The potential of methylxanthine-based therapies in pediatric respiratory tract diseases

  • Ainhoa Oñatibia-Astibia
    Affiliations
    Official College of Pharmacists of Gipuzkoa, 20006 San Sebastian, Spain
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  • Eva Martínez-Pinilla
    Correspondence
    Corresponding author. Laboratory of Cell and Molecular Neuropharmacology, Center for Applied Medical Research (CIMA)-University of Navarra, Pio XII 55, 31008, Pamplona, Spain.
    Affiliations
    Neuroscience Department, Center for Applied Medical Research (CIMA), University of Navarra, 31008 Pamplona, Spain
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  • Rafael Franco
    Affiliations
    Molecular Neurobiology Laboratory, Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Barcelona, 08028 Barcelona, Spain

    CIBERNED, Centro de Investigación en Red, Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, 28049 Madrid, Spain

    Institute of Biomedicine of the University of Barcelona, IBUB, 08028, Barcelona, Spain
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Open ArchivePublished:February 01, 2016DOI:https://doi.org/10.1016/j.rmed.2016.01.022

      Highlights

      • Methylxanthines are natural products used to combat airway diseases.
      • Complementary molecular mechanisms are necessary for methylxanthine efficacy.
      • Methylxanthine are safe and may be used even in the apnea of prematurity.
      • Methylxanthines act locally on airways and centrally on respiratory control centers.

      Abstract

      Caffeine, theophylline and theobromine are the most known methylxanthines as they are present in coffee, tea and/or chocolate. In the last decades, a huge experimental effort has been devoted to get insight into the variety of actions that these compounds exert in humans. From such knowledge it is known that methylxanthines have a great potential in prevention, therapy and/or management of a variety of diseases. The benefits of methylxanthine-based therapies in the apnea of prematurity and their translational potential in pediatric affections of the respiratory tract are here presented.

      Keywords

      Abbreviations:

      AOP (Apnea of prematurity), IBMX (3-isobutyl-1-methylxantine), PDEs (phosphodiesterases), cAMP (cyclic AMP), CNS (central nervous system), CYP1A2 (1A2 isoenzyme of the cytochrome P450), FDA (Food and Drug Administration), IgE (Immunoglobulin E), NAEPP (National Asthma Education and Prevention Program), EPR3 (Expert Panel Report 3), WHO (World Health Organization), IL-10 (Interleukin 10), ECRHS (European Community Respiratory Health Survey), SARs (Slowly Adapting Stretch Receptors), COPD (Chronic Obstructive Pulmonary Disease)

      1. Introduction

      Diseases of the upper or lower respiratory tracts include a group of severe and disabling pathologies that affect a high percentage of the World population. Air pollution and the increase of environmental allergens have contributed in the last decades to a significant increase in respiratory-related morbidity and mortality. The incomplete development of the respiratory system and the immaturity of respiratory control are major issues in prematures and, accordingly, the majority of emergency room visits and hospitalizations in children are related to respiratory problems. Apart from classical treatments, which include a wide range of prescription drugs, natural compounds such as the methylxanthines in coffee and cacao have protective effects on the respiratory system. The present article focuses on the potential of methylxanthines in diseases of the airways with increasing prevalence in children, in particular in asthma, cough and the apnea of prematurity (AOP).

      2. Methylxanthines in natural sources

      Methylxanthines are natural components of cacao-related products or non-alcoholic beverages as coffee, tea or yerba mate. Methylxanthines are also present in cola drinks and in several plant species as Camellia ptilophylla (cocoa tea), Paulliania cupana (guaraná) or Citrus sp., used to prepare dietary supplements [
      • Ashihara H.
      • Suzuki T.
      Distribution and biosynthesis of caffeine in plants.
      ,
      • Kretschmar J.A.
      • Baumann T.
      Caffeine in Citrus flowers.
      ]. Seven natural methylxanthines have been so far identified: aminophylline, 3-isobutyl-1-methylxantine (IBMX), paraxanthine, pentoxifylline, theobromine, theophylline and caffeine. The latter three have been more extensively studied due to their presence in coffee, tea and/or chocolate.
      Caffeine concentration in coffee cups, which depends on the coffee plant varietals and the preparation method, ranges from 57 mg/100 ml in a cup made from grounded and toasted coffee to 2 mg/100 ml in decaffeinated preparations [

      Eufic.org [internet]. The European Food Information Council (EUFIC) [cited 2014 Nov 14]. Available from: http://www.eufic.org/.

      ]. Coffee also has theobromine and theophylline but at a 20-fold lower concentration [
      • Bispo M.S.
      • Veloso M.C.
      • Pinheiro H.L.
      • De Oliveira R.F.
      • Reis J.O.
      • De Andrade J.B.
      Simultaneous determination of caffeine, theobromine, and theophylline by high-performance liquid chromatography.
      ]. Preparations from yerba mate leaves (Ilex paraguariensis), known as mate tea, contain the highest concentration of theophylline followed by black and green tea [
      • Bispo M.S.
      • Veloso M.C.
      • Pinheiro H.L.
      • De Oliveira R.F.
      • Reis J.O.
      • De Andrade J.B.
      Simultaneous determination of caffeine, theobromine, and theophylline by high-performance liquid chromatography.
      ,
      • Hicks M.B.
      • Hsieh Y.H.P.
      • Bell L.N.
      Tea preparation and its influence on methylxanthine concentration.
      ]. Finally, theobromine is the most abundant methylxanthine in cacao (Theobroma cacao) and cacao by-products, where the theobromine amount is directly related with the percentage of cacao, e.g. in dark chocolate more than in milk chocolate [
      • Langer S.
      • Marshall L.J.
      • Day A.J.
      • Morgan M.R.
      Flavanols and methylxanthines in commercially available dark chocolate: a study of the correlation with nonfat cocoa solid.
      ].

      3. Physiological effects of methylxanthines

      3.1 Mechanism of action

      Four different cellular mechanisms of action have been described for methylxanthines: mobilization of intracellular calcium, inhibition of phosphodiesterases (PDEs), modulation of GABAA receptors and antagonism of adenosine receptors (Fig. 1). Recent evidence however points to further modes of action that may be relevant in translational research.
      Figure thumbnail gr1
      Fig. 1Molecular mechanisms and physiological actions of methylxanthines.
      Methylxanthines affect the concentration of the two main second messengers in mammals, cyclic AMP (cAMP) and Ca2+. On the one hand, methylxanthines facilitate the release of Ca2+ from the endoplasmic reticulum in both skeletal and cardiac muscle. On the other hand, methylxanthines increase cAMP levels through the inhibition of PDEs [
      • Butcher R.W.
      • Sutherland E.W.
      Adenosine 3′,5′-phosphate in biological materials. I. Purification and properties of cyclic 3′,5′-nucleotide phosphodiesterase and use of this enzyme to characterize adenosine 3′,5′-phosphate in human urine.
      ]. Due to structural similarity with purine nucleosides, caffeine, theophylline and theobromine can act as competitive inhibitors of adenosine receptors. In humans there are four widely distributed sub-types of these G-protein coupled receptors, A1, A2A, A2B and A3. In fact, every cell in the human body has at least one subtype of adenosine receptors. Cells in many tissues express one or several of these four subtypes. For instance, smooth muscle and endothelial cells may express A1, A2A and A2B receptor subtypes, whereas skeletal muscle mainly expresses A2A and A2B receptor subtypes, A2A in type 1 and A2B in type 2 fibers [
      • Lynge J.
      • Hellsten Y.
      Distribution of adenosine A1, A2A and A2B receptors in human skeletal muscle.
      ]. The A3 subtype is less studied but its expression has been reported in lungs and liver [
      • Salvatore C.A.
      • Jacobson M.A.
      • Taylor H.E.
      • Linden J.
      • Johnson R.G.
      Molecular cloning and characterization of the human A3 adenosine receptor.
      ], brain [
      • Zhou Q.Y.
      • Li C.
      • Olah M.E.
      • Johnson R.A.
      • Stiles G.L.
      • Civelli O.
      Molecular cloning and characterization of an adenosine receptor: the A3 adenosine receptor.
      ], colon [
      • Gessi S.
      • Cattabriga E.
      • Avitabile A.
      • Gafa R.
      • Lanza G.
      • Cavazzini L.
      • et al.
      Elevated expression of A3 adenosine receptors in human colorectal cancer is reflected in peripheral blood cells.
      ], eosinophils [
      • Kohno Y.
      • Ji X.
      • Mawhorter S.D.
      • Koshiba M.
      • Jacobson K.A.
      Activation of A3 adenosine receptors on human eosinophils elevates intracellular calcium.
      ] and dendritic cells [
      • Fossetta J.
      • Jackson J.
      • Deno G.
      • Fan X.
      • Du X.K.
      • Bober L.
      • et al.
      Pharmacological analysis of calcium responses mediated by the human A3 adenosine receptor in monocyte-derived dendritic cells and recombinant cells.
      ]. Adenosine receptor blockade seems to be the responsible for many of the central nervous system (CNS) effects of methylxanthines. Concentrations reached after dietary intake are high enough to block adenosine receptors.

