Search results for: adipose derived stem cells

Authors: Mahmoud M. Zakaria, Omnia F. Elmoursi, Mahmoud M. Gabr, Camelia A. AbdelMalak, Mohamed A. Ghoneim

Abstract:

Many protocols were publicized for differentiation of human mesenchymal stem cells (MSCS) into insulin-producing cells (IPCs) in order to excrete insulin hormone ingoing to treat diabetes disease. Our aim is to evaluate relative gene expression for each independent protocol. Human bone marrow cells were derived from three volunteers that suffer diabetes disease. After expansion of mesenchymal stem cells, differentiation of these cells was done by three different protocols (the one-step protocol was used conophylline protein, the two steps protocol was depending on trichostatin-A, and the three-step protocol was started by beta-mercaptoethanol). Evaluation of gene expression was carried out by real-time PCR: Pancreatic endocrine genes, transcription factors, glucose transporter, precursor markers, pancreatic enzymes, proteolytic cleavage, extracellular matrix and cell surface protein. Quantitation of insulin secretion was detected by immunofluorescence technique in 24-well plate. Most of the genes studied were up-regulated in the in vitro differentiated cells, and also insulin production was observed in the three independent protocols. There were some slight increases in expression of endocrine mRNA of two-step protocol and its insulin production. So, the two-step protocol was showed a more efficient in expressing of pancreatic endocrine genes and its insulin production than the other two protocols.

Keywords: mesenchymal stem cells, insulin producing cells, conophylline protein, trichostatin-A, beta-mercaptoethanol, gene expression, immunofluorescence technique

Procedia PDF Downloads 191

Authors: Parcha Sreenivasa Rao, P. Lakshmi

Abstract:

Diabetes Mellitus is metabolic disorder, characterized by high glucose levels in the blood due to one of the reason i.e., the death of β cells. The lack of β cells leads to the reduced insulin levels. The β cell death generally occurs due to apoptosis induced by the several cytokines. IL-1β, IFN- ϒ and TNF –α cytokines that are generally cause apoptosis to the β cell. The nutrient based apoptosis is generally seen with high glucose and free fatty acids. It is also noted that the β cell death triggered by Fas ligand and its receptor Fas at the surface of the activated CD8+ T- lymphocytes. Reports also reveal that the β cell apoptosis is under control of the transcription factors NF-kB and STAT- 1. The arresting or opposing of the β cell apoptosis can be overcome by the different growth factors like GLP-1, growth hormone, prolactin, VEGF, Dipeptidyl peptidase-4, Vildagliptin, suberoylanilidehydroxamic acid, trichistatin-A, XIAP, Bcl-2, FGF-21. Present investigation explains antiapoptotic property of the β cells derived from the mesenchymal stem cells of umbilical cord.

Keywords: stem cells, umblical cord, diabetes, apoptosis

Procedia PDF Downloads 353

Authors: Ahmad Shah Farhat

Abstract:

Background: Prenatal asphyxia or birth asphyxia is the medical situation resulting from a newborn infant that lasts long enough during the birth process to cause physical harm, usually to the brain. Human umbilical cord blood (UCB) is a well-established source of hematopoietic stem/progenitor cells (HSPCs) for allogeneic stem cell transplantation. These can be used clinically to care for children with malignant diseases. Low O2 can cause in proliferation and differentiation of stem cells. Method: the cord blood of 11 infants with 3-5 Apgar scores or need to cardiac pulmonary Resuscitation as an asphyxia group and ten normal infants with more than 8 Apgar scores as the normal group was collected, and after isolating hematopoietic stem cells, the cells were cultured in enriched media for 14 days to compare the numbers of colonies by microscope. Results: There was a significant difference in the number of RBC precursor colonies (red colonies) in cultured media with 107 cord blood hematopoietic stem cells of infants who were exposed to hypoxemia in two wells of palate. There was not a significant difference in the number of white cell colonies in the two groups in the two wells of the plate. Conclusion: Hypoxia in the perinatal period can cause the increase of hematopoietic stem cells of cord blood, special red precursor stem cells in vitro, like an increase of red blood cells in the body when exposed to low oxygen conditions. Thus, it will be usable.

Keywords: asphyxia, neonre, stem cell, red cell

Procedia PDF Downloads 41

Authors: Laila Montaser, Hala Gabr, Maha El-Bassuony, Gehan Tawfeek

Abstract:

Orthotropic liver transplantation (OLT) is the final procedure of both end stage and metabolic liver diseases. Hepatocyte transplantation is an alternative for OLT, but the sources of hepatocytes are limited. Bone marrow mesenchymal stem cells (BM-MSCs) can differentiate into hepatocyte-like cells and are a potential alternative source for hepatocytes. The MSCs from bone marrow are a promising target population as they are capable of differentiating along multiple lineages and, at least in vitro, have significant expansion capability. MSCs from bone marrow may have the potential to differentiate in vitro and in vivo into hepatocytes. Our study examined whether mesenchymal stem cells (MSCs), which are stem cells originated from human bone marrow, are able to differentiate into functional hepatocyte-like cells in vitro. Our aim was to investigate the differentiation potential of BM-MSCs into hepatocyte-like cells. Adult stem cell therapy could solve the problem of degenerative disorders, including liver disease.

Keywords: bone marrow, differentiation, hepatocyte, stem cells

Procedia PDF Downloads 488

Authors: Ashok K. Gupta

Abstract:

Over the past few decades, since the bio-engineering revolution, autologous cell therapy (ACT) has become a rapidly evolving field. Currently, this form of therapy has broad applications in modern medicine and plastic surgery, ranging from the treatment/improvement of wound healing to life-saving operations. A study was conducted on 50 patients having to disfigure, and deform post burn scars and was treated by injection of extracted, refined adipose tissue grafts with their unique stem cell properties. To compare the outcome, a control of 20 such patients was treated with conventional skin or soft-tissue flaps or skin grafting, and a control of 10 was treated with more advanced microsurgical techniques such as Pre-fabricated flaps/pre laminated flaps / free flaps. Assessment of fat volume and survival post- follow up period was done by radiological aid, using MRI and clinically (Survival of the autograft and objective parameters for scar elasticity were evaluated skin elasticity parameters 3 to 9 months postoperatively). Recently, an enzyme that is involved in collagen crosslinking in fibrotic tissue, lysyl hydroxylase (LH2), was identified. This enzyme is normally active in bone and cartilage but hardly in the skin. It has been found that this enzyme is highly expressed in scar tissue and subcutaneous fat; this is in contrast to the dermis, where the enzyme is hardly expressed. Adipose tissue-derived stem cell injections are an effective method in the treatment of various extensive post-burn scar deformities that makes it possible to re-create the lost sub-dermal tissue for improvement in the function of involved joint movements.

