Umbilical Cord Mesenchymal Cells Research Articles

Plasma Redox Balance in Advanced-Maternal-Age Pregnant Women and Effects of Plasma on Umbilical Cord Mesenchymal Stem Cells.

Pregnancy at advanced maternal age (AMA) is a condition of potential risk for the development of maternal-fetal complications with possible repercussions even in the long term. Here, we analyzed the changes in plasma redox balance and the effects of plasma on human umbilical cord mesenchymal cells (hUMSCs) in AMA pregnant women (patients) at various timings of pregnancy. One hundred patients and twenty pregnant women younger than 40 years (controls) were recruited and evaluated at various timings during pregnancy until after delivery. Plasma samples were used to measure the thiobarbituric acid reactive substances (TBARS), glutathione and nitric oxide (NO). In addition, plasma was used to stimulate the hUMSCs, which were tested for cell viability, reactive oxygen species (ROS) and NO release. The obtained results showed that, throughout pregnancy until after delivery in patients, the levels of plasma glutathione and NO were lower than those of controls, while those of TBARS were higher. Moreover, plasma of patients reduced cell viability and NO release, and increased ROS release in hUMSCs. Our results highlighted alterations in the redox balance and the presence of potentially harmful circulating factors in plasma of patients. They could have clinical relevance for the prevention of complications related to AMA pregnancy.

Anticancer effect of hUC-MSC-derived exosome-mediated delivery of PMO-miR-146b-5p in colorectal cancer

Antisense oligonucleotide (ASO) is a novel therapeutic platform for targeted cancer therapy. Previously, we have demonstrated that miR-146b-5p plays an important role in colorectal cancer progression. However, a safe and effective strategy for delivery of an ASO to its targeted RNA remains as a major hurdle in translational advances. Human umbilical cord mesenchymal cell (hUC-MSC)–derived exosomes were used as vehicles to deliver an anti-miR-146b-5p ASO (PMO-146b). PMO-146b was assembled onto the surface of exosomes (e) through covalent conjugation to an anchor peptide CP05 (P) that recognized an exosomal surface marker, CD63, forming a complex named ePPMO-146b. After ePPMO-146b treatment, cell proliferation, uptake ability, and migration assays were performed, and epithelial-mesenchymal transition progression was evaluated in vitro. A mouse xenograft model was used to determine the antitumor effect and distribution of ePPMO-146b in vivo. ePPMO-146b was taken up by SW620 cells and effectively inhibited cell proliferation and migration. The conjugate also exerted antitumor efficacy in a xenograft mouse model of colon cancer by systematic administration, where PPMO-146b was enriched in tumor tissue. Our study highlights the potential of hUC-MSC-derived exosomes anchored with PPMO-146b as a novel safe and effective approach for PMO backboned ASO delivery.Graphical Schematic illustration of the preparation of an exosomal anchor peptide (CP05)-PMO that conjugately binds to exosomes from hUC-MSCs (ePPMO-146b) and the antitumor effect of ePPMO-146b in CRC, which occurs through the inhibition of Smad signaling and epithelial–mesenchymal transition.

Multiomics reveal human umbilical cord mesenchymal stem cells improving acute lung injury via the lung-gut axis.

