Differential peptide-dependent regulation of growth hormone (GH): A comparative analysis in pituitary cultures of reptiles, birds, and mammals.

Growth hormone (GH) is a pituitary protein that exerts pleiotropic roles in vertebrates. The mechanisms regulating GH synthesis and secretion are finely controlled by hypothalamic neuropeptides and other factors. These processes have been considerably studied in mammals but are still poorly understood in other groups. To better understand the pituitary GH regulation during vertebrate phylogeny, we compared the effects of incubating several peptides on cultures of ex-vivo pituitary fragments obtained from representative specimens of reptiles (iguana), birds (chicken) and mammals (rat). The peptides used were: growth hormone-releasing hormone (GHRH), thyrotropin-releasing hormone (TRH), pituitary adenylate cyclase-activating polypeptide (PACAP), ghrelin, gonadotropin-releasing hormone (GnRH), and somatostatin (SST). In rat pituitary cultures, GH secretion was stimulated by GHRH and TRH, while gh mRNA expression was increased by GHRH and PACAP. In the case of chicken pituitaries, GH release was promoted by GHRH, ghrelin, PACAP, and GnRH, although the latter two had a dual effect since at a shorter incubation time they decreased GH secretion; in turn, gh mRNA expression was significantly stimulated by TRH, PACAP, and GnRH. The most intense effects were observed in iguana pituitary cultures, where GH secretion was significantly augmented by GHRH, PACAP, TRH, ghrelin, and GnRH; while gh mRNA expression was stimulated by GHRH, TRH, and PACAP, but inhibited by ghrelin and SST. Also, in the three species, SST was able to block the GHRH-stimulated GH release. Furthermore, it was found that the expression of Pou1f1 mRNA was increased with greater potency by GHRH and PACAP in the iguana, than in chicken or rat pituitary cultures. Additionally, in-silico analysis of the gh gene promoter structures in the three species showed that the reptilian promoter has more Pit-1 consensus binding sites than their avian and mammalian counterparts. Taken together, results demonstrate that pituitary peptide-mediated GH regulatory mechanisms are differentially controlled along vertebrate evolution.

Host Cells Upregulate Phosphate Transporter PIT1 to Inhibit Ehrlichia chaffeensis Intracellular Growth.

Ehrlichia chaffeensis infects and proliferates inside monocytes or macrophages and causes human monocytic ehrlichiosis (HME), an emerging life-threatening tick-borne zoonosis. After internalization, E. chaffeensis resides in specialized membrane-bound inclusions, E. chaffeensis-containing vesicles (ECVs), to evade from host cell innate immune responses and obtain nutrients. However, mechanisms exploited by host cells to inhibit E. chaffeensis growth in ECVs are still largely unknown. Here we demonstrate that host cells recognize E. chaffeensis Ech_1067, a penicillin-binding protein, and then upregulate the expression of PIT1, which is a phosphate transporter and transports phosphate from ECVs to the cytosol to inhibit bacterial growth. We found that host cells upregulate the PIT1 expression upon E. chaffeensis infection using transcriptome sequencing, qRT-PCR and Western blotting, and PIT1 is localized on the ECV membrane in infected THP-1 cells using confocal microscopy. Silence of PIT1 using shRNA enhances E. chaffeensis intracellular growth. Finally, we found that E. chaffeensis Ech_1067 induces the upregulation of PIT1 expression through the MyD88-NF-κB pathway using recombinant protein for stimulation and siRNA for silence. Our findings deepen the understanding of the innate immune responses of host cells to inhibit bacterial intracellular growth and facilitate the development of new therapeutics for HME.

Homeostatic coordination of cellular phosphate uptake and efflux requires an organelle-based receptor for the inositol pyrophosphate IP8.

Phosphate (Pi) serves countless metabolic pathways and is involved in macromolecule synthesis, energy storage, cellular signaling, and bone maintenance. Herein, we describe the coordination of Pi uptake and efflux pathways to maintain mammalian cell Pi homeostasis. We discover that XPR1, the presumed Pi efflux transporter, separately supervises rates of Pi uptake. This direct, regulatory interplay arises from XPR1 being a binding partner for the Pi uptake transporter PiT1, involving a predicted transmembrane helix/extramembrane loop in XPR1, and its hitherto unknown localization in a subset of intracellular LAMP1-positive puncta (named "XLPVs"). A pharmacological mimic of Pi homeostatic challenge is sensed by the inositol pyrophosphate IP8, which functionalizes XPR1 to respond in a temporally hierarchal manner, initially adjusting the rate of Pi efflux, followed subsequently by independent modulation of PiT1 turnover to reset the rate of Pi uptake. These observations generate a unifying model of mammalian cellular Pi homeostasis, expanding opportunities for therapeutic intervention.

