scholarly journals Platelet-Derived Microparticles Increase Expression of hTERT in Umbilical Cord Mesenchymal Stem Cells

2018 ◽  
Vol 5 (4) ◽  
pp. 31 ◽  
Author(s):  
Maryam Samareh Salavati Pour ◽  
Fatemeh Hoseinpour Kasgari ◽  
Alireza Farsinejad ◽  
Ahmad Fatemi ◽  
Roohollah Mirzaee Khalilabadi
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Life Span ◽  
Real Time ◽  
Umbilical Cord ◽  
Control Group ◽  
Surface Markers

Introduction: Mesenchymal stem cells (MSCs) are widely studied due to their self- renewal potential and capacity to differentiate into multiple tissues. However, they have a limited life span of several divisions in vitro, which alters various cellular characteristics and reduces their application. Aim: We evaluated the effect of platelet-derived microparticles on gene expression of hTERT, one of the main factors involved in aging and cell longevity. Materials and methods: Umbilical cord MSCs were used for this study. Cells were characterized by evaluating morphology via inverted microscope and identifying associated surface markers using flow cytometry. Platelet-derived microparticles were prepared by centrifuging platelet bags at varying speeds, and their concen- trations were determined by Bradford assay. At 30% confluency, MSCs were treated with 50 μg/mL of microparticles for five days. Then, RNA was extracted and cDNA was synthesized. Quantitative expression of hTERT was assessed using real-time polymerase chain reaction (PCR). Results: Fibroblast-like cells were isolated from umbilical cord tissue and MSCs were identified by the presence of mesenchymal surface markers via flow cytometry. Real- time PCR showed that gene expression of hTERT increased by more than three times when treated with platelet-derived microparticles, in comparison to expression of the control group. Conclusion: We concluded that platelet-derived microparticles may be a potentially safe and effective method to increase hTERT gene expression in MSCs, ultimately prolonging their life span in vitro. 

scholarly journals Programming of human umbilical cord mesenchymal stem cells in vitro to promote pancreatic gene expression

2013 ◽  
Vol 8 (3) ◽  
pp. 769-774 ◽  
Author(s):  
HONGWU WANG ◽  
YUPING YANG ◽  
GUYU HO ◽  
XIAOBO LIN ◽  
WEIZHAO WU ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Umbilical Cord ◽  
Human Umbilical Cord

scholarly journals Platelet Microparticles Accelerate Proliferation and Growth of Mesenchymal Stem Cells through Longevity-Related Genes

2021 ◽  
Vol 24 (8) ◽  
pp. 607-614
Author(s):  
Maryam Samareh Salavati Pour ◽  
Fatemeh Hoseinpour Kasgari ◽  
Alireza Farsinejad ◽  
Ahmad Fatemi ◽  
Gholamhossein Hassanshahi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Real Time ◽  
Real Time Pcr ◽  
Treated Group ◽  
Population Doubling ◽  
Control Group ◽  
Population Doubling Time ◽  
Longevity Genes

Background: Due to their self-renewal and differentiation ability, the mesenchymal stem cells (MSCs) have been studied extensively. However, the MSCs lifespan is restricted; they undergo several divisions in vitro that cause several alternations in cellular features and relatively lessens their application. Thus, this study was aimed to assess the effect of platelet-derived microparticles (PMPs), a valuable source of proteins, microRNAs (miRNAs), and growth factors, on the expression of hTERT, c-MYC, p16, p53, and p21 as the most important aging and cell longevity genes alongside with population doubling time (PDT) of PMP-treated cells in comparison to a control group. Methods: Umbilical cord MSCs (UC-MSCs) were used in this study, whereby they reached a confluency of 30%. MSCs were treated by PMPs (50 µg/mL), and then, PDT was determined for both groups. Quantitative expression of hTERT, c-MYC, p16, p53, and p21 was examined through quantitative real-time PCR at various intervals (i.e. after five and thirty days as well as freezing-thawing process). Results: Our results demonstrated that the treated group had a shorter PDT in comparison to the control group (P<0.050). The real-Time PCR data also indicated that PMPs were able to remarkably up-regulate hTERT and c-MYC genes expression while down-regulating the expression of p16, p21, and p53 genes (P<0.050), especially following five days of treatment. Conclusion: According to these data, it appears that PMPs are a safe and effective candidate for prolonging the lifespan of UC-MSCs; however, further investigations are needed to corroborate this finding.

