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Abstract In spite of recent advances in the field of undernutrition, current dietary therapy relying on the supply of high protein high calorie formulas is still plagued with transient recovery of impaired organs resulting in significant relapse of cases. This is partly attributed to the inadequacy of current research models in recapitulating clinical undernutrition for mechanistic exploration. Using 1636 Macaca fascicularis monkeys, a human‐relevant criterion for determining undernutrition weight‐for‐age z‐score (WAZ), with a cutoff point of ≤ −1.83 is established as the benchmark for identifying undernourished nonhuman primates (U‐NHPs). In U‐NHPs, pathological anomalies in multi‐organs are revealed. In particular, severe dysregulation of hepatic lipid metabolism characterized by impaired fatty acid oxidation due to mitochondria dysfunction, but unlikely peroxisome disorder, is identified as the anchor metabolic aberration in U‐NHPs. Mitochondria dysfunction is typified by reduced mito‐number, accumulated long‐chain fatty acids, and disruption of OXPHOS complexes. Soy peptide‐treated U‐NHPs increase in WAZ scores, in addition to attenuated mitochondria dysfunction and restored OXPHOS complex levels. Herein, innovative criteria for identifying U‐NHPs are developed, and unknown molecular mechanisms of undernutrition are revealed hitherto, and it is further proved that soypeptide supplementation reprogramed mitochondrial function to re‐establish lipid metabolism balance and mitigated undernutrition.

Calcium-independent phospholipase A2β (iPLA2β), a member of the phospholipase A2 (PLA2s) superfamily, is encoded by the PLA2G6 gene. Mutations in the PLA2G6 gene have been identified as the primary cause of infantile neuroaxonal dystrophy (INAD) and, less commonly, as a contributor to Parkinson's disease (PD). Recent studies have revealed that iPLA2β deficiency leads to neuroinflammation, iron accumulation, mitochondrial dysfunction, lipid dysregulation, and other pathological changes, forming a complex pathogenic network. These discoveries shed light on potential mechanisms underlying PLA2G6-associated neurodegeneration (PLAN) and offer valuable insights for therapeutic development. This review provides a comprehensive analysis of the fundamental characteristics of iPLA2β, its association with neurodegeneration, the pathogenic mechanisms involved in PLAN, and potential targets for therapeutic intervention. It offers an overview of the latest advancements in this field, aiming to contribute to ongoing research endeavors and facilitate the development of effective therapies for PLAN.

Summary: The ribosome-associated protein quality control (RQC) pathway acts as a translational surveillance mechanism to maintain proteostasis. In mammalian cells, the cytoplasmic RQC pathway involves nuclear export mediator factor (NEMF)-dependent recruitment of the E3 ligase Listerin to ubiquitinate ribosome-stalled nascent polypeptides on the lysine residue for degradation. However, the quality control of ribosome-stalled nuclear-encoded mitochondrial nascent polypeptides remains elusive, as these peptides can be partially imported into mitochondria through translocons, restricting accessibility to the lysine by Listerin. Here, we identify a Listerin-independent organelle-specific mitochondrial RQC pathway that acts on NEMF-mediated carboxy-terminal poly-alanine modification. In the pathway, mitochondrial proteins carrying C-end poly-Ala tails are recognized by the cytosolic E3 ligase Pirh2 and the ClpXP protease in the mitochondria, which coordinately clear ribosome-stalled mitochondrial nascent polypeptides. Defects in this elimination pathway result in NEMF-mediated aggregates and mitochondrial integrity failure, thus providing a potential molecular mechanism of the RQC pathway in mitochondrial-associated human diseases.

Abstract GBA1 variants are important risk factors for Parkinson’s disease (PD). Most studies assessing GBA1-related PD risk have been performed in European-derived populations. Although the coding region of the GBA1 gene in the Chinese population has been analyzed, the sample sizes were not adequate. In this study, we aimed to investigate GBA1 variants in a large Chinese cohort of patients with PD and healthy control and explore the associated clinical characteristics. GBA1 variants in 4034 patients and 2931 control participants were investigated using whole-exome and whole-genome sequencing. The clinical features of patients were evaluated using several scales. Regression analysis, chi-square, and Fisher exact tests were used to analyze GBA1 variants and the clinical symptoms of different groups. We identified 104 variants, including 8 novel variants, expanding the spectrum of GBA1 variants. The frequency of GBA1 variants in patients with PD was 7.46%, higher than that in the control (1.81%) (P

Parkinson's disease (PD) is the most common neurodegenerative movement disease. It is featured by abnormal alpha-synuclein (α-syn) aggregation in dopaminergic neurons in the substantia nigra. Macroautophagy (autophagy) is an evolutionarily conserved cellular process for degradation of cellular contents, including protein aggregates, to maintain cellular homeostasis. Corynoxine B (Cory B), a natural alkaloid isolated from Uncaria rhynchophylla (Miq.) Jacks., has been reported to promote the clearance of α-syn in cell models by inducing autophagy. However, the molecular mechanism by which Cory B induces autophagy is not known, and the α-syn-lowering activity of Cory B has not been verified in animal models. Here, we report that Cory B enhanced the activity of Beclin 1/VPS34 complex and increased autophagy by promoting the interaction between Beclin 1 and HMGB1/2. Depletion of HMGB1/2 impaired Cory B-induced autophagy. We showed for the first time that, similar to HMGB1, HMGB2 is also required for autophagy and depletion of HMGB2 decreased autophagy levels and phosphatidylinositol 3-kinase III activity both under basal and stimulated conditions. By applying cellular thermal shift assay, surface plasmon resonance, and molecular docking, we confirmed that Cory B directly binds to HMGB1/2 near the C106 site. Furthermore, in vivo studies with a wild-type α-syn transgenic drosophila model of PD and an A53T α-syn transgenic mouse model of PD, Cory B enhanced autophagy, promoted α-syn clearance and improved behavioral abnormalities. Taken together, the results of this study reveal that Cory B enhances phosphatidylinositol 3-kinase III activity/autophagy by binding to HMGB1/2 and that this enhancement is neuroprotective against PD.

