Human Cord Research Articles

Interleukin-33 mediated regulation of microRNAs in human cord blood-derived mast cells: Implications for infection, immunity, and inflammation.

Mast cell (MCs) activation is the driving force of immune responses in several inflammatory diseases, including asthma and allergies. MCs are immune cells found throughout the body and are equipped with numerous surface receptors that allow them to respond to external signals from parasites and bacteria as well as to intrinsic signals such as cytokines. Upon activation, MCs release various mediators and proteases that contribute to inflammation. This study aimed to identify microRNAs (miRNAs) that regulate MC response to interleukin-33 and their target genes using a model of human cord blood-derived mast cells (hCBMCs). hCBMCs were induced with 10 and 20 ng of recombinant human interleukin-33 (rhIL-33) for 6 and 24 h, respectively. Total RNA was extracted from these cells and miRNA profiling was performed using high-throughput microarrays. Differential expression of miRNAs and target analysis were performed using Transcriptome Analysis Console and Ingenuity Pathway Analysis. The most significant miRNAs in each condition were miR-6836-5p (fold change = 1.76, p = 3E-03), miR-6883-5p (fold change = -2.13, p = 7E-05), miR-1229-5p (fold change = 2.46, p = 8E-04), and miR-3613-5p (fold change = 66.7, p = 1E-06). Target analysis revealed that these miRNAs regulate mast cell responsiveness and degranulation by modulating the expression of surface receptors, adaptors, and signaling molecules in response to rhIL-33 stimulation. This study is the first miRNA profiling and target analysis of hCBMCs that will further enhance our understanding of the role of miRNAs in the immune response in a timely manner and their relevance for the development of a new therapeutic target for inflammatory disorders.

Spinal cord neural stem cells derived from human embryonic stem cells promote synapse regeneration and remyelination in spinal cord injury model rats.

Spinal cord injury (SCI) is a devastating injury that significantly impairs patients' quality of life. To date, there is no effective treatment to mitigate nerve tissue damage and restore neurological function. Neural stem cells (NSCs) derived from human embryonic stem cells (hESCs) are considered an important cell source for reconstructing damaged neural circuits and enabling axonal regeneration. Recent preclinical studies have shown that NSCs are potential therapeutic cell sources for neuroprotection and neuroregeneration in SCI animal models. NSCs can be derived from different sources and the spinal cord-specific NSCs have a higher potential for the regeneration of SCI. However, the long-term therapeutic efficacy of spinal cord-specific NSCs remains unproven. Here, we generated human spinal cord NSCs (hSCNSCs) and investigated the effects of transplanted hSCNSCs on the repair of the SCI model rats for 60 days. The transplanted hSCNSCs improved BBB scores, reduced the lesion area and promoted an increase in the number of Nestin-positive cells in the spinal cord compared to the model rats. Meanwhile, hSCNSC transplantation promoted the expression of synaptophysin, a synaptic signature protein and MBP, a protein associated with remyelination. Interestingly, BAF45D, a chromatin remodelling factor that contributes to the induction of hSCNSCs with region-specific spinal cord identity, were increased by the hSCNSC transplantation. In addition, conditioned medium derived from the hSCNSCs also promoted regenerative repair of the injured spinal cord. These results demonstrate that hSCNSCs may play a critical role in the regenerative repair of SCI.

