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Effects of PGA and PGB compounds on human respiratory tract smooth muscle in vitro
PROSTAGLANDINS
EFFECTS OF PGA AND PGB COMPOUNDSON HUMAN RESPIRATORY TRACT SMOOTH MUSCLE -IN VITRO
P.Y. Lo,
P.G. Adaikan and S.M.M.
Karhn
Department of Obstetrics and Gynaecology, Kandang Kerbau Hospital, University of Singapore, Singapore 8.
INTRODUCTION Prostaglandins E2 and Fw are present in human pulmonary tissues (1,2). Several studies on the pharmacological effects of these two prostaglandins on human respiratory tract have been reported. Both -in vivo and -in vitro PGFa compound produce constriction and PGE compounds usually produce dilatation of the bronchial muscle (see 3 for references). PGA (4) and PGB(5,6) compounds also occur naturally in mammalian tissues, although their presence in pulmonary tissue has not been reported. Pharmacological studies with these prostaglandins have so far been restricted mainly to smooth muscles of the respiratory tract of laboratory animals. On the guinea pig tracheal chain preparation prostaglandin Al inhibited the contractile effect of acetylcholine but was 30 times less active than prostaglandin E2 (7). In anaesthetised dogs and cats, intravenously administered PGAl and PGA2 increased airway resistance and reduced lung compliance indicating bronchoconstriction of both small and large airways (8,9). Prostaglandin B2 produced a dose dependent increase in airway resistance in anaesthetised dogs (10). On cat tracheal muscle in vitro, prostaglandins BI and B2 inhibited acetylcholine and carbachol induced contraction but were 50 times less active than PGE2 (see 3 for references). In this paper we report the effects of PGAI, PGA2, PGBI and PGB2on human respiratory tract smooth muscle in vitro.
3-30-76
MARCH
1976
VOL. 11 NO. 3
531
PROSTAGLANDINS
MATERIALS AND METHODS Macroscopically normal human trachea and lungs were obtained at post-mortem within 6 hours of death. Circular muscle strips were prepared from three anatomical regions. (1) Tracheal smooth muscle strrps were cut transversely from the posterior region of the trachea which is devoid of cartilage rings. (2) Bronchial tubes with plates of cartilage were dissected out from surrounding tissues and cut spirally to obtaLncircular muscle strips 2-3 cm long. (3) Bronchiole tubes from the distal part of the lung were also cut into 2-3 cm long spiral strips. The strips were mounted in 25 ml organ baths containing KrebsHenseleit solution of the following composition (g/i) sodium chloride, 6.74; potassium chloride, 0.35; calcium chloride (2H20), 0.26; potassium dihydrogen orthophosphate, 0.16; magnesium sulphate (7H20), 0.287; glucose, 1.54; sodium bicarbonate, 1.86; disodium EDTA, 0.01. The bathing fluid was maintained at 37’C and aerated with 95% oxygen and 5% carbon dioxide. The pH of the bathing solution was 7.4. An initial ten&on of 1.5 g was applied to tracheal strips and 0.5 g to bronchial and bronchiolar strips. The tissues were allowed to equilibrate for 45 minutes before addition of drugs. The fourpoint method was used to compare the potencies of PGAI, A2, BI and B2. Prostaglandin Pw was used as a reference standard. In different strips, the drug contact time with the tissue varied between 5-12 minutes and dose cycle time 20-45 minutes.
