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Background: Human induced pluripotent stem cell (iPSC)-derived peripheral sensory neurons present a valuable tool to model human diseases and are a source for applications in drug discovery and regenerative medicine. Clinically, peripheral sensory neuropathies can result in maladies ranging from a complete loss of pain to severe painful neuropathic disorders. Sensory neurons are located in the dorsal root ganglion and are comprised of functionally diverse neuronal types. Low efficiency, reproducibility concerns, variations arising due to genetic factors and time needed to generate functionally mature neuronal populations from iPSCs remain key challenges to study human nociception in vitro. Here, we report a detailed functional characterization of iPSC-derived sensory neurons with an accelerated differentiation protocol ("Anatomic" protocol) compared to the most commonly used small molecule approach ("Chambers" protocol). Anatomic's commercially available RealDRG™ were further characterized for both functional and expression phenotyping of key nociceptor markers.
Methods: Multiple iPSC clones derived from different reprogramming methods, genetics, age, and somatic cell sources were used to generate sensory neurons. Manual patch clamp was used to functionally characterize both control and patient-derived neurons. High throughput techniques were further used to demonstrate that RealDRGs™ derived from the Anatomic protocol are amenable to high throughput technologies for disease modelling.
Results: The Anatomic protocol rendered a purer culture without the use of mitomycin C to suppress non-neuronal outgrowth, while Chambers differentiations yielded a mix of cell types. Chambers protocol results in predominantly tonic firing when compared to Anatomic protocol. Patient-derived nociceptors displayed higher frequency firing compared to control subject with both, Chambers and Anatomic differentiation approaches, underlining their potential use for clinical phenotyping as a disease-in-a-dish model. RealDRG™ sensory neurons show heterogeneity of nociceptive markers indicating that the cells may be useful as a humanized model system for translational studies.
Conclusions: We validated the efficiency of two differentiation protocols and their potential application for functional assessment and thus understanding the disease mechanisms from patients suffering from pain disorders. We propose that both differentiation methods can be further exploited for understanding mechanisms and development of novel treatments in pain disorders.
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http://dx.doi.org/10.1186/s13287-024-03696-2 | DOI Listing |
Mol Ther Nucleic Acids
December 2024
Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine and Science, Rochester, MN 55905, USA.
There is an urgent need for agents that promote health and regeneration of cells and tissues, specifically to treat diseases of the aging nervous system. Age-associated nervous system degeneration and various diseases are driven by many different biochemical stresses, often making it difficult to target any one disease cause. Our laboratory has previously identified DNA aptamers with apparent regenerative properties in murine models of multiple sclerosis by selecting aptamers that bind oligodendrocyte membrane preparations.
View Article and Find Full Text PDFNeurobiol Pain
November 2024
Structural Genomics Consortium (SGC), UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
PIKfyve (1-phosphatidylinositol 3-phosphate 5-kinase), a lipid kinase, plays an important role in generating phosphatidylinositol (3,5)-bisphosphate (PI(3,5)P). SGC-PIKFYVE-1, a potent and selective inhibitor of PIKfyve, has been used as a chemical probe to explore pathways dependent on PIKfyve activity. Based on reported changes in membrane dynamics and ion transport in response to PIKfyve inhibition, we hypothesized that pharmacological inhibition of PIKfyve could modulate pain.
View Article and Find Full Text PDFNeuron
December 2024
Department of Anesthesiology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Institute for Translational Brain Research, MOE Frontiers Center for Brain Science, Fudan University, Shanghai 200032, China. Electronic address:
The lateral parabrachial nucleus (PBN) is critically involved in neuropathic pain modulation. However, the cellular and molecular mechanisms underlying this process remain largely unknown. Here, we report that in mice, the right-sided, but not the left-sided, PBN plays an essential role in the development of hyperalgesia following nerve injury, irrespective of the injury side.
View Article and Find Full Text PDFCurr Biol
December 2024
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA 90095, USA; Molecular Biology Institute, University of California, Los Angeles, Los Angeles, CA 90095, USA. Electronic address:
Skin-penetrating nematodes infect nearly one billion people worldwide. The developmentally arrested infective larvae (iL3s) seek out hosts, invade hosts via skin penetration, and resume development inside the host in a process called activation. Activated infective larvae (iL3as) traverse the host body, ending up as parasitic adults in the small intestine.
View Article and Find Full Text PDFElife
December 2024
Experimental Otology Group, InnerEarLab, Department of Otolaryngology, University Medical Center Göttingen, Göttingen, Germany.
To encode continuous sound stimuli, the inner hair cell (IHC) ribbon synapses utilize calcium-binding proteins (CaBPs), which reduce the inactivation of their Ca1.3 calcium channels. Mutations in the gene underlie non-syndromic autosomal recessive hearing loss DFNB93.
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