Abstract Genome-wide association studies (GWASs) have identified numerous susceptibility loci for Parkinson’s disease (PD), but its genetic architecture remains underexplored in populations of non-European ancestry. To identify genetic variants associated with PD in the Chinese population, we performed a GWAS using whole-genome sequencing (WGS) in 1,972 cases and 2,478 controls, and a replication study in a total of 8209 cases and 9454 controls. We identified one new risk variant rs61204179 (P combined = 1.47 × 10−9) with low allele frequency, four previously reported risk variants (NUCKS1/RAB29-rs11557080, SNCA-rs356182, FYN-rs997368, and VPS13C-rs2251086), as well as three risk variants in LRRK2 coding region (A419V, R1628P, and G2385R) with genome-wide significance (P

Maintaining mitochondrial homeostasis is a potential therapeutic strategy for various diseases, including neurodegenerative diseases, cardiovascular diseases, metabolic disorders, and cancer. Selective degradation of mitochondria by autophagy (mitophagy) is a fundamental mitochondrial quality control mechanism conserved from yeast to humans. Indeed, small-molecule modulators of mitophagy are valuable pharmaceutical tools that can be used to dissect complex biological processes and turn them into potential drugs. In the past few years, pharmacological regulation of mitophagy has shown promising therapeutic efficacy in various disease models. However, with the increasing number of chemical mitophagy modulator studies, frequent methodological flaws can be observed, leading some studies to draw unreliable or misleading conclusions. This review attempts (a) to summarize the molecular mechanisms of mitophagy; (b) to propose a Mitophagy Modulator Characterization System (MMCS); (c) to perform a comprehensive analysis of methods used to characterize mitophagy modulators, covering publications over the past 20 years; (d) to provide novel targets for pharmacological intervention of mitophagy. We believe this review will provide a panorama of current research on chemical mitophagy modulators and promote the development of safe and robust mitophagy modulators with therapeutic potential by introducing high methodological standards.

ObjectiveThere is controversial evidence that FMR1 premutation or “gray zone” (GZ) allele (small CGG expansion, 45–54 repeats) was associated with Parkinson’s disease (PD). We aimed to explore further the association between FMR1 CGG repeat expansions and PD in a large sample of Chinese origin.MethodsWe included a cohort of 2,362 PD patients and 1,072 controls from the Parkinson’s Disease and Movement Disorders Multicenter Database and Collaborative Network in China (PD-MDCNC) in this study and conducted repeat-primed polymerase chain reaction (RP-PCR) for the size of FMR1 CGG repeat expansions.ResultsTwo PD patients were detected with FMR1 premutation (61 and 56 repeats), and the other eleven PD patients were detected with the GZ allele of FMR1 CGG repeat expansions. Those thirteen PD patients responded well to levodopa and were diagnosed with clinically established PD. Specifically, one female PD patient with GZ allele was also found with premature ovarian failure. However, compared to healthy controls, we found no significant enrichment of GZ allele carriers in PD patients or other subgroups of PD cases, including the subgroups of female, male, early-onset, and late-onset PD patients. Furthermore, we did not find any correlation between the FMR1 gene CGG repeat sizes and age at onset of PD.ConclusionIt suggested that FMR1 premutation was related to PD, but the GZ allele of FMR1 CGG repeat expansions was not significantly enriched in PD cases of Chinese origin. Further larger multiple ethnic studies are needed to determine further the role of the FMR1 GZ allele in PD.

2- arachidonoylglycerol (2-AG), an abundant endocannabinoid in the brain, regulates diverse neural functions. Here, the authors identified four loss-of-function mutations in dicylglycerol lipase β (DAGLB) from six patients with early onset Parkinsonism. In mice, loss of DAGLB in dopamine neurons reduced neuronal activity and impaired locomotor function and augmentation of 2-AG levels boosted neuronal activity and rescued locomotor deficits.

ObjectiveParkinson's disease (PD) and dystonia are two closely related movement disorders with overlaps in clinical phenotype. Variants in several dystonia-related genes were demonstrated to be associated with PD; however, genetic evidence for the involvement of dystonia-related genes in PD has not been fully studied. Here, we comprehensively investigated the association between rare variants in dystonia-related genes and PD in a large Chinese cohort.MethodsWe comprehensively analyzed the rare variants of 47 known dystonia-related genes by mining the whole-exome sequencing (WES) and whole-genome sequencing (WGS) data from 3,959 PD patients and 2,931 healthy controls. We initially identified potentially pathogenic variants of dystonia-related genes in patients with PD based on different inheritance models. Sequence kernel association tests were conducted in the next step to detect the association between the burden of rare variants and the risk for PD.ResultsWe found that five patients with PD carried potentially pathogenic biallelic variants in recessive dystonia-related genes including COL6A3 and TH. Additionally, we identified 180 deleterious variants in dominant dystonia-related genes based on computational pathogenicity predictions and four of which were considered as potentially pathogenic variants (p.W591X and p.G820S in ANO3, p.R678H in ADCY5, and p.R458Q in SLC2A1). A gene-based burden analysis revealed the increased burden of variant subgroups of TH, SQSTM1, THAP1, and ADCY5 in sporadic early-onset PD, whereas COL6A3 was associated with sporadic late-onset PD. However, none of them reached statistical significance after the Bonferroni correction.ConclusionOur findings indicated that rare variants in several dystonia-related genes are suggestively associated with PD, and taken together, the role of COL6A3 and TH genes in PD is highlighted.

Abstract Dynamic change of mitochondrial morphology and distribution along neuronal branches are essential for neural circuitry formation and synaptic efficacy. However, the underlying mechanism remains elusive. We show here that Pink1 knockout (KO) mice display defective dendritic spine maturation, reduced axonal synaptic vesicles, abnormal synaptic connection, and attenuated long-term synaptic potentiation (LTP). Drp1 activation via S616 phosphorylation rescues deficits of spine maturation in Pink1 KO neurons. Notably, mice harboring a knockin (KI) phosphor-null Drp1 S616A recapitulate spine immaturity and synaptic abnormality identified in Pink1 KO mice. Chemical LTP (cLTP) induces Drp1S616 phosphorylation in a PINK1-dependent manner. Moreover, phosphor-mimetic Drp1S616D restores reduced dendritic spine localization of mitochondria in Pink1 KO neurons. Together, this study provides the first in vivo evidence of functional regulation of Drp1 by phosphorylation and suggests that PINK1-Drp1S616 phosphorylation coupling is essential for convergence between mitochondrial dynamics and neural circuitry formation and refinement.

Alzheimer's disease (AD), characterized by the accumulation of protein aggregates including phosphorylated Tau aggregates, is the most common neurodegenerative disorder with limited therapeutic agents. Autophagy plays a critical role in the degradation of phosphorylated Tau aggregates, and transcription factor EB (TFEB) is a master regulator of autophagy and lysosomal biogenesis. Thus, small-molecule autophagy enhancers targeting TFEB hold promise for AD therapy. Here, we found that celastrol, an active ingredient isolated from the root extracts of Tripterygium wilfordii (Lei Gong Teng in Chinese) enhanced TFEB-mediated autophagy and lysosomal biogenesis in vitro and in mouse brains. Importantly, celastrol reduced phosphorylated Tau aggregates and attenuated memory dysfunction and cognitive deficits in P301S Tau and 3xTg mice, two commonly used AD animal models. Mechanistical studies suggest that TFEB-mediated autophagy-lysosomal pathway is responsible for phosphorylated Tau degradation in response to celastrol. Overall, our findings indicate that Celastrol is a novel TFEB activator that promotes the degradation of phosphorylated Tau aggregates and improves memory in AD animal models. Therefore, Celastrol shows potential as a novel agent for the treatment and/or prevention of AD and other tauopathies.

