Mesenchymal Stem Cells Induce an Immunosuppressive Microenvironment in Pituitary Tumors.

CONTEXT: The tumor microenvironment (TME) includes diverse cellular components such as mesenchymal stem cells (MSCs) and immune cells, among others. MSC have been isolated from different tumors and they favor tumor cell growth; however, their role in pituitary tumors (PTs) remains unknown. OBJECTIVE: Herein we report the presence of MSCs in 2 adrenocorticotropin (ACTH)-secreting PTs causing Cushing disease (MCU), 2 nonfunctioning adenomas of gonadotrope differentiation (MNF), and 2 nontumoral pituitary glands (MS). METHODS: We have analyzed the transcriptomic profiles by RNA sequencing and compared MSCs in terms of their immunosuppressive effects against lymphoid T-cell and macrophage populations by means of cocultures and flow cytometry. RESULTS: Our transcriptomic analysis revealed molecular differences between MSCs derived from nontumoral pituitaries and MSCs derived from PTs. Two distinct subpopulations of MSC emerged: one displaying immunosuppressive properties and the other with increased proproliferative capabilities, regardless of their origin. MSCs derived from ACTH- and nonfunctioning PTs, but not those derived from nontumoral glands, significantly inhibited the proliferation of activated T cells, favored the generation of regulatory T cells, and promoted M2 macrophage polarization. Such immunosuppressive effects were correlated with an upregulation of programmed death ligand 1 and intracellular expression of macrophage colony-stimulating factor (M-CSF) and interleukin-10. Importantly, MSC derived from ACTH-PTs showed a higher immunosuppressive potential than MSC isolated from nonfunctioning tumors. CONCLUSION: This study demonstrates the presence of at least 2 MSC subpopulations in the pituitary gland and suggests that immunosuppressive effects of MSCs may have important implications in PT growth.

Pituitary autoimmune disease: nuances in clinical presentation.

Pituitary autoimmune disease is considered an autoimmune organ-specific disorder, characterized by a pituitary infiltration of lymphocytes, macrophages, and plasma cells that could lead to loss of pituitary function. Hypophysitis may be secondary to systemic diseases or infections. Primary pituitary hypophysitis is classified into lymphocytic, granulomatous, xanthomatous, mixed forms (lymphogranulomatous, xanthogranulomatous), necrotizing and IgG4 plasmacytic, according to the histological findings. Concerning lymphocytic hypophysitis (LH), it is characterized by lymphocytic infiltration and can be subclassified according to the affected area on: lymphocytic adenohypophysitis, lymphocytic infundibulo-neurohypophysitis and lymphocytic panhypophysitis. LH had always been considered a rare disease. Nevertheless, with improved imaging techniques, especially magnetic resonance imaging (MRI), LH diagnosis has been increased. This disease usually affects young women during pregnancy or postpartum period with headache, visual impairment, ACTH deficiency and a homogenous sellar mass with thickening of pituitary stalk in MRI. Definitive diagnosis depends on histopathological evaluation; nevertheless, a presumptive diagnosis could be done in a typical case. As no specific autoantigen was identified in LH, there is no antipituitary antibody (APA) method available for helping diagnosis. However, APA used in some centers for research could support an autoimmune origin for some hypopituitarism previously named as idiopathic, confirming nuances in clinical presentation of pituitary autoimmune disease. Therapeutic approach should be based on the grade of suspicious and clinical manifestations of LH.

Integrative study of hypothalamus-pituitary-thyroid-immune system interaction: thyroid hormone-mediated modulation of lymphocyte activity through the protein kinase C signaling pathway.

