positive selection. Control macrophage samples were generated from CD14+ peripheral blood monocytes from healthy donors, and differentiated with M-CSF. Macrophages were also isolated from synovial fluid from patients with Psoriatic Arthritis and other arthropathies to allow for comparison of dysregulated genes and pathways across related diseases. RNAs were isolated, processed, and transcriptional profiling of the samples was performed using Affymetrix arrays. Statistical analysis was performed using principle component analysis (PCA) and upregulated genes defined as >2.0-fold increase in expression and an FDR p-value of <0.05. Internal genesets from cytokine-stimulated macrophages, and M1 or M2 polarized macrophages, were used as comparators to further characterize upregulated genes and pathways in RA macrophages. Results: Approximately 1900 genes were significantly upregulated in RA synovial macrophages, and 70% demonstrated overlap with Psoriatic arthritis and other arthropathies. Forty percent of RA-upregulated genes were upregulated in M1 polarized macrophages, vs. 7% in M2 macrophages. Only 10% of RA-regulated genes were upregulated by chronic treatment of control macrophages with TNFa, indicating other factors impact gene expression to a greater extent in RA macrophages. Gene Set Enrichment and Ingenuity Pathway Analysis indicated that several immunoregulatory pathways previously linked to RA pathogenesis were upregulated in RA macrophages (including CD40, and FcR), independent of treatment. Interestingly, the LTb signaling pathway, which was highly upregulated in M1 macrophages was not significantly induced in RA macrophages indicating a unique inflammatory phenotype of these disease macrophages. Conclusion: We performed a comprehensive study focused on identifying differentially regulated genes and pathways in synovial macrophages from patients with RA and related arthropathies. These results suggest a unique inflammatory phenotype of RA macrophages and maintenance of key inflammatory pathways during flare that is independent of treatment. These observations may shed light on novel intervention points for treatment of RA.
  • Author:
    Purdue1 P Edward; HillJon; Goldring Steven R; BinderNikolaus B; SwantekJennifer L; Shen Zhenxin; CrottiTania N; McHughKevin P
    Title:
    The sphingosine-1-phosphate pathway is a key regulator of bone substrate-mediated osteoclast differentiation in inflammatory arthritis
    Source:
    Arthritis Rheum 64 (10), 1063 (2012)
    Abstract:
    Multiple lines of evidence have established that osteoclasts are required for physiologic bone resorption and pathological bone loss in inflammatory disorders. Isolation of differentiated osteoclasts from bone is technically challenging, but with the discovery that M-CSF and RANKL are sufficient for osteoclast differentiation came the ability to generate multinucleated osteoclasts from myeloid precursor cells in vitro with high efficiency. In vivo, analysis of osteoclasts at sites of bone resorption reveals that functionally activated osteoclasts are almost exclusively localized to the immediate bone surface, indicating that cell-substrate interactions contribute to terminal osteoclast differentiation. In this study, we have employed a unique in vitro osteoclastogenesis system using authentic bone substrates and expression profiling and pathway analysis to identify critical bone substrate-mediated pathways of osteoclast formation and activation. We have further validated the sphingosine-1-phosphate (S1P) generation, transport and signaling pathway as a key component of the bone-substrate regulated osteoclast differentiation program in vitro and in vivo. Methods: Murine bone marrow derived macrophages were cultured in the presence or absence of RANKL on plastic, hydroxyapatite, or calvarial bone discs. RNA was isolated at different stages of osteoclast generation and subjected to microarray analysis. Pathway analysis was performed using GSEA and Ingenuity pathway analysis. SphK inhibitors were used to validate involvement of S1P signaling in osteoclastogenesis in vitro. Immunohistochemical analysis of the RA bone-pannus interface was used assess activation of this pathway in osteoclasts in vivo. Results: Microarray analysis revealed unique clusters of RANKL induced and bone substrate-modulated osteoclast genes. Of 1490 genes upregulated by RANKL in differentiated osteoclasts, 5.4% were further upregulated by culture on the bone substrate; an additional 8.5% were downregulated. Pathway analysis identified the S1P pathway as significantly regulated by both RANKL and bone substrate, and that this regulation was dependent upon the presence of integrin beta 3, a key mediator of osteoclast attachment to bone. SphK inhibitors dose-dependently blocked RANKL-induced human and mouse osteoclastogenesis in vitro. Furthermore, SphK1 was highly induced in osteoclasts at sites of RA bone erosions. Conclusion: Interaction of osteoclasts with the bone surface regulates multiple critical pathways of osteoclast formation and activation. Pathway analysis and in vitro/in vivo validation identifies the S1P pathway as a critical regulator of RANKL-bone-dependent osteoclastogenesis. This pathway represents a novel therapeutic target for preventing osteoclast-mediated bone destruction in inflammatory bone loss disorders.
