The loss of antioxidant activities impairs intestinal epithelium homeostasis by altering lipid metabolism.

Reactive oxygens species (ROS) are common byproducts of metabolic reactions and could be at the origin of many diseases of the elderly. Here we investigated the role of ROS in the renewal of the intestinal epithelium in mice lacking catalase (CAT) and/or nicotinamide nucleotide transhydrogenase (NNT) activities. Cat-/- mice have delayed intestinal epithelium renewal and were prone to develop necrotizing enterocolitis upon starvation. Interestingly, crypts lacking CAT showed fewer intestinal stem cells (ISC) and lower stem cell activity than wild-type. In contrast, crypts lacking NNT showed a similar number of ISCs as wild-type but increased stem cell activity, which was also impaired by the loss of CAT. No alteration in the number of Paneth cells (PCs) was observed in crypts of either Cat-/- or Nnt-/- mice, but they showed an evident decline in the amount of lysozyme. Cat deficiency caused fat accumulation in crypts, and a fall in the remarkable high amount of adipose triglyceride lipase (ATGL) in PCs. Notably, the low levels of ATGL in the intestine of Cat -/- mice increased after a treatment with the antioxidant N-acetyl-L-cysteine. Supporting a role of ATGL in the regulation of ISC activity, its inhibition halt intestinal organoid development. These data suggest that the reduction in the renewal capacity of intestine originates from fatty acid metabolic alterations caused by peroxisomal ROS.

Survival of Vascular Accesses in Chronic Hemodialysis Patients.

BACKGROUND/AIMS: Vascular access (VA) is the highest risk factor for blood infections, hospitalization, and mortality of patients undergoing hemodialysis (HD). The risk of mortality while using a catheter is greater than that while using grafts. The objective of this article is to know the survival rate in relation to the type of VA. METHODS: A retrospective cohort of HD patients was studied. The data gathered included age, gender, first VA at the surrogate site, days between the first and second access, number of accesses, and anatomical site of VA placement. Mean differences were estimated using χ2 or Student's t test. Survival was calculated using the Kaplan-Meier curves and included in tables. Statistical significance was established as p < 0.05. The statistical computer software package SPSSw v25 was used for the analysis. RESULTS: A total of 896 patients were included with a mean age of 47.88 years (SD ± 16.52), the duration of the first VA was 398.81 days (±565.79), the mean number of VAs used was 2.26 (±1.15), and the median time undergoing HD was 728.73 days. The duration of catheter placement was 330.42 days, and 728.60 days for fistula use (p = 0.001). The mean number of days of renal replacement was 611.59 days for catheter and 1,495.25 days for internal arteriovenous fistula (IAVF) patients (p = 0.001). CONCLUSIONS: The survival of the initial VA is greater for the IAVF, followed by the tunneled catheters and the lowest by the non-tunneled catheters, which continue to be frequently used in our setting.

Establishing consensus on the perioperative management of cholecystectomy in public hospitals: a Delphi study with an expert panel in Mexico.

BACKGROUND: Several guidelines have put forward recommendations about the perioperative process of cholecystectomy. Despite the recommendations, controversy remains concerning several topics, especially in low- and middle-income countries. The aim of this study was to develop uniform recommendations for perioperative practices in cholecystectomy in Mexico to standardize this process and save public health system resources. METHODS: A modified Delphi method was used. An expert panel of 23 surgeons anonymously completed two rounds of responses to a 29-item questionnaire with 110 possible answers. The consensus was assessed using the percentage of responders agreeing on each question. RESULTS: From the 29 questions, the study generated 27 recommendations based on 20 (69.0%) questions reaching consensus, one that was considered uncertain (3.4%), and six (20.7%) items that remained open questions. In two (6.9%) cases, no consensus was reached, and no recommendation could be made. CONCLUSIONS: This study provides recommendations for the perioperative management of cholecystectomy in public hospitals in Mexico. As a guide for public institutions in low- and middle-income countries, the study identifies recommendations for perioperative tests and evaluations, perioperative decision making, postoperative interventions and institutional investment, that might ensure the safe practice of cholecystectomy and contribute to conserving resources.