      3.2 Methylxanthine metabolism

      Theobromine and theophylline are intermediate molecules of caffeine metabolism that has been studied in animal models but, more predominantly, in man. Following oral administration, gastrointestinal absorption of caffeine is fast and complete, reaching almost 100% of bioavailability. Once it reaches the bloodstream, where it binds to albumin, caffeine is distributed in all tissues by simple diffusion or carrier-mediated transport. Caffeine metabolism is hepatic and mainly mediated by the 1A2 isoenzyme of the cytochrome P450 (CYP1A2), which is the responsible for the between-subject variability of caffeine plasma concentration [
      • Brosen K.
      Drug interactions and the cytochrome P450 system. The role of cytochrome P450 1A2.
      ]. The half-life of caffeine is 4.1 h, but it may be higher during pregnancy, in patients with hepatic diseases or in infants and neonates (up to 100 h) [
      • Begas E.
      • Kouvaras E.
      • Tsakalof A.
      • Papakosta S.
      • Asprodini E.K.
      In vivo evaluation of CYP1A2, CYP2A6, NAT-2 and xanthine oxidase activities in a Greek population sample by the RP-HPLC monitoring of caffeine metabolic ratios.
      ]. The main products of the first steps in caffeine metabolism are paraxanthine, theobromine and theophylline (Fig. 2) [
      • Tassaneeyakul W.
      • Birkett D.J.
      • McManus M.E.
      • Tassaneeyakul W.
      • Veronese M.E.
      • Andersson T.
      • et al.
      Caffeine metabolism by human hepatic cytochromes P450: contributions of 1A2, 2E1 and 3A isoforms.
      ]. Due to its actions on CYP1A2 expression, some lifestyles -as smoking- increase caffeine clearance [
      • Fabes M.S.
      • Fuhr U.
      Time response of cytochrome P450 1A2 activity on cessation of heavy smoking.
      ]. Theobromine, theophylline and the final product, methyluric acid, are excreted via urine. Although theobromine is toxic for a variety of mammals, the incidence of toxicity in humans is virtually absent; toxic events may occur when methylxanthines are taken in combination with dietary supplements for weight loss [
      • Pendleton M.
      • Brown S.
      • Thomas C.
      • Odle B.
      Potential toxicity of caffeine when used as a dietary supplement for weight loss.
      ,
      • Pendleton M.
      • Brown S.
      • Thomas C.M.
      • Odle B.
      Potential toxicity of caffeine when used as a dietary supplement for weight loss.
      ] or with drugs of abuse [
      • Baggott M.J.
      • Childs E.
      • Hart A.B.
      • De Bruin E.
      • Palmer A.A.
      • Wilkinson J.E.
      • et al.
      Psychopharmacology of theobromine in healthy volunteers.
      ].
      Figure thumbnail gr2
      Fig. 2Scheme of caffeine metabolism in man. The name of the enzyme catalyzing each reaction is provided on each arrow. A not yet identified enzyme, 1.13.12.X, belongs to the family of oxidoreductases acting on single donors and using a single oxygen atom (IUPAC, Enzyme Commission). * CYP Cytochrome p450; family 1, 2 and/or 3.

      4. Methylxanthines for AOP therapy

      AOP is the most common problem in premature neonates; it refers to a period of time longer than 20 s with no breathing, often accompanied by hypoxia, bradycardia, cyanosis, pallor and/or severe hypotonia. It usually affects infants born at a gestational age of 37 weeks or less [

      National Institutes of Health Consensus Development Conference on Infantile Apnea and Home Monitoring, Sept 29 to Oct 1, 1986. Pediatrics 1987;79: 292–299.

      ]. The incidence of apnea is inversely related to gestational stage [
      • Lorch S.A.
      • Srinivasan L.
      • Escoobar G.J.
      Epidemiology of apnea and bradycardia resolution in premature infants.
      ]: 7, 15, 54 and 100% at gestational ages of, respectively, 34–35, 32–33, 30–31 and less than 29 weeks. As the infant's lungs and central respiratory control mature, AOP resolves such that it is more of a developmental disorder rather than a disease [
      • Picone S.
      • Bedetta M.
      • Paolillo P.
      Caffeine citrate: when and for how long. A literature review.
      ].
      AOP is traditionally classified as central, obstructive or mixed. Central apnea is characterized by cycles of disrupted breathing during sleep, obstructive apnea exhibits regular respiratory thoracic movements but inadequate nasal air flow, and the mixed type results from a combination of both [
      • Finer N.N.
      • Barrington K.J.
      • Hayes B.J.
      • Hugh A.
      Obstructive, mixed, and central apnea in the neonate: physiologic correlates.
      ]. Central apnea accounts for approximately 40% of all cases of apnea, obstructive apnea accounts for a 10% and the mixed type accounts for a 50% [
      • Zhao J.
      • Gonzalez F.
      • Mu D.
      Apnea of prematurity: from cause to treatment.
      ].
      Several theories regarding the pathogenesis of AOP have been reported but none is fully accepted. Hypercapnia detected by central chemoreceptors may be resumed by the increase of ventilation and by the increase in tidal volume and breathing frequency [
      • Putnam R.W.
      • Conrad S.C.
      • Gdovin M.J.
      • Erlichman J.S.
      • Leiter J.C.
      Neonatal maturation of the hypercapnic ventilatory response and central neural CO2 chemosensitivity.
      ]. Premature infants may not be able to afford these automatic responses due, for instance, to failure in proper respiratory muscle control. In fact, some in vivo studies have demonstrated a differential threshold for hypercapnia in preterm infants and that contraction of diaphragm before that of upper airway muscles favors AOP [
      • Darnall R.A.
      The role of CO2 and central chemoreception in the control of breathing in the fetus and the neonate.
      ,
      • Carlo W.A.
      • Martin R.J.
      • Difiore J.M.
      Differences in CO2 threshold of respiratory muscles in preterm infants.
      ]. It should be noted that the difference between basal pCO2 and pCO2 triggering apneic responses is very small in neonates. In this sense, minor oscillations in breathing in preterm infants may lead to abnormal respiratory homeostasis [
      • Morin D.
      • Di Pasquale E.
      • Hilaire G.
      • Monteau R.
      Possible involvement of serotonin in obstructive apnea of the newborn.
      ,
      • Khan A.
      • Qurashi M.
      • Kwiatkowski K.
      • Cates D.
      • Rigatto H.
      Measurement of the CO2 apneic threshold in newborn infants: possible relevance for periodic breathing and apnea.
      ]. The hyperactive laryngeal chemoreflex, in response to pharyngeal reflux of gastric content, constitutes another possible cause for infantile apnea [
      • Heman-Ackah Y.D.
      • Pernell K.J.
      • Goding Jr., G.S.
      The laryngeal chemoreflex: an evaluation of the normoxic response.
      ,
      • Thach B.T.
      Reflux associated apnea in infants: evidence for a laryngeal chemoreflex.
      ].

