The transcription factor OCT6 promotes the dissolution of the naïve pluripotent state by repressing Nanog and activating a formative state gene regulatory network.

In the mouse embryo, the transition from the preimplantation to the postimplantation epiblast is governed by changes in the gene regulatory network (GRN) that lead to transcriptional, epigenetic, and functional changes. This transition can be faithfully recapitulated in vitro by the differentiation of mouse embryonic stem cells (mESCs) to epiblast-like cells (EpiLCs), that reside in naïve and formative states of pluripotency, respectively. However, the GRN that drives this conversion is not fully elucidated. Here we demonstrate that the transcription factor OCT6 is a key driver of this process. Firstly, we show that Oct6 is not expressed in mESCs but is rapidly induced as cells exit the naïve pluripotent state. By deleting Oct6 in mESCs, we find that knockout cells fail to acquire the typical morphological changes associated with the formative state when induced to differentiate. Additionally, the key naïve pluripotency TFs Nanog, Klf2, Nr5a2, Prdm14, and Esrrb were expressed at higher levels than in wild-type cells, indicating an incomplete dismantling of the naïve pluripotency GRN. Conversely, premature expression of Oct6 in naïve cells triggered a rapid morphological transformation mirroring differentiation, that was accompanied by the upregulation of the endogenous Oct6 as well as the formative genes Sox3, Zic2/3, Foxp1, Dnmt3A and FGF5. Strikingly, we found that OCT6 represses Nanog in a bistable manner and that this regulation is at the transcriptional level. Moreover, our findings also reveal that Oct6 is repressed by NANOG. Collectively, our results establish OCT6 as a key TF in the dissolution of the naïve pluripotent state and support a model where Oct6 and Nanog form a double negative feedback loop which could act as an important toggle mediating the transition to the formative state.

The clinical and biochemical spectrum of ectopic acromegaly.

Ectopic acromegaly is a rare condition caused by extrapituitary central or peripheral neuroendocrine tumours (NET) that hypersecrete GH or, more commonly, GHRH. It affects less than 1% of acromegaly patients and a misdiagnosis of classic acromegaly can lead to an inappropriate pituitary surgery. Four types of ectopic acromegaly have been described: 1) Central ectopic GH-secretion: Careful cross-sectional imaging is required to exclude ectopic pituitary adenomas. 2) Peripheral GH secretion: Extremely rare. 3) Central ectopic GHRH secretion: Sellar gangliocytomas immunohistochemically positive for GHRH are found after pituitary surgery. 4) Peripheral GHRH secretion: The most common type of ectopic acromegaly is due to peripheral GHRH-secreting NETs. Tumours are large and usually located in the lungs or pancreas. Pituitary hyperplasia resulting from chronic GHRH stimulation is difficult to detect or can be misinterpreted as pituitary adenoma in the MRI. Measurement of serum GHRH levels is a specific and useful diagnostic tool. Surgery of GHRH-secreting NETs is often curative.

Acute and subacute clinical markers after sport-related concussion in rugby union players.

OBJECTIVES: The purpose of this study was to investigate the relationship between on-field post-concussion symptoms reported by athletes, on-field neurological signs reported by a trainer or physician, and/or post-concussion symptoms 72 h after brain injury in male rugby players. DESIGN: Cross-sectional study in a Sports Concussion Clinic setting. METHODS: We enrolled 92 adult rugby union players, within the first 72 h after sport concussion. Four scales were measured. Immediate Concussion Sign Checklist (sideline); Immediate Concussion Symptom Checklist (24 h after concussion); Post-Concussion Symptoms Scale and Beck Depression Inventory (in-office 72 h after concussion). RESULTS: Odds ratios revealed that overtly symptomatic athletes were over 2.6 times more likely (p = 0.047) to exhibit post-traumatic amnesia than asymptomatic athletes. There were no differences in terms of on-field loss of consciousness or confusion. Immediate symptoms reported by athletes retrospectively were associated with symptoms reported on the Beck Depression Inventory (odds ratio 2.8; 95 % confidence interval 1.14-6.88), headache (odds ratio 4.9; 95 % confidence interval 1.92-12.79), memory concerns (odds ratio 3.15; 95 % confidence interval 1.06-9.34), pressure in the head (odds ratio 2.8; 95 % confidence interval 1.03-8.08), and visual disturbances (odds ratio 3.9; 95 % confidence interval 1.05-14.50) reported 72 h after sports concussion. CONCLUSIONS: Athletes who were overtly symptomatic after sports concussion were significantly more likely to experience post-traumatic amnesia and two or more on-field concussion signs relative to those athletes who were asymptomatic. Also, players with immediate symptoms reported higher depressive symptoms, somatic symptoms (headache and visual disturbances), and cognitive symptoms.