The E6/E7 oncogenes of human papilloma virus and estradiol regulate hedgehog signaling activity in a murine model of cervical cancer.

Human papilloma virus oncogenes and estradiol are major etiologic factors associated with cervical cancer. In order to understand the mechanism by which these two factors promote carcinogenesis, the role of the Hedgehog (Hh) signaling pathway was evaluated during the normal growth of cervical epithelium and in the presence of E6/E7 oncogenes and exogenous estradiol. Hh signaling activity was determined in live animals (i.e., Gli-Luc reporter levels) during the estrous cycle and was found to be higher in the cervical area during the major growth phases, proestrus-estrus, in comparison to the diestrus phase. The same pattern was observed in transgenic mice expressing the E6/E7 oncogenes, though with notably higher levels than in control mice. Adding estradiol also markedly increased Gli activity in the cervix and the skin. In agreement with the correlation between high bioluminescence and tissue growth in different context, cervical cell proliferation was reduced upon Hh signaling inhibition in mice. Treatment with itraconazole, a putative novel Hh inhibitor, at an early stage of cervical carcinogenesis, did not decrease Hh signaling but it did reduce growth. Therefore, Hh signaling likely contributes to cervical carcinogenesis and itraconazole is effective to reduce growth but by a mechanism involving additional signaling pathways.

Functional determination of the differentiation potential of ventral mesencephalic neural precursor cells during dopaminergic neurogenesis.

The ventral mesencephalic neural precursor cells (vmNPCs) that give rise to dopaminergic (DA) neurons have been identified by the expression of distinct genes (e.g., Lmx1a, Foxa2, Msx1/2). However, the commitment of these NPCs to the mesencephalic DA neuronal fate has not been functionally determined. Evaluation of the plasticity of vmNPCs suggests that their commitment occurs after E10.5. Here we show that E9.5 vmNPCs implanted in an ectopic area of E10.5 mesencephalic explants, retained their specification marker Lmx1a and efficiently differentiated into neurons but did not express the gene encoding tyrosine hydroxylase (Th), the limiting enzyme for dopamine synthesis. A proportion of E10.5-E11.5 implanted vmNPCs behaved as committed, deriving into Th+ neurons in ectopic sites. Interestingly, implanted cells from E12.5 embryos were unable to give rise to a significant number of Th+ neurons. Concomitantly, differentiation assays in culture and in mesencephalic explants treated with Fgf2+LIF detected vmNPCs with astrogenic potential since E11.5. Despite this, a full suspension of E12.5 vmNPCs give rise to DA neurons in a similar proportion as those of E10.5 when they were transplanted into adult brain, but astrocytes were only detected with the former population. These data suggest that the subventricular postmitotic progenitors present in E12.5 ventral mesencephalon are unable to implant in embryonic explants and are the source of DA neurons in the transplanted adult brain. Based on our findings we propose that during DA differentiation committed vmNPCs emerge at E10.5 and they exhaust their neurogenic capacity with the rise of NPCs with astrogenic potential.

Mouse embryonic stem cell-derived cells reveal niches that support neuronal differentiation in the adult rat brain.

A neurogenic niche can be identified by the proliferation and differentiation of its naturally residing neural stem cells. However, it remains unclear whether "silent" neurogenic niches or regions suitable for neural differentiation, other than the areas of active neurogenesis, exist in the adult brain. Embryoid body (EB) cells derived from embryonic stem cells (ESCs) are endowed with a high potential to respond to specification and neuralization signals of the embryo. Hence, to identify microenvironments in the postnatal and adult rat brain with the capacity to support neuronal differentiation, we transplanted dissociated EB cells to conventional neurogenic and non-neurogenic regions. Our results show a neuronal differentiation pattern of EB cells that was dependent on the host region. Efficient neuronal differentiation of EB cells occurred within an adjacent region to the rostral migratory stream. EB cell differentiation was initially patchy and progressed toward an even distribution along the graft by 15-21 days post-transplantation, giving rise mostly to GABAergic neurons. EB cells in the striatum displayed a lower level of neuronal differentiation and derived into a significant number of astrocytes. Remarkably, when EB cells were transplanted to the striatum of adult rats after a local ischemic stroke, increased number of neuroblasts and neurons were observed. Unexpectedly, we determined that the adult substantia nigra pars compacta, considered a non-neurogenic area, harbors a robust neurogenic environment. Therefore, neurally uncommitted cells derived from ESCs can detect regions that support neuronal differentiation within the adult brain, a fundamental step for the development of stem cell-based replacement therapies.

