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Reconstitution of the Mouse Germ Cell Specification Pathway in Culture by Pluripotent Stem Cells

From Cell
By Stuart P. Atkinson

Protocols for the differentiation of pluripotent stem cells to functional therapeutically relevant cells often give relatively low yield. However, new protocols from various groups have addressed this by going back to learn more about the in vivo development of the cell type or tissue that they wish to attain, and utilising this knowledge in new more advanced differentiation protocols which aim to increase yield. In vitro attempts to generate gametes or primordial germ cells in mouse and human (Area reviews in Daley and Saitou et al) have largely been based on the isolation of these cells from spontaneously differentiating embryoid body cultures, an inefficient method that generally does not derive sufficient cells for substantial analysis. Now, researchers from the group of Mitinori Saitou at Kyoto University, Japan have demonstrated the efficient generation of PGC-like cells in mice which have spermatogenic capability. By utilising what information is known regarding temporal and signalling dynamics during PGC development in vivo and in vitro, they have devised a new protocol which leads to the development of PGC-like cells over multiple stages which reflects the development of the epiblast in vivo. The major novel step is the conversion of mouse embryonic stem cells (mESCs) to epiblast-like cells (EpiLCs), a state which is highly similar to cells of the pregastrulating epiblast, but distinct from epiblast stem cells (EpiSCs), which are known to be competent to express Blimp1/Prdm1 and Prdm14 (Kurimoto et al, Ohinita et al, Vincent et al and Yamaji et al), two of the major regulators of PGCs. Importantly, these regulators mediate the generation of functional sperm after ex vivo induction by BMP4 and neonatal intratesticular transplantation (Saitou et al). This study is published in the August edition of Cell (Hayashi et al).

Reconstitution of the Mouse Germ Cell Specification Pathway in Culture by Pluripotent Stem Cells

From Cell
By Stuart P. Atkinson

Protocols for the differentiation of pluripotent stem cells to functional therapeutically relevant cells often give relatively low yield. However, new protocols from various groups have addressed this by going back to learn more about the in vivo development of the cell type or tissue that they wish to attain, and utilising this knowledge in new more advanced differentiation protocols which aim to increase yield. In vitro attempts to generate gametes or primordial germ cells in mouse and human (Area reviews in Daley and Saitou et al) have largely been based on the isolation of these cells from spontaneously differentiating embryoid body cultures, an inefficient method that generally does not derive sufficient cells for substantial analysis. Now, researchers from the group of Mitinori Saitou at Kyoto University, Japan have demonstrated the efficient generation of PGC-like cells in mice which have spermatogenic capability. By utilising what information is known regarding temporal and signalling dynamics during PGC development in vivo and in vitro, they have devised a new protocol which leads to the development of PGC-like cells over multiple stages which reflects the development of the epiblast in vivo. The major novel step is the conversion of mouse embryonic stem cells (mESCs) to epiblast-like cells (EpiLCs), a state which is highly similar to cells of the pregastrulating epiblast, but distinct from epiblast stem cells (EpiSCs), which are known to be competent to express Blimp1/Prdm1 and Prdm14 (Kurimoto et al, Ohinita et al, Vincent et al and Yamaji et al), two of the major regulators of PGCs. Importantly, these regulators mediate the generation of functional sperm after ex vivo induction by BMP4 and neonatal intratesticular transplantation (Saitou et al). This study is published in the August edition of Cell (Hayashi et al).

Complete Meiosis from Human Induced Pluripotent Stem Cells

From the August Edition of Stem Cells
By Stuart P. Atkinson

Multiple studies have shown that mouse and human embryonic stem cells (ESCs) can differentiate in vitro into primordial germ cells (PGCs) and oocyte- or sperm-like cells (Marques-Mari et al). Therapeutic use of these cells would require patient specificity, so generation of such cells from individualised induced pluripotent stem cells (iPSC) would be required; a feat which has yet to be reported. However, in a study published in the August Edition of Stem Cells, Eguizabal et al from the laboratory of Juan Carlos Izpisúa Belmonte at the Center for Regenerative Medicine in Barcelona, Spain the complete differentiation of human iPSCs to post-meiotic cells has now been demonstrated, perhaps leading the way towards the ultimate goal of patient specific gamete formation.

Complete Meiosis from Human Induced Pluripotent Stem Cells

From the August Edition of Stem Cells
By Stuart P. Atkinson

Multiple studies have shown that mouse and human embryonic stem cells (ESCs) can differentiate in vitro into primordial germ cells (PGCs) and oocyte- or sperm-like cells (Marques-Mari et al). Therapeutic use of these cells would require patient specificity, so generation of such cells from individualised induced pluripotent stem cells (iPSC) would be required; a feat which has yet to be reported. However, in a study published in the August Edition of Stem Cells, Eguizabal et al from the laboratory of Juan Carlos Izpisúa Belmonte at the Center for Regenerative Medicine in Barcelona, Spain the complete differentiation of human iPSCs to post-meiotic cells has now been demonstrated, perhaps leading the way towards the ultimate goal of patient specific gamete formation.

