Transplant of GABAergic Precursors Restores Hippocampal Inhibitory Function in a Mouse Model of Seizure Susceptibility

Defects in GABAergic function can cause epilepsy. in the last years, cell-based therapies have attempted to correct these defects with disparate success on animal models of epilepsy. Recently, we demonstrated that medial ganglionic eminence (MGE)-derived cells grafted into the neonatal normal brain migrate and differentiate into functional mature GABAergic interneurons. These cells are able to modulate the local level of GABA-mediated synaptic inhibition, which suggests their suitability for cell-based therapies. However, it is unclear whether they can integrate in the host circuitry and rescue the loss of inhibition in pathological conditions. Thus, as proof of principle, we grafted MGE-derived cells into a mouse model of seizure susceptibility caused by specific elimination of GABAergic interneuron subpopulations in the mouse hippocampus after injection of the neurotoxic saporin conjugated to substance P (SSP-Sap). This ablation was associated with significant decrease in inhibitory postsynaptic currents (IPSC) on CA1 pyramidal cells and increased seizure susceptibility induced by pentylenetetrazol (PTZ). Grafting of GFP(+) MGE-derived cells in SSP-Sap-treated mice repopulates the hippocampal ablated zone with cells expressing molecular markers of mature interneurons. Interestingly, IPSC kinetics on CA1 pyramidal cells of ablated hippocampus significantly increased after transplantation, reaching levels similar to the normal mice. More importantly, this was associated with reduction in seizure severity and decrease in postseizure mortality induced by PTZ. Our data show that MGE-derived cells fulfill most of the requirements for an appropriate cell-based therapy, and indicate their suitability for neurological conditions where a modulation of synaptic inhibition is needed, such as epilepsy.Spanish Ministry of Science and InnovationCIPFCarlos III Institute (Spanish Ministry of Science and Innovation)Generalitat ValencianaAndalusian Ctr Mol Biol & Regenerat Med CABIMER, Dept Cell Therapy & Regenerat Med, Seville, SpainCIPF, Valencia, SpainUniversidade Federal de São Paulo, Dept Physiol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Physiol, São Paulo, BrazilSpanish Ministry of Science and Innovation: SAF 07/61880Spanish Ministry of Science and Innovation: FIS 07/0079Web of Scienc

Analysis of the Ush2a Gene in Medaka Fish (Oryzias latipes)

Patients suffering from Usher syndrome (USH) exhibit sensorineural hearing loss, retinitis pigmentosa (RP) and, in some cases, vestibular dysfunction. USH is the most common genetic disorder affecting hearing and vision and is included in a group of hereditary pathologies associated with defects in ciliary function known as ciliopathies. This syndrome is clinically classified into three types: USH1, USH2 and USH3. USH2 accounts for well over one-half of all Usher cases and mutations in the USH2A gene are responsible for the majority of USH2 cases, but also for atypical Usher syndrome and recessive non-syndromic RP. Because medaka fish (Oryzias latypes) is an attractive model organism for genetic-based studies in biomedical research, we investigated the expression and function of the USH2A ortholog in this teleost species. Ol-Ush2a encodes a protein of 5.445 aa codons, containing the same motif arrangement as the human USH2A. Ol-Ush2a is expressed during early stages of medaka fish development and persists into adulthood. Temporal Ol-Ush2a expression analysis using whole mount in situ hybridization (WMISH) on embryos at different embryonic stages showed restricted expression to otoliths and retina, suggesting that Ol-Ush2a might play a conserved role in the development and/or maintenance of retinal photoreceptors and cochlear hair cells. Knockdown of Ol-Ush2a in medaka fish caused embryonic developmental defects (small eyes and heads, otolith malformations and shortened bodies with curved tails) resulting in late embryo lethality. These embryonic defects, observed in our study and in other ciliary disorders, are associated with defective cell movement specifically implicated in left-right (LR) axis determination and planar cell polarity (PCP)

Exposure to N-Ethyl-N-Nitrosourea in Adult Mice Alters Structural and Functional Integrity of Neurogenic Sites

