Advertisement

Journal of Endocrinological Investigation

, Volume 2, Issue 2, pp 173–182 | Cite as

Radioimmunoassay measurement of prostaglandins E2 and F in human urine

  • G. Ciabattoni
  • F. Pugliese
  • M. Spaldi
  • G. A. Cinotti
  • C. Patrono
Article

Abstract

Techniques are described in detail for the radioimmunological measurement of prostaglandin (PG) E2 and F in human urine. 50 ml urine samples are extracted with an organic solvent system and then purified by silicic acid column chromatography. The overall recovery after extraction and purification, calculated with labeled as well as unlabeled compounds, is in the order of 70%. The column eluates are assayed at 1:12–1:60 dilution in the “standard diluent” of the assay: 8 pg PGE2.or PGF2a/ml of whole urine represents the lowest measurable concentration. A urine blank and a solvent blank were evaluated separately, subjecting 50 ml of urine obtained from an indomethacin treated subject (“PG-free” urine) or 50 ml of distilled water, respectively, to the extraction-purification procedures. Both were found not to interfere with the antigen-antibody reaction. Urinary PG-like immunoreactivity (LI) was characterized in terms of immunochemical and thin-layer chromatographic (TLC) behavior. Both urinary PGE2-LI and PGF-LI behaved as authentic PGE2 and PGF, upon dilution and on TLC. In 33 healthy female subjects (aged 19–58 yr), urinary excretion rates averaged 178 ± 80 (mean ± SD) ng/day for PGE2 and 498 ± 181 ng/day for PGF. In a group of 8 healthy men, both PGE2 and PGF crexcretion rates were higher and more scattered than the female values. When two healthy women were given indomethacin (200 mg/day), urinary PGE2 and PGF dropped to undetectable levels during the 4th day of drug therapy. I ntravenous injection of furosemide (50 mg) in a female volunteer was followed by an immediate rise of urinary sodium, PGE2, PGF and PGE2/PGF ratio and plasma renin activity. Ina 10 year old girl with Bartter’s syndrome, urinary PG excretion rate was elevated with a 3 times higher than normal PGE2/PGF ratio. Indomethacin therapy resulted in a prompt drop of PG excretion rate and of plasma renin activity. These results show that a combi nation of adequate purification steps and antisera with optimal characteristics provides a reliable method for measuring PGE2 and PGF in urine, and therefore a valuable tool to further our knowledge of the physiology and physiopathology of the renal PG-system.

Key-words

Renal prostaglandins human urine radioimmunoassay indomethacin furosemide Bartter’s syndrome 