  • Author:
    Schleputz M; Bernau M; Kanzler SS; Uhlig S; Martin C
    Title:
    Neurally-induced bronchoconstriction in precision-cut lung slices of guinea pigs is modulated by lipopolysaccharide and neurotrophins.
    Source:
    Naunyn Schmiedebergs Arch Pharmakol 386 (1), 71 (2013)
    Abstract:
    Rationale: Inflammation, a hallmark of asthma and COPD, contributes to neural modulation and consequential airway hyperresponsiveness (AHR). To further study this concept the present investigation aimed to characterize the effect of lipopolysaccharide (LPS) as well as neurotrophins and their receptors on neurally-induced bronchoconstriction (BC) in precision-cut lung slices (PCLS) of guinea pigs. Methods: PCLS were prepared from Dunkin Hartley guinea pigs. PCLS were incubated with LPS (500 ng/mL) for 18 h to mimic inflammation and thereafter TNF release was measured by ELISA. Neurotrophic receptors (TrkA, -B, -C, p75NTR, GFR.alpha.) were analyzed by Western-blotting. Electric field stimulation (EFS) of PCLS was performed to induce neural BC. BC was monitored by videomicroscopy and the initial airway area was defined as 100% initial airway area (IAA). Frequency-response curves (1-100 Hz) were conducted on untreated PCLS (control) or on pretreated PCLS, incubated with LPS (500 ng/mL for 18 h), nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), neurotrophin 3 (NT3), or glial derived neurotrophic factor (GDNF) (each at 50 ng/mL for 20 min or 24 h). Results: LPS application in guinea pig PCLS led to a 128-fold induction of TNF release and augmented BC in frequency-response curves (44%-IAA vs. 63%-IAA in control for frequencies .gtoreq. 50 Hz). The expression of TrkA, -B, -C, p75NTR and GFR.alpha. was detected in PCLS. Among the neurotrophins studied NGF and GDNF (44%-IAA and 41%-IAA, respectively, vs. 63%-IAA in control for frequencies .gtoreq. 50 Hz) increased the neurallyinduced BC after short incubation periods. In contrast, after incubation for 24 h only NT3 (46%-IAA for frequencies .gtoreq. 50 Hz) enhanced BC in comparison to the control (61%-IAA for frequencies .gtoreq. 50 Hz). Conclusion: Since TNF levels were elevated after LPS, a pro-inflammatory state could be confirmed in PCLS, leading to AHR after EFS. Distinct neurotrophic factors are capable to induce AHR. The fast effects of NGF and GDNF may be based on direct modulation of neurotransmission, i.e. action potential profiles, whereas NT3 may involve altered transcription based changes of nerve function.