Full regeneration of the tribasal Polypterus fin.

Full limb regeneration is a property that seems to be restricted to urodele amphibians. Here we found that Polypterus, the most basal living ray-finned fish, regenerates its pectoral lobed fins with a remarkable accuracy. Pectoral Polypterus fins are complex, formed by a well-organized endoskeleton to which the exoskeleton rays are connected. Regeneration initiates with the formation of a blastema similar to that observed in regenerating amphibian limbs. Retinoic acid induces dose-dependent phenotypes ranging from inhibition of regeneration to apparent anterior-posterior duplications. As in all developing tetrapod limbs and regenerating amphibian blastema, Sonic hedgehog is expressed in the posterior mesenchyme during fin regeneration. Hedgehog signaling plays a role in the regeneration and patterning processes: an increase or reduction of fin bony elements results when this signaling is activated or disrupted, respectively. The tail fin also regenerates but, in contrast with pectoral fins, regeneration can resume after release from the arrest caused by hedgehog inhibition. A comparative analysis of fin phenotypes obtained after retinoic acid treatment or altering the hedgehog signaling levels during regeneration allowed us to assign a limb tetrapod equivalent segment to Polypterus fin skeletal structures, thus providing clues to the origin of the autopod. We propose that appendage regeneration was a common property of vertebrates during the fin to limb transition.

Utility of a clinical risk scale to predict the requirement of advanced airway management in patients with a diagnosis of deep neck abscess.

OBJECTIVES: To analyze the clinical utility of a clinical risk scale to predict the need for advanced airway management in patients with deep neck abscess. METHODS: Observational, analytical, cross-sectional study. Patients over 18 years old, both genders, with surgical management of a deep neck abscess, between January 1st, 2015 to December 31th, 2021, who were applied the clinical risk scale (https://7-414-5-19.shinyapps.io/ClinicalRiskScore/). The sensitivity, specificity, and predictive values of the scale were calculated based on the identified clinical outcomes. A p<0.05 was considered significant. RESULTS: A sample of 213 patients was obtained, 121 (56.8%) men, of whom 50 (23.5%) required advanced airway management. Dyspnea was the variable with the most statistical weight in our study, (p=0.001) as well as the multiple spaces involvement, (p=0.001) the presence of air corpuscles, (p=0.001) compromise of the retropharyngeal space (p=0.001) and age greater than 55 years (p=0.001). Taking these data into account, were found for the clinical risk scale a sensitivity of 97% and a specificity of 65% (p=0.001, 95% CI 0.856-0.984). CONCLUSIONS: The clinical risk scale developed to predict advanced airway management in patients with a diagnosis of deep neck abscess may be applicable in our environment with high sensitivity and specificity. LEVEL OF EVIDENCE: IV.

[Risk factors at admission associated with intubation in patients with COVID-19]. / Factores de riesgo al ingreso asociados a intubación en pacientes con COVID-19.

Background: Intubation rates up to 33% have been found in patients diagnosed with COVID-19. Some cohorts have reported the presence of dyspnea in 84.1% of intubated patients, being this the only symptom associated with intubation. Oxygen saturation < 90% and increased respiratory rate have also been described as predictors of intubation. Objective: To analyze the risk factors associated with intubation in patients hospitalized for COVID-19 at their admission. Material and methods: An observational, retrospective, analytical, cross-sectional study was carried out. The universe of study consisted of patients over 18 years of age hospitalized due to a diagnosis of SARS-CoV-2 virus infection from April 1, 2020 to April 31, 2021 in the Hospital de Especialidades (Specialties Hospital) "Dr. Bernardo Sepúlveda Gutiérrez" at the National Medical Center. Results: The mean age of intubated patients was 59.17 years (95% confidence interval [95% CI] -9.994 to -3.299, p < 0.001). Overall, 76.7% (230) of patients had a history of one or more preexisting comorbidities, including hypertension in 42.3% (127), obesity in 36.7% (110), and diabetes mellitus in 34.3% (103). Conclusions: The main clinical characteristics of patients hospitalized for COVID-19 in our center who required intubation are very similar to those observed in different centers, including male sex, age over 50 years and obesity, which were the most common.

