Umbilical Cord Blood Cells Research Articles

Changes of dendritic cell and natural killer cell on the cord blood with idiopathic fetal growth restriction

Objective To investigate the characteristics of dendritic cells (DC) and natural killer cells (NK) in umbilical cord blood of pregnant patients diagnosed with idiopathic fetal growth restriction (IFGR). Methods A prospective study cohort of IFGR patients was established who were in the third trimester (28–36 weeks), with a healthy, pregnant woman cohort selected as controls. Umbilical cord blood was collected. Results The study included 50 pregnant women in the IFGR group and 50 pregnant women in the healthy, control group. The incidence of SGA in the IFGR group was 52.0%, and the incidence of preterm birth was 18.0%. The incidence of neonatal complications in neonates with live birth in the IFGR group was 12.0%. The birth weight, body length and placental weight of the newborns in the IFGR group were significantly lower than those in the control group (p < .05). Flow cytometry revealed no significant difference in the proportion or maturity of DC in umbilical cord blood between IFGR group and control group (p > .05). The proportion of NK cells in umbilical cord blood of IFGR group was significantly higher than that of normal control group. The proportion of CD56dimCD16+ NK cells was also significantly higher than that of the normal control group (p < .05), but the expression of NK cell surface killing activator receptor NKG2D and inhibitory receptor NKG2A was not statistically significant (p > .05). Conclusion The number and proportion of DC cells in cord blood may not be the key factors affecting the outcomes observed during FGR pregnancies. However, we found the proportion of NK cells in cord blood to be significantly increased, as well as the ratio of CD56dimCD16 + NK to CD56highCD16-NK to be imbalanced, which may be one of the pathogenesis of the pathological pregnancy leading to IFGR.

Stem Cells for Aging-Related Disorders.

This review captures recent advances in biological and translational research on stem cells. In particular, we discuss new discoveries and concepts regarding stem cell treatment of aging-related disorders. A myriad of stem cell sources exists, from hematopoietic to mesenchymal and neural cell lineages. We examine current applications of exogenous adult bone marrow-derived stem cells as an effective and safe transplantable cell source, as well as the use of electrical stimulation to promote endogenous neurogenesis for Parkinson's disease. We also explore the potential of transplanting exogenous umbilical cord blood cells and mobilizing host resident stem cells in vascular dementia and aging. In addition, we assess the ability of small molecules to recruit resident stem cells in Alzheimer's disease. Finally, we evaluate mechanisms of action recently implicated in stem cell therapy, such as the role of long non-coding RNAs, G-protein coupled receptor 5, and NeuroD1. Our goal is to provide a synopsis of recent milestones regarding the application of stem cells in aging.

Umbilical cord blood therapy modulates neonatal hypoxic ischemic brain injury in both females and males

Preclinical and clinical studies have shown that sex is a significant risk factor for perinatal morbidity and mortality, with males being more susceptible to neonatal hypoxic ischemic (HI) brain injury. No study has investigated sexual dimorphism in the efficacy of umbilical cord blood (UCB) cell therapy. HI injury was induced in postnatal day 10 (PND10) rat pups using the Rice-Vannucci method of carotid artery ligation. Pups received 3 doses of UCB cells (PND11, 13, 20) and underwent behavioural testing. On PND50, brains were collected for immunohistochemical analysis. Behavioural and neuropathological outcomes were assessed for sex differences. HI brain injury resulted in a significant decrease in brain weight and increase in tissue loss in females and males. Females and males also exhibited significant cell death, region-specific neuron loss and long-term behavioural deficits. Females had significantly smaller brains overall compared to males and males had significantly reduced neuron numbers in the cortex compared to females. UCB administration improved multiple aspects of neuropathology and functional outcomes in males and females. Females and males both exhibited injury following HI. This is the first preclinical evidence that UCB is an appropriate treatment for neonatal brain injury in both female and male neonates.

Single-cell chromatin accessibility landscape of human umbilical cord blood in trisomy 18 syndrome