  • Author:
    Miriyala S; Panchatcharam M; Landar A; Ramanujam M; Chatterjee S; Muthuswamy A
    Title:
    The janus of oxidative stress signaling in different pathophysiological conditions
    Source:
    Oxidative Med Cell Longevity art no 708796 (2013)
    Abstract:
    no abstract available
  • Author:
    Hirsch S; Corradini L; Just S; Arndt K; Doods H
    Title:
    The CGRP receptor antagonist BIBN4096BS peripherally alleviates inflammatory pain in rats
    Source:
    Pain Article in Press (2013)
    Abstract:
    Calcitonin gene-related peptide (CGRP) is known to play a major role in the pathogenesis of pain syndromes, in particular migraine pain. Here we focus on its implication in a rat pain model of inflammation, induced by injection of complete Freund adjuvant (CFA). The nonpeptide CGRP receptor antagonist BIBN4096BS reduces migraine pain and trigeminal neuronal activity. Here we demonstrate that the compound reduces inflammatory pain and spinal neuronal activity. Behavioural experiments reveal a reversal of the CFA-induced mechanical hypersensitivity and monoiodoacetate (MIA)-induced weight-bearing deficit in rats after systemic drug administration. To further investigate the mechanism of action of the CGRP antagonist in inflammatory pain, in vivo electrophysiological studies were performed in CFA-injected rats. Recordings from wide dynamic range neurons in deep dorsal horn layers of the lumbar spinal cord confirmed a reduction of neuronal activity after systemic drug application. The same amount of reduction occurred after topical administration onto the paw, with resulting systemic plasma concentrations in the low nanomolar range. However, spinal administration of BIBN4096BS did not modify the neuronal activity in the CFA model. Peripheral blockade of CGRP receptors by BIBN4096BS significantly alleviates inflammatory pain. © 2013 International Association for the Study of Pain.
  • Author:
    Wang J; Shiratori I; Uehori J; Ikawa M; Arase H
    Title:
    Neutrophil infitration during inflammation is regulated by PLRalpha via modulation of integrin activation.
  • Author:
    Moss N; Xiong Z; Burke M; Cogan D; Gao DA; Haverty K; Heim-Riether A; Hickey ER; Nagaraja R; Netherton M; O'Shea K; Ramsden P; Schwartz R; Shih D-T; Ward Y; Young E; Zhang Q
    Title:
    Exploration of cathepsin S inhibitors characterized by a triazole P1-P2 amide replacement.
    Source:
    Bioorg Med Chem Lett 22 (23), 7189-7193 (2012)
  • Author:
    Blech M; Peter D; Fischer P; Bauer MMT; Hafner M; Zeeb M; Nar H
    Title:
    One target - two different binding modes: Structural insights into gevokizumab and canakinumab interactions to interleukin-1beta.
    Source:
    J Mol Biol 425 (1), 94-111 (2013)
    Abstract:
    Interleukin-1beta (IL-1beta) is a key orchestrator in inflammatory and several immune responses. IL-1beta exerts its effects through interleukin-1 receptor type I (IL-1RI) and interleukin-1 receptor accessory protein (IL-1 RAcP), which together form a heterotrimeric signaling-competent complex. Canakinumab and gevokizumab are highly specific IL-1beta monoclonal antibodies. Canakinumab is known to neutralize IL-1beta by competing for binding to IL-1R and therefore blocking signaling by the antigen:antibody complex. Gevokizumab is claimed to be a regulatory therapeutic antibody that modulates IL-1beta bioactivity by reducing the affinity for its IL-1RI:IL-1RAcP signaling complex. How IL-1beta signaling is affected by both canakinumab and gevokizumab was not yet experimentally determined. We have analyzed the crystal structures of canakinumab and gevokizumab antibody binding fragment (Fab) as well as of their binary complexes with IL-1beta. Furthermore, we characterized the epitopes on IL-1beta employed by the antibodies by NMR epitope mapping studies. The direct comparison of NMR and X-ray data shows that the epitope defined by the crystal structure encompasses predominantly those residues whose NMR resonances are severely perturbed upon complex formation. The antigen:Fab co-structures confirm the previously identified key contact residues on IL-1beta and provide insight into the mechanisms leading to their distinct modulation of IL-1beta signaling. A significant steric overlap of the binding interfaces of IL-1R and canakinumab on IL-1beta causes competitive inhibition of the association of IL-1beta and its receptor. In contrast, gevokizumab occupies an allosteric site on IL-1beta and complex formation results in a minor reduction of binding affinity to IL-1RI. This suggests two different mechanisms of IL-1beta pathway attenuation.
  • Author:
    Jenh C-H; Cox MA; Cui L; Reich E-P; Sullivan L; Chen S-C; Kinsley D; Qian S; Kim SH; Rosenblum S; Kozlowski J; Fine JS; Zavodny PJ; Lundell D
    Title:
    A selective and potent CXCR3 antagonist SCH 546738 attenuates the development of autoimmune diseases and delays graft rejection.
