HMGB1 is involved in autophagy inhibition caused by SNCA/α-synuclein overexpression

<p>SNCA/α-synuclein and its rare mutations are considered as the culprit proteins in Parkinson disease (PD). Wild-type (WT) SNCA has been shown to impair macroautophagy in mammalian cells and in transgenic mice. In this study, we monitored the dynamic changes in autophagy process and confirmed that overexpression of both WT and SNCA^A53T inhibits autophagy in PC12 cells in a time-dependent manner. Furthermore, we showed that SNCA binds to both cytosolic and nuclear high mobility group box 1 (HMGB1), impairs the cytosolic translocation of HMGB1, blocks HMGB1<b>-</b>BECN1 binding, and strengthens BECN1<b>-</b>BCL2 binding. Deregulation of these molecular events by SNCA overexpression leads to autophagy inhibition. Overexpression of BECN1 restores autophagy and promotes the clearance of SNCA. siRNA knockdown of <i>Hmgb1</i> inhibits basal autophagy and abolishes the inhibitory effect of SNCA on autophagy while overexpression of HMGB1 restores autophagy. Corynoxine B, a natural autophagy inducer, restores the deficient cytosolic translocation of HMGB1 and autophagy in cells overexpressing SNCA, which may be attributed to its ability to block SNCA<b>-</b>HMGB1 interaction. Based on these findings, we propose that SNCA<b>-</b>induced impairment of autophagy occurs, in part, through HMGB1, which may provide a potential therapeutic target for PD.</p

LC–MS-Based Urinary Metabolite Signatures in Idiopathic Parkinson’s Disease

Increasing evidence has shown that abnormal metabolic phenotypes in body fluids reflect the pathogenesis and pathophysiology of Parkinson’s disease (PD). These body fluids include urine; however, the relationship between, specifically, urinary metabolic phenotypes and PD is not fully understood. In this study, urinary metabolites from a total of 401 clinical urine samples collected from 106 idiopathic PD patients and 104 normal control subjects were profiled by using high-performance liquid chromatography coupled to high-resolution mass spectrometry. Our study revealed significant correlation between clinical phenotype and urinary metabolite profile. Metabolic profiles of idiopathic PD patients differed significantly and consistently from normal controls, with related metabolic pathway variations observed in steroidogenesis, fatty acid beta-oxidation, histidine metabolism, phenylalanine metabolism, tryptophan metabolism, nucleotide metabolism, and tyrosine metabolism. In the fruit fly <i>Drosophila melanogaster</i>, the alteration of the kynurenine pathway in tryptophan metabolism corresponded with pathogenic changes in the alpha-synuclein overexpressed <i>Drosophila</i> model of PD. The results suggest that LC–MS-based urinary metabolomic profiling can reveal the metabolite signatures and related variations in metabolic pathways that characterize PD. Consistent PD-related changes across species may provide the basis for understanding metabolic regulation of PD at the molecular level

Effects of Huanglian-Jie-Du-Tang and Its Modified Formula on the Modulation of Amyloid-β Precursor Protein Processing in Alzheimer's Disease Models