Radical Acceleration of Nuclear Reprogramming By Chromatin Remodeling with the Transactivation Domain of MyoD

From the September Edition of Stem Cells
By Stuart P. Atkinson

Generation of induced pluripotent stem cells (iPSCs) is both slow and inefficient; the route from somatic target cell generally takes a minimum of 4 weeks and only 1 in a thousand target cells being reprogrammed. The proper reconfiguration of the chromatin landscape is deemed a potential obstacle in the reprogramming process; so much so that small molecule inhibitors which promote a more open chromatin configuration are becoming common place in many reprogramming protocols (Huangfu, Maehr et al and Huangfu, Osafune et al). One potential problem with this approach is the lack of specific chromatin changes; instead these inhibitors promote global chromatin changes. Specific chromatin changes occur due to the specific recruitment of epigenetic regulators to specific loci by transcription factors; and so this suggests that currently used transcription factors in reprogramming (Oct4, Sox2, Klf4, Myc and Nanog) may have limited means to reconfigure chromatin. Myod1 is a master transcription factor for skeletal myogenesis and can directly convert one cell type into another, as exemplified by its ability to generate myotubes from pigmented retinal epithelial cells (Choi et al). This suggests that Myod1 has a more potent ability to recruit epigenetic modifiers leading to activation of suppressed genes embedded in closed chromatin. This hypothesis has been now been tested by the laboratory of from the Stem Cell Institute, University of Minnesota, USA and Laboratory of Animal Reproduction, Kinki University, Nara, Japan and is presented in the September Edition of Stem Cells (Hirai et al). Full-length mouse Oct4 (O) was fused with various fragments of mouse Myod1, excluding the basic helix-loop-helix (bHLH) domain to avoid activation of Myod1 target genes, and were co-transduced with a polycistronic retroviral vector encoding mouse Sox2, Klf4, and Myc (SKM) into MEFs derived from Oct4-GFP mice, allowing for the monitoring of the reprogramming process. Using this system, the authors demonstrate that that a specific chimeric Oct4-Myod1 protein can reprogram MEFs to an iPSC state more efficiently than OSKM.

Radical Acceleration of Nuclear Reprogramming By Chromatin Remodeling with the Transactivation Domain of MyoD

From the September Edition of Stem Cells
By Stuart P. Atkinson

Generation of induced pluripotent stem cells (iPSCs) is both slow and inefficient; the route from somatic target cell generally takes a minimum of 4 weeks and only 1 in a thousand target cells being reprogrammed. The proper reconfiguration of the chromatin landscape is deemed a potential obstacle in the reprogramming process; so much so that small molecule inhibitors which promote a more open chromatin configuration are becoming common place in many reprogramming protocols (Huangfu, Maehr et al and Huangfu, Osafune et al). One potential problem with this approach is the lack of specific chromatin changes; instead these inhibitors promote global chromatin changes. Specific chromatin changes occur due to the specific recruitment of epigenetic regulators to specific loci by transcription factors; and so this suggests that currently used transcription factors in reprogramming (Oct4, Sox2, Klf4, Myc and Nanog) may have limited means to reconfigure chromatin. Myod1 is a master transcription factor for skeletal myogenesis and can directly convert one cell type into another, as exemplified by its ability to generate myotubes from pigmented retinal epithelial cells (Choi et al). This suggests that Myod1 has a more potent ability to recruit epigenetic modifiers leading to activation of suppressed genes embedded in closed chromatin. This hypothesis has been now been tested by the laboratory of from the Stem Cell Institute, University of Minnesota, USA and Laboratory of Animal Reproduction, Kinki University, Nara, Japan and is presented in the September Edition of Stem Cells (Hirai et al). Full-length mouse Oct4 (O) was fused with various fragments of mouse Myod1, excluding the basic helix-loop-helix (bHLH) domain to avoid activation of Myod1 target genes, and were co-transduced with a polycistronic retroviral vector encoding mouse Sox2, Klf4, and Myc (SKM) into MEFs derived from Oct4-GFP mice, allowing for the monitoring of the reprogramming process. Using this system, the authors demonstrate that that a specific chimeric Oct4-Myod1 protein can reprogram MEFs to an iPSC state more efficiently than OSKM.

