Background: The golden retriever muscular dystrophy (GRMD) dogs represent the best available animal model for therapeutic trials aiming at the future treatment of human Duchenne muscular dystrophy (DMD). We have obtained a rare litter of six GRMD dogs (3 males and 3 females) born from an affected male and a carrier female which were submitted to a therapeutic trial with adult human stem cells to investigate their capacity to engraft into dogs muscles by local as compared to systemic injection without any immunosuppression.Methods: Human Immature Dental Pulp Stem Cells (hIDPSC) were transplanted into 4 littermate dogs aged 28 to 40 days by either arterial or muscular injections. Two non-injected dogs were kept as controls. Clinical translation effects were analyzed since immune reactions by blood exams and physical scores capacity of each dog. Samples from biopsies were checked by immunohistochemistry (dystrophin markers) and FISH for human probes.Results and Discussion: We analyzed the cells' ability in respect to migrate, engraftment, and myogenic potential, and the expression of human dystrophin in affected muscles. Additionally, the efficiency of single and consecutive early transplantation was compared. Chimeric muscle fibers were detected by immunofluorescence and fluorescentin situhybridisation (FISH) using human antibodies and X and Y DNA probes. No signs of immune rejection were observed and these results suggested that hIDPSC cell transplantation may be done without immunosuppression. We showed that hIDPSC presented significant engraftment in GRMD dog muscles, although human dystrophin expression was modest and limited to several muscle fibers. Better clinical condition was also observed in the dog, which received monthly arterial injections and is still clinically stable at 25 months of age.Conclusion: Our data suggested that systemic multiple deliveries seemed more effective than local injections. These findings open important avenues for further researches.

Pioneer work in male mouse embryonic stem (ES) cells differentiation into germ cells (GC) showed generations of male or female gametes in separate experiments, using genetically manipulated or preselected ES cells. In an attempt to produce both types of gametes from male mouse ES cells without any genetic manipulation or preselection, we induce the differentiation by retinoic acid (RA) within nonadherent embryoid bodies (EB). It seems that gamete-like cell formation occurs in the correct manner based on the expression of early and late GC-specific genes such as Oct-4, Mvh, Stella, Dazl, Piwil 2, Pdrd 1, Rex 14, Rnf 17, Bmp8b, Acrosin, Stra-8, Haprin, LH-R, Gdf9, Zp3, Zp2, Sycp1, and Sycp3. Immunofluorescence analysis of morphologically well-formed GC and presumptive gametes showed positive labeling for SSEA1, Oct-4, EMA-1, FE-J1, Dazl, Fragilis, Mvh, Acrosin, and acetylated α-tubulin. Conventional cytogenetic and FISH analysis indicated a chromosome reduction in ES-derived GC. Our data suggest that ES cells with XY chromosomes can produce under the same experimental conditions both types of presumptive gametes, and this production depends on their positional and temporal information within the EB context.

Human pluripotent stem cells (hPSCs) may significantly improve drug development pipeline, serving as anin vitrosystem for the identification of novel leads, and for testing drug toxicity. Furthermore, these cells may be used to address the issue of differential drug response, a phenomenon greatly influenced by genetic factors. This application depends on the availability of hPSC lines from populations with diverse ancestries. So far, it has been reported that most lines of hPSCs derived worldwide are of European or East Asian ancestries. We have established 23 lines of hPSCs from Brazilian individuals, and we report the analysis of their genomic ancestry. We show that embryo-derived PSCs are mostly of European descent, while induced PSCs derived from participants of a national-wide Brazilian cohort study present high levels of admixed European, African and Native American genomic ancestry. Additionally, we use high density SNP data and estimate local ancestries, particularly those of CYP genes loci. Such information will be of key importance when interpreting variation among cell lines with respect to cellular phenotypes of interest. The availability of genetically admixed lines of hPSCs will be of relevance when setting up futurein vitrostudies of drug response.

Normal mouse pluripotent stem cells were originally derived from the inner cell mass (ICM) of blastocysts and shown to be the in vitro equivalent of those pre-implantation embryonic cells, and thus were called embryonic stem cells (ESCs). More than a decade later, pluripotent cells were isolated from the ICM of human blastocysts. Despite being called human ESCs, these cells differ significantly from mouse ESCs, including different morphology and mechanisms of control of pluripotency, suggesting distinct embryonic origins of ESCs from the two species. Subsequently, mouse pluripotent stem cells were established from the ICM-derived epiblast of post-implantation embryos. These mouse epiblast stem cells (EpiSCs) are morphological and epigenetically more similar to human ESCs. This raised the question of whether cells from the human ICM are in a more advanced differentiation stage than their murine counterpart, or whether the available culture conditions were not adequate to maintain those human cells in their in vivo state, leading to a transition into EpiSC-like cells in vitro. More recently, novel culture conditions allowed the conversion of human ESCs into mouse ESC-like cells called naïve (or ground state) human ESCs, and the derivation of naïve human ESCs from blastocysts. Here we will review the characteristics of each type of pluripotent stem cells, how (and whether) these relate to different stages of embryonic development, and discuss the potential implications of naïve human ESCs in research and therapy.