Insights into the Regulation of Neurogenesis Termination and the Secretomes of Oligodendrocytes

Neurogenesis is a key process of neurodevelopment that requires intricate temporal regulation. While the regulation from embryonic to adult neurogenesis has been more actively characterized, the molecular mechanisms regulating the postnatal termination of neurogenesis and the disappearance of embryonic radial glia, the neural stem cells (NSCs) responsible for neurogenesis, are still largely unknown. In the first chapter, we investigated the role of transcription factor PR domain-containing 16 (Prdm16) using genetic mouse models and single-cell RNA sequencing. This work provides evidence on how conditional deletion of Prdm16 expression from NSCs extends the time span in which cortical neurogenesis occurs and alters the cellular composition of the ventricular sub-ventricular zone. Mechanistically, we determined that Prdm16 induces a postnatal reduction in Vascular Cell Adhesion Molecule 1 (Vcam1). We also showed that the extended presence of radial glia and neurogenesis phenotype was rescued in Prdm16-Vcam1 double conditional knockout mice. These findings demonstrate that inhibition of Vcam1 by Prdm16 promotes the postnatal termination of neurogenesis and the disappearance of embryonic radial glia. Similarly, the function of oligodendrocyte progenitor cells (OPCs) beyond their well-characterized contribution to myelination remains to be well characterized. OPCs and oligodendrocytes are a significant cell population in the brain, comprising around 22-28% of the total cells. OPCs remain present throughout life suggesting potential additional functions beyond their canonical role in generating oligodendrocytes. In the second chapter, I investigated the secreted proteome of OPCs and oligodendrocytes to elucidate additional functions these cells may have in homeostatic and inflammatory conditions. Using an in vitro model of purified rat OPCs and oligodendrocytes, I found that both OPCs and oligodendrocytes secrete various extracellular matrix proteins under physiological conditions suggesting they may play a larger role in extracellular matrix production and remodeling than previously described. Similarly, following cytokine treatment, I found both OPCs and oligodendrocytes secretion of immunoregulators such as C2 and B2M providing evidence that these cells have an active role in the inflammatory response possibly through cross-talk with other glial and immune cells via signaling molecules like C2 and B2M. These findings shed light on OPCs and oligodendrocytes having a more active role in regulating their environment via the proteins they secrete, highlighting their multifaceted function beyond their established myelin-forming roles. Together these findings contribute novel insights into molecular regulators of neurogenesis termination and the functions of OPCs and oligodendrocytes. The novel finding of Prdm16 has a repressor of Vcam1 expression in radial glia fills a critical gap in our understanding of neural stem biology and offers valuable knowledge for future regenerative medicine efforts aimed at treating central nervous system disorders. Likewise, the unbiased secretome of OPCS and oligodendrocytes highlights several avenues for further investigation into these cells as modulators of the ECM and the inflammatory response

Human iPSC-Derived Endothelial Cells and Microengineered Organ-Chip Enhance Neuronal Development

Summary: Human stem cell-derived models of development and neurodegenerative diseases are challenged by cellular immaturity in vitro. Microengineered organ-on-chip (or Organ-Chip) systems are designed to emulate microvolume cytoarchitecture and enable co-culture of distinct cell types. Brain microvascular endothelial cells (BMECs) share common signaling pathways with neurons early in development, but their contribution to human neuronal maturation is largely unknown. To study this interaction and influence of microculture, we derived both spinal motor neurons and BMECs from human induced pluripotent stem cells and observed increased calcium transient function and Chip-specific gene expression in Organ-Chips compared with 96-well plates. Seeding BMECs in the Organ-Chip led to vascular-neural interaction and specific gene activation that further enhanced neuronal function and in vivo-like signatures. The results show that the vascular system has specific maturation effects on spinal cord neural tissue, and the use of Organ-Chips can move stem cell models closer to an in vivo condition. : Sances et al. combine Organ-Chip technology with human induced pluripotent stem cell-derived spinal motor neurons to study the maturation effects of Organ-Chip culture. By including microvascular cells also derived from the same patient line, the authors show enhancement of neuronal function, reproduction of vascular-neuron pathways, and specific gene activation that resembles in vivo spinal cord development. Keywords: organ-on-chip, spinal cord, iPSC, disease modeling, amyotrophic lateral sclerosis, microphysiological system, brain microvascular endothelial cells, spinal motor neurons, vasculature, microfluidic devic

Transplantation of human neural progenitor cells secreting GDNF into the spinal cord of patients with ALS: a phase 1/2a trial.

Amyotrophic lateral sclerosis (ALS) involves progressive motor neuron loss, leading to paralysis and death typically within 3-5 years of diagnosis. Dysfunctional astrocytes may contribute to disease and glial cell line-derived neurotrophic factor (GDNF) can be protective. Here we show that human neural progenitor cells transduced with GDNF (CNS10-NPC-GDNF) differentiated to astrocytes protected spinal motor neurons and were safe in animal models. CNS10-NPC-GDNF were transplanted unilaterally into the lumbar spinal cord of 18 ALS participants in a phase 1/2a study (NCT02943850). The primary endpoint of safety at 1 year was met, with no negative effect of the transplant on motor function in the treated leg compared with the untreated leg. Tissue analysis of 13 participants who died of disease progression showed graft survival and GDNF production. Benign neuromas near delivery sites were common incidental findings at post-mortem. This study shows that one administration of engineered neural progenitors can provide new support cells and GDNF delivery to the ALS patient spinal cord for up to 42 months post-transplantation