Targeting epigenetic and post-translational modifications regulating pyroptosis for the treatment of inflammatory diseases.

Inflammatory diseases, including infectious diseases, diabetes-related diseases, arthritis-related diseases, neurological diseases, digestive diseases, and tumor, continue to threaten human health and impose a significant financial burden despite advancements in clinical treatment. Pyroptosis, a pro-inflammatory programmed cell death pathway, plays an important role in the regulation of inflammation. Moderate pyroptosis contributes to the activation of native immunity, whereas excessive pyroptosis is associated with the occurrence and progression of inflammation. Pyroptosis is complicated and tightly controlled by various factors. Accumulating evidence has confirmed that epigenetic modifications and post-translational modifications (PTMs) play vital roles in the regulation of pyroptosis. Epigenetic modifications, which include DNA methylation and histone modifications (such as methylation and acetylation), and post-translational modifications (such as ubiquitination, phosphorylation, and acetylation) precisely manipulate gene expression and protein functions at the transcriptional and post-translational levels, respectively. In this review, we summarize the major pathways of pyroptosis and focus on the regulatory roles and mechanisms of epigenetic and post-translational modifications of pyroptotic components. We also illustrate these within pyroptosis-associated inflammatory diseases. In addition, we discuss the effects of novel therapeutic strategies targeting epigenetic and post-translational modifications on pyroptosis, and provide prospective insight into the regulation of pyroptosis for the treatment of inflammatory diseases.

Transplantation of human neural stem cell prevents symptomatic motor behavior disability in a rat model of Parkinson's disease.

Parkinson's disease (PD) is a ubiquitous brain cell degeneration disease and presents a significant therapeutic challenge. By injecting 6-hydroxydopamine (6-OHDA) into the left medial forebrain bundle, rats were made to exhibit PD-like symptoms and treated by intranasal administration of a low-dose (2 × 105) or high-dose (1 × 106) human neural stem cells (hNSCs). Apomorphine-induced rotation test, stepping test, and open field test were implemented to evaluate the motor behavior and high-performance liquid chromatography was carried out to detect dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), serotonin, and 5-hydroxyindole-3-acetic acid in the striatum of rats. Animals injected with 6-OHDA showed significant motor function deficits and damaged dopaminergic system compared to the control group, which can be restored by hNSCs treatment. Treatment with hNSCs significantly increased the tyrosine hydroxylase-immunoreactive cell count in the substantia nigra of PD animals. Moreover, the levels of neurotransmitters exhibited a significant decline in the striatum tissue of animals injected with 6-OHDA when compared to that of the control group. However, transplantation of hNSCs significantly elevated the concentration of DA and DOPAC in the injured side of the striatum. Our study offered experimental evidence to support prospects of hNSCs for clinical application as a cell-based therapy for PD.

The association between plasma GPNMB and Parkinson's disease and multiple system atrophy.

AIMS: Parkinson's disease (PD), as the second most common neurodegenerative disorder, often presents diagnostic challenges in differentiation from other forms of Parkinsonism. Recent studies have reported an association between plasma glycoprotein nonmetastatic melanoma protein B (pGPNMB) and PD. METHODS: A retrospective study was conducted, comprising 401 PD patients, 111 multiple system atrophy (MSA) patients, 13 progressive supranuclear palsy (PSP) patients and 461 healthy controls from the Chinese Han population, with an assessment of pGPNMB levels. RESULTS: The study revealed that pGPNMB concentrations were significantly lower in PD and MSA patients compared to controls (area under the receiver operating characteristics curve (AUC) 0.62 and 0.74, respectively, P < 0.0001 for both), but no difference was found in PSP patients compared to controls (P > 0.05). Interestingly, the level of pGPNMB was significantly higher in PD patients than in MSA patients (AUC = 0.63, P < 0.0001). Furthermore, the study explored the association between pGPNMB levels and disease severity in PD and MSA patients, revealing a positive correlation in PD patients but not in MSA patients with both disease severity and cognitive impairment. CONCLUSION: This study successfully replicated prior findings, demonstrating an association between pGPNMB levels and disease severity, and also identified a correlation with cognitive impairment in PD patients of the Chinese Han population. Additionally, this study is the first to identify a significant difference in pGPNMB levels between MSA, PD, and normal controls. The data provide new evidence supporting the potential role of pGPNMB in the diagnosis and differential diagnosis of Parkinsonism.

