Direct Lineage Conversion of Adult Mouse Liver Cells and B Lymphocytes to Neural Stem Cells

Abstract

SummaryOverexpression of transcription factors has been used to directly reprogram somatic cells into a range of other differentiated cell types, including multipotent neural stem cells (NSCs), that can be used to generate neurons and glia. However, the ability to maintain the NSC state independent of the inducing factors and the identity of the somatic donor cells remain two important unresolved issues in transdifferentiation. Here we used transduction of doxycycline-inducible transcription factors to generate stable tripotent NSCs. The induced NSCs (iNSCs) maintained their characteristics in the absence of exogenous factor expression and were transcriptionally, epigenetically, and functionally similar to primary brain-derived NSCs. Importantly, we also generated tripotent iNSCs from multiple adult cell types, including mature liver and B cells. Our results show that self-maintaining proliferative neural cells can be induced from nonectodermal cells by expressing specific combinations of transcription factors.

Endocrine Pancreas Development and Regeneration: Noncanonical Ideas From Neural Stem Cell Biology

“…Hes3 expression in vivo has been implicated as an indicator of the efficacy of a g-secretase inhibitor (that blocks Notch signaling) in models of breast cancer (81). Recent studies have implicated Hes3 (still largely in a correlative manner) in different reprogramming paradigms (mouse embryonic fibroblasts to induced pluripotent cells and direct adult to induced NSCs) by demonstrating Hes3 expression regulation during reprogramming and by correlating Hes3 transduction of adult cells with successful reprogramming to induced NSCs (82,83).…”

“…However, Hes1 expression is much broader than the pattern of ectopic pancreas formation in the Hes1-null mice, suggesting that additional mechanisms could contribute to Ptf1a control or the control of other genes helping to limit ectopic pancreas generation. Hes3 and Hes5, genes with established roles in neural development and specification (5,74,82,83,95,96), are also expressed in the developing stomach and small intestine, respectively (24,97), so they could be candidates as cocontrollers of pancreas organogenesis. Another interesting member of the Hes/Hey gene family is Hes6, a proneural gene implicated in glioma progression (98) …”

Generation of diverse neural cell types through direct conversion

“…This may be in part due to the post-mitotic state of the target cell type (neuronlike cells), with the conversion procedure including a halt in proliferation, thus limiting the ability of these cells to expand once reprogrammed [98][99][100] . In addition to determining methods of increasing conversion efficiency, studies have expanded into investigating whether similar methods could be utilised for generation of proliferative neural stem and progenitor cells, which are both expandable in vitro and capable of generating multiple neural cell types [101] , with initial studies demonstrating the generation of induced neural progenitor [101] and crest [102] cells.…”

“…Furthermore, a number of studies also demonstrated the generation of induced neuroblasts [106] , and induced oligodendrocyte [107,108] and dopaminergic neuron [109] progenitor cells. While the majority of studies utilised fibroblasts as the starting cell type, there have also been reports of the direct conversion of astrocytes [106,110] , Sertoli cells [111] , epitheliallike cells in urine [112] , cord blood-derived stem cells [113] , bone marrow-derived stem cells [114] , liver cells, and B lymphocytes [99] into induced neural stem and progenitor cells, as summarised in Table 3. Generation of induced neural stem and progenitor cells has been achieved using a variety of approaches.…”

“…Similarly, episomal vector delivery of Oct4, Sox2, Klf4, Lin28, and L-Myc in combination with histone deacetylase inhibitor treatment has converted pig fibroblasts into induced neural progenitor cells [120] , and Oct4, Sox2, Klf4, and small hairpin RNA-p53 with a cocktail of small molecules has converted human fibroblasts into induced neural stem cells [119] , however no conversion efficiencies were reported for either study. Interestingly, a secondary system enabling non-viral neural direct conversion has been reported, in which fibroblasts, liver cells, and B lymphocytes were isolated from chimeric mice carrying inducible vectors expressing Brn2, Hes1, Hes3, Klf4, c-Myc, Plagl1, Notch1 (NICD), and Rfx4, with subsequent conversion into induced neural stem cells following transgene induction, however again with no reported conversion efficiencies [99] . Overall, while some nonviral methods achieve conversion efficien cies similar to studies utilising viral vectors [147] , others achieve considerably lower conversion efficiencies [98,112] or have not reported them [99,119,120] , thus necessitating optimisation of non-viral methods in order for them to become a viable alternative.…”