      4.1 Caffeine and theophylline in the therapy of AOP

      The clinical management of AOP includes prevention, supportive measures and pharmacological and oxygen therapies. Supportive measures imply prone position, tactile stimulation, continuous positive airway pressure, and/or mechanical ventilation, whereas prevention comes from an appropriate posture of the neck and the maintenance of body temperature between 36.5 and 37 °C [
      • Henderson-Smart D.J.
      • Steer P.A.
      Caffeine versus theophylline for apnea in preterm infants.
      ,
      • Bhatia J.
      Current options in the management of apnea prematurity.
      ,
      • Paolillo P.
      • Picone S.
      Apnea of prematurity.
      ]. One of the tried interventions, blood transfusion to increase oxygen transport capacity, has been shown ineffective [
      • Paolillo P.
      • Picone S.
      Apnea of prematurity.
      ,
      • Valieva O.A.
      • Strandjord T.P.
      • Mayock D.E.
      • Juul S.E.
      Effects of transfusions in extremely low birth weight infants: a retrospective study.
      ,
      • Westkamp E.
      • Soditt V.
      • Adrian S.
      • Bohnhorst B.
      • Groneck P.
      • Poets C.F.
      Blood transfusion in anemic infants with apnea of prematurity.
      ]. In contrast, both theophylline administration and CO2 inhalation are effective [
      • Bhatia J.
      Current options in the management of apnea prematurity.
      ,
      • Al-Saif S.
      • Alvaro R.
      • Manfreda J.
      • Kwiatkowski K.
      • Cates D.
      • Qurashi M.
      • et al.
      A randomized controlled trial of theophylline versus CO2 inhalation for treating apnea of prematurity.
      ]. A classical therapy to reduce the amount of apneas consists of inhalation of CO2-containing air for 2 h. Despite clinical benefits in reducing the apneic time and rate from 9 s/min and 94 episodes/hour to 3 s/min and 34 episodes/hour, inhalation of CO2-containing air suffers one limitation related with the quickly accommodation to the CO2 concentration in inspired air [
      • Paolillo P.
      • Picone S.
      Apnea of prematurity.
      ,
      • Westkamp E.
      • Soditt V.
      • Adrian S.
      • Bohnhorst B.
      • Groneck P.
      • Poets C.F.
      Blood transfusion in anemic infants with apnea of prematurity.
      ,
      • Al-Saif S.
      • Alvaro R.
      • Manfreda J.
      • Kwiatkowski K.
      • Cates D.
      • Qurashi M.
      • et al.
      A randomized controlled trial of theophylline versus CO2 inhalation for treating apnea of prematurity.
      ].
      Among all the pharmacological treatments, methylxanthines such as caffeine, theophylline and aminophylline, are a good choice to treat AOP [
      • Henderson-Smart D.J.
      • Steer P.A.
      Caffeine versus theophylline for apnea in preterm infants.
      ,
      • Bhatia J.
      Current options in the management of apnea prematurity.
      ]. Kuzenko (1973) and Kuzenko and Paala (1973) reported the benefits of rectal administration of aminophylline in the therapy of AOP [
      • Kuzemko J.A.
      Aminophylline in apnoeic attacks of newborn.
      ,
      • Kuzemko J.A.
      • Paala J.
      Apnoeic attacks in the newborn treated with aminophylline.
      ]. These findings were confirmed later by Bednarek and Roloff [
      • Bednarek F.J.
      • Roloff D.W.
      Treatment of apnea of prematurity with aminophylline.
      ] and Shannon et al. [
      • Shannon D.C.
      • Gotay F.
      • Stein I.M.
      • Rogers M.C.
      • Todres I.D.
      • Moylan F.M.
      Prevention of apnea and bradycardia in low-birthweight infants.
      ] that introduced the therapeutic use of oral administration of theophylline. Several studies have been undertaken to improve the effectiveness of this treatment. The optimal therapeutic doses of theophylline are 4 mg/kg and 1.5 mg/kg in, respectively, neonates and preterm neonates. Optimal plasma concentration should be in the 8–10 μg/ml range, but plasma levels higher than 14–16 μg/ml lead to undesirable side effects like sleeplessness, hyperglycemia, hypertension and/or heart arrhythmias [
      • Roberts R.
      Drug Therapy in Infants.
      ].
      Methylxanthine therapy is a mainstay of treatment of central apnea by stimulating the CNS and respiratory muscle function [
      • Kreutzer K.
      • Bassler D.
      Caffeine for apnea of prematurity: a neonatal success story.
      ,
      • Aubier M.
      • Murciano D.
      • Viires N.
      • Lecocguic Y.
      • Pariente R.
      Respiratory muscle pharmacotherapy.
      ,
      • Van der Heijden H.F.
      • Dekhuijzen P.N.
      • Folgering H.
      • Van Herwaarden C.L.
      Pharmacotherapy of respiratory muscles in chronic obstructive pulmonary disease.
      ]. As a non-selective antagonist of adenosine receptors, caffeine may improve minute ventilation, CO2 sensitivity, contraction of the diaphragmatic muscles and neural respiratory regulation [
      • Henderson-Smart D.J.
      • Steer P.A.
      Caffeine versus theophylline for apnea in preterm infants.
      ,
      • Aranda J.V.
      • Beharry K.
      • Valencia G.B.
      • Natarajan G.
      • Davis J.
      Caffeine impact on neonatal morbidities.
      ]. These benefits of caffeine in decreasing the frequency of apneic episodes in infants were first demonstrated in late 70 s [
      • Aranda J.V.
      • Gorman W.
      • Bergsteinsson H.
      • Gunn T.
      Efficacy of caffeine in treatment of apnea in the low-birth-weight infant.
      ]. However, it was not until 2000, when the Food and Drug Administration (FDA) approved caffeine for the treatment of AOP, that this methylxanthine emerged as a good alternative to theophylline. Caffeine citrate can be administrated orally or intramuscularly with a recommended initial dose of 20 mg/kg, followed by a daily maintenance dose of 5 mg/kg [
      • Arora P.
      Pathogenesis and management of apnea of prematurity.
      ,
      • Steer P.
      • Flenady V.
      • Shearman A.
      • Charles B.
      • Gray P.H.
      • Henderson-Smart D.
      • et al.
      High dose caffeine citrate for extubation of preterm infants: a randomised controlled trial.
      ]. It is well known that methylxanthines cross the blood brain barrier and, accordingly, central actions impact on CO2 sensitivity of chemoreceptors, diaphragm and respiratory muscle contraction and on catecholamine-induced responses [
      • Zhao J.
      • Gonzalez F.
      • Mu D.
      Apnea of prematurity: from cause to treatment.
      ]. As mentioned above, all these anti-AOP actions may be due to blockade of adenosine receptors, especially of A1 and A2A [
      • Martin R.J.
      • Abu-Shaweesh J.M.
      • Baird T.M.
      Apnoea of prematurity.
      ].
      Caffeine, theophylline, and aminophylline are non-selective antagonists of adenosine receptors, being adenosine a centrally-acting respiratory depressant involved in the inhibition of respiratory drive [
      • Andreas S.
      • Kreuzer H.
      Cheyne-Stokes respiration in patients with congestive heart failure.
      ,
      • Koos B.J.
      • Maeda T.
      • Jan C.
      Adenosine A[1] and A[2A] receptors modulate sleep state and breathing in fetal sheep.
      ,
      • Watt A.H.
      • Buss D.C.
      • Routledge P.A.
      Effect of aminophylline on the respiratory depressant action of intravenous adenosine in neonatal rabbits.
      ]. In this sense, some authors demonstrated, in newborn piglets, that methylxanthines acting on adenosine A2A receptors expressed in GABAergic neurons can prevent the inhibition of laryngeal chemoreflex induced by GABA release [
      • Kuzemko J.A.
      • Paala J.
      Apnoeic attacks in the newborn treated with aminophylline.
      ,
      • Abu-Shaweesh J.M.
      Activation of central adenosine A[2A] receptors enhances superior laryngeal nerve stimulation-induced apnea in piglets via a GABAergic pathway.
      ]. The possible role of adenosine A1 receptors in this phenomenon cannot be ruled out.
      Epidemiological evidence shows both theophylline and caffeine as highly successful molecules at reducing apneic episodes of neonates by acting on the CNS [
      • Gannon B.A.
      Theophylline or caffeine: which is best for apnea of prematurity?.
      ]. A comparative study demonstrated that caffeine performs better than theophylline in terms of efficacy, peripheral side effects or drug clearance (Table 1). In fact, theophylline is generally less well tolerated and more prone to cause tachycardia and/or gastrointestinal dysfunction. In contrast, caffeine has a faster onset of action, lesser fluctuations in plasma concentration, longer half-life and fewer peripheral side effects [
      • Picone S.
      • Bedetta M.
      • Paolillo P.
      Caffeine citrate: when and for how long. A literature review.
      ,
      • Aranda J.V.
      • Turmen T.
      Methylxanthines in apnea of prematurity.
      ].
      Table 1Caffeine and theophylline in the treatment of apnea of prematurity.
      Adapted from Ref. 
      • Gannon B.A.
      Theophylline or caffeine: which is best for apnea of prematurity?.
      (t1/2: time required for reducing the plasma concentration of the drug to a half).
      VariableCaffeineTheophylline
      Efficacy++++++
      Peripheral side effects+/−+ + +
      Drug clearance (t1/2, hours)10030
      Plasma level at steady stateStableFluctuating
      Dosing intervalOnce a day1–3 times per day

      5. Methylxanthines for asthma therapy

      Asthma is a chronic respiratory disease that usually starts at a young age. It is characterized by inflammation and constriction of the airways with episodes that can be life-threatening. The main symptoms are wheezing and serious coughing periods that severely affect the ability to breathe [

      nhlbi.nih.gov [internet]. National Institutes of Health. What Is Asthma? [Cited 2014 Nov 12]. Available from:http://www.nhlbi.nih.gov/health/health-topics/topics%20/asthma/.

      ]. Despite asthma was described 2000 years ago, anti-inflammatory drugs were not used until the 1960 s [

      who.int [internet]. World Health Organization. Asthma: Definition. [Cited 2014 Nov 17]. Available from: http://www.who.int/respiratory/asthma/definition.

      ].
      An expert panel commissioned in 2007 by the National Asthma Education and Prevention Program (NAEPP), developed a guideline for the diagnosis and management of asthma, namely the “Expert Panel Report 3” (EPR 3). According to the World Health Organization (WHO) estimation, there are 235 million people with asthma, mainly children. Despite the effective therapies, asthma is a deadly disease, especially in lower-middle income countries where this pathology is under-diagnosed and poorly managed due to the high cost of the available therapies [
      • Walker M.L.
      • Holt K.E.
      • Anderson G.P.
      • Teo S.M.
      • Sly P.D.
      • Holt P.G.
      • et al.
      Elucidation of pathways driving asthma pathogenesis: Development of a systems-level analytic strategy.
      ].
      Main risks are airway immaturity, genetic predisposition and the environmental exposure to substances and particles that may cause allergic reactions and/or irritation. Triggers may be quite diverse: medicines (i.e. aspirin), chemical irritants in the workplace (i.e. detergents or metals), household allergens (i.e. dust or pet dander), outdoor allergens (i.e. pollen and mold), extreme emotional arousal (i.e. anger or fear) and strenuous physical exercise [

      who.int [internet]. World Health Organization. Asthma: Definition. [Cited 2014 Nov 17]. Available from: http://www.who.int/respiratory/asthma/definition.

      ]. Several explanations have been provided such as association of inflammation of airways in asthma with allergic sensitization. The respiratory tract is able to recognize common environmental allergens and generate a specific and exacerbated immunologic response that can induce an immunoglobulin E (IgE)-dependent inflammatory process mediated by histamine, tryptase, leukotrienes and/or prostaglandins [
      • Holgate S.T.
      Pathogenesis of Asthma.
      ,
      • Beasley R.
      • Pekkanen J.
      • Pearce N.
      Has the role of atopy in the development of asthma been over-emphasized?.
      ]. Other factors influencing the immune system maturation and impacting on the prevalence of allergic and autoimmune diseases in childhood are infectious agents [
      • Strachan D.P.
      Family size, infection and atopy: the first decade of the «hygiene hypothesis».
      ].

