Superior Induced Pluripotent Stem Cell Generation through Phactr3-Driven Mechanomodulation of Both Early and Late Phases of Cell Reprogramming.

Human cell reprogramming traditionally involves time-intensive, multistage, costly tissue culture polystyrene-based cell culture practices that ultimately produce low numbers of reprogrammed cells of variable quality. Previous studies have shown that very soft 2- and 3-dimensional hydrogel substrates/matrices (of stiffnesses ≤ 1 kPa) can drive ~2× improvements in human cell reprogramming outcomes. Unfortunately, these similarly complex multistage protocols lack intrinsic scalability, and, furthermore, the associated underlying molecular mechanisms remain to be fully elucidated, limiting the potential to further maximize reprogramming outcomes. In screening the largest range of polyacrylamide (pAAm) hydrogels of varying stiffness to date (1 kPa to 1.3 MPa), we have found that a medium stiffness gel (~100 kPa) increased the overall number of reprogrammed cells by up to 10-fold (10×), accelerated reprogramming kinetics, improved both early and late phases of reprogramming, and produced induced pluripotent stem cells (iPSCs) having more naïve characteristics and lower remnant transgene expression, compared to the gold standard tissue culture polystyrene practice. Functionalization of these pAAm hydrogels with poly-l-dopamine enabled, for the first-time, continuous, single-step reprogramming of fibroblasts to iPSCs on hydrogel substrates (noting that even the tissue culture polystyrene practice is a 2-stage process). Comparative RNA sequencing analyses coupled with experimental validation revealed that a novel reprogramming regulator, protein phosphatase and actin regulator 3, up-regulated under the gel condition at a very early time point, was responsible for the observed enhanced reprogramming outcomes. This study provides a novel culture protocol and substrate for continuous hydrogel-based cell reprogramming and previously unattained clarity of the underlying mechanisms via which substrate stiffness modulates reprogramming kinetics and iPSC quality outcomes.

Muscle Stem Cells Undergo Extensive Clonal Drift during Tissue Growth via Meox1-Mediated Induction of G2 Cell-Cycle Arrest.

Organ growth requires a careful balance between stem cell self-renewal and lineage commitment to ensure proper tissue expansion. The cellular and molecular mechanisms that mediate this balance are unresolved in most organs, including skeletal muscle. Here we identify a long-lived stem cell pool that mediates growth of the zebrafish myotome. This population exhibits extensive clonal drift, shifting from random deployment of stem cells during development to reliance on a small number of dominant clones to fuel the vast majority of muscle growth. This clonal drift requires Meox1, a homeobox protein that directly inhibits the cell-cycle checkpoint gene ccnb1. Meox1 initiates G2 cell-cycle arrest within muscle stem cells, and disrupting this G2 arrest causes premature lineage commitment and the resulting defects in muscle growth. These findings reveal that distinct regulatory mechanisms orchestrate stem cell dynamics during organ growth, beyond the G0/G1 cell-cycle inhibition traditionally associated with maintaining tissue-resident stem cells.

Haematopoietic stem cell induction by somite-derived endothelial cells controlled by meox1.

Haematopoietic stem cells (HSCs) are self-renewing stem cells capable of replenishing all blood lineages. In all vertebrate embryos that have been studied, definitive HSCs are generated initially within the dorsal aorta (DA) of the embryonic vasculature by a series of poorly understood inductive events. Previous studies have identified that signalling relayed from adjacent somites coordinates HSC induction, but the nature of this signal has remained elusive. Here we reveal that somite specification of HSCs occurs via the deployment of a specific endothelial precursor population, which arises within a sub-compartment of the zebrafish somite that we have defined as the endotome. Endothelial cells of the endotome are specified within the nascent somite by the activity of the homeobox gene meox1. Specified endotomal cells consequently migrate and colonize the DA, where they induce HSC formation through the deployment of chemokine signalling activated in these cells during endotome formation. Loss of meox1 activity expands the endotome at the expense of a second somitic cell type, the muscle precursors of the dermomyotomal equivalent in zebrafish, the external cell layer. The resulting increase in endotome-derived cells that migrate to colonize the DA generates a dramatic increase in chemokine-dependent HSC induction. This study reveals the molecular basis for a novel somite lineage restriction mechanism and defines a new paradigm in induction of definitive HSCs.

[Renal cell carcinoma molecular biology. Prognostic and therapeutic usefulness]. / Biología molecular en adenocarcinoma renal (RCC). Su utilidad pronóstica y terapéutica.

