Mass Spectrometric and Immunologic Detection of Prolactin-Derived Vasoinhibin in Human Serum.

Abstract

Background: Circulating levels of the antiangiogenic protein, vasoinhibin, derived from the proteolytic cleavage of prolactin (PRL), in prolactinoma are unknown, as is the molecular nature of its isoforms. Dimerization of recombinant vasoinhibin has been reported.

Methods: Vasoinhibin in a human serum sample was identified by using preparative electrophoresis with subsequent SDS-PAGE and Western blot analysis, as well as mass spectrometry (MS) and ELISA.

Results: MS identified a partial vasoinhibin sequence in a 14-kDa protein band from human serum, which eluted in the 28-kDa fraction from the preparative electrophoresis. Measurement of vasoinhibin levels by ELISA identified a concentration of 284 ng/mL at a PRL level of 9,850 ng/mL. Recombinant human vasoinhibin demonstrated dimerization and multimerization when analyzed directly by SDS-PAGE and Western blot analysis under reducing and non-reducing conditions, as well as after immunoprecipitation.

Conclusions: The vasoinhibin sequence was identified in a higher molecular weight fraction, corroborating experi-mental evidence showing the dimerization and aggregation of recombinant human vasoinhibin. This report is sig-nificant, regarding the higher risk of cardiovascular disease and mortality in male patients with hyperprolactin-emia as well as emerging reports of linking PRL and vasoinhibin levels in patients with prolactinoma with left ventricular dysfunction and Takotsubo syndrome.