Supplementary Material for: MiR-148b, MiR-152/ALCAM Axis Regulates the Proliferation and Invasion of Pituitary Adenomas Cells

Background/Aims: Aberrant expression of miRNA has been found in many tumor tissues to regulate the tumorigenesis by binding to the 3`- untranslated region (3`-UTR) of the target genes. The aim of this study is to investigate the role of miR-148b, miR-152/ALCAM axis in human pituitary adenomas (PAs). Methods: First, we detected the expression level of miR-148b-3p and miR-152 in human PAs samples by using qRT-PCR. Then we studied the role of miR-148b-3p, miR-152 on human PAs cell proliferation, invasion and apoptosis by using MTS assay, Transwell invasion assay and Annexin V/PI Staining Test. To study the relationship between miR-148b-3p, miR-152 and activated leukocyte antigen molecule (ALCAM), we overexpressed miR-148-3p or miR-152 by transfecting specific mimics. Lucifearase reporter assay was then performed to confirm the target. Next, we studied the biological functions of ALCAM in human PAs cells. Finally, the role of miR-148b-3p, miR-152/ALCAM axis in PAs cells was studied. Results: The expression level of miR-148-3p and miR-152 in invasive PAs samples was lower than those in noninvasive samples. Overexpression of miR-148b-3p, miR-152 could repress proliferation and invasion, and promote apoptosis. Moreover, miR-148b-3p and miR-152 could repress activated leukocyte antigen molecule (ALCAM) expression. Knockdown of ALCAM could repress proliferation and invasion and promote apoptosis. By contrary, overexpression of ALCAM promoted proliferation and invasion. Further, the rescue experiments indicated that overexpression of ALCAM significantly restored the proliferation, apoptosis, and invasion influenced by miR-148b-3p and miR-152. Conclusions: Our study suggests that miR-148b-3p, miR-152 may serve as suppressors in PAs through downregulating ALCAM expression. miR-148b, miR-152/ ALCAM axis may be a new therapeutic target in the future.