Keywords: adipose tissue-derived stem cell injections, treatment of various extensive post-burn scar deformities, re-create the lost sub-dermal tissue, improvement in function of involved joint movements

Procedia PDF Downloads 40

Authors: Rasha Ahmadi

Abstract:

Glioblastoma is one of the most frequent brain malignancies, having a high mortality rate and limited survival in individuals with this malignancy. Despite different treatments and surgery, recurrence of glioblastoma cancer stem cells may arise as a subsequent tumor. For this reason, it is crucial to research the markers associated with glioblastoma stem cells and specifically their microenvironment. In this study, using bioinformatics analysis, we analyzed and nominated genes in the microenvironment pathways of glioblastoma stem cells. In this study, an appropriate database was selected for analysis by referring to the GEO database. This dataset comprised gene expression patterns in stem cells derived from glioblastoma patients. Gene clusters were divided as high and low expression. Enrichment databases such as Enrichr, STRING, and GEPIA were utilized to analyze the data appropriately. Finally, we extracted the potential genes 2700 high-expression and 1100 low-expression genes are implicated in the metabolic pathways of glioblastoma cancer progression. Cellular senescence, MAPK, TNF, hypoxia, zimosterol biosynthesis, and phosphatidylinositol metabolism pathways were substantially expressed and the metabolic pathways were downregulated. After assessing the association between protein networks, MSMP, SOX2, FGD4 ,and CNTNAP3 genes with high expression and DMKN and SBSN genes with low were selected. All of these genes were observed in the survival curve, with a survival of fewer than 10 percent over around 15 months. hsa-mir-192-5p, hsa-mir-129-5p, hsa-mir-215-5p, hsa-mir-335-5p, and hsa-mir-340-5p played key function in glioblastoma cancer stem cells microenviroments. We introduced critical genes through integrated and regular bioinformatics studies by assessing the amount of gene expression profile data that can play an important role in targeting genes involved in the energy and microenvironment of glioblastoma cancer stem cells. Have. This study indicated that hsa-mir-192-5p, and hsa-mir-129-5p are appropriate candidates for this.

Keywords: Glioblastoma, Cancer Stem Cells, Biomarker Discovery, Gene Expression Profiles, Bioinformatics Analysis, Tumor Microenvironment

Procedia PDF Downloads 108

Authors: Kumari Rekha, Mathur K Dhananjay, Maheshwari Deepanshu, Nautiyal Nidhi, Shubham Smriti, Laal Deepika, Sinha Swati, Kumar Anupam, Biswas Subhrajit, Shiv Kumar Sarin

Abstract:

Background: Mesenchymal stem cells are multipotent stem cells derived from mesoderm and are used for therapeutic purposes because of their self-renewal, homing capacity, Immunomodulatory capability, low immunogenicity and mitochondrial transfer signaling. MSCs have the ability to regulate the mechanism of both innate as well as adaptive immune responses through the modulation of cellular response and the secretion of inflammatory mediators. Different sources of MSC are UC MSC, BM MSC, Dental Pulp, and Adipose MSC. The most frequent source used is umbilical cord tissue due to its being easily available and free of limitations of collection procedures from respective hospitals. The immunosuppressive role of MSCs is particularly interesting for clinical use since it confers resistance to rejection by the host immune response. Methodology: In this study, T helper cells (TH4), Cytotoxic T cells (CD-8), immunoregulatory cells (CD25 +FOXP3+) are compared from isolated MSC from three different methods, UC Dissociation Kit (Miltenyi), Explant Culture and Collagenase Type-IV. To check the immunomodulatory property, these MSCs were seeded with PBMC(Coculture) in CD3 coated 24 well plates. Cd28 antibody was added in coculture for six days. The coculture was analyzed in FACS Verse flow cytometry. Results: From flow cytometry analysis of coculture, it found that All over T helper cells (CD4+) number p<0.0264 increases in (All Enzymes) MSC rather than explant MSC(p>0.0895) as compared to Collagenase(p>0.7889) in a coculture of Activated T cell and Mesenchymal Stem Cell. Similar T reg cells (CD25+, FOXP3+) expression p<0.0234increases in All Enzymes), decreases in Explant and Collagenase. Experiments have shown that MSCs can also directly prevent the cytotoxic activity of CD8 lymphocytes mainly by blocking their proliferation rather than by inhibiting the cytotoxic effect. And promoting the t-reg cells, which helps in the mediation of immune response in various diseases. Conclusion: MSC suppress Cytotoxic CD8 T cell and Enhance immunoregulatory T reg (CD4+, CD25+, FOXP3+) Cell expression. Thus, MSC maintains a proper balance(ratio) between CD4 T cells and Cytotoxic CD8 T cells.

Keywords: MSC, disease, T cell, T regulatory

Procedia PDF Downloads 80

Authors: Ahmed Khalaf Reyad Raslan

Abstract:

Cancer stem cells (CSCs) describe a class of pluripotent cancer cells that behave analogously to normal stem cells in their ability to differentiate into the spectrum of cell types observed in tumors. The de-differentiation processes, such as an epithelial-mesenchymal transition (EMT), are known to enhance cellular plasticity. Here, we demonstrate a new hypothesis to use hybrid superparamagnetic magnetite nanoparticles (Fe₃O₄)- graphene oxide functionalized surface with Collagen to target the cancer stem cell as an early detection tool for cancer. We think that with the use of magnetic resonance imaging (MRI) and the new hybrid system would be possible to track the cancer stem cells.

Keywords: hydrogel, alginate, reduced graphene oxide, collagen

Procedia PDF Downloads 119

Authors: Natasha Petrovska, Mirjana Pavlovic, Maria M. Larrondo-Petrie

Abstract:

Neural stem cells (NSCs) are multi-potent, self-renewing cells that generate new neurons. Three subtypes of NSCs can be separated regarding the stages of NSC lineage: quiescent neural stem cells (qNSCs), activated neural stem cells (aNSCs) and neural progenitor cells (NPCs), but their gene expression signatures are not utterly understood yet. Single-cell examinations have started to elucidate the complex structure of NSC populations. Nevertheless, there is a lack of thorough molecular interpretation of the NSC lineage heterogeneity and an increasing need for tools to analyze and improve the efficiency and correctness of single-cell sequencing data. Feature selection and ordering can identify and classify the gene expression signatures of these subtypes and can discover novel subpopulations during the NSCs activation and differentiation processes. The aim here is to review the implementation of the feature selection technique on NSC subtypes and the classification techniques that have been used for the identification of gene expression signatures.