<div><p>Huanglian-Jie-Du-Tang (HLJDT) is a famous traditional Chinese herbal formula that has been widely used clinically to treat cerebral ischemia. Recently, we found that berberine, a major alkaloid compound in HLJDT, reduced amyloid-β (Aβ) accumulation in an Alzheimer’s disease (AD) mouse model. In this study, we compared the effects of HLJDT, four single component herbs of HLJDT (Rhizoma coptidis (RC), Radix scutellariae (RS), Cortex phellodendri (CP) and Fructus gardenia (FG)) and the modified formula of HLJDT (HLJDT-M, which is free of RS) on the regulatory processing of amyloid-β precursor protein (APP) in an <i>in vitro</i> model of AD. Here we show that treatment with HLJDT-M and its components RC, CP, and the main compound berberine on N2a mouse neuroblastoma cells stably expressing human APP with the Swedish mutation (N2a-SwedAPP) significantly decreased the levels of full-length APP, phosphorylated APP at threonine 668, C-terminal fragments of APP, soluble APP (sAPP)-α and sAPPβ-Swedish and reduced the generation of Aβ peptide in the cell lysates of N2a-SwedAPP. HLJDT-M showed more significant APP- and Aβ- reducing effects than berberine, RC or CP treatment alone. In contrast, HLJDT, its component RS and the main active compound of RS, baicalein, strongly increased the levels of all the metabolic products of APP in the cell lysates. The extract from FG, however, did not influence APP modulation. Interestingly, regular treatment of TgCRND8 APP transgenic mice with baicalein exacerbated the amyloid plaque burden, APP metabolism and Aβ production. Taken together, these data provide convincing evidence that HLJDT and baicalein treatment can increase the amyloidogenic metabolism of APP which is at least partly responsible for the baicalein-mediated Aβ plaque increase in the brains of TgCRND8 mice. On the other hand, HLJDT-M significantly decreased all the APP metabolic products including Aβ. Further study of HLJDT-M for therapeutic use in treating AD is warranted.</p></div

Extracts of HLJDT constituent herbs alter the processing of APP in N2a-SwedAPP cells.

<p>Conditioned medium and cell lysates were prepared from N2a-SwedAPP cells that were treated with (A) RC, (B) CP, (C) FG or (D) RS at various doses as indicated for 48 h. Western blotting was used to detect sAPPα and sAPPβ-sw in conditioned medium, and to detect Fl-APP, pAPPThr668 and β-actin in cell lysates. Bars represent mean±S.E.M. for three experiments. One-way ANOVA revealed significant differences due to treatment at various doses of components of HLJDT: *p<0.05; **p<0.01; ***p<0.001.</p

Modulation of APP processing by HLJDT and HLJDT-M in N2a-SwedAPPcells.

<p>Conditioned medium and cell lysates were prepared from N2a-SwedAPP cells that were treated with (A) HLJDT or (B) HLJDT-M at various doses as indicated for 48 h. Western blotting was used to detect sAPPα and sAPPβ-sw in conditioned medium, and to detect Fl-APP, pAPPThr668, CTFs and β-actin in cell lysates. Bars represent mean±S.E.M. for three experiments. One-way ANOVA revealed significant differences due to treatment at various doses of HLJDT or HLJDT-M: *p<0.05; **p<0.01; ***p<0.001.</p

Selected components of Huang-Liang-Jie-Du-Tang (HLJDT).

<p>A. Chemical structures of main compounds of Huang-Liang-Jie-Du-Tang (HLJDT). B. Quality analysis of HLJDT (at 245 nm) (A); Fructus gardeniae (FG) (at 245 nm) (B); Rhizoma coptidis (FG) (at 275 nm) (C); Radix scutellariae (RS) (at 275 nm) (D); Cortex phellodendri (CP) (at 275 nm) (E). Peaks of these compounds are shown: 1. Geniposide; 2. Berberine; 3. Palmatine; 4. Baicalein; 5. Baicalin; 6. Wogonin.</p

Regulation of the levels of intracellular Aβ by HLJDT, HLJDT-M and its components in cultured N2a-SwedAPP cells.

<p>Aβ1–40 and Aβ1–42 peptides were analyzed by enzyme-linked immunosorbent assay (ELISA) in N2a-SwedAPP cell lysates 48 h after addition of (A) RC or CP; (B) RS; (C) HLJDT or HLJDT-M; or (D) berberine or baicalein. Bars represent mean±S.E.M. of the level of intracellular Aβ1–40 or Aβ1–42 peptides in three experiments relative to DMSO control (untreated). One-way ANOVA revealed significant differences due to treatment: *p<0.05; **p<0.01; ***p<0.001.</p