Renal cell adenocarcinoma requires different therapeutic pathways because it is one of the most therapy-resistant tumors, on the other hand it is biologically one of the most attractive tumors. Its pathological classification has a genetic base. There is an anomaly of the Von Hippel Lindau gene in 80% of adenocarcinomas, being this fact determinant to know the biological characteristics of tumor initiation and development, as well as the identification of factors susceptible to be used as therapeutic targets. Since 2005 a group of molecules have been used in the treatment of metastatic adenocarcinomas and, even though therapeutic results are significant but not clinically relevant yet, we are sure they are a key way for more efficient future developments. The present study tries to make a tour on the research of the biological anomalies in renal adenocarcinoma with special emphasis in the Von HippelLindau gene.

Biologia molecular en adenocarcinoma renal (RCC). Su utilidad pronóstica y terapéutica / Renal cell carcinoma molecular biology. Prognostic and therapeutic usefulness

El adenocarcinoma renal requiere caminos terapéuticos diferentes porque es uno de los tumores más resistentes a tratamiento, por contra es uno de los tumores biológicamente más atractivos. Su clasificación anatomopatológica tiene un fundamento genético. En el 80% de los adenocarcinomas existe una alteración del gen Von Hippel Lindau y este hecho ha sido determinante para conocer las características biológicas de la aparición y desarrollo tumoral así como la identificación de factores que pueden ser susceptibles de ser utilizados como dianas terapéuticas. Desde 2005 un grupo de moléculas se ha utilizado en el tratamiento de los adenocarcinomas metastásicos y aunque los resultados terapéuticos son significativos pero no todavía clínicamente relevantes, estamos seguros que son un camino clave para desarrollos posteriores más eficientes. El presente estudio pretende hacer un recorrido por la investigación de las alteraciones biológicas en adenocarcinoma renal haciendo especial énfasis en las alteraciones del gen Von Hippel Lindau(AU)

Renal cell adenocarcinoma requires different therapeutic pathways because it is one of the most therapy-resistant tumors, on the other hand it is biologically one of the most attractive tumors. Its pathological classification has a genetic base. There is an anomaly of the Von Hippel Lindau gene in 80% of adenocarcinomas, being this fact determinant to know the biological characteristics of tumor initiation and development, as well as the identification of factors susceptible to be used as therapeutic targets. Since 2005 a group of molecules have been used in the treatment of metastatic adenocarcinomas and, even though therapeutic results are significant but not clinically relevant yet, we are sure they are a key way for more efficient future developments. The present study tries to make a tour on the research of the biological anomaliesin renal adenocarcinoma with special emphasis in the Von HippelLindau gene(AU)

Impact of renal retransplantation on graft and recipient survival.

OBJECTIVES: The aim of this study was to evaluate the influence of retransplantation in graft and recipient survival. METHODS: We carried out a retrospective study in 419 renal transplants and studied the influence of retransplantation in graft and patient survival. A homogeneity study was performed between the two groups with a Student`s T and a chi-square tests. Graft survival analysis was performed with Kaplan-Meyer and log rank tests. RESULTS: Of 419 transplants, 370 (88.3%) were first transplantations, 45 (10.7%) second transplantations and 4(1%) third ones. Mean follow-up of the whole group was 72.5 months (±54.1 SD). There were no differences in follow-up between groups (Mean Follow-up 73.1 months ±54.4 SD in first transplantations vs. 61.6 months ±51.2 SD in repeat transplantation. p >0.05). The actuarial graft survival showed no differences between patients with first transplantation and those with a repeat one. [3 and 5 year SV of 89% (95% CI: 87-91%) and 84%(95% CI: 82-86%) Vs 88% (95% CI; 83-93%) and 85% (95% CI:i; 80-90%) respectively]. After adjusting for all the heterogeneity variables we still did not find differences on graft survival. The actuarial recipient survival showed no differences between patients with first transplantation and those with a repeat one. [3 and 5 year SV of 98% and 96% Vs.97%]. CONCLUSIONS: There are no differences of graft and recipient survival between patients with a first transplantation and those with a repeat one.