Acute lung injury (ALI) and its final severe stage, acute respiratory distress syndrome, are associated with high morbidity and mortality rates in patients due to the lack of effective specific treatments. Gut microbiota homeostasis, including that in ALI, is important for human health. Evidence suggests that the gut microbiota improves lung injury through the lung-gut axis. Human umbilical cord mesenchymal cells (HUC-MSCs) have attractive prospects for ALI treatment. This study hypothesized that HUC-MSCs improve ALI via the lung-gut microflora. To explore the effects of HUC-MSCs on lipopolysaccharide (LPS)-induced ALI in mice and the involvement of the lung-gut axis in this process. C57BL/6 mice were randomly divided into four groups (18 rats per group): Sham, sham + HUC-MSCs, LPS, and LPS + HUC-MSCs. ALI was induced in mice by intraperitoneal injections of LPS (10 mg/kg). After 6 h, mice were intervened with 0.5 mL phosphate buffered saline (PBS) containing 1 × 106 HUC-MSCs by intraperitoneal injections. For the negative control, 100 mL 0.9% NaCl and 0.5 mL PBS were used. Bronchoalveolar lavage fluid (BALF) was obtained from anesthetized mice, and their blood, lungs, ileum, and feces were obtained by an aseptic technique following CO2 euthanasia. Wright's staining, enzyme-linked immunosorbent assay, hematoxylin-eosin staining, Evans blue dye leakage assay, immunohistochemistry, fluorescence in situ hybridization, western blot, 16S rDNA sequencing, and non-targeted metabolomics were used to observe the effect of HUC-MSCs on ALI mice, and the involvement of the lung-gut axis in this process was explored. One-way analysis of variance with post-hoc Tukey's test, independent-sample Student's t-test, Wilcoxon rank-sum test, and Pearson correlation analysis were used for statistical analyses. HUC-MSCs were observed to improve pulmonary edema and lung and ileal injury, and decrease mononuclear cell and neutrophil counts, protein concentrations in BALF and inflammatory cytokine levels in the serum, lung, and ileum of ALI mice. Especially, HUC-MSCs decreased Evans blue concentration and Toll-like receptor 4, myeloid differentiation factor 88, p-nuclear factor kappa-B (NF-κB)/NF-κB, and p-inhibitor α of NF-κB (p-IκBα)/IκBα expression levels in the lung, and raised the pulmonary vascular endothelial-cadherin, zonula occludens-1 (ZO-1), and occludin levels and ileal ZO-1, claudin-1, and occludin expression levels. HUC-MSCs improved gut and BALF microbial homeostases. The number of pathogenic bacteria decreased in the BALF of ALI mice treated with HUC-MSCs. Concurrently, the abundances of Oscillospira and Coprococcus in the feces of HUS-MSC-treated ALI mice were significantly increased. In addition, Lactobacillus, Bacteroides, and unidentified_Rikenellaceae genera appeared in both feces and BALF. Moreover, this study performed metabolomic analysis on the lung tissue and identified five upregulated metabolites and 11 downregulated metabolites in the LPS + MSC group compared to the LPS group, which were related to the purine metabolism and the taste transduction signaling pathways. Therefore, an intrinsic link between lung metabolite levels and BALF flora homeostasis was established. This study suggests that HUM-MSCs attenuate ALI by redefining the gut and lung microbiota.

Recent Advances in InVitro Maturation (IVM): A Systematic Literature Review

In Vitro Maturation (IVM) is one of the Assisted Reproductive Technologies (ART) that involves the retrieval of immature oocytes from antral follicles in the ovaries, either in a non-stimulated or minimally stimulated state. IVM has drawn the attention of fertility specialists due to its reliability, cost-effectiveness, low risk of Ovarian Hyperstimulation Syndrome (OHSS), and acceptable clinical pregnancy rates Objective: Understanding the Process and Factors Influencing In Vitro Maturation of Human Oocytes (IVM) Method: The research methodology employed in this study is the Systematic Literature Review (SLR) method. Data was collected by documenting all articles based on predetermined inclusion and exclusion criteria. A total of 10 national and international journal articles were utilized for this study, obtained from databases such as Google Scholar, PubMed, and Science Direct, using specific keywords including "In Vitro Maturation" and "human oocyte maturation". The literature search and study selection process followed the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. Results: There were no instances of moderate-severe OHSS (ovarian hyperstimulation syndrome) in the IVM group. The fertilization rate in the group receiving growth hormone (GH) was 12.8% higher compared to the control group. Additionally, the blastocyst rate in the GH group was 9.5% higher than that of the control group. Treatment with a concentration of 50 µmol/L ALA (alpha-lipoic acid) significantly accelerated oocyte maturation. It resulted in a significantly higher mitochondrial DNA (mtDNA) copy number in mature oocytes compared to the control group (0 µmol/L ALA). The highest maturation rates were observed in human umbilical cord mesenchymal cells (hUCM) in the in vitro maturation (fIVM) of germinal vesicle (GV) oocytes, while the lowest maturation rates were observed using alpha-Minimum Essential Medium (α-MEM) in the in vitro maturation (vIVM) of GV oocytes. When AMH (anti-Müllerian hormone) was added to the IVM medium, a maturation rate of 100% was achieved in mature oocytes. The inclusion of 50 mmol/L CoQ10 in IVM media was found to reduce the level of oocyte aneuploidy by nearly 50%. The formation rate of metaphase II (MII) stage oocytes was higher in culture medium containing 1.0 mm resveratrol compared to the control group. Subsequently, a total of eight cleavage-stage embryos were frozen after intracytoplasmic sperm injection (ICSI). One year later, when the patient returned for the transfer of cryopreserved embryos, two embryos were thawed and transferred into the uterus. The patient successfully became pregnant, and the twin pregnancy progressed without complications, delivering two healthy full-term baby boys. Conclusion: The occurrence of OHSS was not observed in any of the IVM intervention groups. Supplementation with growth hormone (GH), alpha-lipoic acid (ALA), umbilical cord-derived mesenchymal stem cells (CM-MSC), anti-Müllerian hormone (AMH), and resveratrol in the culture media each demonstrated a significant improvement in human oocyte maturation. Furthermore, resveratrol and coenzyme Q10 were effective in addressing abnormal spindle morphology, irregular chromosome arrangement, and high postmeiotic aneuploidy. It is worth noting that IVM is not exclusively limited to patients with polycystic ovary syndrome (PCOS), as it can yield satisfactory outcomes in patients with autoimmune premature ovarian insufficiency (POI).

