The effect of miR-23b-3p on regulating GH by targeting POU1F1 in Yanbian yellow cattle.

This study aimed to evaluate the effect of miR-23b-3p on growth hormone (GH) in pituitary cells of Yanbian yellow cattle. The mRNA and protein levels of GH and miR-23b-3p target genes were measured by real time fluorescence quantitative PCR (qPCR) and Western blot, respectively. The target relationship of miR-23b-3p was validated by double luciferase reporter gene system. The results showed that GH mRNA and protein levels in pituitary cells of Yanbian yellow cattle were significantly lower in the miR-23b-3p-mi group than in the NC group (P<0.01), while GH mRNA and protein levels were higher in the miR-23b-3p-in group than in the iNC group (P<0.05). The result of bioinformatics analysis and double luciferase reporter gene system validation proved that miR-23b-3p targeted 3'UTR of pituitary specific transcription factor 1 (POU1F1). POU1F1 mRNA and protein levels were lower miR-23b-3p-mi group than in the NC group (P<0.01), while POU1F1 mRNA and protein levels were higher in the miR-23b-3p-in group than in the iNC group (P<0.01). These results demonstrated that miR-23b-3p could regulate GH expression in pituitary cells by regulating POU1F1 gene.

Familial idiopathic basal ganglia calcification with a heterozygous missense variant (c.902C>T/p.P307L) in SLC20A2 showing widespread cerebrovascular lesions.

We describe a postmortem case of familial idiopathic basal ganglia calcification (FIBGC) in a 72-year-old Japanese man. The patient showed progressive cognitive impairment with a seven-year clinical course and calcification of the basal ganglia, thalami, and cerebellar dentate nuclei. A novel heterozygous missense variant in SLC20A2 (c.920C>T/p.P307L), a type III sodium-dependent phosphate transporter (PiT-2), was subsequently identified, in addition to typical neuropathological findings of FIBGC, such as capillary calcification of the occipital gray matter, confluent calcification of the basal ganglia and cerebellar white matter, widespread occurrence of vasculopathic changes, cerebrovascular lesions, and vascular smooth muscle cell depletion. Immunohistochemistry for PiT-2 protein revealed no apparent staining in endothelial cells in the basal ganglia and insular cortex; however, the immunoreactivity in endothelial cells of the cerebellum was preserved. Moreover, Western blot analysis identified preserved PiT-2 immunoreactivity signals in the frontal cortex and cerebellum. The variant identified in the present patient could be associated with development of FIBGC and is known to be located at the large intracytoplasmic part of the PiT-2 protein, which has potential phosphorylation sites with importance in the regulation of inorganic phosphate transport activity. The present case is an important example to prove that FIGBC could stem from a missense variant in the large intracytoplasmic loop of the PiT-2 protein. Abnormal clearance of inorganic phosphate in the brain could be related to the development of vascular smooth muscle damage, the formation of cerebrovascular lesions, and subsequent brain calcification in patients with FIBGC with SLC20A2 variants.

Uraemic extracellular vesicles augment osteogenic transdifferentiation of vascular smooth muscle cells via enhanced AKT signalling and PiT-1 expression.

Extracellular vesicles (EV) function as messengers between endothelial cells (EC) and vascular smooth muscle cells (VSMC). Since chronic kidney disease (CKD) increases the risk for vascular calcifications, we investigated whether EV derived from uraemic milieu-stimulated EC and derived from uraemic rats impact the osteogenic transdifferentiation/calcification of VSMC. For that purpose, human EC were treated with urea and indoxyl sulphate or left untreated. Experimental uraemia in rats was induced by adenine feeding. 'Uraemic' and control EV (EVUR ; EVCTRL ) were isolated from supernatants and plasma by using an exosome isolation reagent. Rat VSMC were treated with a pro-calcifying medium (CM) with or without EV supplementation. Gene expressions, miRNA contents and protein expressions were determined by qPCR and Western blots, respectively. Calcifications were determined by colorimetric assays. Delivery of miRNA inhibitors/mimics to EV and siRNA to VSMC was achieved via transfection. EVCTRL and EVUR differed in size and miRNA contents. Contrary to EVCTRL , EC- and plasma-derived EVUR significantly increased the pro-calcifying effects of CM, including altered gene expressions of osterix, runx2, osteocalcin and SM22α. Further, EVUR enhanced the protein expression of the phosphate transporter PiT-1 in VSMC and induced a phosphorylation of AKT and ERK. Knock down of PiT-1 and individual inhibition of AKT and ERK signalling in VSMC blocked the pro-calcifying effects of EVUR . Similar effects were achieved by inhibition of miR-221/-222 and mimicking of miR-143/-145 in EVUR . In conclusion, EVUR might represent an additional puzzle piece of the complex pathophysiology of vascular calcifications in CKD.