Introducción: se han encontrado tasas de intubación de hasta 33% en pacientes con diagnóstico de COVID-19. Algunas cohortes han informado la presencia de disnea en el 84.1% de los pacientes intubados y este ha sido el único síntoma asociado con la intubación. La saturación de oxígeno < 90% y el aumento de la frecuencia respiratoria también han sido descritos como predictores de intubación. Objetivo: analizar los factores de riesgo asociados a intubación en pacientes con COVID-19 al momento de su admisión hospitalaria. Material y métodos: se realizó un estudio observacional, transversal, analítico y retrospectivo. El universo de estudio consistió en pacientes mayores de 18 años, hospitalizados por diagnóstico de infección por virus SARS-CoV-2 del 1 abril de 2020 al 31 abril de 2021 en el Hospital de Especialidades "Dr. Bernardo Sepúlveda Gutiérrez" del Centro Médico Nacional. Resultados: se analizaron un total de 300 pacientes. La media de edad de los pacientes intubados fue de 59.17 años (p < 0.001, intervalo de confianza del 95% [IC 95%] −9.994 a −3.299). En general, el 76.7% (230) de los pacientes tenía antecedentes de una o más comorbilidades preexistentes, incluida la hipertensión en 42.3% (127), la obesidad en 36.7% (110) y la diabetes mellitus en 34.3% (103). Conclusiones: las principales características clínicas de los pacientes hospitalizados por COVID-19 en nuestro centro que requirieron de intubación son muy similares a las observadas en distintos centros, entre ellas el sexo masculino, la edad mayor de 50 años y la obesidad, que fueron las más prevalentes.

Telencephalic neural precursor cells show transient competence to interpret the dopaminergic niche of the embryonic midbrain.

Neural Precursor Cells (NPCs) generate complex stereotypic arrays of neuronal subtypes in the brain. This process involves the integration of patterning cues that progressively restrict the fate of specific NPCs. Yet the capacity of NPCs to interpret foreign microenvironments during development remains poorly defined. The aim of this work was to test the competence of mouse telencephalic NPCs to respond to the dopaminergic niche of the mesencephalon. Telencephalic NPCs isolated from midgestation mouse embryos (E10.5) and transplanted to age-matched mesencephalic explants efficiently differentiated into neurons but were largely unable to produce midbrain dopaminergic (mDA) neurons. Instead, E10.5 telencephalic NPCs behaved as restricted gabaergic progenitors that maintained ectopic expression of Foxg1 and Pax6. In contrast, E8.5 telencephalic NPCs were able to differentiate into Lmx1a(+)/Foxa2(+)/TH(+) neurons in the dopaminergic niche of the mesencephalic explants. In addition, these early telencephalic NPCs showed region-dependent expression of Nkx6.1, Nkx2.2 and site-specific differentiation into gabaergic neurons within the mesencephalic tissue. Significant dopaminergic differentiation of E8.5 telencephalic NPCs was not observed after transplantation to E12.5 mesencephalic explants, suggesting that inductive signals in the dopaminergic niche rapidly decay after midgestation. Moreover, we employed transplantation of embryonic stem cells-derived precursors to demonstrate that extinction of inductive signals within the telencephalon lags behind the commitment of residing NPCs. Our data indicate that the plasticity to interpret multiple instructive niches is an early and ephemeral feature of the telencephalic neural lineage.

Progressive interdigital cell death: regulation by the antagonistic interaction between fibroblast growth factor 8 and retinoic acid.