IntroductionParkinson's disease (PD) is a progressive movement disorder caused by a loss of dopaminergic neurons. Previous studies have highlighted the importance of mitochondria dynamics in the pathogenesis of PD. Dynamin-1-like (DNM1L) is a gene that encodes dynamin-related protein 1 (DRP1), a GTPase essential for proper mitochondria fission. In the present study, we evaluated the relationship between DNM1L variants and PD in the Chinese population.MethodsA total of 3,879 patients with PD and 2,931 healthy controls were recruited and burden genetic analysis combined with high-throughput sequencing was applied.ResultsWe identified 23 rare variants in the coding region of DNM1L, while no difference in variant burden was shown between the cases and controls. We also identified 201 common variants in the coding and flanking regions and found two significant SNPs, namely, rs10844308 and rs143794289 [odds ratio (OR) = 1.220 and 0.718, p = 0.025 and 0.036, respectively]. We also performed a meta-analysis to correlate the two SNPs with PD risk. However, none of the common variants was significant using logistic regression.ConclusionDespite the critical role of DRP1, our study did not support the relationship between DNM1L variants and PD risk in the Chinese population.

ObjectiveCerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a hereditary cerebral small vessel disease caused by mutations in the NOTCH3 gene. Previous studies have established a link between NOTCH3 variants and Parkinson’s disease (PD) in terms of neuropathology and clinical characteristics. In this study, we aimed to explore the role of NOTCH3 gene in PD in a large Chinese cohort.MethodsA total of 1,917 patients with early-onset or familial PD and 1,652 matched controls were included. All variants were divided into common or rare types by minor allele frequency (MAF) at a threshold of 0.01 (MAF > 0.01 into common variants and others into rare variants). Common variants were subjected to single-variant tests by PLINK, then gene-based analyses were used for rare variants with the optimized sequence kernel association test (SKAT-O). For genotype–phenotype correlation assessment, regression models were conducted to compare clinical features between the studied groups.ResultsThree common variants (rs1044006, rs1043997, and rs1043994) showed a nominal protective effect against PD. However, none of these SNPs survived Bonferroni correction. The results in the validation cohort revealed a significant but opposite association between these variants and PD. The gene-based analyses of rare variants showed no significant associations of NOTCH3 with PD. Although we did not find significant associations in the following genotype–phenotype analysis, the higher clinical scores of motor symptoms in NOTCH3-variant carriers were of interest.ConclusionOur results indicated that NOTCH3 gene may not play an important role in the early-onset or familial PD of Chinese population.

Autism spectrum disorder (ASD) represents a group of neurodevelopmental phenotypes with a strong genetic component. An excess of likely gene-disruptive (LGD) mutations in GIGYF1 was implicated in ASD. Here, we report that GIGYF1 is the second-most mutated gene among known ASD high–confidence risk genes. We investigated the inheritance of 46 GIGYF1 LGD variants, including the highly recurrent mutation c.333del:p.L111Rfs*234. Inherited GIGYF1 heterozygous LGD variants were 1.8 times more common than de novo mutations. Among individuals with ASD, cognitive impairments were less likely in those with GIGYF1 LGD variants relative to those with other high-confidence gene mutations. Using a Gigyf1 conditional KO mouse model, we showed that haploinsufficiency in the developing brain led to social impairments without significant cognitive impairments. In contrast, homozygous mice showed more severe social disability as well as cognitive impairments. Gigyf1 deficiency in mice led to a reduction in the number of upper-layer cortical neurons, accompanied by a decrease in proliferation and increase in differentiation of neural progenitor cells. We showed that GIGYF1 regulated the recycling of IGF-1R to the cell surface. KO of GIGYF1 led to a decreased level of IGF-1R on the cell surface, disrupting the IGF-1R/ERK signaling pathway. In summary, our findings show that GIGYF1 is a regulator of IGF-1R recycling. Haploinsufficiency of GIGYF1 was associated with autistic behavior, likely through interference with IGF-1R/ERK signaling pathway.

Noninvasive diagnosis of Alzheimer’s disease (AD) is important for patients. Significant differences in the methylation of mitochondrial DNA (mtDNA) were found in AD brain tissue. Cell-free DNA (cfDNA) is a noninvasive and economical diagnostic tool. We aimed to characterize mtDNA methylation alterations in the plasma cfDNA of 31 AD patients and 26 age- and sex-matched cognitively normal control subjects. We found that the mtDNA methylation patterns differed between AD patients and control subjects. The mtDNA was predominantly hypomethylated in the plasma cfDNA of AD patients. The hypomethylation sites or regions were mainly located in mt-rRNA, mt-tRNA, and D-Loop regions. The hypomethylation of the D-Loop region in plasma cfDNA of AD patients was consistent with that in previous studies. This study presents evidence that hypomethylation in the non-protein coding region of mtDNA may contribute to the pathogenesis of AD and potential application for the diagnosis of AD.

ObjectivesParkinson’s disease (PD) is a neurodegenerative disorder with the manifestation of motor symptoms and non-motor symptoms. Previous studies have indicated the role of several transmembrane (TMEM) protein family genes in PD pathogenesis.Materials and MethodsIn order to better investigate the genetic role of PD-related TMEM protein family genes in PD, including TMEM230, TMEM59, TMEM108, TMEM163, TMEM175, and TMEM229B, 1,917 sporadic early onset PD (sEOPD) or familial PD (FPD) patients and 1,652 healthy controls were analyzed by whole-exome sequencing (WES) while 1,962 sporadic late-onset PD (sLOPD) and 1,279 healthy controls were analyzed by whole-genome sequencing (WGS). Rare and common variants for each gene were included in the analysis.ResultsOne hundred rare damaging or loss of function variants of six genes were found at the threshold of MAF < 0.1%. Three rare Dmis variants of TMEM230 were specifically identified in PD. Rare missense variants of TMEM59 were statistically significantly associated with PD in the WES cohort, indicating the role of TMEM59 in FPD and sEOPD. Rare missense variants of TMEM108 were suggestively associated with PD in the WGS cohort, indicating the potential role of TMEM108 in sLOPD. The rare variant of the other three genes and common variants of six genes were not significantly associated with PD.ConclusionWe performed a large case-control study to systematically investigate the role of several PD-related TMEM protein family genes in PD. We identified three PD-specific variants in TMEM230, the significant association of TMEM59 with FPD, and sEOPD and the suggestive association of TMEM108 with sLOPD.