Protective effects of spironolactone on vascular calcification in chronic kidney disease.

BACKGROUND: Vascular calcification is common in chronic kidney disease (CKD) and associated with increased cardiovascular mortality. Aldosterone has been implicated as an augmenting factor in the progression of vascular calcification. The present study further explored putative beneficial effects of aldosterone inhibition by the mineralocorticoid receptor antagonist spironolactone on vascular calcification in CKD. METHODS: Serum calcification propensity was determined in serum samples from the MiREnDa trial, a prospective, randomized controlled clinical trial to investigate efficacy and safety of spironolactone in maintenance hemodialysis patients. Experiments were conducted in mice with subtotal nephrectomy and cholecalciferol treatment, and in calcifying primary human aortic smooth muscle cells (HAoSMCs). RESULTS: Serum calcification propensity was improved by spironolactone treatment in patients on hemodialysis from the MiREnDa trial. In mouse models and HAoSMCs, spironolactone treatment ameliorated vascular calcification and expression of osteogenic markers. CONCLUSIONS: These observations support a putative benefit of spironolactone treatment in CKD-associated vascular calcification. Further research is required to investigate possible improvements in cardiovascular outcomes by spironolactone and whether the benefits outweigh the risks in patients with CKD.

Evidence of an intestinal phosphate transporter alternative to type IIb sodium-dependent phosphate transporter in rats with chronic kidney disease.

BACKGROUND: Phosphate is absorbed in the small intestine via passive flow and active transport.NaPi-IIb, a type II sodium-dependent phosphate transporter, is considered to mediate active phosphate transport in rodents. To study the regulation of intestinal phosphate transport in chronic kidney disease (CKD), we analyzed the expression levels of NaPi-IIb, pituitary-specific transcription factor 1 (PiT-1) and PiT-2 and the kinetics of intestinal phosphate transport using two CKD models. METHODS: CKD was induced in rats via adenine orThy1 antibody injection. Phosphate uptake by intestinal brush border membrane vesicles (BBMV) and the messenger RNA (mRNA) expression of NaPi-IIb, PiT-1 and PiT-2 were analyzed. The protein expression level of NaPi-IIb was measured by mass spectrometry (e.g. liquid chromatography tandem mass spectrometry). RESULTS: In normal rats, phosphate uptake into BBMV consisted of a single saturable component and its Michaelis constant (Km) was comparable to that of NaPi-IIb. The maximum velocity (Vmax) correlated with mRNA and protein levels of NaPi-IIb. In the CKD models, intestinal phosphate uptake consisted of two saturable components. The Vmax of the higher-affinity transport, which is thought to be responsible for NaPi-IIb, significantly decreased and the decrease correlated with reduced NaPi-IIb expression. The Km of the lower-affinity transport was comparable to that of PiT-1 and -2. PiT-1 mRNA expression was much higher than that of PiT-2, suggesting that PiT-1 was mostly responsible for phosphate transport. CONCLUSIONS: This study suggests that the contribution of NaPi-IIb to intestinal phosphate absorption dramatically decreases in rats with CKD and that a low-affinity alternative to NaPi-IIb, in particular PiT-1, is upregulated in a compensatory manner in CKD.

