Effects of Rolipram (a New Selective Phosphodiesterase 4

2. Effects of Rolipram (a New Selective Phosphodiesterase 4 Inhibitor) versus Theophylline (a Non-selective Phosphodiesterase Inhibitor) on Bronchial Smooth Muscles and Heart Rate in Guinea Pigs By Hussam A. S. Murad and Khaled A. Mahmoud Department of Pharmacology and Therapeutics Faculty of Medicine , Ain Shams University

3. Introduction Bronchial asthma is characterized by reversible airway obstruction, inflammation and hyperreactivity. Phosphodiesterases inhibitors (PDEIs) increase c-AMP levels in lung causing airway smooth muscle relaxation and inhibition of airway remodeling in asthma . Mammals express multiple isoforms of PDEs.The PDE 4 isoform is primarily expressed in the lung. Theophylline is a non-selective PDEI in airway and other tissues producing bronchodilatation but also nausea, headache, diuresis , cardiovascular and other adverse effects. Rolipram (a selective PDE 4 Inhibitor) has less adverse effects compared to non-selective PDEIs in addition to effective inhibition of eosinophil recruitment in tissues which is of great therapeutic value in treatment of asthma. Aim of the Study The research question : Which produces more bronchodilatation with less chronotropic effect , rolipram or theophylline ? Thus the present study was conducted to compare effects of rolipram versus theophylline on bronchial smooth muscle and heart rate in guinea pigs

4. Methods I. In vitro experiment : Isolated perfused guinea pig tracheal spiral strip: • 2 groups (6 /group)were used. The tracheal spiral strip was suspended in the organ bath. The submaximal dose of histamine was determined as 4ug/ml bath. DMSO (0.1%) followed by graded doses of rolipram (40-320 nM ) or theophylline (3000-24000 nM) were tested on the submaximal dose-induced contractions in the 1st or 2nd groups respectively . • II. In vivo experiments : • A) Determination of airway resistance in anasthetized ovalbumin-sensitized guinea pigs: • 2 groups (6 /group) were used . Each animal was sensitized with ovalbumin . After 4 weeks, the animal was anaesthetized by urethane . Determination of airway resistance was measured using Starling pump. The volume of expired air was measured by an air-tight rubber tube attached to the tracheostomy tube at one end, the other end was connected to Harvard pressure module 275 which was connected to a pen to record changes in airflow.

5. Bronchoconstriction was produced by I.P injection of histamine (300 ug/kg ) and airway resistance was measured for 15 min after injection of histamine as the change from the basal level. DMSO (0.1%) was given I.P. at maximal histamine-induced bronchoconstriction . After 5 min. rolipram (7.7 mg/kg = 0.28 mmol/ kg) or theophylline (67 mg/kg = 3.72 mmol/ kg) was given by I.P. injection to the first and second group respectively . B) Determination of ECG changes in anasthetized guinea pigs: 2 groups (6 /group) were used. The animal was anaesthetized by urethane . and its four limbs were connected to electrodes of Nihon Kohden Cardiofax by needles. The ECG apparatus was switched on the standard lead II and run at a paper speed of 25 mm/ second 1). Normal record was taken as a control then rolipram (7.7 mg/kg = 0.28 mmol/ kg) or theophylline (67 mg/kg = 3.72 mmol/ kg) was given I.P. to first and second group respectively.

6. Results I- Effects of rolipram and theophylline on isolated guinea pig tracheal spiral strip: Rolipram (40 , 80, 160 & 320 nM) produced more significant and more potent (less EC50) concentration-dependent relaxations of histamine-induced contractions than theophylline (3000 , 6000 , 12000 & 24000 nM) Rolipram Theophylline

8. II- Effects of rolipram and theophylline on airway resistance in anasthetized ovalbumin- sensitized guinea pigs: Rolipram (7.7 mg/kg = 0.28 mmol/ kg) produced more significant and more potent reductions than theophylline (67 mg/kg = 3.72 mmol/ kg) in histamine-induced airway resistance.

10. III- Effects of rolipram and theophylline on ECG in anasthetized guinea pigs: Rolipram (7.7 mg/kg = 0.28 mmol/ kg) produced non-significant changes in contrast to theophylline (67 mg/kg = 3.72 mmol/ kg) which produced a significant increase in heart rate.

12. Discussion Rolipram inhibits antigen-induced bronchoconstriction in anasthetized ovalbumin-sensitized guinea pigs more potently than theophylline (on molecular basis, rolipram has a smaller dose). Rolipram also relaxes isolated guinea pig tracheal spiral more potently than theophylline (rolipram has less EC50). PDE4 enzyme provide the dominant PDE activity in lung , monocytes and macrophages suggesting that specific PDE4 inhibitors may allow for selective effects on these inflammatory cells, in addition to bronchodilatation. Rolipram produces direct bronchodilatation and inhibition of function of leucocytes as release of LTC4/D4/E4 from eosinophils. Also rolipram modulates integrin-induced actin assembly at cell periphery impairing cell locomotion and migration through inhibition of PDE4, activation of PKA, increase of K efflux and reduction of Ca influx.

13. Consequently, rolipram inhibits antigen-induced bronchoconstriction, airway plasma leakage (oedema), eosiniphil infiltration and hyperreactivity. In contrast theophylline inhibits only antigen-induced bronchoconstriction and airway plasma leakage (oedema) but neither airway eosiniphil infiltration nor hyperreactivity. In addition, the IC50 values for effect of rolipram and theophylline on PDE4 activity of bronchial tissues are 2 uM & > 100 uM respectively indicating more potency of rolipram. Theophylline, in addition to being a non-selective PDEI, it blocks adenosine 1 receptors causing not only a beneficial bronchodilatation but also harmful especially cardiac adverse effects Conclusion Based on selective PDE4 inhibition, more potency , more inhibition of inflammatory cell functions and less cardiac side effects in bronchodilator doses in guinea pigs, rolipram, might have a more beneficial therapeutic value than the non-selective PDE inhibitor, theophylline, in treatment of asthmatic patients especially those having cardiac problems.

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