BackgroundTrisomy 18 syndrome (Edwards syndrome, ES) is a type of aneuploidy caused by the presence of an extra chromosome 18. Aneuploidy is the leading cause of early pregnancy loss, intellectual disability, and multiple congenital anomalies. The research of trisomy 18 is progressing slowly, and the molecular characteristics of the disease mechanism and phenotype are still largely unclear.ResultsIn this study, we used the commercial Chromium platform (10× Genomics) to perform sc-ATAC-seq to measure chromatin accessibility in 11,611 single umbilical cord blood cells derived from one trisomy 18 syndrome patient and one healthy donor. We obtained 13 distinct major clusters of cells and identified them as 6 human umbilical cord blood mononuclear cell types using analysis tool. Compared with the NC group, the ES group had a lower ratio of T cells to NK cells, the ratio of monocytes/DC cell population did not change significantly, and the ratio of B cell nuclear progenitor and megakaryocyte erythroid cells was higher. The differential genes of ME-0 are enriched in Human T cell leukemia virus 1 infection pathway, and the differential peak genes of ME-1 are enriched in apopotosis pathway. We found that CCNB2 and MCM3 may be vital to the development of trisomy 18. CCNB2 and MCM3, which have been reported to be essential components of the cell cycle and chromatin.ConclusionsWe have identified 6 cell populations in cord blood. Disorder in megakaryocyte erythroid cells implicates trisomy 18 in perturbing fetal hematopoiesis. We identified a pathway in which the master differential regulatory pathway in the ME-0 cell population involves human T cell leukemia virus 1 infection, a pathway that is dysregulated in patients with trisomy 18 and which may increase the risk of leukemia in patients with trisomy 18. CCNB2 and MCM3 in progenitor may be vital to the development of trisomy 18. CCNB2 and MCM3, which have been reported to be essential components of the cell cycle and chromatin, may be related to chromosomal abnormalities in trisomy 18.

Screening Fetal Hemoglobin Inducing Compounds in Human Umbilical Cord Blood-Derived Erythroid Progenitor Cells

Background: Human Umbilical Cord Blood-Derived Erythroid Progenitor Cells-2 (HUDEP-2) cells are an immortalized human CD34+ hematopoietic stem and progenitor cell (HSPC)-derived erythroid precursor cell line that expresses BCL11A and β-globin. HUDEP-1 cells are an immortalized human CD34+ HSPC-derived erythroid precursor cell line that does not express BCL11A, predominantly expresses γ-globin, and exhibits an increase in γ-globin after differentiation. High baseline γ-globin and difficulty in distinguishing between true γ-globin induction and changes in HUDEP-1 differentiation, which increases g-globin, combined with the lack of β-globin expression, make HUDEP-1 cells a suboptimal system in which to study hemoglobin switching. HUDEP-2 cells have been used successfully to demonstrate HbF induction in several gene editing experiments involving deletion of BCL11A enhancer regions and substitution mutations in the γ-globin locus. HUDEP-2 also express very low levels of γ-globin; this has led to the hope that potential HbF inducing compounds can be screened in HUDEP-2 cells, causing them to express significant amounts of γ-globin, and potentially a reduction in β-globin expression, demonstrating hemoglobin switching.Objectives: Determine if HUDEP-2 cell lines are suitable systems in which to examine pharmacologic hemoglobin switching. We will determine if γ-globin induction can be achieved in HUDEP-2 cells from hydroxyurea, metformin, piceatannol, decitabine, or pomalidomide at doses that induce HbF in human HSPCs and raise γ-globin expression in HUDEP-1 cells.Methods: We performed drug treatment on differentiated HUDEP-1 andHUDEP-2 cells as well as hematopoietic stem and progenitor cells (HSPCs) from SCD patients with hydroxyurea, metformin, piceatannol, decitabine and pomalidomide at different doses. The effect of the drugs on HUDEP-1 differentiation was determined by measurement of Band3 by quantitative western blot. After 7 days of drug treatment, HbF was measured by western blot and HPLC in HSPCs, and by RT-qPCR (gamma globin expression) and western blot in HUDEP-1 and HUDEP-2.Results: Hydroxyurea, metformin, piceatannol, decitabine and pomalidomide all significantly increased γ-globin expression in HUDEP-1 cells, and HbF in HSPCs derived from patients with SCD. However, metformin was shown to increase HUDEP-1 differentiation, as measured by a rise in Band 3, which could explain the rise in γ-globin expression. HPLC analysis of the culture of HSPCs derived from patients with SCD showed that drug treatment with 30 µM hydroxyurea, 100 µM Metformin, 25 µM piceatannol or 200 nM decitabine produced a 2.3 fold, 2.1 fold, 3.1 fold, and 2.9 fold increase in HbF respectively. No change was observed in γ-globin expression or HbF in HUDEP-2 cells with any of the 5 drugs tested.Conclusions: γ-globin is not inducible in HUDEP-2 cells using hydroxyurea, metformin, piceatannol, decitabine and pomalidomide, known HbF inducing drugs that are believed to act through different pathways and mechanisms. It is therefore unlikely that HUDEP-2 cells will be effective in screening drug compounds for HbF induction activity. HUDEP-1 cells do show increased γ -globin levels in response to these agents, but care must be taken to show that the rise in γ-globin is not due to further differentiation of HUDEP-1 cells. DisclosuresNo relevant conflicts of interest to declare.