      5.1 Theophylline and doxofylline in the therapy of asthma

      Management of asthma includes both, effective treatment of acute attacks and long-term control medications. Standard therapies to alleviate bronchoconstriction and asthma symptoms while decreasing the frequency and severity of asthma episodes include short-and long-acting β-adrenergic agonists (e.g. formoterol and salmeterol), anti-cholinergic agents and inhaled corticosteroids (e.g. salbutamol, tertbutaline, formoterol, salmeterol, fluticasone, beclomethasone, budesonide, or ciclesonide) [
      • Wong J.J.
      • Lee J.H.
      • Turner D.A.
      • Rehder K.J.
      A review of the use of adjunctive therapies in severe acute asthma exacerbation in critically ill children.
      ]. Other treatments with moderate benefits include leukotriene antagonists (e.g. montelukast) and long-acting muscarinic antagonists (e.g. tiotropium or aclidinium). Although currently available anti-asthma drugs exhibit good results in terms of efficacy, they show some limitations related with adherence, tolerability, adverse side-effects and/or high cost [
      • Chung K.F.
      • Wenzel S.E.
      • Brozek J.L.
      • Bush A.
      • Castro M.
      • Sterk P.J.
      • et al.
      International ERS/ATS guidelines on definition, evaluation and treatment of severe asthma.
      ]. Inhaled corticosteroid therapy, for instance, is not effective in all asthmatic patients; some children do not respond to steroids whereas others show a substantial decrease of the beneficial effects within months after corticosteroids are discontinued [
      • Guilbert T.W.
      • Morgan W.J.
      • Zeiger R.S.
      • Mauger D.T.
      • Boehmer S.J.
      • Szefler S.J.
      • et al.
      Long-term inhaled corticosteroids in preschool children at high risk for asthma.
      ,
      • Warner S.M.
      • Knight D.A.
      Airway modeling and remodeling in the pathogenesis of asthma.
      ,
      • Saglani S.
      • Lui S.
      • Ullmann N.
      • Campbell G.A.
      • Sherburn R.T.
      • Mathie S.A.
      • et al.
      IL-33 promotes airway remodeling in pediatric patients with severe steroid-resistant asthma.
      ]. Adherence to the therapies is also problematic due to undesirable effects, such as decrease in bone density, alterations in adrenal function or cataracts [
      • Partridge M.R.
      • van der Molen T.
      • Myrseth S.E.
      • Busse W.W.
      Attitudes and actions of asthma patients on regular maintenance therapy: the INSPIRE study.
      ,
      • Bosley C.M.
      • Parry D.T.
      • Cochrane G.M.
      Patient compliance with inhaled medication: does combining beta-agonists with corticosteroids improve compliance?.
      ,
      • Lipworth B.J.
      Systemic adverse effects of inhaled corticosteroid therapy: A systematic review and meta-analysis.
      ,
      • Nelson H.S.
      • Weiss S.T.
      • Bleecker E.R.
      • Yancey S.W.
      • Dorinsky P.M.
      SMART Study Group. The Salmeterol Multicenter Asthma Research Trial: a comparison of usual pharmacotherapy for asthma or usual pharmacotherapy plus salmeterol.
      ]. It is known that theobromine and caffeine improve lung function and produce bronchodilatation in asthma patients [
      • Bara A.I.
      • Barley E.A.
      Caffeine for asthma.
      ,
      • Simons F.E.
      • Becker A.B.
      • Simons K.J.
      • Gillespie C.A.
      The bronchodilator effect and pharmacokinetics of theobromine in young patients with asthma.
      ]. Also, it has been demonstrated that patients with asthma and bronchitis may self-administer coffee or chocolate due to perceived symtomatic relief [
      • Zhao J.
      • Gonzalez F.
      • Mu D.
      Apnea of prematurity: from cause to treatment.
      ].
      Theophylline has been used for various decades in the treatment of asthma and is still among the most prescribed drugs; it is effective and relatively inexpensive [
      • Barnes P.J.
      Pathophysiology of asthma.
      ]. Theophylline has a bronchodilator action and may decrease inflammation underlying asthma [
      • Pauwels R.
      The effects of theophylline on airway inflammation.
      ]. Moreover, theophylline also produces a relaxation in airways and of vascular smooth muscle thus decreasing the mean pulmonary arterial pressure [
      • Rabe K.F.
      • Magnussen H.
      • Dent G.
      Theophylline and selective PDE inhibitors as bronchodilators and smooth muscle relaxants.
      ]. These actions are particularly relevant when asthma is not sufficiently controlled by inhaled corticosteroids with or without concomitant long-acting β2-adrenergic agonists. In these situations theophylline is used as an effective add-on therapy [
      • Walker M.L.
      • Holt K.E.
      • Anderson G.P.
      • Teo S.M.
      • Sly P.D.
      • Holt P.G.
      • et al.
      Elucidation of pathways driving asthma pathogenesis: Development of a systems-level analytic strategy.
      ].
      In prospective randomized clinical trials, in cohorts of patients with severe asthma, low-dose addition of theophylline to inhaled corticosteroids results in a better disease management than doubling the dose of corticosteroids or than the use of β2-adrenergic agonists [
      • Barnes P.J.
      Pathophysiology of asthma.
      ,
      • Evans D.J.
      • Taylor D.A.
      • Zetterstrom O.
      • Chung K.F.
      • O'Connor B.J.
      • Barnes P.J.
      A comparison of low-dose inhaled budesonide plus theophylline and high-dose inhaled budesonide for moderate asthma.
      ,
      • Ukena D.
      • Harnest U.
      • Sakalauskas R.
      • Magyar P.
      • Vetter N.
      • Steffen H.
      • et al.
      Comparison of addition of theophylline to inhaled steroid with doubling of the dose of inhaled steroid in asthma.
      ]. PDEs inhibition is the most accepted mechanism of action for theophylline in asthma pathology. Muscle contraction is regulated by intracellular levels of cAMP, which is synthesized by adenylate cyclases and hydrolyzed by PDEs. Rabe et al., in 1995 demonstrated that theophylline is able to relax the inherent tone of human bronchial rings at concentrations similar to those inhibiting cAMP hydrolysis by PDEs [
      • Rabe K.F.
      • Magnussen H.
      • Dent G.
      Theophylline and selective PDE inhibitors as bronchodilators and smooth muscle relaxants.
      ]. Specifically, theophylline induces bronchodilatation by inhibition of PDE3 activity whereas its anti-inflammatory effect may be due to inhibition of PDE4 together with histone deacetylase-2 activation [
      • Barnes P.J.
      Targeting the epigenome in the treatment of asthma and chronic obstructive pulmonary disease.
      ]. Adenosine receptor antagonism [
      • Polosa R.
      • Blackburn M.R.
      Adenosine receptors as targets for therapeutic intervention in asthma and chronic obstructive pulmonary disease.
      ] or increase of interleukin 10 (IL-10) [
      • Mascali J.J.
      • Cvietusa P.
      • Negri J.
      • Borish L.
      Anti-inflammatory effects of theophylline: modulation of cytokine production.
      ] and the nuclear translocation of the nuclear factor kappa-light-chain-enhancer of activated B cells [
      • Ichiyama T.
      • Hasegawa S.
      • Matsubara T.
      • Hayashi T.
      • Furukawa S.
      Theophylline inhibits NF-kappa B activation and I kappa B alpha degradation in human pulmonary epithelial cells.
      ] are further proposed mechanisms. Beneficial effects only take place at high concentrations in therapies that use theophylline alone. In this sense, the conventional dosing strategy consisting of oral administration of 200–400 mg twice per day may cause adverse effects. An excessive inhibition of adenosine receptors could produce gastrointestinal (stomach ache, nausea or diarrhea), CNS (headache, irritability or difficulties concentrating at work) or cardiovascular (irregular heart rate or palpitations) manifestations. A synthetic methylxanthine, doxofylline, which differs from theophylline by the presence of a dioxolone group in position 7, has been developed to limit undesirable side effects. Different studies in animal models as well as in humans have found that this drug shows similar efficacy as theophylline but with less adverse effects [
      • Shukla D.
      • Chakraborty S.
      • Singh S.
      • Mishra B.
      Doxofylline: a promising methylxanthine derivative for the treatment of asthma and chronic obstructive.
      ]. In this sense, the lower affinity to A1 and A2-adenosine receptors that shows doxofylline may be related to these less adverse reactions [
      • Franzone J.S.
      • Cirillo R.
      • Barone D.
      Doxofylline and theophylline are xanthines with partly different mechanisms of action in animals.
      ,
      • Cirillo R.
      • Barone D.
      • Franzone J.S.
      Doxofylline, an antiasthmatic drug lacking affinity for adenosine receptors.
      ,
      • Sankar J.
      • Lodha R.
      • Kabra S.K.S.
      Doxofylline: The next generation methylxanthine.
      ,
      • Goldstein M.F.
      • Chervinsky P.
      Efficacy and safety of doxofylline compared to theophylline in chronic reversible asthma–a double-blind randomized placebo-controlled multicentre clinical trial.
      ,
      • Dini F.L.
      • Cogo R.
      Doxofylline: a new generation xanthine bronchodilator devoid of major cardiovascular adverse effects.
      ].

