Limits...
Myogenic-induced mesenchymal stem cells are capable of modulating the immune response by regulatory T cells.

Joo S, Lim HJ, Jackson JD, Atala A, Yoo JJ - J Tissue Eng (2014)

Bottom Line: Therefore, our aim was to evaluate the effects of mesenchymal stem cell differentiation on the immune characteristics of cells in vitro.Two different 5-aza-2'-deoxycytidine doses (0.5 and 3 µM) were evaluated for their effects on mesenchymal stem cell skeletal myogenic differentiation potential, immune antigen expression, and mixed lymphocytic reactions.Myogenic-induced mesenchymal stem cells do not elicit alloreactive lymphocyte proliferative responses and are able to modulate immune responses.

View Article: PubMed Central - PubMed

Affiliation: Wake Forest Institute for Regenerative Medicine, Wake Forest University Health Sciences, Winston-Salem, NC, USA ; Biomedical Research Institute, Joint Institute for Regenerative Medicine, Kyungpook National University Hospital, Daegu, Republic of Korea.

ABSTRACT
Cell therapy for patients who have intractable muscle disorders may require highly regenerative cells from young, healthy allogeneic donors. Mesenchymal stem cells are currently under clinical investigation because they are known to induce muscle regeneration and believed to be immune privileged, thus making them suitable for allogeneic applications. However, it is unclear whether allogeneic and myogenic-induced mesenchymal stem cells retain their immunomodulatory characteristics. Therefore, our aim was to evaluate the effects of mesenchymal stem cell differentiation on the immune characteristics of cells in vitro. We investigated the immunologic properties of mesenchymal stem cells after myogenic induction. Mesenchymal stem cells were obtained from C57BL/6 mice and the C3H/10T1/2 murine mesenchymal stem cell line. Two different 5-aza-2'-deoxycytidine doses (0.5 and 3 µM) were evaluated for their effects on mesenchymal stem cell skeletal myogenic differentiation potential, immune antigen expression, and mixed lymphocytic reactions. Using a mixed lymphocytic reaction, we determined the optimal splenocyte proliferation inhibition dose. The induction of regulatory T cells was markedly increased by the addition of 3 µM 5-aza-2'-deoxycytidine-treated mesenchymal stem cells. Myogenic-induced mesenchymal stem cells do not elicit alloreactive lymphocyte proliferative responses and are able to modulate immune responses. These findings support the hypothesis that myogenic-induced mesenchymal stem cells may be transplantable across allogeneic barriers.

No MeSH data available.


Related in: MedlinePlus

Proliferative activity of CD3+ spleen cells cocultured with undifferentiated and 5-aza-CdR-treated myogenic-induced MSCs. (a) To assess proliferation, the CFSE-labeled responding spleen cells (1 × 106) from C57BL/6 mice were cultured alone with PHA as positive control or was cultured alone with Mit C as negative control or cocultured with Mit C–treated allogeneic MSCs either myogenic induced (5-aza-CdR-treated) or undifferentiated with PHA. Triplicate cultures were incubated at 37 °C for 5 days. On day 5 of culture, the cells were harvested for staining with anti-CD3 APC. Samples were acquired on a fluorescence-activated cell sorting calibur flow cytometer and data were gated on CD3+ events. CFSE analyses were determined using FlowJo software. Numbers in histograms represent percent divided cells. (b) Precursor frequency of T lymphocytes in the presence of PHA was significantly greater than undifferentiated cells (0µM 5-aza-CdR, p < 0.05), whereas there was no significant difference between undifferentiated cells and those treated with 5-aza-CdR: 0 µM, 0.5 µM, and 3 µM (106 MSCs per treatment group). Data are representative of six independent experiments. Preliminary analysis of data for panels A and B was performed using the proliferation algorithm in the FlowJo software. Statistical analysis was determined by a one-way ANOVA followed by a Dunn’s multiple comparisons test.5-Aza-CdR: 5-aza-2′-deoxycytidine; MSC: mesenchymal stem cell; CFSE: carboxyfluorescein diacetate succinimidyl ester; PHA: phytohemagglutinin; APC: allophycocyanin; Mit C: mitomycin C; ANOVA: analysis of variance; ns: not significant.*p < 0.05.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2 - License 3
getmorefigures.php?uid=PMC3927963&req=5