37 Healthy Foals Produced Using Bone Marrow-Mesenchymal Stem Cells as Nuclear Donors in Horse Cloning

2018 ◽  
Vol 30 (1) ◽  
pp. 158
Author(s):  
R. Olivera ◽  
L. Moro ◽  
R. Jordan ◽  
C. Luzzani ◽  
S. Miriuka ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Umbilical Cord ◽  
Donor Cell ◽  
Control Group ◽  
Nuclear Reprogramming ◽  
Marrow Mesenchymal Stem Cells

Somatic cell nuclear transfer efficiency is based on the capacity of the donor cell to be reset and reprogrammed to an embryonic state. So, the less differentiated the donor cells are, the more easily they could be reprogrammed by a recipient cytoplasm. Failures on appropriate nuclear reprogramming frequently lead to abnormalities associated with the placenta, umbilical cord, birthweight, and limbs. In the present study, we evaluated the efficiency of bone marrow mesenchymal stem cells (BM-MSC) compared with adult fibroblasts (AF) as nuclear donors in horse cloning and evaluated both in vitro and in vivo development of the embryos generated. Moreover, we focused on comparing the health of the foals generated and on the presence of anatomical abnormalities in foals produced from the different treatments. Embryos produced by AI, recovered by uterine flushing, and transferred to recipient mares were used as controls. All variables were analysed by Fisher test (P < 0.05). The cloning procedure was performed according to Olivera et al. (2016 PLoS One 11, e0164049, 10.1371/journal.pone.0164049). Both cleavage and blastocyst rates were higher when MSC were used as nuclear donors (P < 0.05). Cleavage rates were 85.6% (3875/4527) v. 90.2% (3095/3432) and blastocyst rates were 10.9% (492/4527) and 18.1% (622/3432) for AF and MSC groups, respectively. In the AF group, 476 blastocysts were transferred to recipient mares (232 transfers), and in the MSC group, 594 blastocysts were transferred 297 transfers). In the AI control group, 88 embryos were transferred. Pregnancies were diagnosed by transrectal ultrasonography 15 days after embryo transfer in all the groups. Pregnancy rates were similar between both cloning groups (41/232, 17.7% and 37/297, 12.5%for AF and MSC, respectively), but higher in the AI group (71/88, 80.7%). However, significant differences were observed in the birth of viable offsprings among the cloning groups. Despite similar rates of foal delivery (AF, 17/41, 41.5%; MSC, 21/37, 56.7%), a higher proportion of viable foals were obtained from the MSC group (20/37, 54.1%) compared with the AF group (9/41, 22%; P < 0.05). Surprisingly, as in the AI group (63/63, 100%), all of the viable foals obtained using MSC (20/20, 100%) were considered normal and did not show abnormalities associated with cloning. In contrast, in the AF group, only 4/9 (44.4%) were considered normal foals. The defects present in the other 5 foals were related to flexural and angular limb deformities and umbilical cord malformations. These were corrected rapidly with standard treatments or, in the case of the umbilical cords, minor surgery. This study shows for the first time that BM-MSC can be used as nuclear donors in horse cloning and that the foals obtained are as healthy as those produced by AI, showing no abnormalities related to deficiencies in nuclear reprogramming.

scholarly journals The Effects of Indoxyl Sulfate on Human Umbilical Cord-Derived Mesenchymal Stem Cells In Vitro

2016 ◽  
Vol 38 (1) ◽  
pp. 401-414 ◽  
Author(s):  
Wei Wang ◽  
Xueyong Liu ◽  
Wei Wang ◽  
Jinghua Li ◽  
Yuanyuan Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Umbilical Cord ◽  
Indoxyl Sulfate ◽  
Proliferative Potential ◽  
Human Umbilical Cord ◽  
Uremic Toxin ◽  
Ki 67