Impacto del retrasplante renal en la supervivencia del injerto y receptor / Impact of renal retransplantation on graft and recipient survival

OBJETIVO: Nuestro objetivo es valorar si un segundo o tercer trasplante tienen influencia en la supervivencia del injerto renal y en la del receptor.MÉTODOS: Analizamos retrospectivamente 419 trasplantes renales consecutivos realizados entre 1994 y 2010, analizando la influencia del retrasplante en la supervivencia del injerto renal. Se ha realizado un estudio de homogeneidad de los dos grupos mediante Tablas de contingencia para las variables cualitativas y t de student para las cuantitativas. La supervivencia y comparación de supervivencia con Kaplan-Meyer y log-rank..RESULTADOS: De los 419 trasplantes, 370 (88,3%) fueron primeros trasplantes 45(10,7%) segundos trasplantes y 4(1%) terceros. Media de seguimiento de todo el grupo de 72,5 meses (+/- 54,1 DE) y mediana de 68,8 meses( Rango de 0 a 188 meses ).No existen diferencias en el tiempo de seguimiento (Media del grupo de pacientes con un solo trasplante de 73,1 meses +/-54,4DE Vs. 61,6 meses +/-51,2DE del grupo de pacientes retrasplantados. p >0,05).El análisis de la supervivencia actuarial del injerto revela que no existen diferencias estadísticamente significativas entre los pacientes con un primer trasplante y los retrasplantados [SPV 89% (95% IC; 87- 91%) y 84% (95% IC; 82-86%) a los 3 y 5 años frente a 88% (95% IC; 83-93%) a los 3 años y 85% (95% IC; 80-90%) a los 5 años]. Al ajustar por las variables para las que los grupos no fueron homogeneos las diferencias se siguen manteniendo.El análisis de supervivencia de los receptores revela que tampoco existen diferencias entre los dos grupos [SPV del 98% y 96% a los 3 y 5 años en los primeros trasplantes frente a 97% a los 3 años y 5 años en los retrasplantados].CONCLUSIONES: No existen diferencias en la supervivencia del injerto ni en la de los receptores entre pacientes trasplantados por primera vez y aquellos que reciben un retrasplante(AU)

OBJECTIVES: The aim of this study was to evaluate the influence of retransplantation in graft and recipient survival.METHODS: We carried out a retrospective study in 419 renal transplants and studied the influence of retransplantation in graft and patient survival.A homogeneity study was performed between the two groups with a Student`s T and a chi-square tests. Graft survival analysis was performed with Kaplan-Meyer and log rank tests.RESULTS: Of 419 transplants, 370 (88.3%) were first transplantations, 45(10.7%) second transplantations and 4(1%) third ones. Mean follow-up of the whole group was 72.5 months (+/-54.1 SD).There were no differences in follow-up between groups (Mean Follow-up 73.1 months +/-54.4 SD in first transplantations vs. 61.6 months +/-51.2 SD in repeat transplantation. p >0.05). The actuarial graft survival showed no differences between patients with first transplantation and those with a repeat one. [3 and 5-year SV of 89% (95% CI: 87-91%) and 84% (95% CI: 82-86%) Vs 88% (95% CI; 83-93%) and 85% (95% CI; 80-90%) respectively].After adjusting for all the heterogeneity variables we still did not find differences on graft survival.The actuarial recipient survival showed no differences between patients with first transplantation and those with a repeat one. [3 and 5 year SV of 98% and 96% Vs. 97%].CONCLUSIONS: There are no differences of graft and recipient survival between patients with a first transplantation and those with a repeat one(AU)

Association of steroid and xenobiotic receptor (SXR) and multidrug resistance 1 (MDR1) gene expression with survival among patients with invasive bladder carcinoma.

UNLABELLED: What's known on the subject? and What does the study add? SXR and MDR1 are known as responsible for chemo and radiotherapy resistance in some cancers, like kidney cancer (MDR1). Invasive bladder cancer is an aggressive disease, with different behaviour upon its tumoral stage, and also within the same tumoral stage, therefore molecular markers are sought. This study shows a new molecular marker, which has shown as a predictor for bad prognosis cancers, therefore, allowing us for a better patient selection for aggressive therapies. OBJECTIVE: To investigate the prognostic value of steroid and xenobiotic receptor (SXR) and multidrug resistance 1 (MDR1) gene expression in relation to survival among patients with invasive bladder cancer. PATIENTS AND METHODS: The prospective study included 67 patients diagnosed with invasive bladder cancer and treated with radical cystectomy at one of two institutions. SXR and MDR1 gene expression was assessed by real-time quantitative polymerase chain reaction (RT-PCR) in tumoral and normal tissue from frozen surgical specimens. RESULTS: Patients were followed for a mean of 29 months; 31 patients (46%) had progression. In univariate analysis, significant predictors of overall survival (OS) were pathological stage, lymph node (LN) status, histological grade, vascular-lymphatic invasion, and SXR expression. In multivariate analysis, independent predictors of OS were LN status (odds ratio [OR], 2.96; P=0.034), vascular-lymphatic invasion (OR, 2.50; P=0.029), and SXR expression (OR, 1.05, P=0.03). Among the 51 patients with negative LNs (pN0), univariate predictors of OS were SXR expression, MDR1 expression, and pathological stage. In multivariate analysis, SXR expression (OR, 1.06; P=0.01) and MDR1 expression (OR, 3.27; P=0.03) were independently associated with survival. Within the pN0 group, patients with SXR expression had shorter progression-free survival than did those without expression (P=0.004). This association persisted in the N0 subgroup with stage pT3-pT4 disease (P=0.028). However, in the pN1 group SXR expression did not have any influence. CONCLUSIONS: For patients with invasive bladder cancer, SXR expression has value as a predictor of survival independent of the standard pathological predictors. Its maximum importance appears to be in patients with stage pT3-pT4 pN0 disease.