Children with severe neurological symptoms associated with SARS-CoV-2 infection during Omicron pandemic in China.

BACKGROUND: To analyze the clinical characteristics and outcomes of children with severe neurological symptoms associated with the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection during the Omicron pandemic in China. METHODS: This study used a questionnaire to obtain data from pediatric intensive care unit (PICU) centers in seven tertiary hospitals in Northeast China from December 1, 2022, to January 31, 2023. RESULTS: A total of 255 patients were confirmed to have SARS-CoV-2 infection, and 45 patients (17.65 %) were included in this study. Of these, seven (15.6%) patients died, and the median time from admission to death was 35 h (IQR, 14-120 h). Twenty (52.6%) survivors experienced neurological sequelae. Patients with platelet counts lower than 100 × 109/L had a higher incidence of complications such as multiple organ dysfunction, mechanical ventilation rate, and mortality. Cranial magnetic resonance imaging (MRI) always reveals cerebral tissue edema, with some severe lesions forming a softening site. CONCLUSION: Children infected with SARS-CoV-2 often exhibit severe neurological symptoms, and in some cases, they may rapidly develop malignant cerebral edema or herniation, leading to a fatal outcome. An early decrease in platelet count may associated with an unfavorable prognosis. IMPACT: Since early December 2022, China has gradually adjusted its prevention and control policy of SARS-CoV-2; Omicron outbreaks have occurred in some areas for a relatively short period. Due to the differences in ethnicity, endemic strains and vaccination status, there was a little difference from what has been reported about children with SARS-CoV-2 infection with severe neurological symptoms in abroad. This is the first multicenter clinical study in children with nervous system involvement after acute SARS-CoV-2 infection in China, and helpful for pediatricians to have a more comprehensive understanding of the clinical symptoms and prognosis of such disease.

Correlation Between Substantia Nigra Hyperechogenicity and Iron Metabolism in the Postural Instability Gait Difficulty Subtype of Parkinson's Disease.

OBJECTIVE: The correlation between substantia nigra (SN) hyperechogenicity on transcranial sonography (TCS) and serum iron metabolism parameters in patients with the postural instability gait difficulty (PIGD) subtype of Parkinson's disease (PD) was investigated so as to explore the pathological mechanism of SN hyperechogenicity. METHODS: The study enrolled 95 PIGD patients recruited by the Parkinson's Disease Specialty in the Second Affiliated Hospital of Soochow University during June 2019-2021. On the basis of the TCS results, the PIGD patients were assigned to the PD with SN hyperechogenicity (SN+) group (n = 60) and PD without SN hyperechogenicity (SN-) group (n = 35). Meanwhile, 49 sex- and age-matched healthy individuals were included in the control group. All participants underwent blood tests. Differences in the iron metabolism parameters among the three groups and the correlation between SN hyperechogenicity and serum iron metabolism parameters were analyzed. RESULTS: Serum ferritin, ceruloplasmin and transferrin levels were lower in the SN+ and SN- groups than in the control group (all p values <0.001). The serum ceruloplasmin level was lower in the SN+ group (0.23 [0.20, 0.25] g/L) than in the SN- group (0.25 [0.22, 0.29] g/L) (p = 0.001), and the proportion of patients with an abnormal ceruloplasmin level was higher in the SN+ group than in the SN- group (43.3% [26/60] vs. 14.3% [5/35], χ2 = 8.484, p = 0.004). Moreover, the SN hyperechogenicity area was negatively correlated with the serum transferrin level (r = -0.428, p < 0.001). CONCLUSION: Decreased serum ceruloplasmin levels may be associated with SN hyperechogenicity development in PIGD patients. The SN hyperechogenicity area is negatively correlated with the serum transferrin level.

