Assessment of Molecular Markers in Pediatric Ovarian Tumors: Romanian Single-Center Experience.

Pediatric ovarian tumors exhibit unique diagnostic and therapeutic challenges. This study evaluates the expression of SALL4 and OCT3/4 biomarkers in pediatric ovarian tumors and their associations with tumor subtype, stage, and clinical outcome. A retrospective analysis was conducted on 64 patients under 18 years old, examining demographic data, tumor characteristics, immunohistochemical staining, and clinical outcomes. Our results show that SALL4 was significantly expressed in adenocarcinoma, dysgerminoma (DSG), mixed germ cell tumors (GCTs), and immature teratoma, while OCT3/4 was highly expressed in DSG and mixed GCTs. Both markers are associated with a higher tumor grade and stage, indicating a more aggressive disease. The SALL4 positivity expression was correlated with high alpha fetoprotein (AFP) and lactate dehydrogenase (LDH) levels, while OCT3/4 positivity significantly predicted the risk of subsequent metastasis. The mean progression-free survival (PFS) was notably shorter in patients with positive markers. These findings underscore the diagnostic and prognostic value of SALL4 and OCT3/4 in pediatric ovarian tumors, aligning with previous research and supporting their use in clinical practice for better disease management and patient outcomes.

The expression of Oct3/4A mRNA and not its isoforms is upregulated by the HPV16 E7 oncoprotein.

PURPOSE: Oct3/4 a transcription factor is involved in maintaining the characteristics of cancer stem cells. Oct3/4 can be expressed differentially with respect to the progression of cervical cancer (CC). In addition, Oct3/4 can give rise to three isoforms by alternative splicing of the mRNA Oct3/4A, Oct3/4B and Oct3/4B1. The aim of this study was to evaluate the mRNA expression from Oct3/4A, Oct3/4B and Oct3/4B1 in low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL), CC samples, and measure the effect of the HPV16 E7 oncoprotein on the mRNA expression from Oct3/4 isoforms in the C-33A cell line. METHODS: The expression levels of Oct3/4A, Oct3/4B and Oct3/4B1 mRNA were analyzed by reverse transcription quantitative polymerase chain reaction (RT-qPCR) in patients with LSILs, HSILs and CC. Additionally, C-33A cells that expressed the HPV16 E7 oncoprotein were established to evaluate the effect of E7 on the expression of Oct3/4 mRNA isoforms. RESULTS: Oct3/4A (p = 0.02), Oct3/4B (p = 0. 001) and Oct3/4B1 (p < 0. 0001) expression is significantly higher in patients with LSIL, HSIL and CC than in woman with non-IL. In the C-33A cell line, the expression of Oct3/4A mRNA in the presence of the E7 oncoprotein increased compared to that in nontransfected C-33A cells. CONCLUSION: Oct3/4B and Oct3/4B1 mRNA were expressed at similar levels among the different groups. These data indicate that only the mRNA of Oct3/4A is upregulated by the HPV16 E7 oncoprotein.

Upregulation of the Oct3/4 Network in Basal Breast Cancer Is Associated with Its Metastatic Potential and Shows Tissue Dependent Variability.

Adaptive plasticity of Breast Cancer stem cells (BCSCs) is strongly correlated with cancer progression and resistance, leading to a poor prognosis. In this study, we report the expression profile of several pioneer transcription factors of the Oct3/4 network associated with tumor initiation and metastasis. In the triple negative breast cancer cell line (MDA-MB-231) stably transfected with human Oct3/4-GFP, differentially expressed genes (DEGs) were identified using qPCR and microarray, and the resistance to paclitaxel was assessed using an MTS assay. The tumor-seeding potential in immunocompromised (NOD-SCID) mice and DEGs in the tumors were also assessed along with the intra-tumor (CD44+/CD24-) expression using flow cytometry. Unlike 2-D cultures, the Oct3/4-GFP expression was homogenous and stable in 3-D mammospheres developed from BCSCs. A total of 25 DEGs including Gata6, FoxA2, Sall4, Zic2, H2afJ, Stc1 and Bmi1 were identified in Oct3/4 activated cells coupled with a significantly increased resistance to paclitaxel. In mice, the higher Oct3/4 expression in tumors correlated with enhanced tumorigenic potential and aggressive growth, with metastatic lesions showing a >5-fold upregulation of DEGs compared to orthotopic tumors and variability in different tissues with the highest modulation in the brain. Serially re-implanting tumors in mice as a model of recurrence and metastasis highlighted the sustained upregulation of Sall4, c-Myc, Mmp1, Mmp9 and Dkk1 genes in metastatic lesions with a 2-fold higher expression of stem cell markers (CD44+/CD24-). Thus, Oct3/4 transcriptome may drive the differentiation and maintenance of BCSCs, promoting their tumorigenic potential, metastasis and resistance to drugs such as paclitaxel with tissue-specific heterogeneity.

