scholarly journals Life Span of the Chondrocytes from Human Umbilical Cord Derived-Mesenchymal Stem Cell Differentiation

2019 ◽  
Vol 12 (1) ◽  
pp. 65-70
Author(s):  
Ketut Dewi Kumara Wati ◽  
Endah Dhyanti Pratiwi ◽  
Yanni Dirgantara ◽  
Cynthia Retna Sartika ◽  
Meita Dhamayanti ◽  
...  
Keyword(s):  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Life Span ◽  
Mtt Assay ◽  
Basal Medium ◽  
Stem Cell Differentiation ◽  
Morphological Observation ◽  
Blue Staining ◽  
Alcian Blue Staining ◽  
Human Umbilical Cord

Problem in growth might need to be sought deeper inside the cells to enhance better management. There was a lack of study done in chondrocytes to determine growth problem mechanism. We sought to see the chondrocyte’s life span by observing the cells characteristic in a small container over long period. Human mesenchymal stem cell-derived MSC underwent chondrocyte differentiation procedure in a 96 wells-plate using chondrocyte/osteocyte differentiation medium followed by basal medium. A morphological observation and MTT assay was done to observe apoptosis in chondrocyte in the different wells. MTT assay absorbance was used to determine number of apoptosis. Statistical analysis was done to assess the different of mean absorbance values on different days of observation using Anova folowed by post hoc analysis in SPSS IBM package version 24 , with significance determined at P<0.05. Chondrocytes was confirmed with 1% Alcian blue staining and showed characteristic larger then its parent MSC. The MTT assay showed that chondrocyte absorbance reduce from day 3 through day 14, which was different significantly (anova, P=0.03) and day 14 was the most differed compared to day 7 and day 10 (P=0.02, P=0.017 and P=0.013, respectively). The 2D monolayer chondrocyte underwent total apoptosis on day 14 which can set the limit for observation in further long-term observational cell study.

scholarly journals LAA and CoCl2 Pretreated Exosomes Enhance Chondrogenic Differentiation of Mesenchymal Stem Cell

2021 ◽  
Author(s):  
Anggraini Barlian ◽  
Rizka Musdalifah Amsar ◽  
Salindri Prawitasari ◽  
Christofora Hanny Wijaya ◽  
Ika Dewi Ana ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Mesenchymal Stem Cell ◽  
Chondrogenic Differentiation ◽  
Stem Cell Differentiation ◽  
Human Umbilical Cord ◽  
Hypoxic Conditions ◽  
Transmission Electron

Abstract BackgroundCells produce extracellular vesicles, such as exosomes and microvesicles, which are used for intracellular communication. Cell-free therapies could be enhanced by using mesenchymal stem cell-derived exosomes. Preconditioning parental cells affects the properties of their exosomes. This study aimed to investigate the role of L-ascorbic acid (LAA) and CoCl2 in the exosomes produced by human Wharton’s jelly mesenchymal stem cells (hWJ MSC) and its potential to induce chondrogenic differentiation of stem cells was also studied.. MethodThe cells were obtained from umbilical cords and characterized based on mesenchymal stem cell criteria. The cells were cultured in a serum-free medium containing LAA and CoCl2. Exosomes produced by the cells were isolated and their morphology observed with Transmission Electron Microscopy. The presence of CD 63 was confirmed using ELISA. The particle size distribution and exosome concentration were analyzed with Nanoparticle Tracking Analysis (NTA). The ability of exosomes to induce stem cell differentiation into chondrocytes was investigated using the Alcian blue assay and immunocytochemistry.ResultsStem cells were successfully isolated from the human umbilical cord. The cells can differentiate into adipocytes, chondrocytes, and osteocytes. Flowcytometry analysis showed the specific surface marker of mesenchymal stem cells. Exosomes isolated from pretreatment cells showed round-shaped morphology and confirmed the presence of CD 63. NTA analysis revealed that pretreatment of cells with LAA increases exosome yields. LAA supplementation in cell medium under hypoxic conditions induced by CoCl2 produces exosomes that can induce the chondrogeic differentiation of stem cells, confirmed by the presence of glycosaminoglycan and collagen type 2.ConclusionExosomes produced by preconditioning hWJ-MSC with LAA in hypoxic conditions have the potential to enhance human Wharton Jelly stem cell differentiation into chondrocytes.

scholarly journals Role of Heparin Sodium Salt in the Modulation of Human Umbilical Cord-Derived Mesenchymal Stem Cell Differentiation

2019 ◽  
Vol 6 (4) ◽  
pp. 165-171
Author(s):  
Madhumala Gopinath ◽  
Suman Nandy ◽  
Ganesan Jothimani ◽  
Sarubala Malayaperumal ◽  
Birendra Kumar Mishra ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
Mesenchymal Stem Cell ◽  
Umbilical Cord ◽  
Sodium Salt ◽  
Stem Cell Differentiation ◽  
Human Umbilical Cord ◽  
Heparin Sodium ◽  
Mesenchymal Stem Cell Differentiation

scholarly journals Exosomes from human umbilical cord mesenchymal stem cells attenuate the inflammation of severe steroid-resistant asthma by reshaping macrophage polarization

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Bing Dong ◽  
Chao Wang ◽  
Jing Zhang ◽  
Jinrong Zhang ◽  
Yinuo Gu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Quantitative Proteomics ◽  
Macrophage Polarization ◽  
Human Umbilical Cord ◽  
Raw 264.7 ◽  
Steroid Resistant ◽  
Central Protein

