Azelastine Desensitization of Transient Receptor Potential Vanilloid 1: A Potential Mechanism Explaining its Therapeutic Effect in Nonallergic Rhinitis
Restricted accessResearch articleFirst published online May, 2014
Azelastine Desensitization of Transient Receptor Potential Vanilloid 1: A Potential Mechanism Explaining its Therapeutic Effect in Nonallergic Rhinitis
Capsaicin, a prototypic transient receptor potential vanilloid 1 (TRPV1) agonist, has been shown to be more clinically effective in the treatment of nonallergic rhinitis (NAR) compared with other rhinitis subtypes. Azelastine has also been found to be clinically effective in the treatment of NAR but its mechanism(s) of action is still poorly elucidated. This study was designed to determine, using in vitro cell lines, whether topical therapies including azelastine have activity on TRPV1 ion channels similar to capsaicin.
Methods
The effects of capsaicin (1 μM), azelastine (30 μM), bepotastine (10 μM), olopatadine (10 μM), and fluticasone (200 μM) on TRPV1 channels using mice neuronal cells (Cath.a), as surrogates for submucosal sensory neurons, and human nasal epithelial cells (hNEC) were determined and compared. For azelastine, bepotastine, and capsaicin, which elicited an agonist effect on TRPV1, live cell [Ca2+] signaling in Cath.a cells and hNECs expressing TRPV1 were performed in the absence and presence of capsazepine at 10 μM (a TRPV1 antagonist) using wild-type mouse embryonic fibroblasts (wtMEFs) that express TRPV1 ion channels and TRPV1 homozygous null mutant (TRPV1 -/-) knockout MEF cells as controls to establish TRPV1 channel selectivity. As azelastine has previously been found clinically effective in NAR, additional experiments were performed to determine its ability to desensitize TRPV1 ion channels and its effect on regulating intracellular calcium homeostasis.
Results
Cath.a cells treated with azelastine, bepotastine, or capsaicin showed a significant increase in TRPV1-dependant (Ca2+) specific cytosolic fluorescence. Continuous treatment with azelastine or capsaicin resulted in desensitization of TRPV1 channels. In hNECs, azelastine stimulation resulted in Ca2+ shifts from the cytosol to mitochondria and overexpression of hematopoietic cell-specific Lyn substrate 1–associated protein X1, which may thus be effective in cytosolic Ca2+ homeostasis.
Conclusion
Azelastine, similar to capsaicin, exhibits direct activity on TRPV1 ion channels that may represent a novel mechanistic pathway explaining its clinical efficacy in NAR.
WebbD.R., MeltzerE.O., FinnA.F.Jr.Intranasal fluticasone propionate is effective for perennial nonallergic rhinitis with or without eosinophilia.Annals Allergy Asthma Immunol88: 385–390, 2002.
4.
BanovC.H., LiebermanP., and Vasomotor Rhinitis StudyG.Efficacy of azelastine nasal spray in the treatment of vasomotor (perennial nonallergic) rhinitis.Ann Allergy Asthma Immunol86: 28–35, 2001.
5.
BernsteinJ.A.Azelastine hydrochloride: A review of pharmacology, pharmacokinetics, clinical efficacy and tolerability.Curr Med Res Opin23: 2441–2452, 2007.
6.
CiprandiG.Treatment of nonallergic perennial rhinitis.Allergy59(suppl 76): 16–22, 2004.
7.
BernsteinJ.A., DavisB.P., PicardJ.K.A randomized, double-blind, parallel trial comparing capsaicin nasal spray with placebo in subjects with a significant component of nonallergic rhinitis.Ann Allergy Asthma Immunol107: 171–178, 2011.
8.
BlomH.M., Van RijswijkJ.B., GarreldsI.M.Intranasal capsaicin is efficacious in non-allergic, non-infectious perennial rhinitis. A placebo-controlled study.Clin Exp Allergy27: 796–801, 1997.
9.
