Angiotensin peptides regulate angiogenic activity in rat anterior pituitary tumour cell cultures.

INTRODUCTION: Angiogenesis has been shown to be necessary for the development and progression of solid tumours. VEGF is one of the crucial pro-angiogenic cytokines produced by the cells of many of the tumours examined, including various types of anterior pituitary adenomas. Angiotensin II (Ang II) is known to regulate the expression of VEGF in a variety of tissues both in the physiological and pathological conditions. Moreover, an association of the renin-angiotensin system (RAS) with oestrogen-induced vascular changes during the development of rat pituitary PRL-secreting adenoma has already been demonstrated. The aim of the study was to determine the in vitro effects of angiotensin peptides (Ang II, Ang III and Ang IV) on the secretion of VEGF in two anterior pituitary adenoma cell cultures: the culture of the rat pituitary lactosomatotrope tumour cell line (GH3) and the primary culture of rat PRL-secreting tumour induced by diethylstilbestrol (DES).
MATERIAL AND METHODS: GH3 and prolactinoma cells were cultured in an F10 and an F-12 medium respectively and then placed into 24 multiwell plates (10(5) of GH3 cells/well and 10(6) of rat prolactinoma cells/well). After 12 hours of preincubation the cells underwent 24-hour treatment with Ang II, Ang III or Ang IV at final concentrations of 10(-12), 10(-10), 10(-8) or 10(-6)M and, in the case of the GH3 cells, combined treatment with Ang II (10(-10)M) and specific AT1 or AT2 receptor antagonist (losartan or PD123319 respectively at a concentration of 10(-8) or 10(-7) M). The concentration of VEGF in the supernatant collected was determined using specific ELISA assay kits. Statistical evaluation was performed using Student's test and analysis of variance (ANOVA). Differences were considered significant if p < 0.05.
RESULTS: The incubation of both GH3 cells and rat adenoma cells with Ang II, Ang III or Ang IV at concentrations of 10(-12) -10(-8)M resulted in a significant increase in VEGF concentration in the culture medium. Exposure of GH3 cells to Ang III or Ang IV at concentrations of 10(-6)M led to a significant inhibition of cytokine release, and Pearson's correlation curve showed a tendency for Ang II at concentrations of more than 10(-6)M to inhibit VEGF secretion in primary prolactinoma cell culture. The stimulatory influence of Ang II on VEGF secretion in GH3 cell culture was negated by losartan or by PD123319 in both concentrations tested.
CONCLUSIONS: Ang II, Ang III and Ang IV affect the secretion of VEGF in cultures of the rat lactosomatotrope GH3 cell line and primary rat prolactinoma cells. Both AT1 and AT2 receptors mediate the stimulatory action of Ang II on the cytokine release in GH3 cell culture. The mechanism of the observed anti-angiogenic effects of angiotensin peptides remains unexplained.