Inducible expression of Oct-3/4 reveals synergy with Klf4 in targeting Cyclin A2 to enhance proliferation during early reprogramming.

During reprogramming of somatic cells, heightened proliferation is one of the earliest changes observed. While other early events such as mesenchymal-to-epithelial transition have been well studied, the mechanisms by which the cell cycle switches from a slow cycling state to a faster cycling state are still incompletely understood. To investigate the role of Oct-3/4 in this early transition, we created a 4-Hydroxytamoxifen (OHT) dependent Oct-3/4 Estrogen Receptor fusion (OctER). We confirmed that OctER can substitute for Oct-3/4 to reprogram mouse embryonic fibroblasts to a pluripotent state. During the early stages of reprograming, Oct-3/4 and Klf4 individually did not affect cell proliferation but in combination hastened the cell cycle. Using OctER + Klf4, we found that proliferative enhancement is OHT dose-dependent, suggesting that OctER is the driver of this transition. We identified Cyclin A2 as a likely target of Oct-3/4 + Klf4. In mESC, Klf4 and Oct-3/4 bind ∼100bp upstream of Cyclin A2 CCRE, suggesting a potential regulatory role. Using inducible OctER, we show a dose-dependent induction of Cyclin A2 promoter-reporter activity. Taken together, our results suggest that Cyclin A2 is a key early target during reprogramming, and support the view that a rapid cell cycle assists the transition to pluripotency.

OCT3/4 enhances tumor immune response by upregulating the TET1-dependent NRF2/MDM2 axis in bladder cancer.

This study aimed to investigate the function of OCT3/4 on tumor immune escape in bladder cancer. Initially, the expression of OCT3/4, TET1, NRF2 and MDM2 was quantified in tumor tissues and cells, followed by gain- or loss-of-function studies to define their roles in cell migration, invasion and apoptosis and tumorigenicity in nude mice. Bladder cancer presented with abundant expression levels of OCT3/4, TET1, NRF2 and MDM2. We found that OCT3/4 promoted TET1 expression via binding to its promoter and that TET1 recruited MLL protein to NRF2 promoter and upregulated its expression, while NRF2 enhanced MDM2 expression. Upregulated MDM2 accelerated tumor immune escape in bladder cancer in mice. OCT3/4 knockdown suppressed the cell migration and invasion while inducing apoptosis, and consequently prevented tumor growth and immune escape in mice. Collectively, OCT3/4 may promote the progression of tumor immune escape in bladder cancer through acting as a promoter of the TET1/NRF2/MDM2 axis.

Assessment of the association of OCT3/4 with GLUT1 and CD105 in oral squamous cell carcinoma using dual immunohistochemistry.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is the most common cancer affecting the oral and maxillofacial region. This study aimed to investigate the role of cancer stem cells (CSCs) in angiogenesis and hypoxic response in OSCC. METHODS: This retrospective observational study evaluated 56 cases of OSCC using dual immunohistochemistry. Octamer-binding transcription factor 3/4 (OCT3/4) marker was used to evaluate CSC activity. Glucose transporter 1 (GLUT1) marker was used to evaluate the hypoxic response and angiogenesis, while endoglin (CD105) was used to evaluate the late stage of angiogenesis and blood vessel formation. RESULTS: Co-expression of OCT3/4 and GLUT1 was noted in 11 of 12 patients with grade III OSCC. However, we did not observe co-expression of these markers in 13 of 22 patients with grade I OSCC. Although we observed a significant correlation between co-expression of GLUT1 and OCT3/4 and tumor grade, there was no significant correlation between co-expression of OCT3/4 and CD105 and different grades of OSCC. CONCLUSIONS: CSCs could play important roles in the initial stages of hypoxic response and angiogenesis. Our result reported that in higher grades of OSCC, GLUT1 as a first response to hypoxic situations might be a result of CSCs. Further studies are required to discover other biomarkers, their roles, and associated pathways of CSCs in OSCC.

