Orexin B protects dopaminergic neurons from 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced neurotoxicity associated with reduced extracellular signal-regulated kinase phosphorylation.

The loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) is a major pathological hallmark of Parkinson's disease (PD). Orexin B (OXB) has been reported to promote the growth of DA neurons. However, the roles of OXB in the degeneration of DA neurons still remained not fully clear. An in vivo PD model was constructed by administrating 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in mice. Pole test was performed to investigate the motor function of mice and the number of DA neurons was detected by immunofluorescence (IF). A PD cell model was established by treating SH-SY5Y cells with 1-methyl-4-phenylpyridinium (MPP+). OXB was added to the culture medium 2 h after MPP + treatment. Microscopic analysis was carried out to investigate the function of OXB in the cell model of PD 24 h after MPP + challenge. RNA-Seq analysis of the PD cell model was performed to explore the possible mechanisms. Western blot was used to detect the phosphorylation levels of extracellular signal-regulated kinase (ERK). OXB significantly decreased the DA neurons death caused by MPTP, alleviated MPP+-induced neurotoxicity in SH-SY5Y cells, and robustly enhanced the weight and motor ability of PD mice. Besides, RNA-Seq analysis demonstrated that the mitogen-activated protein kinase (MAPK) pathway was involved in the pathology of PD. Furthermore, MPP + led to increased levels of phosphorylation of ERK (p-ERK), OXB treatment significantly decreased the levels of p-ERK in MPP+-treated SH-SY5Y cells. This study demonstrated that OXB exerts a neuroprotective role associated with reduced ERK phosphorylation in the PD model. This suggests that OXB may have therapeutic potential for treatment of PD.

Ma X ，Cao F ，Cui J ，Li X ，Yin Z ，Wu Y ，Wang Q ... -  《-》

Erratum: Eyestalk Ablation to Increase Ovarian Maturation in Mud Crabs.

《Jove-Journal of Visualized Experiments》

TMEM106B Knockdown Exhibits a Neuroprotective Effect in Parkinson's Disease via Decreasing Inflammation and Iron Deposition.

Parkinson's disease (PD) is closely related to iron accumulation and inflammation. Emerging evidence indicates that TMEM106B plays an essential role in PD. But whether TMEM106B could act on neuroinflammation and iron metabolism in PD has not yet been investigated. The aim of this study was to investigate the pathological mechanisms of inflammation and iron metabolism of TMEM106B in PD. 1-methyl-4-phenylpyridinium (MPP+)- and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced SH-SY5Y cells and mice were treated with LV-shTMEM106B and AAV-shTMEM106B to construct PD cellular and mouse models. Pole tests and open-field test (OFT) were performed to evaluate the locomotion of the mice. Immunohistochemistry and iron staining were used to detect TH expression and iron deposition in the SN. Iron staining was used to measure the levels of iron. Western blotting was used to detect the expression of inflammatory factors (tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6)), NOD-like receptor protein 3 (NLRP3) inflammasome, divalent metal transporter 1 (DMT1), and Ferroportin1 (FPN1)). Knockdown of TMEM106B improved motor ability and rescued dopaminergic (DA) neuron loss. TMEM106B knockdown attenuated the increases of TNF-α, IL-6, NLRP3 inflammasome, and DMT1 expression in the MPP+ and MPTP-induced PD models. Furthermore, TMEM106B knockdown also increases the expression of FPN1. This study provides the first evidence that knockdown of TMEM106B prevents dopaminergic neurodegeneration by modulating neuroinflammation and iron metabolism.

Liu Y ，Qin K ，Jiang C ，Gao J ，Hou B ，Xie A ... -  《-》

Atractylenolide-I Attenuates MPTP/MPP(+)‑Mediated Oxidative Stress in Parkinson's Disease Through SIRT1/PGC‑1α/Nrf2 Axis.

