Umbilical Cord Blood Cells Research Articles

Umbilical Cord Blood Cell Transplant Into Injured Spinal Cord With Lithium Carbonate or Placebo Followed by Locomotor Training

Umbilical Cord Blood Cell Transplant Into Injured Spinal Cord With Lithium Carbonate or Placebo Followed by Locomotor Training

PB1994 HEMATOPOIETIC STEM CELL EXPANSION IN THE PRESENCE OF NICOTINE AMIDE AND IN THE MICROFLUIDIC DEVISE AND THE DBM SCAFFOLD

Background:The low number of umbilical cord blood cells is an important barrier to successful bonemarrow transplantation. Therefore, the growth of these cells while maintaining theirfunctional characteristics is of great importanceAims:Using microfluidic technology as well asculturing cells on 3D scaffolds like DBM has greatly contributed to the simulation of themechanical and chemical micro‐environment of the bone marrow tissue. There are alsospecific compounds for proliferation without differentiation of cells, one of them isnicotinamideMethods:In this study, hematopoietic stem cells were cultured 7 days in two groups: 1.In DBM in the microfluidic device 2. In DBM in the static state (control), under the followingConditions:1.negative control with feeder2.cytokine+feeder3.nicotinamide+feeder4.nicotinamide+cytokine+feederAfter 7 days, cell count, real‐time PCR, their purity using flow cytometry, Colony forming unit assayand apoptosis was evaluated. The status of the cells on the scaffold was observed usingelectron microscopy.Results:Our results indicated that more cells were counted in static culturethan microfluidic culture (perfusion), while the ability of cell colonization in microfluidicculture was far higher. The purity of HSCs was higher in microfluidic culture, and fewer cellsentered the apoptotic phase.Summary/Conclusion:Nicotinamide by preventing epigenetic changes resulting from laboratory cell culture, andthe microfluidic device associated with DBM by simulating bone marrow niche is a verya suitable compound for the development of these cells.

Loss of RAF kinase inhibitor protein is involved in myelomonocytic differentiation and aggravates RAS-driven myeloid leukemogenesis.

RAS-signaling mutations induce the myelomonocytic differentiation and proliferation of hematopoietic stem and progenitor cells. Moreover, they are important players in the development of myeloid neoplasias. RAF kinase inhibitor protein (RKIP) is a negative regulator of RAS-signaling. As RKIP loss has recently been described in RAS-mutated myelomonocytic acute myeloid leukemia, we now aimed to analyze its role in myelomonocytic differentiation and RAS-driven leukemogenesis. Therefore, we initially analyzed RKIP expression during human and murine hematopoietic differentiation and observed that it is high in hematopoietic stem and progenitor cells and lymphoid cells but decreases in cells belonging to the myeloid lineage. By employing short hairpin RNA knockdown experiments in CD34+ umbilical cord blood cells and the undifferentiated acute myeloid leukemia cell line HL-60, we show that RKIP loss is indeed functionally involved in myelomonocytic lineage commitment and drives the myelomonocytic differentiation of hematopoietic stem and progenitor cells. These results could be confirmed in vivo, where Rkip deletion induced a myelomonocytic differentiation bias in mice by amplifying the effects of granulocyte macrophage-colony-stimulating factor. We further show that RKIP is of relevance for RAS-driven myelomonocytic leukemogenesis by demonstrating that Rkip deletion aggravates the development of a myeloproliferative disease in NrasG12D-mutated mice. Mechanistically, we demonstrate that RKIP loss increases the activity of the RAS-MAPK/ERK signaling module. Finally, we prove the clinical relevance of these findings by showing that RKIP loss is a frequent event in chronic myelomonocytic leukemia, and that it co-occurs with RAS-signaling mutations. Taken together, these data establish RKIP as novel player in RAS-driven myeloid leukemogenesis.

Cord blood research, banking, and transplantation: achievements, challenges, and perspectives.

