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Infiltration of circulating myeloid cells through CD95L contributes to neurodegeneration in mice.

Gao L, Brenner D, Llorens-Bobadilla E, Saiz-Castro G, Frank T, Wieghofer P, Hill O, Thiemann M, Karray S, Prinz M, Weishaupt JH, Martin-Villalba A - J. Exp. Med. (2015)

Bottom Line: We show that DN death is not mediated by CD95-induced apoptosis because deletion of CD95 in DNs does not influence MPTP-induced neurodegeneration.In contrast, deletion of CD95L in peripheral myeloid cells significantly protects against MPTP neurotoxicity and preserves striatal dopamine levels.Altogether, this study emphasizes the role of the peripheral innate immune response in neurodegeneration and identifies CD95 as potential pharmacological target for neurodegenerative disease.

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Affiliation: Division of Molecular Neurobiology, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany.

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Pharmacological neutralization of CD95L protects mice against MPTP toxicity and alters peripheral immune response. (A) Scheme of MPTP and APG112 treatment. (B) Quantification of total TH+ DNs in the SNpc at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: control versus MPTP: *, P < 0.001; MPTP versus MPTP + APG112: †, P < 0.01. (C) Representative images of TH+ neurons in SNpc of control and APG112-treated mice. Bar, 100 µm. (D) Quantification of striatal DA levels using HPLC at day 6 after last administration of saline or MPTP to control or APG112-treated mice. (E) Calculation of the metabolite ratio [(DOPAC + HVA/DA) × 100] after quantification of striatal DA metabolite levels by HPLC. (D and E) Data are presented as mean ± SEM; n = 8. ANOVA on ranks, Student–Newman–Keuls multiple comparison: *, P < 0.05; **, P < 0.01; ***, P < 0.001. (F) HPLC measurement of striatal MPP+ levels in WT and WT + APG112 mice at 90 min after MPTP injection. Data are presented as mean ± SEM; n = 3. Student’s t test: n.s., not significant. (G) Representative dot plots of blood monocytes and gating scheme of flow cytometry. (H) Quantification of blood monocyte subsets by FACS at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: *, P < 0.05. (B and H) Data are presented as dot plot with median and were pooled from two independent experiments; n = 16–17.
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fig3: Pharmacological neutralization of CD95L protects mice against MPTP toxicity and alters peripheral immune response. (A) Scheme of MPTP and APG112 treatment. (B) Quantification of total TH+ DNs in the SNpc at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: control versus MPTP: *, P < 0.001; MPTP versus MPTP + APG112: †, P < 0.01. (C) Representative images of TH+ neurons in SNpc of control and APG112-treated mice. Bar, 100 µm. (D) Quantification of striatal DA levels using HPLC at day 6 after last administration of saline or MPTP to control or APG112-treated mice. (E) Calculation of the metabolite ratio [(DOPAC + HVA/DA) × 100] after quantification of striatal DA metabolite levels by HPLC. (D and E) Data are presented as mean ± SEM; n = 8. ANOVA on ranks, Student–Newman–Keuls multiple comparison: *, P < 0.05; **, P < 0.01; ***, P < 0.001. (F) HPLC measurement of striatal MPP+ levels in WT and WT + APG112 mice at 90 min after MPTP injection. Data are presented as mean ± SEM; n = 3. Student’s t test: n.s., not significant. (G) Representative dot plots of blood monocytes and gating scheme of flow cytometry. (H) Quantification of blood monocyte subsets by FACS at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: *, P < 0.05. (B and H) Data are presented as dot plot with median and were pooled from two independent experiments; n = 16–17.

Mentions: To test this hypothesis, we treated WT mice with a stable CD95-Fc fusion protein that neutralizes CD95 activity and does not cross the BBB (CD95-Fc hereafter referred to as APG112; Fig. 3 A). Accordingly, APG112 was not detected in the brain tissue of MPTP-treated mice but only endovascular in the brain of MPTP + APG–treated mice (not depicted). Thus, the CD95-Fc main site of action is in the periphery, and therefore, it can be used to distinguish between the contribution of neurodegeneration of peripheral myeloid cells and resident microglia. 6 d after the last MPTP injection, we analyzed brains and blood of saline- and MPTP-treated mice. Mice that received saline showed a significant reduction of DNs upon MPTP intoxication, whereas, similar to CD95Lf/f;LysMcre mice, mice that had been systemically treated with APG112 were resistant to MPTP-induced degeneration of SNpc DNs. Altogether, these data demonstrate that CD95L neutralization is neuroprotective in a mouse model of DN degeneration (Fig. 3, B and C).


Infiltration of circulating myeloid cells through CD95L contributes to neurodegeneration in mice.