Umbilical Cord-Mesenchymal Stem Cell Therapy for COVID-19 Related Acute Respiratory Distress Syndrome: A Mini-Review

Introduction: Acute respiratory distress syndrome (ARDS) is the leading cause of death among coronavirus disease 2019 (COVID-19) patients, mainly due to the cytokine storm and the rearrangements in coagulation and immune responses. Accordingly, the immunomodulatory and regenerative properties of umbilical mesenchymal stem cells (UC-MSCs) have been studied for the treatment of COVID-19. Methods: This mini-review evaluated adults with moderate-severe COVID-19 infection and compared the results of placebo plus standard of care (SOC) therapy with those obtained from the administration of umbilical cord mesenchymal cells (UC-MSCs). We searched the following databases: Cochrane, Central Register of Controlled Trials, and PubMed; subsequently, 8 clinical trials were included in this mini-review. Some statistically significant difference was found in the levels of clinical and inflammatory markers between the intervention and the control groups. Conclusion: Early phase trials have shown the promising efficacy and safety of UC-MSC therapy for COVID-19-associated moderate-to-severe ARDS. Large multicenter phase III randomized controlled clinical trials will further confirm these findings.

Exosome Derived from Human Umbilical Cord Mesenchymal Cell Exerts Immunomodulatory Effects on B Cells from SLE Patients.

Excessive proliferation and activation of B cells, resulting in the production of various autoantibodies, is a crucial link and significant feature of the pathogenesis of systemic lupus erythematosus (SLE), as well as the pathological basis of systemic multiorgan damage. However, whether exosomes derived from human umbilical cord mesenchymal stem cells (hucMSCs-Exo) are involved in the immune regulation of SLE has not been clarified. Therefore, our study aimed to investigate the efficacy of hucMSCs-Exo for treating SLE. hucMSCs-Exo and peripheral blood mononuclear cells (PBMCs) of SLE patients were cocultured in vitro, and B cell apoptosis, activation, proliferation, and inflammation levels were detected by flow cytometry. Subsequently, the expression level of miR-155 in B lymphocytes of SLE patients was detected by qRT-PCR, and the target gene relationship between miR-155 and SHIP-1 was found through bioinformatics and dual luciferase activity experiments, which verified the inhibition of miR-155 in B lymphocytes of SLE patients to regulate immunity. We found that hucMSCs-Exo promoted B cell apoptosis, prevented B cell overactivation, and reduced inflammation. MicroRNA-155 (miR-155) has a powerful regulatory function in B cells. It was demonstrated that hucMSCs-Exo acts synergistically with miR-155 inhibitors to target SHIP-1 to B cells more effectively than exosomes alone. Our results provide insight into how hucMSCs-Exo regulates autoimmunity in patients with lupus and suggest targeting miR-155 for autoimmunity while protecting immunity.