Abbreviations

PG

prostaglandin

TLC

thin layer chromatography

PGI2

prostacyclin

EPA 44069

15 (S)-hydroxy-9 α, 11 α-(epoxymethano)-prosta-5Z, 13E-dienoic acid

EPA 46619

15(S)-hydroxy-11 α, 9α-(epoxymethano)-prosta-5Z, 13E dienoic acid

TXB2

thromboxane B2

PGE-M

7α-hydroxy-5, 11-diketotetranorprostane 1,16 dioic acid

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Frölich J.C., Wilson T.W., Sweetman B.J., Smigel M., Nies A.S., Carr K., Watson J.T., Oates J.A. Urinary prostaglandins. Identification and origin. J.Clin. Invest. 55: 763, 1975.PubMedCentralPubMedCrossRefGoogle Scholar
  2. 2.
    Gill J.R. Jr., Frölich J.C., Bowden R.E., Taylor A.A., Keiser H.R., Seyberth H.W., Oates J.A., Bartter F.C. Bartter’s syndrome: a disease characterized by high urinary prostaglandins and a dependence of hyperreninemia on prostaglandin synthesis. Am. J. Med. 67: 43, 1976.CrossRefGoogle Scholar
  3. 3.
    La Torre E., Patrono C., Fortuna A., Grossi-Belloni D. Role of prostaglandin F human cerebral vasospasm. J. Neurosurg 47: 293, 1974.CrossRefGoogle Scholar
  4. 4.
    Wolfe L.S., Marner O.A. Measurement of prostaglandin F levels in human cerebrospinal fluid in normal and pathologic conditions. Prostaglandins 9: 183, 1975.PubMedCrossRefGoogle Scholar
  5. 5.
    Jaffe B.M., Behrman H.R., Parker C.W. Radioimmunoassay measurement of prostaglandins E, A, and F in human plasma. J.Clin. Invest. 52: 398, 1973.PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Cottee F., Flower R.J., Moncada S., Salmon J.A., Vane J.R. Synthesis of 6-keto-PGF by ram seminal vesicle microsomes. Prostaglandins 14: 413, 1977.PubMedCrossRefGoogle Scholar
  7. 7.
    Patrono C. Radioimmunoassay of prostaglandin F. Symposium on Radioimmunoassay Techniques, Paris, May 15–19, Institut National de la Santé et de la Recherche Médicale Paris, 1972, p. 291.Google Scholar
  8. 8.
    Patrono C. Radioimmunoassay of prostaglandin F in unextracted human plasma. J. Nucl. Biol. Med. 17:25, 1973.PubMedGoogle Scholar
  9. 9.
    Patrono C., Ciabattoni G., Grossi-Belloni D. Release of prostaglandin F and F from superfused platelets: quantitative evaluation of the inhibitory effects of some aspirin-like drugs. Prostaglandins 9: 557, 1975.PubMedCrossRefGoogle Scholar
  10. 10.
    Malvano R., Zucchelli G.C., Rosa U., Salvetti A. Measurement of plasma renin activity by angiotensin I radioimmunoassay: I) anassessment of some methodological aspects. J. Nucl. Biol. Med. 16: 24, 1972.PubMedGoogle Scholar
  11. 11.
    Togna G., Gandolfi C., Andreoni A., Fumagalli A., Passa-rotti C., Faustini F., Patrono C. Inhibition of human platelet aggregation by stable analogs of prostacyclin. Pharmacol. Res. Commun. 9: 909,1977.PubMedCrossRefGoogle Scholar
  12. 12.
    Nicosia Zaraga S., Galli G. A rapid gas chromatographic-mass spectrometry method for prostaglandin analysis at picomole levels Anal. Biochem. 61: 192, 1974.CrossRefGoogle Scholar
  13. 13.
    Berson S.A., Yalow R.S. Radioimmunoassay of ACTH in plasma. J. Clin. Invest. 47: 2725, 1968.PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Vane J.R. Inhibition of prostaglandin synthesis as a mechanism of action for aspirin-like drugs. Nature New. Biol. 237: 232, 1971.CrossRefGoogle Scholar
  15. 15.
    Weber P.C., Scherer B., Larsson C. Increase of free arachidonic acid by furosemide in man as the cause of prostaglandin and renin release. Eur. J. Pharmac. 41: 329, 1977.CrossRefGoogle Scholar
  16. 16.
    Zia P., Speckart P., Zipser R., Horton R. A radioimmunoassay of PGE for human plasma and urine. Prostaglandins 13: 1005, 1977.Google Scholar
  17. 17.
    Beckmann L., Leovey E., Armstrong C., Ramwell P.W. Arachidonic acid and iPGE and iPGF levels in human urine. Clin. Res 25: 426A, 1977.Google Scholar
  18. 18.
    Abe K., Yasujima M., Chiba S., Irokawa N., Ito T., Yoshinaga K. Effect of furosemide on urinary excretion of prostaglandin E in normal volunteers and patients with essential hypertension. Prostaglandins 14: 513, 1977.PubMedCrossRefGoogle Scholar
  19. 19.
    Dray F. Bartter’s syndrome: contrasting patterns of prostaglandin excretion in children and adults. Clin. Sci. Mol. Med. 54: 115, 1978.PubMedGoogle Scholar
  20. 20.
    Tan S.Y., Sweet P., Murlow P.J. Impaired renal production of prostaglandin E2: a newly identified lesion in human essential hypertension. Prostaglandins 75: 139, 1978.CrossRefGoogle Scholar
  21. 21.
    Scherer B., Schnermann J., Sofroniev M. Weber PC Prostaglandin (PG) analysis in urine of humans and rats by different radioimmunoassays: effect on PG-excretion by PG-synthetase inhibitors, laparotomy and furosemide. Prostaglandins 75: 255, 1978.CrossRefGoogle Scholar
  22. 22.
    Pace-Asciak C.R., Wolfe L.S. A novel prostaglandin derivative formed from arachidonic acid by rat stomach homogenate. Biochemistry 10: 3657, 1971.PubMedCrossRefGoogle Scholar
  23. 23.
    Patrono C., Ciabattoni G., Cinotti G.A., Pugliese F., De Salvo A., Castrucci G. Characterization of human urinary prostaglandin-like immunoreactivity. Prostaglandins 15: 700, 1978.Google Scholar
  24. 24.
    Perry D.L., Desiderio D.M. Utility of d 4-PGE2 as an internal standard to quantify endogenous levels of PGE1, PGE2, 190H PGE1( and 190H PGE2 in human seminal fluid by GC-MS-SIM. Prostaglandins 14: 745, 1977.PubMedCrossRefGoogle Scholar
  25. 25.
    Yalow R.S., Berson S.A. Problems of validation of radioimmunoassay In: Odell W.D., Daughaday W.H. (Eds.) Principles of competitive protein-binding assays J.B. Lippincott Company, Philadelphia and Toronto, 1971, p. 374.Google Scholar
  26. 26.
    Weber P.C., Larsson C., Änggård E., Hamberg M., Corey E.J., Nicolau K.C., Samuelsson B. Stimulation of renin release from rabbit renal cortex by arachidonic acid and prostaglandin endoperoxides. Circulat. Res. 39: 868, 1976.CrossRefGoogle Scholar
  27. 27.
    Stone K.J., Hart M. Inhibition of renal PGE2-9-ketoreductase by diuretics Prostaglandins 12: 197, 1976.PubMedCrossRefGoogle Scholar
  28. 28.
    Weber P.C., Larsson C., Scherer B. Prostaglandin E2-9-ketoreductase as a mediator of salt intake-related prostaglandin-renin interaction. Nature 266: 65, 1977.CrossRefGoogle Scholar

Copyright information

© Italian Society of Endocrinology (SIE) 1978

Authors and Affiliations

  • G. Ciabattoni
    • 1
    • 2
  • F. Pugliese
    • 1
    • 2
  • M. Spaldi
    • 1
    • 2
  • G. A. Cinotti
    • 1
    • 2
  • C. Patrono
    • 1
    • 2
  1. 1.Department of PharmacologyCatholic UniversityRomeItaly
  2. 2.Department of Medicine IIUniversity of RomeRomeItaly

Personalised recommendations