The first hematopoietic transplant in which umbilical cord blood (UCB) was used as the source of hematopoietic cells was performed in October 1988. Since then, significant achievements have been reported in terms of our understanding of the biology of UCB-derived hematopoietic stem (HSCs) and progenitor (HPCs) cells. Over 40,000 UCB transplants (UCBTs) have been performed, in both children and adults, for the treatment of many different diseases, including hematologic, metabolic, immunologic, neoplastic, and neurologic disorders. In addition, cord blood banking has been developed to the point that around 800,000 units are being stored in public banks and more than 4 million units in private banks worldwide. During these 30 years, research in the UCB field has transformed the hematopoietic transplantation arena. Today, scientific and clinical teams are still working on different ways to improve and expand the use of UCB cells. A major effort has been focused on enhancing engraftment to potentially reduce risk of infection and cost. To that end, we have to understand in detail the molecular mechanisms controlling stem cell self-renewal that may lead to the development of ex vivo systems for HSCs expansion, characterize the mechanisms regulating the homing of HSCs and HPCs, and determine the relative place of UCBTs, as compared to other sources. These challenges will be met by encouraging innovative research on the basic biology of HSCs and HPCs, developing novel clinical trials, and improving UCB banking both in the public and private arenas.

Mechanisms of mixed umbilical cord blood-derived mesenchymal stem cells inhibiting proliferation and inducing apoptosis of C6 glioma cells in vitro

Objective To study the mechanisms of the mixed umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) inhibiting proliferation and inducing apoptosis of C6 glioma cells in vitro. Methods Cysteinyl aspartate-specific protease (Caspase)-8, B cell lymphoma/leukemia-2 associated X protein (bax), B cell lymphoma/leukemia-2 (bcl-2) and Caspase-3 protein expression in C6 cells was analyzed by immunohistochemistry and Western blotting, respectively. Caspase-8, bax, bcl-2 and Caspase-3 mRNA levels were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). Cell cycle was detected by flow cytometry. Apoptosis of C6 cells was examined by Annexin V-propidium iodide (PI) staining and terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) methods, respectively. Results The mixed UCB-MSCs had a tendency to increase the expression of Caspase-8, bax, Caspase-3 mRNA, and decrease the expression of bcl-2 mRNA. The expression of apoptosis protein and the number of apoptosis cells increased with the increase of the ratio of E/T (E: effect cells namely UCB-MSCs; T: target cells namely C6 cells), showing a proportional dependence relationship; E∶T=(5+ 5)∶1. The relative expression of Caspase-8, bax, bcl-2, and Caspase-3 mRNA in C6 cells was 0.78±0.13, 0.28±0.03, 0.70±0.08, and 0.76±0.12, respectively. When E/T=0∶1, 5∶1, (2.5+ 2.5)∶1, 10∶1, (5+ 5)∶1, the number of apoptosis C6 cells was (2.9±0.6), (3.1±0.5), (14.5±1.8), (14.3±2.2), and (22.8±4.5) cells, respectively. Conclusion After phytohemagglutinin (PHA) stimulation, the mixed UCB-MSCs could inhibit proliferation of C6 cells in vitro, which was related to induction of apoptosis of glioma cells. Key words: Glioma; Umbilical cord blood; Mesenchymal stem cells; Apoptosis; Proliferation

Single-cell transcriptomic landscape of nucleated cells in umbilical cord blood.

BackgroundFor both pediatric and adult patients, umbilical cord blood (UCB) transplant is a therapeutic option for a variety of hematologic diseases, such as blood cancers, myeloproliferative disorders, genetic diseases, and metabolic disorders. However, the level of cellular heterogeneity and diversity of nucleated cells in UCB has not yet been assessed in an unbiased and systemic fashion. In the present study, nucleated cells from UCB were subjected to single-cell RNA sequencing to simultaneously profile the gene expression signatures of thousands of cells, generating a rich resource for further functional studies. Here, we report the transcriptomes of 17,637 UCB cells, covering 12 major cell types, many of which can be further divided into distinct subpopulations.ResultsPseudotemporal ordering of nucleated red blood cells identifies wave-like activation and suppression of transcription regulators, leading to a polarized cellular state, which may reflect nucleated red blood cell maturation. Progenitor cells in UCB also comprise 2 subpopulations with activation of divergent transcription programs, leading to specific cell fate commitment. Detailed profiling of cytotoxic cell populations unveiled granzymes B and K signatures in natural killer and natural killer T-cell types in UCB.ConclusionsTaken together, our data form a comprehensive single-cell transcriptomic landscape that reveals previously unrecognized cell types, pathways, and mechanisms of gene expression regulation. These data may contribute to the efficacy and outcome of UCB transplant, broadening the scope of research and clinical innovations.

