ECE2023 Poster Presentations Pituitary and Neuroendocrinology (123 abstracts)
Somatostatin receptor type 2 (SSTR2) expression regulation by miR-375 in a murine pituitary corticotroph tumor cell model
Claudia Pivonello 1 , Roberta Patalano 2 , Tatiana Montò 2 , Mariarosaria Negri 2 , Feliciana Amatrudo 2 , Donatella Paola Provvisiero 2 , Chiara Simeoli 2 , Nicola Paola 2 , Angelica Larocca 2 , Erminio Massimo Crescenzo 2 , Annamaria Colao 2,3 & Rosario Pivonello 3,4
1Università Federico II di Napoli, Dipartimento di Sanità Pubblica e medicina Preventiva, Naples, Italy; 2Università Federico II di Napoli, Dipartimento di Medicina Clinica e Chirurgia, Naples, Italy; 3Università Federico II di Napoli, UNESCO Chair for Health Education and Sustainable Development, Naples, Italy; 2Università Federico II di Napoli, Dipartimento di Medicina Clinica e Chirurgia, Naples, Italy
Cushing’s disease (CD), caused by an adrenocorticotropic hormone (ACTH)-secreting pituitary tumor, is the most common form of hypercortisolism. Pituitary surgery is the first-line treatment of CD and medical treatment is an alternative second-line approach to control cortisol excess. Long-term exposure to glucocorticoids (GC) downregulates the expression of somatostatin receptor type 2 (SSTR2) but not type 5 (SSTR5) in human and mouse ACTH-secreting tumor cell cultures; thus octreotide, the somatostatin receptor ligand (SRL) with high affinity for SSTR2, failed to show any significant decrease neither in circulating ACTH and, consequently, cortisol levels, nor on tumor growth in CD patients. A recent study showed that in vitro dexamethasone treatment of a murine corticotroph tumor cell model induces an increase in miR-375 expression. The sequence analysis of SSTR2 revealed a binding site for miR-375. This finding supports the hypothesis that GC excess might downregulate SSTR2 expression through an epigenetic mechanism. The aim of the current study was to evaluate whether miR-375 is involved in the regulation of SSTR2 expression in a murine corticotroph tumor cell model AtT20. SSTR2 gene and miR-375 expression levels were evaluated by RT-qPCR in murine corticotroph (AtT20) and compared with somatotroph (GH3) cell lines. To evaluate the involvement of miR-375 in the regulation of SSTR2 protein expression, 24h miR-375 knocking-down was performed in AtT20 and SSTR2 protein levels were evaluated by western blot (WB) and immunofluorescence (IF). Moreover, miR-375 expression levels were also evaluated in 6 human corticotroph pituitary tumors and 2 normal pituitaries, whereas miR-375 circulating levels were evaluated in 21 CD patients and 19 healthy subjects by RT-qPCR. The results demonstrated an inverse SSTR2 and miR-375 expression in AtT20 and GH3 cells, with low levels of SSTR2 messengers and high levels of miR-375 in AtT20 and an opposite expression pattern in GH3 cells. After 24h of miR-375 knocking-down, induced by miR-375 inhibitor, SSTR2 protein expression, evaluated by WB and IF, was significantly increased in AtT20 (P=0,0310 and P=0,0154, respectively), compared to the control. Furthermore, miR-375 showed a higher expression in human corticotroph pituitary tumors than in normal pituitaries, as well as a higher and significant miR-375 circulating levels were observed in CD patients (P<0.0001) compared to healthy subjects. In conclusion, these data suggested that SSTR2 protein expression might be epigenetically downregulated by miR-375 in corticotroph pituitary tumors. Functional assays are mandatory to evaluate the role of SSTR2 in the condition of miR-375 knock-down.
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Endocrine Abstracts
ISSN 1470-3947 (print) | ISSN 1479-6848 (online)
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