      6. Methylxanthines in cough management

      Cough is a protective reflex that helps in keeping airway integrity and prevents lung infections. However, a reduction in the threshold for reflex initiation leads to cough in response to stimuli that are normally innocuous. Persistent coughing is a health problem that impairs life quality and causes chest pain, loss of bladder control or headedness [
      • Mazzone S.B.
      An overview of the sensory receptors regulating cough.
      ,

      nhlbi.nih.gov [internet]. National Institutes of Health. What Is Cough? [Cited 2014 Oct 30]. Available from:http://www.nhlbi.nih.gov/health/health-topics/topics/%20cough/.

      ]. Furthermore, infections that affect the upper (common cold, flu, laryngitis o sinusitis) or the lower (bronchitis and pneumonia) tract often lead to episodes of severe coughing. Non-infectious causes of chronic cough include allergic rhinitis, asthma, hypertension or cardiovascular diseases and require pharmacological treatment of the underlying condition [

      Nhs.uk [internet]. Natioanl Health System. Cough–Causes - NHS Choices. [Cited 2014 Oct 30]. Available from:http://www.nhs.uk/Conditions/Cough/Pages/%20Causes.aspx.

      ]. Despite of the idea that this reflex is a normal body reaction, the prevalence of chronic cough is high (18%). The European Community Respiratory Health Survey (ECRHS) estimated that the incidence is higher in females than in males and is linked to pathologies like asthma or obesity and/or to unhealthy lifestyles like tobacco smoking [
      • Chung K.F.
      • Widdicombe J.G.
      • Boushey H.A.
      Cough: Causes, Mechanisms and Therapy.
      ].
      Along the respiratory tract, neurons expressing C-fibers and rapidly and slowly adapting stretch receptors (SARs) can be activated in response to mechanical and physico-chemical stimuli (acid, temperature), or natural irritants (capsaicin) [
      • Polverino M.
      • Polverino F.
      • Fasolino M.
      • Andò F.
      • Alfieri A.
      • De Blasio F.
      Anatomy and neuro-pathophysiology of the cough reflex arc.
      ,
      • Karlsson J.A.
      • Sant'Ambrogio G.
      • Widdicombe J.
      Afferent neural pathways in cough and reflex bronchoconstriction.
      ]. The reflex impulse arrives to the cerebral cough center, located in the upper brain stem and pons, via the vagus nerve. Vagus, phrenic, and spinal motor nerves constitute the efferent pathway arriving to diaphragm, abdominal wall and expiratory muscles and generate a coordinated response [
      • Dinh Q.T.
      • Heck S.
      • Le D.D.
      • Bals R.
      • Welte T.
      Pathophysiology, diagnostics and therapy of chronic cough: neuronal reflexes and antitussiva.
      ].

      6.1 Theobromine to alleviate persistent coughing

      The pharmacological management of cough is complex. The inhibition of the cerebral cough center by codeine is one of the most effective treatments. This opiate may be used alone or in combination with anti-inflammatory or antipyretic drugs. Due to undesirable effects, codeine prescription is contraindicated in newborn infants and in children undertaking surgery; its use in adults is also decreasing. Apart from eliminating or, at least, reducing the exposition to irritants, expectorants and mucolytic drugs decrease the viscosity of bronchial secretion thus facilitating its release. Acetylcysteine, for instance, is able to break disulfide bonds in mucoproteins, whereas other drugs, ambroxol or carbocysteine, can modulate the viscoelastic properties of the mucus [
      • Macciò A.
      • Madeddu C.
      • Panzone F.
      • Mantovani G.
      Carbocysteine: clinical experience and new perspectives in the treatment of chronic inflammatory diseases.
      ,
      • Wunderer H.
      • Morgenroth K.
      • Weis G.
      The cleaning system of the airways: physiology, pathophysiology and effects of ambroxol.
      ]. Menthol increases the excitability threshold in peripheral reflexogenic areas, distending peripheral blood vessels and alleviating bronchoconstriction [
      • Mikaili P.
      • Mojaverrostami S.
      • Moloudizargari M.
      • Aghajanshakeri S.
      Pharmacological and therapeutic effects of Mentha Longifolia L. and its main constituent, menthol.
      ]. Finally, anti-inflammatory drugs (non-steroidal anti-inflammatory drugs or cyclooxygenase inhibitors) decrease respiratory airway inflammation by inhibiting the production of chemical mediators such as prostaglandin or cyclooxygenase [
      • Lai H.
      • Rogers D.F.
      New pharmacotherapy for airway mucus hypersecretion in asthma and COPD: targeting intracellular signaling pathways.
      ,
      • Gras D.
      • Chanez P.
      • Vachier I.
      • Petit A.
      • Bourdin A.
      Bronchial epithelium as a target for innovative treatments in asthma.
      ,

      Aemps.gob.es [internet]. Agencia Española de Medicamentos y Productos Sanitarios [cited 2014 Nov 15]. Available from: http://aemps.gob.es/.

      ].
      Growing evidence in the last years suggests that theobromine may be a novel and promising antitussive treatment for patients with either acute or chronic coughing. On the other hand, it can be used as a bronchodilator owing to the fact that the administration of methylxanthines increases the airway diameter [
      • Tilley S.L.
      Methylxanthines in asthma.
      ].
      Some in vivo and in vitro studies performed by Usmani et al., in 2005, suggest that theobromine is able to inhibit cough in a direct way, thus behaving as an antitussive drug. The authors observed that concentrations of theobromine showing no signs of adverse effects inhibit citric acid-induced cough in guinea-pigs. Interestingly, the methylxanthine was able to inhibit capsaicin-induced cough in a cohort of patients undertaken a randomized, double-blind, placebo controlled study. Moreover, experiments in isolated human and guinea-pig vagus nerve preparations demonstrated that theobromine directly inhibits sensory afferent nerve activation [
      • Usmani O.S.
      • Belvisi M.G.
      • Patel H.J.
      • Crispino N.
      • Birrell M.A.
      • Korbonits M.
      • et al.
      Theobromine inhibits sensory nerve activation and cough.
      ]. This is in accordance with the recently described ability of methylxanthines to regulate the activation of human intermediate-conductance Ca2+-activated potassium channels that are key in controlling afferent and efferent vagal activity [
      • Shcroder R.L.
      • Jensen B.S.
      • Strobaek D.
      • Olensen S.P.
      • Christophersen P.
      Activation of the human, intermediate-conductance, Ca2+ activated K+ channel by methylxanthines.
      ,
      • Fox A.J.
      • Barnes P.J.
      • Venkatesan P.
      • Belvisi M.G.
      Activation of large conductance potassium channels inhibits the afferent and efferent function of airway sensory nerves in the guinea pig.
      ]. Interestingly, it has been reported that cacao intake may reduce cough. Taken together this observation and the enhanced bioavailability of methylxanthines in natural sources, dark chocolate consumption appears as an attractive way to administer theobromine to patients with cough [
      • Halfdanarson T.R.
      • Jatoi A.
      Chocolate as a cough suppressant: rationale and justification for an upcoming clinical trial.
      ]. Despite theobromine use is a novel alternative to the controversial opioid treatment, more studies are needed to know the effects of daily intake of this methylxanthine, especially in children in whom an appropriate balance between toxicity risk and therapeutic benefit is critical [
      • Chang A.B.
      • Halstead R.A.
      • Petsky H.L.
      Methylxanthines for prolonged non-specific cough in children.
      ].

      7. Methylxanthine-based therapy in other respiratory diseases: COPD and bronchiectasis

      In view of the demonstrated benefits of methylxanthines in serious respiratory tract pathologies such as AOP, asthma and cough, some laboratories are testing the translation potential of these natural compounds or synthetic analogs in other lung diseases, namely chronic obstructive pulmonary disease (COPD) and bronchiectasis.
      COPD is a disorder caused by airway tract inflammation that may lead to serious difficulties in breathing. The WHO considers it the fifth cause of mortality in the World and predicts that could reach the second place in 2030 [

      Who.int [internet]. World Health Organization. [cited 2014 Nov 10]. Available from: http://www.who.int/whosis/whostat/2008/en/.

      ]. The principal causes of COPD are related with unhealthy habits as smoking and long-term exposures to polluted air, chemical vapors or dust. The heavy air pollution in some populated Chinese cities affects lung function and exacerbates COPD in both adults and children [
      • Hu G.
      • Zhong N.
      • Ran P.
      Air pollution and COPD in China.
      ]. However, COPD is usually diagnosed in adult age. Interestingly, theophylline and the synthetic xanthine derivate doxofylline, have been recommended for the treatment of this disorder with positive results and similar proven efficacy [
      • Akram M.F.
      • Nasiruddin M.
      • Ahmad Z.
      • Ali Khan R.
      Doxofylline and theophylline: a comparative clinical study.
      ].
      Bronchiectasis is an uncommon and irreversible stretching and widening of the bronchial tree that can be congenital or secondary to other pathological processes as tuberculosis, tumors or bacterial (Klebsiella pneuominae, Staphilococcus aureus or Pseudomonas aeuroginosa) and viral (adenovirus) infections [

      nhlbi.nih.gov [internet]. National Institutes of Health. What Is Bronchiectasis?. [cited 2014 Nov 12] Available from:http://www.nhlbi.nih.gov/health/health-topics/topics/brn/.