Parkinson’s disease (PD) is the second most common neurodegenerative disease. PARK9 (also known as ATP13A2) is recognized as one of the key genes that cause PD, and a mutation in this gene was first discovered in a rare case of PD in an adolescent. Lewy bodies (LBs) formed by abnormal aggregation of α-synuclein, which is encoded by the SNCA gene, are one of the pathological diagnostic criteria for PD. LBs are also recognized as one of the most important features of PD pathogenesis. In this article, we first summarize the types of mutations in the ATP13A2 gene and their effects on ATP13A2 mRNA and protein structure; then, we discuss lysosomal autophagy inhibition and the molecular mechanism of abnormal α-synuclein accumulation caused by decreased levels and dysfunction of the ATP13A2 protein in lysosomes. Finally, this article provides a new direction for future research on the pathogenesis and therapeutic targets for ATP13A2 gene-related PD from the perspective of ATP13A2 gene mutations and abnormal aggregation of α-synuclein.

Neurological disorders are a group of disorders with motor, sensory or cognitive damage, caused by dysfunction of the central or peripheral nervous system. Cyclin-dependent kinases 5 (Cdk5) is of vital significance for the development of the nervous system, including the migration and differentiation of neurons, the formation of synapses, and axon regeneration. However, when the nervous system is subject to pathological stimulation, aberrant activation of Cdk5 will induce abnormal phosphorylation of a variety of substrates, resulting in a cascade signaling pathway, and thus lead to pathological changes. Cdk5 is intimately related to the pathological mechanism of a variety of neurological disorders, such as A-β protein formation in Alzheimer’s disease, mitochondrial fragmentation in cerebral ischemia, and apoptosis of dopaminergic neurons in Parkinson’s disease. It is worth noting that Cdk5 inhibitors have been reported to have neuroprotective effects by inhibiting related pathological processes. Therefore, in this review, we will briefly introduce the physiological and pathological mechanisms of Cdk5 in the nervous system, focusing on the recent advances of Cdk5 in neurological disorders and the prospect of targeted Cdk5 for the treatment of neurological disorders.

Increasing evidence reveals sex as an important factor in the development of Parkinson’s disease (PD), but associations between genes on the sex chromosomes and PD remain unknown. TAF1 is a gene located on the X chromosome which is known to cause X-linked syndromic mental retardation-33 (MRXS33) and X-linked Dystonia-Parkinsonism (XDP). In this study, we conducted whole-exome sequencing (WES) among 1,917 patients with early-onset or familial PD and 1,652 controls in a Chinese population. We detected a hemizygous frameshift variant c.29_53dupGGA(CAG)2CTACCATCA(CTG)2C (p.A19Dfs*50) in two unrelated male patients. Further segregation analysis showed an unaffected family member carried this variant, which suggested the penetrance of the variant may be age-related and incomplete. To verify the effects of TAF1 on PD, genetic analyses were carried separately by gender. Analysis of rare variants by optimal sequence kernel association (SKAT-O) test showed a nominally significant difference in variant burden between the male PD patients and controls (2.01 vs. 1.38%, p = 0.027). In the female group, none of the variant types showed significant association with PD in this study. In conclusion, we found rare variants in TAF1 may be implicated in PD, but further genetic and functional analyses were needed.

Hepatic encephalopathy (HE) is a brain dysfunction associated with poor quality of life, increased morbidity and mortality. The pathogenesis of HE is still not fully clarified and effective therapeutic strategies are imperative. Among multiple factors that contribute to the pathophysiological process of HE, ammonia neurotoxicity is thought to be central in the pathogenesis of HE. Therefore, in this study, we subjected SH-SY5Y cells to ammonia insult and performed a pooled genome-wide CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) knockout screen to unveil the underlying molecular mechanisms of ammonia neurotoxicity and discover new potential therapeutic targets for HE. We found that EGLN3 (egl-9 family hypoxia-inducible factor 3) UCP3,GTPBP5, OR4D11 and SDR9C7 with 6 unique sgRNAs may contribute to protection against ammonia injury, while EGLN3 may be most related to ammonia resistance. We knocked down EGLN3 by transfecting neurons with specific shRNA lentivirus and confirmed that EGLN3 knockdown decreased ammonia-induced caspase-3 activation and apoptosis. We also demonstrated that EGLN3 knockdown ameliorated ammonia induced decreased expression of Bcl-2, increased expression of Bax and inhibited release of cytochrome c into the cytosol in neurons, suggesting that EGLN3 inhibition protected against ammonia induced apoptosis through mitochondrial dependent apoptosis pathway. Future therapeutic strategies regulating EGLN3 may be applied to the management of HE.

Abstract In recent years, many studies have investigated the correlations between Parkinson’s disease (PD) and vitamin D status, but the conclusion remains elusive. The present review focuses on the associations between PD and serum vitamin D levels by reviewing studies on the associations of PD with serum vitamin D levels and vitamin D receptor (VDR) gene polymorphisms from PubMed, Web of Science, Cochrane Library, and Embase databases. We found that PD patients have lower vitamin D levels than healthy controls and that the vitamin D concentrations are negatively correlated with PD risk and severity. Furthermore, higher vitamin D concentrations are linked to better cognitive function and mood in PD patients. Findings on the relationship between VDR gene polymorphisms and the risk of PD are inconsistent, but the FokI (C/T) polymorphism is significantly linked with PD. The occurrence of FokI (C/T) gene polymorphism may influence the risk, severity, and cognitive ability of PD patients, while also possibly influencing the effect of Vitamin D3 supplementation in PD patients. In view of the neuroprotective effects of vitamin D and the close association between vitamin D and dopaminergic neurotransmission, interventional prospective studies on vitamin D supplementation in PD patients should be conducted in the future.

Precise recognition of early Parkinson’s disease (PD) has always been a challenging task requiring more feasible biomarkers to be integrated to improve diagnostic accuracy. MicroRNAs (miRNAs) of cerebrospinal fluid (CSF) are believed to be potential and promising candidate biomarkers for PD. However, the role of altered miRNAs of CSF play in PD is unclear. Here, we recruited patients with early stages of PD and controls to analyze the expression of miRNA in CSF by the Next Generation Sequencing (NGS). Furthermore, we tested the levels of these miRNA in SH-SY5Y cells treated with MPP+ using real-time quantitative PCR. We found 21 miRNAs were upregulated in CSF of early PD patients and miR-409-3p, one of the identified 21 miRNAs, was further confirmed in SH-SY5Y cells treated with MPP+. Also, more cells survived in the overexpression of the miR-409-3p group when SH-SY5Y cells and mice were treated with MPP+ and MPTP, respectively. Mechanistically, we demonstrated the binding of miR-409-3p and 3’UTR of ATXN3 through a dual luciferase reporter gene assay. Moreover, miR-409-3p mimic reduced the aggregation of polyglutamine-expanded mutant of ATXN3 and apoptosis. Our results provide experimental evidence for miR-409-3p in CSF as a diagnostic marker of PD.

