Bypassing Pluripotency: Scalable Direct Reprogramming of Dermal Fibroblasts into Functional Motor Neurons for Regenerative Therapy

Researchers at the Massachusetts Institute of Technology have developed a highly efficient protocol for the direct conversion of dermal fibroblasts into mature motor neurons, circumventing the intermediate pluripotent stem cell stage traditionally required for neuronal differentiation. This lineage reprogramming approach yields more than ten motor neurons per starting fibroblast in murine cells and achieves successful engraftment and synaptic integration in the host brain, representing a significant advance toward scalable cell-replacement therapies for spinal cord injury and motor neuron diseases such as amyotrophic lateral sclerosis (ALS).

The conventional route for generating patient-specific neurons relies on reprogramming somatic cells into induced pluripotent stem cells (iPSCs) followed by directed differentiation. Although robust, this two-step process is time-consuming (typically requiring several weeks) and frequently results in incomplete maturation or heterogeneous populations due to cells becoming trapped in intermediate transcriptional states. In contrast, direct lineage conversion induces terminal differentiation without reversion to pluripotency, thereby reducing both duration and epigenetic variability.

Employing mouse embryonic fibroblasts, the laboratory of Katie Galloway systematically refined a previously reported cocktail of transcription factors. By iterative elimination of individual factors, the investigators identified a minimal set of three proneural and motor neuron-specifying transcription factors—Neurogenin-2 (NGNgn2), Islet-1 (Isl1), and Lhx3—as sufficient to drive neuronal fate when co-expressed at balanced stoichiometry. Delivery of these three factors via a single polycistronic retroviral or lentiviral vector ensured uniform expression levels across the transduced population, markedly improving reprogramming consistency compared with multi-vector approaches.

To overcome the historically low efficiency of direct conversion (

Electrophysiological recordings and calcium imaging confirmed that the resulting neurons exhibit spontaneous and evoked action potentials, express cholinergic markers characteristic of spinal motor neurons, and display complex dendritic arborization. When transplanted into the striatum of immunocompetent adult mice, the reprogrammed neurons survived for at least two weeks, extended neurites, and formed synapses with host circuitry, as evidenced by juxtaposition of pre- and postsynaptic markers.

Parallel experiments with human dermal fibroblasts demonstrated that a slightly modified transcription factor combination achieves direct conversion, albeit with lower efficiency (estimated 10–30%), over a five-week period. Although further optimization is required, this proof-of-concept in human cells suggests translational feasibility.

The findings, published in two companion articles in Cell Systems (Wang et al., 2025), establish a streamlined, high-yield platform for generating transplantable motor neurons without iPSC intermediacy. By increasing the scalability of autologous or allogeneic neuronal production, this methodology may facilitate preclinical testing and eventual clinical application of cell-replacement strategies for traumatic spinal cord injury and neurodegenerative conditions affecting motor circuitry.

The research was supported by the National Institute of General Medical Sciences and the National Science Foundation Graduate Research Fellowship Program.