Thyroid hormones play critical roles in differentiation, growth and metabolism, but their participation in immune system regulation has not been completely elucidated. Modulation of in vivo thyroid status was used to carry out an integrative analysis of the role of the hypothalamus-pituitary-thyroid (HPT) axis in T and B lymphocyte activity. The participation of the protein kinase C (PKC) signaling pathway and the release of some cytokines upon antigenic stimulation were analyzed. Lymphocytes from hyperthyroid mice displayed higher T-and B-cell mitogen-induced proliferation, and those from hypothyroid mice displayed lower T- and B-cell mitogen-induced proliferation, compared with euthyroid animals. Reversion of hypothyroid state by triiodothyronine (T3) administration recovered the proliferative responses. No differences were found in lymphoid subset balance. Both total PKC content and mitogen-induced PKC translocation were higher in T and B cells from hyperthyroid mice, and lower in cells from hypothyroid mice, compared with controls. Levels of thyroid-stimulating (TSH) and TSH-releasing (TRH) hormones were not directly related to lymphocyte proliferative responses. After immunization with sheep red blood cells (SRBCs) and re-stimulation, in vitro spleen cells from hyper- or hypothyroid mice showed, respectively, increased or decreased production of interleukin (IL)-2 and interferon (IFN)-gamma cytokines. Additionally, an increase in IL-6 and IFN-gamma levels was found in hyperthyroid cells after in vivo injection and in vitro re-stimulation with lipopolysaccharide (LPS). Our results show for the first time a thyroid hormone-mediated regulation of PKC content and of cytokine production in lymphocytes; this regulation could be involved in the altered responsiveness to mitogen-induced proliferation of T and B cells. The results also confirm the important role that these hormones play in regulating lymphocyte reactivity.

Expression of mRNA for interleukin-1beta, interleukin-6, tumor necrosis factor-alpha and macrophage migration inhibitory factor in HPA-axis tissues in Schistosoma mansoni-infected baboons (Papio cynocephalus).

Cytokines may regulate the function of the hypothalamic-pituitary-adrenal axis during schistosomiasis. This possibility was investigated in baboons experimentally infected with Schistosoma mansoni. Serum levels of corticotrophin-releasing hormone, adrenocorticotrophin, cortisol and dehydroepiandrosterone were confirmed to be decreased in infected baboons as previously shown. To explore if this effect is associated with specific expression of cytokines with endocrine activity, and are also associated with the pathology of the disease, Northern blots for interleukin-1beta, interleukin-6, tumor necrosis factor-alpha and macrophage migration inhibitory factor in hypothalamic-pituitary-adrenal axis tissues were performed. Infection induced interleukin-1beta gene expression in the hypothalamus, while interleukin-6 and migration inhibitory factor mRNAs were induced only in the pituitary and adrenal glands. Tumor necrosis factor-alpha gene expression was induced in the hypothalamus and the pituitary gland. Histopathological analysis of the hypothalamic-pituitary-adrenal axis tissues in infected and control baboons revealed no morphological differences between them. These results suggest that specific cytokines expressed in hypothalamic-pituitary-adrenal axis tissues could regulate hormone secretion during schistosomiasis.

Altered functional responses with preserved morphology of gonadotrophic cells in congenitally athymic mice.

Neonatal thymectomy or congenital absence of the thymus induces severe reproductive deficiencies in female mice, which are associated with reduced levels of circulating and pituitary gonadotropins. In contrast, the reproductive function is well preserved in nude males. It was therefore of interest to assess gonadotrophic cell morphology and function in congenitally athymic male mice. Circulating gonadotropins were measured under basal and stressful conditions, taking as a reference their haired counterparts. Adult normal (+/+), heterozygous nude (nu/+), and homozygous (nu/nu) CD-1 mice were subjected to 1-h immobilization stress. Serum levels of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were assessed by RIA at 0, 30, and 60 min poststress. Athymic animals showed significantly lower basal levels of serum LH and FSH than their heterozygous littermates. Immunohistochemical assessment of LH and FSH cell populations revealed a normal morphology and cell number in the athymic animals compared to their normal littermates. Immobilization stress induced a significant reduction in gonadotrophin levels, particularly LH, in normal mice but had only a weak effect in athymic animals. It is concluded that congenital athymia in the adult male mouse is associated with decreased basal levels of serum LH and FSH, in the presence of a normal gonadotroph number and morphology. The anomalous responses of athymic mice to stress do not appear to be due to primary hypopituitarism but, rather, to an altered modulation of pituitary hormone secretion. .

Androgen regulation of immunological and biological activities of pituitary follicle-stimulating hormone isoforms in male rats.