Concomitant Prediction of the Ki67 and PIT-1 Expression in Pituitary Adenoma Using Different Radiomics Models.

OBJECTIVES: To preoperatively predict the high expression of Ki67 and positive pituitary transcription factor 1 (PIT-1) simultaneously in pituitary adenoma (PA) using three different radiomics models. METHODS: A total of 247 patients with PA (training set: n = 198; test set: n = 49) were included in this retrospective study. The imaging features were extracted from preoperative contrast-enhanced T1WI (T1CE), T1-weighted imaging (T1WI), and T2-weighted imaging (T2WI). Feature selection was performed using Spearman's rank correlation coefficient and least absolute shrinkage and selection operator (LASSO). The classic machine learning (CML), deep learning (DL), and deep learning radiomics (DLR) models were constructed using logistic regression (LR), support vector machine (SVM), and multi-layer perceptron (MLP) algorithms. The area under the receiver operating characteristic (ROC) curve (AUC), sensitivity, specificity, accuracy, negative predictive value (NPV) and positive predictive value (PPV) were calculated for the training and test sets. In addition, combined with clinical characteristics, the best CML and the best DL models (SVM classifier), the DL radiomics nomogram (DLRN) was constructed to aid clinical decision-making. RESULTS: Seven CML features, 96 DL features, and 107 DLR features were selected to construct CML, DL and DLR models. Compared to CML and DL model, the DLR model had the best performance. The AUC, sensitivity, specificity, accuracy, NPV and PPV were 0.827, 0.792, 0.800, 0.796, 0.800 and 0.792 in the test set, respectively. CONCLUSIONS: Compared with CML and DL models, the DLR model shows the best performance in predicting the Ki67 and PIT-1 expression in PAs simultaneously.

Histopathology of growth hormone-secreting pituitary tumors: State of the art and new perspectives.

Somatotroph (GH) adenomas/PitNETs typically arise from adenohypophysis and are biochemically active, leading to acromegaly and gigantism. More rarely, they present with ectopic origin and do not present overt biochemical or clinical features (silent variants). Histopathological examination should consider the clinical and radiological background, and include multiple steps assessing tumor morphology, pituitary transcription factors (PTFs), hormone secretion, proliferation markers, granulation, and somatostatin receptors (STRs), aimed at depicting as better as possible tumor origin (in case of non-functioning and/or metastatic tumor), and clinical behavior, including response to treatment. GH-secreting tumors are part of the Pit-1 family tumors and can secrete GH only (pure somatotrophs) or co-secrete prolactin (mixed tumors; in this case, various histological subtypes have been identified). Each subtype presents unique radiological, biochemical, and clinical characteristic. Therefore, the integration of biochemical, clinical, radiological, and histopathological elements is fundamental for proper diagnosis and management of pituitary adenomas/PitNETs, to be performed in referral Centers. In more recent times, the importance of genetic and epigenetic evaluation in the characterization of pituitary tumors (i.e., early identification of aggressive variants) has been outlined by some large studies, with the intention of improving targeted treatments.

Pituitary neuroendocrine tumors with PIT1/SF1 co-expression show distinct clinicopathological and molecular features.