    Source:
    BMC Immunol 13 art.no.2 (2012)
    Abstract:
    Background: The CXCR3 receptor and its three interferon-inducible ligands (CXCL9, CXCL10 and CXCL11) have been implicated as playing a central role in directing a Th1 inflammatory response. Recent studies strongly support that the CXCR3 receptor is a very attractive therapeutic target for treating autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis and psoriasis, and to prevent transplant rejection. We describe here the in vitro and in vivo pharmacological characterizations of a novel and potent small molecule CXCR3 antagonist, SCH 546738.Results: In this study, we evaluated in vitro pharmacological properties of SCH 546738 by radioligand receptor binding and human activated T cell chemotaxis assays. In vivo efficacy of SCH 546738 was determined by mouse collagen-induced arthritis, rat and mouse experimental autoimmune encephalomyelitis, and rat cardiac transplantation models. We show that SCH 546738 binds to human CXCR3 with a high affinity of 0.4 nM. In addition, SCH 546738 displaces radiolabeled CXCL10 and CXCL11 from human CXCR3 with IC 50ranging from 0.8 to 2.2 nM in a non-competitive manner. SCH 546738 potently and specifically inhibits CXCR3-mediated chemotaxis in human activated T cells with IC 90about 10 nM. SCH 546738 attenuates the disease development in mouse collagen-induced arthritis model. SCH 546738 also significantly reduces disease severity in rat and mouse experimental autoimmune encephalomyelitis models. Furthermore, SCH 546738 alone achieves dose-dependent prolongation of rat cardiac allograft survival. Most significantly, SCH 546738 in combination with CsA supports permanent engraftment.Conclusions: SCH 546738 is a novel, potent and non-competitive small molecule CXCR3 antagonist. It is efficacious in multiple preclinical disease models. These results demonstrate that therapy with CXCR3 antagonists may serve as a new strategy for treatment of autoimmune diseases, including rheumatoid arthritis and multiple sclerosis, and to prevent transplant rejection.
  • Author:
    Chen L; Klein T; Leung P S
    Title:
    Effects of combining linagliptin treatment with BI-38335, a novel SGLT2 inhibitor, on pancreatic islet function and inflammation in db/db mice
    Source:
    Curr Mol Med 12 (8), 995-1004 (2012)
    Abstract:
    Dipeptidyl peptidase-4 (DPP-4) inhibitors enhance incretin actions and beta-cell function. Concurrently, sodium-glucose co-transporter 2 (SGLT2) inhibitors block renal glucose reabsorption promoting excretion. In this study, we investigated the effects of linagliptin (a DPP-4 inhibitor) and BI-38335 (an SGLT2 inhibitor), individually and in combination, on glucose homeostasis, islet function, and pancreatic islet morphology in db/db mice. Diabetic and non-diabetic mice received linagliptin (3 mg/kg), BI-38335 (1 mg/kg), the two drugs in combination or control once daily for 8 weeks. Blood glucose homeostasis and insulin sensitivity were assessed. Pancreatic islet function and morphology as well as inflammatory factors and tolllike receptor 2 (TLR2) pathways involved in islet inflammation were investigated. Active treatments markedly reduced blood glucose and glycated hemoglobin A1c (HbA 1c) levels, with the combined treatment showing the greater effects. Insulin resistance was improved in the BI-38335 and combination groups with the enhancement of insulin sensitivity and significant increase of serum adiponectin levels. The combined treatment exhibited greater effects on enhanced islet glucose-stimulated insulin secretion and improved glucose tolerance. Moreover, the combination restored the islet beta-/alpha-cell ratio, reduced beta-cell apoptosis, decreased expression of islet immune cell markers, and suppressed factors related to the TLR2 pathway. In addition, all active treatments reduced serum lipid profiles, though the combination produced more robust effects. Collectively, our data show that combined treatment with BI-38335 and linagliptin work, at least in part, synergistically to benefit islet cell function/architecture and insulin resistance, thus improving glycemic control. © 2012 Bentham Science Publishers.