RESULTS Prostaglandtns Al, A2, BI and B2 produced contractions of circular muscle strips from the trachea, bronchial and bronchiole tubes. Prostaglandin B2 was more potent than PGFa, PGA!, A2 and BI on all three preparations (Table I, Fig. I). The difference in potencies was more marked on the circular muscle from the bronchiole tube where PGB2was about three times more active than PGF20,, Detailed results are given in Table I. Usually one or two prostaglandins were tested on one tissue and no tachyphylaxis was encountered. Because of the long dose cycle time,determination of full dose response curves with individual prostaglandtns was impracticable. However, two or three point dose response curves of PGA and PGBcompounds were performed and found to be parallel to those of FGFW
532
MARCH
1976
VOL. 11 NO. 3
PROSTAGLANDINS
TABLE I Human Respiratory Smooth Muscle Stimulating Potencies of FGBl, PCB2, PCAl and PCA2, relative to PCFZ~~taken as = I
mm WY Prostaglandin
BI B2 Al A2
Mean (2 SE)
0.34 @I. 15) 1.60 (9.45) 0.078(-H). 02) 0.66 (3.51)
RW.F 0.038 0.31 0.015 0.060
-
0.84 3.30 0.19 2.20
Threshold dose(rauge) ug/ml 100 - 1000 20 - 400 50 - 1000 40- 800
Human bronchial tube
FOTENCY Prostaglandiu
B1 B2 Al A2
Threshold dose (range) ug/ml
Tiz&qxT 1.40 1.40 (to. 07) 0.096 1.50 (S. 47)
0.76 1.20 0.063 0.65
-
2.0 1.70 0.13 3.60
20075400 40 -
500 400 800 500
Human bronchiole tube POTENCY Prostaglandin
Bl B2 Al A2
MARCH 1976
Mean (2 SE)
1.00 3.00 0.27 0.53
(+o. 20) (S.77) (5.03) (_to. 31)
Range
0.57 0.78 0.19 0. I6
VOL. 11 NO. 3
-1.90 - 4.80 - 0.35 - 1.50
TImeshold dose (range) ng/ml loo500 50 - 100 400 200
No. of experiments
6 5 4 4
533
PROSTAGLANDINS Bronchial
tuba PtFu
Brmchide
l
we.
In vitro effects of prostaglandins Fw, BI and B2 on circular muscles of human respiratory tract
DISCUSSION In animal studies both bronchodilatatlon and bronchoconstrictlon have been reported with PGA and PGBcompounds in vivo (see Introduction). On cat tracheal muscle in vitro FGBcompounds inhibited acetylcholine and carbachol induced contractions, indicating bronchodilatation, whereas on guinea pig tracheal chain preparation PGBcompounds showed constrictor effect which was more potent than the stimulant actlon of PGF% (11). In the present study, however, PGA1, FGA2, FGBl and pGB2 produced contracttons of circular muscle strips from the trachea-bronchial smooth muscle of man. In partkular, FGB2was shown to be more potent than prostaglandins Al, A2, BI and FW Prostaglandins of the A and B series have previously been showd to be more potent than IGE2 and PGFa in their stimulant effect on other human smooth muscles such as human umbilical cord (12) and human penile muscle (13). Unlike PGFwprostaglandins of the A and B series are not metabolised by prostaglandin 15 hydroxy dehydrogenase, although metabolism by other exzymes may occur (7,14). It is therefore likely that in in vivo situation the bronchoconstrictor effects of PGA and PGBcompounds may be more pronounced. The unnatural Isomer of PGA2 (15”epi-FGA2) produces bronchoconstrictor effect in man (15) and acute bronchoconstrlction haa been encountered in women, when PGA2 or PGB2are used to stimulate second trimester pregnant uterus (16).
534
MARCH
1976
VOL. 11 NO. 3
PROSTAGLANDINS
Prostaglandin Fa produces bronchoconstriction in man (3). This prostaglsndin also has a constrictor effect on the circular strips of human bronchial muscle in vitro (17). A correlation therefore exists between in vitro and in vtvo findings for PGF= PGA2 and PGB2. Similarly, both bzn-chodilator and bronchoconstrictor effects reported in in vivo situations with PGE2 (18, 19) have been confirmed on human bronchial muscle (17) and tracheo-bronchial muscles (20) in vitro.
ACKNOWLEDGEMENTS Kalamazoo, Prostaglandins were supplied by the Upjohn Co., Michigan. The work was carried out under USAID Grant Contract No. AID/ cm/pha-c-73/36 and a research grant from The Wellcome Trust, U. K. We are grateful to Dr. Chao Tzee Cheng, Department of Pathology, Singapore General Hospital for post-mortem specimens.
