Inhibitors of receptor tyrosine kinases do not suppress progesterone-induced [Ca2+]i signalling in human spermatozoa

J. C. Kirkman-Brown (Lead / Corresponding author), L. Lefièvre, C. Bray, Paul M. Stewart, C. L. R. Barratt, S. J. Publicover

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Previous studies have implicated receptor tyrosine kinases in progesterone-induced [Ca2+]i signalling, and consequent induction of the acrosome reaction, in human spermatozoa. We have investigated the effects of tyrosine kinase inhibition on [Ca2+]i responses in large numbers of individual human spermatozoa. Genistein (5, 50 and 250 micromol/l), an inhibitor of receptor-linked tyrosine kinases, significantly inhibited the progesterone-induced acrosome reaction (P < 0.05). However, we could detect no effect of genistein on progesterone-induced [Ca2+]i signalling. In control experiments, application of progesterone induced a significant transient [Ca2+]i response in approximately 77% of cells and a sustained [Ca2+]i ramp/plateau in approximately 48% of cells (n = 26; 5411 cells). In preparations pretreated with genistein (50 micromol/l), significant transient and sustained responses were detected in 69.5 and 39.1% of cells respectively (n = 5; 1109 cells). The amplitudes of both transient and sustained [Ca2+]i responses were similar in control and genistein-pretreated preparations. Tyrphostin A47 (100 micromol/l), another receptor tyrosine kinase inhibitor, also failed to inhibit either the transient or sustained [Ca2+]i response (n = 3; 468 cells). Assessment of tyrosine phosphorylation of two sperm proteins (p105/81) showed greatly increased levels of phosphotyrosine in response to capacitation but a negligible increase in response to progesterone stimulation. Pretreatment with genistein (50 and 250 micromol/l) decreased capacitation-induced tyrosine phosphorylation and resulted in a loss of phosphorylation in response to progesterone treatment. We conclude that neither the transient nor sustained phases of the progesterone-induced [Ca2+]i response require receptor tyrosine kinase signalling. Previous reports of modulation of the progesterone-induced [Ca2+]i signal by tyrosine kinase inhibition probably reflect inhibition of the acrosome reaction.

Original languageEnglish
Pages (from-to)326-332
Number of pages7
JournalMolecular Human Reproduction
Volume8
Issue number4
DOIs
Publication statusPublished - 1 Apr 2002

Fingerprint

Receptor Protein-Tyrosine Kinases
Progesterone
Spermatozoa
Genistein
Acrosome Reaction
Phosphorylation
Protein-Tyrosine Kinases
Tyrosine
Architectural Accessibility
Phosphotyrosine

Keywords

  • Acrosome/enzymology
  • Blotting, Western
  • Calcium/metabolism
  • Calcium Signaling/drug effects
  • Enzyme Inhibitors/pharmacology
  • Genistein/pharmacology
  • Humans
  • In Vitro Techniques
  • Male
  • Progesterone/metabolism
  • Receptor Protein-Tyrosine Kinases/antagonists & inhibitors
  • Signal Transduction/physiology
  • Spermatozoa/enzymology
  • Tyrosine/metabolism
  • Tyrphostins/pharmacology

Cite this

Kirkman-Brown, J. C. ; Lefièvre, L. ; Bray, C. ; Stewart, Paul M. ; Barratt, C. L. R. ; Publicover, S. J. / Inhibitors of receptor tyrosine kinases do not suppress progesterone-induced [Ca2+]i signalling in human spermatozoa. In: Molecular Human Reproduction. 2002 ; Vol. 8, No. 4. pp. 326-332.
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abstract = "Previous studies have implicated receptor tyrosine kinases in progesterone-induced [Ca2+]i signalling, and consequent induction of the acrosome reaction, in human spermatozoa. We have investigated the effects of tyrosine kinase inhibition on [Ca2+]i responses in large numbers of individual human spermatozoa. Genistein (5, 50 and 250 micromol/l), an inhibitor of receptor-linked tyrosine kinases, significantly inhibited the progesterone-induced acrosome reaction (P < 0.05). However, we could detect no effect of genistein on progesterone-induced [Ca2+]i signalling. In control experiments, application of progesterone induced a significant transient [Ca2+]i response in approximately 77{\%} of cells and a sustained [Ca2+]i ramp/plateau in approximately 48{\%} of cells (n = 26; 5411 cells). In preparations pretreated with genistein (50 micromol/l), significant transient and sustained responses were detected in 69.5 and 39.1{\%} of cells respectively (n = 5; 1109 cells). The amplitudes of both transient and sustained [Ca2+]i responses were similar in control and genistein-pretreated preparations. Tyrphostin A47 (100 micromol/l), another receptor tyrosine kinase inhibitor, also failed to inhibit either the transient or sustained [Ca2+]i response (n = 3; 468 cells). Assessment of tyrosine phosphorylation of two sperm proteins (p105/81) showed greatly increased levels of phosphotyrosine in response to capacitation but a negligible increase in response to progesterone stimulation. Pretreatment with genistein (50 and 250 micromol/l) decreased capacitation-induced tyrosine phosphorylation and resulted in a loss of phosphorylation in response to progesterone treatment. We conclude that neither the transient nor sustained phases of the progesterone-induced [Ca2+]i response require receptor tyrosine kinase signalling. Previous reports of modulation of the progesterone-induced [Ca2+]i signal by tyrosine kinase inhibition probably reflect inhibition of the acrosome reaction.",
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Inhibitors of receptor tyrosine kinases do not suppress progesterone-induced [Ca2+]i signalling in human spermatozoa. / Kirkman-Brown, J. C. (Lead / Corresponding author); Lefièvre, L.; Bray, C.; Stewart, Paul M.; Barratt, C. L. R.; Publicover, S. J.