Expression of the C-terminal region of the SSX protein is a useful diagnostic biomarker for spermatocytic tumour.

AIMS: Spermatocytic tumour (ST) is a rare testicular germ cell neoplasm with few confirmatory biomarkers that can be challenging to diagnose. Like normal spermatogonia, STs are known to express SSX proteins. Recently, a novel SSX antibody directed against a conserved C-terminal region of SSX1, SSX2 and SSX4 (SSX_CT) has emerged as a reliable biomarker for these SSX proteins and synovial sarcoma. However, SSX_CT immunostaining has not been demonstrated in ST. The aim of this study was to assess the diagnostic utility of SSX_CT immunohistochemistry in ST and other tumours in the differential diagnosis with ST. METHODS AND RESULTS: SSX_CT, OCT3/4 and c-KIT immunohistochemistry was performed on 15 STs, 38 seminomas, 13 embryonal carcinomas, 12 yolk sac tumours, six choriocarcinomas, four teratomas, seven Sertoli cell tumours, and six lymphomas. Staining was scored as negative, rare, focal, or diffuse. SSX_CT was positive in all (15/15) STs, and diffusely positive in 14 of 15 (93%). SSX_CT was positive in 22 of 38 (58%) seminomas; however, only two cases showed diffuse expression. SSX_CT was negative in all other tumours. OCT3/4 was negative in all STs, but positive in all seminomas and embryonal carcinomas. c-KIT was frequently positive in both STs (12/15; 80%) and seminomas (33/38; 87%). OCT3/4 and c-KIT were negative in all other tumours. CONCLUSIONS: SSX_CT is a valuable and highly sensitive biomarker that supports the diagnosis of ST. Diffuse expression of SSX-CT in STs is also highly specific for ST. Nevertheless, SSX_CT is best used in combination with OCT3/4 when ST is in the differential diagnosis.

Substrates and Inhibitors of Organic Cation Transporters (OCTs) and Plasma Membrane Monoamine Transporter (PMAT) and Therapeutic Implications.

The gene products of the SLC22A gene family (hOCT1, hOCT2, and hOCT3) and of the SLC29A4 gene (hPMAT or hENT4) are all polyspecific organic cation transporters. Human OCTs (including hPMAT) are expressed in peripheral tissues such as small intestine, liver, and kidney involved in the pharmacokinetics of drugs. In the human brain, all four transporters are expressed at the blood-brain barrier (BBB), hOCT2 is additionally expressed in neurons, and hOCT3 and hPMAT in glia. More than 40% of the presently used drugs are organic cations. This chapter lists and discusses all known drugs acting as substrates or inhibitors of these four organic cation transporters, independently of whether the transporter is expressed in the central nervous system (CNS) or in peripheral tissues. Of interest is their involvement in drug absorption, distribution, and excretion as well as potential OCT-associated drug-drug interactions (DDIs), with a focus on drugs that act in the CNS.

Organic Cation Transporters in Brain Catecholamine Homeostasis.