Pituitary neuroendocrine tumors (PitNETs) are classified according to cell lineage, which requires immunohistochemistry for adenohypophyseal hormones and the transcription factors (TFs) PIT1, SF1, and TPIT. According to the current WHO 2022 classification, PitNETs with co-expression of multiple TFs are termed "plurihormonal". Previously, PIT1/SF1 co-expression was prevailingly reported in PitNETs, which otherwise correspond to the somatotroph lineage. However, little is known about such tumors and the WHO classification has not recognized their significance. We compiled an in-house case series of 100 tumors, previously diagnosed as somatotroph PitNETs. Following TF staining, histopathological features associated with PIT1/SF1 co-expression were assessed. Integration of in-house and publicly available sample data allowed for a meta-analysis of SF1-associated clinicopathological and molecular features across a total of 270 somatotroph PitNETs. The majority (74%, 52/70) of our densely granulated somatotroph PitNETs (DGST) unequivocally co-expressed PIT1 and SF1 (DGST-PIT1/SF1). None (0%, 0/30) of our sparsely granulated somatotroph PitNETs (SGST) stained positive for SF1 (SGST-PIT1). Among DGST, PIT1/SF1 co-expression was significantly associated with scarce FSH/LH expression and fewer fibrous bodies compared to DGST-PIT1. Integrated molecular analyses including publicly available samples confirmed that DGST-PIT1/SF1, DGST-PIT1 and SGST-PIT1 represent distinct tumor subtypes. Clinicopathological meta-analyses indicated that DGST-PIT1 respond more favorably towards treatment with somatostatin analogs compared to DGST-PIT1/SF1, while both these subtypes show an overall less aggressive clinical course than SGST-PIT1. In this study, we spotlight that DGST with co-expression of PIT1 and SF1 represent a common, yet underrecognized, distinct PitNET subtype. Our study questions the rationale of generally classifying such tumors as "plurihormonal", and calls for a refinement of the WHO classification. We propose the term "somatogonadotroph PitNET".

SGK3 promotes vascular calcification via Pit-1 in chronic kidney disease.

Rationale: Vascular calcification (VC) is a life-threatening complication in patients with chronic kidney disease (CKD) caused mainly by hyperphosphatemia. However, the regulation of VC remains unclear despite extensive research. Although serum- and glucocorticoid-induced kinase 3 (SGK3) regulate the sodium-dependent phosphate cotransporters in the intestine and kidney, its effect on VC in CKD remains unknown. Additionally, type III sodium-dependent phosphate cotransporter-1 (Pit-1) plays a significant role in VC development induced by high phosphate in vascular smooth muscle cells (VSMCs). However, it remains unclear whether SGK3 regulates Pit-1 and how exactly SGK3 promotes VC in CKD via Pit-1 at the molecular level. Thus, we investigated the role of SGK3 in the certified outflow vein of arteriovenous fistulas (AVF) and aortas of uremic mice. Methods and Results: In our study, using uremic mice, we observed a significant upregulation of SGK3 and calcium deposition in certified outflow veins of the AVF and aortas, and the increase expression of SGK3 was positively correlated with calcium deposition in uremic aortas. In vitro, the downregulation of SGK3 reversed VSMCs calcification and phenotype switching induced by high phosphate. Mechanistically, SGK3 activation enhanced the mRNA transcription of Pit-1 through NF-κB, downregulated the ubiquitin-proteasome mediated degradation of Pit-1 via inhibiting the activity of neural precursor cells expressing developmentally downregulated protein 4 subtype 2 (Nedd4-2), an E3 ubiquitin ligase. Moreover, under high phosphate stimulation, the enhanced phosphate uptake induced by SGK3 activation was independent of the increased protein expression of Pit-1. Our co-immunoprecipitation and in vitro kinase assays confirmed that SGK3 interacts with Pit-1 through Thr468 in loop7, leading to enhanced phosphate uptake. Conclusion: Thus, it is justifiable to conclude that SGK3 promotes VC in CKD by enhancing the expression and activities of Pit-1, which indicate that SGK3 could be a therapeutic target for VC in CKD.

Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

Inorganic phosphate (Pi) is a critical determinant of calcification, and its concentration is regulated by alkaline phosphatase (ALP) and Pit1. ALP is a key regulator of osteogenic calcification and acts by modulating local inorganic phosphate (Pi) concentrations through hydrolyzing pyrophosphate in the extracellular matrix (ECM). Pit1, a sodium-dependent phosphate transporter, regulates calcification via facilitating phosphate uptake within the cells. To investigate whether zinc differentially regulates osteoblastic and vascular calcifications, we examined ALP activity and Pit1 in osteoblastic and vascular smooth muscle cells (VSMCs). Our findings demonstrate that calcification in osteoblastic MC3T3-E1 cells is decreased via diminished ALP action under zinc deficiency. In contrast, zinc-deficiency-induced calcification in VSMCs is independent of ALP action, as demonstrated by very weak ALP activity and expression in calcified VSMCs. In zinc-deficient A7r5 VSMC, P accumulation increased with increasing Na phosphate concentration (3-7 mM) but not with ß-GP treatment, which requires ALP activity to generate Pi. Ca deposition also increased with Na phosphate in a dose-dependent manner; in contrast, ß-GP did not affect Ca deposition. In osteoblastic cells, Pit1 expression was not affected by zinc treatments. In contrast, Pit1 expression is highly upregulated in A7r5 VSMC under zinc deficiency. Using phosphonoformic acid, a competitive inhibitor of Pit1, we showed that calcification is inhibited in both A7r5 and MC3T3-E1 cells, indicating a requirement for Pit1 in both calcifications. Moreover, the downregulation of VSMC markers under zinc deficiency was restored by blocking Pit1. Taken together, our results imply that zinc-deficiency-induced calcification in VSMC is independent of ALP action in contrast to osteoblastic calcification. Moreover, Pit1 expression in VSMCs is a target for zinc deficiency and may mediate the inhibition of VSMC marker expression under zinc deficiency.

