Inhibitory effect of somatostatin receptor 5 activation on hormone secretion in pituitary prolactinoma through cyclic adenosine monophosphate pathway
Objective To explore the involvement of the cyclic adenosine monophosphate(cAMP)signaling pathway in the regulation of hormone secretion inhibition by somatostatin receptor 5(SSTR5)acti-vation in pituitary prolactinoma through in vitro experiments.Methods We utilized our developed SSTR5-overexpressing GH3 cell line as an in vitro model for pituitary prolactinoma.Cells were stimulated using the SSTR5 agonist BIM23052 combined with the cAMP activator Forskolin(5 μmol/L)or pertussis toxin(10 nmol/L),or with cAMP response element binding protein(CREB)overexpression.The activity of PRL-luc,cAMP response element(CRE)-Luc,Pit1-Luc,AP1-Luc,and estrogen response element(ERE)-Luc promoters was assessed using a luciferase reporter gene assay.The Renilla luciferase gene was used as an internal reference to standardize transfection efficiency.Statistical analysis between groups was performed using Student's t-test and one-way ANOVA.Results To verify the effectiveness of SSTR5 overexpression in GH3 cells,we monitored CRE reporter activity.It was found that CRE reporter gene ac-tivity in GH3SSTR5 cells was significantly reduced[(74.87±6.66)％]compared to GH3mock cells[(100.00±5.21)％,t=5.15,P＜0.05],indicating that SSTR5 overexpression inhibits CRE transcrip-tional activity.Pretreatment of GH3SSTR5 cells with various concentrations of BIM23052(0,0.1,1.0,10.0,100.0 nmol/L)combined with Forskolin(5 μmol/L)resulted in CRE reporter gene activities of(100.00±2.23)％,(17.79±2.30)％,(6.25±0.37)％,(25.27±2.99)％,and(44.60±3.25)％,respectively.This suggests that BIM23052 reduces Forskolin-induced CRE transcriptional activity,exhibi-ting a U-shaped dose-response curve,with the maximum inhibitory effect observed at 1.0 nmol/L(t=71.68,P＜0.01).Additionally,since the Pit1 promoter contains CRE elements,BIM23052 also reduced Forskolin-induced Pit1 promoter activity in a dose-dependent manner[(317.91±9.03)％,(278.11±4.94)％,(250.72±14.33)％,(176.21±8.88)％,(162.34±7.54)％,P＜0.01].However,BIM23052 and pertussis toxin treatment did not significantly inhibit other response elements on the Prl pro-moter,such as AP1 or ERE(P＞0.05).Overexpression of CREB showed that Prl promoter activity changed from[Ctrl group(100.00±12.01)％vs.BIM group(63.02±15.35)％,t=5.10,P＜0.01]in the blank transfection group to[Ctrl group(102.17±11.80)％vs.BIM group(106.27±14.62)％,t=0.58,P＞0.05]in the CREB overexpression group,indicating that BIM23052 treatment does not affect Prl promoter activity in CREB-overexpressing GH3 cells.This further confirms that inhibition of CREB ac-tivity is necessary for the SSTR5 agonist to inhibit PRL synthesis.Conclusion In pituitary prolactinoma,SSTR5 regulates prolactin synthesis by inhibiting the cAMP-CREB pathway.
ProlactinomasSomatostatin receptor 5Cyclic adenosine monophosphateCyclic adenosine monophosphate response element binding protein
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