A murine ESC-like state facilitates transgenesis and homologous recombination in human pluripotent stem cells.

Murine pluripotent stem cells can exist in two functionally distinct states, LIF-dependent embryonic stem cells (ESCs) and bFGF-dependent epiblast stem cells (EpiSCs). However, human pluripotent cells so far seemed to assume only an epiblast-like state. Here we demonstrate that human iPSC reprogramming in the presence of LIF yields human stem cells that display morphological, molecular, and functional properties of murine ESCs. We termed these hLR5 iPSCs because they require the expression of five ectopic reprogramming factors, Oct4, Sox2, Klf4, cMyc, and Nanog, to maintain this more naive state. The cells are "metastable" and upon ectopic factor withdrawal they revert to standard human iPSCs. Finally, we demonstrate that the hLR5 state facilitates gene targeting, and as such provides a powerful tool for the generation of recombinant human pluripotent stem cell lines.

[Usefulness of PET scans in diagnosing recurrent prostate cancer. Prostate with PSA level < 5 ng/ml]. / Valor de la PET en la recurrencia del cáncer de próstata con PSA < 5 ng/ml.

INTRODUCTION AND OBJECTIVES: We intend to evaluate the usefulness of PET scans in diagnosing recurrent prostate cancer after a curative attempt using radical treatment. MATERIAL AND METHODS: 92 consecutive prostate cancer patients in biochemical progression following radical surgery (63) or radiation treatment (29) were studied with positron emission tomography (PET). In all cases two scans were performed in the same day (11C-choline and 18F-FDG). PET efficacy was evaluated both globally (by employing the results achieved with both 11C-choline and 18F-FDG) and using both radiotracers independently to detect recurrence in patients with biochemical progression. For this purpose, we used comparison of means for k-independent samples, 2 x 2 and 2 x X contingency tables and ROC curves. RESULTS: 1. Global PET: there is evidence of PET alteration regarding the PSA level (P=.003): the clinical stage (P=.01). There are no statistically significant PET alterations regarding the affected biopsy (uni or bilateral), surgical margins, pathological stage and time to progression. ROC curve PET-PSA is statistically significant (P< .0001) permitting calculation of different cut-off points, with a specificity of 91% (highest) for a PSA of 4.3 ng/ml. 2. PET 18FDG: the area under the ROC curve is statistically significant (P< .0001) with a specificity of 91% for a PSA of 6.51 ng/ml. 3. PET 11choline: the area under the ROC curve is statistically significant (P< .0001) with a specificity of 91% for a PSA of 5.15 ng/ml. CONCLUSIONS: PET is a useful tool for diagnosing prostate cancer recurrence after a curative attempt using radical treatment.

Valor de la PET en la recurrencia del cáncer de próstata con PSA < 5 ng/ml / Usefulness of PET scans in diagnosing recurrent prostate cancer. Prostate with PSA level < 5 ng/ml

Introducción y objetivo: Evaluamos la utilidad de la tomografía por emisión de positrones (PET) en el diagnóstico de la recurrencia del cáncer de próstata tras tratamiento con intención curativa. Material y métodos: Se sometió a 92 pacientes consecutivos en progresión bioquímica tras cirugía radical (63) o radioterapia (29) a una PET. En todos los casos, se realizaron dos escáneres PET en el mismo día (11C-colina y 18F-FDG). Se evalúa la eficacia de la PET de manera global (utilizando los resultados con 11C-colina y 18F-FDG) y de manera independiente para detectar recurrencia en pacientes con progresión bioquímica. Para ello, se utilizan la comparación de medias para k muestras independientes, tablas de contingencia 2 × 2 y 2× X y curvas ROC. Resultados: 1. PET global: hay evidencia de la alteración de la PET en función del antígeno prostático específico (PSA) (p = 0,003), estadio clínico (p = 0,01). No existe una alteración de la PET estadísticamente significativa en función de la afectación de la biopsia (unilateral o bilateral), los márgenes quirúrgicos, el estadio patológico y el tiempo a progresión. La curva ROC PET-PSA es significativa (p < 0,0001) y permite calcular distintos puntos de corte; PSA = 4,3 ng/ml el que presenta una mayor especificidad (91%). 2. PET 18FDG: el área bajo la curva ROC es significativa (p < 0,0001), con una especificidad del 91% para un PSA =6,51 ng/ml. 3. PET 11colina: el área bajo la curva ROC es significativa (p < 0,0001), con una especificidad del 91% para un PSA = 5,15 ng/ml. Conclusiones: La PET es una herramienta útil en el diagnóstico de la recurrencia de cáncer de próstata tras tratamiento radical con intención curativa (AU)