PATH-21. SINGEL CELL TRANSCRIPTOMICS OF SPINAL EPENDYMOMA SUBTYPES IDENTIFIES INTRATUMORAL HETEROGENEITY

Abstract Spinal ependymomas comprise glial neoplasms that are classified by histological and molecular features. Two of the respective types, myxopapillary ependymomas (MPE) and spinal ependymomas (SP-EPN) are largely understudied. For each of these, our group identified clinically relevant subtypes with poorer (MPE-A; SP-EPN-A) and more favorable progression-free survival (MPE-B; SP-EPN-B) employing global methylation and transcriptomic profiling. As cellular origin and pathogenesis of these tumors are largely unknown, therapeutic options for patients with unfavorable subtypes are unavailable. We performed single cell transcriptomic sequencing of 12 formalin-fixed and paraffin-embedded spinal ependymomas including all MPE and SP-EPN subtypes and obtained a median of 10,091 cells per tumor after quality control. Uniform Manifold Approximation and Projection (UMAP) analysis revealed intertumoral heterogeneity as most samples formed separate clusters with tumors of the same subtype located in proximity. The expression of literature-defined neoplastic consensus cell states was examined for each cell of the data set and revealed intratumoral heterogeneity. Tumors expressed predominantly two programs, being a ciliated (ependymal-like) cell state (61% of all neoplastic cells) and the astrocytic state (31 % of all neoplastic cells). Interestingly, third and fourth most abundant states (mesenchymal and hypoxia) were almost exclusively expressed in MPE-A and SP-EPN-A. Furthermore, distinct clones of chromosomal copy number variations were identified across all tumors. These clones can be distinguished in UMAP representation of MPE single cell transcriptomes and to a lesser extend in SP-EPN. Next, we sought to characterize the resemblance of integrated tumor subtypes to normal spinal cord cell populations and observed a high similarity to human spinal cord ependymal cells. All spinal ependymoma subtypes display intratumoral heterogeneity with respect to the transcriptomic cell state resemblance of the neoplastic cells. Together with the transcriptomic similarity to ependymal and astrocytic cells, our data suggest the tumor origin within the astro-ependymal lineage.

Conditioned medium of human umbilical cord-mesenchymal stem cells cultivated with human cord blood serum enhances stem cell stemness and secretome profiles.

The proteins secreted by human umbilical cord mesenchymal stem cells (hUC-MSCs) may enhance tissue regeneration and wound healing. Traditional hUC-MSC cultures may not be enough since they undergo recurrent cellular senescence during large-scale production. This decreases the therapeutic ability of hUC-MSCs by altering genes and proteins that control stemness, proliferation, and protein release. Human cord blood serum (CBS) and the middle-density technique were used to evaluate hUC-MSC regeneration ability. To evaluate early-passage hMSCs for secretome-based therapies, they were expanded and secreted in vitro. After 4 days, hUC-MSCs cultivated at 3000 cells/cm2 and supplemented with 1 ng/ml CBS showed increased growth, cell proliferation, and a much lower population doubling time. CBS treatment reduced CD34, CD45, and HLA-DR levels in human umbilical cord mesenchymal stem cells (hUC-MSCs) by less than 2 %. Positive markers such CD73, CD90, and CD105 were found at >97 %, like control hUC-MSCs. Over extended culture, this combination culture can increase survival, proliferation, and stemness and postpone cell death and hUC-MSC senescence. The protein profile and hUC-MSC secretion were improved to make MSC secretion protein therapeutic. This improves cell-free treatment, proliferation, and wound healing in human skin cells. To improve cell-based transplantation or cosmeceutical manufacturing, this technique can boost hUC-MSC regeneration capacity.

Development of a quantitative prediction algorithm for human cord blood-derived CD34+ hematopoietic stem-progenitor cells using parametric and non-parametric machine learning models

The transplantation of CD34+ hematopoietic stem-progenitor cells (HSPCs) derived from cord blood serves as the standard treatment for selected hematological, oncological, metabolic, and immunodeficiency disorders, of which the dose is pivotal to the clinical outcome. Based on numerous maternal and neonatal parameters, we evaluated the predictive power of mathematical pipelines to the proportion of CD34+ cells in the final cryopreserved cord blood product adopting both parametric and non-parametric algorithms. Twenty-four predictor variables associated with the cord blood processing of 802 processed cord blood units randomly sampled in 2020–2022 were retrieved and analyzed. Prediction models were developed by adopting the parametric (multivariate linear regression) and non-parametric (random forest and back propagation neural network) statistical models to investigate the data patterns for determining the single outcome (i.e., the proportion of CD34+ cells). The multivariate linear regression model produced the lowest root-mean-square deviation (0.0982). However, the model created by the back propagation neural network produced the highest median absolute deviation (0.0689) and predictive power (56.99%) in comparison to the random forest and multivariate linear regression. The predictive model depending on a combination of continuous and discrete maternal with neonatal parameters associated with cord blood processing can predict the CD34+ dose in the final product for clinical utilization. The back propagation neural network algorithm produces a model with the highest predictive power which can be widely applied to assisting cell banks for optimal cord blood unit selection to ensure the highest chance of transplantation success.