RadresaO., ParéM., and AlbertJ.S.Multiple roles of transient receptor potential (TRP) channels in inflammatory conditions and current status of drug development.Curr Top Med Chem13: 367–385, 2013.
10.
StjärneP.Sensory and motor reflex control of nasal mucosal blood flow and secretion; clinical implications in non-allergic nasal hyperreactivity.Acta Physiol Scand Suppl600: 1–64, 1991.
11.
WangH., WangD.H., and GalliganJ.J.P2Y2 receptors mediate ATP-induced resensitization of TRPV1 expressed by kidney projecting sensory neurons.Am J Physiol Regul Integr Comp Physiol298: R1634–R1641, 2010.
12.
Van GervenL., AlpizarY.A., WoutersM.M.Capsaicin treatment reduces nasal hyperreactivity and transient receptor potential cation channel subfamily V, receptor 1 (TRPV1) overexpression in patients with idiopathic rhinitis.J Allergy Clin Immunolpii: S0091–S6749 (13)01340–7, 2013.
13.
TurnerH., FleigA., StokesA.Discrimination of intracellular calcium store subcompartments using TRPV1 (transient receptor potential channel, vanilloid subfamily member 1) release channel activity.Biochem J371: 341–350, 2003.
14.
NIH/3T3 (ATCC® CRL-1658™) ATCC, Manassas, VA.
15.
ZhangL.L., Yan LiuD., MaL.Q.Activation of transient receptor potential vanilloid type-1 channel prevents adipogenesis and obesity.Circ Res100: 1063–1070, 2007.
DewittS., LaffafianI., MorrisM.R., and HallettM.B.Cytosolic Ca2+ measurement and imaging in inflammatory cells.Methods Mol Biol225: 47–59, 2003.
18.
YangY., YangH., WangZ.Functional TRPV1 expression in human corneal fibroblasts.Exp Eye Res107: 121–129, 2013.
19.
SungB., PrasadS., RavindranJ.Capsazepine, a TRPV1 antagonist, sensitizes colorectal cancer cells to apoptosis by TRAIL through ROS-JNK-CHOP-mediated upregulation of death receptors.Free Rad Biol Med53: 1977–1987, 2012.
20.
ReillyC.A., JohansenM.E., LanzaD.L.Calcium-dependent and independent mechanisms of capsaicin receptor (TRPV1)-mediated cytokine production and cell death in human bronchial epithelial cells.J Biochem Mol Toxicol19: 266–275, 2005.
21.
BhaveG., HuH.J., GlaunerK.S.Protein kinase C phosphorylation sensitizes but does not activate the capsaicin receptor transient receptor potential vanilloid 1 (TRPV1).Proc Natl Acad Sci USA100: 12480–12485, 2003.
22.
PrescottE.D., and JuliusD.A modular PIP2 binding site as a determinant of capsaicin receptor sensitivity.Science300: 1284–1288, 2003.
23.
RatheeP.K., DistlerC., ObrejaO.PKA/AKAP/VR-1 module: A common link of Gs-mediated signaling to thermal hyperalgesia.J Neurosci22: 4740–4745, 2002.
24.
LivakK.J., and SchmittgenT.D.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.Methods25: 402–408, 2001.
25.
LiuB., ZhangC., and QinF.Functional recovery from desensitization of vanilloid receptor TRPV1 requires resynthesis of phosphatidylinositol 4,5-bisphosphate.J Neurosci25: 4835–4843, 2005.
26.
Sanz-SalvadorL., Andrés-BorderiaA., Ferrer-MontielA., and Plannells-CasesR.Agonist- and Ca2+-dependent desensitization of TRPV1 channel targets the receptor to lysosomes for degradation.J Biol Chem287: 19462–19471, 2012.
27.
GrzybowskaE.A., ZayatV., KonopínskiR.HAX-1 is a nucleocytoplasmic shuttling protein with a possible role in mRNA processing.FEBS J280: 256–272, 2013.
