Protective effect of the dopamine D(3) receptor agonist (7-OH-PIPAT) against apoptosis in malignant peripheral nerve sheath tumor (MPNST) cells.
Emerging evidence indicates that the dopamine D(3) receptor (D(3)R) mediates protective roles both in neuronal and non-neuronal cell lines. In a previous study we proposed that neurofibromin, a large tumor suppressor protein encoded by the neurofibromatosis type 1 gene (NF1), may increase susceptibility to apoptosis after serum deprivation in malignant peripheral nerve sheath tumor (MPNST) cells, thus acting as a proapoptotic gene. In addition, it has been observed that D(3)Rs are functionally correlated to neurofibromin. In this study, we examined whether 7-OH-PIPAT, a potent dopamine D(3)R agonist, exerts an antiapoptotic role under the same culture conditions and then correlated this effect to changes in NF1 expression. Results showed that serum deprivation caused a significant reduction of cell viability (MTT assay) both after 24 and 48 h (p<0.001). Treatment with increasing concentrations of 7-OH-PIPAT (10(-9)-10(-5) M) induced a progressive increase in cell viability both after 24 and 48 h as compared to vehicle-treated cells, with significant changes at the highest concentrations tested (10(-6) and 10(-5) M). Consistently, at the latter two concentrations, a significant reduction in oligonucleosomes formation was observed, thus suggesting an antiapoptotic role of 7-OH-PIPAT. These results were confirmed by Hoechst 33254 nuclear staining. To investigate whether these effects were correlated to changes in NF1 transcript and protein expression, quantitative real-time PCR, Western blot and immunofluorescence analyses were performed. Results demonstrated that the upregulation of NF1 transcripts and protein levels induced by serum withdrawal were remarkably attenuated by 10(-6) and 10(-5) M agonist treatment within 24 h (p<0.01 and p<0.001, respectively), whereas similar effects were observed already at a lower concentration (10(-7) M) after 48 h treatment (p<0.001). In conclusion, these results suggest that D(3)R might mediate the protective response to serum deprivation in MPNST cells through the inhibition of NF1 gene expression, further underlying a subtle role of these receptors in MPNST development.
Castorina
A
A
Department of Anatomy, Diagnostic Pathology, Legal Medicine, Hygiene and Public Health, 95123 Catania, Italy.
Giunta
S
S
D'Agata
V
V
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection; D018491:Dopamine Agonists / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D018317:Nerve Sheath Neoplasms / Q000378:metabolism / Q000473:pathology*; D025542:Neurofibromin 1 / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D050637:Receptors, Dopamine D3 / Q000819:agonists* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044967:Serum / Q000378:metabolism; D013764:Tetrahydronaphthalenes / Q000494:pharmacology*; D013997:Time Factors
D018491:Dopamine Agonists; C493595:Drd3 protein, rat; D025542:Neurofibromin 1; D012333:RNA, Messenger; D050637:Receptors, Dopamine D3; D013764:Tetrahydronaphthalenes; C081289:7-hydroxy-2-(N-n-propyl-N-(3-iodo-2'-propenyl)-amino)tetralin