PACAP/PAC1-R activation contributes to hyperalgesia in 6-OHDA-induced Parkinson's disease model rats via promoting excitatory synaptic transmission of spinal dorsal horn neurons.

Pain is a common annoying non-motor symptom in Parkinson's disease (PD) that causes distress to patients. Treatment for PD pain remains a big challenge, as its underlying mechanisms are elusive. Pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptor PAC1-R play important roles in regulating a variety of pathophysiological processes. In this study, we investigated whether PACAP/PAC1-R signaling was involved in the mechanisms of PD pain. 6-hydroxydopamine (6-OHDA)-induced PD model was established in rats. Behavioral tests, electrophysiological and Western blotting analysis were conducted 3 weeks later. We found that 6-OHDA rats had significantly lower mechanical paw withdrawal 50% threshold in von Frey filament test and shorter tail flick latency, while mRNA levels of Pacap and Adcyap1r1 (gene encoding PAC1-R) in the spinal dorsal horn were significantly upregulated. Whole-cell recordings from coronal spinal cord slices at L4-L6 revealed that the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) in dorsal horn neurons was significantly increased, which was reversed by application of a PAC1-R antagonist PACAP 6-38 (250 nM). Furthermore, we demonstrated that intrathecal microinjection of PACAP 6-38 (0.125, 0.5, 2 µg) dose-dependently ameliorated the mechanical and thermal hyperalgesia in 6-OHDA rats. Inhibition of PACAP/PAC1-R signaling significantly suppressed the activation of Ca2+/calmodulin-dependent protein kinase II and extracellular signal-regulated kinase (ERK) in spinal dorsal horn of 6-OHDA rats. Microinjection of pAAV-Adcyap1r1 into L4-L6 spinal dorsal horn alleviated hyperalgesia in 6-OHDA rats. Intrathecal microinjection of ERK antagonist PD98059 (10 µg) significantly alleviated hyperalgesia in 6-OHDA rats associated with the inhibition of sEPSCs in dorsal horn neurons. In addition, we found that serum PACAP-38 concentration was significantly increased in PD patients with pain, and positively correlated with numerical rating scale score. In conclusion, activation of PACAP/PAC1-R induces the development of PD pain and targeting PACAP/PAC1-R is an alternative strategy for treating PD pain.

Overlapping Epstein-Barr virus encephalitis and autoimmune glial fibrillary acidic protein astrocytopathy.

We describe three cases of overlapping Epstein-Barr virus (EBV) Encephalitis and Autoimmune Glial Fibrillary Acidic Protein Astrocytopathy (GFAP-A). The three cases all presented with initial symptoms of fever, headache, coma, and posture tremor of the upper limbs, then followed by limb weakness and dysuria. All of the three cases were on ventilators. Case 1 and 2 improved dramatically after intravenous methylprednisoloneand immunoglobulin treatment. However, case 3 presented dyspneic, and died from gastrointestinal hemorrhage. The GFAP-A triggered by EBV intracranial infection could initially masquerade as EBV encephalitis only, and the detection of GFAP antibody is essential for differentiation.

Cystathionine ß-Synthase Suppresses NLRP3 Inflammasome Activation via Redox Regulation in Microglia.

Aims: Cystathionine ß-synthase (CBS) is essential for homocysteine (Hcy) transsulfuration, yielding cysteine as a common precursor of hydrogen sulfide (H2S), glutathione (GSH), and other sulfur molecules, which produce neuroprotective effects in neurological conditions. We previously reported a disruption of microglial CBS/H2S signaling in a Parkinson's disease (PD) mouse model. Yet, it remains unclear whether CBS affects nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) inflammasome activity and other pathologies in PD. Results: Microglial CBS expression decreased after lipopolysaccharide (LPS) stimulation. Elevated GSSG (the oxidized GSH) content and decreased H2S generation were found in the brains of microglial cbs conditional-knockout (cbscKO) mice, whereas serum and brain Hcy levels remained unaltered. Moreover, microglial cbscKO mice were susceptible to NLRP3 inflammasome activation and dopaminergic neuron losses caused by LPS injection into the substantia nigra, whereas cbs overexpression or activation produced opposite effects. In vitro studies showed that cbs overexpression or activation suppressed microglial NLRP3 inflammasome activation and interleukin (IL)-1ß secretion by reducing mitochondrial reactive oxygen species (mitoROS) level. Conversely, ablation of cbs enhanced NLRP3 expression and mitoROS generation and augmented microglial NLRP3 inflammasome activity in response to adenosine triphosphate challenge, which was blocked by the mitoROS scavenger. Innovation and Conclusion: The study demonstrated an elevated GSSG level and reduced H2S generation, which correlated with a susceptible status of microglia in the brain of cbscKO mice. Our findings reveal a critical role of CBS in restraining the microglial NLRP3 inflammasome by controlling redox homeostasis and highlight that activation or upregulation of CBS may become a potential strategy for PD treatment.