Latin American Initiative for Lifestyle Intervention to Prevent Cognitive Decline (LatAm-FINGERS): Study design and harmonization.

INTRODUCTION: Latin American Initiative for Lifestyle Intervention to Prevent Cognitive Decline (LatAm-FINGERS) is the first non-pharmacological multicenter randomized clinical trial (RCT) to prevent cognitive impairment in Latin America (LA). Our aim is to present the study design and discuss the strategies used for multicultural harmonization. METHODS: This 1-year RCT (working on a 1-year extension) investigates the feasibility of a multi-domain lifestyle intervention in LA and the efficacy of the intervention, primarily on cognitive function. An external harmonization process was carried out to follow the FINGER model, and an internal harmonization was performed to ensure this study was feasible and comparable across the 12 participating LA countries. RESULTS: Currently, 1549 participants have been screened, and 815 randomized. Participants are ethnically diverse (56% are Nestizo) and have high cardiovascular risk (39% have metabolic syndrome). DISCUSSION: LatAm-FINGERS overcame a significant challenge to combine the region's diversity into a multi-domain risk reduction intervention feasible across LA while preserving the original FINGER design.

Generation of a human induced pluripotent stem cell line (INEUi001-A) from an amyotrophic lateral sclerosis/frontotemporal dementia patient with a C9ORF72 G4C2 genotype of <2 (GGGGCCG) and 10 repeats.

Human induced pluripotent stem cell (hiPSC) line INEUi001-A was reprogrammed from peripheral blood mononuclear cells (PBMC) using the lentiviral-hSTEMCCA-loxP vector. PBMCs were obtained from a 75- year-old female ALS/FTD disease patient carrying a heterozygous deletion within the C9ORF72 hexanucleotide repeat region resulting in a GGGGCCG sequence (∼1.16 repeats). C9ORF72 genotype was maintained and stemness and pluripotency confirmed in INEUi001-A hiPSC line.

Pathogenesis of Alzheimer's disease: Involvement of the choroid plexus.

The choroid plexus (ChP) produces and is bathed in the cerebrospinal fluid (CSF), which in aging and Alzheimer's disease (AD) shows extensive proteomic alterations including evidence of inflammation. Considering inflammation hampers functions of the involved tissues, the CSF abnormalities reported in these conditions are suggestive of ChP injury. Indeed, several studies document ChP damage in aging and AD, which nevertheless remains to be systematically characterized. We here report that the changes elicited in the CSF by AD are consistent with a perturbed aging process and accompanied by aberrant accumulation of inflammatory signals and metabolically active proteins in the ChP. Magnetic resonance imaging (MRI) imaging shows that these molecular aberrancies correspond to significant remodeling of ChP in AD, which correlates with aging and cognitive decline. Collectively, our preliminary post-mortem and in vivo findings reveal a repertoire of ChP pathologies indicative of its dysfunction and involvement in the pathogenesis of AD. HIGHLIGHTS: Cerebrospinal fluid changes associated with aging are perturbed in Alzheimer's disease Paradoxically, in Alzheimer's disease, the choroid plexus exhibits increased cytokine levels without evidence of inflammatory activation or infiltrates In Alzheimer's disease, increased choroid plexus volumes correlate with age and cognitive performance.

Acute severe hypoxia induces apoptosis of human pluripotent stem cells by a HIF-1α and P53 independent mechanism.

