Human somatic cells subjected to genetic induction with six germ line-related factors display meiotic germ cell-like features.

The in vitro derivation of human germ cells has attracted interest in the last years, but their direct conversion from human somatic cells has not yet been reported. Here we tested the ability of human male somatic cells to directly convert into a meiotic germ cell-like phenotype by inducing them with a combination of selected key germ cell developmental factors. We started with a pool of 12 candidates that were reduced to 6, demonstrating that ectopic expression of the germ line-related genes PRDM1, PRDM14, LIN28A, DAZL, VASA and SYCP3 induced direct conversion of somatic cells (hFSK (46, XY), and hMSC (46, XY)) into a germ cell-like phenotype in vitro. Induced germ cell-like cells showed a marked switch in their transcriptomic profile and expressed several post-meiotic germ line related markers, showed meiotic progression, evidence of epigenetic reprogramming, and approximately 1% were able to complete meiosis as demonstrated by their haploid status and the expression of several post-meiotic markers. Furthermore, xenotransplantation assays demonstrated that a subset of induced cells properly colonize the spermatogonial niche. Knowledge obtained from this work can be used to create in vitro models to study gamete-related diseases in humans.

Understanding Mammalian Germ Line Development with In Vitro Models.

Germ line development is crucial in organisms with sexual reproduction to complete their life cycle. In mammals, knowledge about germ line development is based mainly on the mouse model, in which genetic and epigenetic events are well described. However, little is known about how germ line development is orchestrated in humans, especially in the earliest stages. New findings derived from human in vitro models to obtain germ cells can shed light on these questions. This comprehensive review summarizes the current knowledge about mammalian germ line development, emphasizing the state of the art obtained from in vitro models for germ cell-like cell derivation. Current knowledge of the pluripotency cycle and germ cell specification has allowed different in vitro strategies to obtain germ cells with proven functionality in mouse models. Several reports during the last 10 years show that in vitro germ cell derivation with proven functionality to generate a healthy offspring is possible in mice. However, differences in the embryo development and pluripotency potential between human and mouse make it difficult to extrapolate these results. Further efforts on both human and mouse in vitro models to obtain germ cells from pluripotent stem cells may help to elucidate how human physiological events take place; therefore, therapeutic strategies can also be considered.

Germ line development: lessons learned from pluripotent stem cells.

Current knowledge about mammalian germ line development is mainly based on the mouse model and little is known about how this fundamental process occurs in humans. This review summarizes our current knowledge of genetic and epigenetic germ line development in mammals, mainly focusing on primordial germ cell (PGC) specification events, comparing the differences between mouse and human models. We also emphasize the knowledge derived from the most successful strategies used to generate germ cell-like cells in vitro in both models and major obstacles to obtaining bona fide in vitro-derived gametes are considered.

Germ cell transplantation into mouse testes procedure.