Catecholamines, including dopamine, norepinephrine, and epinephrine, are modulatory transmitters released from specialized neurons throughout the brain. Collectively, catecholamines exert powerful regulation of mood, motivation, arousal, and plasticity. Transporter-mediated uptake determines the peak concentration, duration, and physical spread of released catecholamines, thus playing key roles in determining the magnitude and duration of their modulatory effects. Most studies of catecholamine clearance have focused on the presynaptic high-affinity, low-capacity dopamine (DAT), and norepinephrine (NET) transporters, which are members of the uptake1 family of monoamine transporters. However, recent studies have demonstrated that members of the uptake2 family of monoamine transporters, including organic cation transporter 2 (OCT2), OCT3, and the plasma membrane monoamine transporter (PMAT) are expressed widely throughout the brain. In contrast to DAT and NET, these transporters have higher capacity and lower affinity for catecholamines and are multi-specific, each with the capacity to transport all catecholamines. The expression of these transporters in the brain suggests that they play significant roles in regulating catecholamine homeostasis. This review summarizes studies describing the anatomical distribution of OCT2, OCT3, and PMAT, their cellular and subcellular localization, and their contribution to the regulation of the clearance of catecholamines in the brain.

General Overview of Organic Cation Transporters in Brain.

Inhibitors of Na+/Cl- dependent high affinity transporters for norepinephrine (NE), serotonin (5-HT), and/or dopamine (DA) represent frequently used drugs for treatment of psychological disorders such as depression, anxiety, obsessive-compulsive disorder, attention deficit hyperactivity disorder, and addiction. These transporters remove NE, 5-HT, and/or DA after neuronal excitation from the interstitial space close to the synapses. Thereby they terminate transmission and modulate neuronal behavioral circuits. Therapeutic failure and undesired central nervous system side effects of these drugs have been partially assigned to neurotransmitter removal by low affinity transport. Cloning and functional characterization of the polyspecific organic cation transporters OCT1 (SLC22A1), OCT2 (SLC22A2), OCT3 (SLC22A3) and the plasma membrane monoamine transporter PMAT (SLC29A4) revealed that every single transporter mediates low affinity uptake of NE, 5-HT, and DA. Whereas the organic transporters are all located in the blood brain barrier, OCT2, OCT3, and PMAT are expressed in neurons or in neurons and astrocytes within brain areas that are involved in behavioral regulation. Areas of expression include the dorsal raphe, medullary motoric nuclei, hypothalamic nuclei, and/or the nucleus accumbens. Current knowledge of the transport of monoamine neurotransmitters by the organic cation transporters, their interactions with psychotropic drugs, and their locations in the brain is reported in detail. In addition, animal experiments including behavior tests in wildtype and knockout animals are reported in which the impact of OCT2, OCT3, and/or PMAT on regulation of salt intake, depression, mood control, locomotion, and/or stress effect on addiction is suggested.

Brain Plasma Membrane Monoamine Transporter in Health and Disease.

Precise control of monoamine neurotransmitter levels in the central nervous system (CNS) is crucial for proper brain function. Dysfunctional monoamine signaling is associated with several neuropsychiatric and neurodegenerative disorders. The plasma membrane monoamine transporter (PMAT) is a new polyspecific organic cation transporter encoded by the SLC29A4 gene. Capable of transporting monoamine neurotransmitters with low affinity and high capacity, PMAT represents a major uptake2 transporter in the brain. Broadly expressed in multiple brain regions, PMAT can complement the high-affinity, low-capacity monoamine uptake mediated by uptake1 transporters, the serotonin, dopamine, and norepinephrine transporters (SERT, DAT, and NET, respectively). This chapter provides an overview of the molecular and functional characteristics of PMAT together with its regional and cell-type specific expression in the mammalian brain. The physiological functions of PMAT in brain monoamine homeostasis are evaluated in light of its unique transport kinetics and brain location, and in comparison with uptake1 and other uptake2 transporters (e.g., OCT3) along with corroborating experimental evidences. Lastly, the possibility of PMAT's involvement in brain pathophysiological processes, such as autism, depression, and Parkinson's disease, is discussed in the context of disease pathology and potential link to aberrant monoamine pathways.

Organic Cation Transporters and Nongenomic Glucocorticoid Action.