The clinical presentation of PIT1 positive pituitary neuroendocrine tumor immunonegative for growth hormone, prolactin, and thyroid stimulating hormone with analysis of clinical and immunostaining dissociation.

BACKGROUND: PIT1 is a pituitary transcription factor that is associated with either growth hormone (GH), prolactin (PRL), or thyroid-stimulating hormone (TSH) production. However, PIT1-positive pituitary neuroendocrine tumors (PitNETs) are occasionally immunonegative for GH, PRL, and TSH. This paper describes the clinical presentation of PIT1 positive however immunonegative PitNETs. METHODS: We conducted a retrospective analysis, identifying 228 PIT1-positive PitNET patients between 2017 and 2022. Out of these, ten (4%) tested negative for GH, PRL, and TSH. Functioning PitNETs were defined as those causing hormonal excess symptoms or hormonal overproduction. RESULTS: As for 10 patients immunonegative for all three hormones however PIT1-positive, the mean ( ± standard deviation) age was 46 ± 13 years with 70% women. Six patients exhibited signs of excess GH or PRL, and three had visual problems. Additionally, one patient had secondary hypothyroidism and adrenal insufficiency resulting from the mass effect. All tumors were macroadenoma, with a median volume of 2.1 cm3 (range, 0.8-17.5 cm3). Gross total resection was attained in six patients by trans-sphenoidal surgery. Postoperatively, eight patients experienced clinical improvement: three in vision, two in amenorrhea, two in headache, and one in acromegaly symptoms. Biochemical improvement was observed in six patients, with all experiencing remission in hormonal excess and one showing improvement in secondary hypothyroidism. Stereotactic radiosurgery was performed in three patients. CONCLUSIONS: Patients with functioning PitNETs may exhibit PIT1 staining without GH, PRL, or TSH staining. Hormonally active tumors exist in this patient population; therefore, close endocrine follow-up is necessary despite the lack of staining for GH, PRL, and TSH.

Plurihormonal Pituitary Neuroendocrine Tumours - A Single Centre Experience.

Introduction. Plurihormonal pituitary neuroendocrine tumours (PitNET)/adenomas are pituitary neuroendocrine tumours composed of monomorphous cell populations expressing anterior pituitary transcription factors and/or hormones belonging to more than one cell lineage. Studies dedicated to plurihormonal tumours are rare and quite heterogenous with their results, bearing in mind changes in diagnostic criteria and inconsistent use of antibodies for anterior pituitary transcription factors in the diagnostic immunohistochemical panel. Material and Methods. We retrospectively analysed all patients surgically treated for PitNETs from 2016 to July 2022 in a tertiary healthcare institution. All tumours previously diagnosed PitNETs with the word "plurihormonal" were re-examined and potentially re-classified, according to 2022 WHO classification of endocrine tumours. Results. Among 721 patients surgically treated for PitNET in 5.5 years period, the diagnosis of plurihormonal PitNET was established in 11 tumours (1.3%). All tumours showed diffuse and intensive positivity for anterior pituitary transcription factors PIT1 and SF1. Clinically, all patients presented with acromegaly. Conclusions. Retrospective studies related to newly defined plurihormonal PitNETs with a reassessment of diagnoses are necessary due to their rarity and ambition to investigate their origin and biological behaviour. The fact that the majority of plurihormonal PitNETs are clinically presented with acromegaly and show simultaneous positivity to PIT1 and SF1 transcription factors deserve special attention and need for further research in larger cohorts of these exceptional tumours.

