Therapeutic potential of spheroids of stem cells from human exfoliated deciduous teeth for chronic liver fibrosis and hemophilia A.

PURPOSE: Mesenchymal stem cell (MSC)-based cell therapies have emerged as a promising treatment option for various diseases. Due to the superior survival and higher differentiation efficiency, three-dimensional spheroid culture systems have been an important topic of MSC research. Stem cells from human exfoliated deciduous teeth (SHED) have been considered an ideal source of MSCs for regenerative medicine. Thus, in the present study, we introduce our newly developed method for fabricating SHED-based micro-hepatic tissues, and demonstrate the therapeutic effects of SHED-based micro-hepatic tissues in mouse disease models. METHODS: SHED-converted hepatocyte-like cells (SHED-HLCs) were used for fabricating spherical micro-hepatic tissues. The SHED-HLC-based spheroids were then transplanted both into the liver of mice with CCl4-induced chronic liver fibrosis and the kidney of factor VIII (F8)-knock-out mice. At 4 weeks after transplantation, the therapeutic efficacy was investigated. RESULTS: Intrahepatic transplantation of SHED-HLC-spheroids improved the liver dysfunction in association with anti-fibrosis effects in CCl4-treated mice. Transplanted SHED-converted cells were successfully engrafted in the recipient liver. Meanwhile, renal capsular transplantation of the SHED-HLC-spheroids significantly extended the bleeding time in F8-knock-out mice. CONCLUSIONS: These findings suggest that SHED-HLC-based micro-hepatic tissues might be a promising source for treating pediatric refractory diseases, including chronic liver fibrosis and hemophilia A.

Acquired isolated hypoganglionosis as a distinct entity: results from a nationwide survey.

PURPOSE: Acquired isolated hypoganglionosis (A-IH) is a late-onset intestinal pseudo-obstruction disorder and shows different pathophysiological findings from congenital isolated hypoganglionosis (C-IH). In this study, we retrospectively examined five cases of A-IH and investigated the features of A-IH. METHODS: Five cases of A-IH were extracted from a nationwide retrospective cohort study in 10 years, from which totally 355 cases of Allied Disorders of Hirschsprung's Disease (ADHD) were collected. RESULTS: Ages of onset were between 13 and 17 years in three cases, and 4 years and 4 months in ones. Initial symptoms were abdominal distension and/or chronic constipation in 4 cases, whereas one exhibited intestinal perforation. Affected lesions varied from case to case, extending various length of intestinal tracts. All cases underwent multiple operations (average: 4.6 times), such as enterostomy, resection of dilated intestines, and/or pull-through. Pathological findings showed the decreased numbers of ganglion cells and degeneration of ganglion cells, whereas the size of the plexus was normal. Currently, all cases were alive and almost all eat regular food without requiring parenteral feeding. CONCLUSION: A-IH is rare, but distinct entity characterized by different clinical courses and pathological findings from those of C-IH. The outcome is considered to be favorable after a resection of affected intestine.

Safety, efficacy and pharmacokinetics of palivizumab in off-label neonates, infants, and young children at risk for serious respiratory syncytial virus infection: a multicenter phase II clinical trial.

Background: Pediatric patients with certain rare diseases are at increased risk of severe respiratory syncytial virus (RSV) infection. However, the prophylactic use of anti-RSV antibody (palivizumab) in these patients is not indicated at present in Japan. Methods: This first-in-the-world multicenter, uncontrolled, open-label, phase II clinical trial was carried out between 28 July 2019 and 24 September 2021 at seven medical institutions in Japan to investigate the efficacy, safety, and pharmacokinetics of palivizumab in 23 subjects recruited from among neonates, infants, or children aged 24 months or younger who had any of the following conditions: pulmonary hypoplasia, airway stenosis, congenital esophageal atresia, inherited metabolic disease, or neuromuscular disease. At least four continuous doses of palivizumab were administered intramuscularly at 15 mg/kg at intervals of 30 days. Findings: Twenty-three enrolled subjects completed the study. No subject required hospitalization for RSV. Adverse events (AE) did not notably differ from the event terms described in the latest interview form. Five severe AEs required unplanned hospitalization, but resolved without RSV infection. Therapeutically effective concentrations of palivizumab were maintained throughout the study period. Interpretation: Palivizumab might be well tolerated and effective in preventing serious respiratory symptoms and hospitalization due to severe RSV infection, indicating the prophylactic use in the pediatric patients included in this study. Funding: Japan Agency for Medical Research and Development (AMED), grant numbers 19lk0201097h0001 (to MM), 20lk0201097h0002 (to MM), 21lk0201097h0003 (to MM), and 22lk0201097h0004 (to MM). AMED did not have any role in the execution of this study, analysis and interpretation of the data, or the decision to submit the results.

Efficient derivation of sympathetic neurons from human pluripotent stem cells with a defined condition.

Sympathetic neurons (SNs) are an essential component of the autonomic nervous system. They control vital bodily functions and are responsible for various autonomic disorders. However, obtaining SNs from living humans for in vitro study has not been accomplished. Although human pluripotent stem cell (hPSC)-derived SNs could be useful for elucidating the pathophysiology of human autonomic neurons, the differentiation efficiency remains low and reporter-based cell sorting is usually required for the subsequent pathophysiological analysis. To improve the efficiency, we refined each differentiation stage using PHOX2B::eGFP reporter hPSC lines to establish a robust and efficient protocol to derive functional SNs via neuromesodermal progenitor-like cells and trunk neural crest cells. Sympathetic neuronal progenitors could be expanded and stocked during differentiation. Our protocol can selectively enrich sympathetic lineage-committed cells at high-purity (≈80%) from reporter-free hPSC lines. Our system provides a platform for diverse applications, such as developmental studies and the modeling of SN-associated diseases.

Derivation of mesenchymal stromal cells from pluripotent stem cells through a neural crest lineage using small molecule compounds with defined media.

Neural crest cells (NCCs) are an embryonic migratory cell population with the ability to differentiate into a wide variety of cell types that contribute to the craniofacial skeleton, cornea, peripheral nervous system, and skin pigmentation. This ability suggests the promising role of NCCs as a source for cell-based therapy. Although several methods have been used to induce human NCCs (hNCCs) from human pluripotent stem cells (hPSCs), such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), further modifications are required to improve the robustness, efficacy, and simplicity of these methods. Chemically defined medium (CDM) was used as the basal medium in the induction and maintenance steps. By optimizing the culture conditions, the combination of the GSK3ß inhibitor and TGFß inhibitor with a minimum growth factor (insulin) very efficiently induced hNCCs (70-80%) from hPSCs. The induced hNCCs expressed cranial NCC-related genes and stably proliferated in CDM supplemented with EGF and FGF2 up to at least 10 passages without changes being observed in the major gene expression profiles. Differentiation properties were confirmed for peripheral neurons, glia, melanocytes, and corneal endothelial cells. In addition, cells with differentiation characteristics similar to multipotent mesenchymal stromal cells (MSCs) were induced from hNCCs using CDM specific for human MSCs. Our simple and robust induction protocol using small molecule compounds with defined media enabled the generation of hNCCs as an intermediate material producing terminally differentiated cells for cell-based innovative medicine.