A placental model of SARS-CoV-2 infection reveals ACE2-dependent susceptibility and differentiation impairment in syncytiotrophoblasts.

SARS-CoV-2 infection causes COVID-19. Several clinical reports have linked COVID-19 during pregnancy to negative birth outcomes and placentitis. However, the pathophysiological mechanisms underpinning SARS-CoV-2 infection during placentation and early pregnancy are not clear. Here, to shed light on this, we used induced trophoblast stem cells to generate an in vitro early placenta infection model. We identified that syncytiotrophoblasts could be infected through angiotensin-converting enzyme 2 (ACE2). Using a co-culture model of vertical transmission, we confirmed the ability of the virus to infect syncytiotrophoblasts through a previous endometrial cell infection. We further demonstrated transcriptional changes in infected syncytiotrophoblasts that led to impairment of cellular processes, reduced secretion of HCG hormone and morphological changes vital for syncytiotrophoblast function. Furthermore, different antibody strategies and antiviral drugs restore these impairments. In summary, we have established a scalable and tractable platform to study early placental cell types and highlighted its use in studying strategies to protect the placenta.

Comparative roadmaps of reprogramming and oncogenic transformation identify Bcl11b and Atoh8 as broad regulators of cellular plasticity.

Coordinated changes of cellular plasticity and identity are critical for pluripotent reprogramming and oncogenic transformation. However, the sequences of events that orchestrate these intermingled modifications have never been comparatively dissected. Here, we deconvolute the cellular trajectories of reprogramming (via Oct4/Sox2/Klf4/c-Myc) and transformation (via Ras/c-Myc) at the single-cell resolution and reveal how the two processes intersect before they bifurcate. This approach led us to identify the transcription factor Bcl11b as a broad-range regulator of cell fate changes, as well as a pertinent marker to capture early cellular intermediates that emerge simultaneously during reprogramming and transformation. Multiomics characterization of these intermediates unveiled a c-Myc/Atoh8/Sfrp1 regulatory axis that constrains reprogramming, transformation and transdifferentiation. Mechanistically, we found that Atoh8 restrains cellular plasticity, independent of cellular identity, by binding a specific enhancer network. This study provides insights into the partitioned control of cellular plasticity and identity for both regenerative and cancer biology.

The application of human pluripotent stem cells to model the neuronal and glial components of neurodevelopmental disorders.

Cellular models of neurodevelopmental disorders provide a valuable experimental system to uncover disease mechanisms and novel therapeutic strategies. The ability of induced pluripotent stem cells (iPSCs) to generate diverse brain cell types offers great potential to model several neurodevelopmental disorders. Further patient-derived iPSCs have the unique genetic and molecular signature of the affected individuals, which allows researchers to address limitations of transgenic behavioural models, as well as generate hypothesis-driven models to study disorder-relevant phenotypes at a cellular level. In this article, we review the extant literature that has used iPSC-based modelling to understand the neuronal and glial contributions to neurodevelopmental disorders including autism spectrum disorder (ASD), Rett syndrome, bipolar disorder (BP), and schizophrenia. For instance, several molecular candidates have been shown to influence cellular phenotypes in three-dimensional iPSC-based models of ASD patients. Delays in differentiation of astrocytes and morphological changes of neurons are associated with Rett syndrome. In the case of bipolar disorders and schizophrenia, patient-derived models helped to identify cellular phenotypes associated with neuronal deficits (e.g., excitability) and mutation-specific abnormalities in oligodendrocytes (e.g., CSPG4). Further we provide a critical review of the current limitations of this field and provide methodological suggestions to enhance future modelling efforts of neurodevelopmental disorders. Future developments in experimental design and methodology of disease modelling represent an exciting new avenue relevant to neurodevelopmental disorders.

An improved reprogrammable mouse model harbouring the reverse tetracycline-controlled transcriptional transactivator 3.

Reprogrammable mouse models engineered to conditionally express Oct-4, Klf-4, Sox-2 and c-Myc (OKSM) have been instrumental in dissecting molecular events underpinning the generation of induced pluripotent stem cells. However, until now these models have been reported in the context of the m2 reverse tetracycline-controlled transactivator, which results in low reprogramming efficiency and consequently limits the number of reprogramming intermediates that can be isolated for downstream profiling. Here, we describe an improved OKSM mouse model in the context of the reverse tetracycline-controlled transactivator 3 with enhanced reprogramming efficiency (>9-fold) and increased numbers of reprogramming intermediate cells albeit with similar kinetics, which we believe will facilitate mechanistic studies of the reprogramming process.