REFERENCES 1.
ingg&d, E. (1965). The isolation and determination of prostaglandins in the lungs of sheep, guinea pig, monkey and man. Biochem. Pharmacol. 1507. -’14
2.
Karim, S. M. M., Sandler, M. and Williams, E.D. (1967). Distribution of prostaglandins in human tissues. Br. J. Pharmac. 31, 340-344.
3.
Smith, A. P. (1976). Prostaglandins and the respiratory system. In : Advances in Prostaglandiu Research : Physiological, Pharmacological and Pathological Aspects. P. 83-102 (S. M. M. Karim, editor), Medical and Technical Publications, Lancaster.
4.
Hamberg, M. and Samuelsson, B. (1967). New groups of naturally occurring prostaglandins. In : Prostaglandins, Nobel Symposium II (eds. S. Bergstrom and B. Samuelsson), Stockholm, Almqvtst and Wiksell, New York, Interscience, P. 63-70.
5.
Eisenbarth, G. S. and Lebovitz, H. E. (1974). Prostaglandin inhibition of cartilage chondromucoprotein synthesis : concept of ‘Intrinsic Activity “. Prostaglandtns z, 11-20.
6.
Wong, P. Y. and Wuthier, R. E. (1974). Isolation and identification of prostaglandin PGBl in growth cartilage. Prostaglandins, g, 125-132.
7.
Horton, E. W. and Jones, R. L. (1969). Prostaglandins AI, A2 and 19hydroxy AI; their act ions of smooth muscle and their inactivation on
MARCH
1976
VOL. 11 NO. 3
535
PROSTAGLANDINS
passage through the pulmonary and hepatic portal vascular beds. Br. J. Pharmacol. -37, 705-722. 8.
Ballantyne, A., Jones, R.L. and Ungar, A. (1973). The actions of prostaglandins Al and A2 on airway resistance and compliance in the cat. Br. J. Pharmacol. -47, 63OP-631P.
9.
Hirose, R. and Said,S.I. (1971). Respiratory effects of prostaglandins Al, El and F20. Clin. Res. 2, 512.
10. Greenberg, S., Howard, L. and Wilson, W. R. (1973). Comparative effects of prostaglandins A2 and B2 on vascular and airway resistances and adrenergic neurotransmission. Can. J. Physiol. Pharmacol. 2, 699-705. 11. Lo, P. Y. and Karim, S. M. M. (1976). Effects of A and B prostaglandins on guinea pig tracheal chain. In Preparation. 12. Adaikan, P.G. and Karim, S. M.M. (1974). Effects of Al, AZ, Bl, B2, E2 and Fa on human umbilical cord vessels. Prostaglandins 8, 411-416. 13. Karim, S. M. M. and Adaikan, P. G. (1975). In : Advances in Prostaglandin Research : Prostaglandins and Reproduction, P. 26-28 (S. M. M. Karim, eidtor), Medical and Technical Publications, Lancaster. 14. Attallah, A. A., prostaglandin A. tion. Life Sci.,
Payskkapan, W. and Lee, J.B. (1974). Metabolism of 1. The kidney cortex as a major site of FGA2 degrada-14, 1521-1534.
15. Smith, A. P. (1974). Effects of three prostaglandin analogues on airway tone in asthmatics and normal subjects. Int. Res. Commun. System, 2, 1457. 16. Karim, S. M. M. (1976). Effect of prostaglandin B2 on the pregnant human uterus. In Preparation. 17. Gardiner, P. J. (1975). The effects of some natural prostaglandins on isolated human circular bronchial muscle. Prostaglandins, s, 607-616. 18. Cuthbert, M. F. (1971). Bronchodilator activity of aerosols of prostaglandins El and E2 in asthmatic subjects. Proc. R. Sot. Med. -64, 15-16. 19. Mathe, A.A., Hedqvist, P., Holmgren, A. and Svanborg, N. (1973). Bronchial hyperractivity of prostaglandin I?- and histamine in patients with asthma. Br. Med. J. I, 193-196. 20. Lo, P.Y., Adaikan, P. G. and Karim, S.M. M. (1976).The effect of PGE2 and its metabolites on human airway smooth muscle in vitro. In Preparation. 536
MARCH
1976
VOL. 11 NO. 3
EFFECTS OF PGA AND PGB COMPOUNDSON HUMAN RESPIRATORY TRACT SMOOTH MUSCLE -IN VITRO
P.Y. Lo,
P.G. Adaikan and S.M.M.