Clinical features of anti-pituitary-specific transcription factor-1 (PIT-1) hypophysitis: a new aspect of paraneoplastic autoimmune condition.

The pathogenesis of anti-pituitary-specific transcription factor-1 (PIT-1) hypophysitis was gradually revealed as cases emerged. Our comprehensive analysis, including all reported cases, identified a new instance of anti-PIT-1 hypophysitis postimmune checkpoint inhibitor therapy. All 9 patients exhibited extremely low growth hormone (GH), prolactin (PRL), and thyroid-stimulating hormone (TSH) levels; 2 had a slightly atrophic pituitary gland; 4 had thymoma, and 5 had malignant neoplasms of diffuse large B-cell lymphoma (DLBCL) and other origins. Patients with thymoma showed multiple autoimmune diseases. HLA-A*24:02 and/or A*02:06 were present in six and DR53 in 5 cases analyzed. High anti-PIT-1 antibody titers and ectopic PIT-1 expression in the cytosol and nucleus of the tumor tissues were observed in patients with thymoma or DLBCL, whereas it was exclusively observed in the nuclei of a bladder cancer patient. These findings provide new insights into the pathophysiology of paraneoplastic autoimmune hypophysitis.

Phosphate (Pi) Transporter PIT1 Induces Pi Starvation in Salmonella-Containing Vacuole in HeLa Cells.

Salmonella enterica serovar Typhimurium (S. Typhimurium), an important foodborne pathogen, causes diarrheal illness and gastrointestinal diseases. S. Typhimurium survives and replicates in phagocytic and non-phagocytic cells for acute or chronic infections. In these cells, S. Typhimurium resides within Salmonella-containing vacuoles (SCVs), in which the phosphate (Pi) concentration is low. S. Typhimurium senses low Pi and expresses virulence factors to modify host cells. However, the mechanism by which host cells reduce the Pi concentration in SCVs is not clear. In this study, we show that through the TLR4-MyD88-NF-κB signaling pathway, S. Typhimurium upregulates PIT1, which in turn transports Pi from SCVs into the cytosol and results in Pi starvation in SCVs. Immunofluorescence and western blotting analysis reveal that after the internalization of S. Typhimurium, PIT1 is located on SCV membranes. Silencing or overexpressing PIT1 inhibits or promotes Pi starvation, Salmonella pathogenicity island-2 (SPI-2) gene expression, and replication in SCVs. The S. Typhimurium ΔmsbB mutant or silenced TLR4-MyD88-NF-κB pathway suppresses the expression of the SPI-2 genes and promotes the fusion of SCVs with lysosomes. Our results illustrate that S. Typhimurium exploits the host innate immune responses as signals to promote intracellular replication, and they provide new insights for the development of broad-spectrum therapeutics to combat bacterial infections.

Somatostatin receptor expression and clinical outcome of multilineage pituitary tumours expressing PIT1 and SF1.

The application of transcription factor immunohistochemistry to pituitary neuroendocrine tumour (PitNET) assessment has allowed identification of tumours that do not conform to a single lineage. Multilineage pituitary transcription factor 1 (PIT1) and steroidogenic factor 1 (SF1) PitNETs are a rare and relatively newly described tumour subtype. These tumours express both transcription factors and may also express combinations of hormones corresponding to both lineages. Histological and clinical characteristics can vary, and overall clinical behaviour and prognosis is not known. We describe the clinical outcomes and somatostatin receptor status (SSTR) of a series of nine cases identified from our cohort of pituitary tumours at Westmead Hospital. Eight PitNETs (88.9%) expressed growth hormone and caused acromegaly at presentation. Of the seven macrotumours that caused acromegaly, one had cavernous sinus invasion. The Ki-67 labeling index score ranged from 0.6% to 3.6%. About 88% of tumours that secreted excess growth hormone exhibited strong immunostaining for SSTR 2 and all tumours displayed weak immunoreactivity for SSTR5. In 62.5% of patients with acromegaly, cure was achieved after surgical resection. Somatostatin receptor ligands resulted in clinical remission in cases where medical treatment was initiated. There was no new tumour recurrence or regrowth over an overall mean follow-up period of 62.5 months.