APP intracellular domain acts as a transcriptional regulator of miR-663 suppressing neuronal differentiation.

Amyloid precursor protein (APP) is best known for its involvement in the pathogenesis of Alzheimer's disease. We have previously demonstrated that APP intracellular domain (AICD) regulates neurogenesis; however, the mechanisms underlying AICD-mediated regulation of neuronal differentiation are not yet fully characterized. Using genome-wide chromatin immunoprecipitation approaches, we found that AICD is specifically recruited to the regulatory regions of several microRNA genes, and acts as a transcriptional regulator for miR-663, miR-3648 and miR-3687 in human neural stem cells. Functional assays show that AICD negatively modulates neuronal differentiation through miR-663, a primate-specific microRNA. Microarray data further demonstrate that miR-663 suppresses the expression of multiple genes implicated in neurogenesis, including FBXL18 and CDK6. Our results indicate that AICD has a novel role in suppression of neuronal differentiation via transcriptional regulation of miR-663 in human neural stem cells.

[Influence of donor age on graft survival]. / Edad del donante y su influencia en la supervivencia del injerto.

INTRODUCTION: In 2007 in Spain 43% of donors were older than 60 years. This produces a worse graft quality and probably a worse survival. OBJECTIVE: Our objective is to analyze the influence of donor age on graft survival. MATERIAL AND METHODS: We analyze retrospectively 216 renal consecutive transplants realized between 2000 and 2008. A univaried and multivaried study (Cox regression) was performed and Kaplan-Meyer test with log rank for graft survival. RESULTS: Follow-up mean of 40 months (+/-33,4 SD). The univaried analysis of graft survival showed that donor age had a significative influence on graft survival. (OR=1,03; 95% CI 1,01-1,05) (p: 0,009). Studying the relation between donor and recipient age we find an inverse correlation (Pearson's Correlation: 0,55. p<0,0001), but there are significative differences after the adjustment for recipient age. (OR: 1,02; 95% CI 1,01-1,04) (p: 0,04). Optimal cut-point value determined by the ROC analysis was 60 years. The graft survival of donors over 60 years is 79% (95% CI; 74-84%) and 71% (95% CI; 65-77%) at 3 and 5 years in contrast with 94% (95% CI; 94-96%) and 90% (95% CI; 88-92 in donors under 60. (p: 0,002). The multivaried study of the influential factors on graft survival reveals that donor age dichotomized in older or younger than 60, the presence of a surgical immediate reintervention and a delayed graft function were independent influence factors. CONCLUSIONS: Donor age over 60 years has a negative and independent prognostic influence on graft survival.

[Surgical complications in kidney transplantation and their influence on graft survival]. / Complicaciones quirúrgicas en el trasplante renal y su influencia en la supervivencia del injerto.

OBJECTIVES: To analyze surgical complications in kidney transplantation and their influence on graft survival. MATERIALS AND METHODS: A retrospective analysis was made of the early and late surgical complications occurring in 216 consecutive kidney transplants performed at our institution and their influence on graft survival. RESULTS: At least one surgical complication occurred in 82 (38%) of the 216 transplantations, and 68 (31%) required some type of repeat surgery, 23 in the early postoperative period and 45 more than 3 months after surgery. Mean follow-up was 48 months (SD +/-33.4), and median follow-up 48 months (range, 0-166 months). No recipient or donor factors predisposing to surgical complications were found. Graft survival was significantly shorter in patients with surgical complications [3- and 5-year survival rates of 86% (95% CI 83-89) and 78% (95% CI 73-82) as compared to 92% (95% CI 90-94) and 88% (95% CI 85-91), p=0.004]. Early repeat surgery, venous thrombosis, and wound infection were among the complications having an independent influence on graft survival. A multivariate analysis of graft survival in the whole group showed early repeat surgery to be a factor with an independent prognostic value (OR: 4.7; 95% CI 2.2-10, p<0.0001). Delayed function and donor age older than 60 years were the other independent influential factors. CONCLUSION Surgical complications have an influence on graft survival. The need for early repeat surgery, delayed function, and donor age older than 60 years are independent predictors of graft survival.

