A Novel Immunosuppressor, (5R)-5-Hydroxytriptolide, Alleviates Movement Disorder and Neuroinflammation in a 6-OHDA Hemiparkinsonian Rat Model
Su Ruijun2,3,4, Sun Min2,3,4, Wang Wei2,3,4, Zhang Jianliang2,3, Zhang Li2,3,4, Zhen Junli2,3,4, Qian Yanjing2,3,4, Zheng Yan1,3,4,*, Wang Xiaomin2,3,4,*
1Department of Physiology, 2Department of Neurobiology, and 3Key Laboratory for Neurodegenerative Disorders of the Ministry of Education, Capital Medical University, Beijing 100069, China. 4Beijing Institute for Brain Disorders, Beijing100069, China.
Parkinson’s disease (PD) is one of the most common age-related neurodegenerative diseases. Promising therapies for PD still need to be explored. Immune dysfunction has been found to be involved in PD pathogenesis. Here, a novel immunosuppressor, (5R)-5-hydroxytriptolide (LLDT8), was used to treat 6-hydroxydopamine (6-OHDA)-induced hemiparkinson rats. We found that oral administration of LLDT8 significantly alleviated apomorphine-induced rotations at a dose of 125 µg/kg, and improved performance in cylinder and rotarod tests at a lower dose of 31.25 µg/kg, in 6-OHDA hemiparkinsonian rats. Moreover, loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) of the 6-OHDA rat was attenuated in response to LLDT8 treatment in a dose-dependent manner. In addition, inflammatory factors IL-1β, IL-6 and TNF-α, were significantly inhibited in LLDT8-treated hemiparkisonian rats, compared with vehicle. Notably, the level of dopamine (DA) in the striatum of PD rats was restored by LLDT8 treatment. Furthermore, we also detected that the disequilibrium of peripheral lymphocytes was reversed by LLDT8 administration. Taken together, the results imply that the immunosuppressor, LLDT8, can rescue dopaminergic neurodegeneration in 6-OHDA hemiparkinsonian rats, thus providing a potential therapeutic strategy for PD.
Su Ruijun,Sun Min,Wang Wei, et al. A Novel Immunosuppressor, (5R)-5-Hydroxytriptolide, Alleviates Movement Disorder and Neuroinflammation in a 6-OHDA Hemiparkinsonian Rat Model[J]. Aging and disease,
2017, 8(1): 31-43.
Figure 1. Chemical structure of (5R)-5-hydroxytriptolide. Formula: C20H24O7. Molecular weight: 376.39.
Figure 2. LLDT8 administration enhanced movement performance in a 6-OHDA rat model. (A) The schedule of experimental presentation. (B-D) The behavioral performance of the PD rats was determined by apomorphine-induced rotations (B), rotarod (C) and cylinder test (D). (E) Representative graph of the open field test. (F-J) Fp (Floor plane) -total movement distance (cm) (F), Fp-rest time (s) (G), Fp-velocity (cm/s) (H), Fp-moves (times) (I) and Vp(Vertical plane)-total movement distance (cm) (J) were recorded in the open field test. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham, *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, or a Tamhane’s T2 test when variances were not equal, n=8-9.
Figure 3. PD-like pathology in the 6-OHDA rat model was attenuated by LLDT8 treatment. (A) IHC staining for TH+ neurons in SNpc (as indicated by the dotted bracket). Scale bar = 500 μm. (B) The ratio (left/right) of stereological quantification of numbers of TH+ neurons in SNpc after systemic administration of different doses of LLDT8 (31.25, 62.5 and 125 μg/kg) or rasagiline, for 5 weeks. (C) IHC staining for TH+ fibers in striatum (STR) treated with LLDT8 for 5 weeks. Scale bar = 2.0 mm. (D) The ratio (left/right) of TH+ fiber density in STR after systemic administration of different doses of LLDT8 (31.25, 62.5, and 125 μg/kg) or rasagiline, for 5 weeks. (E) The scan picture of TH+ protein and (F) the statistical TH+ protein results of all experimental groups treated with LLDT8. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham, *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, n=4-6.
Figure 4. The level of DA in striatum of the PD rat model was restored by LLDT8 treatment in a dose-dependent manner. The model rats were treated by different doses of LLDT8 (31.25, 62.5, and 125 μg/kg) or Rasagiline (0.3mg/㎏) once daily for 5 weeks. (A) The concentrations of DA, and (B) its metabolite dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) in the striatum of all experimental rats were measured by HPLC, the metabolic rate of DA was calculated and used in statistical analysis. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham, *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, n=8-9.
Figure 5. The glial response in the PD rat model treated with LLDT8. The representative IHC staining for Mac1+ microglia (A) and GFAP+ astrocytes (C) in SNpc. Scale bar = 500 μm. The quantitative percentage of area occupied by Mac1+ cells (B) and GFAP+ cells (D) in the indicated region of SNpc. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham, *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, n=4-6.
Figure 6. Dose-dependent effects of LLDT8 on inflammation factors in the PD rat model. The concentrations of (A) IL-1β, (B) IL-6 and (C) TNF-α in ipsilateral striatum of all experimental animals. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham, *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, n=8-9.
Figure 7. Treatment of LLDT8 balanced the peripheral immune cells of the PD rat model. (A-D) Gating strategy is represented for each cell type assessed using flow cytometry. Representative flow cytometry plots showing the lymphocytes subsets in PD rat blood are shown. P1 is represented for total lymphocytes (A); P3: T lymphocytes (B); Q1-1: B cells (C); Q1: natural killer (NK) cells (D). (E) The counts of total lymphocytes and T cells, (F) the counts of NK and B cells were statistically analyzed. Data are shown as mean ± SEM. #P < 0.05, ##P < 0.01 vs. sham; *P < 0.05, **P < 0.01 vs. vehicle using one-way ANOVA with post hoc LSD t-test, n=8-9.
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