      ,
      • Coleman L.T.
      • Kramer S.S.
      • Markowitz R.I.
      • Kravitz R.M.
      Bronchiectasis in children.
      ]. Currently, it has no cure and the therapeutic management focuses on relieving symptoms and improving the life quality of patients. The aims of the treatment depend on the initial event, e.g. a specific antibiotic will be prescribed if the cause is a bacterium. However, if the bronchial damage is extensive, oxygen therapy and hydration or respiratory exercises are suggested along with bronchodilators or corticosteroids. Based on the proven benefits in other respiratory diseases and on its anti-inflammatory properties, theophylline is gaining attention for bronchiectasis therapy. The results of clinical trials performed with methylxanthines are not yet conclusive and more placebo controlled clinical studies are needed [

      Steele K, Greenstone M, Lasserson JA. Oral methyl-xanthines for bronchiectasis. Cochrane Database. 1, CD002734.

      ]. To assess the effect of theophylline in bronchiectasis, one clinical trial whose primary outcome is the score of the St. George's respiratory questionnaire is currently recruiting patients [see Ref No NCT01684683 in: https://clinicaltrials.gov].

      8. Conclusions

      AOP, asthma and cough are examples of common respiratory problems that affect preterm infants and children and whose incidence in the past decades has increased dramatically. Acute respiratory events in the early life are likely to be important in the development of chronic lung diseases that affect the physical health of the children, but also their social, emotional, and academic performance. Classical therapies including corticosteroid or opiate administration and inhalation of CO2-containing air, display clinical benefits but suffer from limitations related with adherence, adverse side effects and/or high cost. In this context, methylxanthines arise as natural compounds capable of stimulate CNS and respiratory muscle function. Individually administered or as add-on therapy to other interventions, solid evidence proves the methylxanthine efficacy in a variety of respiratory pediatric diseases. The excellent bioavailability and the experience from using methylxanthine-containing nutrients may benefit a more extended use in children for which pleasant taste and medication adherence are correlated.

      Conflicts of interests

      The authors declare no conflict of interest.

      References

        • Ashihara H.
        • Suzuki T.
        Distribution and biosynthesis of caffeine in plants.
        Front. Biosci. 2004; 1: 1864-1876
        • Kretschmar J.A.
        • Baumann T.
        Caffeine in Citrus flowers.
        Phytochemistry. 1999; 52: 19-23
      1. Eufic.org [internet]. The European Food Information Council (EUFIC) [cited 2014 Nov 14]. Available from: http://www.eufic.org/.

        • Bispo M.S.
        • Veloso M.C.
        • Pinheiro H.L.
        • De Oliveira R.F.
        • Reis J.O.
        • De Andrade J.B.
        Simultaneous determination of caffeine, theobromine, and theophylline by high-performance liquid chromatography.
        J. Chromatogr. Sci. 2002; 40: 45-48
        • Hicks M.B.
        • Hsieh Y.H.P.
        • Bell L.N.
        Tea preparation and its influence on methylxanthine concentration.
        Food Res. Intenat. 1996; 29: 325-330
        • Langer S.
        • Marshall L.J.
        • Day A.J.
        • Morgan M.R.
        Flavanols and methylxanthines in commercially available dark chocolate: a study of the correlation with nonfat cocoa solid.
        J. Agric. Food Chem. 2011; 59: 8435-8441
        • Butcher R.W.
        • Sutherland E.W.
        Adenosine 3′,5′-phosphate in biological materials. I. Purification and properties of cyclic 3′,5′-nucleotide phosphodiesterase and use of this enzyme to characterize adenosine 3′,5′-phosphate in human urine.
        J. Biol. Chem. 1962; 237: 1244-1250
        • Lynge J.
        • Hellsten Y.
        Distribution of adenosine A1, A2A and A2B receptors in human skeletal muscle.
        Acta Physiol. Scand. 2000; 169: 283-290
        • Salvatore C.A.
        • Jacobson M.A.
        • Taylor H.E.
        • Linden J.
        • Johnson R.G.
        Molecular cloning and characterization of the human A3 adenosine receptor.
        Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10365-11069
        • Zhou Q.Y.
        • Li C.
        • Olah M.E.
        • Johnson R.A.
        • Stiles G.L.
        • Civelli O.
        Molecular cloning and characterization of an adenosine receptor: the A3 adenosine receptor.
        Proc. Natl. Acad. Sci. USA. 1992; 89: 7432-7436
        • Gessi S.
        • Cattabriga E.
        • Avitabile A.
        • Gafa R.
        • Lanza G.
        • Cavazzini L.
        • et al.
        Elevated expression of A3 adenosine receptors in human colorectal cancer is reflected in peripheral blood cells.
        Clin. Cancer Res. 2004; 10: 5895-5901
        • Kohno Y.
        • Ji X.
        • Mawhorter S.D.
        • Koshiba M.
        • Jacobson K.A.
        Activation of A3 adenosine receptors on human eosinophils elevates intracellular calcium.
        Blood. 1996; 88: 3569-3574
        • Fossetta J.
        • Jackson J.
        • Deno G.
        • Fan X.
        • Du X.K.
        • Bober L.
        • et al.
        Pharmacological analysis of calcium responses mediated by the human A3 adenosine receptor in monocyte-derived dendritic cells and recombinant cells.
        Mol. Pharmacol. 2003; 63: 342-350
        • Brosen K.
        Drug interactions and the cytochrome P450 system. The role of cytochrome P450 1A2.
        Clin. Pharmacokinet. 1995; 29: 20S-25S
        • Begas E.
        • Kouvaras E.
        • Tsakalof A.
        • Papakosta S.
        • Asprodini E.K.
        In vivo evaluation of CYP1A2, CYP2A6, NAT-2 and xanthine oxidase activities in a Greek population sample by the RP-HPLC monitoring of caffeine metabolic ratios.
        Biomed. Chromatogr. 2007; 21: 190-200
        • Tassaneeyakul W.
        • Birkett D.J.
        • McManus M.E.
        • Tassaneeyakul W.
        • Veronese M.E.
        • Andersson T.
        • et al.
        Caffeine metabolism by human hepatic cytochromes P450: contributions of 1A2, 2E1 and 3A isoforms.
        Biochem. Pharmacol. 1994; 18: 1767-1776
        • Fabes M.S.
        • Fuhr U.
        Time response of cytochrome P450 1A2 activity on cessation of heavy smoking.
        Clin. Pharmacol. Ther. 2004; 76: 178-184
        • Pendleton M.
        • Brown S.
        • Thomas C.
        • Odle B.
        Potential toxicity of caffeine when used as a dietary supplement for weight loss.
        J. Diet. Suppl. 2012; 9: 293-298
        • Pendleton M.
        • Brown S.
        • Thomas C.M.
        • Odle B.
        Potential toxicity of caffeine when used as a dietary supplement for weight loss.
        J. Diet. Suppl. 2013; 10: 1-5
        • Baggott M.J.
        • Childs E.
        • Hart A.B.
        • De Bruin E.
        • Palmer A.A.
        • Wilkinson J.E.
        • et al.
        Psychopharmacology of theobromine in healthy volunteers.
        Psychopharmacol. Berl. 2013; 228: 109-118
      2. National Institutes of Health Consensus Development Conference on Infantile Apnea and Home Monitoring, Sept 29 to Oct 1, 1986. Pediatrics 1987;79: 292–299.