Multi-Phase Ex Vivo Generation of Platelet-like Particles from CD34+ Cord Blood Cell-Derived Megakaryocytes

Currently, transfusable platelet units are solely derived from volunteer donors, but limited shelf-life and risk of infectious disease transmission present several logistical challenges for platelet storage and distribution at hospitals and blood centers. Thus, there is a growing interest in producing transfusable platelets in culture. Ex vivo platelet production is currently limited by the small quantity of input cells available for megakaryocytic (MK) differentiation, and low efficiency of generating platelets in vitro from differentiated MK cells. We present a serum-free, stroma-free, multi-phase culture process for production of CD41+/CD42+ platelet-like-particles (PLPs) from CD34+-selected umbilical cord blood (CB) cells. Our method greatly expands the number of input CD34+ cells towards the MK lineage, thus increasing the total number of CD41+/CD42+ mature MK cells and PLPs per input CD34+ cell by several fold compared to cultures without pre-expansion.CD34+ CB cells were expanded using histone deacetylase inhibitor valproic acid (VPA) and cytokines (primary culture),transferred to low oxygen conditions for further expansion and MK commitment (secondary culture), and then transferred to a final stage where MK cells mature and are fragmented under shear conditions (tertiary culture). We demonstrate that expanded pools of CD34+ CB cells differentiate into mature MK cells and produce functional CD41+/CD42+ PLPs under in vitro shear conditions.The length of the primary pre-expansion culture had a strong effect on CD41+/CD42+ presentation in subsequent culture steps. We evaluated the growth and differentiation of CD34+ cells for 16 independent CB units pre-expanded for 5 or 7 days in the presence of VPA. While 7-day VPA expansion decreased the peak percentage of CD41+/CD42+ cells in comparison to 5-day VPA expansion (E7: 21 ± 1.6% vs. E5: 30 ± 1.9% CD41+/CD42+ p=0.001), total MK cell production tended to be greater for 7-day vs. 5-day VPA expansion due to increased cell proliferation (E7: 381 ± 90 vs. E5: 290 ± 52 CD41+/CD42+ MK cells per input CD34+ cell). In comparison, unexpanded cultures produced 118 ± 29 CD41+/CD42+ cells per input CD34+ cell. Also, DNA content, as measured by percent high ploidy CD41+ cells, was positively correlated with length of pre-expansion (7-day VPA expanded: 13 ± 0.2%, 5-day VPA: 10 ± 0.5%, unexpanded: 7.1 ± 0.4% percent ≥8N CD41+ cells, p<0.001).In secondary and tertiary culture, we evaluated the effect of shear force on proliferation, MK maturation, and PLP generation. Culturing cells in an orbital shaker increased the percentage of CD41+/CD42+ cells for VPA treated cultures (shear 35 ± 2.8% vs. static 24 ± 2.6%, p = 0.02), but decreased the number of total cells in culture (shear 1270 ± 298 vs. static 2030 ± 323 total nucleated cells per CD34+ input cell). Individual MK cells within a heterogenous bulk culture were shown to release CD41+/CD42+ PLPs asynchronously over the 6-day tertiary phase. Time-course studies of PLP sampling every 24 hours revealed that 2200± 640 CD41+/CD42+ PLPs were generated per input CD34+ cell (7-day VPA expansion, n=3) at peak concentration. The CD41+/CD42+ PLP production rate of 7-day VPA cultures was greater than that for unexpanded CB CD34+ cells (1100± 120 CD41+/CD42+ PLPs per input CD34+ cell, n=3). We analyzed the morphology and functionality of generated PLPs and showed that generated PLPs became PAC-1+ and CD62P+ when activated by thrombin receptor agonists. PLPs also showed typical formations of stretched microtubule coils and actin stress fibers when spread on fibrinogen-coated glass surfaces upon activation. Interestingly, PLPs released by 7-day VPA cultures were found to be smaller than PLPs released by unexpanded and 5-day VPA cultures, and more comparable in size to donor platelets. Our study presents a proof-of-concept that the number of functional PLPs can be significantly increased via a multi-phase MK production process. DisclosuresNo relevant conflicts of interest to declare.

Umbilical Cord Blood Cells Express mRNA for Neuronal Genes MAP2 and NEFL.

