A stably self-renewing adult blood-derived induced neural stem cell exhibiting patternability and epigenetic rejuvenation

Abstract

Recent reports suggest that induced neurons (iNs), but not induced pluripotent stem cell (iPSC)-derived neurons, largely preserve age-associated traits. Here, we report on the extent of preserved epigenetic and transcriptional aging signatures in directly converted induced neural stem cells (iNSCs). Employing restricted and integration-free expression of SOX2 and c-MYC, we generated a fully functional, bona fide NSC population from adult blood cells that remains highly responsive to regional patterning cues. Upon conversion, low passage iNSCs display a profound loss of age-related DNA methylation signatures, which further erode across extended passaging, thereby approximating the DNA methylation age of isogenic iPSC-derived neural precursors. This epigenetic rejuvenation is accompanied by a lack of age-associated transcriptional signatures and absence of cellular aging hallmarks. We find iNSCs to be competent for modeling pathological protein aggregation and for neurotransplantation, depicting blood-to-NSC conversion as a rapid alternative route for both disease modeling and neuroregeneration.

Take the shortcut – direct conversion of somatic cells into induced neural stem cells and their biomedical applications

“…While full rejuvenation to a pluripotent state resets the epigenetic signatures of the cellular source almost completely [46,47] (and reviewed in this issue [16]), little is known about epigenetic memory in iNSCs. A recent study included a comprehensive genome-wide methylome analysis of reprogrammed iNSCs [48]. Besides the expression of COL3A1, the authors did not observe any other features of fibroblasts, suggesting that the residual fibroblast memory was not sufficient to hamper neural stem cell identity and function in this case.…”

“…While Kim et al [11] reported passage of their mouse iNSCs for 3-5 times only, more recent studies demonstrated maintenance of NSC characteristics for more than 30 passages [13,14,28,34,36,38,58,73]. The neural expansion media were supplemented with either LIF and small molecules like CHIR99021, SB431542, purmorphamine, A83-01, and/or ascorbic acid [25,26,48], or basic fibroblast growth factor (FGF2) and epidermal growth factor (EGF) [14,30,32,34,36,38,[57][58][59][60]62], or even a combination of them [33,35,41]. Among the various publications reporting iNSC generation, there is extensive accordance in terms of expression of bona fide neural stem cell markers, such as SOX1, SOX2, PAX6, NESTIN, CD133, and BLBP.…”

Mechanisms of cellular rejuvenation

“…In this study, the authors used forced expression of solely Sox2 and c-Myc in addition to defined culturing conditions to generate multipotent NSCs that are able to differentiate into neurons, astrocytes, and oligodendrocytes. Interestingly, both in gene expression and methylome analysis highly passaged iNSCs were similar to iPSC-derived NSCs, whereas low passages iNSCs were not fully rejuvenated [51]. Analysis of global gene expression and DNA methylation profile revealed rejuvenation of the iNSCs (Fig.…”

“…Lineage conversion is possible between many distant cell types including blood cells, cardiomyocytes, hepatocytes, pancreatic cells, or neurons [50]. Unlike in a similar approach, where short administration of OSKM (3-6 days) followed by culturing in neural medium was sufficient to directly generate NSCs from fibroblasts [52], the cells in the Oct4-free conversion protocol never went through a pluripotent state [51]. In this study, the authors used forced expression of solely Sox2 and c-Myc in addition to defined culturing conditions to generate multipotent NSCs that are able to differentiate into neurons, astrocytes, and oligodendrocytes.…”

“…We can only speculate about potential mechanisms: aging factors might be degraded, diluted through cell divisions, removed by asymmetric cell divisions or, alternatively, only few cells free of aging factors in an aged population would successfully reprogram. In fact, the study from Sheng et al [51] suggests that cell division is the key to successfully rejuvenate cells. However, the theory of an 'elite' cell was recently rejected based on a study using single-cell mapping by lentiviral delivery of cell tags [53].…”