OBJECTIVE: To illustrate the step-by-step protocol followed to assay germ cell transplantation into the seminiferous epithelium of mouse testes. DESIGN: Video presentation of an animal model for research in reproductive and regenerative medicine. SETTING: Research laboratory. ANIMAL(S): Male nude mice (NU-Foxn1(nu)). INTERVENTION(S): Mice were chemically sterilized with alkylant compounds (busulfan) followed by gonadal microsurgery to inject donor germ cells. MAIN OUTCOME MEASURE(S): Donor cells should be labeled with reporter genes, such as green fluorescent protein (GFP), lactose operon (LacZ), or alternatively design an effective strategy with specific antibodies to track them within the recipient testes. Sperm detection in the ejaculate can also be used as a read out. However, in this case detection of the donor genotype in the sperm is mandatory to elucidate their origin. RESULT(S): In the present study we describe the complete protocol for germ cell transplant by efferent duct injection, including the preparation of recipient mice, surgery for the germ cell transplant, and analysis of recipient testes. The main strength of this technique is that it constitutes the gold standard for a functional test of the germ cell potential as only spermatogonial stem cells are able to properly colonize the seminal lumen. Both fresh and frozen/thawed testicular cells are suitable for this technique as donor germ cells. Also, enrichment of living spermatogonial stem cells, previous to the transplant, seems to improve the efficiency of colonization. For proper colonization of germ cells, the niche should be available and thus mouse strains that lack endogenous spermatogenesis such as W/W(v) mutant mice are usually used. In the case of nonmatched donor cells, seminiferous epithelium of immune-suppressed recipient mice should be germ cell depleted before the transplant. One limitation of this technique is that the procedure can take up to 3 months. Also, in contrast to the full recovery of spermatogenesis in mouse-to-mouse transplants, xenotransplantation of germ cells from phylogenetically distant species, such as humans into mouse recipients, results in colonization of donor cells and spermatogonial expansion, but fail in their spermatogenic progression due to evolutive incompatibilities with the recipient niche. Xenografting of pieces of donor testis tissue under the skin of mouse hosts is an alternative approach that is currently being investigated to try to solve this limitation. CONCLUSION(S): Transplantation of spermatogonial stem cells into the seminal lumen of mouse testes is a functional assay that defines this cellular subpopulation by its ability to colonize it. This technique can be used as a model to elucidate the insights of spermatogonial stem cells, to produce transgenic animals by genetically manipulating donor cells before transplantation, but also it has potential applications in fertility preservation in cattle and humans as it is feasible in large animals, as recent reports have demonstrated with rhesus monkeys, that recovered spermatogenesis after allogenic transplantation, and even from human cadaver testes. Therefore spermatogonial stem cells isolated from prepuberal boys, who are treated with alkylant chemotherapy, could be returned to their testis to regenerate spermatogenesis in the future.

Factores asociados al ingreso hospitalario y evaluación de un programa de gestión de casos para trastorno mental grave mediante un estudio de cohortes históricas / Factors associated with hospital admission and evaluation of a case management program for severe mental disorder by a historical cohort study

Introducción: Las personas con trastorno mental grave (TMG) presentan serias dificultades para desarrollar una vida normalizada, por lo que son necesarios programas de atención comunitaria que mejoren sus condiciones de vida e integración social. Este trabajo pretende evaluar el funcionamiento de un programa de gestión de casos (PGC) en Segovia (España). Metodología: Se realiza una primera fase descriptiva valorando el funcionamiento del PGC en 2011 mediante variables sociodemográficas, asistenciales y clínicas. Se estudian los factores asociados a la ocurrencia de ingreso hospitalario. Finalmente, mediante un diseño de cohortes históricas, se evalúa el riesgo de ingreso del PGC comparando con una cohorte no expuesta. Se emplean técnicas estadísticas bi y multivariantes con cálculo de riesgos relativos e intervalos de confianza. Resultados: En 2011 se atiende a 82 pacientes en el PGC, principalmente hombres de mediana edad. La evolución clínica media es de 19 años y la permanencia media en el PGC superior a los 6 años. El 78% pertenecen al espectro diagnóstico de la esquizofrenia. El ingreso afecta al 27% de los pacientes. Ser mujer, ser atendido por equipos de salud mental I-II, el aumento de visitas domiciliarias y el abandono del seguimiento son los factores predictores de ingreso, mientras la mayor evolución clínica es factor protector. No se detecta efecto protector del PGC frente al ingreso hospitalario en los diferentes análisis del estudio de cohortes históricas. Conclusiones: Es necesario evaluar de forma sistemática los programas de atención comunitaria dirigidos al TMG con el fin de realizar ajustes y modificaciones tendentes a la mejora de su efectividad clínica (AU)

Introduction: People with severe mental disorder (SMD) have serious difficulties in developing a normal life, so community care programs to improve their living conditions and social integration are necessary. This work evaluates the performance of a case management program (CMP) in Segovia (Spain).Methodology: We conduct a first descriptive phase evaluating the performance of the CMP in 2011 by sociodemographic, health services and clinical variables. We study the factors associated with the occurrence of hospital admission. Finally, using a historical cohort design, we assess the risk of hospital admission of CMP compared to unexposed cohort. Bi and multivariate statistical techniques are employed to perform the analysis with the calculation of relative risks and confidence intervals. Results: In 2011, 82 patients are cared for in the CMP, mainly middle-aged men. The average clinical course is 19 years and the average stay in the CMP over 6 years. 78%belong to the diagnosis of schizophrenia spectrum. Income affects 27% of patients. Women, mental health teams I-II, increased home visits and abandonment of monitoring are predictors of income, while the highest level of clinical course is protective. No protective effect of income is detected for the CMP in the different analyzes of the historical cohort study. Conclusions: It is necessary to systematically assess community care programs directed at SMD to make adjustments and modifications aiming at improving their clinical effectiveness (AU)