Corticosteroid hormones exert powerful influences on neuronal physiology and behavior by activating intracellular glucocorticoid receptors (GR) and mineralocorticoid receptors (MR), which act as ligand-gated transcription factors, altering gene expression. In addition to these genomic effects on physiology and behavior, which are usually delayed by minutes to hours, corticosteroid hormones also initiate rapid effects through diverse nongenomic mechanisms. One such mechanism involves the direct inhibition by corticosteroid hormones of monoamine transport mediated by the "uptake2" transporter, organic cation transporter 3 (OCT3), a high-capacity, low-affinity transporter for norepinephrine, epinephrine, dopamine, serotonin, and histamine. In this review we describe studies that demonstrate OCT3 expression and corticosterone-sensitive monoamine transport in the brain and present evidence supporting the hypothesis that corticosterone exerts rapid, nongenomic actions on glia and neurons, ultimately modulating physiology and behavior, by inhibiting OCT3-mediated monoamine clearance. We also describe the corticosteroid sensitivity of the other members of the uptake2 family and examine their potential contributions to nongenomic effects of corticosteroids in the brain.

Genetic and Epigenetic Regulation of Organic Cation Transporters.

Organic cation transporters (OCTs) of the solute carrier family (SLC) 22 are the subject of intensive research because they mediate the transport of many clinically-relevant drugs such as the antidiabetic agent metformin, the opioid tramadol, and the antimigraine agent sumatriptan. OCT1 (SLC22A1) and OCT2 (SLC22A2) are highly expressed in human liver and kidney, respectively, while OCT3 (SLC22A3) shows a broader tissue distribution. As suggested from studies using knockout mice, particularly OCT2 and OCT3 appear to be of relevance for brain physiological function and drug response. The knowledge of genetic factors and epigenetic modifications affecting function and expression of OCTs is important for a better understanding of disease mechanisms and for personalized treatment of patients. This review briefly summarizes the impact of genetic variants and epigenetic regulation of OCTs in general. A comprehensive overview is given on the consequences of OCT2 and OCT3 knockout in mice and the implications of genetic OCT2 and OCT3 variants on central nervous system function in humans.

Cruciform Formable Sequences within Pou5f1 Enhancer Are Indispensable for Mouse ES Cell Integrity.

DNA can adopt various structures besides the B-form. Among them, cruciform structures are formed on inverted repeat (IR) sequences. While cruciform formable IRs (CFIRs) are sometimes found in regulatory regions of transcription, their function in transcription remains elusive, especially in eukaryotes. We found a cluster of CFIRs within the mouse Pou5f1 enhancer. Here, we demonstrate that this cluster or some member(s) plays an active role in the transcriptional regulation of not only Pou5f1, but also Sox2, Nanog, Klf4 and Esrrb. To clarify in vivo function of the cluster, we performed genome editing using mouse ES cells, in which each of the CFIRs was altered to the corresponding mirror repeat sequence. The alterations reduced the level of the Pou5f1 transcript in the genome-edited cell lines, and elevated those of Sox2, Nanog, Klf4 and Esrrb. Furthermore, transcription of non-coding RNAs (ncRNAs) within the enhancer was also upregulated in the genome-edited cell lines, in a similar manner to Sox2, Nanog, Klf4 and Esrrb. These ncRNAs are hypothesized to control the expression of these four pluripotency genes. The CFIRs present in the Pou5f1 enhancer seem to be important to maintain the integrity of ES cells.

SLC22 Transporters in the Fly Renal System Regulate Response to Oxidative Stress In Vivo.