Stimulation of rats fetuses growth by transplantation of rats bone marrow cells or human umbilical cord blood cells during gastrulation

Background & Aim In humans and animals with interstitial transplantation and hemochorial type of the placenta decidual cells (DC) differentiates in the uterus, and form the maternal part of the placenta. In the absence of DC development stops at the stage of merosome. Previously, we have shown that non-pregnant rats bone marrow cells transplantation (BMT) stimulates the development of DC in pseudo-pregnant rats. It is of interest to study the effect of pregnant rats bone-marrow cells on fetal development during transplantation in the early stages of pregnancy, as well as whether human umbilical cord blood cells have a similar effect. Methods, Results & Conclusion Bone marrow mononuclear cells (BMMC) of 4-13 days of pregnancy rats were administered intravenously to does of the same gestation period. The weight and morphology of the fetuses were evaluated on the 18th day of pregnancy. It was found that BMMC transplantation on 7-9 days of pregnancy significantly increased the weight of fetuses to 835 ± 15 mg compared to the weight of non-pregnant does (745 ± 11 mg) and the weight of does fetuses, which were administered BMMC of diestrus does (772 ± 9 m). Administration of BMMC on 11-12 days of pregnancy reduced fetal weight to 587 ± 5 mg. Also the umbilical cord blood mononuclear cells (UCBMC) was used in the study, which were administered subcutaneously in doses of 26 or 71 × 106 cells per 250 g of animal weight. Fetal weight on the 18th day of pregnancy was the maximum (856 ± 8 mg) with the administration of UCBMC on the 9th day of pregnancy. With the administration of UCBMC on day 11 of pregnancy fetal weight was reduced to 803 ± 19 mg. The dose reduction of the UCBMC to 26 × 106 resulted in the weight of the fetus on the 9th day of pregnancy decrease to 813 ± 17 mg. Thus, both rats BMMC and a human UCBMC stimulate 18th day of pregnancy fetus weight after administration to the female rats on the 7th, 8th and 9th days of pregnancy in the period of gastrulation. No morphological abnormalities were found in the fetuses. There is also no increase in post-implantation death, which indicates that the increase in fetal weight is not the result of compensatory growth. Similar results of transplantation of different types of cells indicate a general mechanism of fetal growth stimulation. It is possible that the result develops due to stimulation of decidual cell function but particular mechanisms of it are still need future researches.

SUN-036 SLC16A11 Gene Expression In Mothers With And Without Gestational Diabetes And Their Offspring Umbilical Cord Blood.