Background/Aims: Indoxyl sulfate, an important protein-bound uremic toxin, can damage stem cells, thus hampering stem cell-based regenerative medicine approaches targeting chronic kidney diseases (CKD). Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) are thought to have promising clinical application because of their high proliferative potential and ease of isolation than MSCs from other sources. In the present study, we aimed to determine the harmful effects of indoxyl sulfate on the phenotype and functional potential of hUC-MSCs in vitro. Methods: The toxicity and cell viability was examined by Trypan blue exclusion and MTT assay. The cellular surface markers and the percentage of apoptotic cells by Annexin-V/PI staining were analyzed by flow cytometry. Proliferation was evaluated based on cell number counting and Ki-67 immunostaining. Cell senescence was measured using senescence-associated β-Galactosidase activity. The ability to stimulate the development of CD4+CD25+FoxP3+ regulatory T cells was assessed by incubating hUC-MSCs with peripheral blood mononuclear cells from the healthy volunteers. Results: Our results demonstrated that the immunophenotype of hUC-MSCs was not affected by indoxyl sulfate flow cytometry. However, a significant decrease in cell numbers and fraction of Ki-67 positive proliferating cells, along with a significant increase in cellular senescence were detected in hUC-MSCs after exposure to indoxyl sulfate. Additionally, their ability to stimulate CD4+CD25+FoxP3+ regulatory T cell production was compromised when hUC-MSCs were pretreated with indoxyl sulfate. Conclusion: Taken together, our study clearly demonstrated that the molecular alterations and functional incompetence in hUC-MSCs under the challenge of indoxyl sulfate in vitro.

scholarly journals Characterization and plasticity of wharton's jelly mesenchymal stem cells of goat

2021 ◽  
Vol 37 ◽  
pp. e37002
Author(s):  
Gustavo Cardoso da Silva Neves ◽  
Napoleão Martins Argôlo Neto ◽  
Maíra Soares Ferraz ◽  
Clautina Ribeiro de Moraes Da Costa ◽  
Andressa Rêgo Da Rocha ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Umbilical Cord ◽  
Growth Curve ◽  
Invasive Procedure ◽  
Wharton’S Jelly ◽  
Wharton's Jelly ◽  
Undifferentiated State

Mesenchymal stem cells (MSCs), obtained from several anatomical sites, have already been described, characterized and used in therapeutic models for tissue repair. The umbilical cord mesenchymal stem cells, represented by cells from arteries and veins walls, as well as Wharton's jelly are easy to be obtained, highly available, require no invasive procedure, do not present risk to donors and do not present ethical limitation. The aim of this research was to analyze the plasticity of Wharton's jelly mesenchymal stem cells (WJ-MSCs) of goat, evaluating their behavior in vitro and characterizing them immunophenotypically. Thus, tests were performed on colony forming units, viability and cell growth curve, flow cytometry analysis and plasticity potential. Goat umbilical cord matrix cells exhibited fibroblastoid morphology with colony formation and self-renewal ability, always maintaining their undifferentiated state up to the eighth passage (P8). The growth curve kinetics exhibited the LAG, LOG, and DECAY phases, without displaying a PLATEAU phase. The plasticity assay demonstrated positive differentiation for osteogenic, adipogenic and chondrogenic lines, characterized by the synthesis of intracytoplasmic granules or extracellular matrix with the presence of calcium, lipids and proteoglycans. Flow cytometry demonstrated the expression of CD90 and CD105; absence of CD14 expression.  It is concluded that the cell population isolated from the Wharton's  jelly of goat constitutes a representative sample of mesenchymal stem cells, with great possibilities in the field of regenerative and reproductive medicine.

scholarly journals Evaluation of Kohlrabi (Brassica oleracea Var. Gongylodes) Extract Effect on Mesenchymal Stem Cells Viability and Apoptosis

2020 ◽  
Vol 8 (2) ◽  
pp. 93-102
Author(s):  
Naser Kalhor Qom ◽  
◽  
Mohsen Sheykhhasan ◽  
Ali Kowsari ◽  
◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Cell Viability ◽  
Brassica Oleracea ◽  
Real Time ◽  
Real Time Pcr ◽  
Surface Markers ◽  
Expression Levels ◽  
Significant Difference

Background: Cell viability and apoptosis are two crucial factors that may determine cell fate. There are several factors, such as hypoxia, which may be effective in cell processes. Because of its unique features, such as its antioxidant, anti-inflammatory, and anti-apoptosis mechanisms, kohlrabi (Brassica oleracea var. gongylodes) extract may be used in the amelioration of cell viability and a decrease in cell apoptosis. In this study, we evaluate the effect of kohlrabi extract on the viability and apoptosis of adipose-derived mesenchymal stem cells (AD-MSCs). Materials and Methods: In this study, extract from kohlrabi and mesenchymal stem cells from adipose tissue were isolated in a laboratory under sterile conditions. Expression of mesenchymal stem cell (MSC) surface markers, including CD44, CD90, and CD105 was evaluated by flow cytometry method. Besides, CD34 was used as a negative marker. MTT assay was carried out to determine the cell viability. Evaluation of BCL2 and BAX expression levels was performed by real-time PCR. Results: MSC surface markers were verified by flow cytometry. The obtained results demonstrated a significant difference between the cell viability of the kohlrabi-extract treated and control group over time (P=0.03). In addition, the real-time PCR analysis showed that expression levels of BCL2 significantly increased in hypoxic condition after treatment with leaf extract (P=0.019). However, there was no significant expression change in the BAX gene. Conclusion: Our study illustrates that kohlrabi extract may have positive effects on cell survival while having inhibitory effects on apoptosis.