Roles of protein post-translational modifications in glucose and lipid metabolism: mechanisms and perspectives.

The metabolism of glucose and lipids is essential for energy production in the body, and dysregulation of the metabolic pathways of these molecules is implicated in various acute and chronic diseases, such as type 2 diabetes, Alzheimer's disease, atherosclerosis (AS), obesity, tumor, and sepsis. Post-translational modifications (PTMs) of proteins, which involve the addition or removal of covalent functional groups, play a crucial role in regulating protein structure, localization function, and activity. Common PTMs include phosphorylation, acetylation, ubiquitination, methylation, and glycosylation. Emerging evidence indicates that PTMs are significant in modulating glucose and lipid metabolism by modifying key enzymes or proteins. In this review, we summarize the current understanding of the role and regulatory mechanisms of PTMs in glucose and lipid metabolism, with a focus on their involvement in disease progression associated with aberrant metabolism. Furthermore, we discuss the future prospects of PTMs, highlighting their potential for gaining deeper insights into glucose and lipid metabolism and related diseases.

Plasma sDPP4 (Soluble Dipeptidyl Peptidase-4) and Cognitive Impairment After Noncardioembolic Acute Ischemic Stroke.

BACKGROUND: DPP4 (dipeptidyl peptidase-4) inhibitors have been proven to promote neuronal regeneration, reverse the development of cognitive deficits. However, the association of circulating soluble form (sDPP4 [soluble DPP4]) with poststroke cognitive impairment (PSCI) is unclear. We aimed to investigate the association between plasma sDPP4 levels and PSCI in patients with ischemic stroke. METHODS: A total of 600 noncardioembolic stroke patients were included based on a preplanned ancillary study from the CATIS (China Antihypertensive Trial in Acute Ischemic Stroke). We used the Montreal Cognitive Assessment to evaluate cognitive function at 3 months follow-up after ischemic stroke. Binary logistic regression analyses were performed to investigate the association of plasma sDPP4 levels with subsequent PSCI. We further calculated integrated discrimination improvement and category-free net reclassification improvement to investigate the incremental prognostic effect of plasma sDPP4 beyond the basic model with conventional risk factors. RESULTS: Plasma sDPP4 was inversely associated with PSCI after ischemic stroke, and the adjusted odds ratio (95% CI) for the highest versus lowest quartile of sDPP4 was 0.49 (0.29-0.81; P for trend=0.011). Each 1-SD increase of logarithm-transformed plasma sDPP4 concentration was associated with 17% (odds ratio, 0.83 [95% CI, 0.70-0.99]) lower risk of PSCI. Adding plasma sDPP4 to the basic model notably improved risk reclassification for PSCI, as shown by a category-free net reclassification improvement of 19.10% (95% CI, 2.52%-35.68%; P=0.03) and integrated discrimination improvement of 0.79% (95% CI, 0.13%-1.46%; P=0.02). CONCLUSIONS: Higher plasma sDPP4 levels were associated with decreased risk of cognitive impairment after noncardioembolic ischemic stroke.

Protein glycosylation in urine as a biomarker of diseases.