BACKGROUND: Studies have identified the intrauterine environment as a regulating factor of the methylation profiles of DNA of newborns. In Mexicans a SLC16A11 risk haplotypes is associated with type 2 diabetes mellitus. It is in our interest to evaluate SLC16A11 expression from the earliest stages of human development in the cells of umbilical cord blood of newborn offspring of mothers with and without gestational diabetes mellitus (GDM), and thus identify indirectly epigenetic modifications due to maternal hyperglycemic status. To date is scarce this kind of studies. OBJECTIVE: To determine the SLC16A11 gene expression, in the cells of umbilical cord blood of newborn offspring of mothers with or without GDM. MATERIAL AND METHODS: Pregnant patients submitting to cesarean, with or without GDM, were invited to participate in at Gyneco-obstetric hospital of IMSS, Mexico City. RNA of peripheral blood cells (PBC) from mothers with or without GDM, and from the umbilical cord blood cells of their newborn offspring, were extracted. Briefly, to evaluate differences in SLC16A11 gene expression, cDNA was generated by using retro-transcriptase reaction and then subjected to end point PCR with specific oligonucleotides. Variables were analyzed with t-Student test using the SPSS program. Values of p < 0.05 were considered statistically significant. RESULTS: Mean age of mothers was 30 ± 4.25 years. GDM mothers were heavier (32.90 ± 3.7 vs 30.12 ± 3.4 kg/m2, p=0.032), but there were no differences in glycemic control (HbA1c)(5.4 ± 0.6 vs 5.2 ± 0.6, p=0.333). Somatometric measurements of offspring showed no significant differences (weight: 3013.33 ± 675.54 vs 3135.71g ± 740.59, p=0.626 and length: 48.50 ± 3.03 vs 49.17cm ± 1.56, p=0.368). Analysis SLC16A11 gene expression did not showed differences in mothers, (p=0.059), and in offspring (p=0.315). Because there is a SLC16A11 gene expression, this could suggest that the SLC16A11 gene promoter could not be methylated. CONCLUSIONS: No modifications in SLC16A11 gene expression were present in GDM, which suggests that GDM does not modify the molecular mechanism in SLC16A11 RNA expression.

Забезпечення публічного інтересу у сфері донорства крові

Забезпечення публічного інтересу у сфері донорства крові

Combining Human Umbilical Cord Blood Cells With Erythropoietin Enhances Angiogenesis/Neurogenesis and Behavioral Recovery After Stroke.

Disruption of blood flow in the brain induces stroke, the leading cause of death and disability worldwide. However, so far the therapeutic options are limited. Thus, the therapeutic efficacy of cell-based approaches has been investigated to develop a potential strategy to overcome stroke-induced disability. Human umbilical cord blood cells (hUCBCs) and erythropoietin (EPO) both have angiogenic and neurogenic properties in the injured brain, and their combined administration may exert synergistic effects during neurological recovery following stroke. We investigated the therapeutic potential of hUCBC and EPO combination treatment by comparing its efficacy to those of hUCBC and EPO alone. Adult male Sprague-Dawley rats underwent transient middle cerebral artery occlusion (MCAO). Experimental groups were as follows: saline (injected once with saline 7 d after MCAO); hUCBC (1.2 × 107 total nucleated cells, injected once via the tail vein 7 d after MCAO); EPO (500 IU/kg, injected intraperitoneally for five consecutive days from 7 d after MCAO); and combination of hUCBC and EPO (hUCBC+EPO). Behavioral measures (Modified Neurological Severity Score [mNSS] and cylinder test) were recorded to assess neurological outcomes. Four weeks after MCAO, brains were harvested to analyze the status of neurogenesis and angiogenesis. In vitro assays were also conducted using neural stem and endothelial cells in the oxygen-glucose deprivation condition. Performance on the mNSS and cylinder test showed the most improvement in the hUCBC+EPO group, while hUCBC- and EPO-alone treatments showed superior outcomes relative to the saline group. Neurogenesis and angiogenesis in the cortical region was the most enhanced in the hUCBC+EPO group, while the findings in the hUCBC and EPO treatment alone groups were better than those in the saline group. Astrogliosis in the brain tissue was reduced by hUCBC and EPO treatment. The reduction was largest in the hUCBC+EPO group. These results were consistent with in vitro assessments that showed the strongest neurogenic and angiogenic effect with hUCBC+EPO treatment. This study demonstrates that combination therapy is more effective than single therapy with either hUCBC or EPO for neurological recovery from subacute stroke. The common pathway underlying hUCBC and EPO treatment requires further study.