Abstract Background Severe, steroid-resistant asthma (SSRA) is a serious clinical problem in asthma management. Affected patients have severe clinical symptoms, worsened quality of life, and do not respond to steroid, a mainstay steroid treatment of asthma. Thus, effective therapies are urgently needed. Exosomes derived from mesenchymal stem cell (MSC-Exo) has become attractive candidates for the lung inflammatory diseases through its immunomodulatory effects. In this study, we explored the therapeutic effects of MSC-Exo in SSRA and identified the therapeutic mechanism of MSC-Exo. Method Exosomes from human umbilical cord mesenchymal stem cell (hUCMSC) were isolated and characterized by transmission electron microscopy, nanoparticle tracking analysis and flow cytometry analysis. Effects of MSC-Exo on airway hyper responsiveness (AHR), inflammation, histopathology, and macrophage polarization in SSRA in mice were evaluated. Systematic depletion of macrophages determined the role of macrophages in the therapeutic effect of SSRA in mice. LPS-stimulated RAW 264.7 cell model was constructed to determine the underlying mechanism of MSC-Exo on macrophage polarization. qRT-PCR, Western blotting, immunofluorescence, and flow cytometry were performed to evaluate the expression of M1 or M2 markers. Tandem mass tags (TMT)-labeled quantitative proteomics were applied to explore the central protein during the regulation effect of MSC-Exo on macrophage polarization. Knockdown and overexpression of TRAF1 were used to further clarify the role of the central protein on macrophage polarization. Result We successfully isolated and characterized exosomes from hUCMSCs. We verified that the intratracheal administration of MSC-Exo reversed AHR, histopathology changes, and inflammation in SSRA mice. Systematic depletion of macrophages weakened the therapeutic effect of MSC-Exo. We found that MSC-Exo treatment inhibited M1 polarization and promoted M2 polarization in LPS-stimulated RAW 264.7 cells. Subsequently, tumor necrosis factor receptor-associated factor 1 (TRAF1) was determined as the central protein which may be closely related to the regulation of macrophage polarization from TMT-labeled quantitative proteomics analysis. Knockdown and overexpression of TRAF1 demonstrated that the effect of MSC-Exo treatment on macrophage polarization, NF-κB and PI3K/AKT signaling was dependent on TRAF1. Conclusion MSC-Exo can ameliorate SSRA by moderating inflammation, which is achieved by reshaping macrophage polarization via inhibition of TRAF1.

scholarly journals Human umbilical cord mesenchymal stem cell-derived exosomal miR-27b attenuates subretinal fibrosis via suppressing epithelial–mesenchymal transition by targeting HOXC6

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Dongli Li ◽  
Junxiu Zhang ◽  
Zijia Liu ◽  
Yuanyuan Gong ◽  
Zhi Zheng
Keyword(s):  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Umbilical Cord ◽  
Mechanism Of Action ◽  
Epithelial Mesenchymal Transition ◽  
Human Umbilical Cord ◽  
Mesenchymal Transition ◽  
Rpe Cells ◽  
Subretinal Fibrosis

Abstract Background and aim Subretinal fibrosis resulting from neovascular age-related macular degeneration (nAMD) is one of the major causes of serious and irreversible vision loss worldwide, and no definite and effective treatment exists currently. Retinal pigmented epithelium (RPE) cells are crucial in maintaining the visual function of normal eyes and its epithelial–mesenchymal transition (EMT) is associated with the pathogenesis of subretinal fibrosis. Stem cell-derived exosomes have been reported to play a crucial role in tissue fibrosis by transferring their molecular contents. This study aimed to explore the effects of human umbilical cord-derived mesenchymal stem cell exosomes (hucMSC-Exo) on subretinal fibrosis in vivo and in vitro and to investigate the anti-fibrotic mechanism of action of hucMSC-Exo. Methods In this study, human umbilical cord-derived mesenchymal stem cells (hucMSCs) were successfully cultured and identified, and exosomes were isolated from the supernatant by ultracentrifugation. A laser-induced choroidal neovascularization (CNV) and subretinal fibrosis model indicated that the intravitreal administration of hucMSC-Exo effectively alleviated subretinal fibrosis in vivo. Furthermore, hucMSC-Exo could efficaciously suppress the migration of retinal pigmented epithelial (RPE) cells and promote the mesenchymal–epithelial transition by delivering miR-27b-3p. The latent binding of miR-27b-3p to homeobox protein Hox-C6 (HOXC6) was analyzed by bioinformatics prediction and luciferase reporter assays. Results This study showed that the intravitreal injection of hucMSC-Exo effectively ameliorated laser-induced CNV and subretinal fibrosis via the suppression of epithelial–mesenchymal transition (EMT) process. In addition, hucMSC-Exo containing miR-27b repressed the EMT process in RPE cells induced by transforming growth factor-beta2 (TGF-β2) via inhibiting HOXC6 expression. Conclusions The present study showed that HucMSC-derived exosomal miR-27b could reverse the process of EMT induced by TGF-β2 via inhibiting HOXC6, indicating that the exosomal miR-27b/HOXC6 axis might play a vital role in ameliorating subretinal fibrosis. The present study proposed a promising therapeutic agent for treating ocular fibrotic diseases and provided insights into the mechanism of action of hucMSC-Exo on subretinal fibrosis.

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