“…However, as the present study did not involve direct co-culture of the two cell types (but rather the secretome), we decided to also characterize the effects of secretomes from MSC in their proliferative phase (non-irradiated). Although some reports have shown irradiated MSC maintain their immunomodulatory function in vitro [25,26], we still hypothesized that radiation injury, or entry into a non-replicative G 0 cell cycle, may have detrimental effects on the ASC-support engendered by our secretome; thus we sought to test secretomes from both irradiated and non-irradiated MSC.10.1080/20013078.2018.1463778-F0001Figure 1.ASC survival ex vivo with irradiated, non-irradiated secretomes and secretomes treated with Cleanascite. (a) Representative flow cytometry analysis of blood ASC (CD19 + CD27 hi CD38 hi ) from PMBC isolated 7 days after vaccination from a healthy adult.…”
Extracellular vesicles (EVs) from bone marrow (BM)-derived mesenchymal stromal cells (BM-MSC) are novel mechanisms of cell-cell communication over short and long distances. BM-MSC have been shown to support human antibody secreting cells (ASC) survival ex vivo, but whether the crosstalk between the MSC-ASC interaction can occur via EVs is not known. Thus, we evaluated the role of EVs in ASC survival and IgG secretion. EVs were isolated from irradiated and non-irradiated primary BM-MSC and were quantified. They were further characterized by electron microscopy (EM) and CD63 and CD81 immuno-gold EM staining. Human ASC were isolated via fluorescence-activated cell sorting (FACS) and cultured ex vivo with the EV fractions, the EV-reduced fractions, or conventional media. IgG Elispots were used to measure the survival and functionality of the ASC. Contents of the EV fractions were evaluated by proteomics. We saw that both irradiated and non-irradiated MSC secretome preparations afforded vesicles of a size consistent with EVs. Both preparations appeared comparable in EM morphology and CD63 and CD81 immuno-gold EM. Both irradiated and non-irradiated EV fractions supported ASC function, at 88% and 90%, respectively, by day 3. In contrast, conventional media maintained only 4% ASC survival by day 3. To identify the specific factors that provided in vitro ASC support, we compared proteomes of the irradiated and non-irradiated EV fractions with conventional media. Pathway analysis of these proteins identified factors involved in the vesicle-mediated delivery of integrin signalling proteins. These findings indicate that BM-MSC EVs provide an effective support system for ASC survival and IgG secretion.
“…However, as the present study did not involve direct co-culture of the two cell types (but rather the secretome), we decided to also characterize the effects of secretomes from MSC in their proliferative phase (non-irradiated). Although some reports have shown irradiated MSC maintain their immunomodulatory function in vitro [25,26], we still hypothesized that radiation injury, or entry into a non-replicative G 0 cell cycle, may have detrimental effects on the ASC-support engendered by our secretome; thus we sought to test secretomes from both irradiated and non-irradiated MSC.10.1080/20013078.2018.1463778-F0001Figure 1.ASC survival ex vivo with irradiated, non-irradiated secretomes and secretomes treated with Cleanascite. (a) Representative flow cytometry analysis of blood ASC (CD19 + CD27 hi CD38 hi ) from PMBC isolated 7 days after vaccination from a healthy adult.…”
Extracellular vesicles (EVs) from bone marrow (BM)-derived mesenchymal stromal cells (BM-MSC) are novel mechanisms of cell-cell communication over short and long distances. BM-MSC have been shown to support human antibody secreting cells (ASC) survival ex vivo, but whether the crosstalk between the MSC-ASC interaction can occur via EVs is not known. Thus, we evaluated the role of EVs in ASC survival and IgG secretion. EVs were isolated from irradiated and non-irradiated primary BM-MSC and were quantified. They were further characterized by electron microscopy (EM) and CD63 and CD81 immuno-gold EM staining. Human ASC were isolated via fluorescence-activated cell sorting (FACS) and cultured ex vivo with the EV fractions, the EV-reduced fractions, or conventional media. IgG Elispots were used to measure the survival and functionality of the ASC. Contents of the EV fractions were evaluated by proteomics. We saw that both irradiated and non-irradiated MSC secretome preparations afforded vesicles of a size consistent with EVs. Both preparations appeared comparable in EM morphology and CD63 and CD81 immuno-gold EM. Both irradiated and non-irradiated EV fractions supported ASC function, at 88% and 90%, respectively, by day 3. In contrast, conventional media maintained only 4% ASC survival by day 3. To identify the specific factors that provided in vitro ASC support, we compared proteomes of the irradiated and non-irradiated EV fractions with conventional media. Pathway analysis of these proteins identified factors involved in the vesicle-mediated delivery of integrin signalling proteins. These findings indicate that BM-MSC EVs provide an effective support system for ASC survival and IgG secretion.