        • Lorch S.A.
        • Srinivasan L.
        • Escoobar G.J.
        Epidemiology of apnea and bradycardia resolution in premature infants.
        Pediatrics. 2011; 128: 366-373
        • Picone S.
        • Bedetta M.
        • Paolillo P.
        Caffeine citrate: when and for how long. A literature review.
        J. Matern. Fetal Neonatal Med. 2012; 25: 11-14
        • Finer N.N.
        • Barrington K.J.
        • Hayes B.J.
        • Hugh A.
        Obstructive, mixed, and central apnea in the neonate: physiologic correlates.
        J. Pediatr. 1992; 121: 943-950
        • Zhao J.
        • Gonzalez F.
        • Mu D.
        Apnea of prematurity: from cause to treatment.
        Eur. J. Pediatr. 2009; 170: 1097-1105
        • Putnam R.W.
        • Conrad S.C.
        • Gdovin M.J.
        • Erlichman J.S.
        • Leiter J.C.
        Neonatal maturation of the hypercapnic ventilatory response and central neural CO2 chemosensitivity.
        Respir. Physiol. Neurobiol. 2005; 15: 165-179
        • Darnall R.A.
        The role of CO2 and central chemoreception in the control of breathing in the fetus and the neonate.
        Respir. Physiol. Neurobiol. 2010; 173: 201-212
        • Carlo W.A.
        • Martin R.J.
        • Difiore J.M.
        Differences in CO2 threshold of respiratory muscles in preterm infants.
        J. Appl. Physiol. 1985; 65: 2434-2439
        • Morin D.
        • Di Pasquale E.
        • Hilaire G.
        • Monteau R.
        Possible involvement of serotonin in obstructive apnea of the newborn.
        Biol. Neonate. 1994; 65: 176-181
        • Khan A.
        • Qurashi M.
        • Kwiatkowski K.
        • Cates D.
        • Rigatto H.
        Measurement of the CO2 apneic threshold in newborn infants: possible relevance for periodic breathing and apnea.
        J. Appl. Physiol. 2005; 98: 1171-1176
        • Heman-Ackah Y.D.
        • Pernell K.J.
        • Goding Jr., G.S.
        The laryngeal chemoreflex: an evaluation of the normoxic response.
        Laryngoscope. 2009; 119: 370-379
        • Thach B.T.
        Reflux associated apnea in infants: evidence for a laryngeal chemoreflex.
        Am. J. Med. 1997; 103: 120S-124S
        • Henderson-Smart D.J.
        • Steer P.A.
        Caffeine versus theophylline for apnea in preterm infants.
        Cochrane Database Syst. Rev. 2010; 20 (CD000273)
        • Bhatia J.
        Current options in the management of apnea prematurity.
        Clin. Pediatr. [Phila]. 2000; 39: 327-336
        • Paolillo P.
        • Picone S.
        Apnea of prematurity.
        J. Neonatal Perinat. Med. 2013; 2: 1-7
        • Valieva O.A.
        • Strandjord T.P.
        • Mayock D.E.
        • Juul S.E.
        Effects of transfusions in extremely low birth weight infants: a retrospective study.
        J. Pediatr. 2009; 155: 331-337
        • Westkamp E.
        • Soditt V.
        • Adrian S.
        • Bohnhorst B.
        • Groneck P.
        • Poets C.F.
        Blood transfusion in anemic infants with apnea of prematurity.
        Bio Neonate. 2002; 82: 228-232
        • Al-Saif S.
        • Alvaro R.
        • Manfreda J.
        • Kwiatkowski K.
        • Cates D.
        • Qurashi M.
        • et al.
        A randomized controlled trial of theophylline versus CO2 inhalation for treating apnea of prematurity.
        J. Pediatr. 2008; 153: 513-518
        • Kuzemko J.A.
        Aminophylline in apnoeic attacks of newborn.
        Lancet. 1973; 30: 1509
        • Kuzemko J.A.
        • Paala J.
        Apnoeic attacks in the newborn treated with aminophylline.
        Arch. Dis. Child. 1973; 48: 404-406
        • Bednarek F.J.
        • Roloff D.W.
        Treatment of apnea of prematurity with aminophylline.
        Pediatrics. 1976; 58: 335-339
        • Shannon D.C.
        • Gotay F.
        • Stein I.M.
        • Rogers M.C.
        • Todres I.D.
        • Moylan F.M.
        Prevention of apnea and bradycardia in low-birthweight infants.
        Pediatrics. 1975; 55: 594
        • Roberts R.
        Drug Therapy in Infants.
        WB Saunders, Philadelphia, United States of America1984: 119-137
        • Kreutzer K.
        • Bassler D.
        Caffeine for apnea of prematurity: a neonatal success story.
        Neonatology. 2014; 105: 332-336
        • Aubier M.
        • Murciano D.
        • Viires N.
        • Lecocguic Y.
        • Pariente R.
        Respiratory muscle pharmacotherapy.
        Bull. Eur. Physiopathol. Respir. 1984; 20: 459-466
        • Van der Heijden H.F.
        • Dekhuijzen P.N.
        • Folgering H.
        • Van Herwaarden C.L.
        Pharmacotherapy of respiratory muscles in chronic obstructive pulmonary disease.
        Respir. Med. 1996; 90: 513-522
        • Aranda J.V.
        • Beharry K.
        • Valencia G.B.
        • Natarajan G.
        • Davis J.
        Caffeine impact on neonatal morbidities.
        J. Matern. Fetal Neonatal Med. 2010; 23: 20-23
        • Aranda J.V.
        • Gorman W.
        • Bergsteinsson H.
        • Gunn T.
        Efficacy of caffeine in treatment of apnea in the low-birth-weight infant.
        J. Pediatr. 1977; 90: 467-472
        • Arora P.
        Pathogenesis and management of apnea of prematurity.
        J. Neonatal Biol. 2012; 1: 1-2
        • Steer P.
        • Flenady V.
        • Shearman A.
        • Charles B.
        • Gray P.H.
        • Henderson-Smart D.
        • et al.
        High dose caffeine citrate for extubation of preterm infants: a randomised controlled trial.
        Arch. Dis. Child. 2004; 89: 499-503
        • Martin R.J.
        • Abu-Shaweesh J.M.
        • Baird T.M.
        Apnoea of prematurity.
        Paediatr. Respir. Rev. 2004; 5: S377-S382
        • Andreas S.
        • Kreuzer H.
        Cheyne-Stokes respiration in patients with congestive heart failure.
        Z Kardiol. 1998; 87: 15-21
        • Koos B.J.
        • Maeda T.
        • Jan C.
        Adenosine A[1] and A[2A] receptors modulate sleep state and breathing in fetal sheep.
        J. Appl. Physiol. 2001; 91: 343-350
        • Watt A.H.
        • Buss D.C.
        • Routledge P.A.
        Effect of aminophylline on the respiratory depressant action of intravenous adenosine in neonatal rabbits.
        Life Sci. 1987; 5: 29-34
        • Abu-Shaweesh J.M.
        Activation of central adenosine A[2A] receptors enhances superior laryngeal nerve stimulation-induced apnea in piglets via a GABAergic pathway.
        J. Appl. Physiol. 2007; 103: 1205-1211
        • Gannon B.A.
        Theophylline or caffeine: which is best for apnea of prematurity?.
        Neonatal Netw. 2000; 19: 33-36
        • Aranda J.V.
        • Turmen T.
        Methylxanthines in apnea of prematurity.
        Clin. Perinatol. 1979; 6: 87-108
      3. nhlbi.nih.gov [internet]. National Institutes of Health. What Is Asthma? [Cited 2014 Nov 12]. Available from:http://www.nhlbi.nih.gov/health/health-topics/topics%20/asthma/.

      4. who.int [internet]. World Health Organization. Asthma: Definition. [Cited 2014 Nov 17]. Available from: http://www.who.int/respiratory/asthma/definition.