Regenerative and Anti-Senescence Potential of Extracts from Different Parts of Black Persimmon in an In Vitro Model of Vascular Endothelium.

Antioxidants are essential for mitigating oxidative stress and maintaining vascular health. Endothelial colony-forming cells (ECFCs) are pivotal in endothelial regeneration and angiogenesis and serve as a model to study the diversity of endothelial cells across various organs. This study evaluated the effects of peel, pulp, and seed extracts from Diospyros digyna Jacq. fruit (black persimmon) on human cord blood-derived ECFCs (CB-ECFCs) to determine how the distinct antioxidant profiles of the fruit's different parts influence cellular functions. The extracts did not affect endothelial marker expression, cell proliferation, or nitric oxide production, indicating no cytotoxic or inflammatory effects. However, functional assays revealed that the seed extract significantly enhanced tube formation, increasing closed tubular networks by 1.5-fold. All extracts promoted cell migration, with the seed extract demonstrating the most substantial effect, surpassing even vascular endothelial growth factor (VEGF). Additionally, the seed extract exhibited the strongest reduction in cellular senescence, both before and after oxidative stress induction with H2O2. These findings underscore the potential of black persimmon extracts, especially from the seed, to enhance the regenerative capabilities of CB-ECFCs and reduce cellular senescence without affecting the normal endothelial phenotype. This positions them as promising candidates for developing endothelial cell therapies and advancing vascular regeneration.

Acute and chronic impact of interleukin-33 stimulation on chemokines and growth factors in human cord blood-derived mast cells.

Mast cells (MCs) are multifaceted immune cells that are capable of recognizing and responding to various stimuli by releasing an array of cytokines. We aimed to use human cord blood-derived mast cells (hCBMCs) as a model to evaluate different conditions under which chemokines and growth factors are expressed and secreted as mediators upon stimulation with the alarmin interleukin-33 (IL-33). hCBMCs were stimulated with 10 ng/mL or 20 ng/mL of recombinant human IL-33 (rhIL-33) for 6 h (acute) or 24 h (chronic). The mRNA expression of chemokines and growth factors was analyzed using microarrays, and the mediators released in the supernatant were evaluated using a multiplex assay. The mRNA expression levels of C-C chemokine ligands (CCL) CCL1, CCL5, granulocyte macrophage colony-stimulating factor (GM-CSF), and macrophage inflammatory protein (MIP)-4/CCL18 were upregulated under all conditions. In contrast, C-X-C motif chemokine ligand (CXCL) CXCL8 and CCL24 levels increased only under acute (6 h) and prolonged (24 h) conditions, respectively. Moreover, high levels of CXCL8, MIP-1α, and MIP-1β were secreted during acute inflammation, whereas the release of GM-CSF and CXCL9 proteins increased under all four conditions. This study highlights the sentinel role of MCs in mounting a specific immune response against a pathogenic-like stimulus in a timely and dose-dependent manner and is relevant for improving inflammatory treatment options.

Astrocytic Ryk signaling coordinates scarring and wound healing after spinal cord injury.

Wound healing after spinal cord injury involves highly coordinated interactions among multiple cell types, which is poorly understood. Astrocytes play a central role in creating a border against the non-neural lesion core. To do so, astrocytes undergo dramatic morphological changes by first thickening the processes and then elongating and overlap them. We show here show that the expression of a cell-surface receptor, Ryk, is induced in astrocytes after injury in both rodent and human spinal cord. Astrocyte-specific knockout of Ryk dramatically elongated the reactive astrocytes and accelerated the formation of the border and reduced the size of the scar. Astrocyte-specific knockout of Ryk also accelerated the injury responses of multiple cell types, including the resolution of neuroinflammation. Single cell transcriptomics analyses revealed a broad range of changes cell signaling among astrocytes, microglia, fibroblasts, endothelial cell, etc, after astrocyte-specific Ryk knockout, suggesting that Ryk not only regulates the injury response of astrocytes but may also regulate signals which coordinate the responses of multiple cell types. The elongation is mediated by NrCAM, a cell adhesion molecule induced by astrocyte-specific conditional knockout of Ryk after spinal cord injury. Our findings suggest a promising therapeutic target to accelerate wound healing and promote neuronal survival and enhance functional recovery.

