Angiotension expression and its effects on Rho signaling may explain the link between hypertension and fibroids

Abstract

OBJECTIVE: A key feature of uterine fibroids is an excess amount of fibrotic extracellular matrix accompanied by altered mechanical homeostasis and by altered Rho signaling. Uterine fibroids are associated with hypertension and there is increasing evidence that angiotensin II (AngII), a major regulator of blood pressure and cardiovascular homeostasis, is involved in tissue remodeling and fibrosis of the lung and heart. We sought to examine whether AngII altered the Rho-signaling pathway in fibroids. DESIGN: Experimental investigation. MATERIALS AND METHODS: Western blot analysis of whole tissue lysates from fibroid and myometrial surgical samples (n=6) were probed for the AngII receptor type 1 (ATR1). Immortalized uterine fibroid and myometrial cells (Malik et al. 2008) were cultured to 60% confluency in 10% Fetal Bovine Serum DMEM/F12. FBS was then reduced to 0.5% for 16 hours and cell lines were then treated for 30 minutes with AngII or an ATR1-blocker Losartan or for 3 minutes with a known RhoA activator lysophospatidic acid (LPA). Active RhoA levels were quantified using the G-LISA method. RESULTS: ATR1 was over-expressed 3-fold (3.5 +/-1.3 integrated density) in fibroid tissue extracts relative to matched myometrium. Untreated uterine fibroid cells had 1.6-fold higher levels of active RhoA relative to untreated myometrial cells. Fibroid cells stimulated with AngII demonstrated increased expression of active RhoA (2.6-fold) but treated myometrial cells remained unchanged. As an internal control, LPA increased active RhoA in myometrial cells (6.2-fold) more than fibroid cells (5.4-fold). Pretreatment with Losartan decreased the active RhoA expression by 35% in fibroid cells. CONCLUSIONS: These data suggest that angiotensin signaling is altered in uterine fibroids. ATR1 expression was increased in fibroid tissue compared to matched myometrium. The positive relationship between angiotensin stimulation and active Rho expression may have relevance to the association of uterine fibroids and hypertensive disease.