Overexpression of TNF-alpha and FasL Induces Apoptosis of Normal and Tumoral Anterior Pituitary Cells.

CANDOLFI M; YU J; LIU G; FERRARI L; BARCIA C; LIU C; PUNTEL M; PISERA D; CASTRO MG; SEILICOVICH A

Congreso; 87º Annual Meeting Endocrine Society; 2005

Our previous work showed that TNF-a and FasL induce apoptosis of anterior pituitary cells and that both proteins may be involved in anterior pituitary cell renewall in female rats. To further analize the effect of these proapoptotic factors, we overexpressed TNF-a and FasL in primary cultures of anterior pituitary cells and in pituitary cell lines. Anterior pituitary cells from female Wistar and Sprague Dawley rats, as well as GH3 and AtT20 cells, were infected with first generation adenoviral vectors encoding TNF-a, FasL or, as a control, b-Galactosidase (b-Gal), under the control of the human cytomegalovirus promoter. Succesful expression of the encoded transgenes were determined by immunocytochemistry. Although we observed basal expression of TNF-a and FasL in control cultures, FACS cell cycle analysis showed that the overexpression of TNF-a and FasL increases the percentage of hypodiploid anterior pituitary cells from both, Wistar and Sprague Dawley rats. Nuclear morphology and TUNEL staining of anterior pituitary cultures from Sprague Dawley rats, revealed that cells undergo an apoptotic death process (TUNEL positive cells: C: 4.0 %; TNF-á: 12.4 %, p<0.01; FasL: 28.5 %, p<0.01, Chi2 test). The proapoptotic effect of TNF-a and FasL was also observed in lactotrope and somatotrope subpopulations from Wistar rats (hypodiploid lactotropes: C: 4.5 ± 2.0 %; TNF-a: 30.8 ± 8.4 %, p<0.01; FasL: 42.2 ± 6.3 , p<0.01, hypodiploid somatotropes: C: 1.6 ± 0.4 %; TNF-a: 8.4 ± 2.6 %, p<0.01; FasL: 16.4 ± 6.4 %, p<0.01, one-way ANOVA). The expression of b-Gal was detected in these cultures but did not affect anterior pituitary cell viability. In the somatolactotrope cell line GH3, we detected strong immunoreactivity for TNFR1 and Fas. TNF-a but not FasL expression was also observed in control cultures of GH3. The overexpression of TNF-a and FasL in this cell line increased the percentage of TUNEL positive cells (C: 3.1 %; TNF-a: 95.1 %, p<0.01; FasL: 75.0 %, p<0.01, Chi2 test). Also, the overexpression of TNF-a and FasL in the corticotrope cell line AtT20 induced apoptosis as detected by the TUNEL method (C: 7.6 %; TNF-a: 91.6 %, p<0.01; FasL: 81.1 %, p<0.01, Chi2 test). In conclusion, these results suggest that TNFR and Fas activity is present not only in the normal anterior pituitary, but also in tumoral pituitary cells. The overexpression of proapoptotic factors could be a useful tool in the therapy of pituitary adenomas.