Parkinson's disease (PD) is typically marked by motor dysfunction accompanied by loss of dopaminergic (DA) neurons and aggravated oxidative stress in the substantia nigra pars compacta (SNpc). Atractylenolide-I (ATR-I) is a potent antioxidant sesquiterpene with neuroprotective properties. However, whether ATR-I plays a neuroprotective role against oxidative stress in PD remains unclear. The objective of this study was to explore the mechanism of antioxidant action of ATR-I in PD models both in vivo and in vitro. Here, we show that ATR-I alleviated motor deficits in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mice. Moreover, ATR-I treatment effectively reduced DA neuron loss and increased tyrosine hydroxylase expression in the SNpc of MPTP-induced mice. Additionally, ATR-I inhibited oxidative stress (as manifested by elevated superoxide dismutase and glutathione peroxidase activities, and reduced malondialdehyde content) in MPTP-induced mice and attenuated reactive oxygen species levels in 1-methyl-4-phenylpyridinum (MPP+)-treated SH-SY5Y cells. Finally, ATR-I upregulated expressions of silent information regulator 1 (SIRT1), peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), NF-E2-related factor-2 (Nrf2), and heme oxygenase-1 in MPTP-induced mice and MPP+-treated SH-SY5Y cells, but had little effect on these factors in the presence of the SIRT1 inhibitor EX527. Taken together, these findings indicated that the important antioxidant role of ATR-I in MPTP/MPP+-mediated oxidative stress and the pathogenesis of PD through the SIRT1/PGC-1α/Nrf2 axis, highlighting its potential as a therapeutic option for PD.

Gao Y ，Li S ，Zhang S ，Zhang Y ，Zhang J ，Zhao Y ，Chang C ，Gao X ，Chen L ，Yang G ... -  《-》

Uncaria rhynchophylla alkaloid extract exerts neuroprotective activity against Parkinson's disease via activating mitophagy with the involvement of UCHL1.

Uncaria rhynchophylla (Miq.) Miq. ex Havil. (UR), a traditional Chinese medicinal plant, plays an active role in neuroprotection. Clinical medication and modern pharmacological studies have proved the efficacy of UR against Parkinson's disease (PD), with alkaloids being recognized as the main bioactive components. But the therapeutic effect and mechanism of U. rhynchophylla alkaloid extract (URA) against PD need further exploration. The study aimed to investigate the therapeutic effect and potential mechanism of URA on PD. LC-MS methodology was used to evaluate the chemical constituents of URA. The anti-PD activity of URA in vivo was measured on the mouse and rat models of PD. Neuroprotective effect of URA on PC12 cells was measured by MTT assay. Dopamine (DA) and its metabolites were detected by LC-MS for probing the protection ability on dopaminergic neurons. The differentially expressed proteins between model group and URA administrated group were analyzed by proteomics, suggesting oxidative phosphorylation as possible pathway of URA. Considering the critical role of mitochondria in oxidative phosphorylation, JC-1 staining, MitoSOX staining, transmission electron microscopy (TEM) observation and adenosine triphosphate (ATP) levels detection were used to analyze the effects of URA on mitochondrial morphology and function. Biolayer interferometry (BLI) was used to search for the possible UCHL1-bonding compounds in URA. URA significantly mitigated the behavioural defects by improving coordination, shortening the time to climb down the whole pole (T-LA) and increasing the forelimbs' muscle strength of MPTP-induced PD mice and 6-OHDA-induced PD rats. In addition, URA improved tyrosine hydroxylase expression in dopaminergic neurons by immunohistochemistry (IHC) staining, and thus increased the neurotransmitter levels of DA and relevant metabolites. Furthermore, URA promoted mitophagy as reflected by a significant decrease in reactive oxygen species (ROS) generation, an increase in ATP levels and clearance of damaged mitochondria. Subsequently, Ubiquitin C-terminal hydrolase 1 (UCHL1), which is associated with the mitochondrial dysfunction in PD, is suggested to be a promising target based on the proteomics result, and proved by the blocked protective effects of URA by UCHL1 inhibitor. Furthermore, hirsuteine (HTE) was identified as a potential active compound of URA binding to UCHL1 by BLI, and the binding capacity and site were verified by surface plasmon resonance (SPR) and molecular docking. This work demonstrates that URA exerts effective neuroprotective activity against PD via activation of mitophagy with the involvement of UCHL1, and HTE may be a potential active compound of URA.

Hou X ，Liang X ，Zhao X ，Shi Y ，Zhuo F ，Tong X ，Yang X ，Zhai Q ，Wang J ，Guo Q ，Tu P ，Zeng K ，Zhang Q ... -  《-》