PDGFRα promotes organ resistance to immune-mediated damage

Abstract Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that exhibits great clinical heterogeneity. Development and clinical expression of autoimmune diseases is strongly influenced by genetic variants. The single nucleotide polymorphism (SNP) rs1364989, located close to PDGFRA, has been associated with the development of lupus nephritis through an unknown mechanism. Because PDGF, through its α and β receptors, regulates tissue response to inflammation, we hypothesized that rs1364989 could modulate the behavior of renal cells in the context of immune-mediated glomerulonephritis. To determine whether rs1364989 affects the expression of PDGFRα, we analyzed umbilical cord mesenchymal cells from carriers of risk and protective alleles. We observed that the presence of the risk allele was associated with reduced PDGFRα expression. Next, we analyzed whether inflammatory mediators could modulate PDGFRα expression in in vitro and in vivo settings. We documented that its levels increase during acute inflammation in a murine model of glomerulonephritis. In vitro, this effect could be reproduced by IL-17. Absence of PDGFRα increased kidney damage during immune complex-induced glomerulonephritis. An analogous effect was observed in joints of mice with collagen-induced arthritis. Our results demonstrate that the SLE-associated allele of rs1364989 reduces the expression of PDGFRα and that this receptor may activate a pathway that protects tissues in response to local inflammation. These findings highlight the importance of tissue resilience as a factor that can protect an organ from inflammatory damage in the setting of autoimmunity. This work was supported by the Consejo Nacional de Ciencia y Tecnología (CONACYT), Mexico

Human umbilical cord mesenchymal stem cells-derived exosomes cells have a therapeutic potential in multiple sclerosis: a preclinical study

Human umbilical cord mesenchymal stem cells-derived exosomes cells have a therapeutic potential in multiple sclerosis: a preclinical study

Secreted responses of umbilical cord mesenchymal cells to full thickness swine burns are refined by the zone of injury

Background & Aim Primary treatment of battlefield burns is often inadequate due to delayed field evacuation or limited care in mass casualty incidents. Untreated burns expand in depth and surface area and elicit a hyper-immune response until the damaged tissue is removed. Third degree (full-thickness) burns penetrate the dermis and comprise 3 concentric zones of injury. The central zone of coagulation is irrevocably damaged while the surrounding zone of stasis is at risk. Tissue in the outermost zone of hyperemia can be saved despite an acute inflammatory response and increased perfusion. Mesenchymal stromal cells (MSCs) reportedly improve outcomes as post-surgical burn treatments. We assessed in vitro whether MSCs may also benefit burn sequelae if applied prior to surgical intervention. Methods, Results & Conclusion Nine anaesthetized swine received 3rddegree burns on the inner flank by a heated brass bar. After 30 minutes, the burn was excised ∼1 cm outside the zone of hyperemia. Control (unburned) samples were also excised. The burn center (zones of coagulation and stasis) and margin (zone of hyperemia) were isolated by dissection. Human umbilical cord MSCs (UC-MSCs), were co-incubated with 0.4 +/- 0.005g of tissue using a 6-well Transwell system for 24, 48 or 72 hours (h) in xeno-free media. Secreted proteins were assessed by multiplex ELISA of 115 analytes. Within 24h of exposure to any burn tissue, UC-MSCs increased output of inflammation mediators including fractalkine, GCP-2, GRO-β, I-309, IL-19, MCP-3, MIP3-α, MMP-2 and TSLP. IL-2Ra and MIG were specifically elevated in response to the necrotic center tissue, followed by β-NGF and IL-3 at 48h and SCF and SCGF-β at 72h. Intriguingly, the burn margin stimulated production of wound healing molecules in addition to inflammation mediators. G-CSF, HGF, IL-1β, IL-1RA, LIF and TECK increased significantly at 24 hours, followed by IL-20, IL-35, MDC, sCD163 and sTNF-R1 at 72h. Thus, UC-MSCs exhibit refined soluble responses that may improve outcomes for severe burn patients that initially receive sub-optimal treatment and delayed definitive care. The observed zone-specific responses suggest that the molecular mechanisms of MSC treatments may differ if the cells are applied before or after surgical debridement and depending on their localization within the wound. UC-MSCs delivered to unsalvageable tissue can produce a response focused on managing inflammation, while UC-MSCs delivered to the viable peripheral zone additionally promote wound healing.

