Biology subjects

Pineda, J. R.

Publications and source records attributed to Pineda, J. R..

4 recordsLinked to original sources

bioRxiv2026

Human dental pulp stem cells grafted into C57BL/6J hippocampus differentiate towards immature neuronal like cells displaying action potential firing activity

Stem cell therapy represents a promising strategy for the replacement and functional restoration of damaged neural tissue in neurodegenerative conditions. Human dental pulp stem cells (hDPSCs) have emerged as potential candidates for neuroregeneration due to their ease of isolation, neural crest origin, neurotrophic and anti-inflammatory capacity, and demonstrated ability to differentiate in vitro into neuronal-like cells exhibiting electrophysiological activity. Although the immunomodulatory and neuroprotective properties of hDPSCs have been reported in multiple models of brain disease, their capacity to functionally integrate into host neuronal circuits remain poorly understood. In this study, we have grafted green fluorescent protein (GFP)-transduced, neural preconditioned hDPSCs into the CA1 region of the hippocampus of C57BL/6J mice. One month after transplantation, GFP+-hDPSCs survived in the brains of non-immunosuppressed mice and remained localized within the grafted area. Notably, the transplanted cells underwent in situ differentiation and exhibited a neuroblast-like phenotype, characterized by positive doublecortin expression and immature neuronal-like electrophysiological properties, like high membrane input resistance, low capacitance, and the ability to generate single action potentials after stimulation. Together, these findings provide the first evidence that hDPSCs can survive and integrate into the hippocampal network of the mouse brain at one-month post graft, supporting their potential use for future therapeutic applications in acute brain lesions and neurodegenerative disorders.

neuroscience↗

bioRxiv2024

Functional differentiation of Human Dental Pulp Stem Cells into neuron-like cells exhibiting electrophysiological activity

BackgroundHuman dental pulp stem cells (hDPSCs) constitute a promising alternative for central nervous system (CNS) cell therapy. Unlike other human stem cells, hDPSCs can be differentiated, without genetic modification, to neural cells that secrete neuroprotective factors. However, a better understanding of their real capacity to give rise to functional neurons and integrate into synaptic networks is still needed. For that, ex vivo differentiation protocols must be refined, especially to avoid the use of fetal animal serum. MethodsIn this study, we sought to improve existing differentiation protocols for obtaining functional neuron-like cells from hDPSCs. We compared the effects of the absence or presence of fetal serum during the initial expansion phase as a step prior to switching cultures to neurodifferentiation media. We improved hDPSC neurodifferentiation by adding retinoic acid (RA) and potassium chloride (KCl) pulses for 21 or 60 days and characterized the results by immunofluorescence, digital morphometric analysis, RT-qPCR and electrophysiology. ResultsWe found that neural markers like Nestin, GFAP, S100{beta} and p75NTR were expressed differently in neurodifferentiated hDPSC cultures depending on the presence or absence of serum during the initial cell expansion phase. In addition, hDPSCs previously grown as spheroids in serum-free medium exhibited in vitro expression of neuronal markers such as doublecortin (DCX), neuronal nuclear antigen (NeuN), Ankyrin-G and MAP2 after neurodifferentiation. Presynaptic vGLUT2, Synapsin-I, and excitatory glutamatergic and inhibitory GABAergic postsynaptic scaffold proteins and receptor subunits were also present in these neurodifferentiated hDPSCs. Treatment with KCl and RA increased the amount of both voltage-gated Na+ and K+ channel subunits in neurodifferentiated hDPSCs at the transcript level. Consistently, these cells displayed voltage-dependent K+ and TTX-sensitive Na+ currents as well as spontaneous electrophysiological activity and repetitive neuronal action potentials with a full baseline potential recovery. ConclusionOur study demonstrates, for the first time, that hDPSCs can be differentiated to neuronal-like cells that display functional excitability and thus evidence the potential of these easily accessible human stem cells for nerve tissue engineering. Our results highlight the importance of choosing an appropriate culture protocol to successfully neurodifferentiate hDPSCs.

cell biology↗

bioRxiv2022

HB-EGF and zinc activate EGFR to induce reactive neural stem cells in the mouse hippocampus after seizures

Hippocampal seizures mimicking mesial temporal lobe epilepsy (MTLE) cause a profound disruption of the adult neurogenic niche in mice. Seizures provoke neural stem cells to switch to a reactive phenotype (reactive-neural stem cells, React-NSCs)) characterized by multibranched hypertrophic morphology, massive activation to enter mitosis, symmetric division and final differentiation into reactive astrocytes. As a result, neurogenesis is chronically impaired. Here we, using a mouse model of MTLE, show that the epidermal growth factor receptor (EGFR) signalization pathway is key for the induction of React-NSCs and that its inhibition exerts a beneficial effect on the neurogenic niche. We show that during the initial days after the induction of seizures by a single intrahippocampal injection of kainic acid, a strong release of zinc and heparin-binding epidermal growth factor, both activators of the EGFR signalization pathway in neural stem cells, is produced. Administration of the EGFR inhibitor gefitinib, a chemotherapeutic in clinical phase IV, prevents the induction of React-NSCs and preserves neurogenesis. SignificanceIn mouse models of MTLE-HS, seizures cause a profound disruption of the hippocampal neurogenic niche and neurogenesis results chronically impaired, in agreement with what occurs in the human MTLE-HS hippocampus. Thus, the normal cognitive functions associated with neurogenesis are altered, but also the endogenous regenerative capacity that could compensate the high rate of neurons in the granule cell layer of the dentate gyrus. We provide here for the first time a molecular mechanism (the EGFR transduction pathway) regulating the induction of React-NSCs.

neuroscience↗

bioRxiv2018

Human Dental Pulp Stem Cells Grown In Neurogenic Media Differentiate Into Endothelial Cells And Promote Neovasculogenesis In The Mouse Brain.

Dental Pulp Stem Cells (DPSCs) have a demonstrated capacity to acquire neuronal-like phenotypes, suggesting their use in brain cell therapies. In the present work, we wanted to address the phenotypic fate of adult DPSCs cultured in Neurocult media (Stem Cell Technologies), a cell culture medium without serum routinely used for the expansion of adult neural stem cells (NSCs). Our results showed for the first time, that non-genetically modified adult DPSCs cultured with Neurocult generated neurosphere-like dentospheres expressing the NSC markers Nestin and GFAP, but also the vascular endothelial cell marker CD31. One month post-intracranial graft into athymic nude mice, human CD31+ or Nestin+ DPSC-derived cells were found tightly associated with brain blood vessels increasing their laminin staining. These results suggest that DPSCs integrated and contributed to an increased generation of neovasculature within brain tissue and that Neurocult medium constituted a fast and efficient way to obtain endothelial cells from human DPSCs.

bioengineering↗