Overexpression of OLIG2 and MYT1L Transcription Factors Enhance the Differentiation Potential of Human Mesenchymal Stem Cells into Oligodendrocytes.

BACKGROUND: Demyelinating diseases represent a broad spectrum of disorders and are characterized by the loss of specialized glial cells (oligodendrocytes), which eventually leads to neuronal degeneration. Stem cell-based regenerative approaches provide therapeutic options to regenerate demyelination-induced neurodegeneration. OBJECTIVES: The current study aims to explore the role of oligodendrocyte-specific transcription factors (OLIG2 and MYT1L) under suitable media composition to facilitate human umbilical-cord-derived mesenchymal stem cells (hUC-MSCs) differentiation toward oligodendrocyte for their potential use to treat demyelinating disorders. METHODOLOGY: hUC-MSCs were isolated, cultured, and characterized based on their morphological and phenotypic characteristics. hUC-MSCs were transfected with OLIG2 and MYT1L transcription factors individually and in synergistic (OLIG2 + MYT1L) groups using a lipofectamine-based transfection method and incubated under two different media compositions (normal and oligo induction media). Transfected hUC-MSCs were assessed for lineage specification and differentiation using qPCR. Differentiation was also analyzed via immunocytochemistry by determining the expression of oligodendrocyte-specific proteins. RESULTS: All the transfected groups showed significant upregulation of GFAP and OLIG2 with downregulation of NES, demonstrating the MSC commitment toward the glial lineage. Transfected groups also presented significant overexpression of oligodendrocyte-specific markers (SOX10, NKX2.2, GALC, CNP, CSPG4, MBP, and PLP1). Immunocytochemical analysis showed intense expression of OLIG2, MYT1L, and NG2 proteins in both normal and oligo induction media after 3 and 7 days. CONCLUSIONS: The study concludes that OLIG2 and MYT1L have the potential to differentiate hUC-MSCs into oligodendrocyte-like cells, which is greatly facilitated by the oligo induction medium. The study may serve as a promising cell-based therapeutic strategy against demyelination-induced neuronal degeneration.

Carbonised Human Hair Incorporated in Agar/KGM Bioscaffold for Tissue Engineering Application: Fabrication and Characterisation.

Carbon derived from biomass waste usage is rising in various fields of application due to its availability, cost-effectiveness, and sustainability, but it remains limited in tissue engineering applications. Carbon derived from human hair waste was selected to fabricate a carbon-based bioscaffold (CHAK) due to its ease of collection and inexpensive synthesis procedure. The CHAK was fabricated via gelation, rapid freezing, and ethanol immersion and characterised based on their morphology, porosity, Fourier transforms infrared (FTIR), tensile strength, swelling ability, degradability, electrical conductivity, and biocompatibility using Wharton's jelly-derived mesenchymal stem cells (WJMSCs). The addition of carbon reduced the porosity of the bioscaffold. Via FTIR analysis, the combination of carbon, agar, and KGM was compatible. Among the CHAK, the 3HC bioscaffold displayed the highest tensile strength (62.35 ± 29.12 kPa). The CHAK also showed excellent swelling and water uptake capability. All bioscaffolds demonstrated a slow degradability rate (<50%) after 28 days of incubation, while the electrical conductivity analysis showed that the 3AHC bioscaffold had the highest conductivity compared to other CHAK bioscaffolds. Our findings also showed that the CHAK bioscaffolds were biocompatible with WJMSCs. These findings showed that the CHAK bioscaffolds have potential as bioscaffolds for tissue engineering applications.

Impact of Graphene Derivatives as Artificial Extracellular Matrices on Mesenchymal Stem Cells.

Thanks to stem cells' capability to differentiate into multiple cell types, damaged human tissues and organs can be rapidly well-repaired. Therefore, their applicability in the emerging field of regenerative medicine can be further expanded, serving as a promising multifunctional tool for tissue engineering, treatments for various diseases, and other biomedical applications as well. However, the differentiation and survival of the stem cells into specific lineages is crucial to be exclusively controlled. In this frame, growth factors and chemical agents are utilized to stimulate and adjust proliferation and differentiation of the stem cells, although challenges related with degradation, side effects, and high cost should be overcome. Owing to their unique physicochemical and biological properties, graphene-based nanomaterials have been widely used as scaffolds to manipulate stem cell growth and differentiation potential. Herein, we provide the most recent research progress in mesenchymal stem cells (MSCs) growth, differentiation and function utilizing graphene derivatives as extracellular scaffolds. The interaction of graphene derivatives in human and rat MSCs has been also evaluated. Graphene-based nanomaterials are biocompatible, exhibiting a great potential applicability in stem-cell-mediated regenerative medicine as they may promote the behaviour control of the stem cells. Finally, the challenges, prospects and future trends in the field are discussed.