Hematopoietic tissue-derived cells including stem cells have been shown to differentiate into or fuse with solid organ tissue-specific cells. Among hematopoietic tissue cells, umbilical cord blood (UCB)-derived stem cells have the advantage of a higher engraftment potential. It was the purpose of this study to gather evidence on a molecular level of UCB-derived cells and stem cells generating or fusing with solid-organ tissue specific cells such as CNS-specific cells as a model for potential treatment of injured tissue.Fresh UCB was obtained by standard UCB banking procedures. Apheresis-derived mononuclear cells (adult MNC) were obtained from normal stem cells donors who underwent a 4-day G-CSF mobilization treatment. UCB cells (4 individual UCB harvests) and adult MNC (4 individuals) were selected for CD133/1+ or CD14+ or CD3+ cells by immunomagnetobead separation (autoMACS; Miltenyi Biotec). Each specimen contained between 8 and 20 × 106 selected cells at a purity of >90%.Real-time PCR was performed on the ABI Prism 7900 using the Assays-on-Demand for various neuronal genes such as Neurofilament (NEFL), Microtubule-associated protein 2 (MAP2), βTubulin 3 (TUBB3) and Nestin (NES), and the cylophilin Pre-Developed Assay Reagent (Applied Biosystems) without multiplexing. The ddCt method was used to calculate fold-difference in neuronal gene expression in the cell specimens relative to Stratagene's normal lymph node RNA.Neuronal cell-specific NEFL and MAP2 expression was found in both, UCB and adult MNC, but not TUBB3 or NES expression. The MAP2 expression was in favor of CD133−, CD14− and CD3− selected cells, whereas NEFL expression was significantly higher in CD3+ selected cells as compared with CD3− cells in both, UCB cells and adult MNC. [Display omitted] These preliminary results indicate that UCB and mobilized adult blood contain cell subsets that express mRNA for neuronal cell specific genes. NEFL expression seems to be higher in CD3-selected cells, whereas MAP2 and NEFL gene expression in progenitor cell subsets (CD133+) was found to be less.As shown by us in a clinical sex-mismatched UCB transplant setting and by others hematopoietic tissue-derived neuronal-like cells are identified in CNS tissue. Circulating cells expressing neuronal-specific genes could contribute to homeostasis of CNS tissue as known for the hematopoietic tissue.

Teaching tolerance: Diverse cellular interactions enable healthy maturation

The fetal immune system is distinguishable from the adult immune system by a higher degree of tolerance to foreign antigens. This tolerance is important for fetal development within the 'foreign' maternal environment, and during birth when barrier surfaces are first colonized by microbiota. Immune responses against the wave of newly colonizing microbiota would cause massive damage to barrier tissues, so neonates need suppressed immune responses and instead rely on maternal antibodies for protection. It is becoming clear that the early-life establishment of tolerance could impact immune homeostasis and predisposition to autoimmune diseases throughout life. However, it is not well understood how and when perinatal tolerogenic immune responses switch towards adult-like effector immune responses. Here, we present a new report on the differences between cells from perinatal umbilical cord blood (UCB) and adult peripheral blood mononuclear cells (PBMC), which give mechanistic insights into fetal tolerogenic responses.

Restoration of Aged Hematopoietic Cells by Their Young Counterparts Through Instructive Microvesicle Release

This study addresses the potential to reverse age-associated morbidity by establishing methods to restore the aged hematopoietic system. Parabiotic animal models indicated that young secretome could restore aged tissues, leading us to establish a heterochronic transwell system with aged mobilized peripheral blood (MPB), co-cultured with young MPB or umbilical cord blood (UCB) cells. Functional studies and omics approaches indicate that the miRNA cargo of microvesicles (MVs) restores the aged hematopoietic system. The in vitro findings were validated in immune deficient (NSG) mice carrying an aged hematopoietic system, improving aged hallmarks such as increased lymphoid:myeloid ratio, decreased inflammation and cellular senescence. Elevated MYC and E2F pathways, and decreased p53 were key to hematopoietic restoration. These processes require four restorative miRs that target the genes for transcription/differentiation, namely PAX and phosphatase PPMIF. These miRs when introduced in aged cells were sufficient to restore the aged hematopoietic system in NSG mice. The aged MPBs were the drivers of their own restoration, as evidenced by the changes from distinct baseline miR profiles in MPBs and UCB to comparable expressions after exposure to aged MPBs. Restorative natural killer cells eliminated dormant breast cancer cells in vivo, indicating the broad relevance of this cellular paradigm - preventing and reversing age-associated disorders such as clearance of early malignancies and enhanced responses to vaccine and infection.