Follicle-stimulating hormone (FSH) is involved in the regulation and maintenance of gametogenesis. It exists in multiple molecular forms with different oligosaccharide structures which in turn are influenced by the hormonal milieu. Previous studies from our laboratory demonstrated that antiandrogen administration to immature male rats altered the biological activity and the distribution profile of pituitary FSH isoforms. The aim of this study was to examine possible modifications in pituitary FSH polymorphism throughout sexual development (10-, 32- and 75-day-old rats). In addition, the effect of androgen deprivation by castration (32-day-old rats) and its replacement with a nonaromatizable androgen - dihydrotestosterone - on pituitary FSH polymorphism was determined. Concanavalin A affinity chromatography was used to isolate groups of FSH isoforms according to their carbohydrate inner structure. Radioimmunoassay and Sertoli cell bioassay were used to evaluate FSH immuno- and bioactivities. Androgen rise in serum was accompanied by a marked increase in pituitary bio- and immuno-FSH content in 32- and 75-day-old rats. However, FSH pituitary content did not vary despite the significant increment observed in serum FSH levels after castration and decrease to control levels after androgen replacement. The distribution profile of immuno- and bioactive FSH changed throughout sexual maturation. The proportion of pituitary FSH isoforms bearing complex oligosaccharide structures (triantennary, bisecting, complete and truncated biantennary) increased with age, with a concomitant decrease in the proportion of isoforms bearing incomplete carbohydrate chains. The distribution profile observed in castrated 32-day-old rats was similar to that determined in 10-day-old animals. Androgen replacement restored the distribution profile to normal. These results suggest that androgens regulate the incorporation of sugar residues to the carbohydrate chains of pituitary FSH favoring the biosynthesis of complex-type oligosaccharide structures.

Influence of the pituitary gland on the immune response in young rats.

The response of hypophysectomized (HYPOX) and sham-operated (S-HYPOX) female and male Wistar young rats (8 weeks old) to antigenic stimulation was compared. Humoral antigenic responses against hemocyanin were measured by ELISA. [3H]thymidine incorporation into cultured spleen cells was used to determine proliferative response to concanavalin A (ConA) or antigenic stimulation. Anti-hemocyanin serum titers in the HYPOX animals was about half of that observed in control S-HYPOX rats. Similarly, the cellular proliferative response was significantly decreased in HYPOX animals when compared to S-HYPOX rats; the blastogenic response to hemocyanin in UC rats (which did not receive the antigen injection) was close to zero. S-HYPOX control rats responded to direct ConA stimulation as UC controls. Body weight and the weight of pituitary target organs (adrenal, thyroid, ovary and testes) was about 1/4 of that of controls. Hypophysectomy also resulted in a striking reduction in spleen weight. These results indicate that the pituitary gland is involved in cellular and humoral immune regulation in young rats.

Influence of the pituitary gland on the immune response in young rats

The response of hypophysectomized (HYPOX) and sham-operated (S-HYPOX) female and male Wistar young rats (8 weeks old) to antigenic stimulation was compared. Humoral antigenic responses against hemocyanin were measured by ELISA. [3H]thyimidine incorporation into cultured splen cells was used to determine proliferative response to concanavalin A (ConA) or antigenic stimulation. Anti-hemocyanin serum titers in the HYPOX animals was about half of that observed in control S-HYPOX rats. Similarly, the cellular proliferative response was significantly decreased in HYPOX animals when compared to S-HYPOX rats; the blastogenic response to hemocyanin in UC rats (which did not receive the antigen injection) was close to zero. S-HYPOX control rats responded to direct ConA stimulation as UC controls. Body weight and the weight of pituitary target organs (adrenal, thyroid, ovary and testes) was about 1/4 of that of controls. Hypophysectomy also resulted in a striking reduction in spleen weight. These results indicate that the pituitary gland is involved in cellular and humoral immune regulation in young rats. (AU)

Influence of the pituitary gland on the immune response in young rats

The response of hypophysectomized (HYPOX) and sham-operated (S-HYPOX) female and male Wistar young rats (8 weeks old) to antigenic stimulation was compared. Humoral antigenic responses against hemocyanin were measured by ELISA. [3H]thyimidine incorporation into cultured splen cells was used to determine proliferative response to concanavalin A (ConA) or antigenic stimulation. Anti-hemocyanin serum titers in the HYPOX animals was about half of that observed in control S-HYPOX rats. Similarly, the cellular proliferative response was significantly decreased in HYPOX animals when compared to S-HYPOX rats; the blastogenic response to hemocyanin in UC rats (which did not receive the antigen injection) was close to zero. S-HYPOX control rats responded to direct ConA stimulation as UC controls. Body weight and the weight of pituitary target organs (adrenal, thyroid, ovary and testes) was about 1/4 of that of controls. Hypophysectomy also resulted in a striking reduction in spleen weight. These results indicate that the pituitary gland is involved in cellular and humoral immune regulation in young rats.