Effects of Maternal Hypertensive Disorders on the Expression of Arginine Vasopressin Receptors in Offspring

Children born to women with hypertensive disorders inpregnancy have an elevated risk of developing hypertensive disorders later inlife. The mechanisms for this fetal programming are unclear. We have previouslyidentified a critical role for arginine vasopressin (AVP) in the development ofpreeclampsia. Additionally, we demonstrated differences in the expression ofAVP receptors on circulating cells in women who developed preeclampsia comparedto women who did not experience preeclampsia. Because AVP is a hormone thatfunctions to regulate blood pressure and volume homeostasis, we hypothesizedthat differences in expression of vasopressin receptors and related genes arealso altered in children born to women with preeclampsia.We measured the expression of receptors AVPR1a, AVPR1b, AVPR2, OXTR, and CUL5 as well as LNPEP, the major aminopeptidase that cleavesAVP. Expression in umbilical cord blood cells from women with 1) no hypertensivedisorders in pregnancy (control subjects) (N=20) 2) pregnant women with chronichypertension (N=20) and 3) women with preeclampsia (N=10) in the indexpregnancy were compared. Coded umbilical cord blood cells and associated clinicaldata were obtained from the University of Iowa Maternal Fetal Tissue Bank (IRB#200910784). RNA was purified from the cells and used for real‐time quantitativePCR to assess variation in receptor expression in children born to women withhypertensive disorders of pregnancy relative to control subjects. In childrenborn to women with chronic hypertension, AVPR2 (−8.2 Fold Change), LNPEP (−13Fold Change), CUL5 (−14 Fold Change) AVPR1b (−3.1 Fold Change) and OXTR (−4.3Fold Change) were all significantly under‐expressed with controls. Samples fromchildren born to women with preeclampsia had significantly lower expression of AVPR1a(−4.7 Fold Change), AVPR1b (−2.5 Fold Change), AVPR2 (−2.5 Fold Change), andOXTR (−2.8 Fold Change) when compared to non‐preeclamptic samples. Thesedifferences in gene expression may be related to the future risk ofhypertension later in life for children born to women with hypertensivediseases in pregnancy. In future work, we will interrogate different cellpopulations. In addition, we will determine whether maternal peripheral bloodvasopressin receptor expression correlates with child cord blood receptorexpression.This work was supported by the Iowa Center for Research byUndergraduates (LD), NIH CTSA grant UL1TR002537, American Heart AssociationPostdoctoral Fellowship (SS), and the American Heart Association StrategicallyFocused Research Network Grant (MS, DS, SS).Support or Funding InformationThis work was supported by the Iowa Center for Research by Undergraduates (LD), NIH CTSA grant UL1TR002537, American Heart Association Postdoctoral Fellowship (SS), and the American Heart Association Strategically Focused Research Network Grant (MS, DS, DB, KW, SS).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Human umbilical cord blood cells restore vascular integrity in injured rat brain and modulate inflammation in vitro.

Aim: Traumatic brain injury is a complex condition consisting of a mechanical injury with neurovascular disruption and inflammation with limited clinical interventions available. A growing number of studies report systemic delivery of human umbilical cord blood (HUCB) as a therapy for neural injuries. Materials & methods: HUCB cells from five donors were tested to improve blood-brain barrier integrity in a traumatic brain injury rat model at a dose of 2.5×107 cells/kg at 24 or 72h postinjury and for immunomodulatory activity in vitro. Results & Conclusion: We observed that cells delivered 72h postinjury significantly restored blood-brain barrier integrity. HUCB cells reduced the amount of TNF-α and IFN-γ released by activated primary rat splenocytes, which correlated with the expression of COX2 and IDO1.

Human Umbilical Cord Therapy Improves Long-Term Behavioral Outcomes Following Neonatal Hypoxic Ischemic Brain Injury.