Parkinson's disease (PD) is a neurodegenerative disease characterized by progressive death of dopaminergic neurons in the substantia nigra and the formation of Lewy bodies (LBs). Mutations in PD-related genes lead to neuronal pathogenesis through various mechanisms, with known examples including SNCA/α-synuclein (PAKR1), Parkin (PARK2), PINK1 (PARK6), DJ-1 (PARK7), and LRRK2 (PARK8). Molecular chaperones/co-chaperones are proteins that aid the folding of other proteins into a functionally active conformation. It has been demonstrated that chaperones/co-chaperones interact with PD-related proteins and regulate their function in PD. HSP70, HSP90 and small heat shock proteins can prevent neurodegeneration by regulating α-syn misfolding, oligomerization and aggregation. The function of chaperones is regulated by co-chaperones such as HSP110, HSP40, HOP, CHIP, and BAG family proteins. Parkin, PINK1 and DJ-1 are PD-related proteins which are associated with mitochondrial function. Molecular chaperones regulate mitochondrial function and protein homeostasis by interacting with these PD-related proteins. This review discusses critical molecular chaperones/co-chaperones and PD-related proteins which contribute to the pathogenesis of PD, hoping to provide new molecular targets for therapeutic interventions to thwart the disease progression instead of only bringing symptomatic relief. Moreover, appreciating the critical role of chaperones in PD can also help us screen efficient biomarkers to identify PD at an early stage.

Background: Recently, many studies have shown that low vitamin D (VD) levels may be related to an increased risk of Parkinson’s disease (PD), but the underlying mechanisms remain unclear.Objective: To explore the relationship between PD and VD levels, as well as to analyze the effects of VD on spontaneous brain activity and explore the possible mechanism of its involvement in PD risk.Methods: In a cross-sectional study, we quantified the difference in VD levels between 330 PD patients and 209 healthy controls (HC) to explore the correlation between VD and PD risk. We also acquired resting-state Functional Magnetic Resonance Imaging (rs-fMRI) data from 46 PD patients and 21 HC. The PD patients were divided into three groups according to 25(OH)D levels: PD patients with VD deficiency (PD + VDD), PD patients with VD insufficiency (PD + VDI), and PD patients with normal VD (PD + NVD). The effect of VD status on spontaneous neuronal activity in the whole brain was analyzed by measuring the fraction amplitude of low-frequency fluctuation (fALFF).Results: Compared with HC, the PD patients had lower serum 25(OH)D levels (23.60 ± 7.27 vs. 25.60 ± 5.78, P < 0.001). The 25(OH)D level may have a potential dose-dependent effect on the risk of PD (Ptrend = 0.007). A high risk of PD was associated with VD deficiency [25(OH)D < 20 ng/mL, OR = 2.319], and the lowest quartile of 25(OH)D concentration was associated with a high risk of PD (OR = 1.941). In the rs-fMRI study, PD + VDD patients had wider brain regions with altered fALFF than other PD groups when compared with the corresponding HC groups. Both PD + VDD and PD + VDI showed higher fALFF in the cuneus, left precuneus, calcarine cortex and right lingual, as well as lower fALFF in the left middle temporal gyrus. PD + VDD patients also showed higher fALFF in the left superior, middle and inferior frontal gyri, as well as the left precentral gyrus than HC. Among PD patients, there was only a statistically significant difference in fALFF between the PD + VDD and PD + NVD groups. Compared with the PD + NVD group, PD + VDD patients exhibited higher fALFF in the left precentral and left postcentral gyrus, as well as the left inferior parietal lobule.Conclusion: These results demonstrate that PD patients had lower serum VD levels than HC, and VD may have a potential dose-dependent effect on PD risk. Lower serum VD levels can affect the spontaneous neuronal activity of default-mode network (DMN) and visual pathway neurons in PD patients, providing a possible mechanism for its effect on PD risk.

Metabolic associated fatty liver disease (MAFLD) is associated with obesity, type 2 diabetes mellitus, and other metabolic syndromes. Farnesoid X receptor (FXR, NR1H4) plays a prominent role in hepatic lipid metabolism. This study combined the expression of liver genes in FXR knockout (KO) mice and MAFLD patients to identify new pathogenic pathways for MAFLD based on genome-wide transcriptional profiling. In addition, the roles of new target genes in the MAFLD pathogenic pathway were also explored. Two groups of differentially expressed genes were obtained from FXR-KO mice and MAFLD patients by transcriptional analysis of liver tissue samples. The similarities and differences between the two groups of differentially expressed genes were analyzed to identify novel pathogenic pathways and target genes. After the integration analysis of differentially expressed genes, we identified 134 overlapping genes, many of which have been reported to play an important role in lipid metabolism. Our unique analysis method of comparing differential gene expression between FXR-KO mice and patients with MAFLD is useful to identify target genes and pathways that may be strongly implicated in the pathogenesis of MAFLD. The overlapping genes with high specificity were screened using the Gene Expression Omnibus (GEO) database. Through comparison and analysis with the GEO database, we determined that BHMT2 and PKLR could be highly correlated with MAFLD. Clinical data analysis and RNA interference testing in vitro confirmed that BHMT2 may a new regulator of lipid metabolism in MAFLD pathogenesis. These results may provide new ideas for understanding the pathogenesis of MAFLD and thus provide new targets for the treatment of MAFLD.

PM2.5, also known as fine particles, refers to particulate matter with a dynamic diameter of ≦2.5 μm in air pollutants, that carries metals (Zn, Co, Cd) which can pass through the alveolar epithelium and enter the circulatory system and tissues. PM2.5 can cause serious health problems, such as non-alcoholic fatty liver and hepatocellular carcinoma, although the underlying mechanisms of its toxic effect are poorly understood. Here, we exposed L02 cells to PM2.5 and performed a pooled genome−wide clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) to assess loss of function and identify new potential PM2.5targets. Enrichr and KEGG pathway analyses were performed to identify candidate genes associated with PM2.5 toxicity. Results revealed that four key genes, namely ATPase Na+/K+ transporting subunit alpha 2 (ATP1A2), metallothionein 1M (MT1M), solute carrier family 6 members 19 (SLC6A19) and transient receptor potential cation channel subfamily V member 6 (TRPV6) were associated with PM2.5 toxicity, mainly in regulating the mineral absorption pathway. Downregulating these genes increased cell viability and attenuated apoptosis in cells exposed to PM2.5. Conversely, overexpressing TRPV6 exacerbated cell apoptosis caused by PM2.5, while a reactive oxygen species (ROS) inhibitor N-acetyl-l-cysteine (NAC) alleviated PM2.5-induced apoptosis. In conclusion, ATP1A2, MT1M, SLC6A19 and TRPV6 may be contributing to absorption of metals in PM2.5 thereby inducing apoptosis mediated by ROS. Therefore, they hold potential as therapeutic targets for PM2.5-related diseases.