Human embryonic and induced pluripotent stem cells are self-renewing pluripotent stem cells (hPSCs) that can differentiate into a wide range of specialized cells. Although moderate hypoxia (5% O2) improves hPSC self-renewal, pluripotency, and cell survival, the effect of acute severe hypoxia (1% O2) on hPSC viability is still not fully elucidated. In this sense, we explore the consequences of acute hypoxia on hPSC survival by culturing them under acute (maximum of 24 h) physical severe hypoxia (1% O2). After 24 h of hypoxia, we observed HIF-1α stabilization concomitant with a decrease in cell viability. We also observed an increase in the apoptotic rate (western blot analysis revealed activation of CASPASE-9, CASPASE-3, and PARP cleavage after hypoxia induction). Besides, siRNA-mediated downregulation of HIF-1α and P53 did not significantly alter hPSC apoptosis induced by hypoxia. Finally, the analysis of BCL-2 family protein expression levels disclosed a shift in the balance between pro- and anti-apoptotic proteins (evidenced by an increase in BAX/MCL-1 ratio) caused by hypoxia. We demonstrated that acute physical hypoxia reduced hPSC survival and triggered apoptosis by a HIF-1α and P53 independent mechanism.

Noxa and Mcl-1 expression influence the sensitivity to BH3-mimetics that target Bcl-xL in patient-derived glioma stem cells.

The recurrence of Glioblastoma is partly attributed to the highly resistant subpopulation of glioma stem cells. A novel therapeutic approach focuses on restoring apoptotic programs in these cancer stem cells, as they are often deregulated. BH3-mimetics, targeting anti-apoptotic Bcl-2 family members, are emerging as promising compounds to sensitize cancer cells to antineoplastic treatments. Herein, we determined that the most abundantly expressed anti-apoptotic Bcl-2 family members, Bcl-xL and Mcl-1, are the most relevant in regulating patient-derived glioma stem cell survival. We exposed these cells to routinely used chemotherapeutic drugs and BH3-mimetics (ABT-263, WEHI-539, and S63845). We observed that the combination of BH3-mimetics targeting Bcl-xL with chemotherapeutic agents caused a marked increase in cell death and that this sensitivity to Bcl-xL inhibition correlated with Noxa expression levels. Interestingly, whereas co-targeting Bcl-xL and Mcl-1 led to massive cell death in all tested cell lines, down-regulation of Noxa promoted cell survival only in cell lines expressing higher levels of this BH3-only. Therefore, in glioma stem cells, the efficacy of Bcl-xL inhibition is closely associated with Mcl-1 activity and Noxa expression. Hence, a potentially effective strategy would consist of combining Bcl-xL inhibitors with chemotherapeutic agents capable of inducing Noxa, taking advantage of this pro-apoptotic factor.

Case Report: Progression of a Silent Corticotroph Tumor to an Aggressive Secreting Corticotroph Tumor, Treated by Temozolomide. Changes in the Clinic, the Pathology, and the ß-Catenin and α-SMA Expression.

Clinically silent corticotroph tumors are usually macroadenomas that comprise 20% of ACTH tumors. They frequently progress to aggressive tumors with high recurrence, invasiveness, and on rare occasions, they may become hormonally active causing Cushing's disease. Trustable biomarkers that can predict their aggressive course, as well as their response to traditional or new therapies, are paramount. Aberrant ß-Catenin expression and localization have been proposed as responsible for several malignancies including pituitary tumors. Nevertheless, the role of ß-Catenin in the aggressive transformation of silent corticotropinomas and their response to Temozolomide salvage treatment have not been explored yet. In this work, we present a case of a silent corticotroph tumor that invaded cavernous sinus and compressed optic chiasm and, after a first total resection and tumor remission it recurred six years later as an aggressive ACTH-secreting tumor. This lesion grew with carotid compromise and caused Cushing's signs. It required multiple medical treatments including Cabergoline, Ketoconazole, TMZ, and radiotherapy. Besides, other two surgeries were needed until it could be controlled. Interestingly, we found α-SMA vascular area reduction and differential ß-Catenin cell localization in the more aggressive tumor stages characterized by high Ki-67 indexes and p53 expression. Our results may indicate a role of angiogenesis and ß-Catenin trigged events in the pituitary tumor progression, which could in turn affect the response to TMZ and/or conventional treatments. These molecular findings in this unusual case could be useful for future management of aggressive pituitary tumors.

ß-Catenin is reduced in membranes of human prolactinoma cells and it is inhibited by temozolomide in prolactin secreting tumor models.