Diverse actions of cord blood cell therapy for hypoxic-ischemic encephalopathy.

Perinatal hypoxic‐ischemic encephalopathy (HIE) is a major cause of neonatal death and permanent neurological deficits. However, effective treatments have not yet been established, except therapeutic hypothermia, which is not effective for severe HIE; therefore, developing a novel therapy for HIE is of the utmost importance. Stem cell therapy has recently been identified as a novel therapy for HIE. Among the various stem cell sources, ethical hurdles can be avoided by using stem cells that originate from non‐embryonic or non‐neural tissues, such as umbilical cord blood cells (UCBCs), which are readily available and can be exploited for autologous transplantations. Human UCBs are a rich source of stem and progenitor cells. Many recent studies have reported the treatment effect of UCBCs. Additionally, phase I clinical trials have already been conducted, showing this therapy’s safety and feasibility. One advantage of stem cell therapies, including UCBC administration, is that they exert treatment effects through multifaceted mechanisms. According to the findings of several publications, replacement of lost cells, namely, engraftment and differentiation into neuronal cells, is not likely to be the main mechanism. However, the association between UCBCs and various mechanism of action, such as neurogenesis, angiogenesis, and anti‐inflammation, has been suggested in many studies, and most mechanisms are due to growth factors secreted from UCBCs. These diverse actions of UCBC treatment are expected to exert a substantial effect on HIE, which has a complex injury mechanism.

Umbilical cord blood and cord tissue mesenchymal stromal cells in children with cerebral palsy

Background & Aim: Umbilical cord blood (CB) and mesenchymal stromal cells (MSC) have shown safety in children with cerebral palsy (CP). While early phase clinical trials suggest potential functional benefit, small sample sizes, heterogeneous populations, and variable cell doses have impaired accurate assessment of motor gains following treatment. Aim: Describe change in gross motor function in young children with CP after treatment with high-dose allogeneic unrelated donor CB or allogeneic, third party human cord tissue-derived MSC (hCT-MSC) 12 months post treatment. Methods, Results & Conclusion: We conducted a phase 2 randomized trial of 90 children ages 2-4 years with hypertonic CP due to hypoxic ischemic encephalopathy, periventricular leukomalacia, or in utero stroke/bleed. Randomization, stratified by etiology and severity (Gross Motor Function Classification System (GMFCS) level), was to: (a) 10×107 total nucleated cells (TNC)/kg allogeneic CB at baseline, (b) three doses of 2×106 cells/kg hCT-MSC given at baseline, 3, and 6 months, or (c) Natural History in which 10×107 TNC/kg allogeneic CB was given at one year. Infusions were intravenous and premedicated with diphenhydramine and methylprednisolone without immunosuppression. Primary outcome was change in motor function one-year post enrollment, measured by the Gross Motor Function Measure-66 (GMFM-66). Ninety children (median 3.5 years) were randomized and completed baseline and 6-month evaluations. Due to the COVID pandemic, only 68 completed 12-month assessments. The only adverse events (AEs) related to the cell products were 8 transient infusion reactions (3 CB, 5 hCT-MSC). An additional 95 non-severe AEs and 33 severe AEs were unrelated to the products. At 6 months, there was no statistical difference in change in GMFM-66 scores between Natural History (n=31) and either treatment group (CB n=31, hCTMSC n=28). At 12 months, after adjustment for baseline GMFCS level, GMFM-66 score, and etiology of CP, the mean GMFM-66 score of the hCT- MSC group (n=23) was 1.4 points higher than Natural History (n=25;95% CI: -1.1, 4.0;p=0.27) and the CB group (n=20) was 3.3 points higher than Natural History (95% CI: 0.59, 5.93;p=0.02). High dose allogeneic CB, but not hCT-MSC, infusion is associated with gross motor improvement in young children with CP, consistent with the dose effect in a prior study of autologous CB. A phase 3 randomized placebo-controlled study should be performed to confirm the CB observation.