Background: Hypoxic ischemic (HI) insult in term babies at labor or birth can cause long-term neurodevelopmental disorders, including cerebral palsy (CP). The current standard treatment for term infants with hypoxic ischemic encephalopathy (HIE) is hypothermia. Because hypothermia is only partially effective, novel therapies are required to improve outcomes further. Human umbilical cord blood cells (UCB) are a rich source of stem and progenitor cells making them a potential treatment for neonatal HI brain injury. Recent clinical trials have shown that UCB therapy is a safe and efficacious treatment for confirmed cerebral palsy. In this study, we assessed whether early administration of UCB to the neonate could improve long-term behavioral outcomes and promote brain repair following neonatal HI brain injury.Methods: HI brain injury was induced in postnatal day (PND) 7 rat pups via permanent ligation of the left carotid artery, followed by a 90 min hypoxic challenge. UCB was administered intraperitoneally on PND 8. Behavioral tests, including negative geotaxis, forelimb preference and open field test, were performed on PND 14, 30, and 50, following brains were collected for assessment of neuropathology.Results: Neonatal HI resulted in decreased brain weight, cerebral tissue loss and apoptosis in the somatosensory cortex, as well as compromised behavioral outcomes. UCB administration following HI improved short and long-term behavioral outcomes but did not reduce long-term histological evidence of brain injury compared to HI alone. In addition, UCB following HI increased microglia activation in the somatosensory cortex compared to HI alone.Conclusion: Administration of a single dose of UCB cells 24 h after HI injury improves behavior, however, a single dose of cells does not modulate pathological evidence of long-term brain injury.

DNA·RNA triple helix formation can function as a cis-acting regulatory mechanism at the human β-globin locus

We have identified regulatory mechanisms in which an RNA transcript forms a DNA duplex·RNA triple helix with a gene or one of its regulatory elements, suggesting potential auto-regulatory mechanisms in vivo. We describe an interaction at the human β-globin locus, in which an RNA segment embedded in the second intron of the β-globin gene forms a DNA·RNA triplex with the HS2 sequence within the β-globin locus control region, a major regulator of globin expression. We show in human K562 cells that the triplex is stable in vivo. Its formation causes displacement from HS2 of major transcription factors and RNA Polymerase II, and consequently in loss of factors and polymerase that bind to the human ε- and γ-globin promoters, which are activated by HS2 in K562 cells. This results in reduced expression of these genes. These effects are observed when a small length of triplex-forming RNA is introduced into cells, or when a full-length intron-containing human β-globin transcript is expressed. Related results are obtained in human umbilical cord blood-derived erythroid progenitor-2 cells, in which β-globin expression is similarly affected by triplex formation. These results suggest a model in which RNAs conforming to the strict sequence rules for DNA·RNA triplex formation may participate in feedback regulation of genes in cis.

Umbilical cord blood versus mesenchymal stem cells for inflammation-induced preterm brain injury in fetal sheep.

Chorioamnionitis and fetal inflammation are principal causes of neuropathology detected after birth, particularly in very preterm infants. Preclinical studies show that umbilical cord blood (UCB) cells are neuroprotective, but it is uncertain if allogeneic UCB cells are a feasible early intervention for preterm infants. In contrast, mesenchymal stem cells (MSCs) are more readily accessible and show strong anti-inflammatory benefits. We aimed to compare the neuroprotective benefits of UCB versus MSCs in a large animal model of inflammation-induced preterm brain injury. We hypothesized that MSCs would afford greater neuroprotection. Chronically instrumented fetal sheep at 0.65 gestation received intravenous lipopolysaccharide (150 ng; 055:B5, n = 8) over 3 consecutive days; or saline for controls (n = 8). Cell-treated animals received 108 UCB mononuclear cells (n = 7) or 107 umbilical cord MSCs (n = 8), intravenously, 6 h after the final lipopolysaccharide dose. Seven days later, cerebrospinal fluid and brain tissue was collected for analysis. Lipopolysaccharide induced neuroinflammation and apoptosis, and reduced the number of mature oligodendrocytes. MSCs reduced astrogliosis, but UCB did not have the same effect. UCB significantly decreased cerebral apoptosis and protected mature myelinating oligodendrocytes, but MSCs did not. UCB appears to better protect white matter development in the preterm brain in response to inflammation-induced brain injury in fetal sheep.

GM-CSF Promotes the Expansion and Differentiation of Cord Blood Myeloid-Derived Suppressor Cells, Which Attenuate Xenogeneic Graft-vs.-Host Disease.