28.
VafiadakiE., PapaloukaV., ArvanitisD.A.The role of SERCA2a/PLN complex, Ca(2+) homeostasis, and anti-apoptotic proteins in determining cell fate.Pflugers Arch457: 687–700, 2009.
29.
VafiadakiE., SanoudouD., ArvanitisD.A.Phospholamban interacts with HAX-1, a mitochondrial protein with anti-apoptotic function.J Mol Biol367: 65–79, 2007.
30.
SchneiderC.A., RasbandW.S., and EliceiriK.W.NIH Image to ImageJ: 25 Years of image analysis.Nat Methods9: 671–675, 2012.
31.
MillerB.A., and ZhangW.TRP channels as mediators of oxidative stress.Adv Exp Med Biol704: 531–544, 2011.
32.
ChaudhuryS., BalM., BeluginS.AKAP150-mediated TRPV1 sensitization is disrupted by calcium/calmodulin.Mol Pain7: 34, 2011.
33.
LukacsV., YudinY., HammondG.R.Distinctive changes in plasma membrane phosphoinositides underlie differential regulation of TRPV1 in nociceptive neurons.J Neurosci33: 11451–11463, 2013.
34.
SteinA.T., Ufret-VincentyC.A., HuaL.Phosphoinositide 3-kinase binds to TRPV1 and mediates NGF-stimulated TRPV1 trafficking to the plasma membrane.J Gen Physiol128: 509–522, 2006.
35.
Ufret-VincentyC.A., KleinR.M., HuaL.Localization of the PIP2 sensor of TRPV1 ion channels.J Biol Chem286: 9688–9698, 2011.
36.
LiebermanP., MeltzerE.O., LaForceC.F.Two-week comparison study of olopatadine hydrochloride nasal spray 0.6% versus azelastine hydrochloride nasal spray 0.1% in patients with vasomotor rhinitis.Allergy Asthma Proc32: 151–158, 2011.
37.
SmithP.K., and CollinsJ.Olopatadine 0.6% nasal spray protects from vasomotor challenge in patients with severe vasomotor rhinitis.American J Rhinol Allergy25: e149–e152, 2011.
38.
O'ConnorT.M., O'ConnellJ., O'BrienD.I.The role of substance P in inflammatory disease.J Cellular Physiol201: 167–180, 2004.
39.
KalinerM.A., OppenheimerJ., and FarrarJ.R.Comprehensive review of olopatadine: The molecule and its clinical entities.Allergy Asthma Proc31: 112–119, 2010.
40.
BaraniukJ.N., and MerckS.J.Neuroregulation of human nasal mucosa.Ann N Y Acad Sci170: 604–609, 2009.
41.
SekiN., ShirasakiH., KikuchiM., and HimiT.Capsaicin induces the production of IL-6 in human upper respiratory epithelial cells.Life Sci80: 1592–1597, 2007.
42.
ZhouY., ShapiroM., DongQ.A calcium-activated chloride channel blocker inhibits goblet cell metaplasia and mucus overproduction.Novartis Found Symp248: 150–165, 2002.
43.
KettenmannH., HanischU.K., NodaM., and VerkhratskyA.Physiology of microglia.Physiol Rev91: 461–553, 2011.
44.
MercadoJ., Gordon-ShaagA., ZagottaW.N., and GordonS.E.Ca2+-dependent desensitization of TRPV2 channels is mediated by hydrolysis of phosphatidylinositol 4,5-bisphosphate.J Neurosci30: 13338–13347, 2010.
45.
MandadiS., NumazakiM., TominagaM.Activation of protein kinase C reverses capsaicin-induced calcium-dependent desensitization of TRPV1 ion channels.Cell Calcium35: 471–478, 2004.
46.
KushnarevaY.E., GerencserA.A., BossyB.Loss of OPA1 disturbs cellular calcium homeostasis and sensitizes for excitotoxicity.Cell Death Differ20: 353–365, 2013.