scholarly journals Comparative in vitro study of the cytotoxicity of gelatine and alginate to human umbilical cord mesenchymal stem cells

2019 ◽  
Vol 52 (1) ◽  
pp. 36
Author(s):  
Nike Hendrijantini
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Umbilical Cord ◽  
Colorimetric Assay ◽  
In Vitro Study ◽  
Human Umbilical Cord ◽  
Significant Difference ◽  
Diphenyl Tetrazolium Bromide

Background: Mesenchymal stem cells (MSCs) and scaffold combination constitute a promising approach currently adopted for tissue engineering. Umbilical cord-derived mesenchymal stem cells (hUC-MSCs) are easily obtained and non-invasive. Gelatine and alginate constitute a biocompatible natural polymer scaffold. At present, a cytotoxicity comparison of gelatine and alginate to hUC-MSCs is not widely conducted Purpose: This study aimed to compare the cytotoxicity of gelatine and alginate in hUC-MSCs in vitro. Methods: Isolation and culture were performed on hUC-MSCs derived from healthy full-term neonates. Flow Cytometry CD90, CD105 and CD73 phenotype characterization was performed in passage 4. 3-(4,5-dimethythiazol- 2-yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay was performed to measure the cytotoxicity. The three sample groups were: (T1) hUC-MSCs with α-MEM (alpha-minimum essential medium) solution as control; (T2) hUC-MSCs with gelatine; (T3) hUC-MSCs with alginate Results: Flow cytometry of hUC-MSCs displayed positive CD90, CD105 and CD73 surface markers. Gelatine and alginate had no effect on the viability of hUC-MSCs and no statistically significant difference (p>0.05) of cytotoxicity between gelatine and alginate to hUC-MSCs. Conclusion: Gelatine and alginate proved to be non-toxic to hUC-MSCs in vitro.

scholarly journals Platelet Rich in Growth Factors (PRGF): A Suitable Replacement for Fetal Bovine Serum (FBS) in Mesenchymal Stem Cell Culture

2018 ◽  
Vol 5 (4) ◽  
pp. 12
Author(s):  
Fatemeh Hoseinpour Kasgari ◽  
Maryam Samareh Salavati Pour ◽  
Alireza Farsinejad ◽  
Ahmad Fatemi ◽  
Roohollah Mirzaee Khalilabadi
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Growth Factors ◽  
Real Time ◽  
Umbilical Cord ◽  
Real Time Pcr ◽  
Surface Markers ◽  
Differentiation Potential ◽  
Significant Difference ◽  
Tert Gene

 ntroduction: Due to high differentiation potential and self-renewality, Mesenchymal stem cells are now widely considered by researchers in several diseases. FBS is used as a supplement in culture media for proliferation, differentiation, and other culture processes of MSCs, which is associated with transmission risk of a variety of infections as well as immune responses. PRGF derived from platelets contains growth factors causing the growth and proliferation of MSCs. This study was conducted to compare the effect of PRGF in comparison to FBS on the expression of h-TERT gene, in umbilical cord-derived MSCs. Materials and Methods: This study is an experimental research. Four expired platelet concentrate bags were obtained from Kerman blood transfusion center, and PRGF was prepared by multiple centrifugation rounds of the platelet bag. Calcium chloride was added as an anticoagulant to PRGF in order to prevent gelatinization of the culture medium. On the other hand, mesenchymal stem cells were isolated from the umbilical cord as a primary culture. The phenotype of the cells was confirmed by flow cytometry, and the cells were randomly cultured as control (using FBS) and experimental (using PRGF) groups. The expression of the gene involved in increasing cell longevity (h-TERT) was investigated by real-time PCR technique after six days. Results: Mesenchymal stem cells were successfully isolated from the umbilical cord. Morphologically, the mesenchymal cells cultured in the experimental group (using PRGF) were similar to the cells in the control medium. The cells exhibited a high expression level of CD73, CD90, and CD105, while the surface markers of hematopoietic cells such as CD45 and CD34 were slightly expressed. Therefore, there was no significant difference in the expression of cell surface markers between control and experimental groups. In this study, using the real-time PCR technique, it was shown that PRGF derived from the platelet could increase the expression of h- TERT gene in the umbilical cord mesenchymal stem cells compared with the control. (P = 0.034)  

scholarly journals Study of Human Amniotic Membrane Mesenchymal Stem Cells Using Gelatin and Alginate as Nontoxic Scaffolds

