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. 2022 Jan 18;10(2):205. doi: 10.3390/biomedicines10020205

Figure 5.

Figure 5

CD4+ and CD8+ murine and human T-cell proliferation is significantly reduced by rArg-I. Doubling rates of murine splenocytes and human peripheral blood mononuclear cells (PBMCs) were analyzed utilizing Cell Trace Yellow and Violet labelling, respectively. Cell viability was checked 5–6 days after the onset of the experiment. (A,D,G,J) The proliferation of murine and human T cells are indicated as percentage of control (no vehicle or rArg-I incubation). (B,E,H,K) Corresponding representative histograms. (A,B) Murine T-cell activation resulted in CD4+ T-cell proliferation. rArg-I addition in concentrations of 500 ng/mL or more, reduced proliferation. n = 5 biological repeats. [F(1,21) = 37.65, p < 0.0001, two-way ANOVA]. (D,E) Supplementation of rArg-I suppressed murine CD8+ T-cell proliferation. n = 5 biological repeats. [F(1,21) = 28.06, p < 0.0001, two-way ANOVA]. (G,H) CD4+ human PBMC proliferation was ablated when rArg-I concentrations equal to or higher than 50 ng/mL were added. n = 5–7 biological repeats. [F(1,30) = 1356, p < 0.0001, two-way ANOVA]. (J,K) A similar reduction was found in the human CD8+ PBMC cell population. n = 5–7 biological repeats. [F(1,30) = 6124, p < 0.0001, two-way ANOVA]. In all conditions, the vehicle solution had no impact on the doubling rate. (C,F) rArg-I treatment did not affect murine T-cell survival. n = 5 biological repeats. (I) Human blood-derived CD4+ T cells showed significantly decreased viability when incubated with rArg-I compared to the vehicle when concentrations higher than 100 ng/mL were applied. n = 5–7 biological repeats. [F(1,30) = 34.63, p < 0.01, two-way ANOVA]. (L) Human CD8+ PBMCs showed no sign of decreased viability for rArg-I. n = 5–7 biological repeats. (C,F,I,L) The mean survival of control samples not exposed to vehicle or rArg-I is indicated by the dotted line. The cell viability is indicated as the percentage of FVD eFluor506-negative cells within the CD4+ or CD8+ T gates, respectively. The vehicle solution had no impact on the cell survival for both murine and human cells. Grey-filled bars are vehicle controls, black-filled bars represent rArg-I-treated cells. Data are shown as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.