POU1F1 transcription factor promotes breast cancer metastasis via recruitment and polarization of macrophages.

Cancer progression requires cells surrounding tumors be reeducated and activated to support tumor growth. Oncogenic signals from malignant cells directly influence stromal composition and activation, but the factors mediating this communication are still not well understood. We have previously shown that the transcription factor POU class 1 homeobox 1 (POU1F1), also known as Pit-1, induces profound changes on neoplastic cell-autonomous processes favoring metastasis in human breast cancer. Here we describe for the first time Pit-1-mediated paracrine actions on macrophages in the tumor microenvironment by using cell lines in vitro, zebrafish and mouse models in vivo, and samples from human breast cancer patients. Through the release of CXCL12, Pit-1 in tumor cells was found to mediate the recruitment and polarization of macrophages into tumor-associated macrophages (TAMs). In turn, TAMs collaborated with tumor cells to increase tumor growth, angiogenesis, extravasation and metastasis to lung. Our data reveal a new mechanism of cooperation between tumor cells and macrophages favoring metastasis and poor clinical outcome in human breast cancer, which suggests that Pit-1 and CXCL12 should be further studied as potential prognostic and therapeutic indicators. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Required enhancer-matrin-3 network interactions for a homeodomain transcription program.

Homeodomain proteins, described 30 years ago, exert essential roles in development as regulators of target gene expression; however, the molecular mechanisms underlying transcriptional activity of homeodomain factors remain poorly understood. Here investigation of a developmentally required POU-homeodomain transcription factor, Pit1 (also known as Pou1f1), has revealed that, unexpectedly, binding of Pit1-occupied enhancers to a nuclear matrin-3-rich network/architecture is a key event in effective activation of the Pit1-regulated enhancer/coding gene transcriptional program. Pit1 association with Satb1 (ref. 8) and ß-catenin is required for this tethering event. A naturally occurring, dominant negative, point mutation in human PIT1(R271W), causing combined pituitary hormone deficiency, results in loss of Pit1 association with ß-catenin and Satb1 and therefore the matrin-3-rich network, blocking Pit1-dependent enhancer/coding target gene activation. This defective activation can be rescued by artificial tethering of the mutant R271W Pit1 protein to the matrin-3 network, bypassing the pre-requisite association with ß-catenin and Satb1 otherwise required. The matrin-3 network-tethered R271W Pit1 mutant, but not the untethered protein, restores Pit1-dependent activation of the enhancers and recruitment of co-activators, exemplified by p300, causing both enhancer RNA transcription and target gene activation. These studies have thus revealed an unanticipated homeodomain factor/ß-catenin/Satb1-dependent localization of target gene regulatory enhancer regions to a subnuclear architectural structure that serves as an underlying mechanism by which an enhancer-bound homeodomain factor effectively activates developmental gene transcriptional programs.

Biochemical and structural characterization of a novel cooperative binding mode by Pit-1 with CATT repeats in the macrophage migration inhibitory factor promoter.

Overexpression of the proinflammatory cytokine macrophage migration inhibitory factor (MIF) is linked to a number of autoimmune diseases and cancer. MIF production has been correlated to the number of CATT repeats in a microsatellite region upstream of the MIF gene. We have characterized the interaction of pituitary-specific positive transcription factor 1 (Pit-1) with a portion of the MIF promoter region flanking a microsatellite polymorphism (-794 CATT5-8). Using fluorescence anisotropy, we quantified tight complex formation between Pit-1 and an oligonucleotide consisting of eight consecutive CATT repeats (8xCATT) with an apparent Kd of 35 nM. Using competition experiments we found a 23 base pair oligonucleotide with 4xCATT repeats to be the minimum DNA sequence necessary for high affinity interaction with Pit-1. The stoichiometry of the Pit-1 DNA interaction was determined to be 2:1 and binding is cooperative in nature. We subsequently structurally characterized the complex and discovered a completely novel binding mode for Pit-1 in contrast to previously described Pit-1 complex structures. The affinity of Pit-1 for the CATT target sequence was found to be highly dependent on cooperativity. This work lays the groundwork for understanding transcriptional regulation of MIF and pursuing Pit-1 as a therapeutic target to treat MIF-mediated inflammatory disorders.