Evidence that hematopoietic stem cells in human umbilical cord blood is infectable by dengue virus: proposing a vertical transmission candidate

BackgroundRecent studies have shown that dengue virus (DENV) can efficiently infect bone marrow hematopoietic stem cells (HSCs) as well as the placenta of pregnant women. Although mother-to-infant vertical transmission of DENV through the placenta has been well documented, the evidence of cell-associated vertical transmission is still unknown. Whether DENV can infect umbilical cord blood (UCB) cells before reaching the fetus remains to be explored. Here, we proposed that human UCB cells were permissive to the DENV infection and DENV infected CD133+ and CD34+ HSCs are reservoir of the virus that could be reactivated upon re-culturing in suitable cells. MethodsHuman UCB cells were freshly obtained and subjected to DENV infection. Multicolor flow cytometry (MFCM) was used to demonstrate the phenotypes of the infected HSC populations. Immunofluorescence analysis (IFA) and T-distributed Stochastic Neighbor Embedding (t-SNE) were used to show the association of the DENV antigen, non-structural protein1 (NS1) with HSCs. Key findingsUCB cells were highly permissive to DENV infection. DENV altered the phenotype of the infected HSC population, increased the expression of HSCs, and affected the balance of transcription factors (TFs, GATA1/2/3). IFA revealed the association of the DENV antigen, non-structural protein1 (NS1), with CD34+ and CD133+ cells. T-distributed Stochastic Neighbor Embedding (t-SNE) analysis revealed heterogeneity in the distribution of CD133+NS1+, and CD34+ NS1+ cells. DENV particles were recovered from CD133+ and CD34+ cells even when virus production in the supernatant was negligible. SignificanceWe predict that infection of CD133+ and CD34+ cells in the UCB serve as reservoirs for the amplification of DENV in UCB prior to the virus reaching the fetus and facilitate vertical transmission.

Umbilical cord blood-derived microglia-like cells to model COVID-19 exposure

Microglia, the resident brain immune cells, play a critical role in normal brain development, and are impacted by the intrauterine environment, including maternal immune activation and inflammatory exposures. The COVID-19 pandemic presents a potential developmental immune challenge to the fetal brain, in the setting of maternal SARS-CoV-2 infection with its attendant potential for cytokine production and, in severe cases, cytokine storming. There is currently no biomarker or model for in utero microglial priming and function that might aid in identifying the neonates and children most vulnerable to neurodevelopmental morbidity, as microglia remain inaccessible in fetal life and after birth. This study aimed to generate patient-derived microglial-like cell models unique to each neonate from reprogrammed umbilical cord blood mononuclear cells, adapting and extending a novel methodology previously validated for adult peripheral blood mononuclear cells. We demonstrate that umbilical cord blood mononuclear cells can be used to create microglial-like cell models morphologically and functionally similar to microglia observed in vivo. We illustrate the application of this approach by generating microglia from cells exposed and unexposed to maternal SARS-CoV-2 infection. Our ability to create personalized neonatal models of fetal brain immune programming enables non-invasive insights into fetal brain development and potential childhood neurodevelopmental vulnerabilities for a range of maternal exposures, including COVID-19.

Umbilical cord blood cells in the treatment of patients with schizophrenia in remission

Schizophrenia is one of the most severe chronic relapsing mental diseases that significantly affect the level of social adaptation and quality of life of patients, often leads to their disability. Despite the success of modern psychopharmacology, achieving sustainable remission in schizophrenia remains a difficult task. The purpose of the study were the assessments of the safety and tolerability of intravenous administration of allogeneic AB0/ Rh-compatible mononuclear cord blood cells, as well as to study changes in cognitive performance in patients with schizophrenia in remission after treatment with umbilical cord blood cells. The study involved 30 patients with schizophrenia (men; average age 32,4 ± 9,7 years) in a state of hypochondria remission with a predominance of cognitive disorders against the background of prominent negative changes (F20.01-F20.04 according to ICD-1 0). Design is a prospective, placebo-controlled trial of efficiency and safety. The study consisted of 2 phases. In the pilot phase (3 months), the tolerability of a single cryopreserved concentrate of human cord blood injection containing mononuclear cells in a dose of 260± 20 million cells was estimated. The duration and severity of the effect was compared with placebo. In the clinical phase (48 months), patients received 4 injections of cord blood cell suspension in the same dose with intervals of 14 ± 3 days. The efficacy and safety of exposure were assessed using psychopathological, psychometric (scale of positive and negative symptoms of schizophrenia - PANSS) and psychological (The MATRIX Consensus Cognitive Battery) methods. The obtained results allow to conclude that the influence of human cord blood mononuclear cells on cognitive functions is realized due to the expressed metabolic (nootropic) and psychostimulating effects and restoration of normal neurotransmitters ratio. The effects are manifested in the form of activation of intellectual activity, acceleration of information processing, correction of memory functions, increase in the level of attention and vigilance, as well as a noticeable increase in "social intelligence” and, as a result, improvement in the quality of life. The effect of applying cord blood cells to enhance cognitive functions is characterized by resistance and duration of at least 4 years.

Human CD36hi monocytes induce Foxp3+ CD25+ T cells with regulatory functions from CD4 and CD8 subsets.