Abstract Background We previously performed targeted sequencing of autism risk genes in probands from the Autism Clinical and Genetic Resources in China (ACGC) (phase I). Here, we expand this analysis to a larger cohort of patients (ACGC phase II) to better understand the prevalence, inheritance, and genotype–phenotype correlations of likely gene-disrupting (LGD) mutations for autism candidate genes originally identified in cohorts of European descent. Methods We sequenced 187 autism candidate genes in an additional 784 probands and 85 genes in 599 probands using single-molecule molecular inversion probes. We tested the inheritance of potentially pathogenic mutations, performed a meta-analysis of phase I and phase II data and combined our results with existing exome sequence data to investigate the phenotypes of carrier parents and patients with multiple hits in different autism risk genes. Results We validated recurrent, LGD, de novo mutations (DNMs) in 13 genes. We identified a potential novel risk gene (ZNF292), one novel gene with recurrent LGD DNMs (RALGAPB), as well as genes associated with macrocephaly (GIGYF2 and WDFY3). We identified the transmission of private LGD mutations in genes predominantly associated with DNMs and showed that parental carriers tended to share milder autism-related phenotypes. Patients that carried DNMs in two or more candidate genes show more severe phenotypes. Conclusions We identify new risk genes and transmission of deleterious mutations in genes primarily associated with DNMs. The fact that parental carriers show milder phenotypes and patients with multiple hits are more severe supports a multifactorial model of risk.

Background and Objective:Olfactory dysfunction (hyposmia) is an important non-motor symptom of Parkinson’s disease (PD). To investigate the potential prognostic value of hyposmia as a marker for disease progression, we prospectively assessed clinical manifestations and longitudinal changes of hyposmic PD patients and normosmic ones.MethodsOlfactory function was evaluated with the Sniffin’ Sticks in PD patients at baseline. One hundred five hyposmic PD patients and 59 normosmic PD patients were enrolled and followed up for 2 years. They were subsequently evaluated at baseline and during follow-up periods with neurological and neuropsychological assessments. Clinical manifestations and disease progressions were compared between hyposmic and normosmic patients. In addition, the relationship between disease progressions and olfactory function was analyzed.ResultsOur study suggested that hyposmic PD patients and normosmic ones were similar in gender, age, education levels, age of onset, disease duration, and clinical features at baseline. Hyposmic PD patients exhibited more severe Unified Parkinson’s Disease Rating Scale Part II–III (UPDRS II-III) scores, higher levodopa equivalent dose (LED) needs, and poorer Mini-Mental State Examination (MMSE) score at follow-up visits compared to those in normosmic PD patients. Hyposmia also showed greater rates in the increase of LED needs, improvement of UPDRS III score, and deterioration of MMSE score. Both improvement of UPDRS III score and decline of MMSE score were associated with poorer odor identification.ConclusionOur prospective study demonstrated that hyposmic PD patients showed a relatively worse clinical course compared with normosmic patients. Olfactory dysfunction is a useful predictor of disease progression.

Objective: To conduct an investigation into the reliability of assessing the olfactory function of patients with Parkinson's disease (PD) in a clinical setting of crowding patients in populated countries, such as China, by the hyposmia rating scale (HRS) and compare other non-motor features between patients with PD with olfactory dysfunction (PD-OD) and patients with PD without olfactory dysfunction (PD-NOD), according to the result of olfactory function assessed by the Sniffin' Sticks test.Methods: A total of 320 patients with clinically confirmed or clinically possible PD were recruited. Olfactory function of all participants was assessed with the HRS and the Sniffin' Sticks test. Demographic data and clinical information were collected, and patients were evaluated using standardized assessment protocols. With reference to the Sniffin' Sticks test, the specificity, sensitivity, coincidence rate, and kappa value of the HRS was computed, and then its reliability was evaluated. We divided patients into PD-OD and PD-NOD groups based on the results of olfactory function assessed by the Sniffin' Sticks test. Clinical manifestations were compared between PD-OD and PD-NOD.Results: The percentage of patients with OD determined by the Sniffin' Sticks test was 65.6%, and the percentage of those with OD was 55.6% when using the HRS measured olfactory function. With reference to the Sniffin' Sticks test, the specificity, sensitivity, coincidence rate, and kappa value of the HRS were 82.73, 75.71, 78.13%, and 0.55, respectively. The area under the receiver operating characteristic curve for the HRS was 0.793. There were no differences in demographic characteristics between the PD-OD and PD-NOD groups. The patients with hyposmia had more severe non-motor symptoms.Conclusion: The HRS is of great value as a self-assessment scale for evaluating olfactory function, especially in PD patients over 55 years old. Moreover, PD patients with hyposmia have more severe non-motor features than PD patients without hyposmia, mainly in terms of mood and constipation.

Wnt signaling through the Frizzled (FZD) family of serpentine receptors is essential for embryogenesis and homeostasis, and stringent control of the FZD protein level is critical for stem cell regulation. Through CRISPR/Cas9 genome-wide screening in human cells, we identified TMEM79/MATTRIN, an orphan multi-span transmembrane protein, as a specific inhibitor of Wnt/FZD signaling. TMEM79 interacts with FZD during biogenesis and promotes FZD degradation independent of ZNRF3/RNF43 ubiquitin ligases (R-spondin receptors). TMEM79 interacts with ubiquitin-specific protease 8 (USP8), whose activating mutations underlie human tumorigenesis. TMEM79 specifically inhibits USP8 deubiquitination of FZD, thereby governing USP8 substrate specificity and promoting FZD degradation. Tmem79 and Usp8 genes have a pre-bilaterian origin, and Tmem79 inhibition of Usp8 and Wnt signaling is required for anterior neural development and gastrulation in Xenopus embryos. TMEM79 is a predisposition gene for Atopic dermatitis, suggesting deregulation of Wnt/FZD signaling a possible cause for this most common yet enigmatic inflammatory skin disease.