Karhn
Department of Obstetrics and Gynaecology, Kandang Kerbau Hospital, University of Singapore, Singapore 8.
INTRODUCTION Prostaglandins E2 and Fw are present in human pulmonary tissues (1,2). Several studies on the pharmacological effects of these two prostaglandins on human respiratory tract have been reported. Both -in vivo and -in vitro PGFa compound produce constriction and PGE compounds usually produce dilatation of the bronchial muscle (see 3 for references). PGA (4) and PGB(5,6) compounds also occur naturally in mammalian tissues, although their presence in pulmonary tissue has not been reported. Pharmacological studies with these prostaglandins have so far been restricted mainly to smooth muscles of the respiratory tract of laboratory animals. On the guinea pig tracheal chain preparation prostaglandin Al inhibited the contractile effect of acetylcholine but was 30 times less active than prostaglandin E2 (7). In anaesthetised dogs and cats, intravenously administered PGAl and PGA2 increased airway resistance and reduced lung compliance indicating bronchoconstriction of both small and large airways (8,9). Prostaglandin B2 produced a dose dependent increase in airway resistance in anaesthetised dogs (10). On cat tracheal muscle in vitro, prostaglandins BI and B2 inhibited acetylcholine and carbachol induced contraction but were 50 times less active than PGE2 (see 3 for references). In this paper we report the effects of PGAI, PGA2, PGBI and PGB2on human respiratory tract smooth muscle in vitro.
3-30-76
MARCH
1976
VOL. 11 NO. 3
531
PROSTAGLANDINS
MATERIALS AND METHODS Macroscopically normal human trachea and lungs were obtained at post-mortem within 6 hours of death. Circular muscle strips were prepared from three anatomical regions. (1) Tracheal smooth muscle strrps were cut transversely from the posterior region of the trachea which is devoid of cartilage rings. (2) Bronchial tubes with plates of cartilage were dissected out from surrounding tissues and cut spirally to obtaLncircular muscle strips 2-3 cm long. (3) Bronchiole tubes from the distal part of the lung were also cut into 2-3 cm long spiral strips. The strips were mounted in 25 ml organ baths containing KrebsHenseleit solution of the following composition (g/i) sodium chloride, 6.74; potassium chloride, 0.35; calcium chloride (2H20), 0.26; potassium dihydrogen orthophosphate, 0.16; magnesium sulphate (7H20), 0.287; glucose, 1.54; sodium bicarbonate, 1.86; disodium EDTA, 0.01. The bathing fluid was maintained at 37’C and aerated with 95% oxygen and 5% carbon dioxide. The pH of the bathing solution was 7.4. An initial ten&on of 1.5 g was applied to tracheal strips and 0.5 g to bronchial and bronchiolar strips. The tissues were allowed to equilibrate for 45 minutes before addition of drugs. The fourpoint method was used to compare the potencies of PGAI, A2, BI and B2. Prostaglandin Pw was used as a reference standard. In different strips, the drug contact time with the tissue varied between 5-12 minutes and dose cycle time 20-45 minutes.