47.
SimmenT.Hax-1: A regulator of calcium signaling and apoptosis progression with multiple roles in human disease.Expert Opin Ther Targets15: 741–751, 2011.
48.
DragoI., De StefaniD., RizzutoR., and PozzanT.Mitochondrial Ca2+ uptake contributes to buffering cytoplasmic Ca2+ peaks in cardiomyocytes.Proc Natl Acad Sci U S A109: 12986–12991, 2012.
49.
FadeelB., and GrzybowskaE.HAX-1: A multifunctional protein with emerging roles in human disease.Biochim Biophys Acta1790: 1139–1148, 2009.
50.
HajnóczkyG., CsordásG., and YiM.Old players in a new role: Mitochondria-associated membranes, VDAC, and ryanodine receptors as contributors to calcium signal propagation from endoplasmic reticulum to the mitochondria.Cell Calcium32: 363–377, 2002.
51.
SennN., JeanclosE., and GarayR.Action of azelastine on intracellular Ca2+ in cultured airway smooth muscle.Eur J Pharmacol205: 29–34, 1991.
52.
NakamuraT., NishizawaY., SatoT., and YamatoC.Effect of azelastine on the intracellular Ca2+ mobilization in guinea pig peritoneal macrophages.Eur J Pharmacol148: 35–41, 1988.
53.
EhreC., ZhuY., AbdullahL.H.nPKCepsilon, a P2Y2-R downstream effector in regulated mucin secretion from airway goblet cells.Am J Physiol Cell Physiol293: C1445–C1454, 2007.
54.
BteshJ., FischerM.J., StottK., and McNaughtonP.A.Mapping the binding site of TRPV1 on AKAP79: Implications for inflammatory hyperalgesia.J Neurosci33: 9184–9193, 2013.
55.
SzolcsányiJ., and PintérE.Transient receptor potential vanilloid 1 as a therapeutic target in analgesia.Expert Opin Ther Targets17: 641–657, 2013.
BabesA., CiobanuA.C., NeacsuC., and BabesR.M.TRPM8, a sensor for mild cooling in mammalian sensory nerve endings.Curr Pharm Biotechnol12: 78–88, 2011.
58.
BandellM., StoryG.M., HwangS.W.Noxious cold ion channel TRPA1 is activated by pungent compounds and bradykinin.Neuron41: 849–857, 2004.
59.
LuttrellD.K., and LuttrellL.M.Signaling in time and space: G protein-coupled receptors and mitogen-activated protein kinases.Assay Drug Dev Technol1: 327–338, 2003.
60.
KalerG., OttoM., OkunA., and OkunI.Serotonin antagonist profiling on 5HT2A and 5HT2C receptors by nonequilibrium intracellular calcium response using an automated flow-through fluorescence analysis system, HT-PS 100.J Biomol Screen7: 291–301, 2002.
61.
BernsteinJ.A., LevinL.S., Al-ShuikE., and MartinV.T.Clinical characteristics of chronic rhinitis patients with high vs low irritant trigger burdens.Ann Allergy Asthma Immunol109: 173–178, 2012.
62.
TouskaF., MarsakovaL., TeisingerJ., and VlachovaV.A “cute” desensitization of TRPV1.Curr Pharm Biotechnol12: 122–129, 2011.
63.
NozadC.H., MichaelL. M., Betty LewD., and MichaelC. F.(2010). Non-allergic rhinitis: a case report and review.Clin Mol Allergy8: 1.
64.
PatestasM.A., GartnerL.P.Autonomic nervous system. In: A textbook of neuroanatomy.Malden, MA: Blackwell Publishing, 2006; Ch9: 118–133.
65.
SalibR. J., HarriesP. G., NairS. B., and HowarthP. H.(2008). Mechanisms and mediators of nasal symptoms in non-allergic rhinitis.Clin Exp Allergy38(3): 393–404.