Gao L, Brenner D, Llorens-Bobadilla E, Saiz-Castro G, Frank T, Wieghofer P, Hill O, Thiemann M, Karray S, Prinz M, Weishaupt JH, Martin-Villalba A - J. Exp. Med. (2015)

Pharmacological neutralization of CD95L protects mice against MPTP toxicity and alters peripheral immune response. (A) Scheme of MPTP and APG112 treatment. (B) Quantification of total TH+ DNs in the SNpc at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: control versus MPTP: *, P < 0.001; MPTP versus MPTP + APG112: †, P < 0.01. (C) Representative images of TH+ neurons in SNpc of control and APG112-treated mice. Bar, 100 µm. (D) Quantification of striatal DA levels using HPLC at day 6 after last administration of saline or MPTP to control or APG112-treated mice. (E) Calculation of the metabolite ratio [(DOPAC + HVA/DA) × 100] after quantification of striatal DA metabolite levels by HPLC. (D and E) Data are presented as mean ± SEM; n = 8. ANOVA on ranks, Student–Newman–Keuls multiple comparison: *, P < 0.05; **, P < 0.01; ***, P < 0.001. (F) HPLC measurement of striatal MPP+ levels in WT and WT + APG112 mice at 90 min after MPTP injection. Data are presented as mean ± SEM; n = 3. Student’s t test: n.s., not significant. (G) Representative dot plots of blood monocytes and gating scheme of flow cytometry. (H) Quantification of blood monocyte subsets by FACS at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: *, P < 0.05. (B and H) Data are presented as dot plot with median and were pooled from two independent experiments; n = 16–17.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC4387281&req=5

fig3: Pharmacological neutralization of CD95L protects mice against MPTP toxicity and alters peripheral immune response. (A) Scheme of MPTP and APG112 treatment. (B) Quantification of total TH+ DNs in the SNpc at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: control versus MPTP: *, P < 0.001; MPTP versus MPTP + APG112: †, P < 0.01. (C) Representative images of TH+ neurons in SNpc of control and APG112-treated mice. Bar, 100 µm. (D) Quantification of striatal DA levels using HPLC at day 6 after last administration of saline or MPTP to control or APG112-treated mice. (E) Calculation of the metabolite ratio [(DOPAC + HVA/DA) × 100] after quantification of striatal DA metabolite levels by HPLC. (D and E) Data are presented as mean ± SEM; n = 8. ANOVA on ranks, Student–Newman–Keuls multiple comparison: *, P < 0.05; **, P < 0.01; ***, P < 0.001. (F) HPLC measurement of striatal MPP+ levels in WT and WT + APG112 mice at 90 min after MPTP injection. Data are presented as mean ± SEM; n = 3. Student’s t test: n.s., not significant. (G) Representative dot plots of blood monocytes and gating scheme of flow cytometry. (H) Quantification of blood monocyte subsets by FACS at day 6 after last administration of saline or MPTP of control or APG112-treated mice. ANOVA followed by Newman–Keuls post-hoc test: *, P < 0.05. (B and H) Data are presented as dot plot with median and were pooled from two independent experiments; n = 16–17.
Mentions: To test this hypothesis, we treated WT mice with a stable CD95-Fc fusion protein that neutralizes CD95 activity and does not cross the BBB (CD95-Fc hereafter referred to as APG112; Fig. 3 A). Accordingly, APG112 was not detected in the brain tissue of MPTP-treated mice but only endovascular in the brain of MPTP + APG–treated mice (not depicted). Thus, the CD95-Fc main site of action is in the periphery, and therefore, it can be used to distinguish between the contribution of neurodegeneration of peripheral myeloid cells and resident microglia. 6 d after the last MPTP injection, we analyzed brains and blood of saline- and MPTP-treated mice. Mice that received saline showed a significant reduction of DNs upon MPTP intoxication, whereas, similar to CD95Lf/f;LysMcre mice, mice that had been systemically treated with APG112 were resistant to MPTP-induced degeneration of SNpc DNs. Altogether, these data demonstrate that CD95L neutralization is neuroprotective in a mouse model of DN degeneration (Fig. 3, B and C).

Bottom Line: We show that DN death is not mediated by CD95-induced apoptosis because deletion of CD95 in DNs does not influence MPTP-induced neurodegeneration.In contrast, deletion of CD95L in peripheral myeloid cells significantly protects against MPTP neurotoxicity and preserves striatal dopamine levels.Altogether, this study emphasizes the role of the peripheral innate immune response in neurodegeneration and identifies CD95 as potential pharmacological target for neurodegenerative disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Molecular Neurobiology, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany.

Show MeSH
Related in: MedlinePlus