Neural like cells and acetyl-salicylic acid alter rat brain structure and function following transient middle cerebral artery occlusion.

Introduction Transient cerebral ischemia is a pandemic neurological disorder and the main aim of medical intervention is to reduce complications. Human umbilical cord mesenchymal cells (hUCMs) are capable of differentiating into neural-like cells (NLC) in vitro, therefore we investigated the neuroprotective potential of these cells in comparison to aspirin and in combination (NLC-Aspirin) on spatial memory and neural morphologic changes in male rats submitted to transient cerebral ischemia. Methods Ten days after the intervention, the improvement in learning and memory were assessed in the animals by Morris Water Maze. Thence, the animals were examined for the presence of PKH26 labeled cells in the ischemic area of the brain, the infarct volume and neural changes in the brain tissue. Results Significant spatial memory deficits in the ischemic animals were detected compared with the control animals. The learning and memory were significantly improved (p ≤ 0.05) in the aspirin and NLC groups compared with the ischemic animals. Co-treatment of aspirin and NLCs did not improve the outcome. Moreover, infarction volume and neural changes were significantly altered when aspirin or NLCs were administered. Conclusions Our data suggest the significant neuroprotective potential of aspirin and neural-like cells derived from hUCM cells in the treatment of brain ischemic stroke. Further studies are required to evaluate possible underlying mechanisms, and to evaluate the possible interactions between aspirin and stem cells in a joint treatment aimed at the recovery of cognitive impairments.

Hypobaric Hypoxia Induces Autophagy of Umbilical Cord Mesenchymal Stem Cells

To investigate the autophagy activity changes of umbilical cord mesenchymal cells (MSC) under hypobaric hypoxia and the effect of hypobaric hypoxia on cell viability. Umbilical cord mesenchymal cells were cultured in the chamber of hypobaric hypoxia with an air pressure of 41.1 kPa and an oxygen density of 1%. At 0, 4, 8, 16, 24 and 48 hours, the cells were harvested for Western blot and real-time PCR to observe the expression level of the autophagy marker protein LC3B. And the cell viability under hypobaric hypoxia was evaluated after treatment with autophagy inhibitors HCQ (8 μg/ml) and 3MA (5 mmol/L). LC3B expression in MSC at protein and mRNA levels were up-regulated significantly after being cultured under hypobaric hypoxia condition for 8 hours. And compared with the control group, inhibition of autophagy reduced cell viability while increased Caspase-3 expression and the incidence of apoptosis. Hypobaric hypoxia activates autophagy in MSC, and the activation of autophagy might play a protective role for cell survival.

Isolation, molecular characterization, and in vitro differentiation of bovine Wharton jelly–derived multipotent mesenchymal cells

Extrafetal tissues are a noncontroversial and inexhaustible source of mesenchymal stem cells that can be harvested noninvasively at low cost. In the veterinary field, as in man, stem cells derived from extrafetal tissues express plasticity, reduced immunogenicity, and have high anti-inflammatory potential making them promising candidates for treatment of many diseases. Umbilical cord mesenchymal cells have been isolated and characterized in different species and have recently been investigated as potential candidates in regenerative medicine. In this study, cells derived from bovine Wharton jelly (WJ) were isolated for the first time by enzymatic methods, frozen/thawed, cultivated for at least 10 passages, and characterized. Wharton jelly–derived cells readily attached to plastic culture dishes displaying typical fibroblast-like morphology and, although their proliferative capacity decreased to the seventh passage, these cells showed a mean doubling time of 34.55 ± 6.33 hours and a mean frequency of one colony-forming unit fibroblast like for every 221.68 plated cells. The results of molecular biology studies and flow cytometry analyses revealed that WJ-derived cells showed the typical antigen profile of mesenchymal stem cells and were positive for CD29, CD44, CD105, CD166, Oct-4, and c-Myc. They were negative for CD34 and CD14. Remarkably, WJ-derived cells showed differentiation ability. After culture in induced media, WJ-derived cells were able to differentiate into osteogenic, adipogenic, chondrogenic, and neurogenic lines as shown by positive staining and expression of specific markers. On polymerase chain reaction analysis, these cells were negative for MHC-II and positive for MHC-I, thus reinforcing the role of extrafetal tissue as an allogenic source for bovine cell–based therapies. These results provide evidence that bovine WJ-derived cells may have the potential to differentiate to repair damaged tissues and reinforce the importance of extrafetal tissues as stem cell sources in veterinary regenerative medicine. A more detailed evaluation of their immunologic properties is necessary to better understand their potential role in cellular therapy.