In: Molecular Human Reproduction, Vol. 8, No. 4, 01.04.2002, p. 326-332.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Inhibitors of receptor tyrosine kinases do not suppress progesterone-induced [Ca2+]i signalling in human spermatozoa

AU - Kirkman-Brown, J. C.

AU - Lefièvre, L.

AU - Bray, C.

AU - Stewart, Paul M.

AU - Barratt, C. L. R.

AU - Publicover, S. J.

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N2 - Previous studies have implicated receptor tyrosine kinases in progesterone-induced [Ca2+]i signalling, and consequent induction of the acrosome reaction, in human spermatozoa. We have investigated the effects of tyrosine kinase inhibition on [Ca2+]i responses in large numbers of individual human spermatozoa. Genistein (5, 50 and 250 micromol/l), an inhibitor of receptor-linked tyrosine kinases, significantly inhibited the progesterone-induced acrosome reaction (P < 0.05). However, we could detect no effect of genistein on progesterone-induced [Ca2+]i signalling. In control experiments, application of progesterone induced a significant transient [Ca2+]i response in approximately 77% of cells and a sustained [Ca2+]i ramp/plateau in approximately 48% of cells (n = 26; 5411 cells). In preparations pretreated with genistein (50 micromol/l), significant transient and sustained responses were detected in 69.5 and 39.1% of cells respectively (n = 5; 1109 cells). The amplitudes of both transient and sustained [Ca2+]i responses were similar in control and genistein-pretreated preparations. Tyrphostin A47 (100 micromol/l), another receptor tyrosine kinase inhibitor, also failed to inhibit either the transient or sustained [Ca2+]i response (n = 3; 468 cells). Assessment of tyrosine phosphorylation of two sperm proteins (p105/81) showed greatly increased levels of phosphotyrosine in response to capacitation but a negligible increase in response to progesterone stimulation. Pretreatment with genistein (50 and 250 micromol/l) decreased capacitation-induced tyrosine phosphorylation and resulted in a loss of phosphorylation in response to progesterone treatment. We conclude that neither the transient nor sustained phases of the progesterone-induced [Ca2+]i response require receptor tyrosine kinase signalling. Previous reports of modulation of the progesterone-induced [Ca2+]i signal by tyrosine kinase inhibition probably reflect inhibition of the acrosome reaction.

AB - Previous studies have implicated receptor tyrosine kinases in progesterone-induced [Ca2+]i signalling, and consequent induction of the acrosome reaction, in human spermatozoa. We have investigated the effects of tyrosine kinase inhibition on [Ca2+]i responses in large numbers of individual human spermatozoa. Genistein (5, 50 and 250 micromol/l), an inhibitor of receptor-linked tyrosine kinases, significantly inhibited the progesterone-induced acrosome reaction (P < 0.05). However, we could detect no effect of genistein on progesterone-induced [Ca2+]i signalling. In control experiments, application of progesterone induced a significant transient [Ca2+]i response in approximately 77% of cells and a sustained [Ca2+]i ramp/plateau in approximately 48% of cells (n = 26; 5411 cells). In preparations pretreated with genistein (50 micromol/l), significant transient and sustained responses were detected in 69.5 and 39.1% of cells respectively (n = 5; 1109 cells). The amplitudes of both transient and sustained [Ca2+]i responses were similar in control and genistein-pretreated preparations. Tyrphostin A47 (100 micromol/l), another receptor tyrosine kinase inhibitor, also failed to inhibit either the transient or sustained [Ca2+]i response (n = 3; 468 cells). Assessment of tyrosine phosphorylation of two sperm proteins (p105/81) showed greatly increased levels of phosphotyrosine in response to capacitation but a negligible increase in response to progesterone stimulation. Pretreatment with genistein (50 and 250 micromol/l) decreased capacitation-induced tyrosine phosphorylation and resulted in a loss of phosphorylation in response to progesterone treatment. We conclude that neither the transient nor sustained phases of the progesterone-induced [Ca2+]i response require receptor tyrosine kinase signalling. Previous reports of modulation of the progesterone-induced [Ca2+]i signal by tyrosine kinase inhibition probably reflect inhibition of the acrosome reaction.

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KW - Blotting, Western

KW - Calcium/metabolism

KW - Calcium Signaling/drug effects

KW - Enzyme Inhibitors/pharmacology

KW - Genistein/pharmacology

KW - Humans

KW - In Vitro Techniques

KW - Male

KW - Progesterone/metabolism

KW - Receptor Protein-Tyrosine Kinases/antagonists & inhibitors

KW - Signal Transduction/physiology

KW - Spermatozoa/enzymology

KW - Tyrosine/metabolism

KW - Tyrphostins/pharmacology

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DO - 10.1093/molehr/8.4.326

M3 - Article

VL - 8

SP - 326

EP - 332

JO - Molecular Human Reproduction

JF - Molecular Human Reproduction

SN - 1360-9947

IS - 4

ER -