Single-Cell Phenotypic Characterization of Human Pituitary GHomas and Non-Functioning Adenomas Based on Hormone Content and Calcium Responses to Hypothalamic Releasing Hormones.

Laura Senovilla,Daniel A De Luis,Lucã­A Nãºã±Ez,Enrique Romero,Carlos Villalobos,Josã© Marã­A De Campos,Javier Garcã­A-Sancho

doi:10.3389/fonc.2015.00124

Abstract

Human pituitary tumors are generally benign adenomas causing considerable morbidity due to excess hormone secretion, hypopituitarism, and other tumor mass effects. Pituitary tumors are highly heterogeneous and difficult to type, often containing mixed cell phenotypes. We have used calcium imaging followed by multiple immunocytochemistry to type growth hormone secreting (GHomas) and non-functioning pituitary adenomas (NFPAs). Individual cells were typed for stored hormones and calcium responses to classic hypothalamic releasing hormones (HRHs). We found that GHomas contained growth hormone cells either lacking responses to HRHs or responding to all four HRHs. However, most GHoma cells were polyhormonal cells responsive to both thyrotropin-releasing hormone (TRH) and GH-releasing hormone. NFPAs were also highly heterogeneous. Some of them contained ACTH cells lacking responses to HRHs or polyhormonal gonadotropes responsive to LHRH and TRH. However, most NFPAs were made of cells storing no hormone and responded only to TRH. These results may provide new insights on the ontogeny of GHomas and NFPAs.

Highlights

Pituitary adenomas are non-metastasizing neoplasms of adenohypophyseal cells occurring with an incidence extrapolated from autopsies larger than 20%
We have previously shown that a series of human pituitary adenomas including prolactinomas, pituitary adenomas associated to multiple endocrine neoplasia type I (MEN I) disease, pituitary tumors linked to Cushing’s disease, and a few non-functioning pituitary adenomas (NFPAs) contained cells storing more than one anterior pituitary (AP) hormone and/or showing responses to more than one hypothalamic releasing hormone (HRH) [17]
We aimed here at characterizing the phenotype of individual cells from human GHomas and additional NFPAs according to the hormones stored and functional responses to the four classic HRHs including thyrotropin-releasing hormone (TRH), GH-releasing hormone (GHRH), corticotrophin-releasing hormone (CRH), and gonadotropin-releasing hormone

Summary

Introduction

Pituitary adenomas are non-metastasizing neoplasms of adenohypophyseal cells occurring with an incidence extrapolated from autopsies larger than 20%. We have previously shown that a series of human pituitary adenomas including prolactinomas, pituitary adenomas associated to multiple endocrine neoplasia type I (MEN I) disease, pituitary tumors linked to Cushing’s disease, and a few NFPA contained cells storing more than one AP hormone (multihormonal cells) and/or showing responses to more than one hypothalamic releasing hormone (HRH) (multiresponsive cells) [17] For this end, we used fluorescence imaging of individual cells loaded with fura to record the rises in cytosolic Ca2+ concentration ([Ca2+]cyt) induced by specific HRHs added and removed sequentially. We aimed here at characterizing the phenotype of individual cells from human GHomas and additional NFPAs according to the hormones stored and functional (calcium) responses to the four classic HRHs including TRH, GHRH, corticotrophin-releasing hormone (CRH), and gonadotropin-releasing hormone For this end, we used a combination of multiple immunofluorescence and calcium imaging applied on the same individual cells reported previously [17, 18]

Materials and Methods

Results

Discussion