INTRODUCTION: Prolactinomas are the most frequent pituitary tumor subtype. Despite most of them respond to medical treatment, a proportion are resistant and become a challenge in clinical management. Wnt/ß-Catenin pathway has been implicated in several cancers including pituitary tumors and other sellar region malignancies. Interestingly, Wnt/ß-Catenin inhibition augments the cytotoxicity of the chemotherapeutic agent Temozolomide (TMZ) in different cancers. TMZ is now being implemented as rescue therapy for aggressive pituitary adenoma treatment. However, the molecular mechanisms associated with TMZ action in pituitary tumors remain unclear. OBJECTIVES: Our aims in the present study were to evaluate differential ß-Catenin expression in human resistant prolactinomas and Wnt/ß-Catenin signaling activation and involvement in Prolactin (PRL) secreting experimental models treated with TMZ. RESULTS: We first evaluated by immunohistochemistry ß-Catenin localization in human resistant prolactinomas in which we demonstrated reduced membrane ß-Catenin in prolactinoma cells compared to normal pituitaries, independently of the Ki-67 proliferation indexes. In turn, in vivo 15 mg/kg of orally administered TMZ markedly reduced PRL production and increased prolactinoma cell apoptosis in mice bearing xenografted prolactinomas. Intratumoral ß-Catenin strongly correlated with Prl and Cyclin D1, and importantly, TMZ downregulated both ß-Catenin and Cyclin D1, supporting their significance in prolactinoma growth and as candidates of therapeutic targets. When tested in vitro, TMZ directly reduced MMQ cell viability, increased apoptosis and produced G2/M cell cycle arrest. Remarkably, ß-Catenin activation and VEGF secretion were inhibited by TMZ in vitro. CONCLUSIONS: We concluded that dopamine resistant prolactinomas undergo a ß-Catenin relocalization in relation to normal pituitaries and that TMZ restrains experimental prolactinoma tumorigenicity by reducing PRL production and ß-Catenin activation. Together, our findings contribute to the understanding of Wnt/ß-Catenin implication in prolactinoma maintenance and TMZ therapy, opening the opportunity of new treatment strategies for aggressive and resistant pituitary tumors.

Protocol for morphometric analysis of neurons derived from human pluripotent stem cells.

The analysis of morphological features of neurons derived from human pluripotent stem cells (hPSCs) is important to describe neuronal phenotypes and changes observed throughout development. Using free and easily accessible tools, we describe a protocol for the morphometric quantification of hPSCs-derived neurons in two- and three-dimensions in vitro cultures. We detail the analysis of soma area and main and secondary dendrites lengths of GFP-transfected neurons and the measurement of area and perimeter of immunostained neurospheres.

Characteristics of children ≤36 months of age with DIPG: A report from the international DIPG registry.

BACKGROUND: Children ≤36 months with diffuse intrinsic pontine glioma (DIPG) have increased long-term survival (LTS, overall survival (OS) ≥24 months). Understanding distinguishing characteristics in this population is critical to improving outcomes. METHODS: Patients ≤36 months at diagnosis enrolled on the International DIPG Registry (IDIPGR) with central imaging confirmation were included. Presentation, clinical course, imaging, pathology and molecular findings were analyzed. RESULTS: Among 1183 patients in IDIPGR, 40 were eligible (median age: 29 months). Median OS was 15 months. Twelve patients (30%) were LTS, 3 (7.5%) very long-term survivors ≥5 years. Among 8 untreated patients, median OS was 2 months. Patients enrolled in the registry but excluded from our study by central radiology review or tissue diagnosis had median OS of 7 months. All but 1 LTS received radiation. Among 32 treated patients, 1-, 2-, 3-, and 5-year OS rates were 68.8%, 31.2%, 15.6% and 12.5%, respectively. LTS had longer duration of presenting symptoms (P = .018). No imaging features were predictive of outcome. Tissue and genomic data were available in 18 (45%) and 10 patients, respectively. Among 9 with known H3K27M status, 6 had a mutation. CONCLUSIONS: Children ≤36 months demonstrated significantly more LTS, with an improved median OS of 15 months; 92% of LTS received radiation. Median OS in untreated children was 2 months, compared to 17 months for treated children. LTS had longer duration of symptoms. Excluded patients demonstrated a lower OS, contradicting the hypothesis that children ≤36 months with DIPG show improved outcomes due to misdiagnosis.