Immunoregulation role of the erythroid cells.

Erythroid cells are the most abundant cells in the human body. In addition to their established function in gas-transportation, erythroid cells at various stages of differentiation have recently been shown to have immunomodulatory roles. Red blood cells may serve as modulators of innate and adaptive immunity, while their immature counterparts, CD71+ erythroid cells (CECs) have important immunomodulatory functions in various contexts. CECs are abundant in human cord blood and placenta, where they contribute to fetomaternal tolerance. CECs also accumulate in patients with infections, tumors, and anemia, and effectively suppress T cells by producing high levels of arginase, reactive oxygen species, programmed death-ligand 1, transforming growth factor β, and/or interleukin-10. Here, we systematically summarize the immunomodulatory functions of erythroid cells and propose some potential therapeutic applications based on their characteristics.

From banked human cord blood to induced pluripotent stem cells: New opportunities and promise in induced pluripotent stem cell banking (Review).

Umbilical cord blood (CB) is a valuable source of haematopoietic stem/progenitor cells (HSCs) and is known for the therapeutic use of these cells in treating blood disorders. However, challenges such as a high running cost and the increasing availability of treatment alternatives have made the effort to sustain CB banks difficult. This prompts the need to revisit the current CB banking initiatives to retain the relevance in this ever‑changing era parallel to the fast‑pacing development of cell‑based therapeutic technology. Cellular reprogramming has shown to have successfully converted adult somatic cells into human induced pluripotent stem cells (hiPSCs), which promise wider applications in regenerative medicine, personalized treatment and tissue engineering. CB is the youngest, primitive adult cell source that has not been affected by any prior, acquired disorders. Hence, using CB as a source of candidate cells for generating hiPSCs may be a new opportunity for banking, albeit with challenges. The present review summarizes the rise and fall of CB usage and banking for clinical therapy, the considerations in reprogramming CB into hiPSCs, the safety concerns regarding the use of hiPSC‑derived cells in clinical transplantation and the prospect of using CB‑derived hiPSCs.

Independent control of neurogenesis and dorsoventral patterning by NKX2-2.

Human neurogenesis is disproportionately protracted, lasting >10 times longer than in mouse, allowing neural progenitors to undergo more rounds of self-renewing cell divisions and generate larger neuronal populations. In the human spinal cord, expansion of the motor neuron lineage is achieved through a newly evolved progenitor domain called vpMN (ventral motor neuron progenitor) that uniquely extends and expands motor neurogenesis. This behavior of vpMNs is controlled by transcription factor NKX2-2, which in vpMNs is co-expressed with classical motor neuron progenitor (pMN) marker OLIG2. In this study, we sought to determine the molecular basis of NKX2-2-mediated extension and expansion of motor neurogenesis. We found that NKX2-2 represses proneural gene NEUROG2 by two distinct, Notch-independent mechanisms that are respectively apparent in rodent and human spinal progenitors: in rodents (and chick), NKX2-2 represses Olig2 and the motor neuron lineage through its tinman domain, leading to loss of Neurog2 expression. In human vpMNs, however, NKX2-2 represses NEUROG2 but not OLIG2, thereby allowing motor neurogenesis to proceed, albeit in a delayed and protracted manner. Interestingly, we found that ectopic expression of tinman-mutant Nkx2-2 in mouse pMNs phenocopies human vpMNs, repressing Neurog2 but not Olig2, and leading to delayed and protracted motor neurogenesis. Our studies identify a Notch- and tinman-independent mode of Nkx2-2-mediated Neurog2 repression that is observed in human spinal progenitors, but is normally masked in rodents and chicks due to Nkx2-2's tinman-dependent repression of Olig2.