Keywords: feature selection, feature similarity, neural stem cells, genes, feature selection methods

Procedia PDF Downloads 111

Authors: Jer Ping Ooi, Shah Rizal Bin Kasim, Nor Aini Saidin

Abstract:

The objective of this study was to synthesize nano-sized β-tricalcium phosphate (β-TCP) powder and assess its cytotoxic effects on human osteoblast (hFOB1.19) by using four cytotoxicity assays, namely, lactose dehydrogenase (LDHe), tetrazolium hydroxide (XTT), neutral red (NR), and sulforhodamine B (SRB) assays. β-tricalcium phosphate (β-TCP) is a calcium phosphate compound commonly used as an implant material. To date, bulk-sized β-TCP is reported to be readily tolerated by the osteogenic cells and body based on in vitro, in vivo experiments and clinical studies. However, to what extent of nano-sized β-TCP will react in models as compared to bulk β-TCP is yet to be investigated. Thus, in this project, the cells were treated with nano β-TCP powder within a range of concentrations from 0 to 1000 μg/mL for 24, 48, and 72 h. The cytotoxicity tests showed that loss of cell viability ( > 50%) was high for hFOB1.19 cells in all assays. Cell cycle and apoptosis analysis of hFOB1.19 cells revealed that 50 μg/mL of the compound led to 30.5% of cells being apoptotic after 72 h of incubation, and the percentage was increased to 58.6% when the concentration was increased to 200 μg/mL. When the incubation time was increased from 24 to 72 h, the percentage of apoptotic cells increased from 17.3% to 58.6% when the hFOB1.19 were exposed with 200 μg/mL of nano β-TCP powder. Thus, both concentration and exposure duration affected the cytotoxicity effects of the nano β-TCP powder on hFOB1.19. We hypothesize that these cytotoxic effects on hFOB1.19 are related to the nano-scale size of the β-TCP.

Keywords: β-tricalcium phosphate, hFOB1.19, adipose-derived mesenchymal stem cells, cytotoxicity

Procedia PDF Downloads 290

Authors: Arefeh Jafarian, Mohammad Taghikani, Saied Abroun, Amir Allahverdi, Masoud Soleimani

Abstract:

Introduction: The miRNAs have key roles in control of pancreatic islet development and insulin secretion. In this regards, current study investigated the pancreatic differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) by up-regulation of miR-375 and down-regulation of miR-9 by lentiviruses containing miR-375 and anti-miR-9. Findings: After 21 days of induction, islet-like clusters containing insulin producing cells (IPCs) were confirmed by dithizone (DTZ) staining. The IPCs and β cell specific related genes and proteins were detected using qRT-PCR and immunofluorescence on days 7, 14 and 21 of differentiation. Glucose challenge test was performed at different concentrations of glucose as well as extracellular and intracellular insulin and C-peptide were assayed using ELISA kit. In derived IPCs by miR-375 alone are capable to express insulin and other endocrine specific transcription factors, the cells lack the machinery to respond to glucose. The differentiated hMSCs by miR-375 and anti-miR-9 lentiviruses could secrete insulin and c-peptide in a glucose-regulated manner. Conclusion: It was found that over-expression of miR-375 led to a reduction in levels of Mtpn protein in derived IPCs, while treatment with anti-miR-9 following miR-375 over-expression had synergistic effects on MSCs differentiation and insulin secretion in a glucose-regulated manner. The researchers reported that silencing of miR-9 increased OC-2 protein in IPCs that may contribute to the observed glucose-regulated insulin secretion. These findings highlight miRNAs functions in stem cells differentiation and suggest that they could be used as therapeutic tools for gene-based therapy in diabetes mellitus.

Keywords: diabetes, differentiation, MSCs, insulin producing cells, miR-375, miR-9

Procedia PDF Downloads 292

Authors: Lina Liang, Kai Xu, Jing Zhang

Abstract:

Purpose: Age-related macular degeneration (AMD) is a major leading cause of visual impairment and blindness with no cure currently established. Cell replacement is discussed as a potential therapy for AMD. Besides intravitreal injection and subretinal injection, intravenous administration has been explored as an alternative route. This study is to observe the effect of BSYJ, a traditional Chinese medicine on the homing and differentiation of mesenchymal stem cells transplanted via tail vein injection in an age-related macular degeneration mouse model. Methods: Four-week-old C57BL/6J mice were injected with 40 mg/kg NaIO₃ to induce age-related macular degeneration model. At the second day after NaIO₃ injection, 1×10⁷ GFP labeled bone marrow-derived mesenchymal stem cells (GFP-MSCs) were transplanted via tali vein injection into the experimental mice. Then the mice were randomly divided into two groups, gavaged with either BSYJ solution (BSYJ group, n=12) or distilled water (DW group, n=12). 12 age-matched healthy C57BL/6J mice were fed regularly as normal control. At day 7, day 14, and day 28 after treatment, retina flat mounting was used to detect the homing of mesenchymal stem cells at the retina. Double-labeling immunofluorescence was used to determine the differentiation of mesenchymal stem cells. Results: At 7, 14, 28 days after treatment, the numbers of GFP-MSCs detected by retina flatmount were 10.2 ± 2.5, 14.5 ± 3.4 and 18.7 ± 5.8, respectively in the distilled water group, while 15.7 ± 3.8, 32.3 ± 3.5 and 77.3 ± 6.4 in BSYJ group, the differences between the two groups were significant (p < 0.05). At 28 days after treatment, it was shown by double staining immunofluorescence that there were more GFP positive cells in the retina of BSYJ group than that of the DW group, but none of the cells expressed RPE specific genes such as RPE65 and CRALBP, or photoreceptor genes such as recoverin and rhodopsin either in BSYJ group or DW group. However, GFAP positive cells were found among the cells labeled with GFP, and the double labeling cells were much more in the BSYJ group than the distilled water group. Conclusion: BSYJ could promote homing of mesenchymal stem cells at the retina of age-related macular degeneration model mice induced by NaIO₃, and the differentiation towards to glial cells. Acknowledgement: National Natural Foundation of China (No: 81473736, 81674033,81973912).

Keywords: BSYJ, differentiation, homing, mesenchymal stem cells

Procedia PDF Downloads 107

Authors: A. Reum Son, Jin Seon Kwon, Seung Hun Park, Hai Bang Lee, Moon Suk Kim

Abstract:

These days, stem cell therapy is focused on for promising source of treatment in clinical human disease. As a supporter of stem cells, in situ-forming hydrogels with growth factors and cells appear to be a promising approach in tissue engineering. To examine osteogenic differentiation of hTMSCs which is one of mesenchymal stem cells in vivo in an injectable hydrogel, we use a methoxy polyethylene glycol-polycaprolactone blockcopolymer (MPEG-PCL) solution with osteogenic factors. We synthesized MPEG-PCL hydrogel and measured viscosity to check sol-gel transition. In order to demonstrate osteogenic ability of hTMSCs, we conducted in vitro osteogenesis experiment. Then, to confirm the cell cytotoxicity, we performed WST-1 with hTMSCs and MPEG-PCL. As the result of in vitro experiment, we implanted cell and hydrogel mixture into animal model and checked degree of osteogenesis with histological analysis and amount of expression genes. Through these experimental data, MPEG-PCL hydrogel has sol-gel transition in temperature change and is biocompatible with stem cells. In histological analysis and gene expression, hTMSCs are very good source of osteogenesis with hydrogel and will use it to tissue engineering as important treatment method. hTMSCs could be a good adult stem cell source for usability of isolation and high proliferation. When hTMSCs are used as cell therapy method with in situ-formed hydrogel, they may provide various benefits like a noninvasive alternative for bone tissue engineering applications.