A Case of an Immature PIT1-Lineage Pituitary Neuroendocrine Tumor of the Nasopharynx.

Pituitary neuroendocrine tumors (PitNETs) located in the nasopharynx are a rare occurrence. This case report highlights a case of a 64-year-old female diagnosed with a PitNET incidentally found in her nasopharynx. The tumor was initially seen on MRI, excised, and analyzed with immunohistochemistry, ultimately confirming an immature pituitary-specific positive transcription factor 1 (PIT1)-positive-lineage pituitary neuroendocrine tumor. The tumor contained thyrotropes, somatotrophs, and lactotrophs expressing thyroid stimulating hormone, growth hormone, and prolactin, respectively. These tumors have the potential to exhibit aggressive behavior and can disrupt the surrounding tissue. Furthermore, they can be clinically silent or, conversely, secrete multiple hormones, causing hyperthyroidism, hyperprolactinemia, and acromegaly. For these reasons, they are deemed high risk. Treatment includes surgical excision with or without anti-hormone medications prior to surgery. Medications such as somatostatin analogs are used to decrease tumor size and reduce excessive hormone excretion.

Tumor immune microenvironment in pituitary neuroendocrine tumors (PitNETs): increased M2 macrophage infiltration and PD-L1 expression in PIT1-lineage subset.

PURPOSE: Tumor immune microenvironment in pituitary neuroendocrine tumors (PitNETs) and application of current immunotherapy for refractory PitNETs remains debated. We aim to evaluate the immune landscape in different lineages of PitNETs and determine the potential role of pituitary transcription factors in reshaping the tumor immune microenvironment (TIME), thus promoting the application of current immunotherapy for aggressive and metastatic PitNETs. METHODS: Immunocyte infiltration and expression patterns of immune checkpoint molecules in different lineages of PitNETs were estimated via in silico analysis and validated using an IHC validation cohort. The correlation between varying immune components with clinicopathological features was assessed in PIT1-lineage PitNETs. RESULTS: Transcriptome profiles from 210 PitNETs/ 8 normal pituitaries (NPs) and immunohistochemical validations of 77 PitNETs/6 NPs revealed a significant increase in M2-macrophage infiltration in PIT1-lineage PitNETs, compared with the TPIT-lineage, SF1-lineage subsets and NPs. While CD68 + macrophage, CD4 + T cells, and CD8 + T cells were not different among them. Increased M2-macrophage infiltration was associated with tumor volume (p < 0.0001, r = 0.57) in PIT1-lineage PitNETs. Meanwhile, differentially expressed immune checkpoint molecules (PD-L1, PD1, and CTLA-4) were screened and validated in IHC cohorts. The results showed that PD-L1 was highly expressed in PIT1-lineage subsets, and PD-L1 overexpression showed a positive correlation with tumor volume (p = 0.04, r = 0.29) and cavernous sinus invasion (p < 0.0001) in PIT1-lineage PitNETs. CONCLUSION: PIT1-lineage PitNETs exhibit a distinct immune profile with enrichment of M2 macrophage infiltration and PD-L1 expression, which may contribute to its clinical aggressiveness. Application of current immune checkpoint inhibitors and M2-targeted immunotherapy might be more beneficial to treat aggressive and metastatic PIT-lineage PitNETs.

Plurihormonal (TSH-GH-prolactin Secreting) PIT-1-Positive pituitary Macroadenoma/Pituitary Neuroendocrine Tumor (PitNET) Associated with Graves' Disease: A Case Report.

BACKGROUND: Differential diagnosis of primary and secondary hyperthyroidism can be challenging. Moreover, although extremely rare, the two conditions can coexist. CASE PRESENTATION: A 58-year-old woman presented with symptoms of thyrotoxicosis, gradual changes in face shape, headache and progressive enlargement of hands and feet in the last year. When she was 46 years old, she was diagnosed with Graves' disease, and treated with 4-year methimazole therapy. Since 2016, a progressive increase of free-T4 and free-T3 with normal-TSH and positive TrAb was noticed. RESULTS: At biochemical examination, fT3 was 5.3pg/ml (n. v. 2.5 - 3.9 pg/ml), fT4 was 20.6 pg/ml (n.v 6-12 pg/ml), IGF1 was 698 ng/ml (57 - 240 ng/ml*), GH (basally and after OGTT), and prolactin were significantly increased; while TSH was 1.8 (n.v. 0.35-4.0 mcUI/ml). A pituitary MRI demonstrated a large sellar tumor with suprasellar extension. The patient underwent endoscopic transsphenoidal surgery. Histological examination revealed a plurihormonal (GH-PRLTSH- secreting) PIT-1-positive pituitary adenoma/pituitary neuroendocrine tumor (PitNET). At 3- month follow-up, the pituitary function was normal, and no residual tumor was detected at the MRI. CONCLUSION: We report a rare case of Graves' disease coexisting with a plurihormonal PIT-1-positive pituitary adenoma/PitNET.