Myeloid-derived suppressor cells (MDSCs) are increased in tumor patients. Studies have shown generation of MDSCs from human peripheral blood mononuclear cells (PBMCs) by various cytokine combinations. However, large scale expansion of human MDSCs has not been demonstrated or applied in clinic settings. We investigated which cytokine combinations among GM-CSF/SCF, G-CSF/SCF, or M-CSF/SCF efficiently expand and differentiate human MDSCs following culture CD34+ cells of umbilical cord blood (CB). GM-CSF/SCF showed the greatest expansion of MDSCs. Up to 108 MDSCs (HLA-DRlowCD11b+CD33+) could be produced from 1 unit of CB following 6 weeks of continuous culture. MDSCs produced from culture of CD34+ cells with GM-CSF/SCF for 6 weeks had the greatest suppressive function of T cell proliferation and had the highest expression of immunosuppressive molecules including iNOS, arginase 1 and IDO compared to those differentiated with G-CSF/SCF or M-CSF/SCF. MDSCs secreted IL-10, TGB-β, and VEGF. The infusion of expanded MDSCs significantly prolonged the survival and decreased the GVHD score in a NSG xenogeneic model of GVHD. Injected MDSCs increased IL-10 and TGF-β but decreased the level of TNF-α and IL-6 in the serum of treated mice. Notably, FoxP3 expressing regulatory T (Treg) cells were increased while IFN-γ (Th1) and IL-17 (Th17) producing T cells were decreased in the spleen of MDSC treated mice compared to untreated GVHD mice. Our results demonstrate that human MDSCs are generated from CB CD34+ cells using GM-CSF/SCF. These MDSCs exhibited potent immunosuppressive function, suggesting that they are useable as a treatment for inflammatory diseases such as GVHD.

Orphan designation: Allogeneic ex-vivo-expanded umbilical cord blood-derived haematopoietic CD34+ progenitor cells and allogeneic non-expanded umbilical cord blood-derived haematopoietic mature myeloid and lymphoid cells, Treatment in haematopoietic stem cell transplantation

Orphan designation: Allogeneic ex-vivo-expanded umbilical cord blood-derived haematopoietic CD34+ progenitor cells and allogeneic non-expanded umbilical cord blood-derived haematopoietic mature myeloid and lymphoid cells, Treatment in haematopoietic stem cell transplantation

Biomechanical Stability and Osteogenesis in a Tibial Bone Defect Treated by Autologous Ovine Cord Blood Cells-A Pilot Study.

The aim of this study was to elucidate the impact of autologous umbilical cord blood cells (USSC) on bone regeneration and biomechanical stability in an ovine tibial bone defect. Ovine USSC were harvested and characterized. After 12 months, full-size 2.0 cm mid-diaphyseal bone defects were created and stabilized by an external fixateur containing a rigidity measuring device. Defects were filled with (i) autologous USSC on hydroxyapatite (HA) scaffold (test group), (ii) HA scaffold without cells (HA group), or (iii) left empty (control group). Biomechanical measures, standardized X-rays, and systemic response controls were performed regularly. After six months, bone regeneration was evaluated histomorphometrically and labeled USSC were tracked. In all groups, the torsion distance decreased over time, and radiographies showed comparable bone regeneration. The area of newly formed bone was 82.5 ± 5.5% in the control compared to 59.2 ± 13.0% in the test and 48.6 ± 2.9% in the HA group. Labeled cells could be detected in lymph nodes, liver and pancreas without any signs of tumor formation. Although biomechanical stability was reached earliest in the test group with autologous USSC on HA scaffold, the density of newly formed bone was superior in the control group without any bovine HA.

The use of the concentrate of nucleus containing cells of umbilical cord blood for treatment of asthenic disorders (schizoasthenia) in patients with schizophrenia with negative symptoms

To assess the safety and efficacy of umbilical cord blood (UCB) cells in the treatment of schizoasthenia and comorbid negative disorders. The study was conducted in men (n=15) with remission of schizophrenia with a predominance of asthenic disorders (ICD-10 items F20.6 and F21.5). Patients received intravenous infusions of UCB cells at the average dose of 250 mln viable cells per infusion (4 injections at 2 week intervals). The efficacy and safety were assessed using the Positive and Negative Symptoms of Schizophrenia (PANSS), the General Clinical Impression Scale (CGI), Asthenia Scale MFI-20 3, the MATRIX Consensus Cognitive Battery (MCCB), the Clinical evaluation of adverse events (side effects). The infusions of UCB cells have a positive effect on asthenic disorders, increase the level of general activity and minimize cognitive impairments. The authors assume that UCB cells potentiate the activity of typical and atypical antipsychotics.