Keywords: injectable hydrogel, stem cell, osteogenic differentiation, tissue engineering

Procedia PDF Downloads 423

Authors: Mohammad Nasb, Muhammad Rehan, Chen Hong

Abstract:

Background Meniscus defects critically alter knee function and lead to degenerative changes. Regenerative medicine applications including stem cell transplantation have showed a promising efficacy in finding alternatives to overcome traditional treatment limitations. However, stem cell therapy remains limited due to the substantially reduced viability and inhibitory microenvironment. Since tissue growth and repair are under the control of biochemical and mechanical signals, several approaches have recently been investigated (e.g., low intensity pulsed ultrasound [LIPUS]) to promote the regeneration process. This study employed LIPUS to improve growth and osteogenic differentiation of mesenchymal stem cells derived from human embryonic stem cells to improve the regeneration of meniscus tissue. Methodology: The Mesenchymal stromal cells (MSCs) were transplanted into the epicenter of the injured meniscus in rabbits, which were randomized into two main groups: a treatment group (n=32 New Zealand rabbits) including 4 subgroups of 8 rabbits in each subgroup (LIPUS treatment, MSC treatment, LIPUS with MSC and control), and a second group (n=9) to track implanted cells and their progeny using green fluorescence protein (GFP). GFP consists of the MSC and LIPUS-MSC combination subgroups. Rabbits were then subjected to histological, immunohistochemistry, and MRI assessment. Results: The quantity of the newly regenerated tissue in the combination treatment group that had Ultrasound irradiation after mesenchymal stem cells were better at all end points. Likewise, Tissue quality scores were also greater in knees treated with both approaches compared with controls and single treatment at all end points, achieving significance at twelve and twenty-four weeks [p < 0.05], and [p = 0.008] at twelve weeks. Differentiation into type-I and II collagen-expressing cells were higher in the combination group at up to twenty-four weeks. Conclusions: the combination of mesenchymal stem cells and LIPUS showed greater adhering to the sites of meniscus injury, differentiate into cells resembling meniscal fibrochondrocytes, and improve both quality and quantity of meniscal regeneration.

Keywords: stem cells, regenerative medicine, osteoarthritis, knee

Procedia PDF Downloads 92

Authors: Dipali Dhawan

Abstract:

Cancerous epithelial cells are confined to a primary site by the continued expression of adhesion molecules and the intact basal lamina. However, as the cancer progresses some cells are believed to undergo an epithelial-mesenchymal transition (EMT) event, leading to increased motility, invasion and, ultimately, metastasis of the cells from the primary tumour to secondary sites within the body. These disseminated cancer cells need the ability to self-renew, as stem cells do, in order to establish and maintain a heterogeneous metastatic tumour mass. Identification of the specific subpopulation of cancer stem cells amenable to the process of metastasis is highly desirable. In this study, we have isolated and characterized cancer stem cells from luminal and basal breast cancer cell lines (MDA-MB-231, MDA-MB-453, MDA-MB-468, MCF7 and T47D) on the basis of cell surface markers CD44 and CD24; as well as Side Populations (SP) using Hoechst 33342 dye efflux. The isolated populations were analysed for epithelial and mesenchymal markers like E-cadherin, N-cadherin, Sfrp1 and Vimentin by Western blotting and Immunocytochemistry. MDA-MB-231 cell lines contain a major population of CD44+CD24- cells whereas MCF7, T47D and MDA-MB-231 cell lines show a side population. We observed higher expression of N-cadherin in MCF-7 SP cells as compared to MCF-7NSP (Non-side population) cells suggesting that the SP cells are mesenchymal like cells and hence express increased N-cadherin with stem cell-like properties. There was an expression of Sfrp1 in the MCF7- NSP cells as compared to no expression in MCF7-SP cells, which suggests that the Wnt pathway is expressed in the MCF7-SP cells. The mesenchymal marker Vimentin was expressed only in MDA-MB-231 cells. Hence, understanding the breast cancer heterogeneity would enable a better understanding of the disease progression and therapeutic targeting.

Keywords: cancer stem cells, epithelial to mesenchymal transition, biomarkers, breast cancer

Procedia PDF Downloads 491

Authors: Shih-Ping Liu, Cheng-Hsuan Chang, Yu-Chuen Huang, Shih-Yin Chen, Woei-Cherng Shyu

Abstract:

Embryonic stem cells (ES) and induced pluripotnet stem cells (iPS) are both pluripotent stem cells. For mouse stem cells culture technology, leukemia inhibitory factor (LIF) was used to maintain the pluripotency of stem cells in vitro. However, LIF is an expensive reagent. The goal of this study was to find out a pure compound extracted from Chinese herbal medicine that could maintain stem cells pluripotency to replace LIF and improve the iPS generation efficiency. From 20 candidates traditional Chinese medicine we found that Cordycepsmilitaris triggered the up-regulation of stem cells activating genes (Oct4 and Sox2) expression levels in MEF cells. Cordycepin, a major active component of Cordycepsmilitaris, also could up-regulate Oct4 and Sox2 gene expression. Furthermore, we used ES and iPS cells and treated them with different concentrations of Cordycepin (replaced LIF in the culture medium) to test whether it was useful to maintain the pluripotency. The results showed higher expression levels of several stem cells markers in 10 μM Cordycepin-treated ES and iPS cells compared to controls that did not contain LIF, including alkaline phosphatase, SSEA1, and Nanog. Embryonic body formation and differentiation confirmed that 10 μM Cordycepin-containing medium was capable to maintain stem cells pluripotency after four times passages. For mechanism analysis, microarray analysis indicated extracellular matrix and Jak/Stat signaling pathway as the top two deregulated pathways. In ECM pathway, we determined that the integrin αVβ5 expression levels and phosphorylated Src levels increased after Cordycepin treatment. In addition, the phosphorylated Jak2 and phosphorylated Sat3 protein levels were increased after Cordycepin treatment and suppressed with the Jak2 inhibitor, AG490. The expression of cytokines associated with Jak2/Stat3 signaling pathway were also up-regulated by Q-PCR and ELISA assay. Lastly, we used Oct4-GFP MEF cells to test iPS generation efficiency following Cordycepin treatment. We observed that 10 Μm Cordycepin significantly increased the iPS generation efficiency in day 21. In conclusion, we demonstrated Cordycepin could maintain the pluripotency of stem cells through both of ECM and Jak2/Stat3 signaling pathway and improved iPS generation efficiency.