Human body fluids have become an indispensable resource for clinical research, diagnosis and prognosis. Urine is widely used to discover disease-specific glycoprotein biomarkers because of its recurrently non-invasive collection and disease-indicating properties. While urine is an unstable fluid in that its composition changes with ingested nutrients and further as it is excreted through micturition, urinary proteins are more stable and their abnormal glycosylation is associated with diseases. It is known that aberrant glycosylation can define tumor malignancy and indicate disease initiation and progression. However, a thorough and translational survey of urinary glycosylation in diseases has not been performed. In this article, we evaluate the clinical applications of urine, introduce methods for urine glycosylation analysis, and discuss urine glycoprotein biomarkers. We emphasize the importance of mining urinary glycoproteins and searching for disease-specific glycosylation in various diseases (including cancer, neurodegenerative diseases, diabetes, and viral infections). With advances in mass spectrometry-based glycomics/glycoproteomics/glycopeptidomics, characterization of disease-specific glycosylation will optimistically lead to the discovery of disease-related urinary biomarkers with better sensitivity and specificity in the near future.

Pramipexole inhibits astrocytic NLRP3 inflammasome activation via Drd3-dependent autophagy in a mouse model of Parkinson's disease.

Inflammation is one of the pathogenic processes in Parkinson's disease (PD). Dopamine receptor agonist pramipexole (PPX) is extensively used for PD treatment in clinics. A number of studies show that PPX exerts neuroprotection on dopaminergic (DA) neurons, but the molecular mechanisms underlying the protective effects of PPX on DA neurons are not fully elucidated. In the present study, we investigated whether PPX modulated PD-related neuroinflammation and underlying mechanisms. PD model was established in mice by bilateral striatum injection of lipopolyssaccharide (LPS). The mice were administered PPX (0.5 mg·kg-1·d-1, i.p.) 3 days before LPS injection, and for 3 or 21 days after surgery, respectively, for biochemical and histological analyses. We showed that PPX administration significantly alleviated the loss of DA neurons, and suppressed the astrocyte activation and levels of proinflammatory cytokine IL-1ß in the substantia nigra of LPS-injected mice. Furthermore, PPX administration significantly decreased the expression of NLRP3 inflammasome-associated proteins, i.e., cleaved forms of caspase-1, IL-1ß, and apoptosis-associated speck-like protein containing a caspase recruit domain (ASC) in the striatum. These results were validated in LPS+ATP-stimulated primary mouse astrocytes in vitro. Remarkably, we showed that PPX (100-400 µM) dose-dependently enhanced the autophagy activity in the astrocytes evidenced by the elevations in LC3-II and BECN1 protein expression, as well as the increase of GFP-LC3 puncta formation. The opposite effects of PPX on astrocytic NLRP3 inflammasome and autophagy were eliminated by Drd3 depletion. Moreover, we demonstrated that both pretreatment of astrocytes with autophagy inhibitor chloroquine (40 µM) in vitro and astrocyte-specific Atg5 knockdown in vivo blocked PPX-caused inhibition on NLRP3 inflammasome and protection against DA neuron damage. Altogether, this study demonstrates an anti-neuroinflammatory activity of PPX via a Drd3-dependent enhancement of autophagy activity in astrocytes, and reveals a new mechanism for the beneficial effect of PPX in PD therapy.

BAG3 promotes autophagy and suppresses NLRP3 inflammasome activation in Parkinson's disease.