RESULTS Prostaglandtns Al, A2, BI and B2 produced contractions of circular muscle strips from the trachea, bronchial and bronchiole tubes. Prostaglandin B2 was more potent than PGFa, PGA!, A2 and BI on all three preparations (Table I, Fig. I). The difference in potencies was more marked on the circular muscle from the bronchiole tube where PGB2was about three times more active than PGF20,, Detailed results are given in Table I. Usually one or two prostaglandins were tested on one tissue and no tachyphylaxis was encountered. Because of the long dose cycle time,determination of full dose response curves with individual prostaglandtns was impracticable. However, two or three point dose response curves of PGA and PGBcompounds were performed and found to be parallel to those of FGFW
532
MARCH
1976
VOL. 11 NO. 3
PROSTAGLANDINS
TABLE I Human Respiratory Smooth Muscle Stimulating Potencies of FGBl, PCB2, PCAl and PCA2, relative to PCFZ~~taken as = I
mm WY Prostaglandin
BI B2 Al A2
Mean (2 SE)
0.34 @I. 15) 1.60 (9.45) 0.078(-H). 02) 0.66 (3.51)
RW.F 0.038 0.31 0.015 0.060
-
0.84 3.30 0.19 2.20
Threshold dose(rauge) ug/ml 100 - 1000 20 - 400 50 - 1000 40- 800
Human bronchial tube
FOTENCY Prostaglandiu
B1 B2 Al A2
Threshold dose (range) ug/ml
Tiz&qxT 1.40 1.40 (to. 07) 0.096 1.50 (S. 47)
0.76 1.20 0.063 0.65
-
2.0 1.70 0.13 3.60
20075400 40 -
500 400 800 500
Human bronchiole tube POTENCY Prostaglandin
Bl B2 Al A2
MARCH 1976
Mean (2 SE)
1.00 3.00 0.27 0.53
(+o. 20) (S.77) (5.03) (_to. 31)
Range
0.57 0.78 0.19 0. I6
VOL. 11 NO. 3
-1.90 - 4.80 - 0.35 - 1.50
TImeshold dose (range) ng/ml loo500 50 - 100 400 200
No. of experiments
6 5 4 4
533
PROSTAGLANDINS Bronchial
tuba PtFu
Brmchide
l
we.
In vitro effects of prostaglandins Fw, BI and B2 on circular muscles of human respiratory tract
DISCUSSION In animal studies both bronchodilatatlon and bronchoconstrictlon have been reported with PGA and PGBcompounds in vivo (see Introduction). On cat tracheal muscle in vitro FGBcompounds inhibited acetylcholine and carbachol induced contractions, indicating bronchodilatation, whereas on guinea pig tracheal chain preparation PGBcompounds showed constrictor effect which was more potent than the stimulant actlon of PGF% (11). In the present study, however, PGA1, FGA2, FGBl and pGB2 produced contracttons of circular muscle strips from the trachea-bronchial smooth muscle of man. In partkular, FGB2was shown to be more potent than prostaglandins Al, A2, BI and FW Prostaglandins of the A and B series have previously been showd to be more potent than IGE2 and PGFa in their stimulant effect on other human smooth muscles such as human umbilical cord (12) and human penile muscle (13). Unlike PGFwprostaglandins of the A and B series are not metabolised by prostaglandin 15 hydroxy dehydrogenase, although metabolism by other exzymes may occur (7,14). It is therefore likely that in in vivo situation the bronchoconstrictor effects of PGA and PGBcompounds may be more pronounced. The unnatural Isomer of PGA2 (15”epi-FGA2) produces bronchoconstrictor effect in man (15) and acute bronchoconstrlction haa been encountered in women, when PGA2 or PGB2are used to stimulate second trimester pregnant uterus (16).
534
MARCH
1976
VOL. 11 NO. 3
PROSTAGLANDINS
Prostaglandin Fa produces bronchoconstriction in man (3). This prostaglsndin also has a constrictor effect on the circular strips of human bronchial muscle in vitro (17). A correlation therefore exists between in vitro and in vtvo findings for PGF= PGA2 and PGB2. Similarly, both bzn-chodilator and bronchoconstrictor effects reported in in vivo situations with PGE2 (18, 19) have been confirmed on human bronchial muscle (17) and tracheo-bronchial muscles (20) in vitro.
ACKNOWLEDGEMENTS Kalamazoo, Prostaglandins were supplied by the Upjohn Co., Michigan. The work was carried out under USAID Grant Contract No. AID/ cm/pha-c-73/36 and a research grant from The Wellcome Trust, U. K. We are grateful to Dr. Chao Tzee Cheng, Department of Pathology, Singapore General Hospital for post-mortem specimens.
REFERENCES 1.
ingg&d, E. (1965). The isolation and determination of prostaglandins in the lungs of sheep, guinea pig, monkey and man. Biochem. Pharmacol. 1507. -’14
2.