Comparison between isolation protocols highlights intrinsic variability of human umbilical cord mesenchymal cells

Mesenchymal stem cells (MSCs) though multipotent exhibit limited lifespan in vitro, with progressive reduction in capacity for self-renewal leading to irreversible arrest of cell division, which limits their use for therapeutic purposes. Human umbilical cord wall MSCs are easy to process and proliferate rapidly in culture, but variability of individual samples and impact upon in vitro expansion and aging processes is unknown. We compared isolation protocols to determine which one yields the highest number of viable cells with the best proliferation capacity. Three different protocols were tested: two were enzymatic procedures and one explant method. Isolated cells were evaluated in terms of proliferation, differentiation capacity, and phenotype. All samples were processed using one or more protocols. After passage 2 adherent cells displayed standard phenotypic and differentiation characteristics of MSCs, but our results show that isolating cells directly from Wharton's jelly is more advantageous. Cells obtained from explants presented similar characteristics to those from enzymatic protocols, but always reached proliferation arrest earlier, irrespective of initial population doubling times. From the same sample, cells obtained with enzymatic protocol ii reached later passages while exhibiting shorter doubling times in culture than cells from other protocols, that is, took longer to reach senescence. More important, each individual MSC sample exhibited different population doubling rates and reached senescence at different passages, irrespective of protocol. Thus, even when in strict conformity with procedures and quality control, each cord sample shows a unique behavior, a finding that should be taken into account when planning for therapeutic approaches.

Transplantation of differentiated umbilical cord mesenchymal cells under kidney capsule for control of type I diabetes in rat.

Nowadays, stem cells have been introduced as an appropriate source of regenerative medicine for treatment of type I diabetes. Human umbilical cord matrix-derived mesenchymal cells (hUCMC) have successfully been differentiated into insulin producing cells. The isolated hUCM cells were characterized by the expression of stem cell surface markers and by differentiation into adipocytes and osteocytes. The hUCMCs were cultured with different concentrations of neural conditional medium (NCM) and were induced to differentiate into insulin producing cells (IPCs). As 60% NCM concentration resulted in higher nestin and PDX1 expression, the cells were first exposed to 60% NCM and were then induced for IPCs differentiation. PDX1 and insulin gene expression was evaluated in the treated cells. Also, the secretion capacity of the IPCs was assessed by glucose challenge test. IPCs were transferred under the rat kidney capsule. Blood glucose level, weight gain and immunohistochemistry assessments were done in the treated animals. hUCMC expressed mesenchymal cell surface markers and successfully differentiated into adipocytes and osteocytes. Higher NCM concentration resulted in higher PDX1 and nestin expression. The IPCs expressed insulin and PDX1. IPCs were detectable under the kidney capsule 2 months after injection. IPCs transplantation resulted in a sharp decline of blood sugar level and less weight loss. Differentiated hUCM cells could alleviate the insulin deprivation in the rat model of type I diabetes. In addition, higher NCM concentration leads to more differentiation into IPCs and more nestin and PDX1 expression. Kidney capsule can serve as a suitable nominee for IPCs transplantation.

Evaluation of neurogenic potential of human umbilical cord mesenchymal cells; a time- and concentration-dependent manner.

Background:Retinoic acid as one of the most important regulators for cell differentiation was examined in this study for differentiation of human umbilical mesenchymal cells (hUCM). Methods: After isolation, hUCM were evaluated for mesenchymal stem cell properties by flow cytometry and alkaline phosphatase assay. Also, doubling time of the cells and their differentiation potential into adipogenic and osteogenic cells were tested. hUCM were then cultured with different concentrations of retinoic acid, and on days 1, 7, and 12, the percentage of differentiated cells was determined by immunostaining for nestin, anti-microtubule associated protein 2 (MAP2), glutamic acid decarboxylase (GAD), and gamma-aminobutyric acid (GABA) markers. Results:The isolated cells were negative for the hematopoietic markers and positive for the mesenchymal markers. They showed the population doubling time 60 ± 3 hours and differentiated into osteogenic and adipogenic cells. A descending trend in nestin and an ascending trend in MAP2, GAD, and GABA expression were observed from the first day until the last day between different concentrations of retinoic acid. Conclusion: hUCM cells may have the potential to differentiate into neural cells in the presence of different incubation period and concentration of retinoic acid.

