Mariana Haedo

CELLULAR AND MOLECULAR SPECIFICITY OF PITUITARY GLAND PHYSIOLOGY

The anterior pituitary gland has the ability to respond to complex signals derived from central and peripheral systems. Perception of these signals and their integration are mediated by cell interactions and cross-talk of multiple signaling transduction pathways and transcriptional regulatory networks that cooperate for hormone secretion, cell plasticity, and ultimately specific pituitary responses that are essential for an appropriate physiological response. We discuss the physiopathological and molecular mechanisms related to this integrative regulatory system of the anterior pituitary gland and how it contributes to modulate the gland functions and impacts on body homeostasis.

Regulation of Pituitary Function by Cytokines

Research performed on the pituitary has proven that cytokines play an important role in maintaining pituitary physiology, affecting not only cell proliferation but also hormone secretion. The effects of cytokines can be autocrine or paracrine. This review gives an overview on the effects of the most studied cytokines in the pituitary. Special interest is focused on interleukin-6 (IL-6) because it has the distinctive characteristic of stimulating pituitary tumor cell growth, but has the opposite effect on normal pituitary cells. On the other hand, IL-6 is a cytokine of interest in the pituitary because recent work has shown that it promotes and maintains senescence in certain types of tumors. Given that the majority of pituitary adenomas are microadenomas and the fact that clinically inapparent pituitary tumors are quite common, senescence, perhaps mediated by IL-6, is an attractive mechanism for explaining the benign nature of pituitary tumors.

RSUME is implicated in HIF-1-induced VEGF-A production in pituitary tumour cells.

The recently cloned small RWD-domain containing protein RSUME was shown to increase protein levels of hypoxia-inducible factor-1α (HIF-1α). The latter is the oxygen-regulated subunit of HIF-1, the most important transcription factor of the cellular adaptive processes to hypoxic conditions. It is also a major regulator of vascular endothelial growth factor-A (VEGF-A), which is critically involved in the complex process of tumour neovascularisation. In this study, the expression and role of RSUME in pituitary tumours was studied. We found that RSUME mRNA was up-regulated in pituitary adenomas and significantly correlated with HIF-1α mRNA levels. Hypoxia (1% O(2)) or treatment with hypoxia-mimicking CoCl(2) enhanced RSUME and HIF-1α expression, induced translocation of HIF-1α to the nuclei and stimulated VEGF-A production both in pituitary tumour cell lines and primary human pituitary adenoma cell cultures. When RSUME expression was specifically down-regulated by siRNA, the CoCl(2)-induced increase VEGF-A secretion was strongly reduced which was shown to be a consequence of the RSUME knockdown-associated reduction of HIF-1α synthesis. Thus, RSUME plays an important role in initiating pituitary tumour neovascularisation through regulating HIF-1α levels and subsequent VEGF-A production and may therefore be critically involved in pituitary adenoma progression.

Pituitary tumors: Cell type-specific roles for BMP-4

BMP-4 plays a crucial role not only in the formation of the anterior pituitary during embryo development but also in the pathogenesis of pituitary tumors in adults. In tumor cells, BMP-4 promotes prolactin secretion and lactotroph cell proliferation through a Smad-estrogen receptor crosstalk but it inhibits ACTH production and cell proliferation of corticotrophs. In addition, BMP-4 increases GH secretion in rat pituitary tumor somatolactotroph GH3 cells and FSHbeta subunit gene transcription in the murine gonadotroph cell line, LbetaT2. Therefore, BMP-4 has a differential role on different types of pituitary tumors: it promotes pituitary prolactinoma while it inhibits corticotroph pathogenesis in Cushings disease. The modulation of BMP-4 also plays an important role in the therapeutic mechanism of action of bromocriptine, somatostatin analogs and retinoic acid.

Intrapituitary Expression and Regulation of the gp130 Cytokine Interleukin-6 and Its Implication in Pituitary Physiology and Pathophysiology

Interleukin (IL)‐6, a member of the gp130 cytokine family, is sometimes designated as an “endocrine” cytokine because of its strong regulatory influence on hormone production. Systemically acting IL‐6 derived from immune cells is a potent stimulator of the hypothalamus–pituitary–adrenal axis and therefore plays an important role in modulating immune–neuroendocrine interactions during inflammatory or infectious processes. However, IL‐6 is also produced within the anterior pituitary by so‐called folliculostellate (FS) cells and is also synthesized in and released by tumor cells in pituitary adenomas. Growth factors (e.g., transforming growth factor‐beta), neuropeptides (e.g., pituitary adenylate cyclase‐activating polypeptide), or hormones (e.g., glucocorticoids) regulate IL‐6 production both in FS and pituitary tumor cells. Interestingly, components of the innate immune system, such as toll‐like receptor 4 and nucleotide‐binding oligomerization domains (NODs), are expressed in FS and pituitary tumor cells. Therefore, cell‐wall components of bacteria (lipopolysaccharide, muramyl dipeptide, diamino pimelic acid) stimulate IL‐6 production in normal and tumoral pituitary. The intrinsic IL‐6 production by FS cells in normal anterior pituitary may participate in immune–neuroendocrine interactions during inflammatory processes. In pituitary adenomas, IL‐6 stimulates hormone secretion, tumor cell proliferation, and the production of angiogenic factors, such as vascular endothelial growth factor‐A, suggesting an important role of IL‐6 in the pathophysiology and progression of pituitary adenomas.