Karim, S. M. M., Sandler, M. and Williams, E.D. (1967). Distribution of prostaglandins in human tissues. Br. J. Pharmac. 31, 340-344.
3.
Smith, A. P. (1976). Prostaglandins and the respiratory system. In : Advances in Prostaglandiu Research : Physiological, Pharmacological and Pathological Aspects. P. 83-102 (S. M. M. Karim, editor), Medical and Technical Publications, Lancaster.
4.
Hamberg, M. and Samuelsson, B. (1967). New groups of naturally occurring prostaglandins. In : Prostaglandins, Nobel Symposium II (eds. S. Bergstrom and B. Samuelsson), Stockholm, Almqvtst and Wiksell, New York, Interscience, P. 63-70.
5.
Eisenbarth, G. S. and Lebovitz, H. E. (1974). Prostaglandin inhibition of cartilage chondromucoprotein synthesis : concept of ‘Intrinsic Activity “. Prostaglandtns z, 11-20.
6.
Wong, P. Y. and Wuthier, R. E. (1974). Isolation and identification of prostaglandin PGBl in growth cartilage. Prostaglandins, g, 125-132.
7.
Horton, E. W. and Jones, R. L. (1969). Prostaglandins AI, A2 and 19hydroxy AI; their act ions of smooth muscle and their inactivation on
MARCH
1976
VOL. 11 NO. 3
535
PROSTAGLANDINS
passage through the pulmonary and hepatic portal vascular beds. Br. J. Pharmacol. -37, 705-722. 8.
Ballantyne, A., Jones, R.L. and Ungar, A. (1973). The actions of prostaglandins Al and A2 on airway resistance and compliance in the cat. Br. J. Pharmacol. -47, 63OP-631P.
9.
Hirose, R. and Said,S.I. (1971). Respiratory effects of prostaglandins Al, El and F20. Clin. Res. 2, 512.
10. Greenberg, S., Howard, L. and Wilson, W. R. (1973). Comparative effects of prostaglandins A2 and B2 on vascular and airway resistances and adrenergic neurotransmission. Can. J. Physiol. Pharmacol. 2, 699-705. 11. Lo, P. Y. and Karim, S. M. M. (1976). Effects of A and B prostaglandins on guinea pig tracheal chain. In Preparation. 12. Adaikan, P.G. and Karim, S. M.M. (1974). Effects of Al, AZ, Bl, B2, E2 and Fa on human umbilical cord vessels. Prostaglandins 8, 411-416. 13. Karim, S. M. M. and Adaikan, P. G. (1975). In : Advances in Prostaglandin Research : Prostaglandins and Reproduction, P. 26-28 (S. M. M. Karim, eidtor), Medical and Technical Publications, Lancaster. 14. Attallah, A. A., prostaglandin A. tion. Life Sci.,
Payskkapan, W. and Lee, J.B. (1974). Metabolism of 1. The kidney cortex as a major site of FGA2 degrada-14, 1521-1534.
15. Smith, A. P. (1974). Effects of three prostaglandin analogues on airway tone in asthmatics and normal subjects. Int. Res. Commun. System, 2, 1457. 16. Karim, S. M. M. (1976). Effect of prostaglandin B2 on the pregnant human uterus. In Preparation. 17. Gardiner, P. J. (1975). The effects of some natural prostaglandins on isolated human circular bronchial muscle. Prostaglandins, s, 607-616. 18. Cuthbert, M. F. (1971). Bronchodilator activity of aerosols of prostaglandins El and E2 in asthmatic subjects. Proc. R. Sot. Med. -64, 15-16. 19. Mathe, A.A., Hedqvist, P., Holmgren, A. and Svanborg, N. (1973). Bronchial hyperractivity of prostaglandin I?- and histamine in patients with asthma. Br. Med. J. I, 193-196. 20. Lo, P.Y., Adaikan, P. G. and Karim, S.M. M. (1976).The effect of PGE2 and its metabolites on human airway smooth muscle in vitro. In Preparation. 536
MARCH
1976
VOL. 11 NO. 3