2019 ◽  
Vol 5 ◽  
pp. 1
Author(s):  
Nike Hendrijantini ◽  
Poedjo Hartono ◽  
Helen Susilowati ◽  
Cindy K. Hartono ◽  
Reni P. Daniati ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Amniotic Membrane ◽  
Colorimetric Assay ◽  
Control Group ◽  
Human Amniotic Membrane ◽  
Average Percentage ◽  
Significant Difference

Perinatal mesenchymal stem cells (MSCs), for example, from amniotic membrane, have advantages over adult sources, such as bone marrow, in terms of ease of availability, cell naivety, tissue stem cell abundance, high capacity of proliferation, and less donor site morbidity, because it does not require invasive procedures. Natural polymer scaffolds, such as gelatin and alginate, are biocompatible. Combination of stem cells from amniotic membrane (hAMSCs) and gelatin or alginate as scaffold can be promising. However, cytotoxicity comparison of gelatin and alginate to hAMSCs has not been widely studied. This study was aimed to compare cytotoxicity of gelatin and alginate on hAMSCs in vitro. Isolation and culture were performed on hAMSCs of the healthy full-term pregnancy. In passage 4, Flow Cytometry CD90, CD105, and CD73 phenotype characterization was done. Colorimetric assay of 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) was performed to measure the cytotoxicity. There were three sample groups: (control group) hAMSCs with alpha-minimum essential medium (α-MEM) solution as control; (gelatin group) hAMSCs with gelatin; (alginate group) hAMSCs with alginate. Each group consisted of 12 samples. Flow cytometry of hAMSCs expressed 28.78% CD90, 36.95% CD105, and 44.41% CD73 surface markers. No sample depicted toxicity in either gelatin or alginate group, and this is indicated by the average percentage of living cells in gelatin 97.26% and in alginate 98.43%. No statistically significant difference (ρ=0.057) of cytotoxicity was found between gelatin and alginate to hAMSCs. Gelatin and alginate were nontoxic to hAMSCs in vitro.

scholarly journals Bone Regeneration by Nanohydroxyapatite/Chitosan/Poly(lactide-co-glycolide) Scaffolds Seeded with Human Umbilical Cord Mesenchymal Stem Cells in the Calvarial Defects of the Nude Mice

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Fei Wang ◽  
Xiao-Xia Su ◽  
Yu-Cheng Guo ◽  
Ang Li ◽  
Yin-Cheng Zhang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Bone Regeneration ◽  
Umbilical Cord ◽  
Nude Mice ◽  
Fluorescence Labeling ◽  
Control Group ◽  
Human Umbilical Cord ◽  
Calvarial Defects

In the preliminary study, we have found an excellent osteogenic property of nanohydroxyapatite/chitosan/poly(lactide-co-glycolide) (nHA/CS/PLGA) scaffolds seeded with human umbilical cord mesenchymal stem cells (hUCMSCs)in vitroand subcutaneously in the nude mice. The aim of this study was to further evaluate the osteogenic capacity of nHA/CS/PLGA scaffolds seeded with hUCMSCs in the calvarial defects of the nude mice. Totally 108 nude mice were included and divided into 6 groups: PLGA scaffolds + hUCMSCs; nHA/PLGA scaffolds + hUCMSCs; CS/PLGA scaffolds + hUCMSCs; nHA/CS/PLGA scaffolds + hUCMSCs; nHA/CS/PLGA scaffolds without seeding; the control group (no scaffolds) (n=18). The scaffolds were implanted into the calvarial defects of nude mice. The amount of new bones was evaluated by fluorescence labeling, H&E staining, and Van Gieson staining at 4 and 8 weeks, respectively. The results demonstrated that the amount of new bones was significantly increased in the group of nHA/CS/PLGA scaffolds seeded with hUCMSCs (p<0.01). On the basis of previous studiesin vitroand in subcutaneous implantation of the nude mice, the results revealed that the nHA and CS also enhanced the bone regeneration by nHA/CS/PLGA scaffolds seeded with hUCMSCs in the calvarial defects of the nude mice at early stage.

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