MSH6/2 and PD-L1 Expressions Are Associated with Tumor Growth and Invasiveness in Silent Pituitary Adenoma Subtypes.

Mismatch repair genes mutS homologs 6/2 (MSH6/2) expressions are involved in tumor growth and programmed cell death 1 ligand 1 (PD-L1) expression in tumor immunity, but the direct association with pituitary adenomas (PAs) is not well understood. We aimed to clarify the effects of MSH6/2 and PD-L1 expression on tumor proliferation and invasiveness in nonfunctioning (NF) PAs. We performed immunohistochemistry to classify the NFPAs into gonadotroph adenoma (GAs), silent corticotroph adenomas (SCAs), null cell adenoma (NCAs), and pituitary transcription factor 1 (PIT1) lineage PAs. We evaluated MSH6/2 and PD-L1 mRNA expressions in NFPAs by real-time PCR (n = 73), and statistically analyzed the expressions and clinicopathological factors. We also investigated the effect of MSH6 knockout on PD-L1 expression in AtT-20ins and GH3. MSH6/2 expressions were significantly lower in invasive NFPAs than in non-invasive NFPAs, and lower in SCAs and NCAs than in GAs. MSH6/2 expressions were positively associated with PD-L1 expression. PD-L1 expression was significantly lower in invasive NFPAs than in non-invasive NFPAs, and lower in SCAs and NCAs than in GAs. Although MSH6/2 expressions also tended to be lower in PIT1 lineage PAs than in GAs, PIT1 lineage PAs expressed PD-L1 equivalently to GA, which was unlike SCAs and NCAs. MSH6 knockout in AtT-20ins and GH3 significantly decreased PD-L1 expression (75% and 34% reduction, respectively) with cell proliferation promotion. In conclusion, differences in MSH6/2 and PD-L1 expressions of SCAs, NCAs, and PIT1-lineage PAs from those of GAs appear to contribute to their clinically aggressive characteristics, such as more proliferation and invasiveness.

Intestinal Response to Acute Intragastric and Intravenous Administration of Phosphate in Rats.

BACKGROUND/AIMS: Phosphate (Pi) homeostasis is controlled by the intestine and kidneys whose capacities to transport Pi are under endocrine control. Several studies point to intestinal absorption as a therapeutic target to modulate Pi homeostasis. The small intestine is responsible for almost all Pi absorption in the gut, a process involving Na+-dependent and independent mechanisms. Three Na+-dependent Pi cotransporters have been described in the gastrointestinal tract: NaPi-IIb (a SLC34 member) and Pit-1 and Pit-2 (SLC20 transporters). We recently analysed the acute hormonal and renal response to intragastric (i.g) and intravenous (i.v) Pi-loading. This study demonstrated that the kidney quickly adapts to Pi-loading, with changes manifesting earlier in the i.v than i.g intervention. The aim of this work was to extend the previous studies in order to investigate the acute adaptation of intestinal transport of Pi and expression of intestinal Na+/Pi-cotransporters in response to acute Pi-loading. METHODS: Duodenal and jejunal mucosa was collected 40 minutes and/or 4 hours after administration (i.g and i.v) of either NaCl or Pi to anaesthetized rats. Uptakes of Pi and protein expression of Na+/Pi cotransporters were measured in brush border membrane vesicles (BBMV); the cotransporters' mRNA abundance was quantified by real-time PCR in total RNA extracted from whole mucosa. RESULTS: Pi-loading did not modify transport of Pi in duodenal and jejunal BBMV 4 hours after treatment. Administration of Pi did not alter either the intestinal expression of NaPi-IIb and Pit-2 mRNAs, whereas Pit-1 mRNA expression was only regulated (diminished) in duodenum collected 4 hours after i.g Pi-loading. NaPi-IIb protein expression was decreased in duodenum 4 hours upon i.v Pi infusion, whereas the duodenal and jejunal abundance of the cotransporter was unaffected by i.g administration of Pi. CONCLUSION: Together, these data suggest that the intestine responds acutely to Pi-loading, though this response seems slower than the renal adaptation.