fig3-2041731414524758: Proliferative activity of CD3+ spleen cells cocultured with undifferentiated and 5-aza-CdR-treated myogenic-induced MSCs. (a) To assess proliferation, the CFSE-labeled responding spleen cells (1 × 106) from C57BL/6 mice were cultured alone with PHA as positive control or was cultured alone with Mit C as negative control or cocultured with Mit C–treated allogeneic MSCs either myogenic induced (5-aza-CdR-treated) or undifferentiated with PHA. Triplicate cultures were incubated at 37 °C for 5 days. On day 5 of culture, the cells were harvested for staining with anti-CD3 APC. Samples were acquired on a fluorescence-activated cell sorting calibur flow cytometer and data were gated on CD3+ events. CFSE analyses were determined using FlowJo software. Numbers in histograms represent percent divided cells. (b) Precursor frequency of T lymphocytes in the presence of PHA was significantly greater than undifferentiated cells (0µM 5-aza-CdR, p < 0.05), whereas there was no significant difference between undifferentiated cells and those treated with 5-aza-CdR: 0 µM, 0.5 µM, and 3 µM (106 MSCs per treatment group). Data are representative of six independent experiments. Preliminary analysis of data for panels A and B was performed using the proliferation algorithm in the FlowJo software. Statistical analysis was determined by a one-way ANOVA followed by a Dunn’s multiple comparisons test.5-Aza-CdR: 5-aza-2′-deoxycytidine; MSC: mesenchymal stem cell; CFSE: carboxyfluorescein diacetate succinimidyl ester; PHA: phytohemagglutinin; APC: allophycocyanin; Mit C: mitomycin C; ANOVA: analysis of variance; ns: not significant.*p < 0.05.

Mentions: The ability of MSCs to inhibit activation of lymphocytes by allogeneic spleen cells from C57BL/6 mice and PHA was examined in a CFSE cell proliferation assay. Undifferentiated and 5-aza-CdR-treated MSCs were tested in this assay. Mit C–treated lymphocytes did not proliferate at all (negative control, Figure 3(a)). Stimulation with PHA induced obvious proliferation of lymphocytes (positive control, Figure 3(a)). Both undifferentiated and 3 µM 5-aza-CdR-treated MSCs induced exerted dose-dependent inhibition of lymphocyte activation (Figure 3(a)). MSCs, either undifferentiated or 3 µM 5-aza-CdR-treated MSCs, completely inhibited the proliferation of PHA-stimulated lymphocytes independently for a ratio MSCs:splenocytes equal to 1:1. By contrast, 0.5 µM 5-aza-CdR-treated MSCs did not inhibit lymphocyte activation until the 1:1 ratio.


Myogenic-induced mesenchymal stem cells are capable of modulating the immune response by regulatory T cells.

Joo S, Lim HJ, Jackson JD, Atala A, Yoo JJ - J Tissue Eng (2014)

Proliferative activity of CD3+ spleen cells cocultured with undifferentiated and 5-aza-CdR-treated myogenic-induced MSCs. (a) To assess proliferation, the CFSE-labeled responding spleen cells (1 × 106) from C57BL/6 mice were cultured alone with PHA as positive control or was cultured alone with Mit C as negative control or cocultured with Mit C–treated allogeneic MSCs either myogenic induced (5-aza-CdR-treated) or undifferentiated with PHA. Triplicate cultures were incubated at 37 °C for 5 days. On day 5 of culture, the cells were harvested for staining with anti-CD3 APC. Samples were acquired on a fluorescence-activated cell sorting calibur flow cytometer and data were gated on CD3+ events. CFSE analyses were determined using FlowJo software. Numbers in histograms represent percent divided cells. (b) Precursor frequency of T lymphocytes in the presence of PHA was significantly greater than undifferentiated cells (0µM 5-aza-CdR, p < 0.05), whereas there was no significant difference between undifferentiated cells and those treated with 5-aza-CdR: 0 µM, 0.5 µM, and 3 µM (106 MSCs per treatment group). Data are representative of six independent experiments. Preliminary analysis of data for panels A and B was performed using the proliferation algorithm in the FlowJo software. Statistical analysis was determined by a one-way ANOVA followed by a Dunn’s multiple comparisons test.5-Aza-CdR: 5-aza-2′-deoxycytidine; MSC: mesenchymal stem cell; CFSE: carboxyfluorescein diacetate succinimidyl ester; PHA: phytohemagglutinin; APC: allophycocyanin; Mit C: mitomycin C; ANOVA: analysis of variance; ns: not significant.*p < 0.05.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2 - License 3
Show All Figures
getmorefigures.php?uid=PMC3927963&req=5