The complete cohort of molecules involved in interdigital cell death (ICD) and their interactions are yet to be defined. Bmp proteins, retinoic acid (RA) and Fgf8 have been previously identified as relevant factors in the control of ICD. Here we determined that downregulation of Fgf8 expression in the ectoderm overlying the interdigital areas is the event that triggers ICD, whereas RA is the persistent cell death-inducing molecule that acts on the distal mesenchyme by a mechanism involving the induction of Bax expression. Inhibition of the mitogen-activated protein kinase (Mapk) pathway prevents the survival effect of Fgf8 on interdigital cells and the accompanying Erk1/2 phosphorylation and induction of Mkp3 expression. Fgf8 regulates the levels of RA by both decreasing the expression of Raldh2 and increasing the expression of Cyp26b1, whereas RA reduces Fgfr1 expression and Erk1/2 phosphorylation. In the mouse limb, inhibition of Bmp signaling in the mesenchyme does not affect ICD. However, noggin in the distal ectoderm induces Fgf8 expression and reduces interdigit regression. In the chick limb, exogenous noggin reduces ICD, but, when applied to the distal mesenchyme, this reduction is associated with an increase in Fgf8 expression. In agreement with the critical decline in Fgf8 expression for the activation of ICD, distal interdigital cells acquire a proximal position as interdigit regression occurs. We identified proliferating distal mesenchymal cells as those that give rise to the interdigital cells fated to die. Thus, ICD is determined by the antagonistic regulation of cell death by Fgf8 and RA and occurs through a progressive, rather than massive, cell death mechanism.

Dll1 haploinsufficiency causes brain abnormalities with functional relevance.

Introduction: The Notch pathway is fundamental for the generation of neurons during development. We previously reported that adult mice heterozygous for the null allele of the gene encoding the Delta-like ligand 1 for Notch (Dll1lacZ ) have a reduced neuronal density in the substantia nigra pars compacta. The aim of the present work was to evaluate whether this alteration extends to other brain structures and the behavioral consequences of affected subjects. Methods: Brains of Dll1 +/lacZ embryos and mice at different ages were phenotypically compared against their wild type (WT) counterpart. Afterwards, brain histological analyses were performed followed by determinations of neural cell markers in tissue slices. Neurological deficits were diagnosed by applying different behavioral tests to Dll1 +/lacZ and WT mice. Results: Brain weight and size of Dll1 +/lacZ mice was significantly decreased compared with WT littermates (i.e., microcephaly), a phenotype detected early after birth. Interestingly, enlarged ventricles (i.e., hydrocephalus) was a common characteristic of brains of Dll1 haploinsufficient mice since early ages. At the cell level, general cell density and number of neurons in several brain regions, including the cortex and hippocampus, of Dll1 +/lacZ mice were reduced as compared with those regions of WT mice. Also, fewer neural stem cells were particularly found in the adult dentate gyrus of Dll1 +/lacZ mice but not in the subventricular zone. High myelination levels detected at early postnatal ages (P7-P24) were an additional penetrant phenotype in Dll1 +/lacZ mice, observation that was consistent with premature oligodendrocyte differentiation. After applying a set of behavioral tests, mild neurological alterations were detected that caused changes in motor behaviors and a deficit in object categorization. Discussion: Our observations suggest that Dll1 haploinsufficiency limits Notch signaling during brain development which, on one hand, leads to reduced brain cell density and causes microcephaly and hydrocephalus phenotypes and, on the other, alters the myelination process after birth. The severity of these defects could reach levels that affect normal brain function. Therefore, Dll1 haploinsufficiency is a risk factor that predisposes the brain to develop abnormalities with functional consequences.

Asociación del signo de Moore con la presencia de absceso profundo de cuello de espacio retrofaríngeo.