        • Walker M.L.
        • Holt K.E.
        • Anderson G.P.
        • Teo S.M.
        • Sly P.D.
        • Holt P.G.
        • et al.
        Elucidation of pathways driving asthma pathogenesis: Development of a systems-level analytic strategy.
        Front. Immunol. 2014; 5https://doi.org/10.3389/fimmu.2014.00447
        • Holgate S.T.
        Pathogenesis of Asthma.
        Clin Exp Allergy. 2008; 38: 872-897
        • Beasley R.
        • Pekkanen J.
        • Pearce N.
        Has the role of atopy in the development of asthma been over-emphasized?.
        Pediatr. Pulmonol. 2001; 23: 149-150
        • Strachan D.P.
        Family size, infection and atopy: the first decade of the «hygiene hypothesis».
        Thorax. 2000; 55: S2-S10
        • Wong J.J.
        • Lee J.H.
        • Turner D.A.
        • Rehder K.J.
        A review of the use of adjunctive therapies in severe acute asthma exacerbation in critically ill children.
        Expert Rev. Respir. Med. 2014; 8: 423-441
        • Chung K.F.
        • Wenzel S.E.
        • Brozek J.L.
        • Bush A.
        • Castro M.
        • Sterk P.J.
        • et al.
        International ERS/ATS guidelines on definition, evaluation and treatment of severe asthma.
        Eur. Respir. J. 2014; 43: 343-373
        • Guilbert T.W.
        • Morgan W.J.
        • Zeiger R.S.
        • Mauger D.T.
        • Boehmer S.J.
        • Szefler S.J.
        • et al.
        Long-term inhaled corticosteroids in preschool children at high risk for asthma.
        N. Engl. J. Med. 1985-1997; 200: 354
        • Warner S.M.
        • Knight D.A.
        Airway modeling and remodeling in the pathogenesis of asthma.
        Curr. Opin. Allergy Clin. Immunol. 2008; 8: 44-48
        • Saglani S.
        • Lui S.
        • Ullmann N.
        • Campbell G.A.
        • Sherburn R.T.
        • Mathie S.A.
        • et al.
        IL-33 promotes airway remodeling in pediatric patients with severe steroid-resistant asthma.
        J. Allergy Clin. Immuno. 2013; 132: 676-685
        • Partridge M.R.
        • van der Molen T.
        • Myrseth S.E.
        • Busse W.W.
        Attitudes and actions of asthma patients on regular maintenance therapy: the INSPIRE study.
        BMC Pulm. Med. 2006; 6: 13
        • Bosley C.M.
        • Parry D.T.
        • Cochrane G.M.
        Patient compliance with inhaled medication: does combining beta-agonists with corticosteroids improve compliance?.
        Eur. Respir. J. 1994; 7: 504-509
        • Lipworth B.J.
        Systemic adverse effects of inhaled corticosteroid therapy: A systematic review and meta-analysis.
        Arch. Intern Med. 1999; 159: 941-955
        • Nelson H.S.
        • Weiss S.T.
        • Bleecker E.R.
        • Yancey S.W.
        • Dorinsky P.M.
        SMART Study Group. The Salmeterol Multicenter Asthma Research Trial: a comparison of usual pharmacotherapy for asthma or usual pharmacotherapy plus salmeterol.
        Chest. 2006; 129: 15-26
        • Bara A.I.
        • Barley E.A.
        Caffeine for asthma.
        Cochrane Database Syst. Rev. 2001; 4 (CD001112)
        • Simons F.E.
        • Becker A.B.
        • Simons K.J.
        • Gillespie C.A.
        The bronchodilator effect and pharmacokinetics of theobromine in young patients with asthma.
        J. Allergy Clin. Immunol. 1985; 76: 703-707
        • Barnes P.J.
        Pathophysiology of asthma.
        Br. J. Clin. Pharmacol. 1996; 42: 3-10
        • Pauwels R.
        The effects of theophylline on airway inflammation.
        Chest. 1987; 92: 32S-37S
        • Rabe K.F.
        • Magnussen H.
        • Dent G.
        Theophylline and selective PDE inhibitors as bronchodilators and smooth muscle relaxants.
        Eur. Respir. J. 1995; 8: 637-642
        • Evans D.J.
        • Taylor D.A.
        • Zetterstrom O.
        • Chung K.F.
        • O'Connor B.J.
        • Barnes P.J.
        A comparison of low-dose inhaled budesonide plus theophylline and high-dose inhaled budesonide for moderate asthma.
        N. Engl. J. Med. 1997; 337: 1412-1418
        • Ukena D.
        • Harnest U.
        • Sakalauskas R.
        • Magyar P.
        • Vetter N.
        • Steffen H.
        • et al.
        Comparison of addition of theophylline to inhaled steroid with doubling of the dose of inhaled steroid in asthma.
        Eur. Respir. J. 1997; 10: 2754-2760
        • Barnes P.J.
        Targeting the epigenome in the treatment of asthma and chronic obstructive pulmonary disease.
        Proc. Am. Thorac. Soc. 2009; 6: 693-696
        • Polosa R.
        • Blackburn M.R.
        Adenosine receptors as targets for therapeutic intervention in asthma and chronic obstructive pulmonary disease.
        Trends Pharmacol. Sci. 2009; 30: 528-535
        • Mascali J.J.
        • Cvietusa P.
        • Negri J.
        • Borish L.
        Anti-inflammatory effects of theophylline: modulation of cytokine production.
        Ann. Allergy Asthma Immunol. 1996; 77: 34-38
        • Ichiyama T.
        • Hasegawa S.
        • Matsubara T.
        • Hayashi T.
        • Furukawa S.
        Theophylline inhibits NF-kappa B activation and I kappa B alpha degradation in human pulmonary epithelial cells.
        Naunyn Schmiedeb. Arch. Pharmacol. 2001; 364: 558-561
        • Shukla D.
        • Chakraborty S.
        • Singh S.
        • Mishra B.
        Doxofylline: a promising methylxanthine derivative for the treatment of asthma and chronic obstructive.
        Expert Opin. Pharmacother. 2009; 10: 2343-2356
        • Franzone J.S.
        • Cirillo R.
        • Barone D.
        Doxofylline and theophylline are xanthines with partly different mechanisms of action in animals.
        Drugs Exp. Clin. Res. 1988; 14: 479-489
        • Cirillo R.
        • Barone D.
        • Franzone J.S.
        Doxofylline, an antiasthmatic drug lacking affinity for adenosine receptors.
        Arch. Int. Pharmacodyn. Ther. 1998; 295: 221-237
        • Sankar J.
        • Lodha R.
        • Kabra S.K.S.
        Doxofylline: The next generation methylxanthine.
        India J. Pediatr. 2008; 75: 251-254
        • Goldstein M.F.
        • Chervinsky P.
        Efficacy and safety of doxofylline compared to theophylline in chronic reversible asthma–a double-blind randomized placebo-controlled multicentre clinical trial.
        Med. Sci. Monit. 2002; 8: CR297-304
        • Dini F.L.
        • Cogo R.
        Doxofylline: a new generation xanthine bronchodilator devoid of major cardiovascular adverse effects.
        Curr. Med. Res. Opin. 2001; 16: 258-268
        • Mazzone S.B.
        An overview of the sensory receptors regulating cough.
        Cough. 2005; 4: 1-2
      5. nhlbi.nih.gov [internet]. National Institutes of Health. What Is Cough? [Cited 2014 Oct 30]. Available from:http://www.nhlbi.nih.gov/health/health-topics/topics/%20cough/.

      6. Nhs.uk [internet]. Natioanl Health System. Cough–Causes - NHS Choices. [Cited 2014 Oct 30]. Available from:http://www.nhs.uk/Conditions/Cough/Pages/%20Causes.aspx.

        • Chung K.F.
        • Widdicombe J.G.
        • Boushey H.A.
        Cough: Causes, Mechanisms and Therapy.
        Blackwell Publishing, Massachusetts, Unites States of America2003
        • Polverino M.
        • Polverino F.
        • Fasolino M.
        • Andò F.
        • Alfieri A.
        • De Blasio F.
        Anatomy and neuro-pathophysiology of the cough reflex arc.
        Multidiscip. Respir. Med. 2012; 7https://doi.org/10.1186/2049-6958-7-5
        • Karlsson J.A.
        • Sant'Ambrogio G.
        • Widdicombe J.
        Afferent neural pathways in cough and reflex bronchoconstriction.
        J. Appl. Physiol. 1988; 65: 1007-1023
        • Dinh Q.T.
        • Heck S.
        • Le D.D.
        • Bals R.
        • Welte T.
        Pathophysiology, diagnostics and therapy of chronic cough: neuronal reflexes and antitussiva.
        Pneumologie. 2013; 67: 327-334
        • Macciò A.
        • Madeddu C.
        • Panzone F.
        • Mantovani G.
        Carbocysteine: clinical experience and new perspectives in the treatment of chronic inflammatory diseases.
        Expert Opin. Pharmacother. 2009; 10: 693-703
        • Wunderer H.
        • Morgenroth K.
        • Weis G.
        The cleaning system of the airways: physiology, pathophysiology and effects of ambroxol.
        Med. Monatsschr Pharm. 2009; 32: 42-47
        • Mikaili P.
        • Mojaverrostami S.
        • Moloudizargari M.
        • Aghajanshakeri S.
        Pharmacological and therapeutic effects of Mentha Longifolia L. and its main constituent, menthol.
        Anc. Sci. Life. 2013; 33: 131-138
        • Lai H.
        • Rogers D.F.
        New pharmacotherapy for airway mucus hypersecretion in asthma and COPD: targeting intracellular signaling pathways.
        J. Aerosol Med. Pulm. Drug Deliv. 2010; 23: 219-231
        • Gras D.
        • Chanez P.
        • Vachier I.
        • Petit A.
        • Bourdin A.
        Bronchial epithelium as a target for innovative treatments in asthma.
        Pharmacol. Ther. 2013; 140: 290-305
      7. Aemps.gob.es [internet]. Agencia Española de Medicamentos y Productos Sanitarios [cited 2014 Nov 15]. Available from: http://aemps.gob.es/.

        • Tilley S.L.
        Methylxanthines in asthma.
        Handb. Exp. Pharmacol. 2011; 200: 439-456
        • Usmani O.S.
        • Belvisi M.G.
        • Patel H.J.
        • Crispino N.
        • Birrell M.A.
        • Korbonits M.
        • et al.
        Theobromine inhibits sensory nerve activation and cough.
        FASEB J. 2005; 19: 231-233
        • Shcroder R.L.
        • Jensen B.S.
        • Strobaek D.
        • Olensen S.P.
        • Christophersen P.
        Activation of the human, intermediate-conductance, Ca2+ activated K+ channel by methylxanthines.
        Pflugers Arch. 2000; 440: 809-818
        • Fox A.J.
        • Barnes P.J.
        • Venkatesan P.
        • Belvisi M.G.
        Activation of large conductance potassium channels inhibits the afferent and efferent function of airway sensory nerves in the guinea pig.
        J. Clin. Invest. 1997; 99: 513-519
        • Halfdanarson T.R.
        • Jatoi A.
        Chocolate as a cough suppressant: rationale and justification for an upcoming clinical trial.
        Support Cancer Ther. 2007; 4: 119-122
        • Chang A.B.
        • Halstead R.A.
        • Petsky H.L.
        Methylxanthines for prolonged non-specific cough in children.
        Cochrane Database. 2005; 20 (CD005310)
      8. Who.int [internet]. World Health Organization. [cited 2014 Nov 10]. Available from: http://www.who.int/whosis/whostat/2008/en/.

        • Hu G.
        • Zhong N.
        • Ran P.
        Air pollution and COPD in China.
        J. Thorac. Dis. 2015 Jan; 7: 59-66
        • Akram M.F.
        • Nasiruddin M.
        • Ahmad Z.
        • Ali Khan R.
        Doxofylline and theophylline: a comparative clinical study.
        J. Clin. Diagn Res. 2012; 6: 1681-1684
      9. nhlbi.nih.gov [internet]. National Institutes of Health. What Is Bronchiectasis?. [cited 2014 Nov 12] Available from:http://www.nhlbi.nih.gov/health/health-topics/topics/brn/.

        • Coleman L.T.
        • Kramer S.S.
        • Markowitz R.I.
        • Kravitz R.M.
        Bronchiectasis in children.
        J. Thorac. Imaging. 1995; 10 (Winter): 268-279
      10. Steele K, Greenstone M, Lasserson JA. Oral methyl-xanthines for bronchiectasis. Cochrane Database. 1, CD002734.