Receptor for advanced glycation end products: a key molecule in the genesis of chronic kidney disease vascular calcification and a potential modulator of sodium phosphate co-transporter PIT-1 expression.

BACKGROUND: Chronic kidney disease (CKD) is associated with increased cardiovascular mortality, frequent vascular calcification (VC) and accumulation of uraemic toxins. Advanced glycation end products and S100 proteins interact with the receptor for advanced glycation end products (RAGE). In the present work, we aimed to investigate the role(s) of RAGE in the CKD-VC process. METHODS: Apoe-/- or Apoe-/-Ager (RAGE)-/- male mice were assigned to CKD or sham-operated groups. A high-phosphate diet was given to a subgroup of Apoe-/-and Apoe-/-Ager-/- CKD mice. Primary cultures of Ager+/+ and Ager-/- vascular smooth muscle cells (VSMCs) were established and stimulated with either vehicle, inorganic phosphate (Pi) or RAGE ligands (S100A12; 20 µM). RESULTS: After 12 weeks of CKD we observed a significant increase in RAGE ligand (AGE and S100 proteins) concentrations in the serum of CKD Apoe-/- mice. Ager messenger RNA (mRNA) levels were 4-fold higher in CKD vessels of Apoe-/- mice. CKD Apoe-/- but not CKD Apoe-/- or Ager-/- mice displayed a marked increase in the VC surface area. Similar trends were found in the high-phosphate diet condition. mRNA levels of Runx2 significantly increased in the Apoe-/- CKD group. In vitro, stimulation of Ager+/+VSMCs with Pi or S100A12 induced mineralization and osteoblast transformation, and this was inhibited by phosphonoformic acid (Pi co-transporters inhibitor) and Ager deletion. In vivo and in vitro RAGE was necessary for regulation of the expression of Pit-1, at least in part through production of reactive oxygen species. CONCLUSION: RAGE, through the modulation of Pit-1 expression, is a key molecule in the genesis of VC.

Functional characterization of a human POU1F1 mutation associated with isolated growth hormone deficiency: a novel etiology for IGHD.

POU1F1, a pituitary-specific POU-homeo domain transcription factor, plays an essential role in the specification of the somatotroph, lactotroph and thyrotroph lineages and in the activation of GH1, PRL and TSHß transcription. Individuals with mutations in POU1F1 present with combined deficiency of GH, PRL and TSH. Here, we identified a heterozygous missense mutation with evidence of pathogenicity, at the POU1F1 locus, in a large family in which an isolated growth hormone deficiency segregates as an autosomal dominant trait. The corresponding p.Pro76Leu mutation maps to a conserved site within the POU1F1 transactivation domain. Bandshift assays revealed that the mutation alters wild-type POU1F1 binding to cognate sites within the hGH-LCR and hGH1 promoter, but not to sites within the PRL promoter, and it selectively increases binding affinity to sites within the hGH-LCR. Co-immunoprecipitation studies reveal that this substitution enhances interactions of POU1F1 with three of its cofactors, PITX1, LHX3a and ELK1, and that residue 76 plays a critical role in these interactions. The insertion of the mutation at the mouse Pou1f1 locus results in a dramatic loss of protein expression despite normal mRNA concentrations. Mice heterozygous for the p.Pro76Leu mutation were phenotypically normal while homozygotes demonstrated a dwarf phenotype. Overall, this study unveils the involvement of POU1F1 in dominantly inherited isolated GH deficiency and demonstrates a significant impact of the Pro76Leu mutation on DNA-binding activities, alterations in transactivating functions and interactions with cofactors. Our data further highlight difficulties in modeling human genetic disorders in the mouse despite apparent conservation of gene expression pathways and physiologic functions.