The fetal and neonatal immune systems are uniquely poised to generate tolerance to self, maternal and environmental antigens encountered in the womb and shortly after birth. However, the tolerogenic nature of fetal and neonatal immunity can be detrimental in the context of pathogens, leading to overwhelming bacterial infections or chronic viral infections. A variety of mechanisms contribute to fetal and neonatal tolerance, including a propensity to generate Foxp3+ regulatory T cells (Treg cells). However, the mechanism(s) of fetal Foxp3+ T-cell differentiation, the specific antigen-presenting cells required and factors that inhibit Treg generation after the neonatal period are poorly understood. Here, we demonstrate that a subset of CD14+ monocytes expressing the scavenger molecule, CD36, can generate CD4+ and CD8+ T cells that coexpress Foxp3 and T-bet from both umbilical cord blood. These Foxp3+ T-bet+ T cells potently suppress T-cell proliferation and ameliorate xenogeneic graft-versus-host disease. CD14+ CD36+ monocytes provide known Treg-inducing signals: membrane-bound transforming growth factor-beta and retinoic acid. Unexpectedly, adult peripheral blood monocytes are also capable of inducing Foxp3+ T cells from both cord blood and adult peripheral naïve T cells. The induction of Foxp3+ T cells in umbilical cord blood by monocytes was inhibited by the lymphoid fraction of adult peripheral blood cells. These studies highlight a novel immunoregulatory role of monocytes and suggest that antigen presentation by CD36hi monocytes may contribute to the peripheral development of Foxp3+ T-bet+ T cells with regulatory functions in both neonates and adults.

Abstract PO007: Role of PI3K pathway in reprogramming the tumor niche of angiosarcoma

Abstract Angiosarcoma (AS) is a highly aggressive, rare soft-tissue sarcoma that forms malignant blood vessels. Approximately half of patients have metastatic or unresectable disease with a median overall survival of less than 6 months, and tumor-related mortality is high. Inflamed tissue is frequently seen in angiosarcomas, and immune cells are key components that contribute to the landscape of the tumor microenvironment. Hemangiosarcoma (HSA) occurs commonly in companion dogs, and it shares clinical and morphological features with human AS. Recurrent mutations in TP53 and genes involved in PI3K pathway such as PIK3CA, and PIK3R1 were observed in both human AS and canine HSA, and the mutational landscape was associated with distinct molecular tumor subtypes; in particular, differing immunophenotypes. Recently, we have shown that human AS and canine HSA establish convergent transcriptional programs driven by angiogenic fusion genes and TP53 mutation. However, these vascular tumors are genomically complex, and molecular mechanisms that establish the tumor microenvironment are incompletely known. In this study, we analyzed RNA-seq transcriptomic data generated from human AS tissues (n=13) and canine HSA tissues (n=76) and HSA cell lines (n=11). The data showed that the tumors of both species enriched gene signatures associated with maintenance of hematopoietic stem cells. Then, we found that canine HSA cells supported expansion and differentiation of human CD34+ umbilical cord blood cells using long-term culture-initiating cell and colony-forming unit assays. We also developed xenograft models of canine HSA that facilitate myelopoiesis and macrophage infiltration in the tumor tissue. Our data showed that gene signatures of monocytes and naïve macrophages were detectable in canine HSA cells. In human ASs, macrophage-associated genes, in particular M2 macrophage signatures were significantly enriched, compared to normal tissues. Collectively, our data suggest that human AS and canine HSA have cell-autonomous capacity to govern hematopoietic progenitors and immune cells, potentially establishing the tumor immune niche. We engineered HSA cells to induce homozygous H1047R mutations in PIK3CA gene using CRISPR-Cas9 system. Our ongoing work is to determine if regulating PI3K pathway in the tumor cells contributes to the molecular programs that create the tumor niche of the vascular malignancy. Citation Format: Sophia Wenthe, Ashley J. Schulte, Mathew G. Angelos, Dan S. Kaufman, Jaime F. Modiano, Jong Hyuk Kim. Role of PI3K pathway in reprogramming the tumor niche of angiosarcoma [abstract]. In: Proceedings of the AACR Virtual Special Conference on the Evolving Tumor Microenvironment in Cancer Progression: Mechanisms and Emerging Therapeutic Opportunities; in association with the Tumor Microenvironment (TME) Working Group; 2021 Jan 11-12. Philadelphia (PA): AACR; Cancer Res 2021;81(5 Suppl):Abstract nr PO007.