[Factors associated with hospital admission and evaluation of a case management program for severe mental disorder by a historical cohort study]. / Factores asociados al ingreso hospitalario y evaluación de un programa de gestión de casos para trastorno mental grave mediante un estudio de cohortes históricas.

INTRODUCTION: People with severe mental disorder (SMD) have serious difficulties in developing a normal life, so community care programs to improve their living conditions and social integration are necessary. This work evaluates the performance of a case management program (CMP) in Segovia (Spain). METHODOLOGY: We conduct a first descriptive phase evaluating the performance of the CMP in 2011 by sociodemographic, health services and clinical variables. We study the factors associated with the occurrence of hospital admission. Finally, using a historical cohort design, we assess the risk of hospital admission of CMP compared to unexposed cohort. Bi and multivariate statistical techniques are employed to perform the analysis with the calculation of relative risks and confidence intervals. RESULTS: In 2011, 82 patients are cared for in the CMP, mainly middle-aged men. The average clinical course is 19 years and the average stay in the CMP over 6 years. 78% belong to the diagnosis of schizophrenia spectrum. Income affects 27% of patients. Women, mental health teams I-II, increased home visits and abandonment of monitoring are predictors of income, while the highest level of clinical course is protective. No protective effect of income is detected for the CMP in the different analyzes of the historical cohort study. CONCLUSIONS: It is necessary to systematically assess community care programs directed at SMD to make adjustments and modifications aiming at improving their clinical effectiveness.

Overcoming challenges of ovarian cancer stem cells: novel therapeutic approaches.

Understanding the genetic and molecular mechanisms of ovarian cancer has been the focus of research efforts working toward the greater goal of improving cancer therapy for patients with residual disease after initial treatment with conventional surgery and neoadjuvant chemotherapy. The focus of this review will be centered on new therapeutic strategies based on Cancer Stem Cells studies of chemoresistant subpopulations, the prevention of metastasis, and individualized therapy in order to find the most successful combination of treatments to effectively treat human ovarian cancer. We reviewed recent literature (1993-2011) of novel treatment approaches to ovarian cancer stem cells. As the focus of ovarian cancer investigation has centered on the cancer stem cell model and the complexities that it presents in the development of effective treatments, the future of treating ovarian cancer lies in utilizing individualized treatment systems that include enhancing existing treatments, aiming for novel therapy targets, managing the plasticity of stem cells to induce cellular differentiation, and regulating oncogenic signaling pathways.

Specific unsaturated fatty acids enforce the transdifferentiation of human cancer cells toward adipocyte-like cells.

Differentiation therapy pursues the discovery of novel molecules to transform cancer progression into less aggressive phenotypes by mechanisms involving enforced cell transdifferentiation. In this study, we examined the identification of transdifferentiating adipogenic programs in human cancer cell lines (HCCLs). Our findings showed that specific unsatturated fatty acids, such as palmitoleic, oleic and linoleic acids, trigger remarkable phenotypic modifications in a large number of human cancer cell lines (HCCLs), including hepatocarcinoma HUH-7, ovarian carcinoma SK-OV-3, breast adenocarcinoma MCF-7 and melanoma MALME-3M. In particular, we characterized a massive biogenesis of lipid droplets (LDs) and up-regulation of the adipogenic master regulator, PPARG, resulting in the transdifferentiation of HCCLs into adipocyte-like cells. These findings suggest the possibility of a novel strategy in cancer differentiation therapy via switching the identity of HCCLs to an adipogenic phenotype through unsaturated fatty acid-induced transdifferentiation.