Several SLC22 transporters in the human kidney and other tissues are thought to regulate endogenous small antioxidant molecules such as uric acid, ergothioneine, carnitine, and carnitine derivatives. These transporters include those from the organic anion transporter (OAT), OCTN/OCTN-related, and organic cation transporter (OCT) subgroups. In mammals, it has been difficult to show a clear in vivo role for these transporters during oxidative stress. Ubiquitous knockdowns of related Drosophila SLC22s-including transporters homologous to those previously identified by us in mammals such as the "Fly-Like Putative Transporters" FLIPT1 (SLC22A15) and FLIPT2 (SLC22A16)-have shown modest protection against oxidative stress. However, these fly transporters tend to be broadly expressed, and it is unclear if there is an organ in which their expression is critical. Using two tissue-selective knockdown strategies, we were able to demonstrate much greater and longer protection from oxidative stress compared to previous whole fly knockdowns as well as both parent and WT strains (CG6126: p < 0.001, CG4630: p < 0.01, CG16727: p < 0.0001 and CG6006: p < 0.01). Expression in the Malpighian tubule and likely other tissues as well (e.g., gut, fat body, nervous system) appear critical for managing oxidative stress. These four Drosophila SLC22 genes are similar to human SLC22 transporters (CG6126: SLC22A16, CG16727: SLC22A7, CG4630: SLC22A3, and CG6006: SLC22A1, SLC22A2, SLC22A3, SLC22A6, SLC22A7, SLC22A8, SLC22A11, SLC22A12 (URAT1), SLC22A13, SLC22A14)-many of which are highly expressed in the kidney. Consistent with the Remote Sensing and Signaling Theory, this indicates an important in vivo role in the oxidative stress response for multiple SLC22 transporters within the fly renal system, perhaps through interaction with SLC22 counterparts in non-renal tissues. We also note that many of the human relatives are well-known drug transporters. Our work not only indicates the importance of SLC22 transporters in the fly renal system but also sets the stage for in vivo studies by examining their role in mammalian oxidative stress and organ crosstalk.

Hepatocellular Carcinomas With Mutational Activation of Beta-Catenin Require Choline and Can Be Detected by Positron Emission Tomography.

BACKGROUND & AIMS: In one-third of hepatocellular carcinomas (HCCs), cancer cells have mutations that activate ß-catenin pathway. These cells have alterations in glutamine, bile, and lipid metabolism. We investigated whether positron emission tomography (PET) imaging allows identification of altered metabolic pathways that might be targeted therapeutically. METHODS: We studied mice with activation of ß-catenin in liver (Apcko-liv mice) and male C57Bl/6 mice given injections of diethylnitrosamine, which each develop HCCs. Mice were fed a conventional or a methionine- and choline-deficient diet or a choline-deficient (CD) diet. Choline uptake and metabolism in HCCs were analyzed by micro-PET imaging of mice; livers were collected and analyzed by histologic, metabolomic, messenger RNA quantification, and RNA-sequencing analyses. Fifty-two patients with HCC underwent PET imaging with 18F-fluorodeoxyglucose, followed by 18F-fluorocholine tracer metabolites. Human HCC specimens were analyzed by immunohistochemistry, quantitative polymerase chain reaction, and DNA sequencing. We used hepatocytes and mouse tumor explants for studies of incorporation of radiolabeled choline into phospholipids and its contribution to DNA methylation. We analyzed HCC progression in mice fed a CD diet. RESULTS: Livers and tumors from Apcko-liv mice had increased uptake of dietary choline, which contributes to phospholipid formation and DNA methylation in hepatocytes. In patients and in mice, HCCs with activated ß-catenin were positive in 18F-fluorocholine PET, but not 18F-fluorodeoxyglucose PET, and they overexpressed the choline transporter organic cation transporter 3. The HCC cells from Apcko-liv mice incorporated radiolabeled methyl groups of choline into phospholipids and DNA. In Apcko-liv mice, the methionine- and choline-deficient diet reduced proliferation and DNA hypermethylation of hepatocytes and HCC cells, and the CD diet reduced long-term progression of tumors. CONCLUSIONS: In mice and humans, HCCs with mutations that activate ß-catenin are characterized by increased uptake of a fluorocholine tracer, but not 18F-fluorodeoxyglucose, revealed by PET. The increased uptake of choline by HCCs promotes phospholipid formation, DNA hypermethylation, and hepatocyte proliferation. In mice, the CD diet reverses these effects and promotes regression of HCCs that overexpress ß-catenin.

[Paraganglioma of urinary bladder: a clinicopathological features analysis of 23 cases].