High sodium promotes the secretion and synthesis of PTH through PiT-1-IKKß pathway in parathyroid gland in vitro.

Parathyroid hormone (PTH) is secreted by the parathyroid glands (PTGs) and is an important hormone regulating mineral metabolism. Previous studies reported that high sodium diet will cause the increase in serum PTH, but the specific mechanism is unknown. Consequently, the present study aims to investigate the effects and mechanisms of high sodium on PTH synthesis and secretion from PTGs. We developed a tissue culture model using normal rat PTGs, discovered that sodium elicited and promoted concentration-dependent and time-dependent PTH secretion. Changes in sodium-associated transporters from PTGs incubated with high sodium were thoroughly examined. Increased expression of sodium-phosphate cotransporter Slc20a1 (also known as PiT-1) was observed. Further tests revealed that PiT-1 activated the NF-κB signaling pathway, resulting in increased IKKß phosphorylation, IKBα degradation, and increased p65 phosphorylation followed by nuclear entry, which led to increased PTH transcription. Meanwhile, IKKß phosphorylated SNAP23, promoting exocytosis and eventually led to increased PTH secretion. In conclusion, our findings indicate that PiT-1 plays an important role in the increased secretion and synthesis of PTH directly induced by high sodium under physiological conditions, and may provide a potential therapeutic target for secondary hyperparathyroidism (SHPT).

Multilineage Pituitary Neuroendocrine Tumors (PitNETs) Expressing PIT1 and SF1.

PitNETs are usually restricted in their cytodifferentiation to only one of 3 lineages dictated by expression of the pituitary transcription factors (TFs) PIT1, TPIT, or SF1. Tumors that show lineage infidelity and express multiple TFs are rare. We searched the pathology files of 4 institutions for PitNETs with coexpression of PIT1 and SF1. We identified 38 tumors in 21 women and 17 men, average age 53 (range 21-79) years. They represented 1.3 to 2.5% of PitNETs at each center. Acromegaly was the presentation in 26 patients; 2 had central hyperthyroidism associated with growth hormone (GH) excess and one had significantly elevated prolactin (PRL). The remainder had mass lesions with visual deficits, hypopituitarism, and/or headaches. Tumor size ranged from 0.9 to 5 cm; all 7 lesions smaller than 1 cm were associated with acromegaly. Larger lesions frequently invaded the cavernous sinuses. Four cases represented a second attempt at surgical resection. PIT1 was usually diffusely positive but 5 cases had variable (patchy or focal) staining. SF1 reactivity was variable in intensity but diffuse in all but 2 cases. GATA3 data, available in 14 cases, identified diffuse positivity in 5 and focal staining in 1. GH was expressed in all but 5 tumors, PRL and thyrotropin (TSH) were expressed in 14 and 13, respectively, follicle-stimulating hormone (FSH) in 11 of 18, and luteinizing hormone (LH) in 4 of 17. Keratin staining patterns were diffuse perinuclear/membranous in 27, variable perinuclear in 4, and negative in 3; scattered fibrous bodies were seen in 5 and diffuse fibrous bodies in 1. Ki67 labeling index ranged from < 1 to 7.9%. In 3 cases, these tumors represented one of multiple synchronous PitNETs; a separate corticotroph tumor was seen in 2 patients and one patient had 2 additional discrete lesions, a sparsely granulated lactotroph, and a pure gonadotroph tumor comprising a triple tumor. PitNETs expressing PIT1 and SF1 represent multilineage PitNETs. These rare tumors have variable clinical and morphological features, most often presenting as large tumors with GH excess and occasionally as one of multiple synchronous PitNETs of distinct lineages.

High expression of transcription factor POU2F1 confers improved survival on smokers with lung adenocarcinoma: a retrospective study of two cohorts.