Background: Neuroinflammation mediated by microglia plays a key role in the pathogenesis of Parkinson's disease (PD), and our previous studies showed this was significantly inhibited by enhanced autophagy. In the autophagy pathway, Bcl2-associated athanogene (BAG)3 is a prominent co-chaperone, and we have shown BAG3 can regulate autophagy to clear the PD pathogenic protein α-synuclein. However, the connection between BAG3 and microglia mediated neuroinflammation is not clear. Methods: In this study, we explored whether BAG3 regulated related neuroinflammation and its original mechanism in PD. An inflammatory model of PD was established by injecting adeno-associated virus (AAV)-BAG3 into the bilateral striatum of C57BL/6 male mice to induce overexpression of BAG3, followed by injection of lipopolysaccharide (LPS). The striatum was extracted at 3 days after injection of LPS for Western blotting and reverse transcription quantitative polymerase chain reaction (RT-qPCR), and immunohistochemical staining was performed at 21 days after injection. At the same time, LPS was used to induce activation of BV2 cells to verify the effect of BAG3 in vitro. Results: Overexpression of BAG3 reduced LPS-induced pyroptosis by reducing activation of caspase-1, the NOD-like receptor family, and the pyrin domain-containing 3 (NLRP3) inflammasome, and by release of interleukin (IL)-1ß and tumor necrosis factor (TNF)-α. The LPS-induced inflammatory environment inhibits autophagy, and overexpression of BAG3 can restore autophagy, which may be the mechanism by which BAG3 reduces neuronal inflammation in PD. Conclusions: Our results demonstrate BAG3 promotes autophagy and suppresses NLRP3 inflammasome formation in PD.

G2019S LRRK2 Mutation Enhances MPP+-Induced Inflammation of Human Induced Pluripotent Stem Cells-Differentiated Dopaminergic Neurons.

Induced pluripotent stem cells (iPSCs) offer an unprecedented opportunity to mimic human diseases of related cell types, but it is unclear whether they can successfully mimic age-related diseases such as Parkinson's disease (PD). We generated iPSCs lines from three patients with familial PD associated with the G2019S mutation in the LRRK2 gene and one age-matched healthy individual (control). During long-term culture, dopaminergic (DA) neurons differentiated from iPSCs of G2019S LRRK2 PD patients exhibited morphological changes, including a reduced number of neurites and neurite arborization, which were not evident in DA neurons differentiated from control iPSCs. To mimic PD pathology in vitro, we used 1-methyl-4-phenylpyridium (MPP+) to damage DA neurons and found that DA neurons differentiated from patients with G2019S LRRK2 mutation significantly reduced the survival rate and increased apoptosis compared with the controls. We also found that the mRNA level of inflammatory factors [interleukin (IL)-1ß, tumor necrosis factor-α, cyclooxygenase-2, IL-6, and inducible NO synthase] with G2019S LRRK2 mutation were higher than control group after exposure to MPP+. Our study provides an in vitro model based on iPSCs that captures the patients' genetic complexity and investigates the pathogenesis of familial PD cases in a disease-associated cell type.

Multiple stress fractures of unilateral femur: A case report.

BACKGROUND: Stress fractures of the femoral neck are not common in clinical practice, and simultaneous stress fractures of the femoral neck and proximal femur of the unilateral femur are even more rare. We introduce a case of this type of fracture that was treated in our department, analyze the causes, and review similar stress fractures reported in the literature to provide references for the diagnosis and treatment of such conditions. CASE SUMMARY: A 62-year-old female, with a free medical history, was admitted to the hospital mainly due to pain in the right hip and worsening pain in the right thigh. The patient had no obvious history of trauma. X-ray and computed tomography showed fracture of the femoral neck and proximal femur. The patient had undergone surgery 1 year prior to address a fracture of the left proximal femur that had occurred in a traffic accident. Our first consideration was stress fracture of the femoral neck; however, simultaneous stress fractures of the femoral neck and proximal femur of the unilateral femur were seen. The femoral neck stress fracture was a tension fracture, with obvious displacement and varus deformity of the hip. Considering that the patient was an elderly female, we performed total hip arthroplasty. Follow-up X-rays showed that the stress fracture of the proximal femur had mostly healed after 3 mo. CONCLUSION: Muscle fatigue and hip varus deformity provide an anatomical basis for the occurrence of femoral neck stress fractures.

Identifying the hub gene and immune infiltration of Parkinson's disease using bioinformatical methods.