  • Author:
    Wagner C L; Visvanathan S; Elashoff M; Mcinnes I B; Mease P J; Krueger G G; Murphy F T; Papp K; Gomez-Reino JJ; Mack M; Beutler A; Gladman D; Kavanaugh A
    Title:
    Markers of inflammation and bone remodelling associated with improvement in clinical response measures in psoriatic arthritis patients treated with golimumab
    Source:
    Ann Rheum Dis 72 (1), 83-88 (2013)
    Abstract:
    Objective: To determine serum biomarker associations with clinical response to golimumab treatment in patients with psoriatic arthritis (PsA). Methods: GO-REVEAL was a randomised, placebo-controlled study of golimumab in patients with active PsA. Samples were collected from 100 patients at baseline, week 4 and week 14, and analysed for serum-based biomarkers and protein profiling (total 92 markers); data were correlated with clinical measures at week 14. Results: Serum levels of a subset of proteins (apolipoprotein C III, ENRAGE, IL-16, myeloperoxidase, vascular endothelial growth factor, pyridinoline, matrix metalloproteinase 3, C-reactive protein (CRP), carcinoembryonic antigen, intercellular adhesion molecule 1 and macrophage inflammatory protein 1?) at baseline or week 4 were strongly associated with American College of Rheumatology 20% improvement (ACR20) response and/or disease activity score in 28 joints (DAS28) at week 14. A smaller subset of proteins was significantly associated with a 75% improvement in the psoriasis area and severity index score (PASI75) at week 14, (adiponectin, apolipoprotein CIII, serum glutamic oxaloacetic transaminase, and tumour necrosis factor ?). Subsets of proteins were identified as potentially predictive of clinical response for each of the clinical measures, and the power of these biomarker panels to predict clinical response to golimumab treatment was stronger than for CRP alone. Conclusions: This analysis provides insight into several panels of markers that may have utility in identifying PsA patients likely to have ACR20, DAS28, or PASI75 responses following golimumab treatment. Copyright Article author (or their employer) 2012.
  • Author:
    Teunissen M B M; Zheng L; De Groot M; De Rie M A; Fine J S; Chen S C
    Title:
    Rise in dermal CD11c + dendritic cells associates with early-stage development of psoriatic lesions
    Source:
    Arch Dermatol Res 304 (6), 443-449 (2012)
    Abstract:
    There is limited information available regarding the phenotype and function of leukocytes involved in the earliest stages of psoriatic lesion development. In this study, we examined the presence of different types of leukocytes in psoriatic point lesions collected at three 1-week interval time points from a recent and simultaneously formed group of point lesions. The cells were quantified and compared with K16 expression and epidermal thickness, both typically increased in this disease and considered as hallmarks. We found a significant correlation between K16 + cell increment and the increase in epidermal thickness in the timeframe of 14 days. The change in CD3 +, CD4 +, and CD8 + T-cell numbers in the dermis showed a significant association with these two features from d7 to d14, whereas in the epidermis only CD8 + T cells demonstrated a significant correlation. Remarkably, the relationship between T cells and disease progression was preceded by a significant correlation of CD11c + dendritic cells (DCs) with K16 expression and epidermal thickness from baseline onwards. Interestingly, there was also a numeric correlation of CD11c + DCs with the CD3 + T-cell shifts from d7 to d14. A significant correlation was also found between dermal CD14 + cells and K16 expression from d7 to d14. BDCA-2 + plasmacytoid DCs were absent in nonlesional skin, but found at low numbers in most lesions. The change in plasmacytoid DC or neutrophil numbers did not correlate with lesion development. In conclusion, our study suggests a relevant role for T cells, and in particular dermal CD11c + DCs, in the earliest stage of psoriatic lesion development. © The Author(s) 2012.
  • Author:
    Teunissen MBM; Zheng L; de Groot M; de Rie MA; Fine JS; Chen S-C
    Title:
    Rise in dermal CD11c + dendritic cells associated with early-stage development of psoriatic lesions.
    Source:
    Arch Dermatol Res, 1-7 (2012)