Objective: To investigate the clinicopathological features, diagnosis, differential diagnosis and immunohistochemical (IHC) characteristics of paraganglioma of urinary bladder (PUB). Methods: The clinical and pathological data of 23 cases of PUB were collected at the Second Affiliated Hospital of Fujian Medical University (7 cases); Fujian Provincial Hospital (8 cases); Fujian Medical University Union Hospital (6 cases); and First Affiliated Hospital of Fujian Medical University (2 cases) from May 2010 to November 2018. IHC staining for CK, GATA3, CD56, Syn, CgA, S-100 protein, HMB45, SDHB, OCT3/4 and Ki-67 was done using EliVision method; and the relevant literature was reviewed. Results: There were 14 women and 9 men, aged ranged from 21 to 73 years (median 51 years). Clinically, patients presented with headache, vertigo, palpitation, hypertensive crisis during micturition, hypertension, blurred vision, gross hematuria and paroxysmal pallor. The tumor sizes ranged from 0.9 to 6 cm (mean2.5 cm). Macroscopically, most tumors were exophytic and well delineated within the lamina propria or muscularis propria. The tumors were firm and nodular and showed grayish-tan cut surface. Histologically,the tumor growth pattern was expansive or showed interpenetrating infiltrative growth within the lamina propria or muscularis propria; the tumor cells were typically arranged in distinctive nests (Zellballen) with organoid arrangement; pseudo-rosette were seen in some cases. The cells were rounded or polygonal and had rich, acidophilic or amphophilic cytoplasm and may contain pigmented granules and vacuoles; the nuclei were central or eccentric, with small nucleoli, although occasionally some nuclei were pleomorphic and hyperchromatic. Spindled sustentacular cells could be seen around the nests of tumor cells in some cases. There were abundant vessels that were fissure-like, hemangioma-like or dilated. By IHC, the tumor cells were positive for GATA3 (2/23), OCT3/4 (2/23), CD56 (22/23), Syn (23/23), CgA (22/23), S-100 (sustentacular cell, 23/23) and SDHB (23/23); and negative for CK and HMB45; Ki-67 index was 1%-5%. At follow-up, there was no recurrence or metastasis in 18 cases. Conclusions: The diagnosis of PUB relies on the morphologic and IHC features; but there may be histomorphologic heterogeneity. The most important differential diagnosis is invasive urothelial carcinoma. The tumor cells may show aberrant cytoplasmic expression of OCT3/4; there is no clear correlation between SDHB and OCT3/4 expression in the group.

Deletion of organic cation transporter Oct3 promotes hepatic fibrosis via upregulation of TGFß.

Organic cation transporters (OCT) are responsible for the intracellular uptake and detoxification of a broad spectrum of endogenous and exogenous substrates. OCTs are downregulated in cholestasis, fibrosis, and hepatocellular carcinoma, but the underlying molecular mechanisms and downstream effects of OCT deletion are unknown. Oct3-knockout (Oct3-/-; FVB.Slc22a3tm10pb) and wild-type (WT; FVB) mice were subject to escalating doses of carbon tetrachloride (CCl4) or thioacetamide (TAA) for 6 wk to induce advanced parenchymal liver fibrosis. Secondary biliary fibrosis was generated by bile duct ligation. Liver fibrosis was assessed by hydroxyproline determination, quantitative Sirius red morphometry, and quantitative real-time PCR for fibrosis and inflammation-related genes. Ductular reaction was assessed by bile duct count per field of view in hematoxylin and eosin staining. General gene expression analyses were performed in liver tissue from untreated Oct3-/- and WT mice. Finally, primary murine hepatocytes were treated with the nonselective OCT inhibitor quinine, and transforming growth factor-ß1 (Tgfß1) protein expression was quantified by quantitative real-time PCR and Western blot. Oct3-/- mice developed significantly more fibrosis after bile duct ligation and CCl4 treatment compared with WT mice. Ductular reaction was enhanced in the long-term model. Concomitantly, Oct1 mRNA expression was downregulated during cholestatic and chemically (TAA and CCl4) induced fibrogenesis. The downregulation of Oct1 mRNA in fibrotic liver tissue reversed within 4 wk after TAA cessation. Gene expression analysis by next-generation sequencing revealed an enrichment of Tgfß1 target genes in Oct3-/- mice. Tgfß1 mRNA expression was significantly upregulated after chemically induced fibrosis (P < 0.001) in Oct3-/- compared with WT mice. Accordingly, in primary murine hepatocytes functional inhibition of OCT led to an upregulation of Tgfß1 mRNA expression. Loss of Oct3 promotes fibrogenesis by affecting Tgfß-mediated homeostasis in mice with chronic biliary and parenchymal liver damage and fibrosis.NEW & NOTEWORTHY We show for the first time that organic cation transporter 3 (Oct3) is not only downregulated in fibrosis but loss of Oct3 also leads to an upregulation of transforming growth factor-ß contributing to fibrosis progression.

