Objective: The aim of this study was to quantitatively estimate SSTR2 in clinically non-functioning pituitary adenomas (CNFAs) with use of RNA in-situ hybridization (ISH) and quantitative real-time RT-PCR and correlate the results of both methods.
Methods: A standard histological and immunohistochemical examination was performed on the resected pituitary tumour. Afterwards in-situ hybridization for somatostatine 2 receptor (SSTR2) RNA was performed manually using the RNAscope® 2.0 FFPE Assay (Advanced Cell Diagnostics, Inc., Hayward, CA, USA) on formalin fixed and paraffin embedded tissue sections. Small part of the same tumour resected during operation and stored in RNAlater was used for quantitative real-time RT-PCR.
Results: A 25 adenomas with SSTR2 mRNA expression in qRT-PCR were chosen for further evaluation with RNA ISH. SSTR2 mRNA was expressed in all adenomas from 1413148 680 copies/5 μl cDNA; the median of relative quantity (after normalization to housekeeping gene GUS) for SSTR2 was 111%. In contrast to qRT-PCR immunostaining was positive only in nine adenomas with the use of RNA ISH. Positive cases were subsequently semi-quantitatively assessed according to the manufacturers scoring guideline as follows: 1 adenoma with 1+, 5 with 2+ and 3 with 3+. No adenoma scored 4+ although high expression of SSTR2 mRNA was present. We did not find any correlation between data (Spearmans rank correlation coefficient 0.243).
Conclusion: Use of somatostatine analogues or dopastatins remains controversial in CNFAs. Verification of SSTR presence before use of drug treatment should be useful. Both RNA ISH and qRT-PCR have their pitfalls. In our case, we did show no correlation between these methods. RNA ISH is definitely less sensitive and specific. We think that interpretation of results for SSTR2 expression in RNA ISH and qRT-PCR should be very careful.
Project is supported by Ministry of Health Project No. NT/11344-4/2010.
27 Apr - 01 May 2013
European Society of Endocrinology