The effects of Sonic hedgehog (Shh) signaling pathway activation on S-type neuroblastoma (NB) cell lines and its role in NB tumorigenesis were investigated. Immunohistochemistry was used to detect the expression of Shh pathway components-Patched1 (PTCH1) and Gli1 in 40 human primary NB samples. Western blotting and RT-PCR were used to examine the protein expression and mRNA levels of PTCH1 and Gli1 in three kinds of S-type NB cell lines (SK-N-AS, SK-N-SH and SHEP1), respectively. Exogenous Shh was administrated to activate Shh signaling pathway while cyclopamine was used as a selective antagonist of Shh pathway. S-type NB cell lines were treated with different concentrations of Shh or/and cyclopamine for different durations. Cell viability was measured by using MTT method. Apoptosis rate and cell cycle were assayed by flow cytometry. The xenograft experiments were used to evaluate the role of Shh pathway in tumor growth in immunodeficient mice. High-level expression of PTCH1 and Gli1 was detected in both NB samples and S-type NB cell lines. Cyclopamine decreased the survival rate of the three cell lines while Shh increased it, and the inhibition effects of cyclopamine could be partially reversed by shh pre-treatment. Cyclopamine induced the cell apoptosis and the cell cycle arrest in G(0)/G(1) phase, while Shh induced the reverse effects and could partially prevent effects of cyclopamine. Cyclopamine could also inhibit the growth of NB in vivo. Our studies revealed that activation of the Shh pathway is important for survival and proliferation of S-type NB cells in vivo and in vitro through affecting cell apoptosis and cell cycle, suggesting a new therapeutic approach to NB.

The effects of Sonic hedgehog (Shh) signaling pathway activation on S-type neuroblastoma (NB) cell lines and its role in NB tumorigenesis were investigated. Immunohistochemistry was used to detect the expression of Shh pathway components-- Patchedl (PTCH1) and Glil in 40 human primary NB samples. Western blotting and RT-PCR were used to examine the protein expression and mRNA levels of PTCH1 and Glil in three kinds of S-type NB cell lines (SK-N-AS, SK-N-SH and SHEPI), re-spectively. Exogenous Shh was administrated to activate Shh signaling pathway while cyclopamine was used as a selective antagonist of Shh pathway. S-type NB cell lines were treated with different concen-trations of Shh or/and cyclopamine for different durations. Cell viability was measured by using MTT method. Apoptosis rate and cell cycle were assayed by flow cytometry. The xenograft experiments were used to evaluate the role of Shh pathway in tumor growth in immunodeficient mice. High-level expres-sion of PTCH1 and Gill was detected in both NB samples and S-type NB cell lines. Cyclopamine de-creased the survival rate of the three cell lines while Shh increased it, and the inhibition effects of cyclopaminc could be partially reversed by shh pre-treatment. Cyclopamine induced the cell apoptosis and the cell cycle arrest in G0/G1 phase, while Shh induced the reverse effects and could partially pre-vent effects of cyclopamine. Cyclopamine could also inhibit the growth of NB in vivo. Our studies re-vealed that activation of the Shh pathway is important for survival and proliferation of S-type NB cells in vivo and in vitro through affecting cell apoptosis and cell cycle, suggesting a new therapeutic ap-proach to NB.

AIM: To find specific metabolic markers for women entering peri-menopausal period and patients with menopausal syndrome based on