Evolution of the POU1F1 transcription factor in mammals: Rapid change of the alternatively-spliced ß-domain.

The POU1F1 (Pit-1) transcription factor is important in regulating expression of growth hormone, prolactin and TSH ß-subunit, and controlling development of the anterior pituitary cells in which these hormones are produced. POU1F1 is a conserved protein comprising three main domains, an N-terminal transcription activation domain (TAD), a POU-specific domain and a C-terminal homeodomain. Within the TAD, a ß-domain can be inserted by alternative splicing, giving an extended 'ß-variant' with altered properties. Here sequence data from over 100 species were used to assess the variability of POU1F1 in mammals. This showed that the POU-specific domain and homeodomain are very strongly conserved, and that the TAD is somewhat less conserved, as are linker and hinge regions between these main domains. On the other hand, the ß-domain is very variable, apparently evolving at a rate not significantly different from that expected for unconstrained, neutral evolution. In several species stop and/or frameshift mutations within the ß-domain would prevent expression of the ß-variant as a functional protein. In most species expression of the ß-variant is low (<5% of total POU1F1 expression). The rate of evolution of POU1F1 in mammals shows little variation, though the lineage leading to dog does show an episode of accelerated change. This comparative genomics study suggests that in most mammalian species POU1F1 variants produced by alternative splicing may have little physiological significance.

Alteration of ghrelin/obestatin ratio in adolescence with polycystic ovarian syndrome.

Ghrelin, an endoggenous for the growth hormone secretagogue receptor, has been shown to participate in the regulation of energy homeostasis and pituitary hormone secretion. Obestatin, encoded by the same gene as ghrelin, is described as a physiological opponent of ghrelin. Ghrelin and obestatin are altered in polycystic ovary syndrome (PCOS), which is characterized by insulin resistance and pituitary hormone secretion disorder. The aim of this study was to evaluate ghrelin/obestatin imbalance in relation to insulin resistance and pituitary hormone in adolescence with PCOS. This restrospective case-control study included 33 adolescence with PCOS and 38 control adolescence. Ghrelin and obestatin concentrations in serum were determined by RIA, and the serum fasting glucose and Insulin were determined by the glucose oxidase color method and INS-EASIA. The serum LH and FSH were measured by highly specific hemiluminescence immunoassays. We found that the serum ghrelin levels and ghrelin/obestatin ratio were significant lower in PCOS group than in control group, and the serum obestatin levels were significant higher in PCOS group than in control group. The ghrelin/obestatin ratios were negatively correlation with LH/FSH ratio and insulin resistant index in PCOS group. The findings of this study suggest that ghrelin/obestatin imbalance may play a role in pathogenesis of adolescent PCOS.

FGFR2 regulates Mre11 expression and double-strand break repair via the MEK-ERK-POU1F1 pathway in breast tumorigenesis.

The association between breast cancer risk and genetic variants of fibroblast growth factor receptor 2 (FGFR2) has been identified and repeatedly confirmed; however, the mechanism underlying FGFR2 in breast tumorigenesis remains obscure. Given that breast tumorigenesis is particularly related to DNA double-strand-break-repair (DSBR), we examined the hypothesis that FGFR2 is involved in DSBR. Our results show that expression of Mre11, a vital exonuclease in DSBR, is downregulated by FGFR2, which is further linked to decreased DSBR. Analysis of the Mre11 promoter revealed that POU1F1 mediates FGFR2-induced Mre11 downregulation. Furthermore, ERK, downstream of FGFR2, directly interacts with and phosphorylates POU1F1, increasing POU1F1 binding capacity to the Mre11 promoter and repressing Mre11 expression, which consequently affects DSBR and sensitizes breast cancer cells to chemotherapeutic treatments. The importance of the FGFR2-Mre11-DSBR link in cancer progression is suggested by the finding that genotypes of FGFR2 and Mre11 are associated with survival of breast cancer patients and that FGFR2 expression correlates with cancer prognosis specifically in patients receiving chemotherapy. This study yields important insight into the role of FGFR2 in breast tumorigenesis and may facilitate development of a useful therapeutic approach for breast cancer.