BACKGROUND: Previous studies have shown that prenatal exposure to the mutagen N-ethyl-N-nitrosourea (ENU), a N-nitroso compound (NOC) found in the environment, disrupts developmental neurogenesis and alters memory formation. Previously, we showed that postnatal ENU treatment induced lasting deficits in proliferation of neural progenitors in the subventricular zone (SVZ), the main neurogenic region in the adult mouse brain. The present study is aimed to examine, in mice exposed to ENU, both the structural features of adult neurogenic sites, incorporating the dentate gyrus (DG), and the behavioral performance in tasks sensitive to manipulations of adult neurogenesis. METHODOLOGY/PRINCIPAL FINDINGS: 2-month old mice received 5 doses of ENU and were sacrificed 45 days after treatment. Then, an ultrastructural analysis of the SVZ and DG was performed to determine cellular composition in these regions, confirming a significant alteration. After bromodeoxyuridine injections, an S-phase exogenous marker, the immunohistochemical analysis revealed a deficit in proliferation and a decreased recruitment of newly generated cells in neurogenic areas of ENU-treated animals. Behavioral effects were also detected after ENU-exposure, observing impairment in odor discrimination task (habituation-dishabituation test) and a deficit in spatial memory (Barnes maze performance), two functions primarily related to the SVZ and the DG regions, respectively. CONCLUSIONS/SIGNIFICANCE: The results demonstrate that postnatal exposure to ENU produces severe disruption of adult neurogenesis in the SVZ and DG, as well as strong behavioral impairments. These findings highlight the potential risk of environmental NOC-exposure for the development of neural and behavioral deficits

EPIdemiology of Surgery-Associated Acute Kidney Injury (EPIS-AKI) : Study protocol for a multicentre, observational trial

More than 300 million surgical procedures are performed each year. Acute kidney injury (AKI) is a common complication after major surgery and is associated with adverse short-term and long-term outcomes. However, there is a large variation in the incidence of reported AKI rates. The establishment of an accurate epidemiology of surgery-associated AKI is important for healthcare policy, quality initiatives, clinical trials, as well as for improving guidelines. The objective of the Epidemiology of Surgery-associated Acute Kidney Injury (EPIS-AKI) trial is to prospectively evaluate the epidemiology of AKI after major surgery using the latest Kidney Disease: Improving Global Outcomes (KDIGO) consensus definition of AKI. EPIS-AKI is an international prospective, observational, multicentre cohort study including 10 000 patients undergoing major surgery who are subsequently admitted to the ICU or a similar high dependency unit. The primary endpoint is the incidence of AKI within 72 hours after surgery according to the KDIGO criteria. Secondary endpoints include use of renal replacement therapy (RRT), mortality during ICU and hospital stay, length of ICU and hospital stay and major adverse kidney events (combined endpoint consisting of persistent renal dysfunction, RRT and mortality) at day 90. Further, we will evaluate preoperative and intraoperative risk factors affecting the incidence of postoperative AKI. In an add-on analysis, we will assess urinary biomarkers for early detection of AKI. EPIS-AKI has been approved by the leading Ethics Committee of the Medical Council North Rhine-Westphalia, of the Westphalian Wilhelms-University Münster and the corresponding Ethics Committee at each participating site. Results will be disseminated widely and published in peer-reviewed journals, presented at conferences and used to design further AKI-related trials. Trial registration number NCT04165369

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Preparation of porcelain tile granulates by more environmentally sustainable processes