CD34+ Derived Microparticles Regulate Apoptosis in Normal and Acute Myeloid Leukemia Cells

Introduction Microparticles (MPs) are small vesicles 100nm-1μm derived from the apoptotic or stimulated cells. The mechanism of their production is distinctive from exosomes or apoptotic bodies. MPs have been detected in the blood in many pathological conditions associated mainly with endothelial injury, thrombosis and inflammation. Our previous study showed that MPs originated also from CD34+ cells of umbilical cord blood, which is an alternative source for hemopoeitic stem cell transplantation. MPs considered as markers of cell activation, as well as apoptosis. Apoptosis is a complex interaction network regulated either through death receptor like FAS or through the internal pathway of Bcl2, Bax. The two pathways activate the executor of cell death program, caspases. The aim of this study is to elucidate the role of MPs in apoptosis of hemopoietic cells.Methods Umbilical cord blood units (UCB) were collected after informed consent. The units that were used in this study, were rejected as not appropriate for transplantation due to low volume. The HL60 promyelocytic leukemia cell line was cultured in RPMI (Life Technologies) supplemented with 10% fetal calf serum (FCS) and 1% penicillin-strepromycin. CD34+ MPs were isolated from the plasma of UCBs after centrifugation and magnetic bead MACS purification (Miltenyi Biotec). The number of CD34+ MPs was estimated by flow cytometry using CD34-PE and Annexin V-FITC Abs. Mononuclear cells (MNC) were collected after density gradient centrifugation on lymphoprep (Fresenius) and were cultured for 3 and 6 days in the presence of CD34+ MPs. Viability assays were performed using 7-AAD in flow cytometry. In another set of experiments different numbers of CD34+ MPs were used in MNC cultures. RNA was extracted from MNC using Qiagen RNA extraction kit and reverse transcribed into cDNA using Superscript II reverse transcriptase (Invitrogen) with random primers (Promega). RT-PCR was performed using Platinum Pfx polymerase (Invitrogen). The primers for FAS, BCL2, BAX, caspase 3, survivin and GAPDH genes were used. The PCR products were analysed in an agarose gel electrophoresis.Results Cell viability increased in UCB derived MNC (UCB-MNC) incubated with CD34+ MPs (800 /ml) after 3 day vs. 6 days of culture. The UCB-MNC viability was higher using 800/ml CD34+ MPs vs. 400/ml. In contrast, CD34+ MPs (800 /ml vs. 400/ml) did not affect the viability in one day MNC culture. Purified CD34+ MPs were applied to UCB-MNC cultures and by RT-PCR was shown increased expression of BCL2 gene as well as FAS and caspase-3 genes. The promyelocytc cell line HL60 has been used in order to analyze the effect of CD34+ MPs in leukemic cells. The expression of Bcl2 was decreased in HL60 cells co-incubated with CD34+ MPs. This result shows an opposite effect of CD34+ MPs in the apoptotic gene Bcl2 for the HL60 cells indicating that there are different mechanisms of MP function in various cell types.Conclusions In this study we have identified and monitored the time- and dose-dependent effect of CD34-derived microparticles in the viability of UCB mononuclear cells. Additionally, CD34+ MPs function is accosiated with the high expression of the pro- and anti-apoptotic Bcl2 and apoptotic FAS. In contrast CD34+ MPs decreases the expression of Bcl2 in the promyelocytic leukemia cell line HL60. Therefore the stem cell derived microparticles might serve as a potential regulator of apoptosis in normal and malignant hematopoietic cells. DisclosuresNo relevant conflicts of interest to declare.