[Pelvic lymph node dissection (extended vs standard) and prostate cancer]. / Linfadenectomía (ampliada vs estándar) y cáncer de próstata.

The role and the potential benefit, if any, of pelvic lymphadenectomy in prostate cancer are still controversially discussed. It is generally accepted that PLND at time of radical prostatectomy is the only reliable diagnostic procedure to achieve as much individual histological staging information as possible to trigger postoperative adjuvant management. However, the extent of pelvic lymph node dissection (limited vs. extended) and the most suitable candidates for this procedure are still a matter of intense debate. The aim of this review is to critically evaluate the current status on lymph node dissection in prostate cancer.

[Gleason score 8-10 prostatic adenocarcinoma: prognostic influence in the biochemical progression free survival]. / Adenocarcinoma de próstata gleason clínico 8-10: influencia pronóstica en la supervivencia libre de progresión bioquímica.

OBJECTIVE: Hereby, we analyze the characteristics of the clinical Gleason 8-10 group of patients with in our series diagnosed of Prostate Cancer and treated by means of radical prostatectomy, and we try to ascertain which are the influence factors within this group upon progression and progression free survival. MATERIAL AND METHODS: From the global series of 781 patients with T1-T2 prostate cancer treated by means of radical prostatectomy between 1990 and 2004, we study 108 with a Gleason score on the biopsy of 8-10. Median PSA was 12 ng/ml and 50% were T2. Variables related to biochemical progression and progression free survival have been studied, comparing the group of Gleason 8-10 with the rest and analyzing, within the Gleason 8-10 group which are the related variables with progression and progression free survival, trying to find a predictive model. Contingency tables and logistic regression have been employed. For the survival analysis, Kaplan Meyer curves, log-rank and Cox models. RESULTS: Actual State: 62.7% (490/781) are alive and free of biochemical progression, 24.8% (194/781) are alive with biochemical progression, 2.9% (23/781) are dead by cancer and 1.9% (15/781) are dead by other cause and 7.6% (59/781) are lost. Biochemical progression study of the whole series (781 patients) Clinical Gleason score 8-10 is a influence factor on the univariate study (OR2,61 IC 95%: 1.7-4). In the progression free survival study (PFS) of the whole series (781 patients) the PFS in Clinical Gleason 8-10 at 3 and 5 years is 56 +/- 5% y 35 +/- 7%, significantly worse than the rest of the group (p < 0.0001). In the multivariate study of the influence factors on the PFS includes Clinical Gleason Score 8-10 as an independent prognostic factor (OR: 2.6 IC 95%: 1.6-4.12) p = 0.003, together with the clinical stage (OR: 1.,81 IC 95%: 1.18-2.78) p < 0.006, the PSA (OR: 1.03 IC 95%: 1.025-1.046) p < 0.0001 and the side of tumor on the biopsy (OR: 1.5 IC 95%: 1.01-2.24) p = 0.045. In the clinical Gleason score 8-10 group the influent factors on the PFS are. PSA (OR: 1.02 IC 95%: 1.003-1.04) and pathological stage (OR: 3.84 IC 95%: 1.77-8.27). Patients with a pT2 have a significantly better survival than those pT3 at 3 and 5 years (80 +/- 6%; 54 +/- 13% y 40 +/- 7%; 27 +/- 7%) (p < 0.0001). The best cut point for the PSA is 11 ng/ml. Patients with a PSA < 11 ng/ml have a 3 and 5 years survival better than those with >11 ng/ml PSA (74 +/- 7%, 30 +/- 22% y 40 +/- 7%, 26 +/- 7%) (p < 0.0001). CONCLUSIONS: Clinical Gleason Score 8-10 is a negative independent prognostic factor on the progression free survival, but its prognosis is better if they present a PSA prior surgery lower than 11 ng/ml and the pathological stage is a pT2.

[Extracorporeal circulation and hypothermy surgery in tumors with vena cava extension: 20 years experience at the University Clinic of Navarra]. / Cirugía con circulación extracorpórea e hipotermia en tumores con extensión a vena cava: 20 años de experiencia de la clínica Universitaria de Navarra.

