Comparative Analysis of Wharton’s Jelly Mesenchymal Stem Cell (WJ-MSCs) Isolated Using Explant and Enzymatic Methods

Abstract

Wharton’s Jelly is one of the best sources for mesenchymal stem cells. Human Wharton’s Jelly Mesenchymal Stem Cells (hWJ-MSCs) have high proliferation, multi-lineage differentiation potential, and do not produce any teratogen or carcinogen. These characteristics make hWJ-MSCs become suitable for regenerative medicine. Some methods were developed to isolate hWJ-MSCs from umbilical cord, such as explant method and enzymatic method. This study aims to characterize hWJ-MSCs which are isolated by two different methods, explant attachment method and enzymatic method. hWJ-MSCs isolation was performed through explant method and enzymatic method using trypsin, hyaluronidase and collagenase type 1 with certain ratio of concentration. Isolated hWJ-MSCs was characterized using flow cytometer to detect the expression of CD44, CD90, CD105, CD73 and negative lineage. MSCs differentiation assay was performed to analyze adipogenic, chondrogenic and osteogenic cells lineage. We successfully isolated hWJ-MSCs from umbilical cord through enzymatic and explant methods. Immunophenotyping assay through flow cytometry analysis showed high purity of WJ-MSCs. The isolated hWJ-MSCs from both methods showed positive expression of CD44, CD90, CD105, and CD73. The isolated hWJ-MSCs exhibited capacity to differentiate into adipocyte, chondrocyte, and osteocyte cells. hWJ-MSCs isolated through explant and enzymatic method have high proliferation capacity and be able to differentiate into three different lineage cells. Both methods explant attachment and enzymatic methods are efficiently produced hWJ-MSCs.

Osteogenic potential of a 3D printed silver nanoparticle-based electroactive scaffold for bone tissue engineering using human Wharton's jelly mesenchymal stem cells

“…2(d)). Even though ISCT mentioned that ≥95% of MSCs should express CD105, there are several studies demonstrating that expression of the CD105 marker by MSCs could be varied, such as 88.1%, 39 90.76%, 36 92.94%, 51 and 94.54%, 52 depending on the isolated methods, cell source, culture time in vitro , and differentiation status. 36,53 Thus, the isolated cells still comply with the ISCT's criteria even though the cell population that emits CD105 does not reach 95%.…”

“…Subculture is carried out until it reaches passage 4 for further testing. 36,37 To ensure that the cells isolated from human Wharton's jelly were truly hWJ-MSCs cells and meet the requirements of the International Society for Cell Therapy (ISCT) as hWJ-MSCs, characterization was carried out. Cells were characterized using an inverted microscope to determine the attachment of cells to the substrate and the morphology of the isolated cells, analyzing cell surface marker expression using flow cytometry (BD FACSCanto II, BD Biosciences, Erembodegem, Belgium), and differential staining to determine multilineage differentiation potential.…”

A GMP-compliant manufacturing method for Wharton’s jelly-derived mesenchymal stromal cells

“…While differences in primary cell culture time and yield can be observed, there are no significant disparities in terms of cell viability, morphology, proliferation, surface marker expression, and differentiation capacity after passaging. These findings suggest that both methods result in comparable outcomes for maintaining the desired characteristics of MSCs [21][22][23][24][25][26]. Apart from the WJ-MSCs isolation, several ongoing studies are focused on developing GMP-compliant production processes for clinical-grade WJ-MSCs, involving process design, manufacturing protocols, quality control, and characterization research [18,[27][28][29][30][31].…”

Isolation and Characterization of Fibroblast from Normal and Thalassemia Foreskin

“…The epidermis and dermis layers of the foreskin were separated, and the dermis part of the foreskin was chopped (1 -2 mm). The tissue is placed in a 6-well plate, and let stand for 1 h (until the tissue adheres) [17]. Added to each well 1 mL of complete medium containing MEM-α (Biowest, L0475-500), 20 % Fetal Bovine Serum (FBS) (Biowest, S181B-500), 1 % Nanomycopulitine Concentrate 20× (Biowest, L-X16-100), 1 % ABAM, 1 % Amphotericin B (Biowest, L0009-100), 1 % MEM Non Essential Amino Acids 100× (Biowest, X0557-100), 1 % L-Glutamine 100× (Biowest, X0550-100), 1 % MEM Vitamins 100× (Biowest, X0556-100), 1 % MEM Non Essential Amino Acids 100× (Biowest, X0557-100), and 0.1 % Gentamicin (Gibco, 2517932).…”

“…Explants were incubated for 1 week in a 5 % CO2 incubator at a temperature of 37 °C (Thermo Scientific, 8000DH) [18]. Observation of cell growth was carried out with an inverted microscope (Olympus, CKX41-F32FL), then cells in 6-well plates were harvested and transferred into a T75 tissue culture flask ± 2 weeks after isolation [19,17].…”