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Glutathione-related inflammatory signature in hepatocytes differentiated from the progenitor mesenchymal stem cells.

ABSTRACT: N-acetylcysteine (NAC) as a glutathione inducer is known for its anti-inflammatory effects in inflammatory conditions. The aim of the present study was to know if supplementation of the culture medium with NAC can improve anti-inflammatory activities of hepatocytes during their differentiation from mesenchymal stem cells (MSCs). For this, in vitro hepatic differentiation of MSCs was performed in culture medium supplemented with NAC and selected pro- and anti-inflammatory factors were monitored for two weeks. Treatment of the MSCs undergoing hepatic differentiation with NAC (0.1 and 1.0 mM) caused a significant (~5-fold) increase in proliferation rate of MSCs, whereas the rate of hepatic differentiation was declined in NAC-treated cells as compared to those untreated with NAC. Under these circumstances, NAC caused a significant increase in total glutathione in cell lysate during 2 weeks of differentiation as compared to untreated group. NAC-related increase in glutathione was associated with significant alterations in tumor necrosis factor-? (TNF-?), interleukin (IL)-6, IL-8 and IL-10 levels secreted in the culture medium. A substantial decrease in the IL-6, IL-8 and TNF-? levels in the culture medium supplemented with NAC was obvious in hepatocytes recovered 14 days after differentiation. In contrast, the secretary IL-10 was significantly increased as a result of NAC treatments. These data suggest that NAC supplementation can improve anti-inflammatory activities of the hepatocytes derived from MSCs. NAC function mediated by glutathione synthesis can also help in modulation of proliferation of the stem cells and their differentiation into hepatocyte-like cells.

SUBMITTER: Allameh A

PROVIDER: S-EPMC7287236 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Glutathione-related inflammatory signature in hepatocytes differentiated from the progenitor mesenchymal stem cells.

Allameh Abdolamir A Ahmadi-Ashtiani Hamid Reza HR Maleki Narges N Maleki Narges N

Heliyon 20200608 6

N-acetylcysteine (NAC) as a glutathione inducer is known for its anti-inflammatory effects in inflammatory conditions. The aim of the present study was to know if supplementation of the culture medium with <i>NAC</i> can improve anti-inflammatory activities of hepatocytes during their differentiation from mesenchymal stem cells (MSCs). For this, <i>in vitro</i> hepatic differentiation of MSCs was performed in culture medium supplemented with NAC and selected pro- and anti-inflammatory factors we ...[more]

PMID: 32551386

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Project description:BackgroundBecause specific marker molecules for phenotypical identification of mesenchymal stem and progenitor cells are missing, the assessment of the in vitro-differentiation capacity is a prerequisite to characterize these cells. However, classical differentiation protocols are often cell-consuming and time intensive. Therefore, the establishment of novel strategies for differentiation is one topic of current efforts in stem cell biology. The goal of this study was to demonstrate the practicability of a new differentiation test using plastic adherent cell isolates from different tissues.ResultsWe introduced the mesenchymal microsphere method as a feasible time- and cell saving screening method to analyse multilineage differentiation properties of adult progenitor cells in a three-dimensional system. For this purpose we isolated, characterized and analyzed new sources of adult murine mesenchymal progenitor cells from perirenal adipose tissue and mediastinal stromal tissue in comparison to bone marrow progenitor cells. The proliferation capacity of the cells was demonstrated by determination of the daily doubling index. Although the flow cytometry analysis of undifferentiated cells revealed differences in the expression of CD marker molecules, all isolates have the capacity for multilineage differentiation following the mesenchymal microsphere protocol as well as the classical "micro mass body" protocol for chondrogenic and the monolayer cultivation protocol for osteogenic and adipogenic differentiation. Differentiation was characterized using histochemical and immunhistochemical staining as well as RT-PCR.ConclusionsWe were able to show that the mesenchymal microsphere method is an efficient test system for chondro-, osteo- and adipogenic differentiation of adult progenitor cells. The advantage of this system in comparison to classical protocols is that approximately 7 times lower cell numbers are necessary. Since classical culture procedures are time intensive because high cell numbers have to be obtained, the new differentiation method may also save cells and time in future clinical applications using human mesenchymal stromal cells.

Project description:BackgroundMesenchymal stem cells (MSCs) are promising targets for therapeutic use in regenerative medicine and tissue engineering. In the previous study, we have found that MSCs could be reverted to a primitive stem cell population after in vitro induction of osteogenic and de-osteogenic differentiation (de-osteogenic differentiated MSCs, De-Os-MSCs). De-Os-MSCs showed improved cell survival and osteogenic potential. However, the underlying mechanism and its potential effect on fracture healing has not been explored.MethodsMSCs were isolated from the rat bone marrow. MicroRNAs were cloned into lentiviral vectors and transduced into MSCs to observe the effects on osteogenesis. The expression levels of marker genes were evaluated by quantitative RT-PCR. Ectopic bone formation model was used to evaluate the bone regeneration ability of mir-92b transduced MSCs in vivo. An open femur fracture model was established, and MSCs or De-Os-MSCs were administrated to the fracture sites. Histological, biomechanical and microCT analysis were used to evaluate the quality of bone.ResultsIn the present study, we found that mir-92b was significantly increased in the secretions of De-Os-MSCs. And mir-92b could promote the osteogenic differentiation potential of MSCs by activating pERK and JNK signaling pathways. The ectopic bone formation assay showed that MSCs overexpressing mir-92b formed more bone like tissues in vivo. Most importantly, we found local administration of De-Os-MSCs could accelerate fracture healing using an open femur fracture model in rats. The quality of bone property was much better as shown by microCT and biomechanical testing.ConclusionTaken together, our study demonstrated that mir-92b promoted osteogenesis of MSCs, which was partially accounted for the enhanced osteogenic differentiation potential of De-Os-MSCs. And De-Os-MSCs had shown better regenerative capacity in accelerating fracture healing when they were locally given.The translational potential of this articleDe-Os-MSCs could be used to accelerate fracture healing, and reduce the occurrence of delayed unions and non-unions.

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Glutathione-related inflammatory signature in hepatocytes differentiated from the progenitor mesenchymal stem cells.

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Glutathione-related inflammatory signature in hepatocytes differentiated from the progenitor mesenchymal stem cells.

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