fig3-2041731414524758: Proliferative activity of CD3+ spleen cells cocultured with undifferentiated and 5-aza-CdR-treated myogenic-induced MSCs. (a) To assess proliferation, the CFSE-labeled responding spleen cells (1 × 106) from C57BL/6 mice were cultured alone with PHA as positive control or was cultured alone with Mit C as negative control or cocultured with Mit C–treated allogeneic MSCs either myogenic induced (5-aza-CdR-treated) or undifferentiated with PHA. Triplicate cultures were incubated at 37 °C for 5 days. On day 5 of culture, the cells were harvested for staining with anti-CD3 APC. Samples were acquired on a fluorescence-activated cell sorting calibur flow cytometer and data were gated on CD3+ events. CFSE analyses were determined using FlowJo software. Numbers in histograms represent percent divided cells. (b) Precursor frequency of T lymphocytes in the presence of PHA was significantly greater than undifferentiated cells (0µM 5-aza-CdR, p < 0.05), whereas there was no significant difference between undifferentiated cells and those treated with 5-aza-CdR: 0 µM, 0.5 µM, and 3 µM (106 MSCs per treatment group). Data are representative of six independent experiments. Preliminary analysis of data for panels A and B was performed using the proliferation algorithm in the FlowJo software. Statistical analysis was determined by a one-way ANOVA followed by a Dunn’s multiple comparisons test.5-Aza-CdR: 5-aza-2′-deoxycytidine; MSC: mesenchymal stem cell; CFSE: carboxyfluorescein diacetate succinimidyl ester; PHA: phytohemagglutinin; APC: allophycocyanin; Mit C: mitomycin C; ANOVA: analysis of variance; ns: not significant.*p < 0.05.
Mentions: The ability of MSCs to inhibit activation of lymphocytes by allogeneic spleen cells from C57BL/6 mice and PHA was examined in a CFSE cell proliferation assay. Undifferentiated and 5-aza-CdR-treated MSCs were tested in this assay. Mit C–treated lymphocytes did not proliferate at all (negative control, Figure 3(a)). Stimulation with PHA induced obvious proliferation of lymphocytes (positive control, Figure 3(a)). Both undifferentiated and 3 µM 5-aza-CdR-treated MSCs induced exerted dose-dependent inhibition of lymphocyte activation (Figure 3(a)). MSCs, either undifferentiated or 3 µM 5-aza-CdR-treated MSCs, completely inhibited the proliferation of PHA-stimulated lymphocytes independently for a ratio MSCs:splenocytes equal to 1:1. By contrast, 0.5 µM 5-aza-CdR-treated MSCs did not inhibit lymphocyte activation until the 1:1 ratio.

Bottom Line: Therefore, our aim was to evaluate the effects of mesenchymal stem cell differentiation on the immune characteristics of cells in vitro.Two different 5-aza-2'-deoxycytidine doses (0.5 and 3 µM) were evaluated for their effects on mesenchymal stem cell skeletal myogenic differentiation potential, immune antigen expression, and mixed lymphocytic reactions.Myogenic-induced mesenchymal stem cells do not elicit alloreactive lymphocyte proliferative responses and are able to modulate immune responses.

View Article: PubMed Central - PubMed

Affiliation: Wake Forest Institute for Regenerative Medicine, Wake Forest University Health Sciences, Winston-Salem, NC, USA ; Biomedical Research Institute, Joint Institute for Regenerative Medicine, Kyungpook National University Hospital, Daegu, Republic of Korea.

ABSTRACT
Cell therapy for patients who have intractable muscle disorders may require highly regenerative cells from young, healthy allogeneic donors. Mesenchymal stem cells are currently under clinical investigation because they are known to induce muscle regeneration and believed to be immune privileged, thus making them suitable for allogeneic applications. However, it is unclear whether allogeneic and myogenic-induced mesenchymal stem cells retain their immunomodulatory characteristics. Therefore, our aim was to evaluate the effects of mesenchymal stem cell differentiation on the immune characteristics of cells in vitro. We investigated the immunologic properties of mesenchymal stem cells after myogenic induction. Mesenchymal stem cells were obtained from C57BL/6 mice and the C3H/10T1/2 murine mesenchymal stem cell line. Two different 5-aza-2'-deoxycytidine doses (0.5 and 3 µM) were evaluated for their effects on mesenchymal stem cell skeletal myogenic differentiation potential, immune antigen expression, and mixed lymphocytic reactions. Using a mixed lymphocytic reaction, we determined the optimal splenocyte proliferation inhibition dose. The induction of regulatory T cells was markedly increased by the addition of 3 µM 5-aza-2'-deoxycytidine-treated mesenchymal stem cells. Myogenic-induced mesenchymal stem cells do not elicit alloreactive lymphocyte proliferative responses and are able to modulate immune responses. These findings support the hypothesis that myogenic-induced mesenchymal stem cells may be transplantable across allogeneic barriers.

No MeSH data available.


Related in: MedlinePlus