BACKGROUND: Deep neck abscesses can cause life-threatening complications. They are diagnosed by physical examination, and contrasted tomography as the gold standard. There are no studies about the association of Moore's sign with infections of the retropharyngeal space. OBJECTIVE: To determine the usefulness of Moore's sign in the diagnosis of deep retropharyngeal abscess. METHOD: Observational, analytical, cross-sectional, study of patients with deep neck abscess, from May 1, 2019, to August 30, 2021, with report of Moore's sign. RESULTS: 87 patients were included, 49 (56.3%) males (p = 0.45). Of those who developed complications, 77.8% had a negative Moore's sign (p = 0.001). Of those admitted to the ICU, 72% had a negative Moore's sign (p = 0.001). The sensitivity of the absence of the sign with retropharyngeal involvement was 95.4%, and the specificity was 86.3%. By logistic regression, it was found that those with retropharyngeal involvement are 467 times more likely to present a negative sign (p < 0.05). CONCLUSIONS: The presence of abscess in the retropharynx is associated with complications and a worse prognosis. The evaluation of Moore's sign can be a useful tool to suspect compromise of this space.

ANTECEDENTES: Los abscesos profundos de cuello pueden ocasionar complicaciones letales. Se diagnostican por exploración física, y la tomografía contrastada es el método de referencia. No existen estudios de asociación del signo de Moore con infecciones del espacio retrofaríngeo. OBJETIVO: Determinar la utilidad del signo de Moore en el diagnóstico de absceso profundo en el espacio retrofaríngeo. MÉTODO: Estudio observacional, transversal y analítico, de pacientes con absceso profundo de cuello, del 1 de mayo de 2019 al 30 de agosto de 2021, con reporte de signo de Moore. RESULTADOS: Se incluyeron 87 pacientes, de los cuales 49 (56.3%) eran de sexo masculino (p = 0.45). De los que desarrollaron complicaciones, el 77.8%, tenían el signo de Moore negativo (p = 0.001). De los que ingresaron a la unidad de cuidados intensivos, el 72% tenían negativo el signo de Moore (p = 0.001). La sensibilidad de la ausencia del signo con afección del espacio retrofaríngeo fue del 95.4%, y la especificidad del 86.3%. Por regresión logística se encontró que aquellos con afección del espacio retrofaríngeo tienen 467 veces más posibilidades de presentar signo negativo (p < 0.05). CONCLUSIONES: La presencia de un absceso en el espacio retrofaríngeo se asocia a complicaciones y peor pronóstico. La evaluación del signo de Moore puede ser una herramienta útil para sospechar compromiso de ese espacio.

Retroperitoneal Teratoma in a Transplanted Patient: Case Report and Review.

BACKGROUND: The development of de novo neoplasms in solid organ transplantation is multifactorial. In addition to common factors in the general population, there are specific factors of the disease related or not to chronic renal failure and factors inherent to the transplant itself such as immunosuppression. OBJECTIVE: The aim of this study is to describe the case of a kidney recipient with a retroperitoneal teratoma, his satisfactory treatment, and a brief literature review. METHODS: The case of 59-year-old male patient who received a living donor transplant in 2011, with conventional immunosuppression, graft protocol biopsy per year reported as normal, and follow-up without eventualities is described. The patient's symptoms began in December 2020 with abdominal pain resistant to analgesics, asthenia, and adynamic. Contrast tomography showed a retroperitoneal tumor 25.8 × 16.9 × 19 cm; tumor markers: alpha fetoprotein, 2.16 ng/mL; cancer antigen 19-9, 524.5 UI/ml; and carcinoembryonic antigen, 67.53 ng/mL. Resection of a 25 × 25 × 20 cm retroperitoneal tumor between the vena cava and aorta with 2 L of mucus content was performed. The patient was discharged from the hospital on the second day, with uresis 1 mL/kg/h, and at one month with adequate renal function, and 0.94 mg/dL of serum creatinine. A definitive histologic report was compatible with retroperitoneal mature teratoma. CONCLUSION: Primary retroperitoneal mature teratoma is rarely evidenced in adult patients, usually asymptomatic, and the definitive diagnosis always is established after histologic evaluation. Surgical resection is the main treatment with the complete removal of the tumor and long-term monitoring is needed because of the risk of malignancy.

Mortality Due to COVID-19 in Renal Transplant Recipients, Related to Variants of SARS-CoV-2 and Vaccination in Mexico.