Background and Purpose: Olfactory dysfunction is one of the most common non-motor symptoms in Parkinson's disease (PD) preceding the motor symptoms for years. This study aimed to evaluate different olfactory domains in PD patients in comparison with healthy controls and to explore the relationships among olfactory deficit and other clinical manifestations in patients with PD.Methods: Sniffin' Sticks test, which detects olfactory threshold, discrimination, and identification (TDI), were conducted in 500 PD patients and 115 controls. Furthermore, demographic and clinical data including motor and other non-motor symptoms were collected.Results: In the single olfactory model, the identification test showed the area under the receiver operating characteristic (ROC) curve (AUC = 0.818), followed by threshold test (AUC = 0.731) and discrimination test (AUC = 0.723). Specifically, the identification test has a similar discriminative power as the TDI score (0.818 and 0.828, respectively, p = 0.481). In the integrated olfactory model involved with other non-motor manifestations, identification test scores performed as good as the TDI score in differentiating PD patients from controls (0.916 and 0.918, respectively, p = 0.797). In PD patients, age and cognition together explained 7.5% of the variance of the threshold score, while age, cognition, and gender accounted for the 15.2% explained variance of the discrimination score, while cognition, age, the ability of daily living, and gender together interpreted 11.1% of the variance of the identification score.Conclusion: Our results indicated that the identification domain was the most practical olfactory factor in differentiating PD patients, and the combination of several different manifestations was better than a single symptom. Furthermore, the olfactory identification score may be associated with the ability of daily living.

BACKGROUND:Echinococcosis is a chronic zoonosis caused by tapeworms of the genus Echinococcus. Treatment of the disease is often expensive and complicated, sometimes requiring extensive surgery. Ultrasonographic imaging is currently the main technique for diagnosis, while immunological analysis provides additional information. Confirmation still needs pathological analysis. However, these diagnostic techniques generally detect infection in late stages of the disease. An accurate, early and non-invasive molecular diagnostic method is still unavailable. METHODOLOGY/PRINCIPAL FINDINGS:We sequenced the cell-free DNA (cfDNA) from plasma of echinococcosis patients and confirmed the presence of Echinococcus DNA. To improve detection sensitivity, we developed a method based on targeted next-generation sequencing of repeat regions. Simulation experiments demonstrate that the targeted sequencing is sensitive enough to detect as little as 0.1% of an Echinococcus genome in 1 mL of plasma. Results obtained using patient plasma shows that the Area Under the Curve (AUC) of the method is 0.862, with a detection sensitivity of 62.50% and specificity of 100%, corresponding to a Youden-index of 0.625. CONCLUSIONS/SIGNIFICANCE:This study provides evidence that hydatid cysts release cfDNA fragments into patient plasma. Using the repeat region targeted sequencing method, highly specific detection of Echinococcus infection was achieved. This study paves a new avenue for potential non-invasive screening and diagnosis of echinococcosis.

Background/Aims: We have reported the neuroprotective properties of Heat shock protein B8(HSPB8) against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced injury by inhibiting the mitochondrial apoptotic pathway. However, the exact underlying mechanism of its protective effect on mitochondrial function remains unknown. Here we examined whether the beneficial effect of HSPB8 on OGD/R-induced cell death is associated with mitophagy in mouse neuroblastoma Neuro2a (N2a) cells. Methods: Using the mouse transient middle cerebral artery occlusion (tMCAO) model and mouse neuroblastoma Neuro2a (N2a) cell cultures subjected to OGD/R, we employed western-blot, RT-PCR and immunostaining to analyze the change of expression pattern of HSPB8 and mitophagic flux after brain I/R both in vivo and in vitro. Moreover, via overexpressing HSPB8 or knocking down HSPB8 expression with siRNA in N2a cell, we evaluated the effect of HSPB8 on mitochondrial function during OGD/R. The impact of HSPB8 on mitophagic pathway was also assessed. Finally, mitotophagy inhibitors (CQ and Mdivi-1) were adopted to verify the involvement of mitophagy in HSPB8- induced neuroprotection. Results: HSPB8 could be up-regulated by brain I/R both in vivo and in vitro. Mitophagy enhancement coincided with induction of HSPB8 during I/R. Overexpression of HSPB8 reinforced I/R-induced mitophagy in OGD/R-treated mouse N2a cells and HSPB8 silence suppressed mitophagy process. Inhibition of mitophagy compromised neuroprotection conferred by HSPB8 overexpression. Conclusions: HSPB8 promoted OGD/R-induced mitophagy, which restored the mitochondrial function and contributed to the decrease in cell apoptosis after OGD/R. Therefore, HSPB8 could be a favorable neuroprotective agent for cerebral I/R related disorders.

Abstract 6-OHDA plus ascorbic acid (AA) has long been used to induce Parkinson’s disease in rodents, while only 6-OHDA is commonly used to induce cell damage in cellular PD models. AA was believed to act as an anti-oxidant to prevent the degradation of 6-OHDA; however, some studies suggested that AA dramatically enhanced the selectivity and toxicity of 6-OHDA. To understand the mechanisms by which 6-OHDA/AA induces cell death, we established a 6-OHDA/AA cell toxicity model in human dopaminergic neuroblastoma SH-SY5Y cells. We confirmed that the toxicity of 6-OHDA was dramatically increased in the presence of AA, and the toxicity can be prevented by a flavonoid, baicalein. Mechanistically, our research reveals that 6-OHDA/AA induces cell death mainly through the interruption of intracellular calcium homeostasis, which leads to calpain activation and mitochondrial damage. Baicalein prevents 6-OHDA/AA-induced intracellular calcium elevation as well as consequent mitochondria damage. Taken together, our study confirms that 6-OHDA/AA is a more sensitive model for inducing neuronal lesion in vitro and reveals the central role of intracellular calcium in 6-OHDA/AA-induced cell death. Our studies further show that baicalein prevents 6-OHDA/AA-induced cell death by inhibiting intracellular calcium elevation.

Fibrotic animal models are critical for the pathogenesis investigations and drug explorations in systemic sclerosis (SSc). The bleomycin (BLM)-induced mouse model is the classical and most widely used fibrosis model. However, traditional subcutaneous injection of BLM rarely induced diffuse skin and lung lesions. Hypochlorous acid (HOCl)-induced mice are a more representative model that have diffuse cutaneous lesions, lung fibrosis and renal involvement. However, the fibrotic and immunological features of this model are not fully elucidated. Here, we injected BALB/c mice subcutaneously with HOCl used at different concentrations of HOCl (1:55, 1:70, and 1:110 NaClO: KH2PO4, hereafter named HOCl55, HOCl70, and HOCl110, respectively) for 6 weeks to induce fibrosis, and also used HOCl110 at different time course (4, 5, and 6 weeks). Morphological changes were observed via HE and Masson's trichrome staining. Immunohistochemistry or real-time PCR was used to detect inflammatory infiltrates, important fibrosis pathways and pro-inflammatory mediator expression. Flow cytometry was used to detect the alteration of immune cells in mouse spleen. Skin and lung fibrosis were most obvious in the HOCl55 group compared to lower concentration groups. In the HOCl110 group, dominant inflammatory infiltrates were found after 5 weeks, and significant fibrosis was found after 6 weeks. Then we explored the fibrosis and immunological profiles in the HOCl110 (6 weeks) group. Important fibrosis pathway proteins such as TGF-β, NF-κB, Smad3, p-Smad3, STAT3, and p-STAT3 were significantly elevated at week 6 in the HOCl110 group. Increased infiltration of CD4+T cells, CD8+T cells, CD20+B cells, and myofibroblasts was found both in skin and lung tissues. However, decreased CD4+T cells, CD8+T cells, monocytes and macrophages and increased CD19+B cells were found in the spleen tissues. The mRNA expression of fibrosis mediators such as IL-1β, IL-6, IL-17, IL-33, TNF-α, and CTGF was also upregulated in skin and lung tissues. In conclusion, HOCl induced fibrosis mouse model displayed systemic immune cell infiltration, pro-inflammatory mediator release, vasculopathy and fibrosis, which better mimicked human SSc than BLM-induced mice.