PO-0370 Germ Cells Induced From Human Umbilical Cord Mesenchymal Cell-derived Induced Pluripotent Stem Cells By Bmp4

<h3>Objective</h3> To reprogramme the induced pluripotent stem (iPS) cells from Human umbilical cord mesenchymal cells (HuMSCs) and induce the iPS cells into germ cells by BMP4. <h3>Methods</h3> OCT4, SOX2, Klf4,c-myc, Nanog, Lin28 were transfected into HuMSCs with lentivirus to reprogram HuMSCs into iPS cells. Morphological observation, Alkaline Phosphatase staining, karyotype analysis, RT-PCR, immunofluorescence staining, tumour formation <i><i>in vivo</i></i> and embryniod body formation <i><i>in vitro</i></i> were performed to examine the pluripotency of the iPS cell lines. Then we induced one of the iPS cells lines into germ cells by BMP4. Gene expression was measured by qRT-PCR at days 0, 3, 7, 10 and 14. Early-stage germ specific protein VASA and meiosis specific protein SYCP3 were assesssed by immunofluorescence staining. <h3>Result</h3> We obtained two iPS cell lines completely reprogrammed, HuMSC-iPS1 and HuMSC-iPS2. HuMSC-iPS1 expresses germ cell markers at undifferentiated state. BMP4 can upregulate germ cell markers at different time points highly while the spontaneous differentiation just upregulate DPPA3, DAZL and VASA modestly at day 3. However, all of these genes were downregulated at day 14. VASA and SYCP3 immunofluorescence staining indicates there is a high VASA expression in BMP4 induced group in contrast to low expression in the spontaneous group at day 7. Meanwhile, there is a modest SYCP3 fluorescence in BMP4 induced group in contrast to no immunofluoresence in the spontaneous group. <h3>Conclusion</h3> This system can reprogram HuMSCs into iPS cells effectively. The MSC- iPS1 can differentiate into early germ cells spontaneously while the germ cells induced by BMP4 can enter meiosis.

Expression of epithelial markers by human umbilical cord stem cells. A topographical analysis

IntroductionHuman umbilical cord stem cells have inherent differentiation capabilities and potential usefulness in regenerative medicine. However, the epithelial differentiation capability and the heterogeneity of these cells have not been fully explored to the date. MethodsWe analyzed the expression of several undifferentiation and epithelial markers in cells located in situ in different zones of the umbilical cord –in situ analysis– and in primary ex vivo cell cultures of Wharton's jelly stem cells by microarray and immunofluorescence. ResultsOur results demonstrated that umbilical cord cells were heterogeneous and had intrinsic capability to express in situ stem cell markers, CD90 and CD105 and the epithelial markers cytokeratins 3, 4, 7, 8, 12, 13, 19, desmoplakin and zonula occludens 1 as determined by microarray and immunofluorescence, and most of these markers remained expressed after transferring the cells from the in situ to the ex vivo cell culture conditions. However, important differences were detected among some cell types in the umbilical cord, with subvascular zone cells showing less expression of stem cell markers and cells in Wharton's jelly and the amnioblastic zones showing the highest expression of stem cells and epithelial markers. ConclusionsThese results suggest that umbilical cord mesenchymal cells have intrinsic potential to express relevant epithelial markers, and support the idea that they could be used as alternative cell sources for epithelial tissue engineering.