Debrided Skin as a Source of Autologous Stem Cells for Wound Repair

From the August Edition of Stem Cells
Paper commentary by Stuart P. Atkinson

Tissue resident adult stem cells, such as mesenchymal stem cells (MSCs) or adipose-derived stem cells (ASCs), have previously demonstrated a capacity to repair extensively injured tissues (Picinich et al, Horwitz and Dominici). However, major traumatic injuries such as large surface area burns, which constitute 5%–10% of military casualties, limit the availability of autologous stem cell populations for wound repair and such injuries also require extensive reconstruction. The process of wound debridement; the medical removal of a patient's dead, damaged, or infected tissue to improve the healing potential of the remaining healthy tissue, typically involves the removal of subcutaneous layers and associated tissue structures, including portions of intact hypodermal adipose tissue. This led the group of Robert J. Christy at the United States Army Institute of Surgical Research, Fort Sam Houston, Texas, USA to investigate the potential of debrided skin to be a source of viable autologous stem cells for use in wound treatments. Their report (Natesan et al) is published in the August Edition of Stem Cells.

Debrided Skin as a Source of Autologous Stem Cells for Wound Repair

From the August Edition of Stem Cells
Paper commentary by Stuart P. Atkinson

Tissue resident adult stem cells, such as mesenchymal stem cells (MSCs) or adipose-derived stem cells (ASCs), have previously demonstrated a capacity to repair extensively injured tissues (Picinich et al, Horwitz and Dominici). However, major traumatic injuries such as large surface area burns, which constitute 5%–10% of military casualties, limit the availability of autologous stem cell populations for wound repair and such injuries also require extensive reconstruction. The process of wound debridement; the medical removal of a patient's dead, damaged, or infected tissue to improve the healing potential of the remaining healthy tissue, typically involves the removal of subcutaneous layers and associated tissue structures, including portions of intact hypodermal adipose tissue. This led the group of Robert J. Christy at the United States Army Institute of Surgical Research, Fort Sam Houston, Texas, USA to investigate the potential of debrided skin to be a source of viable autologous stem cells for use in wound treatments. Their report (Natesan et al) is published in the August Edition of Stem Cells.

New insight into how a somatic past shapes the future of human iPSCs

From Nature Cell Biology
Paper commentary by Carla Mellough

The differences between human embryonic stem cells (hESC) and their somatically derived counterparts, induced pluripotent stem cells (iPSC), have been under close scrutiny following the accumulation of various reports indicating greater disparity between the two cell types than originally envisaged (for example see iPSC don’t forget their origins). Conflicting results have been reported and remain unresolved, for example the transcriptional signature of iPSCs, although ascribed to partial memory retention of their somatic origin, does not always correlate with the differences in gene expression between iPSCs and hESCs. This has led to some of the biases being attributed to interlaboratory methodological variation. Functional disparity between differentiating hESCs and iPSCs has also been reported, with iPSC derivatives showing limited differentiation potential and early senescence and perhaps indicating that iPSCs do not hold a comparable clinical value to hESCs. This, alongside numerous reports highlighting the remarkable similarity of both cell types has resulted in some confusion regarding the applicability of iPSCs to translational research. For this reason, elucidation of the true likeness between iPSCs and hESCs has been somewhat limited. A recent study published in Nature Cell Biology from various centres at the University of California by Ohi et al.attempts to address these limitations by systematically comparing human iPSC lines derived from multiple somatic cells types and under the same methodology, in parallel.

New insight into how a somatic past shapes the future of human iPSCs

From Nature Cell Biology
Paper commentary by Carla Mellough

The differences between human embryonic stem cells (hESC) and their somatically derived counterparts, induced pluripotent stem cells (iPSC), have been under close scrutiny following the accumulation of various reports indicating greater disparity between the two cell types than originally envisaged (for example see iPSC don’t forget their origins). Conflicting results have been reported and remain unresolved, for example the transcriptional signature of iPSCs, although ascribed to partial memory retention of their somatic origin, does not always correlate with the differences in gene expression between iPSCs and hESCs. This has led to some of the biases being attributed to interlaboratory methodological variation. Functional disparity between differentiating hESCs and iPSCs has also been reported, with iPSC derivatives showing limited differentiation potential and early senescence and perhaps indicating that iPSCs do not hold a comparable clinical value to hESCs. This, alongside numerous reports highlighting the remarkable similarity of both cell types has resulted in some confusion regarding the applicability of iPSCs to translational research. For this reason, elucidation of the true likeness between iPSCs and hESCs has been somewhat limited. A recent study published in Nature Cell Biology from various centres at the University of California by Ohi et al.attempts to address these limitations by systematically comparing human iPSC lines derived from multiple somatic cells types and under the same methodology, in parallel.

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