Hepatic cytochrome P450 enzyme activities correlate with non-alcoholic fatty liver disease (NAFLD) and hepatic steatosis. The decreased activity of CYP3A4, an important drug-metabolizing enzyme, is associated with the progression of NAFLD. CYP3A4 is predicted as a target gene of miR-200a-3p and miR-150-5p by MicroInspector and TargetScan algorithms analyses. Here, we found decreased CYP3A4 and increased miR-200a-3p and miR-150-5p in LO2 cells with free fatty acid (FFA)-induced steatosis. Dual-luciferase assay confirmed that both miR-200a-3p and miR-150-5p targeted the 3′-untranslated region (3′-UTR) of CYP3A4 and that such interaction was abolished by miRNA binding site mutations in 3′-UTR of CYP3A4. Using miR-200a-3p and miR-150-5p mimics and inhibitors, we further confirmed that endogenous CYP3A4 was regulated posttranscriptionally by miR-200a-3p or miR-150-5p. Moreover, miR-200a-3p and miR-150-5p inhibitors attenuated FFA-induced steatosis in LO2 cells, and such effect was dependent on CYP3Y4 expression. These results suggest that miR-200a-3p and miR-150-5p, through directly targeting 3′-UTR of CYP3A4, contribute to the development of FFA-induced steatosis.

Background and Objectives: Studies examining the risk factors for dyskinesia in Parkinson's disease (PD) have been inconsistent, and racial differences exist. Since there have been no systematic studies of the characteristics of dyskinesia in the Mainland Chinese population, we sought to elucidate the risk factors for dyskinesia.Methods: A total of 1974 PD patients from Mainland China were systematically investigated by univariable and multivariable analyses. PD patients with and without dyskinesia were stratified into 4 groups according to levodopa equivalent daily dose (LEDD) and analyzed by a Cox proportional hazards model. A longitudinal study of 87 patients with dyskinesia was classified into 3 groups according to the duration from onset of PD to the initiation of levodopa, and comparisons among groups were analyzed by the Mann-Whitney test.Results: Early age of onset, long disease duration, being female, high LEDD, low UPDRS III scores (ON-state) and high Hoehn-Yahr stage (ON-state) were predictors of dyskinesia. Dyskinesia was levodopa dosage-dependent, and the incidence increased remarkably when LEDD exceeded 300 mg/d (p < 0.05). The emergence of dyskinesia had no association with the initiation time of levodopa, and if the latter was more than 4 years, the duration of time on chronic levodopa free of motor complications was significantly shortened.Conclusions: We found risk factors for the prediction of dyskinesia. Our data shows that physicians should be cautious if the LEDD exceeds 300 mg/d. The development of dyskinesia was not correlated with the time of levodopa initiation.

Nitric oxide (NO) is an important mediator of inflammation response and the production of NO has been linked to a variety of diseases, including tumors, inflammation and central nervous system diseases. In macrophages, a high level of NO is generated by iNOS during inflammatory responses triggered by cytokines or pathogens. Autophagy, a cellular bulk degradation process via lysosome, has been implicated in many disease conditions including inflammation. In this study, we have reported the previously unknown role of autophagy in regulating iNOS levels in macrophages, both under basal and Lipopolysaccharides (LPS)-induced conditions. Our data showed that iNOS levels accumulated upon autophagy inhibition and decreased upon autophagy induction. iNOS interacted and co-localized with autophagy receptor p62/SQSTM1, especially under LPS-stimulated condition in macrophages. Moreover, the immunostaining data revealed that iNOS also co-localizes with the autophagosome marker LC3 and lysosome marker LAMP1, especially under lysosomal inhibition conditions, indicating iNOS is an autophagy substrate. Finally, we showed that autophagy negatively regulated the generation of NO in macrophages, which is consistent with the changes of iNOS levels. Collectively, our study revealed a previously unknown mechanism by which autophagy regulates iNOS levels to modulate NO production during inflammation.

Mutations in PTEN-induced kinase 1 (PINK1) gene cause PARK6 familial Parkinsonism, and loss of the stability of PINK1 may also contribute to sporadic Parkinson's disease (PD). Degradation of PINK1 occurs predominantly through the ubiquitin proteasome system (UPS), however, to date, few of the proteins have been found to regulate the degradation of PINK1. Using the yeast two-hybrid system and pull-down methods, we identified bcl-2-associated athanogene 5 (BAG5), a BAG family member, directly interacted with PINK1. We showed that BAG5 stabilized PINK1 by decreasing the ubiquitination of PINK1. Interestingly, BAG5 rescued MPP(+)- and rotenone-induced mitochondria dysfunction by up-regulating PINK1 in vitro. In PINK1-null mice and MPTP-treated mice, BAG5 significantly increased in the substantia nigra pars compacta (SNpc) although PINK1 was decreased. Our findings indicated that BAG5, as a key protein to stabilize PINK1, is a promising therapeutic tool for preventing mitochondrial dysfunction following oxidative stress.

Mutations in Connexin-31 (Cx31) are associated with multiple human diseases including erythrokeratodermia variabilis (EKV). The molecular action of Cx31 pathogenic mutants remains largely elusive. We report here that expression of EKV pathogenic mutant Cx31R42P induces cell death with necrotic characteristics. Inhibition of hemichannel activity by a connexin hemichannel inhibitor or high extracellular calcium suppresses Cx31R42P-induced cell death. Expression of Cx31R42P induces ER stress resulting in reactive oxygen species (ROS) production, in turn, to regulate gating of Cx31R42P hemichannels and Cx31R42P induced cell death. Moreover, Cx31R42P hemichannels play an important role in mediating ATP release from the cell. In contrast, no hemichannel activity was detected with cells expressing wildtype Cx31. Together, the results suggest that Cx31R42P forms constitutively active hemichannels to promote necrotic cell death. The Cx31R42P active hemichannels are likely resulted by an ER stress mediated ROS overproduction. The study identifies a mechanism of EKV pathogenesis induced by a Cx31 mutant and provides a new avenue for potential treatment strategy of the disease.