Umbilical cord blood CD34+ cells administration improved neurobehavioral status and alleviated brain injury in a mouse model of cerebral palsy

PurposeCerebral palsy (CP) is the most common neuromuscular disease in children, and currently, there is no cure. Several studies have reported the benefits of umbilical cord blood (UCB) cell treatment for CP. However, these studies either examined the effects of UCB cell fraction with a short experimental period or used neonatal rat models for a long-term study which displayed an insufficient immunological reaction and clearance of human stem cells. Here, we developed a CP model by hypoxia-ischemic injury (HI) using immunodeficient mice and examined the effects of human UCB CD34+ hematopoietic stem cells (HSCs) on CP therapy over a period of 8 weeks.MethodsSixty postnatal day-9 (P9) mouse pups were randomly divided into 4 groups (n = 15/group) as follows: (1) sham operation (control group), (2) HI-induced CP model, (3) CP model with CD34+ HSC transplantation, and (4) CP model with CD34- cell transplantation. Eight weeks after insult, the sensorimotor performance was analyzed by rotarod treadmill, gait dynamic, and open field assays. The pathological changes in brain tissue of mice were determined by HE staining, Nissl staining, and MBP immunohistochemistry of the hippocampus in the mice.ResultsHI brain injury in mice pups resulted in significant behavioral deficits and loss of neurons. Both CD34+ HSCs and CD34- cells improved the neurobehavioral statuses and alleviated the pathological brain injury. In comparison with CD34- cells, the CD34+ HSC compartments were more effective.ConclusionThese findings indicate that CD34+ HSC transplantation was neuroprotective in neonatal mice and could be an effective therapy for CP.

Potential Use of Mesenchymal Multipotent Cells for Hemopoietic Stem Cell Transplantation: Pro and Contra.

The potential of mesenchymal multipotent (stem) cells (MSC) to modify immune reactions and mediate hematopoiesis boosted great interest for their use in allogeneic hemopoietic stem cell transplantation. Because of MSC production of a wide range of cytokines and growth factors, these cells are included in the therapy of graft-versus-host disease (GVHD). A number of clinical studies have demonstrated safety and efficacy of MSC-based therapy in acute GVHD. Japan and some other countries approved biomedical cell products on the base of allogeneic bone marrow (BM) MSCs as medical agents for acute GVHD treatment. Besides, MSCs may form BM stroma and improve hematopoiesis. Simultaneous transplantation of hematopoietic stem cells and MSCs effectively improved engraftment and prevented GVHD in transplantation of umbilical cord blood and human leukocyte antigens-incompatible BM stem cells. The review presents the analysis of clinical studies of MSCs in allogeneic hematopoietic stem cell transplantation and discusses different approaches for improvement of MSC-based GVHD treatment and prophylaxis.

Sequential Ensemble-Decision Aliquot Ranking Isolation and Fluorescence In Situ Hybridization Identification of Rare Cells from Blood by Using Concentrated Peripheral Blood Mononuclear Cells.

Isolation and analysis of circulating rare cells is a promising approach for early detection of cancer and other diseases and for prenatal diagnosis. Isolation of rare cells is usually difficult due to their heterogeneity as well as their low abundance in peripheral blood. We previously reported a two-stage ensemble-decision aliquot ranking platform (S-eDAR) for isolating circulating tumor cells from whole blood with high throughput, high recovery rate (>90%), and good purity (>70%), allowing detection of low surface antigen-expressing cancer cells linked to metastasis. However, due to the scarcity of these cells, large sample volumes and large quantities of antibodies were required to isolate sufficient cells for downstream analysis. Here, we drastically increased the number of nucleated cells analyzed by first concentrating peripheral blood mononuclear cells (PBMCs) from whole blood by density gradient centrifugation. The S-eDAR platform was capable of isolating rare cells from concentrated PBMCs (108/mL, equivalent to processing ∼20 mL of whole blood in the 1 mL sample volume used by our instrument) at a high recovery rate (>85%). We then applied the S-eDAR platform for isolating rare fetal nucleated red blood cells (fNRBCs) from concentrated PBMCs spiked with umbilical cord blood cells and confirmed fNRBC recovery by immunostaining and fluorescence in situ hybridization, demonstrating the potential of the S-eDAR system for isolating rare fetal cells from maternal PBMCs to improve noninvasive prenatal diagnosis.

215 - A Randomized Trial Comparing Umbilical Cord Blood and Cord Tissue Mesenchymal Stromal Cells in Young Children with Cerebral Palsy

215 - A Randomized Trial Comparing Umbilical Cord Blood and Cord Tissue Mesenchymal Stromal Cells in Young Children with Cerebral Palsy