Effect of SOM230 (Pasireotide) on Corticotropic Cells: Action in Dogs with Cushing’s Disease

SOM230 (pasireotide) is a multiligand somatostatin (SRIF) analog able to bind to somatostatin receptor (SSTR) subtypes 1, 2, 3 and 5, and trigger antisecretory and antiproliferative signaling cascades. Canines have become in vivo models to test the pharmacological treatment of corticotropinomas because they frequently develop Cushing’s disease in a spontaneous manner, due to adrenocorticotropic hormone (ACTH)-producing pituitary adenomas. Different levels of expression of SSTR2 and SSTR5 have been shown in both mouse AtT20 cells and canine tumoral corticotropinoma cells. The objective of this study was to evaluate whether SOM230 controls both tumor cell growth and hormone synthesis, therefore controlling the disease. SOM230 was tested in dogs suffering from Cushing’s disease (10 animals were treated continuously during 6 months, and another 10 were treated with 3 cycles consisting of 2 months of treatment followed by a 2-month rest period). A significant decrease in ACTH, urinary cortisol creatinine ratio, adenoma size (magnetic nuclear resonance) and improvement of clinical signs were obtained, without side effects. AtT20 cells treated with SOM230 suppressed pro-opiomelanocortin (POMC) promoter activity through SSTR2, via the Gi α-subunit, and reduced Nur77/Nurr1 transcriptional activity. We conclude that SOM230, in addition to its well-described antisecretory effects, inhibits, as shown in AtT20 cells, ACTH synthesis at the POMC transcriptional level, an effect mediated mainly through SSTR2, and limits tumor growth. The controlled Cushing’s disease in the dogs that received the treatment indicates that SOM230 has a potential therapeutic use in humans suffering from Cushing’s disease.

Differential gene expression in models of pituitary prolactin-producing tumoral cells.

Although several genes and signalling pathways have been identified as important effectors in the development of pituitary tumours, our understanding of pituitary tumorigenesis remains incomplete and is the focus of much current research. Use of the mRNA differential display technique in prolactinomas from D2-receptor knockout mice and in stable GH3 cell line clones with enhanced tumorigenicity in vivo has led to the identification of two genes that are involved in the pathogenic process – BMP-4 and RSUME. Bone morphogenetic protein-4 (BMP-4) has been found to have a crucial role in prolactinoma development and also in signalling crosstalk with oestrogens. In contrast, BMP-4 has an inhibitory role in corticotrophinomas. RSUME (RWD-containing sumoylation enhancer) was identified from a transformed lactosomatotrophic cell line that had increased tumorigenic and angiogenic potential. Expression of RSUME was induced under hypoxic conditions and it has a potential role during vascularization. The differential expression and action of BMP-4 in prolactinomas and corticotrophinomas highlights the importance of studying a gene with contrasting actions in two cell lineages of the same organ in order to understand the pituitary transformation process. Both BMP-4 and RSUME may be interesting targets for inhibiting steps involved in pituitary tumorigenesis.

Autocrine IL-6 mediates pituitary tumor senescence

Cellular senescence is a stable proliferative arrest state. Pituitary adenomas are frequent and mostly benign, but the mechanism for this remains unknown. IL-6 is involved in pituitary tumor progression and is produced by the tumoral cells. In a cell autonomous fashion, IL-6 participates in oncogene-induced senescence in transduced human melanocytes. Here we prove that autocrine IL-6 participates in pituitary tumor senescence. Endogenous IL-6 inhibition in somatotroph MtT/S shRNA stable clones results in decreased SA-β-gal activity and p16INK4a but increased pRb, proliferation and invasion. Nude mice injected with IL-6 silenced clones develop tumors contrary to MtT/S wild type that do not, demonstrating that clones that escape senescence are capable of becoming tumorigenic. When endogenous IL-6 is silenced, cell cultures derived from positive SA-β-gal human tumor samples decrease the expression of the senescence marker. Our results establish that IL-6 contributes to maintain senescence by its autocrine action, providing a natural model of IL-6 mediated benign adenoma senescence.

Cytokines and Genes in Pituitary Tumorigenesis: RSUME Role in Cell Biology

Cytokines of the IL-6 or gp130 family regulate many cellular responses and play regulatory roles in numerous tissues, and are placed as auto-paracrine regulators of pituitary function acting in normal and tumoral anterior pituitary cells. Especially, IL-6 has a regulatory role in the hormone secretion and growth of the anterior pituitary and is involved in adenoma pathogenesis. Recently, IL-6 has been shown to mediate oncogene-induced senescence (OIS). IL-6 might participate in such a process in adenomas pituitary as well. From pituitary tumoral gp130 overexpressing cells, an unknown protein, RSUME, has been cloned. RSUME is induced by hypoxia in pituitary tumors and regulate pathways involved in angiogenic and tumorigenic processes (NF-kappaB/IkappaB and HIF-1alpha pathways). Thus, it could have an important role in the development of the pituitary tumors.

Novel medical therapies for pituitary tumors.

Despite considerable progress, there is still no medical treatment available for some kinds of pituitary tumors, in particular hormone inactive adenomas and corticotroph pituitary tumors. Surgical removal or at least debulking of the tumor is the only option to treat these kinds of tumors apart from rarely applied radiotherapy. Moreover, treatment resistance is present in a considerable proportion of patients bearing pituitary tumors, for which medical treatment regimens are already available (prolactinomas, somatotroph adenomas). Thus, novel or improved medical treatment strategies would be desirable. Here, we summarize preclinical and clinical findings about the hormone and growth-suppressive action of various drugs, which will probably lead to novel future medical treatment concepts for pituitary tumors.