Background: Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide and its most important risk factor is tobacco smoking. While smoking is associated with inferior outcome in NSCLC patients, smoking also correlates with a higher tumor mutational burden. In contrast to adenocarcinomas (ADC) of non-smokers, that frequently harbor targetable gain-of-function mutations, NSCLC smokers largely present with non-targetable loss-of-function mutations of genes associated with DNA-damage repair. The transcription factor Pit-1, Oct1/2, Unc-86 (POU) domain class 2 transcription factor 1 (POU2F1) is a widely expressed bipotential stabilizer of repressed and inducible transcriptional states and frequently deregulated in cancer. Methods: Via immunohistochemistry, we evaluated POU2F1 protein expression on a tissue micro array of 217 operable stage I-III NSCLC patients. Findings were reproduced in a gene expression database of 1144 NSCLC patients, filtered for POU2F1 mRNA expression. After retroviral overexpression of POU2F1 in A549 cells, we evaluated for clonogenic growth and proliferation. Additionally, CRISPR-Cas9 mediated POU2F1 knockdown in A549 cells was likewise analyzed. Results: High protein expression of POU2F1 in 217 NSCLC patients resulted in improved outcome of smokers with ADC [hazard ratio (HR) 0.30 (0.09-0.99), P=0.035]. Moreover, gene expression analysis confirmed favorable outcome of high POU2F1 mRNA expression in smokers with ADC [HR 0.41 (0.24-0.69), P<0.001]. Other than that, retrovirally induced overexpression of POU2F1 in A549 cells significantly reduced both, clonogenic growth as well as proliferation of NSCLC cells, whereas CRISPR-Cas9 mediated knockdown of the protein did not have any impact. Conclusions: Our data suggest that high expression of POU2F1 mediates a less aggressive cancer phenotype in smokers with ADC NSCLC. Pharmacological induction of genes and signaling pathways controlled by POU2F1 may provide novel avenues for future targeted NSCLC therapies in smokers.

Cytodifferentiation of pituitary tumors influences pathogenesis and cavernous sinus invasion.

OBJECTIVE: Pituitary tumors (PTs) continue to present unique challenges given their proximity to the cavernous sinus, whereby invasive behavior can limit the extent of resection and surgical outcome, especially in functional tumors. The aim of this study was to elucidate patterns of cavernoinvasive behavior by PT subtype. METHODS: A total of 169 consecutive first-time surgeries for PTs were analyzed; 45% of the tumors were functional. There were 64 pituitary transcription factor-1 (PIT-1)-expressing, 62 steroidogenic factor-1 (SF-1)-expressing, 38 T-box transcription factor (TPIT)-expressing, and 5 nonstaining PTs. The gold standard for cavernous sinus invasion (CSI) was based on histopathological examination of the cavernous sinus medial wall and intraoperative exploration. RESULTS: Cavernous sinus disease was present in 33% of patients. Of the Knosp grade 3 and 4 tumors, 12 (19%) expressed PIT-1, 7 (11%) expressed SF-1, 8 (21%) expressed TPIT, and 2 (40%), were nonstaining (p = 0.36). PIT-1 tumors had a significantly higher predilection for CSI: 53% versus 24% and 18% for TPIT and SF-1 tumors, respectively (OR 6.08, 95% CI 2.86-13.55; p < 0.001). Microscopic CSI-defined as Knosp grade 0-2 tumors with confirmed invasion-was present in 44% of PIT-1 tumors compared with 7% and 13% of TPIT and SF-1 tumors, respectively (OR 11.72, 95% CI 4.35-35.50; p < 0.001). Using the transcavernous approach to excise cavernous sinus disease, surgical biochemical remission rates for patients with acromegaly, prolactinoma, and Cushing disease were 88%, 87%, and 100%, respectively. The granule density of PIT-1 tumors and corticotroph functional status did not influence CSI. CONCLUSIONS: The likelihood of CSI differed by transcription factor expression; PIT-1-expressing tumors had a higher predilection for invading the cavernous sinus, particularly microscopically, compared with the other tumor subtypes. This elucidates a unique cavernoinvasive behavior absent in cells from other lineages. Innovative surgical techniques, however, can mitigate tumor behavior and achieve robust, reproducible biochemical remission and gross-total resection rates. These findings can have considerable implications on the surgical management and study of PT biology and behavior.