- Email: [email protected]
MEMBRANE STABILISING ACTIVITY AND INHIBITION OF HUMAN SPERM MOTILITY
402 MEMBRANE STABILISING ACTIVITY AND INHIBITION OF HUMAN SPERM MOTILITY
SiR,—Dr Henry and Mr Cassidy (June 21, p 1414) proposed that stabilising activity (MSA) is a major cause of fatal
membrane
poisoning. They also stated that there has been little comparison of the degree of MSA of different agents. We have compared the MSA of drugs, on the basis of their ability to inhibit human sperm motility. Human sperm motility can be assessed by a transmembrane migration method,! which measures the proportion of sperm that moves across the 5 pm pores of a ’Nuclepore’ membrane in 2 h of incubation at 37°C. Water-soluble drugs are first dissolved in phosphate-buffered saline (Delbecco A, pH 73) and then mixed with freshly ejaculated human semen. Water-insoluble drugs are dissolved in chloroform and then pipetted into glass tubes; these tubes are heated on a sand plate to allow the chloroform to evaporate before the semen is addedThe ECso is the concentration of drug that decreases sperm motility to 50% of that of the control sample. The
ECso
of 27 chemicals with MSA
I
are
shown in the table.
5.
Hong CY, Chiang BN, Ku J, Wu P. Screening the in vitro sperm immobilizing effect of some anticancer drugs. Human Toxicol 1985; 4: 461-64. 6. Verma RB, Abrams SML, Turner P. Effect of indoramin on human sperm motility Br J Clin Pharmacol 1983; 15: 127-28. 7. Penhall RK, Hong CY, Muhiddin KA. The effect of flecainide on human sperm motility. Br J Clin Pharmacol 1982; 14: 147P. 8. Zaman S, Lamb JM, Esberger DA, Pearson RM. Inhibition of sperm motility by opiate drugs. Br J Clin Pharmacol 1984; 18: 302P.
NATURE OF ATRIAL NATRIURETIC PEPTIDE IN PLASMA FROM PATIENTS WITH CONGESTIVE HEART FAILURE
SiR,—The raised plasma levels of atrial natriuretic peptide (ANP) found in patients with congestive heart failure (CHF)1 may reflect compensation in response to volume expansion in CHF.2 Gerbes and colleagues3 have suggested that incomplete processing of ANP precursor may result in high plasma levels of immunoreactive material with reduced diuretic activity in patients with cardiovascular diseases and that evaluation of the molecular weight pattern of ANP may be essential for the interpretation of plasma ANP levels. To test this hypothesis we extracted ANP in plasma (16 ml) pooled from three patients with CHF class III or IV by affinity chromatography on agarose-linked ANP antibody. We analysed the ANP by reverse-phase high-performance liquid chromatography (HPLC)4 on an octadecylsilane column with a linear gradient system from 10 to 60% acetonitrile over 80 min. The flow rate was 1 ml/min and 1 ml fractions were collected, lyophilised, reconstituted with the assay buffer, and assayed for ANP.4,5 When an extract of atrial tissue obtained during surgery for mitral valve replacement was analysed by affmity chromatography and HPLC three major peaks of immunoreactive ANP were observed (figure, B). The retention time of peak I coincided with that of synthetic a-ANP. On sodium dodecylsulphate/polyacrylamide gel electrophoresis without reducing agents, immunoreactive ANP in these three peaks migrated in the regions of molecular weights of 3000, 6000, and 14 000. Thus, peak I may correspond to A-ANP, peak II to P-ANP, and peak III to y-ANP (proANP).6The recovery of cx-ANP was 81%, while that of y-ANP was 52%. Analysis of the plasma extract by HPLC revealed peaks of immunoreactive ANP (figure, A) but the retention time of the main peak coincided with that of A-ANP and little, if any, activity in the region of y-ANP was observed. These results suggest that the major form of the circulating ANP in patients with CHF is a-ANP, as it is in healthy individuals.’
t
*Data derived from authors’ studies when
no
references cited.
Sotalol, a p-adrenoceptor blocking drug, without MSA,3potassium cyanide, a mitochondrial respiration inhibitorand colchicine, a microtubule inhibitory have little sperm immobilising effect. Since the sperm immobilising ability of 0-adrenoceptor blocking agents correlates well with lipid solubility of the drug, which in turn determines MSA,3ahuman sperm motility bioassay is a simple means of assessing the MSA of drugs. Department of Medicine, Taiwan Veterans General Hospital, and Yang-Ming Medical College,
Taipei, Taiwan, Republic of China
C. Y. HONG P. Wu C. C. SHIEH B. N. CHIANG
1. Hong CY, Chaput de Saintonge DM, Turner P. A simple method to measure drug effect on human sperm motility. Br J Clin Pharmacol 1981; 11: 385-87. 2. Hong CY, Shieh CC, Wu P, Chiang BN. Effect of phosphatidylcholine, lysophosphatidylcholine, arachidonic acid and docosahexaenoic acid on human sperm motility. Int J Androl (in press). 3. Hong CY, Turner P. Influence of lipid solubility on the sperm immobilising effect of beta-adrenoceptor blocking drugs. Br J Clin Pharmacol 1982; 14: 269-72. 4 Hong CY, Chiang BN, Wei YH. Mitochondrial respiration inhibitors and human sperm motility. implication in the future developments of spermicides Br J Clin Pharmacol 1983; 16: 487-90.
