Estrogenic profiles in lungs from a rat model of pulmonary arterial hypertension measured by liquid chromatography triple quadrupole mass spectrometry.

Pulmonary arterial hypertension (PAH) is a rare, progressive, and ultimately deadly cardiovascular disease. A large disparity in the prevalence and progression of PAH between the sexes exists, which may be underpinned by differences in estrogen signaling. Downstream metabolites of estrogen can possess both protective and harmful effects in health and disease, affecting cellular proliferation and angiogenesis. Understanding estrogenic profiles within the lung requires new analytical approaches. A novel approach to the analysis of estrogens by liquid chromatography-triple quadrupole mass spectrometry in lung tissue was developed and applied to a rodent PAH model induced by SUGEN-Hypoxia, versus vehicle-treated normoxic controls. Both male and female rats were studied (n = 6/group). The protocol was developed, optimized, and validated for a panel of 12 estrogens and their metabolites using solid phase extraction followed by derivatization with 1-(5-fluoro-2,4-dinitrophenyl)-4,4-dimethylpiperazine (MPPZ). Acceptable ranges for recovery (90-104%) and ion suppression (91-109%) of estrogens extracted from lung tissue were achieved. Method validation showed limits of detection of 50 pg/g of tissue (10 pg/mL) and quantitation of 165 pg/g of tissue (33 pg/mL) for most analytes. When the finalized method was applied to rodent tissue samples, four of the analytes were detected, namely estradiol, estrone, 16α-hydroxyestradiol and 16α-hydroxyestrone. Higher amounts of estradiol and estrone were present in lung tissue of female controls compared with males. However, the amounts of estrogen metabolites in lung did not differ in either sex after the induction of PAH. Thus, a novel method to assess estrogenic compounds in fibrous lung tissue was developed, which can be implemented in further studies of sex-specific lung diseases.