Development of a novel explant culture method for the isolation of mesenchymal stem cells from human breast tumor

Therapeutic targeting of stem cells needs to be strategically developed to control tumor growth and prevent metastatic burden successfully. Breast cancer presents a unique clinical problem because of the variety of cellular subtypes present, including cancer stem cells (CSCs). The development of 3D stem-like properties of human breast tumor spheroids in stem cell factor conditioned media was investigated in orthotopic xenografts for enhanced tumorgenicity in the athymic nude rat model. MCF-7, ZR-75-1, and MDA-MB-231 breast cancer cell lines were cultured in serum-free, stem cell factor-supplemented medium under non-adherent conditions and passaged to generate 3rd generation spheroids. The spheroids were co-cultured with fetal lung fibroblast (FLF) cells before orthotopic heterotransplantation into the mammary fat pads of athymic nude rats. Excised xenografts were assessed histologically by H&E staining and immunohistochemistry for breast cancer marker (ERB1), proliferation marker (Ki67), mitotic marker (pHH3), hypoxia marker (HIF-2α), CSC markers (CD47, CD44, CD24, and CD133), and vascularization markers (CD31, CD34). Breast cancer cells cultured in stem cell factor supplemented medium generated 3D spheroids exhibited increased stem-like characteristics. The 3D stem-like spheroids co-cultured with FLF as supporting stroma reproducibly and efficiently established orthotopic breast cancer xenografts in the athymic nude rat.

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Mesenchymal stem cells isolated from peripheral blood and umbilical cord Wharton’s jelly

Srpski arhiv za celokupno lekarstvo ◽

10.2298/sarh1304178t ◽

2013 ◽

Vol 141 (3-4) ◽

pp. 178-186 ◽

Cited By ~ 34

Author(s):

Drenka Trivanovic ◽

Jelena Kocic ◽

Slavko Mojsilovic ◽

Aleksandra Krstic ◽

Vesna Ilic ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Umbilical Cord ◽

Peripheral Blood ◽

Myogenic Differentiation ◽

Smooth Muscle Actin ◽

Culture Method ◽

Good Alternative ◽

Explant Culture ◽

Differentiation Potential

Introduction. Mesenchymal stem cells (MSCs) are a promising tool for regenerative medicine, but due to the heterogeneity of their populations, different sources and isolation techniques, the characteristics defining MSCs are inconsistent. Objective. The aim of this study was to compare the characteristics of MSCs derived from two different human tissues: peripheral blood (PB-MSCs) and umbilical cord Wharton?s Jelly (UC-MSCs). Methods. The PB-MSC and UC-MSC were isolated by adherence to plastic after gradient-density separation or an explant culture method, respectively, and compared regarding their morphology, clonogenic efficiency, proliferating rates, immunophenotype and differentiation potential. Results. MSCs derived from both sources exhibit similar morphology, proliferation capacity and multilineage (osteogenic, chondrogenic, adipogenic and myogenic) differentiation potential. Differences were observed in the clonogenic capacity and the immunophenotype, since UC-MSCs showed higher CFU-F (colony-forming units-fibroblastic) cloning efficiency, as well as higher embryonic markers (Nanog, Sox2, SSEA4) expression. When additional surface antigens were analyzed by flow cytometry (CD44, CD90, CD105, CD33, CD34, CD45, CD11b, CD235a) or immunofluorescent labeling (vimentin, STRO-1 and ?-smooth muscle actin), most appeared to have similar epitope profiles irrespective of MSC source. Conclusion. The results obtained demonstrated that both MSCs represent good alternative sources of adult MSCs that could be used in cell therapy applications.

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Comparative Assessment of Oral Mesenchymal Stem Cells Isolated from Healthy and Diseased Tissues

Microscopy and Microanalysis ◽

10.1017/s1431927615014749 ◽

2015 ◽

Vol 21 (5) ◽

pp. 1249-1263 ◽

Cited By ~ 5

Author(s):

Emöke Páll ◽

Adrian Florea ◽

Olga Soriţău ◽

Mihai Cenariu ◽

Adrian S. Petruţiu ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Cell Lines ◽

Specific Antigen ◽

Human Mesenchymal Stem Cells ◽

Culture Method ◽

Explant Culture ◽

Population Doubling ◽

Population Doubling Time ◽

Differentiation Potential

AbstractThe aim of the present study was to isolate human mesenchymal stem cells (MSCs) from palatal connective and periodontal granulation tissues and to comparatively evaluate their properties. MSCs were isolated using the explant culture method. Adherence to plastic, specific antigen makeup, multipotent differentiation potential, functionality, and ultrastructural characteristics were investigated. The frequency of colony-forming unit fibroblasts for palatal-derived mesenchymal stem cells (pMSCs) was significantly higher than that of granulation tissue-derived mesenchymal stem cells (gtMSCs). A significantly higher population doubling time and lower migration potential were recorded for gtMSCs than for pMSCs. Both cell lines were positive for CD105, CD73, CD90, CD44, and CD49f, and negative for CD34, CD45, and HLA-DR, but the level of expression was different. MSCs from both sources were relatively uniform in their ultrastructure. Generally, both cell lines possessed a large, irregular-shaped euchromatic nucleus, and cytoplasm rich in mitochondria, lysosomes, and endoplasmic reticulum. The periphery of the plasma membrane displayed many small filopodia. MSCs from both cell lines were successfully differentiated into osteogenic, adiopogenic, and chondrogenic lineages. Both healthy and diseased tissues may be considered as valuable sources of MSCs for regenerative medicine owing to the high acceptance and fewer complications during harvesting.

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Post-calving umbilical cord tissue offcut: A potential source for the isolation of bovine mesenchymal stem cells

Veterinary World ◽

10.14202/vetworld.2020.2772-2779 ◽

2020 ◽

Vol 13 (12) ◽

pp. 2772-2779

Author(s):

Parishma Debbarma ◽

Tanmay Mondal ◽

Camelia Manna ◽

Kuldeep Kumar ◽

Joydip Mukherjee ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Umbilical Cord ◽

Cultured Cells ◽

Culture Method ◽

Explant Culture ◽

Isolated Cells ◽

Isolation Technique ◽

Umbilical Cord Tissue

Background and Aim: Veterinary health care is an emergent area in animal sciences and innovative therapeutic approaches happen to be imperative in the present days. In view of the importance of cattle health and production, it is necessary to take up contemporary approach of stem cell therapy in this sector also. This study aimed to standardize an explant culture method of bovine umbilical tissue offcut to isolate mesenchymal stem cells (MSCs) because considerable efforts are required for ensuring easy accessibility and availability of MSCs in bulk quantity, as well as in establishing and characterizing the cell lines. Materials and Methods: The umbilical cord (UC) tissue matrix offcut was collected after calving. A simplified in vitro cell isolation technique was followed to collect the emerged out cells from the explants of UC. Further, we expanded these isolated cells in vitro, observed its growth kinetics, and characterized to confirm as per the criterion of bovine MSCs. Results: A considerable exponential growth rate of the UC-derived cells was noticed. In addition to their confirmation as MSCs, the cells also exhibited plastic adherent property and maintained the spindle-shaped morphology throughout the in vitro culture. The cultured cells were found positive MSC-specific surface markers CD105, CD90, and CD73 and were negative for hematopoietic cell marker CD45. Cytochemical studies revealed the ability of the cells to differentiate into osteogenic, chondrogenic, and adipogenic lineages. Conclusion: This simplified method of isolation and culture of bovine multipotent MSCs from the UC offcut collected after calving could be extrapolated for the greater availability of the cells for prospective therapeutic applications.

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