Induced pluripotent stem cells can be reprogrammed from human umbilical cord mesenchymal cells by six transcription factors

Objective To establish and identify the induced pluripotent stem cell(iPSC) line reprogrammed from human umbilical cord mesenchymal cells(HuMSCs). Methods HuMSCs were cultured by adhesion method, and OCT4, SOX2, KLF4, c-Myc, NANOG, LIN-28 were transfected into HuMSCs with lentiviral victor to reprogramme HuMSCs into iPSC.Morphological observation, pluripotency genes(SOX2, TDGF1, THY-1, OCT4, REX1 and TERF1) expression, alkaline phosphatase detection, karyotype analysis, embryonic stem cells(ESC) specific proteins(NANOG, OCT4, SSEA-4, TRA-1-81) immunofluorescence staining, differentiated into teratomas in vivo(inject the iPSC into SCID mice) and embryniod bodies in vitro were performed to exam the pluripotency of the iPSC. Results Four days after being infected by lentivirus, the HuMSCs became round-shape; 10 days after infection, some embryonic stem(ESC)-like colonies appeared.Fourteen days after infection, picked up the regularly shaped colonies and cultured several passages.About 1.25% HuMSCs were reprogrammed into iPSC.The iPSC presented clone-like growth like ESC.All the cells were positive to alkaline phosphatase staining and expressed the pluripotency genes.The iPSC also expressed the ESC specific proteins, and karyotype analysis showed normal chromosome caryotype(46, XY). Furthermore, the iPSC could form embryoid bodies in vitro, expressed alpha fetoprotein(AFP), smooth muscle actin(SMA) and β-tubulin.The iPSC could alsoform teratomas in vivo. Conclusion OCT4, SOX2, KLF4, c-Myc, NANOG, LIN-28 can reprogram HuMSCs into iPSC efficiently. Key words: Umbilical cord mesenchymal stem cells; Induced pluripotent stem cells; Transcription factor; Lentiviral vector

Effect of umbilical cord MSC infusion on the pulmonary infection in haploidentical hematopoietic stem cell transplantation

This study was purposed to investigate the effect of umbilical cord mesenchymal cells (UC-MSC) infusion on the pulmonary infection in haploidentical hematopoietic stem cell transplantation (hi-HSCT). The infection of 83 patients underwent hi-HSCT was detected and analysed, among them 42 patients received haploidentical hi-HSCT, 41 received hi-HSCT combined with UC-MSC infusion. The results showed that 31 cases (73.81% ± 6.78%) were infected by cytomegalovirus and 21 cases in patients received hi-HSCT experienced pulmonary infections, including infections of fungal, virus, bacteria, tubercle bacillus, PCP and so on, the incidence rate was (50 ± 7.72)%; the infection of cytomegalovirus (CMV) was found in 31 cases, the incidence rate was (78.05 ± 6.46)%. In patients received hi-HSCT combined with UC-MSC, only 15 patients experienced pulmonary infection, the incidence rate was (36.59 ± 7.52)%, and the infection of cytomegalovirus (CMV) was observed in 32 patients, the incidence rate was (78.05 ± 6.46)%. There was no obvious statistical difference between two groups(P > 0.05). It is concluded that the UC-MSC infusion not increases the infection rate in hi-HSCT.

Gene expression profile changes induced upon umbilical cord mesenchymal cell infusion therapy in a rat model of hepatic cirrhosis

To investigate changes in gene expression that occur upon treatment with human umbilical cord mesenchymal stem cells (UC-MSCs) for hepatic cirrhosis using a rat model system. Hepatic cirrhosis was induced in Sprague-Dawley rats by subcutaneous injection of carbon tetrachloride and oral administration of alcohol.UC-MSCs were isolated from human umbilical cord and the cells' immunophenotype and differentiation towards osteogenic and adipogenic lineages were confirmed.The UC-MSC sample or vehicle alone (phosphate buffered saline, PBS) was transplanted by intravenous injection.Histopathological staining and serological testing were used to compare the liver morphology and function among the different groups.The gene expression in the PBS group and UC-MSC group were detected by gene microarray and differences between the groups were statistically analyzed by t-test. Transplantation of the UC-MSCs improved liver function in the hepatic cirrhosis rats.Comparison of the gene expression profiles of the PBS group and the UC-MSC group showed that the latter had up-regulation of the genes related to the complement and coagulation cascades and down-regulation of the genes related to cell proliferation, cell cycle, and collagen synthesis. UC-MSC therapy might improve liver function in cirrhosis by increasing the expression of genes related to the complement and coagulation cascades and by decreasing genes involved in cell proliferation and collagen deposition.