Valoración del eje hipófisis-hipotálamo-adrenal en pacientes con corticoterapia crónica a dosis supresivas / Evaluation of the hypotha lamus-pituitary-adrenal axis in patients with chronic corticotherapy in suppresive dosis

Se efectuó la curva de cortisol en 32 individuos sanos con edad promedio de 38.7 años, siendo la cifra matutina de las ocho horas de 10 mg/100ml o mayor. Posteriormente, se realizó el mismo estudio a 42 enfermos, edad promedio 41.4 años, que estaban bajo corticoterapia mayor de 10 mg diarios de prednisona durante as de tres meses en forma ininterrunpida y la cual se suspendió 48 h previas al estudio. Se encontró que 27 de ellos (64.2 por ciento) tenían cifras de cortisol matutino menores a las del grupo testigo y los enfermos de artritis reumatoide eran de los de enfermedad sistémica podría ser un factor más en la supresión del eje H-H-A de los pacientes bajo dosis mayor de 10 mg de prednisona en duración mayor de tres meses continuos

Cytological differences in the localization of glucocorticoid receptor-like immunoreactivity in the normal and transplanted pituitary pars intermedia.

Glucocorticoid receptor-like immunoreactivity (GCRI) was found in the normal pituitary pars intermedia (PI) when immunohistochemistry was used. Since in previous studies we described two kinds of cells in the denervated (grafted) PI, i.e., "light cells" (overactive cells which do not contain detectable melanocyte-stimulating hormone) and "dark cells" (hypoactive cells which contain the hormone), it was decided to investigate whether different patterns of distribution of the receptors could be detected in the grafted gland when compared with the intact PI. Intact glands showed the receptors located in the nucleus. In transplanted glands, it was observed that light cells showed receptors in both the nuclei and the cytoplasm; on the other hand, dark cells displayed them in the nuclei only, as is the case in all cells of the normal PI. We had previously interpreted dark cells as dopamine-indifferent, whereas light cells were considered dopamine-sensitive. The changes in the distribution of GCR after denervation by grafting, which only affected the light cells, support the view of other authors that GCR of the pars intermedia are under the influence of dopamine and reinforce our opinion that dark cells are dopamine-indifferent.

Imuno-histoquímica hipofisária: metodologia e aplicaçäo / Immunohistochemistry of the pituitary gland: methodology and use

Os autores estudaram a aplicaçäo do método da peroxidase-antiperoxidase para detectar hormônio de crescimento (GH), prolactina (PRL) e corticotrofina (ACTH) em cortes de tecidos normais e de pituitária adenomatosa utilizando anticorpos de radioimunoensaio (RIA). Quatro glândulas pituitárias normais obtidas em autópsia e 21 adenomas pituitárias normais obtidos durante cirurgia de pacientes com acromegalia (16), hiperprolactinemia (2) e doença de Cushing (3) foram estudados. Anti-soros obtidos de carneiros imunizados com preparaçöes altamente purificadas de GH humano, prolactina ovina e ACTH porcino. Reatividades cruzadas de anti-GH com PRL, anti-PRL com GH e anti-ACTH 1-39 com ACTH 1-24 e ACTH 18-39 no RIA foram: 0,6%, 0,03%, 3,1% e 5,7% respectivamente. Detecçäo de cada hormônio foi bem sucedida em todos os cortes estudados de tecido pituitário normal. Melhores colaboraçöes foram obtidas usando diluiçöes primárias do anti-soro mais baixas que aquela rotineiramente empregadas em RIA. Anti-GH mostrou näo ter reatividade cruzada significante com PRL em cortes de tecido pituitário. Anti-PRL, contudo, mostrou alguma reatividade cruzada com GH, mas isto pôde ser evitado por absorçäo prévia do anti-soro com GH. Nenhuma diferença na coloraçäo pôde ser detectada fosse usado ou näo o anti-ACTH depois da imuno-absorçäo com ACTH 1-24. Todos os adenomas pituitários de pacientes acromegálicos mostraram células imunorreativas para GH, porém poucos adenomas tiveram todas ou mesmo a maioria das células positivas para GH. Todos os adenomas obtidos de pacientes com hiperprolactinemia e doença de Cushing coraram-se com anti-soro contra PRL e ACTH, respectivamente. Estes resultados validam o uso dos anti-soros mencionados na imuno-histoquímica da pituitária