Diabetic patients treated with metformin during early stages of Alzheimer's disease show a better integral performance: data from ADNI study.

We evaluated the effect of the antidiabetic drug metformin on patients enrolled in the ADNI study considering patients with mild cognitive impairment (MCI) due to Alzheimer's disease (AD). Employing data from this observational study, we performed a principal component analysis focusing on the cognitive sphere by evaluating data from neuropsychological tests included in a modified version of the Alzheimer's Disease Cooperative Study-Preclinical Alzheimer Cognitive Composite (ADCS-PACC). Second, we included the levels of amyloid-ß, tau, and phosphorylated tau in CSF. We found that MCI metformin-treated patients were globally characterized as subjects with a better cognitive performance and CSF biomarkers profile than the mean population of MCI patients. On the other hand, control subjects and type 2 diabetes patients (T2D) were paired by age, gender, ApoE allele, and years of education, defining three groups: MCI, MCI + T2D, and MCI + T2D + metformin. We evaluated the effect of T2D and metformin treatment employing the PACC score and composites defined from standardized ADNI variables to evaluate the memory and learning function. We found that MCI + T2D patients had a worse cognitive performance than MCI patients, but this deleterious effect was not observed in MCI + T2D + metformin patients. These cognitive variations were associated with changes in cortical thickness and hippocampal volume. Finally, no differences were found in metabolic plasmatic parameters (glycemia, cholesterol, triglycerides). Our study-employing different strategies for data analysis from the global study ADNI-shows a beneficial effect of metformin treatment on cognitive performance, CSF biomarkers profile, and neuroanatomical measures in MCI due to AD patients.

Neuropsychological profile of Alzheimer's disease based on amyloid biomarker findings results from a South American cohort.

OBJECTIVE: Increased life expectancy and exponential growth of adults suffering from Alzheimer's disease (AD) worldwide, has led to biomarkers incorporation for diagnosis in early stages. Use of neuropsychological testing remains limited. This study aimed to identify which neuropsychological tests best indicated underlying AD pathophysiology. METHODS: One hundred and forty-one patients with MCI (Mild Cognitive Impairment) were studied. A neuropsychological test battery based on the Uniform Data Set (UDS) from the Alzheimer's Disease Centers program of the National Institute on Aging (NIA) was performed and amyloid markers recorded; according to presence or absence of amyloid identified by positive PIB-PET findings, or low CSF Aß42 levels, patients were separated into MCI amyloid-(n:58) and MCI amyloid + (n = 83) cases. RESULTS: Statistical differences were found in all memory tests between groups. Delayed recall score at thirty minutes on the Rey Auditory Verbal Learning Test (AVLT) was the best predictor of amyloid pathology presence (AUC 0.68), followed by AVLT total learning (AUC 0.66) and AVLT Recognition (AUC 0.59) scores, providing useful cut off values in the clinical setting. CONCLUSIONS: Use of neuropsychological testing, specifically AVLT scores with cutoff values, contributed to the correct diagnosis of MCI due to AD in this SouthAmerican cohort.

Accuracy of central neuro-imaging review of DIPG compared with histopathology in the International DIPG Registry.

BACKGROUND: Diffuse intrinsic pontine glioma (DIPG) remains a clinico-radiologic diagnosis without routine tissue acquisition. Reliable imaging distinction between DIPG and other pontine tumors with potentially more favorable prognoses and treatment considerations is essential. METHODS: Cases submitted to the International DIPG registry (IDIPGR) with histopathologic and/or radiologic data were analyzed. Central imaging review was performed on diagnostic brain MRIs (if available) by two neuro-radiologists. Imaging features suggestive of alternative diagnoses included nonpontine origin, <50% pontine involvement, focally exophytic morphology, sharply defined margins, and/or marked diffusion restriction throughout. RESULTS: Among 286 patients with pathology from biopsy and/or autopsy, 23 (8%) had histologic diagnoses inconsistent with DIPG, most commonly nondiffuse low-grade gliomas and embryonal tumors. Among 569 patients with centrally-reviewed diagnostic MRIs, 40 (7%) were classified as non-DIPG, alternative diagnosis suspected. The combined analysis included 151 patients with both histopathology and centrally-reviewed MRI. Of 77 patients with imaging classified as characteristic of DIPG, 76 (99%) had histopathologic diagnoses consistent with DIPG (infiltrating grade II-IV gliomas). Of 57 patients classified as likely DIPG with some unusual imaging features, 55 (96%) had histopathologic diagnoses consistent with DIPG. Of 17 patients with imaging features suggestive of an alternative diagnosis, eight (47%) had histopathologic diagnoses inconsistent with DIPG (remaining patients were excluded due to nonpontine tumor origin). Association between central neuro-imaging review impression and histopathology was significant (p < 0.001), and central neuro-imaging impression was prognostic of overall survival. CONCLUSIONS: The accuracy and important role of central neuro-imaging review in confirming the diagnosis of DIPG is demonstrated.