“…For that first part of our study, we aimed to evaluate the biological response of aMSCs to radiation exposure in comparison to 4T1 cells, a mesenchymal-like cancer cell model that has considerable ionizing radiationresistant Cancer Stem Cells (CSC) subpopulation [109,182], and to mouse fibroblast (NIH3T3-wt), a normal cell model. Different mechanisms have been reported as main pathways of such radio-resistance, such as cell cycle (CC) arrest (G2/M arrest) and activation of double stranded DNA (dsDNA) damage repair machinery; namely the homologous recombination repair (HRR) and nonhomologous end-joining repair (NHEJR) [95][96][97][98][99]. These mechanisms were also demonstrated to be responsible for the RT resistance of cancer stem cells (CSC), also known as cancer initiating cells, which have been linked to cancer disease recurrence and aggression [192,193].…”
Section: Chapter 7 Discussionmentioning
confidence: 99%
“…In addition, the surface antigen CD105 presence is important for normal cellular DNA repair [94]. Different mechanisms have been reported explaining such radio-resistance such as, cell cycle (CC) arrest (G2/M arrest) and activation of double stranded DNA (dsDNA) damage repair; namely the homologous recombination repair (HRR) and non-homologous end joining repair (NHEJR) [95][96][97][98][99]. These mechanisms were also evidenced to be responsible for the IR resistance of cancer stem cells (CSC), also known as cancer initiating cells, which have been linked to cancer disease recurrence and aggression [100][101][102][103].…”
“…3 Since HSPCs are more sensitive to radiation, mesenchymal stem cells (MSCs) luckily have an intrinsic radiation-resistant phenotype. 4,5 Although effective radiation countermeasure approaches were initiated six decades ago, very few have unanimous endorsement by health professionals. Radiation protectors [such as WR-2721, Toll-like receptor ligands, flavonoids, Ex-RAD® (Onconova Therapeutics Inc., Newton, PA) and vitamin E] are prophylactic agents that are administered before exposure to prevent radiation-induced cellular and molecular damage.…”
Dr Lan Yang and Dr Ying-Min Liang contributed equally to this workObjective: The aim of this study was to find a new radiation protector, platelet factor 4 (PF4) and to identify its effect on haemopoietic microenvironment in vitro and in vivo. Methods: Radiation damage on bone marrow mesenchymal stem cells ex and in vitro was set up as models. Growth curve analysis, clonogenic survival assay, FACSCalibur™ (BD Immunocytometry Systems, San Jose, CA), 5-ethynyl-29-deoxyuridine immunofluorescence staining and quantitative reverse transcription-polymerase chain reaction were employed to assess the characterization of bone marrow mesenchymal stem cells (BMSCs), proliferation, apoptosis, cell cycle and gene expression. Results: A dose-and time-dependent enhancement of cell viability and survival was observed for PF4 treatment along with 500 cGy g-radiation in vitro. The same phenomena were noted in vivo, including enhancement of adherence and proliferation ability while inhibition of cell apoptosis, which were associated with a short-term decrease in the G0/G1 ratio owing to S phase arrest. These were accompanied with enhanced Bcl-2 expression and p53/p21 loss. Conclusion: These results uncover that PF4 might be a novel therapeutic approach, which could reduce DNA damage and increase survival of BMSCs, in part, by inhibiting p53/p21 axis and facilitating DNA damage repair. Advances in knowledge: This study explores the feasibility of a new radioprotector and hence may be clinically important.
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