Introduction and objectives: We intend to evaluate the usefulness of PET scans in diagnosing recurrent prostate cancer after a curative attempt using radical treatment. Material and methods: 92 consecutive prostate cancer patients in biochemical progression following radical surgery (63) or radiation treatment (29) were studied with positron emission tomography (PET). In all cases two scans were performed in the same day (11C-cholineand 18F-FDG). PET efficacy was evaluated both globally (by employing the results achieved with both 11C-choline and 18F-FDG) and using both radiotracers independently to detect recurrence in patients with biochemical progression. For this purpose, we used comparison of means for k-independent samples, 2 × 2 and 2 × X contingency tables and ROC curves. Results: 1. Global PET: there is evidence of PET alteration regarding the PSA level (P=0.003): the clinical stage (P=0.01). There are no statistically significant PET alterations regarding the affected biopsy (uni or bilateral), surgical margins, pathological stage and time to progression. ROC curve PET-PSA is statistically significant (P<0.0001) permitting calculation of different cut-off points, with a specificity of 91% (highest) for a PSA of 4.3 ng/ml. 2. PET18FDG: the area under the ROC curve is statistically significant (P<0.0001) with a specificity of91% for a PSA of 6.51 ng/ml. 3. PET 11choline: the area under the ROC curve is statistically significant (P<0.0001) with a specificity of 91% for a PSA of 5.15 ng/ml. Conclusions: PET is a useful tool for diagnosing prostate cancer recurrence after a curative attempt using radical treatment (AU)

Cumplimiento del tratamiento hormonal sustitutivo en mujeres menopáusicas / Compliance of hormone replacement therapy in menopausal women

FUNDAMENTO: El cumplimiento de un tratamiento es necesario para conseguir su efectividad. Se pretende conocer el cumplimiento del tratamiento hormonal sustitutivo (THS) en mujeres menopáusicas. MÉTODO: Estudio descriptivo observacional realizado en tres Áreas de Salud de la Comunidad Valenciana. Período de seguimiento desde 1989 hasta 1999. Se incluyeron mujeres que acudieron a una unidad de menopausia y comenzaron con el THS. La información se obtuvo de la historia clínica y de una encuesta telefónica realizada a las pacientes. Se analizaron la edad de la mujer, la edad de inicio y el tipo de la menopausia, la edad de comienzo del THS, el nivel educativo, el motivo de prescripción, el tiempo de utilización, el grado de información, los efectos secundarios y las causas de abandono. Se utilizó el método de Kaplan-Meier para conocer el cumplimiento y el riesgo proporcional de Cox para conocer las variables que influyen en el cumplimiento. RESULTADOS: Se incluyeron 363 mujeres. Existe un 75 por ciento de probabilidades de que las mujeres alcancen un tiempo de cumplimiento de 5 años, la mediana se corresponde con un cumplimiento de 11 años (intervalo de confianza [IC] del 95 por ciento, 9-13). Las mujeres que han padecido efectos secundarios (odds ratio ajustada [ORa], 2,60; IC del 95 por ciento, 1,84-3,68) presentan mayor tasa de abandono, mientras que éste es menor en las que han tenido beneficios (ORa, 1,77; IC del 95 por ciento, 1,22-2,53), aquellas cuya menopausia ha sido quirúrgica (ORa, 1,60; IC del 95 por ciento, 1,12-2,28) y las que tienen menos de 55 años al comienzo del THS (ORa, 2,61; IC del 95 por ciento, 1,67-4,07). CONCLUSIONES: El cumplimiento del THS en mujeres menopáusicas es alto y está condicionado por el hecho de tener beneficios y/o efectos secundarios, por la edad al inicio del tratamiento y el tipo de menopausia (AU)