This study examines the feasibility of manufacturing glazed porcelain tiles with a more environmentally friendly manufacturing process, by reducing water and thermal energy consumption. The process studied in this paper is dry milling in a pendulum mill, with subsequent granulation (in order to obtain a press powder with similar flowability to that of spraydried powders). 
 The different morphology of the new granulate with respect to the standard spray-dried granulate modifies the microstructure of the green compacts and thus, their behaviour and fired tile properties. In order to obtain porcelain tiles with the required properties (water absorption, mechanical strength,…) changes have been made in the raw materials mixture and in the processing variables. 
 Finally, porcelain tiles measuring 50x50 cm have been manufactured at industrial scale with the new granulate using a conventional firing cycle, obtaining quality levels identical to those provided by the spray-dried granulate. These results open the possibility of preparing porcelain tile body compositions through a manufacturing process alternative to the standard one, more environmentally friendly and with lower costs.<br><br>En el presente trabajo se ha estudiado la viabilidad de fabricar gres porcelánico esmaltado utilizando un sistema de preparación de la composición del soporte más respetuoso con el medio ambiente, lo que implica una reducción importante de los consumos de agua y de energía térmica. El proceso que se estudia en el presente trabajo es el consistente en la molienda vía seca en molino pendular y en la posterior granulación (para obtener un polvo de prensas con fluidez similar a la de los polvos atomizados).
 La distinta morfología de los nuevos gránulos obtenidos respecto al polvo atomizado actual, modifica la microestuctura en crudo de las piezas y, con ello, el comportamiento y propiedades finales de las baldosas obtenidas. Por ello, ha sido necesario modificar la mezcla de materias primas y ajustar algunas variables de operación para obtener baldosas de gres porcelánico con las propiedades requeridas (absorción de agua, resistencia mecánica, etc.). 
 Finalmente, se han fabricado industrialmente piezas de gres porcelánico (50x50 cm<sup>2</sup>) con el nuevo granulado, mediante un ciclo de cocción convencional, obteniendo unos niveles de calidad idénticos a los que proporciona el polvo atomizado. Estos resultados abren la posibilidad de preparar las composiciones de soporte de gres porcelánico mediante un sistema alternativo al actual, más respetuoso con el medio ambiente y con un menor coste

Cell fusion contributes to pericyte formation after stroke

6 páginas, 2 figuras.Recent reports have shown that bone marrow-derived cells (BMDCs) contribute to the formation of vasculature after stroke. However, the mechanism by which mural cells are formed from BMDC remains elusive. Here, we provide direct evidence that the cell fusion process contributes to the formation of pericytes after stroke. We generated mouse bone marrow chimeras using a cre/lox system that allows the detection of fusion events by X-gal staining. In these mice, we detected X-gal-positive cells that expressed vimentin and desmin, specific markers of mature murine pericytes. Electron microscopy confirmed that fused cells possessed basal lamina and characteristics of pericytes. Furthermore, induction of stroke increased significantly the presence of fused cells in the ischemic area. These cells expressed markers of developing pericytes such as NG2. We conclude that cell fusion participates actively in the generation of vascular tissue through pericyte formation under normal as well as pathologic conditions.This work was supported by grants from Spanish Ministry of Health (FIS 04/2744) and Regenerative Medicine Programme from the CIPF. M.P-G and IZ were recipients of PhD fellowships from CIPF and Generalitat Valenciana, respectively.Peer reviewe

Enhanced Hematovascular Contribution of SCL 3′ Enhancer Expressing Fetal Liver Cells Uncovers Their Potential to Integrate in Extramedullary Adult Niches

13 páginas, 6 figuras, 1 tabla.-- et al.Fetal liver (FL) hematopoietic progenitors have superior blood engraftment competence compared with adult bone marrow (BM), however less is known about FL in vivo vascular capacity. Here we show in transplantation assays that FL cells possess enhanced vascular endothelial potential compared with adult bone marrow. We generated high-level hematopoietic chimeras using donor cells from mice transgenic for the stem cell leukaemia 3′ enhancer human placental alkaline phosphatase (SCL3′Enh-PLAP) reporter construct, active in vascular endothelium, and blood progenitor and stem cells. Long-term lineage tracing analysis revealed PLAP+ vascular-like patches in FL-derived chimeras, whereas adult BM-derived chimeras presented only rare and scattered PLAP+ cells. PLAP+ vascular-like patches were formed following transplantation into both newborn and adult recipient mice, although their frequency was reduced in adult recipients. Confocal analysis of multiple labeled tissues revealed that whereas most liver and heart PLAP+ vascular patch-associated cells were endothelial, PLAP+ vascular patches in the kidney contained endothelial, hematopoietic, and putative hemangioblastic cells. Moreover, fluorescence-activated cell sorting assays showed that only FL PLAPbright+ donor cells can generate PLAP+ vascular patches upon transplantation. Taken together, these data demonstrate superior vascular contribution potential of FL cells, and not only provide new insights into the developmental pathways controlling endothelial development but also may prove informative when addressing the mechanisms involved in vascular regeneration and hemangiogenic recovery in a clinical context.This work was supported by the Spanish Ministry of Education and Science Grant SAF64679, Junta de Andalucia Grants PAI-BIO-295 and PAIDI-CTS1614, CONSOLIDER INGENIO 2010 Grant, the Leukemia Research Fund, fellowship CONACYT-179065 to A.M.G.-O., fellowship I3P-CSIC to C.Q., and fellowship UPO to A.C. A.M.G.-O. and A.C. contributed equally to this work.Peer reviewe