HPLC
analysis of immunoreactive ANP in human plasma patients with CHF (A) and in atrial tissue (B).
from
SiR,—Dr Henry and Mr Cassidy (June 21, p 1414) proposed that stabilising activity (MSA) is a major cause of fatal
membrane
poisoning. They also stated that there has been little comparison of the degree of MSA of different agents. We have compared the MSA of drugs, on the basis of their ability to inhibit human sperm motility. Human sperm motility can be assessed by a transmembrane migration method,! which measures the proportion of sperm that moves across the 5 pm pores of a ’Nuclepore’ membrane in 2 h of incubation at 37°C. Water-soluble drugs are first dissolved in phosphate-buffered saline (Delbecco A, pH 73) and then mixed with freshly ejaculated human semen. Water-insoluble drugs are dissolved in chloroform and then pipetted into glass tubes; these tubes are heated on a sand plate to allow the chloroform to evaporate before the semen is addedThe ECso is the concentration of drug that decreases sperm motility to 50% of that of the control sample. The
ECso
of 27 chemicals with MSA
I
are
shown in the table.
5.
Hong CY, Chiang BN, Ku J, Wu P. Screening the in vitro sperm immobilizing effect of some anticancer drugs. Human Toxicol 1985; 4: 461-64. 6. Verma RB, Abrams SML, Turner P. Effect of indoramin on human sperm motility Br J Clin Pharmacol 1983; 15: 127-28. 7. Penhall RK, Hong CY, Muhiddin KA. The effect of flecainide on human sperm motility. Br J Clin Pharmacol 1982; 14: 147P. 8. Zaman S, Lamb JM, Esberger DA, Pearson RM. Inhibition of sperm motility by opiate drugs. Br J Clin Pharmacol 1984; 18: 302P.
NATURE OF ATRIAL NATRIURETIC PEPTIDE IN PLASMA FROM PATIENTS WITH CONGESTIVE HEART FAILURE
SiR,—The raised plasma levels of atrial natriuretic peptide (ANP) found in patients with congestive heart failure (CHF)1 may reflect compensation in response to volume expansion in CHF.2 Gerbes and colleagues3 have suggested that incomplete processing of ANP precursor may result in high plasma levels of immunoreactive material with reduced diuretic activity in patients with cardiovascular diseases and that evaluation of the molecular weight pattern of ANP may be essential for the interpretation of plasma ANP levels. To test this hypothesis we extracted ANP in plasma (16 ml) pooled from three patients with CHF class III or IV by affinity chromatography on agarose-linked ANP antibody. We analysed the ANP by reverse-phase high-performance liquid chromatography (HPLC)4 on an octadecylsilane column with a linear gradient system from 10 to 60% acetonitrile over 80 min. The flow rate was 1 ml/min and 1 ml fractions were collected, lyophilised, reconstituted with the assay buffer, and assayed for ANP.4,5 When an extract of atrial tissue obtained during surgery for mitral valve replacement was analysed by affmity chromatography and HPLC three major peaks of immunoreactive ANP were observed (figure, B). The retention time of peak I coincided with that of synthetic a-ANP. On sodium dodecylsulphate/polyacrylamide gel electrophoresis without reducing agents, immunoreactive ANP in these three peaks migrated in the regions of molecular weights of 3000, 6000, and 14 000. Thus, peak I may correspond to A-ANP, peak II to P-ANP, and peak III to y-ANP (proANP).6The recovery of cx-ANP was 81%, while that of y-ANP was 52%. Analysis of the plasma extract by HPLC revealed peaks of immunoreactive ANP (figure, A) but the retention time of the main peak coincided with that of A-ANP and little, if any, activity in the region of y-ANP was observed. These results suggest that the major form of the circulating ANP in patients with CHF is a-ANP, as it is in healthy individuals.’
t
*Data derived from authors’ studies when
no
references cited.
Sotalol, a p-adrenoceptor blocking drug, without MSA,3potassium cyanide, a mitochondrial respiration inhibitorand colchicine, a microtubule inhibitory have little sperm immobilising effect. Since the sperm immobilising ability of 0-adrenoceptor blocking agents correlates well with lipid solubility of the drug, which in turn determines MSA,3ahuman sperm motility bioassay is a simple means of assessing the MSA of drugs. Department of Medicine, Taiwan Veterans General Hospital, and Yang-Ming Medical College,
Taipei, Taiwan, Republic of China
C. Y. HONG P. Wu C. C. SHIEH B. N. CHIANG
1. Hong CY, Chaput de Saintonge DM, Turner P. A simple method to measure drug effect on human sperm motility. Br J Clin Pharmacol 1981; 11: 385-87. 2. Hong CY, Shieh CC, Wu P, Chiang BN. Effect of phosphatidylcholine, lysophosphatidylcholine, arachidonic acid and docosahexaenoic acid on human sperm motility. Int J Androl (in press). 3. Hong CY, Turner P. Influence of lipid solubility on the sperm immobilising effect of beta-adrenoceptor blocking drugs. Br J Clin Pharmacol 1982; 14: 269-72. 4 Hong CY, Chiang BN, Wei YH. Mitochondrial respiration inhibitors and human sperm motility. implication in the future developments of spermicides Br J Clin Pharmacol 1983; 16: 487-90.
HPLC
analysis of immunoreactive ANP in human plasma patients with CHF (A) and in atrial tissue (B).
from