Multihormonal pituitary adenoma concomitant with Pit-1 and Tpit lineage cells causing acromegaly associated with subclinical Cushing's disease: a case report.

BACKGROUND: A functional pituitary adenoma can produce multiple anterior-pituitary hormones, such as growth hormone (GH) -producing adenomas (GHoma) with prolactin or thyrotropin stimulating hormone production in the same lineage. However, it is very rare that acromegaly shows subclinical Cushing's disease (SCD) beyond the lineage. Here we describe the involvement of intratumoral coexistence with 2 types of hormone-producing cells associated with different lineage in acromegaly concomitant with SCD. CASE PRESENTATION: In our study, we performed clinical evaluation of the patient showing acromegaly with SCD. To elucidate the mechanisms of this pathology, we analyzed immunohistochemistry and gene expression of anterior-pituitary hormones and transcriptional factors in the resected pituitary tumor. On immunohistochemical staining, most of the tumor cells were strongly stained for GH antibody, while some cells were strongly positive for adrenocorticotropic hormone (ACTH). Gene expression analysis of a transsphenoidal surgery sample of the pituitary gland revealed that ACTH-related genes, such as POMC, Tpit, and NeuroD1 mRNA, had higher expression in the tumor tissue than the nonfunctional adenoma but lower expression compared to an adenoma of typical Cushing's disease. Further, double-labeling detection methods with a fluorescent stain for ACTH and GH demonstrated the coexistence of ACTH-positive cells (GH-negative) among the GH-positive cells in the tumor. Additionally, Pit-1 expression was reduced in the ACTH-positive cells from tumor tissue primary culture. CONCLUSION: Here we described a case of a pituitary tumor diagnosed with acromegaly associated with SCD. We performed quantitative-expression analyses of transcriptional factors of the tumor tissue and immunohistochemistry analysis of tumor-derived primary culture cells, which suggested that the multihormonal pituitary adenoma concomitant with Pit-1 and Tpit lineage cells caused acromegaly associated with SCD.

Increased PIT1 and PIT2 Expression in Streptozotocin (STZ)-induced Diabetic Mice Contributes to Uptake of iAs(V).

OBJECTIVE: This study aimed to investigate the susceptibility of mice with streptozotocin(STZ)-induced diabetes mellitus (TIDM) to the uptake of pentavalent inorganic arsenic (iAsV) and the possible molecular mechanism. METHODS: TIDM was induced in mice by STZ. TIDM and normal mice were treated with 15.0 mg/kg Na2HAsO4·12H2O by intragastric administration. Then, the concentrations of arsenic in various tissues were measured by atomic fluorescence spectrometry. The gene expression levels of Pit1 and Pit2 were quantified by real-time RT-PCR, and their protein levels were detected by Western blotting in mouse heart, kidney, and liver tissues. RESULTS: The concentrations of arsenic in STZ-induced TIDM mouse tissues were higher at 2 h after intragastric administration of Na2HAsO4·12H2O. Compared with the levels in normal mice, PIT1 and PIT2, which play a role in the uptake of iAsV, were upregulated in the livers and hearts of TIDM mice. PIT1 but not PIT2 was higher in TIDM mouse kidneys. The upregulation of Pit1 and Pit2 expression could be reversed by insulin treatment. CONCLUSION: The increased uptake of iAsV in TIDM mouse tissues may be associated with increased PIT1 and/or PIT2 expression.

Over-Expression of POU Class 1 Homeobox 1 Transcription Factor (Pit-1) Predicts Poor Prognosis for Breast Cancer Patients.