Keywords: cordycepin, iPS cells, Jak2/Stat3 signaling pathway, molecular biology

Procedia PDF Downloads 406

Authors: Urvi Panwar, Kanchan Mishra, Kanjaksha Ghosh, ShankerLal Kothari

Abstract:

Background: Day-by-day the increasing prevalence of neurodegenerative diseases have become a global issue to manage them by medical sciences. The adult neural stem cells are rare and require an invasive and painful procedure to obtain it from central nervous system. Mesenchymal stem cell (MSCs) therapies have shown remarkable application in treatment of various cell injuries and cell loss. MSCs can be derived from various sources like adult tissues, human bone marrow, umbilical cord blood and cord tissue. MSCs have similar proliferation and differentiation capability, but the human umbilical cord-derived mesenchymal stem cells (hUCMSCs) are proved to be more beneficial with respect to cell procurement, differentiation to other cells, preservation, and transplantation. Material and method: Human umbilical cord is easily obtainable and non-controversial comparative to bone marrow and other adult tissues. The umbilical cord can be collected after delivery of baby, and its tissue can be cultured using explant culture method. Cell culture medium such as DMEMF12+10% FBS and DMEMF12+Neural growth factors (bFGF, human noggin, B27) with antibiotics (Streptomycin/Gentamycin) were used to culture and differentiate mesenchymal stem cells into neural cells, respectively. The characterisations of MSCs were done with Flow Cytometer for surface markers CD90, CD73 and CD105 and colony forming unit assay. The differentiated various neural cells will be characterised by fluorescence markers for neurons, astrocytes, and oligodendrocytes; quantitative PCR for genes Nestin and NeuroD1 and Western blotting technique for gap43 protein. Result and discussion: The high quality and number of MSCs were isolated from human umbilical cord via explant culture method. The obtained MSCs were differentiated into neural cells like neurons, astrocytes and oligodendrocytes. The differentiated neural cells can be used to treat neural injuries and neural cell loss by delivering cells by non-invasive administration via cerebrospinal fluid (CSF) or blood. Moreover, the MSCs can also be directly delivered to different injured sites where they differentiate into neural cells. Therefore, human umbilical cord is demonstrated to be an inexpensive and easily available source for MSCs. Moreover, the hUCMSCs can be a potential source for neural cell therapies and neural cell regeneration for neural cell injuries and neural cell loss. This new way of research will be helpful to treat and manage neural cell damages and neurodegenerative diseases like Alzheimer and Parkinson. Still the study has a long way to go but it is a promising approach for many neural disorders for which at present no satisfactory management is available.

Keywords: bone marrow, cell therapy, explant culture method, flow cytometer, human umbilical cord, mesenchymal stem cells, neurodegenerative diseases, neuroprotective, regeneration

Procedia PDF Downloads 179

Authors: Brian M. Mehling, Louis Quartararo, Marine Manvelyan, Paul Wang, Dong-Cheng Wu

Abstract:

Numerous investigations suggest that Mesenchymal Stem Cells (MSCs) in general represent a valuable tool for therapy of symptoms related to chronic inflammatory diseases. Blue Horizon Stem Cell Therapy Program is a leading provider of adult and children’s stem cell therapies. Uniquely we have safely and efficiently treated more than 600 patients with documenting each procedure. The purpose of our study is primarily to monitor the immune response in order to validate the safety of intravenous infusion of human umbilical cord blood derived MSCs (UC-MSCs), and secondly, to evaluate effects on biomarkers associated with chronic inflammation. Nine patients were treated for conditions associated with chronic inflammation and for the purpose of anti-aging. They have been given one intravenous infusion of UC-MSCs. Our study of blood test markers of 9 patients with chronic inflammation before and within three months after MSCs treatment demonstrates that there is no significant changes and MSCs treatment was safe for the patients. Analysis of different indicators of chronic inflammation and aging included in initial, 24-hours, two weeks and three months protocols showed that stem cell treatment was safe for the patients; there were no adverse reactions. Moreover data from follow up protocols demonstrates significant improvement in energy level, hair, nails growth and skin conditions. Intravenously administered UC-MSCs were safe and effective in the improvement of symptoms related to chronic inflammation. Further close monitoring and inclusion of more patients are necessary to fully characterize the advantages of UC-MSCs application in treatment of symptoms related to chronic inflammation.

Keywords: chronic inflammatory diseases, intravenous infusion, stem cell therapy, umbilical cord blood derived mesenchymal stem cells (UC-MSCs)

Procedia PDF Downloads 407

Authors: Chuan Yin

Abstract:

Cancer stem cells (CSCs) represent a subpopulation of cancer cells that possess the characteristics associated with normal stem cells. CD133 is a phenotype of melanoma CSCs responsible for melanoma metastasis and drug resistance. Although adriamycin (ADR) is commonly used drug in melanoma therapy, but it is ineffective in the treatment of melanoma CSCs. In this study, we constructed CD133 antibody conjugated ADR immunoliposomes (ADR-Lip-CD133) to target CD133+ melanoma CSCs. The results showed that the immunoliposomes possessed a small particle size (~150 nm), high drug encapsulation efficiency (~90%). After 72 hr treatment on the WM266-4 melanoma tumorspheres, the IC50 values of the drug formulated in ADR-Lip-CD133, ADR-Lip (ADR liposomes) and ADR are found to be 24.42, 57.13 and 59.98 ng/ml respectively, suggesting that ADR-Lip-CD133 was more effective than ADR-Lip and ADR. Significantly, ADR-Lip-CD133 could almost completely abolish the tumorigenic ability of WM266-4 tumorspheres in vivo, and showed the best therapeutic effect in WM266-4 melanoma xenograft mice. It is noteworthy that ADR-Lip-CD133 could selectively kill CD133+ melanoma CSCs of WM266-4 cells both in vitro and in vivo. ADR-Lip-CD133 represent a potential approach in targeting and killing CD133+ melanoma CSCs.