Transplacental SARS-CoV-2 protein ORF8 binds to complement C1q to trigger fetal inflammation

Prenatal SARS-CoV-2 infection is associated with higher rates of pregnancy and birth complications, despite that vertical transmission rates are thought to be low. Here, multi-omics analyses of human placental tissues, cord tissues/plasma, and amniotic fluid from 23 COVID-19 mother-infant pairs revealed robust inflammatory responses in both maternal and fetal compartments. Pronounced expression of complement proteins (C1q, C3, C3b, C4, C5) and inflammatory cytokines (TNF, IL-1α, and IL-17A/E) was detected in the fetal compartment of COVID-19-affected pregnancies. While ~26% of fetal tissues were positive for SARS-CoV-2 RNA, more than 60% of fetal tissues contained SARS-CoV-2 ORF8 proteins, suggesting transplacental transfer of this viral accessory protein. ORF8-positive fetal compartments exhibited increased inflammation and complement activation compared to ORF8-negative COVID-19 pregnancies. In human placental trophoblasts in vitro, exogenous ORF8 exposure resulted in complement activation and inflammatory responses. Co-immunoprecipitation analysis demonstrated that ORF8 binds to C1q specifically by interacting with a 15-peptide region on ORF8 (C37-A51) and the globular domain of C1q subunit A. In conclusion, an ORF8-C1q-dependent complement activation pathway was identified in COVID-19-affected pregnancies, likely contributing to fetal inflammation independently of fetal virus exposure.

Quantification of porcine lower thoracic spinal cord morphology with intact dura mater using high-resolution μCT.

Spinal cord stimulation (SCS) is approved by the Food and Drug Administration for treating chronic intractable pain in the back, trunk, or limbs through stimulation of the dorsal column. Numerous studies have used swine as an analog of the human spinal cord to better understand SCS and further improve its efficacy. We performed high-resolution imaging of the porcine spinal cord with intact dura mater using micro-computed tomography (μCT) to construct detailed 3-dimensional (3D) visualizations of the spinal cord and characterize the morphology of the dorsal and ventral rootlets. We obtained spinal cords from Yorkshire/Landrace crossbred swine (N=7), stained samples with osmium tetroxide, and performed μCT imaging of the T12-T15 levels at isotropic voxel resolutions ranging from 3.3 to 50μm. We measured the anatomical morphology using the 3D volumes and compared our results to measurements previously collected from swine and human spinal cords via microdissection techniques in prior literature. While the porcine thoracic-lumbar spinal cord is a popular model for SCS, we highlight multiple notable differences compared to previously published T8-T12 human measurements including rootlet counts (porcine dorsal/ventral: 12.2±2.6, 26.6±3.4; human dorsal/ventral: 5.3±1.3, 4.4±2.4), rootlet angles (porcine ventral-rostral: 161±1°, ventral-caudal: 155±6°, dorsal-rostral: 148±9°, dorsal-caudal: 142±6°; human ventral-rostral: 170±3°, ventral-caudal: 22±10°, dorsal-rostral: 171±3°, dorsal-caudal: 15±7°), and the presence and count of dorsal rootlet bundles. Detailed measurements and highlighted differences between human and porcine spinal cords can inform variations in modeling and electrophysiological experiments between the two species. In contrast to other approaches for measuring the spinal cord and rootlet morphology, our method keeps the dura intact, reducing potential artifacts from dissection.

Towards non-invasive imaging through spinal-cord generated magnetic fields.

Non-invasive imaging of the human spinal cord is a vital tool for understanding the mechanisms underlying its functions in both healthy and pathological conditions. However, non-invasive imaging presents a significant methodological challenge because the spinal cord is difficult to access with conventional neurophysiological approaches, due to its proximity to other organs and muscles, as well as the physiological movements caused by respiration, heartbeats, and cerebrospinal fluid (CSF) flow. Here, we discuss the present state and future directions of spinal cord imaging, with a focus on the estimation of current flow through magnetic field measurements. We discuss existing cryogenic (superconducting) and non-cryogenic (optically-pumped magnetometer-based, OPM) systems, and highlight their strengths and limitations for studying human spinal cord function. While significant challenges remain, particularly in source imaging and interference rejection, magnetic field-based neuroimaging offers a novel avenue for advancing research in various areas. These include sensorimotor processing, cortico-spinal interplay, brain and spinal cord plasticity during learning and recovery from injury, and pain perception. Additionally, this technology holds promise for diagnosing and optimizing the treatment of spinal cord disorders.