BACKGROUND: SARS-CoV-2 infection in transplant patients has shown greater lethality and vaccination in this group of patients has shown less information. The objective of this study is to show the statistics in Mexico of lethality in kidney recipients infected with COVID-19 in relation to vaccination and variants of the coronavirus. METHODS: This is a bibliographic search of kidney transplant recipient patients since the start of the pandemic in Mexico to determine lethality after SARS-CoV-2 compared to the general population and in relation to patients, the 4 most important infectious peaks in the country due to identified variants, and also before and after vaccination. RESULTS: The global lethality is 26.91% from the beginning of the pandemic to April 9, 2022 in kidney recipients in Mexico (130 deaths of 483 infected kidney transplant recipients) compared to the national lethality of 5.60%. Variant B. 1.1.220 represented the highest lethality with 30.43% and the lowest lethality was Omicron with 16.41%. The lethality prior to vaccination was 30.94% and 23.46% after it. CONCLUSION: Both some variants and vaccination have influenced a lower lethality due to COVID-19 in Mexico in kidney transplant patients; It is important to consider global recommendations, such as a third or fourth dose, a combination of mRNA vaccines and vectors in order to reduce lethality in this group of patients.

Interdigital cell death function and regulation: new insights on an old programmed cell death model.

Interdigital cell death (ICD) is the oldest and best-studied model of programmed cell death (PCD) in vertebrates. The classical view of ICD function is the separation of digits by promotion of tissue regression. However, in addition, ICD can contribute to digit individualization by restricting interdigital tissue growth. Depending on the species, the relative contribution of either regression or growth-restricting functions of ICD to limb morphogenesis may differ. Under normal conditions, most cells appear to die by apoptosis during ICD. Accordingly, components of the apoptotic machinery are found in the interdigits, though their role in the initiation and execution of cell death is yet to be defined. Fgf8 has been identified as a survival factor for the distal mesenchymal cells of the limb such that ICD can initiate following specific downregulation of Fgf8 expression in the ectoderm overlying the interdigital tissue. On the other hand, Bmps may promote cell death directly by acting on the interdigital tissue, or indirectly by downregulating Fgf8 expression in the ectoderm. In addition, retinoic acid can activate ICD directly or through a Bmp-mediated mechanism. Interactions at different levels between these factors establish the spatiotemporal patterning of ICD activation. Defining the regulatory network behind ICD activation will greatly advance our understanding of the mechanisms controlling PCD in general.

The levels of reprogramming factors influence the induction and maintenance of pluripotency: the case of CD1 mouse strain cells.

The amount of proteins of the regulatory pluripotency network can be determinant for somatic cell reprogramming into induced pluripotent stem cells (iPSCs) as well as for the maintenance of pluripotent stem cells (PSCs). Here, we report a transposon-based reprogramming system (PB-Booster) that allowed high expression levels of a polycistronic transgene containing Myc, Klf4, Oct4 and Sox2 (MKOS) and showed increased reprogramming efficiency of fresh mouse embryonic fibroblasts (MEFs) into iPSCs under low, but not under high, MKOS expression levels. In contrast, MEFs after 2 passages derived into a similar number of iPSC colonies as fresh MEFs at a high MKOS dose, but this number was reduced at a low MKOS dose. Timing of reprogramming was not affected by MKOS expression levels but, importantly, exogenous MKOS expression in established PSCs caused a significant cell loss. At high but not at low MKOS expression levels, MEFs of the CD1 strain produced more initial cell clusters than iPSCs and, although reprogrammed at a similar efficiency as MEFs of the 129/Sv strain, iPSCs could not be maintained in the absence of exogenous MKOS. In CD1-iPSCs, Oct4, Nanog, Rex1 and Esrrb expression levels were reduced when compared with the levels in PSCs derived from the 129/Sv strain. Culture of CD1-iPSCs in medium with MEK and GSK3ß inhibitors allowed their self-renewal in the absence of exogenous MKOS, but the expression levels of Oct4, Nanog, Rex1 and Esrrb were only partially increased. Despite the reduced levels of those pluripotency factors, CD1-iPSC kept high capacity for contribution to chimeric mouse embryos. Therefore, levels of regulatory pluripotency factors influence reprogramming initiation and PSC maintenance in vitro without affecting their differentiation potential in vivo.