Amyloid and anatomical correlates of executive functioning in middle-aged offspring of patients with late-onset Alzheimer's disease.

A traditional hallmark of cognitive impairment associated with late-onset Alzheimer´s disease (LOAD) is episodic memory impairment. However, early alterations have been identified in brain regions associated with executive function in asymptomatic, middle-age offspring of patients with LOAD (O-LOAD) compared to those with no family history. We hypothesized that executive function among O-LOAD would correlate with structural and amyloid brain imaging differently from those without a family history of LOAD (control subjects, CS). Executive function, cortical thickness, and in-vivo Aß deposits were quantified in 30 O-LOAD and 25 CS. Associations were observed among O-LOAD only. Cortical thickness in the left lateral orbitofrontal cortex was positively associated with Design Fluency. The Stroop Color and Word Test, correlated positively with right rostral mid-frontal cortex thickness. Trails Making Test-B was inversely related to left medial orbitofrontal thickness. Tower of London total time was positively associated with ß-amyloid deposition in the right precuneus. These results support previous evidence that early executive dysfunction might reflect subtle, early changes in persons at risk of LOAD and suggests that executive function alterations deserve further exploration in the LOAD literature.

Pseudotumoral Presentation of Cerebral Amyloid-Beta Angiopathy: Case Report and Review of Literature.

OBJECTIVE: Cerebral amyloid angiopathy-related inflammation (CAA-RI) is a rare and potentially treatable encephalopathy that usually affects people older than 50 years old and has an acute or subacute clinical presentation characterized by rapidly evolving cognitive decline, focal deficits and seizures. In a small subset of patients the disease can adopt a pseudotumoral form in the neuroimages that represents a very difficult diagnostic challenge. METHODS: Here in we report a patient with a tumour-like presentation of histopathologically confirmed CAA-RI. RESULTS: We also conducted a search and reviewed the clinical and radiological features of 41 cases of pseudotumoral CAA-RI previously reported in the literature in order to identify those characteristics that should raise diagnostic suspicions of the disease, there by avoiding unnecessary surgical treatments. CONCLUSION: The therapy of CAA-RI with steroids is usually effective and clinical and radiological remission can be achieved in the first month in approximately 70% of cases.

celldeath: A tool for detection of cell death in transmitted light microscopy images by deep learning-based visual recognition.

Cell death experiments are routinely done in many labs around the world, these experiments are the backbone of many assays for drug development. Cell death detection is usually performed in many ways, and requires time and reagents. However, cell death is preceded by slight morphological changes in cell shape and texture. In this paper, we trained a neural network to classify cells undergoing cell death. We found that the network was able to highly predict cell death after one hour of exposure to camptothecin. Moreover, this prediction largely outperforms human ability. Finally, we provide a simple python tool that can broadly be used to detect cell death.

Generation of a human induced pluripotent stem cell line from a familial Alzheimer's disease PSEN1 T119I patient.

Human induced pluripotent stem cells (hiPSC) line FLENIi001-A was reprogrammed from dermal fibroblasts using the lentiviral-hSTEMCCA-loxP vector. Fibroblasts were obtained from a skin biopsy of a 72-year-old Caucasian male familial Alzheimer's disease patient carrying the T119I mutation in the PSEN1 gene. PSEN1 genotype was maintained and stemness and pluripotency confirmed in the FLENIi001-A hiPSC line.