M. Lange

Secretion of Polypeptide Growth Factors by Human Nonfunctioning Pituitary Adenoma Cells in Culture

The growth-promoting activities of tumor-conditioned media (TU-CM) obtained from 23 cultured human nonfunctioning pituitary adenomas were studied in vitro. TU-CM obtained from adenoma cell cultures increased both cell counts (range: 108-179%; control = growth in serum-free medium = 100%) and 3H-thymidine incorporation (112-139%) of rat pituitary cell cultures, indicating that TU-CM contains growth-stimulating substances. TU-CM was also able to stimulate the growth of normal fibroblasts (3H-thymidine incorporation: 164-178%; cell counts: 145-157%) and endothelial cells (3H-thymidine incorporation: 131-149%; cell counts: 181-217%), suggesting the presence of - possibly angiogenic-growth factors that act on these cell types. However, the growth of hormone-producing cells was also stimulated, since TU-CM increased 3H-thymidine incorporation into rat pituitary cells in the presence of D-Val-MEM, a medium specifically inhibiting growth of fibroblasts. Addition of neutralizing antibodies against transforming growth factor alpha (TGF-alpha), epidermal growth factor (EGF), insulin-like growth factor I (IGF-I) and basic fibroblast growth factor (bFGF), either alone or in different combinations, reduced the growth-promoting activity of TU-CM on rat pituitary cells (range: 96-71%; control = growth effect of TU-CM without antibodies = 100%), strongly indicating the presence of these growth factors in TU-CM. All 4 antibodies together completely inhibited the growth-stimulatory activity of TU-CM, strongly suggesting that these growth factors play the major role among growth-stimulating substances in TU-CM. This is the first study giving evidence that TGF-alpha, EGF, IGF-I and bFGF are secreted by nonfunctioning adenoma cells indicating that the growth factors could be involved in growth regulation of pituitary adenomas by paracrine or autocrine mechanisms.

Toll-like receptor-4 is expressed in meningiomas and mediates the antiproliferative action of paclitaxel.

Meningiomas are the second most common type of brain and CNS tumors by histology. Surgery and radiotherapy are main treatment options, but meningiomas may be impossible to adequately resect or may regrow after surgery. In spite of many experimental attempts, there is no generally accepted chemotherapeutic approach. We have studied in a series of meningiomas the expression of the Toll‐like receptor 4 (TLR4), which apart from its major role as a key factor of the innate immune system, is believed to play a role in tumorigenesis. All meningiomas studied expressed TLR4 mRNA and protein at variable degree. Paclitaxel, a ligand of TLR4, exhibited a dose‐ and time‐dependent growth suppression in both monolayer and spheroid meningioma cell cultures. The knockdown of TLR4 with siRNA in meningioma cell cultures abrogated the inhibitory effect of paclitaxel. The suppressive action of paclitaxel on meningioma cell growth was enhanced in the presence of fluvastatin or the mitogen‐actvated protein kinase (ERK1/2) inhibitor PD98059. At least part of the growth suppressive effect was mediated by the induction of apoptosis in meningioma cells by paclitaxel alone or in combination with fluvastatin. In conclusion, our in vitro results suggest that paclitaxel alone or in combination with other inhibitors of cell growth (statins, MAPK inhibitors) could provide a potential tool for the treatment of TLR4 expressing meningiomas.

Expression of VEGF receptors in normal and adenomatous human pituitary

Vascular endothelial growth factor (VEGF) is a potent endothelial cell mitogen and exerts its action through different tyrosine kinase receptors: VEGFR-1 (Flt-1), VEGFR-2 (Flk-1/KDR) and neuropilin-1, a co-receptor of VEGFR-2, which are almost specifically expressed in endothelial cells and VEGFR-3 (Flt-4), which is detected mainly in lymphatic vessels. However, it has been shown that VEGF receptors may also be expressed by non-endothelial cells, especially by tumor cells. We have observed that pituitary adenoma cells produce variable amounts of VEGF and have studied the expression of VEGF receptors in normal human pituitary and pituitary adenomas by RT-PCR and immunohistochemistry. Protein expression of VEGFR-1, VEGFR-2, VEGFR-3 and neuropilin-1 was found in normal pituitary and in the pituitary adenomas studied so far. The expression was variable and no correlation between VEGFR-1 and VEGFR-2 expression and different parameters as tumor grade, vessels number and proliferation index (PI) was found. In normal pituitary, VEGFR-1 immunoreactivity was observed to co-localize with ACTH-, FSH-, GH- and LH- secreting cells, but not with endothelial cells, suggesting that VEGF may affect function of endocrine cell types by paracrine mechanisms. Moreover, VEGFR-1 immunostaining was observed mainly in pituitary adenoma cells. In contrast, VEGFR-2, VEGFR-3 and neuropilin-1 immunoreactivity was detected only in vascular formations, in both normal pituitary and pituitary adenomas and interestingly, we observed a correlation between neuropilin-1 expression and vessel count and proliferation index of the tumors analyzed. This finding could be consistent with the observation that neuropilin-1 acts as a co-receptor for VEGFR-2, enhancing the activity of VEGF. According on its receptors localization, VEGF seems to have a role both on tumor cells and on tumor vessels, even if not directly on vessel proliferation. Further studies are anyway necessary to better understand the possible connection between VEGF and its receptors and pituitary adenomas development.