OBJECTIVE: We present our 20 years experience treating patients with vena cava extension in whom an extracorporeal circulation, hypothermia, cardio circulatory arrest (ECC-H-CCA) in order to perform, together with a tumoral resection, a thrombus resection. MATERIAL AND METHODS: From 1985 to 2005 a total of 28 retroperitoneal tumor were treated: 25 renal cancers, a Wilms tumor, a paratesticular rabdomiosarcoma, and a pheocromocitoma. All of them had an extension by means of thrombus above the suprahepatics veins. All of them were treated by means of ECC-H-CCA for thrombus extraction. A descriptive study of the serie is performed as well as a Kaplan Meyer survival study. RESULTS: Surgical complications were present within 10 patients (35%), with a surgical mortality of two patients (7%): one intra-operatively because a massive embolism of the lungs and the other because of a lung embolism on the 4th post-operative day. Global actuarial survival was 29.1+/-10% at three years and 17.5+/-8% at five years. Analyzing only who do not have metastatic lesions, nor lymph nodes at diagnosis their three year survival was 50.9+/-16.3% and 32.2+/-16% at five years. Mean while those who have any metastatic lesion at diagnosis their three and five years survival was 20.8+/-12% and 10.4+/-9% respectively. CONCLUSIONS: The employ of surgical techniques with ECC-H-CCA with in oncological pathology associated with vena cava thrombus is justified and its employment does not worsen the survival; it is indicated because its results, allowing a complete tumoral resection in a safe and reproducible fashion.

Long-term protection in hamsters against human parainfluenza virus type 3 following mucosal or combinations of mucosal and systemic immunizations with chimeric alphavirus-based replicon particles.

No licensed vaccines are available to protect against parainfluenza virus type 3 (PIV3), a significant health risk for infants. In search of a safe vaccine, we used an alphavirus-based chimeric vector, consisting of Sindbis virus (SIN) structural proteins and Venezuelan equine encephalitis virus (VEE) replicon RNA, expressing the PIV3 hemagglutinin-neuraminidase (HN) glycoprotein (VEE/SIN-HN). We compared different routes of intramuscular (i.m.), intranasal (i.n.), or combined i.n. and i.m. immunizations with VEE/SIN-HN in hamsters. Six months after the final immunization, all hamsters were protected against live PIV3 i.n. challenge in nasal turbinates and lungs. This protection appeared to correlate with antibodies in serum, nasal turbinates and lungs. This is the first report demonstrating mucosal protection against PIV3 for an extended time following immunizations with an RNA replicon delivery system.

[Steroid and xenobiotic receptor (SXR), multidrug resistance gene (MDR1) and GSTs, SULTs and CYP polymorphism expression in invasive bladder cancer, analysis of their expression and correlation with other prognostic factors]. / Expresion del receptor de esteroides y xenobioticos (SXR) y del gen de multirresistencia drogas (MDR1) y de los polimorfismos de las enzimas Gsts, SULTs y CYP en tumores vesicales profundos, analisis de su expresion y correlación con otros factores pronósticos.

INTRODUCTION: Steroid and Xenobiotic Receptor (SXR) has demonstrated its activation by numerous drugs, including cytochrome P450 potent inducers like rifampicina or cotrimazol. The role of SXR is well known, and lies regulating in a positive manner cytochrome P450 3A4 (CYP3A4) transcription and the multidrug resistance gene (MDR1), it's considered a key in the xenobiotic detoxification mechanism, being involved in all phases of the detoxification process. Enzymes involved in Policyclic Aromatic hidrocarbures (PAH) metabolism and degradation are polymorphic in humans, including glutation S-transferases (GSTs), N-acetiltransferases (NATs), sulfotransferases (SULTs)1A1 and cytochrome p450 (CYP)1B1. OBJECTIVES: The objectives we've planned are: 1. Analyze the expression of the transcription factor SXR and MDR1 in bladder by means of RT-PCR real time, both in normal bladder and in tumoral bladder. 2. Analyze the relation between clinical and pathological factors with the expression of SXR and MDR1. 3. Analyze the expression of the polymorphims CYP1B1, GSTM1 GSTT1 and SULT1A1 and their correlation with different clinic-pathological and molecular factors. MATERIAL AND METHODS: In a prospective way the size of the sample was estimated. In 67 patients from two institutions (Hospital Universitario Miguel Servet (49 HUMS) and Clinica Universitaria de Navarra (18 CUN)), diagnosed of invasive bladder cancer and treated by means of radical cystectomy, were determined the expression of both SXR and MDR1 by means of real time PCR, as well as the polymorphisms CYP1B1, GSTM1 GSTT1 y SULT1A1 by means of RFLP (Restriction fragment length polymorphism). Correlations with other prognostic factors by contingency tables were performed. RESULTS: Average follow up was 23.7 months with a median of 28.26 months. Of the 67 patients studied, 31 patients (46.3) presented disease progression, in form of local recurrence or in distant metastasis or both. With a average time to progression of 12.4 months and a median of 10 months, with a range of 1.1 month to 31.9 month. 36 patients (53.7%) did not have any evidence of disease progression during follow up. The Steroid and Xenobiotic Receptor as well as the Multidrug Resistance Gene (MDR1) are expressed in both normal bladder (0.94DeltaCt y 0.94DeltaCt) and tumoral bladder in the cystectomy specimen (1.09 DeltaCt y 0.45 DeltaCt). We've analyzed their expression in a quantitative manner and in a qualitative manner. The expression of SXR correlates with the presence of ca. in situ (p=0.024), vasculo-lymphatic invasion (p=0.05) mean while MDR1 correlates with presence of vasculo-lymphatic invasion (p=0.05) Both factors are correlate between each others (p=0.011). Polymorphisms: CYP1B1, GSTM1, GSTT1 and SULT1A1, are expressed in these patients but their expression doesn't correlates with any prognostic factor CONCLUSIONS: Both SXR and MDR1 are expressed in normal bladder as well as in tumoral bladder. And their expression correlates with different prognostic factors with influence in the survival described in the literature.