Small 6q16.1 Deletions Encompassing POU3F2 Cause Susceptibility to Obesity and Variable Developmental Delay with Intellectual Disability.

Genetic studies of intellectual disability and identification of monogenic causes of obesity in humans have made immense contribution toward the understanding of the brain and control of body mass. The leptin > melanocortin > SIM1 pathway is dysregulated in multiple monogenic human obesity syndromes but its downstream targets are still unknown. In ten individuals from six families, with overlapping 6q16.1 deletions, we describe a disorder of variable developmental delay, intellectual disability, and susceptibility to obesity and hyperphagia. The 6q16.1 deletions segregated with the phenotype in multiplex families and were shown to be de novo in four families, and there was dramatic phenotypic overlap among affected individuals who were independently ascertained without bias from clinical features. Analysis of the deletions revealed a ∼350 kb critical region on chromosome 6q16.1 that encompasses a gene for proneuronal transcription factor POU3F2, which is important for hypothalamic development and function. Using morpholino and mutant zebrafish models, we show that POU3F2 lies downstream of SIM1 and controls oxytocin expression in the hypothalamic neuroendocrine preoptic area. We show that this finding is consistent with the expression patterns of POU3F2 and related genes in the human brain. Our work helps to further delineate the neuro-endocrine control of energy balance/body mass and demonstrates that this molecular pathway is conserved across multiple species.

Upregulated OCT3 has the potential to improve the survival of colorectal cancer patients treated with (m)FOLFOX6 adjuvant chemotherapy.

PURPOSE: To investigate the influence of organic cation transporter 3 (OCT3) expression on the effect of the combination regimen of 5-fluorouracil, folinic acid and oxaliplatin ((m)FOLFOX6) in colorectal cancer (CRC) patients. METHODS: This is a retrospective study conducted at a single centre (Sichuan Academy of Medical Sciences & Sichuan Provincial People's Hospital, China). Patients with stage IIb-IV resectable CRC who were being postoperatively treated with (m)FOLFOX6 as a first-line adjuvant chemotherapy regimen for at least 5 cycles and had resected primary tumour samples available were eligible for the study. Patients who preoperatively received chemotherapy and/or radiotherapy or were treated with targeted drugs or other anticancer drugs were excluded from the study. Immunohistochemical staining and digital image analysis were used to assess OCT3 expression in tumour samples. According to OCT3 expression level, the receiver operating characteristic curve (ROC curve) was used to divide the patients into two groups. Cox proportional risk regression was performed with the forward LR (forward stepwise regression based on maximum likelihood estimation) method using SPSS17.0 software. The primary endpoint was the 2-year progression-free survival. RESULTS: In total, 57 patients were included between 2014 and 2016 according to the inclusion and exclusion criteria (22 had low OCT3 expression, and 35 had high OCT3 expression). The mean age was 55.7 (30-74) years, and 37 of the total patients were male. According to TNM stage, 5 patients had stage IV disease, 44 patients had stage III disease, and 8 patients had stage II disease. Through Cox regression analysis, we found that among patients receiving the (m)FOLFOX6 regimen, those with higher OCT3 expression had a higher two-year progression-free survival rate than those with lower OCT3 expression (P = 0.038). The hazard ratio of patients with high OCT3 expression compared with patients with low OCT3 expression was 0.247. Besides, it was found that the age of patients was negatively correlated with expression level of OCT3, which can explain why patients over 70 years do not benefit from oxaliplatin-containing chemotherapy. CONCLUSIONS: High OCT3 expression in CRC tissues may be a protective factor for CRC patients treated with (m)FOLFOX6.