Revision of leaking bleb with sterile cord wall amniotic membrane graft

To report the use of a devitalized, dehydrated human cord wall amniotic membrane to repair a late bleb leak with a scleral defect occurring long after a trabeculectomy. A 32-year-old woman presented with a bleb leak and underlying scleral thinning at the site of a trabeculectomy performed 10 years previously, inducing severe ocular hypotony. The bleb leak was persistent after multiple surgeries, including a conjunctival flap and lyophilized amniotic membrane grafting. A lyophilized cord wall amniotic membrane graft (SclerFIX®, TBF Genie Tissulaire, Mions, France) was used to repair the bleb. At 6 months postoperatively, the graft was intact, the bleb was formed, and no leakage was noted. The intraocular pressure was 12 mmHg. Cord wall amniotic membrane grafting appears to be useful for revision of a leaking bleb. It is simple and quickly accessible, requiring no additional surgery. Further studies with larger sample sizes would be ideal to confirm this result.

Further biological characterization of small molecules UM171 and SR1: In vitro effects on three hematopoietic cell populations from human cord blood

Small molecules UM171 and SR1 have already been taken into clinically-oriented protocols for the ex vivo expansion of hematopoietic stem (HSCs) and progenitor (HPCs) cells. In order to gain further insight into their biology, in the present study we have assessed their effects, both individually and in combination, on the in vitro long-term proliferation and expansion of HSCs and HPCs contained within three different cord blood-derived cell populations: MNCs (CD34+ cells = 0.8 %), LIN− cells (CD34+ cells = 41 %), and CD34+ cells (CD34+ cells >98 %). Our results show that when added to cultures in the absence of recombinant stimulatory cytokines, neither molecule had any effect. In contrast, when added in the presence of hematopoietic cytokines, UM171 and SR1 had significant stimulatory effects on cell proliferation and expansion in cultures of LIN− and CD34+ cells. No significant effects were observed in cultures of MNCs. The effects of both molecules were more pronounced in cultures with the highest proportion of CD34+ cells, and the greatest effects were observed when both molecules were added in combination. In the absence of small molecules, cell numbers reached a peak by days 25–30, and then declined; whereas in the presence of UM171 or/and SR1 cell numbers were sustained up to day 45 of culture. Our results indicate that besides CD34+ cells, LIN− cells could also be used as input cells in clinically-oriented expansion protocols, and that using both molecules simultaneously would be a better approach than using only one of them.

In vitro regenerative effects of a pooled pathogen-reduced lyophilized human cord blood platelet lysate for wound healing applications.

Cord blood platelets, easily obtained from blood units not suitable for haematopoietic stem cell transplantation, represent an abundant source of growth factors for use in wound healing. Although several protocols have been described for platelet lysate production, no standard manufacturing protocol is available. The use of pooled cord blood platelets could thus facilitate standardization. In this study, the effect of varying concentrations (up to 20%) of a pooled pathogen-reduced lyophilized cord blood platelet lysate (PRL-CBPL) was investigated in different cell types involved in the wound healing process. The effect of heparin addition was also evaluated. In parallel, a comparison was performed with a single donor cord blood platelet lysate (SD-CBPL). The effect of PRL-CBPL on the viability and proliferation of different cell lines (L929 mouse fibroblasts and HaCaT keratinocytes) and human primary cells (fibroblasts-NHDF, coronary artery smooth muscle cells-HCASMC and coronary artery endothelial cells-HCAEC), on HaCaT migration and the chemotactic effect on human monocytes (THP-1) was evaluated. PRL-CBPL showed a lower PDGF-AB amount compared to SD-CBPL. Differing concentrations of both CBPL were necessary to influence cell viability and proliferation. 3% was the optimal concentration for L929 and HaCaT as well as for NHDF and HCASMC, while HCAEC required 10%. The effect of added heparin was more evident on SD-CBPL and in particular on NHDF and HCASMC proliferation. Keratinocyte scratch closure was obtained with 3 and 5% PRL-CBPL and SD-CBPL respectively. Both CBPLs caused an increase in the number of migrated THP-1 monocytes in a concentration-dependent manner up to 20% with a higher monocyte migration for SD-CBPL with respect to PRL-CBPL and in cells treated with heparin. The data obtained suggest that PRL-CBPL is an effective standardized alternative to SD-CBPL.

