Fig. 8

Adoptive transfer of Treg diminishes neuroinflammation in the nigrostriatal tract of hαSyn PD mice; CD28SA decreases early T cell activation in brain and modulates early IL-2 response in peripheral lymphatic organs of hαSyn PD mice; delayed treatment with CD28SA fails to reduce SN dopaminergic neurodegeneration in hαSyn PD mice. A-D Immunohistochemical stainings and quantification for CD4⁺ (A, B) and CD8⁺ (C, D) T cells in the SN and striatum of hαSyn-injected mice with either PBS or Treg delivery. Statistical analysis by unpaired two-tailed t-test: SN CD4⁺: t(14) = 2.367, P = 0.0329; striatum CD4⁺: t(14) = 2.601, P = 0.0209; SN CD8⁺: t(14) = 2.835, P = 0.0132; striatum CD8⁺: t(14) = 5.2, P = 0.0001. *P < 0.05, **P < 0.01, ***P < 0.001. Scale bars: 20 µm each. E Representative FACS analysis images showing the percentages of CD8⁺CD69⁺ T cells in brain of EV- or hαSyn-injected mice with either PBS or with CD28SA treatment, ten days after AAV injection (3 days after PBS or CD28SA delivery). F Percentages of CD8⁺CD69⁺ in brain as analyzed by FACS. G, H Concentrations of IL-2 in CLN (G) and spleen (H) of EV- or hαSyn-injected mice treated either with PBS or with CD28SA. Statistical analysis by one-way ANOVA followed by Tukey’s multiple comparisons test: CD8⁺CD69⁺ brain, F(3, 20) = 4.073, P = 0.0207); IL-2 CLN, F(3, 20) = 8.186, P = 0.0009; statistical analysis by Kruskal–Wallis test followed by Dunn’s multiple comparisons test: IL-2 spleen, H = 12.7, P = 0.0053. *P < 0.05, **P < 0.01. Data are shown as mean ± SEM. n = number of biologically independent samples, each sample consisting of pools of two animals. I-K, Representative images of TH and Nissl-stained neurons in the SN of the indicated groups of mice (I) and estimated cell number by unbiased stereology for TH⁺ dopaminergic SN neurons (J) and Nissl⁺ total SN neurons (K). Statistical analysis by unpaired two-tailed t-test. All data are shown as mean ± SEM. n = number of biologically independent animals. Scale bar: 50 µm