Sindbis virus replicon particles encoding calreticulin linked to a tumor antigen generate long-term tumor-specific immunity.

Alphavirus vectors have emerged as a promising strategy for the development of cancer vaccines and gene therapy applications. In this study, we used the replication-defective vaccine vector SIN replicon particles from a new packaging cell line (PCL) to develop SIN replicon particles encoding calreticulin (CRT) linked to a model tumor antigen, human papillomavirus type 16 (HPV16) E7 protein. The linkage of CRT to E7 in SIN replicon particles resulted in a significant increase in E7-specific CD8(+) T-cell precursors and a strong antitumor effect against E7-expressing tumors in vaccinated mice. SINrep5-CRT/E7 replicon particles enhanced presentation of E7 through the major histocompatibility complex (MHC) class I pathway by infecting dendritic cells (DCs) directly and pulsing DCs with lysates of cells infected by SINrep5-CRT/E7 replicons. Vaccination of immunocompromised (BALB/c nu/nu) mice with SINrep5-CRT/E7 replicon particles also generated significant reduction of lung tumor nodules, suggesting that antiangiogenesis may contribute to the antitumor effect of SINrep5-CRT/E7 replicon particles. Furthermore, SINrep5-CRT/E7 replicon particles generated long-term in vivo tumor protection effects and antigen-specific memory immunities. We concluded that the CRT strategy used in the context of SIN replicon particles facilitated the generation of a highly effective vaccine for cancer prophylaxis and immunotherapy.

[Pheochromocytoma with thrombus in cava]. / Feocromocitoma con trombo en cava.

Pheochromocytoma, a paraganglioma of suprarenal location, is a catecholamine-secreting chromaffin cell tumour. Spread of these tumours to the vena cava is rare and the thrombus only reaches the right atrium in exceptional cases. We present the case of a patient who, without previous symptomatology, presented with a clinical picture of multiorganic dysfunction with primary manifestation of a suprarenal tumour with vascular spread to the right atrium affecting the right suprahepatic vein.

[Incidentally detected renal cancer. Prognostic factors]. / Tumores renales incidentales. Factores pronósticos.

OBJECTIVE: To study the clinical and pathological characteristics of incidental renal tumors treated in our center. MATERIAL AND METHODS: A retrospective review is conducted of 318 nephrectomies comparing the clinico-pathological variables of renal tumors diagnosed incidentally with those of symptomatic renal tumors. The factors influencing disease-free survival are analyzed in both groups. RESULTS: In our experience, although incidental renal tumors presented better survival than symptomatic ones owing to their better pathological state and tumor grade, incidental diagnosis was not an independent influencing factor in the multivariate study. Only when patients were studied who did not present metastases on diagnosis did incidental diagnosis become an influencing factor very close to statistical significance. CONCLUSIONS: Incidental diagnosis is not an independent prognostic factor.