Keywords: cancer stem cells, melanoma, immunoliposomes, CD133

Procedia PDF Downloads 357

Authors: Mohammad Afzal Khan, Fatimah Alanazi, Hala Abdalrahman Ahmed, Talal Shamma, Kilian Kelly, Mohammed A. Hammad, Abdullah O. Alawad, Abdullah Mohammed Assiri, Dieter Clemens Broering

Abstract:

Lung transplantation is a life-saving surgical replacement of diseased lungs in patients with end-stage respiratory malfunctions. Despite the remarkable short-term recovery, long-term lung survival continues to face several significant challenges, including chronic rejection and severe toxic side-effects due to global immunosuppression. Stem cell-based immunotherapy has been recognized as a crucial immunoregulatory regimen in various preclinical and clinical studies. Despite initial therapeutic outcomes, conventional stem cells face key limitations. The Cymerus™ manufacturing facilitates the production of a virtually limitless supply of consistent human induced pluripotent stem cell (iPSC)-derived mesenchymal stem cells, which could play a key role in selective immunosuppression and graft repair during rejection. Here, we demonstrated the impact of iPSC-derived human MSCs on the development of immune-tolerance and long-term graft survival in mouse orthotopic airway allografts. BALB/c→C57BL/6 allografts were reconstituted with iPSC-derived MSCs (2 million/transplant/ at d0), and allografts were examined for regulatory T cells (Tregs), oxygenation, microvascular blood flow, airway epithelium and collagen deposition during rejection. We demonstrated that iPSC-derived MSC treatment leads to significant increase in tissue expression of hTSG-6 protein, followed by an upregulation of mouse Tregs and IL-5, IL-10, IL-15 cytokines, which augments graft microvascular blood flow and oxygenation, and thereby maintained a healthy airway epithelium and prevented the subepithelial deposition of collagen at d90 post-transplantation. Collectively, these data confirmed that iPSC-derived MSC-mediated immunosuppression has potential to establish immune-tolerance and rescue allograft from sustained hypoxic/ischemic phase and subsequently limits long-term airway epithelial injury and collagen progression, which therapeutically warrant a study of Cymerus iPSC-derived MSCs as a potential management option for immunosuppression in transplant recipients.

Keywords: stem cell therapy, immunotolerance, regulatory T cells, hypoxia and ischemia, microvasculature

Procedia PDF Downloads 133

Authors: Melanie Macgregor-Ramiasa, Isabel Hopp, Patricia Murray, Krasimir Vasilev

Abstract:

Stem cells based therapies are one of the greatest promises of new-age medicine due to their potential to help curing most dreaded conditions such as cancer, diabetes and even auto-immune disease. However, establishing suitable in vitro culture materials allowing to control the fate of stem cells remain a challenge. Amongst the factor influencing stem cell behavior, substrate chemistry and nanotopogaphy are particularly critical. In this work, we used plasma assisted surface modification methods to produce model substrates with tailored nanotopography and controlled chemistry. Three different sizes of gold nanoparticles were bound to amine rich plasma polymer layers to produce homogeneous and gradient surface nanotopographies. The outer chemistry of the substrate was kept constant for all substrates by depositing a thin layer of our patented biocompatible polyoxazoline plasma polymer on top of the nanofeatures. For the first time, protein adsorption and stem cell behaviour (mouse kidney stem cells and mesenchymal stem cells) were evaluated on nanorough plasma deposited polyoxazoline thin films. Compared to other nitrogen rich coatings, polyoxazoline plasma polymer supports the covalent binding of proteins. Moderate surface nanoroughness, in both size and density, triggers cell proliferation. In association with polyoxazoline coating, cell proliferation is further enhanced on nanorough substrates. Results are discussed in term of substrates wetting properties. These findings provide valuable insights on the mechanisms governing the interactions between stem cells and their growth support.

Keywords: nanotopography, stem cells, differentiation, plasma polymer, oxazoline, gold nanoparticles

Procedia PDF Downloads 250

Authors: Khaled A. shady, Fagr B. Bazeed, Nashwa K. Abousamra, Ihab H. Elberawe, Ashraf E. shaalan, Mohamed A. Sobh

Abstract:

Physical activity activates a variety of adult stem cells which might be released into the circulation or might be activated in their organ-resident state. A variety of stimuli such as metabolic, mechanical, and hormonal stimuli might by responsible for the mobilization. This study was done to know the changes in hematopoietic stem cells and endothelial progenitor in athletes in the 24 hours following 30 min of aerobic exercise. Methods: Ten healthy male's athlete's (age 20.7± 0.61 y) performed moderate running with 30 min at 80% of velocity of The IAT. Blood samples taken pre-, and immediately, 30 min, 2h, 6h and 24h post-exercise were analyzed for hematopoietic stem cells (HSCs ), endothelial progenitor cells (EPCs(, vascular endothelial growth factor (VEGF), nitric oxide (NO), lactic acid (LA), and white blood cells . HSCs and EPCs were quantified by flow cytometry. Results: After 30min of aerobic exercise significant increases in HSCs, EPC, VEGF, NO, LA and WBCs (p ˂ 0.05). This increase will be at different rates according to the timing of taking blood sample and was in the maximum rate of increase after 30 min of aerobic exercise. HSCs, EPC, NO and WBCs were in the maximum rate of increase 2h post exercise. In addition, VEGF was in the maximum rate of increase immediately post exercise and LA concentration not affected after exercise. Conclusion: These data suggest that HSCs and EPCs increased after aerobic exercise due to increase of VEGF which play an important role in mobilization of stem cells and promotes NO increase which contributes to increase EPCs.

Keywords: physical activity, hematopoietic stem cells, mobilization, athletes

Procedia PDF Downloads 68

Authors: S. A. Hashemvarzi, A. Heidarianpour, Z. Fallahmohammadi, M. Pourghasem, M. Kaviani

Abstract:

Introduction: Parkinson’s disease (PD) is a progressive neurodegenerative in the central nervous system characterized by the loss of dopaminergic neurons in the substantia nigra resulting in loss of dopamine release in the striatum. Non-drug treatment options such as Stem cell transplantation and exercise have been considered for treatment of Parkinson's disease. Purpose: The purpose of this study was to evaluate the effect of aerobic exercise after bone marrow stem cells transplantation on recovery of dopaminergic neurons and promotion of angiogenesis markers in the striatum of parkinsonian rats. Materials and Methods: 42 male Wistar rats were divided randomly into six groups: Normal (N), Sham (S), Parkinson’s (P), Stem cells transplanted Parkinson’s (SP), Exercised Parkinson’s (EP) and Stem cells transplanted + Exercised Parkinson’s (SEP). To create a model of Parkinson's, the striatum was destroyed by injection of 6-hydroxy-dopamine into the striatum through stereotaxic apparatus. Stem cells were derived from the bone marrow of femur and tibia of male rats with 6-8 weeks old. After cultivation, approximately 5×105 cells in 5 microliter of medium were injected into the striatum of rats through the channel. Aerobic exercise was included 8 weeks of running on the treadmill with a speed of 15 meters per minute. At the end, all subjects were decapitated and striatum tissues were separately isolated for measurement of vascular endothelial growth factor (VEGF), dopamine (DA) and tyrosine hydroxylase (TH) levels. Results: VEGF, DA and TH levels in the striatum of parkinsonian rats significantly increased in treatment groups (SP, EP and SEP), especially in SEP group compared to P group after treatment (P<0.05). Conclusion: The findings implicate that the BMSCs transplantation in combination with exercise would have synergistic effects leading to functional recovery, dopaminergic neurons recovery and promotion of angiogenesis marker in the striatum of parkinsonian rats.