Enhanced hemato-vascular contribution of SCL 3′ enhancer expressing fetal liver cells uncovers their potential to integrate in extramedullary adult niches

Fetal liver (FL) hematopoietic progenitors have superior blood engraftment competence compared with adult bone marrow (BM), however less is known about FL in vivo vascular capacity. Here we show in transplantation assays that FL cells possess enhanced vascular endothelial potential compared with adult bone marrow. We generated high‐level hematopoietic chimeras using donor cells from mice transgenic for the stem cell leukaemia 3′ enhancer human placental alkaline phosphatase (SCL3′Enh‐PLAP) reporter construct, active in vascular endothelium, and blood progenitor and stem cells. Long‐term lineage tracing analysis revealed PLAP+ vascular‐like patches in FL‐derived chimeras, whereas adult BM‐derived chimeras presented only rare and scattered PLAP+ cells. PLAP+ vascular‐like patches were formed following transplantation into both newborn and adult recipient mice, although their frequency was reduced in adult recipients. Confocal analysis of multiple labeled tissues revealed that whereas most liver and heart PLAP+ vascular patch‐associated cells were endothelial, PLAP+ vascular patches in the kidney contained endothelial, hematopoietic, and putative hemangioblastic cells. Moreover, fluorescence‐activated cell sorting assays showed that only FL PLAPbright+ donor cells can generate PLAP+ vascular patches upon transplantation. Taken together, these data demonstrate superior vascular contribution potential of FL cells, and not only provide new insights into the developmental pathways controlling endothelial development but also may prove informative when addressing the mechanisms involved in vascular regeneration and hemangiogenic recovery in a clinical context

Enhanced Hematovascular Contribution of SCL 3′ Enhancer Expressing Fetal Liver Cells Uncovers Their Potential to Integrate in Extramedullary Adult Niches

13 páginas, 6 figuras, 1 tabla.-- et al.Fetal liver (FL) hematopoietic progenitors have superior blood engraftment competence compared with adult bone marrow (BM), however less is known about FL in vivo vascular capacity. Here we show in transplantation assays that FL cells possess enhanced vascular endothelial potential compared with adult bone marrow. We generated high-level hematopoietic chimeras using donor cells from mice transgenic for the stem cell leukaemia 3′ enhancer human placental alkaline phosphatase (SCL3′Enh-PLAP) reporter construct, active in vascular endothelium, and blood progenitor and stem cells. Long-term lineage tracing analysis revealed PLAP+ vascular-like patches in FL-derived chimeras, whereas adult BM-derived chimeras presented only rare and scattered PLAP+ cells. PLAP+ vascular-like patches were formed following transplantation into both newborn and adult recipient mice, although their frequency was reduced in adult recipients. Confocal analysis of multiple labeled tissues revealed that whereas most liver and heart PLAP+ vascular patch-associated cells were endothelial, PLAP+ vascular patches in the kidney contained endothelial, hematopoietic, and putative hemangioblastic cells. Moreover, fluorescence-activated cell sorting assays showed that only FL PLAPbright+ donor cells can generate PLAP+ vascular patches upon transplantation. Taken together, these data demonstrate superior vascular contribution potential of FL cells, and not only provide new insights into the developmental pathways controlling endothelial development but also may prove informative when addressing the mechanisms involved in vascular regeneration and hemangiogenic recovery in a clinical context.This work was supported by the Spanish Ministry of Education and Science Grant SAF64679, Junta de Andalucia Grants PAI-BIO-295 and PAIDI-CTS1614, CONSOLIDER INGENIO 2010 Grant, the Leukemia Research Fund, fellowship CONACYT-179065 to A.M.G.-O., fellowship I3P-CSIC to C.Q., and fellowship UPO to A.C. A.M.G.-O. and A.C. contributed equally to this work.Peer reviewe