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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and <t>CD3+CD8+</t> T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.
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The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and CD3+CD8+ T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.

Journal: International Journal of Molecular Sciences

Article Title: Proinflammatory Cytokine Preconditioning Enhances the Therapeutic Potency of Different Types of MSCs in Inflammation

doi: 10.3390/ijms27094090

Figure Lengend Snippet: The effects of naïve and Cytomix-stimulated MSCs on T cell proliferation. The gating strategy is shown in . ( A – C ) BM-MSCs, iMSCs WT and iMSCs B2M KO inhibition on the expansion of CD3+, CD3+CD4+ and CD3+CD8+ T cells via CM. Cytomix enhanced the three types of MSCs’ inhibition on the EI of T cells. ( D – F ) All types of MSCs significantly inhibited the division of CD3+, CD3+CD4+ and CD3+CD8+ T cells when co-cultured with PBMCs, in a dose-dependent manner. ( G – I ) All of the MSCs, naïve or Cytomix stimulated, significantly reduced the EI of CD3+, CD3+CD4+ or CD3+CD8+ T cells, at the lowest ratio of MSCs, with iMSCs WT both naïve and Cytomix-stimulated presenting the strongest inhibition. ( J ) Co-culturing all types of MSCs, naïve or Cytomix-stimulated, increased the production of G-CSF in the media, with a dose-dependent effect. ( K ) All types of MSCs significantly decreased the level of IFN-γ at the ratios of 1:20 and 1:10, while the three types of naïve MSCs significantly decreased it at 1:50. ( L ) Naïve and Cytomix-stimulated iMSC WTs significantly increased the level of TNF-α at the ratios of 1:50, 1:20 and 1:10, while Cytomix-stimulated iMSC B2M KO significantly increased it at the ratio of 1:50 and 1:20. ( M ) Naïve BM-MSCs significantly increased the level of IL-10 production at the ratios of 1:20 and 1:10. Representative results of four independent experiments are shown. Results are shown as mean ± SD ( n = 3 in CM; n = 4 in co-culture, each group), and analysed by two-way ANOVA with Šídák’s multiple comparisons test to compare naïve vs. Cytomix in each type of MSCs (no significance in all of the settings), or Tukey’s multiple comparisons test to compare different ratios of MSCs to PBMCs in each MSC group, or Dunnett’s multiple comparisons test to compare different MSC groups vs. vehicle in the setting of the same ratio. */**/***/**** represent comparisons of each type of MSC vs. vehicle; #/## represent comparisons of different ratios of each cell type with different levels of p < 0.05/0.01. EI: expansion index; DP: division percentage.

Article Snippet: The PBMCs were stained with anti-human CD3-APC-Vio770, CD4-APC and CD8-PE-Vio770 antibodies (Cat# 130-113-136, 130-113-222, 130-110-680; Miltenyi Biotec) for 30 min on ice in the dark.

Techniques: Inhibition, Cell Culture, Co-Culture Assay