Keywords: stem cells, treadmill training, neurotrophic factors, Parkinson

Procedia PDF Downloads 321

Authors: Ahmed Gaber Abdel Raheem, Nashwa Ahmed Mohamed

Abstract:

Introduction: Sensorineural hearing loss (SNHL) is largely caused by the degeneration of the cochlea. Therapeutic options for SNHL are limited to hearing aids and cochlear implants. The cell transplantation approach to the regeneration of hair cells has gained considerable attention because stem cells are believed to accumulate in the damaged sites and have the potential for the repair of damaged tissues. The aim of the work: was to assess the use of bone marrow transplantation in repair of damaged inner ear hair cells in rats after the damage had been inflicted by Amikacin injection. Material and Methods: Thirty albino rats were used in this study. They were divided into three groups. Each group ten rats. Group I: used as control. Group II: Were given Amikacin- intratympanic injection till complete loss of hearing function. This could be assessed by Distortion product Otoacoustic Emission (DPOAEs) and / or auditory brain stem evoked potential (ABR). GroupIII: were given intra-peritoneal injection of bone marrow stem cell after complete loss of hearing caused by Amikacin. Clinical assessment was done using DPOAEs and / or auditory brain stem evoked potential (ABR), before and after bone marrow injection. Histological assessment of the inner ear was done by light and electron microscope. Also, Detection of stem cells in the inner ear by immunohistochemistry. Results: Histological examination of the specimens showed promising improvement in the structure of cochlea that may be responsible for the improvement of hearing function in rats detected by DPOAEs and / or ABR. Conclusion: Bone marrow stem cells transplantation might be useful for the treatment of SNHL.

Keywords: amikacin, hair cells, sensorineural hearing loss, stem cells

Procedia PDF Downloads 426

Authors: Prabhu Thangaraju, Manoj Deepak, A. Sivakumar

Abstract:

Removal of inter vertebral disc along with spinal fusion has many disadvantages such as causing stress fractures. If it is possible regenerate the spine it would be possible avoid the complications of the surgery and achieve better results. Our study involves the use of mesenchymal stem cells in regenerating the discs. Our study involved 10 patients who presented with degenerative disc disease between 2008-2011 in our hospital. After adequate pre-operative check prepared mesenchymal stem cells were injected into the disc spaces. These patients were subjected to conservative therapy for a minimum of six weeks before they were accepted into the study. They were followed up regularly for a minimum of 2years with serial radiographs and MRI. 8 out of the 10 patients had completed reduction in the pain. The T2 weighted MRI images in 9 out of the 10 patients showed a bright signal compared the previous Images which indicated that there was improvement in the hydration levels. From the case study of 10 patients who were subjected to mesenchymal cell therapy in our hospital, we can conclude that the use of mesenchymal cells in treatment of intervertebral disc degeneration in a safe and effective option.

Keywords: mesenchymal stem cells, intervertebral disc, the spine, disc degeneration

Procedia PDF Downloads 347

Authors: H. M. Imam, H. M. Rezk, A. F. Tohamy

Abstract:

Background: Human umbilical cord blood (HUCB) is considered as a unique source for stem cells. HUCB contain different types of progenitor cells which could differentiate into hepatocytes. Aims: To investigate the potential of rat's liver damage repair using human umbilical cord mesenchymal stem cells (hUCMSCs). We investigated the feasibility for hUCMSCs in recovery from liver damage. Moreover, investigating fibrotic liver repair and using the CCl4-induced model for liver damage in the rat. Methods: Rats were injected with 0.5 ml/kg CCl4 to induce liver damage and progressive liver fibrosis. hUCMSCs were injected into the rats through the tail vein; Stem cells were transplanted at a dose of 1×106 cells/rat after 72 hours of CCl4 injection without receiving any immunosuppressant. After (6 and 8 weeks) of transplantation, blood samples were collected to assess liver functions (ALT, AST, GGT and ALB) and level of Procollagen III as a liver fibrosis marker. In addition, hepatic tissue regeneration was assessed histopathologically and immunohistochemically using antihuman monoclonal antibodies against CD34, CK19 and albumin. Results: Biochemical and histopathological analysis showed significantly increased recovery from liver damage in the transplanted group. In addition, HUCB stem cells transdifferentiated into functional hepatocytes in rats with hepatic injury which results in improving liver structure and function. Conclusion: Our findings suggest that transplantation of hUCMSCs may be a novel therapeutic approach for treating liver fibrosis. Therefore, hUCMSCs are a potential option for treatment of liver cirrhosis.

Keywords: carbon tetra chloride, liver fibrosis, mesenchymal stem cells, rat

Procedia PDF Downloads 314

Authors: Remi Safi, Aline Hamade, Najat Bteich, Jamal El Saghir, Mona Diab Assaf, Marwan El-Sabban, Fadia Najjar

Abstract:

Estrogen is considered a risk factor for breast cancer since it promotes breast-cell proliferation. The jaesckeanadiol-3-p-hydroxyphenylpropanoate, a hemi-synthetic analogue of the natural phytoestrogen ferutinin (jaesckeanadiol-p-hydroxybenzoate), is designed to be devoid of estrogenic activity. This analogue induces a cytotoxic effect 30 times higher than that of ferutinin towards MCF-7 breast cancer cell line. We compared these two compounds with respect to their effect on proliferation, cell cycle distribution and cancer stem-like cells in the MCF-7 cell line. Treatment with ferutinin (30 μM) and its analogue (1 μM) produced a significant accumulation of cells at the pre G0/G1 cell cycle phase and triggered apoptosis. Importantly, this compound retains its anti-proliferative activity against breast cancer stem/progenitor cells that are naturally insensitive to ferutinin at the same dose. These results position ferutinin analogue as an effective compound inhibiting the proliferation of estrogen-dependent breast cancer cells and consistently targeting their stem-like cells.

Keywords: ferutinin, hemi-synthetic analogue, breast cancer, estrogen, stem/progenitor cells

Procedia PDF Downloads 162

Authors: Abdolamir Allameh, Shahnaz Esmaeili, Mina Allameh, Safoura Khajeniazi

Abstract:

Stem cells with therapeutic applications can be isolated from human placenta/umblical cord blood (UCB) as well as the cord tissue (UC). Stem cells in culture are vulnerable to oxidative stress, particularly when subjected to differentiation process. The aim of this study was to examine the chnages in the rate of oxidation that occurs to cellular macromolecules during hepatic differentiation of mononuclear cells (MSCs). In addition, the impact of the hepatic differentiation process of MSC on cellular and biological activity of the cells will be undertaken. For this purpose, first mononuclear cells (MNCs) were isolated from human UCB which was obtained from a healthy full-term infant. The cells were cultured at a density of 3×10⁵ cells/cm² in DMEM- low-glucose culture media supplemented with 20% FBS, 2 mM L-glutamine, 100 μg/ml streptomycin and 100 U/ml penicillin. Cell cultures were then incubated at 37°C in a humidified 5% CO₂ incubator. After removing non-adherent cells by replacing culture medium, fibroblast-like adherent cells were resuspended in 0.25% trypsin-EDTA and plated in 25 cm² flasks (1×10⁴/ml). Characterization of the MSCs was routinely done by observing their morphology and growth curve. MSCs were subjected to a 2-step hepatocyte differentiation protocol in presence of hepatocyte growth factor (HGF), dexamethazone (DEX) and oncostatin M (OSM). The hepatocyte-like cells derived from MSCs were checked every week for 3 weeks for changes in lipid peroxidation, protein carbonyl formation and DNA oxidation i.e., 8-hydroxy-2'-deoxyguanosine (8-OH-dG) assay. During the 3-week differentiation process of MSCs to hepatocyte-like cells we found that expression liver-specific markers such as albumin, was associated with increased levels of lipid peroxidation and protein carbonyl formation. Whereas, undifferentiated MSCs has relatively low levels of lipid peroxidation products. There was a significant increase ( p < 0.05) in lipid peroxidation products in hepatocytes on days 7, 14, and 21 of differentiation. Likewise, the level of protein carbonyls in the cells was elevated during the differentiation. The level of protein carbonyls measured in hepatocyte-like cells obtained 3 weeks after differentiation induction was estimated to be ~6 fold higher compared to cells recovered on day 7 of differentiation. On the contrary, there was a small but significant decrease in DNA damage marker (8-OH-dG) in hepatocytes recovered 3 weeks after differentiation onset. The level of 8-OHdG which was in consistent with formation of reactive oxygen species (ROS). In conclusion, this data suggest that despite the elevation in oxidation of lipid and protein molecules during hepatocyte development, the cells were normal in terms of DNA integrity, morphology, and biologically activity.

Keywords: adult stem cells, DNA integrity, free radicals, hepatic differentiation

Procedia PDF Downloads 129

Authors: Manali Jain, Neeta Singh, Raunaq Fatima, Soniya Nityanand, Mandakini Pradhan, Chandra Prakash Chaturvedi

Abstract:

Isolated Congenital Heart Defect (ICHD) is the major cause of neonatal death worldwide among all forms of CHDs. A significant proportion of fetuses with ICHD die in the neonatal period if no treatment is provided. Recently, stem cell therapies have emerged as a potential approach to ameliorate ICHD in children. ICHD is characterized by cardiac structural abnormalities during embryogenesis due to alterations in the cardiomyogenic properties of a pool of cardiac progenitors/ stem cells associated with fetal heart development. The stem cells present in the amniotic fluid (AF) are of fetal origin and may reflect the physiological and pathological changes in the fetus during embryogenesis. Therefore, in the present study, the cardiomyogenic potential of AF-MSCs derived from fetuses with ICHD (ICHD AF-MSCs) has been evaluated and compared with that of AF-MSCs of structurally normal fetuses (normal AF-MSCs). Normal and ICHD AF-MSC were analyzed for the expression of cardiac progenitor markers viz., stage-specific embryonic antigen-1 (SSEA-1), vascular endothelial growth factor 2 (VEGFR-2) and platelet-derived growth factor receptor-alpha (PDGFR-α) by flow cytometry. The immunophenotypic characterization revealed that ICHD AF-MSCs have significantly lower expression of cardiac progenitor markers VEGFR-2 (0.14% ± 0.6 vs.48.80% ± 0.9; p <0.01), SSEA-1 (70.86% ± 2.4 vs. 88.36% ±2.7; p <0.01), and PDGFR-α (3.92% ± 1.8 vs. 47.59% ± 3.09; p <0.01) in comparison to normal AF-MSCs. Upon induction with 5’-azacytidine for 21 days, ICHD AF-MSCs showed a significantly down-regulated expression of cardiac transcription factors such as GATA-4 (0.4 ± 0.1 vs. 6.8 ± 1.2; p<0.01), ISL-1 (2.3± 0.6 vs. 14.3 ± 1.12; p<0.01), NK-x 2-5 (1.1 ± 0.3 vs. 14.1 ±2.8; p<0.01), TBX-5 (0.4 ± 0.07 vs. 4.4 ± 0.3; p<0.001), and TBX-18 (1.3 ± 0.2 vs. 4.19 ± 0.3; p<0.01) when compared with the normal AF-MSCs. Furthermore, immunocytochemical staining revealed that both types of AF-MSCs could differentiate into cardiovascular lineages and express cardiomyogenic, endothelial, and smooth muscle actin markers, viz., cardiac troponin (cTNT), CD31, and alpha-smooth muscle actin (α-SMA). However, normal AF-MSCs showed an enhanced expression of cTNT (p<0.001), CD31 (p<0.01), and α-SMA (p<0.05), compared to ICHD AF-MSCs. Overall, these results suggest that the ICHD-AF-MSCs have a defective cardiomyogenic differentiation potential and that the defects in these stem cells may have a role in the pathogenesis of ICHD.

Keywords: amniotic fluid, cardiomyogenic potential, isolated congenital heart defect, mesenchymal stem cells

Procedia PDF Downloads 77

Authors: Kenichi Yamahara, Makiko Ohshima, Shunsuke Ohnishi, Hidetoshi Tsuda, Akihiko Taguchi, Toshihiro Soma, Hiroyasu Ogawa, Jun Yoshimatsu, Tomoaki Ikeda

Abstract:

We have previously reported the therapeutic potential of rat fetal membrane(FM)-derived mesenchymal stem cells (MSCs) using various rat models including hindlimb ischemia, autoimmune myocarditis, glomerulonephritis, renal ischemia-reperfusion injury, and myocardial infarction. In this study, 1) we isolated and characterized MSCs from human amnion and chorion; 2) we examined their differences in the expression profile of growth factors and cytokines; and 3) we investigated the therapeutic potential and difference of these MSCs using murine hindlimb ischemia and acute graft-versus-host disease (GVHD) models. Isolated MSCs from both amnion and chorion layers of FM showed similar morphological appearance, multipotency, and cell-surface antigen expression. Conditioned media obtained from amnion- and chorion-derived MSCs inhibited cell death caused by serum starvation or hypoxia in endothelial cells and cardiomyocytes. Amnion and chorion MSCs secreted significant amounts of angiogenic factors including HGF, IGF-1, VEGF, and bFGF, although differences in the cellular expression profile of these soluble factors were observed. Transplantation of human amnion or chorion MSCs significantly increased blood flow and capillary density in a murine hindlimb ischemia model. In addition, compared to human chorion MSCs, human amnion MSCs markedly reduced T-lymphocyte proliferation with the enhanced secretion of PGE2, and improved the pathological situation of a mouse model of GVHD disease. Our results highlight that human amnionand chorion-derived MSCs, which showed differences in their soluble factor secretion and angiogenic/immuno-suppressive function, could be ideal cell sources for regenerative medicine.

Keywords: amnion, chorion, fetal membrane, mesenchymal stem cells

Procedia PDF Downloads 395