Valproic acid-induced teratogenicity is driven by senescence and prevented by Rapamycin in human spinal cord and animal models.

Valproic acid (VPA) is an effective and widely used anti-seizure medication but is teratogenic when used during pregnancy, affecting brain and spinal cord development for reasons that remain largely unclear. Here we designed a genetic recombinase-based SOX10 reporter system in human pluripotent stem cells that enables tracking and lineage tracing of Neural Crest cells (NCCs) in a human organoid model of the developing neural tube. We found that VPA induces extensive cellular senescence and promotes mesenchymal differentiation of human NCCs. We next show that the clinically approved drug Rapamycin inhibits senescence and restores aberrant NCC differentiation trajectory after VPA exposure in human organoids and in developing zebrafish, highlighting the therapeutic promise of this approach. Finally, we identify the pioneer factor AP1 as a key element of this process. Collectively our data reveal cellular senescence as a central driver of VPA-associated neurodevelopmental teratogenicity and identifies a new pharmacological strategy for prevention. These results exemplify the power of genetically modified human stem cell-derived organoid models for drug discovery.

Decellularized Biohybrid Nerve Promotes Motor Axon Projections.

Developing nerve grafts with intact mesostructures, superior conductivity, minimal immunogenicity, and improved tissue integration is essential for the treatment and restoration of neurological dysfunctions. A key factor is promoting directed axon growth into the grafts. To achieve this, biohybrid nerves are developed using decellularized rat sciatic nerve modified by in situ polymerization of poly(3,4-ethylenedioxythiophene) (PEDOT). Nine biohybrid nerves are compared with varying polymerization conditions and cycles, selecting the best candidate through material characterization. These results show that a 1:1 ratio of FeCl3 oxidant to ethylenedioxythiophene (EDOT) monomer, cycled twice, provides superior conductivity (>0.2mScm-1), mechanical alignment, intact mesostructures, and high compatibility with cells and blood. To test the biohybrid nerve's effectiveness in promoting motor axon growth, human Spinal Cord Spheroids (hSCSs) derived from HUES 3 Hb9:GFP cells are used, with motor axons labeled with green fluorescent protein (GFP). Seeding hSCS onto one end of the conduit allows motor axon outgrowth into the biohybrid nerve. The construct effectively promotes directed motor axon growth, which improves significantly after seeding the grafts with Schwann cells. This study presents a promising approach for reconstructing axonal tracts in humans.

Reliability of task-based fMRI in the dorsal horn of the human spinal cord

Abstract The application of functional magnetic resonance imaging (fMRI) to the human spinal cord is still a relatively small field of research and faces many challenges. Here we aimed to probe the limitations of task-based spinal fMRI at 3T by investigating the reliability of spinal cord blood oxygen level dependent (BOLD) responses to repeated nociceptive stimulation across 2 consecutive days in 40 healthy volunteers. We assessed the test–retest reliability of subjective ratings, autonomic responses, and spinal cord BOLD responses to short heat-pain stimuli (1 s duration) using the intraclass correlation coefficient (ICC). At the group level, we observed robust autonomic responses as well as spatially specific spinal cord BOLD responses at the expected location, but no spatial overlap in BOLD response patterns across days. While autonomic indicators of pain processing showed good-to-excellent reliability, both β-estimates and z-scores of task-related BOLD responses showed poor reliability across days in the target region (gray matter of the ipsilateral dorsal horn). When taking into account the sensitivity of gradient-echo echo planar imaging (GE-EPI) to draining vein signals by including the venous plexus in the analysis, we observed BOLD responses with fair reliability across days. Taken together, these results demonstrate that heat-pain stimuli as short as 1 s are able to evoke a robust and spatially specific BOLD response, which is, however, strongly variable within participants across time, resulting in low reliability in the dorsal horn gray matter. Further improvements in data acquisition and analysis techniques are thus necessary before event-related spinal cord fMRI as used here can be reliably employed in longitudinal designs or clinical settings.