The embryonic midbrain directs neuronal specification of embryonic stem cells at early stages of differentiation.

Specific neuronal differentiation of Embryonic Stem Cells (ESCs) depends on their capacity to interpret environmental cues. At present, it is not clear at which stage of differentiation ESCs become competent to produce multiple neuronal lineages in response to the niche of the embryonic brain. To unfold the developmental potential of ESC-derived precursors, we transplanted these cells into the embryonic midbrain explants, where neurogenesis occurs as in normal midbrain development. Using this experimental design, we show that the transition from ESCs to Embryoid Body (EB) precursors is necessary to differentiate into Lmx1a(+)/Ptx3(+)/TH(+) dopaminergic neurons around the ventral midline of the midbrain. In addition, EB cells placed at other dorsal-ventral levels of the midbrain give rise to Nkx6.1(+) red nucleus (RN) neurons, Nkx2.2(+) ventral interneurons and Pax7(+) dorsal neurons at the correct positions. Notably, differentiation of ESCs into Neural Precursor Cells (NPCs) prior to transplantation markedly reduces specification at the Lmx1a, Nkx6.1 and Pax7 expression domains, without affecting neuronal differentiation. Finally, exposure to Fgf8 and Shh in vitro promotes commitment of some ESC-derived NPCs to differentiate into putative Lmx1a(+) dopaminergic neurons in the midbrain. Our data demonstrate intrinsic developmental potential differences among ESC-derived precursor populations.

Adult Neurogenesis in the Context of Brain Repair and Functional Relevance.

Urodeles and some fishes possess a remarkable capacity to regenerate their limbs/fins, a property that correlates with their additional ability to regenerate large areas of the brain and/or produce a variety of new neurons during adulthood. In contrast, neurogenesis in adult mammals is apparently restricted to two main regions, the subventricular zone of lateral ventricles and the subgranular zone of the hippocampus. There, astrocyte-like neural stem cells (NSCs) reside and derive into new neurons. Although it is becoming apparent that other brain regions carry out neurogenesis, in many cases, its functional significance is controversial, particularly, because very few putative NSCs capable of deriving into new neurons have been found. Hence, is renewal of certain neurons a requirement for a healthy brain? Are there specific physiological conditions that stimulate neurogenesis in a particular region? Does the complexity of the brain demand reduced neurogenesis? In this study, we review the production of new neurons in the vertebrate adult brain in the context of a possible functional relevance. In addition, we consider the intrinsic properties of potential cellular sources of new neurons, as well as the contribution of the milieu surrounding them to estimate the reparative capacity of the brain upon injury or a neurodegenerative condition. The conclusion of this review should bring into debate the potential and convenience of promoting neuronal regeneration in the adult human brain.

The nuclear receptor NR4A1 is regulated by SUMO modification to induce autophagic cell death.

NR4A is a nuclear receptor protein family whose members act as sensors of cellular environment and regulate multiple processes such as metabolism, proliferation, migration, apoptosis, and autophagy. Since the ligand binding domains of these receptors have no cavity for ligand interaction, their function is most likely regulated by protein abundance and post-translational modifications. In particular, NR4A1 is regulated by protein abundance, phosphorylation, and subcellular distribution (nuclear-cytoplasmic translocation), and acts both as a transcription factor and as a regulator of other interacting proteins. SUMOylation is a post-translational modification that can affect protein stability, transcriptional activity, alter protein-protein interactions and modify intracellular localization of target proteins. In the present study we evaluated the role of SUMOylation as a posttranslational modification that can regulate the activity of NR4A1 to induce autophagy-dependent cell death. We focused on a model potentially relevant for neuronal cell death and demonstrated that NR4A1 needs to be SUMOylated to induce autophagic cell death. We observed that a triple mutant in SUMOylation sites has reduced SUMOylation, increased transcriptional activity, altered intracellular distribution, and more importantly, its ability to induce autophagic cell death is impaired.