Correction to: MicroRNAomic Transcriptomic Analysis Reveal Deregulation of Clustered Cellular Functions in Human Mesenchymal Stem Cells During in Vitro Passaging.

The original version of this article unfortunately contained a mistake.

MicroRNAomic Transcriptomic Analysis Reveal Deregulation of Clustered Cellular Functions in Human Mesenchymal Stem Cells During in Vitro Passaging.

Clinical trials using human mesenchymal stem/stromal cells (hMSCs) for cell replacement therapy showed varied outcomes, where cells' efficacy has been perceived as the limiting factor. In particular, the quality and number of the expanded cells in vitro. In this study, we aimed to determine molecular signatures of hMSCs derived from the pulp of extracted deciduous teeth (SHED) and Wharton's jelly (WJSCs) that associated with cellular ageing during in vitro passaging. We observed distinct phenotypic changes resembling proliferation reduction, cell enlargement, an increase cell population in G2/M phase, and differentially expressed of tumor suppressor p53 in passage (P) 6 as compared to P3, which indicating in vitro cell senescence. The subsequent molecular analysis showed a set of diverse differentially expressed miRNAs and mRNAs involved in maintaining cell proliferation and stemness properties. Considering the signaling pathway related to G2/M DNA damage regulation is widely recognized as part of anti-proliferation mechanism controlled by p53, we explored possible miRNA-mRNA interaction in this regulatory pathway based on genomic coordinates retrieved from miRanda. Our work reveals the potential reason for SHED underwent proliferation arrest due to the direct impinge on the expression of CKS1 by miRNAs specifically miR-22 and miR-485-5p which lead to down regulation of CDK1 and Cyclin B. It is intended that our study will contribute to the understanding of these miRNA/mRNA driving the biological process and regulating different stages of cell cycle is beneficial in developing effective rejuvenation strategies in order to obtain quality stem cells for transplantation.

Generation of Sheffield (Shef) human embryonic stem cell lines using a microdrop culture system.

The conventional method for the derivation of human embryonic stem cells (hESCs) involves inner cell mass (ICM) co-culture with a feeder layer of inactivated mouse or human embryonic fibroblasts in an in vitro fertilisation culture dish. Growth factors potentially involved in primary derivation of hESCs may be lost or diluted in such a system. We established a microdrop method which maintained feeder cells and efficiently generated hESCs. Embryos were donated for stem cell research after fully informed patient consent. A feeder cell layer was made by incubating inactivated mouse embryonic fibroblasts (MEFs) feeder cells in a 50 microl drop of medium (DMEM/10% foetal calf serum) under mineral oil in a small tissue culture dish. MEFs formed a confluent layer and medium was replaced with human embryonic stem medium supplemented with 10% Plasmanate (Bayer) and incubated overnight. Cryopreserved embryos were thawed and cultured until the blastocyst stage and the zona pellucida removed with pronase (2 mg/ml; Calbiochem). A zona-free intact blastocyst was placed in the feeder microdrop and monitored for ES derivation with medium changed every 2-3 d. Proliferating hESCs were passaged into other feeder drops and standard feeder preparation by manual dissection until a stable cell line was established. Six hESC lines (Shef 3-8) were derived. From a total of 46 blastocysts (early to expanded), five hESC lines were generated (Shef 3-7). Shef 3-6 were generated on MEFs from 25 blastocysts. Shef7 was generated on human foetal gonadal embryonic fibroblasts from a further 21 blastocysts. From our experience, microdrop technique is more efficient than conventional method for derivation of hESCs and it is much easier to monitor early hESC derivation. The microdrop method lends itself to good manufacturing practice derivation of hESCs.