BACKGROUND: The POU class 1 homeobox 1 transcription factor (POU1F1, also known as Pit-1) was reported to be associated with tumor progression and metastasis. The purpose of this study was to evaluate the prognostic value of Pit-1 in breast cancer patients. MATERIAL AND METHODS: The relative expression levels of Pit-1 in breast cancer patients were detected by quantitative real-time PCR (qRT-PCR). Chi-square analysis was used to analyze the association between Pit-1 expression and clinical features. The Kaplan-Meier method was used to estimate the overall survival of the patients and Cox regression analysis was used to analyze the prognostic value of Pit-1. RESULTS: Increased expression of Pit-1 was detected in the tumor tissues compared with the normal tissues (1.086 vs. 0.541) and the abnormal expression was associated with tumor size, clinical stage, tumor grade, and lymph node metastasis (P<0.05). High expression level of Pit-1 was significantly associated with poor overall survival of the patients (P=0.001) and Cox regression analysis indicated that Pit-1 might be a prognostic factor for breast cancer prognosis (HR=1.955, 95% CI=1.295-3.035, P=0.003). CONCLUSIONS: Pit-1 may be a potential prognostic biomarker for breast cancer patients and it is associated with tumor progression.

Associations of Leptin and Pituitary-Specific Transcription Factor Genes' Polymorphisms with Reproduction and Production Traits in Dairy Buffalo.

This study aimed to detect the genetic variability in Leptin and Pit-1 genes using polymerase chain reaction-restriction fragment length polymorphism and DNA sequencing also to explore their possible associations with reproductive and productive traits of Egyptian buffaloes. Regarding Leptin gene, three genotypes (AA, AG and GG) were identified with frequency of 0.54, 0.40 and 0.06, respectively, and the genotypes were distributed according to the Hardy-Weinberg equilibrium. Allele A was comparatively higher than G with frequency of 0.74 and 0.26, respectively. For Pit-1 gene, the association could not be performed due to the monomorphism (BB). The results showed that AA genotypes were found to be superior in most of production and reproduction traits. AA genotypes yielded more milk (2332.34 kg, p = 0.04) with higher fat% (6.10, p = 0.004) and fat yield (155.75 kg, p = 0.06), reach peak milk production at 42.19 days and required 2.19 services for conception in comparison with GG genotypes. Birthweight of animals with AA genotype was lesser than with GG genotype (39.35 and 43.67 kg, p = 0.02, respectively). The days open is numerically better in AA genotype animals (99.35 days), but the difference between the three genotypes was non-significant. The distinct significant associations reported in this study suggested that Leptin is reputable candidate genetic marker, which might be used to enhance animals' genetic potential for milk production in conjunction with reproduction.

Indoxyl sulfate promotes vascular smooth muscle cell calcification via the JNK/Pit-1 pathway.

We determined the effect of indoxyl sulfate (IS) on Pit-1 expression and the role of Pit-1 in IS-induced osteoblastic differentiation and calcification of vascular smooth muscle cells (VSMCs). To assess osteoblastic differentiation and Pit-1 expression, VSMCs were incubated with various concentrations of IS for different durations. Phosphonoformic acid (PFA), a competitive inhibitor of Pit-1, was used to verify the role of Pit-1. Western blot analysis and quantitative real-time polymerase chain reaction (PCR) were performed to assess Pit-1 protein and mRNA levels, respectively. To evaluate calcification, calcium content was measured. After IS treatment, we observed osteoblastic differentiation and calcification of VSMCs and up-regulation of Pit-1 expression. Moreover, the effect of IS on osteoblastic differentiation and Pit-1 expression was partly dose- and time-dependent. PFA abrogated the IS-induced osteoblastic differentiation and calcification of VSMCs to a certain extent. The c-Jun N-terminal kinase (JNK) pathway was activated after treatment with IS, whereas inhibition of the JNK pathway partially attenuated the effect of IS on both the stimulation of Pit-1 expression and calcium deposition. Our study is the first to demonstrate that IS promotes Pit-1 expression in part by activation of the JNK pathway that is involved in the mechanism of IS-induced osteoblastic differentiation and matrix mineralization.