BACKGROUND: Parkinson's disease (PD) is a common neurodegenerative disorder that affects 1%-2% of the population over 60 years old. Immune response dysfunction in the brain contributes to the occurrence and development of PD. This study aimed to uncover the potential diagnostic genes for PD and characterize the immune cell infiltrates. METHODS: We downloaded the microarray data of patients with PD samples from the Gene Expression Omnibus (GEO) database. Weighted Gene Co-Expression Network Analysis (WGCNA) was used to identify the modules linked to PD in the GSE20163 dataset. Meanwhile, differentially expressed genes (DEGs) between the healthy control samples and PD samples were also identified. Then the PD-related genes were integrated based on the genes in the key module and DEGs. Functional enrichment analysis was used to explore the molecular mechanisms of these PD-related genes. Protein-protein interaction (PPI) network and least absolute shrinkage and selection operator (LASSO) analysis were used to further screen candidate genes for PD. Gene set enrichment analysis (GSEA) was applied to explore the biological functions of these candidate genes. The infiltration of immune cells was detected by single-sample gene set enrichment analysis (ssGSEA) algorithm in the GSE20163 dataset, and Pearson analysis was used to investigate the correlation of candidate genes with immune cells and immune checkpoint proteins. The expression of candidate genes in clinical samples was verified by qPCR. RESULTS: Altogether, we found a unique gene module related to PD, where 109 DEGs were identified in the GSE20163 dataset. Following these results, we screened 68 genes associated with PD. Gene Expression Omnibus (GEO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses suggested that these genes were markedly enriched in the pathway of synthesis and transport of neurons. Three candidate genes (SLC18A2, CALB1, and SYNGR3) were further identified in PD patients through PPI network and LASSO analysis. The receiver operating characteristic (ROC) curve indicated that the three candidate genes had a good performance in distinguishing the PD samples from healthy control samples. The proportions of the aDC, DC, NK CD56dim cells, and follicular helper T cells (TFH) were obviously different between the healthy control and PD samples. Moreover, CTLA4, LAG3, CEACAM1, and CD27 were highly expressed in the PD group. GSEA analysis for candidate genes revealed that they were all closely related to the neurogenic disease. Additionally, the three candidate genes were all strongly correlated with the above immune cells and immune checkpoint proteins. The qPCR results validated the expression differences of SLC18A2 and SYNGR3 in the clinical PD and control samples. CONCLUSION: The three candidate genes may be a useful tool for diagnosing PD patients. These findings provide a reference for exploring new therapeutic targets and strategies for PD treatment.

Protection against glutathione depletion-associated oxidative neuronal death by neurotransmitters norepinephrine and dopamine: Protein disulfide isomerase as a mechanistic target for neuroprotection.

Oxidative stress is extensively involved in neurodegeneration. Clinical evidence shows that keeping the mind active through mentally-stimulating physical activities can effectively slow down the progression of neurodegeneration. With increased physical activities, more neurotransmitters would be released in the brain. In the present study, we investigated whether some of the released neurotransmitters might have a beneficial effect against oxidative neurodegeneration in vitro. Glutamate-induced, glutathione depletion-associated oxidative cytotoxicity in HT22 mouse hippocampal neuronal cells was used as an experimental model. We showed that norepinephrine (NE, 50 µM) or dopamine (DA, 50 µM) exerted potent protective effect against glutamate-induced cytotoxicity, but this effect was not observed when other neurotransmitters such as histamine, γ-aminobutyric acid, serotonin, glycine and acetylcholine were tested. In glutamate-treated HT22 cells, both NE and DA significantly suppressed glutathione depletion-associated mitochondrial dysfunction including mitochondrial superoxide accumulation, ATP depletion and mitochondrial AIF release. Moreover, both NE and DA inhibited glutathione depletion-associated MAPKs activation, p53 phosphorylation and GADD45α activation. Molecular docking analysis revealed that NE and DA could bind to protein disulfide isomerase (PDI). In biochemical enzymatic assay in vitro, NE and DA dose-dependently inhibited the reductive activity of PDI. We further revealed that the protective effect of NE and DA against glutamate-induced oxidative cytotoxicity was mediated through inhibition of PDI-catalyzed dimerization of the neuronal nitric oxide synthase. Collectively, the results of this study suggest that NE and DA may have a protective effect against oxidative neurodegeneration through inhibition of protein disulfide isomerase and the subsequent activation of the MAPKs‒p53‒GADD45α oxidative cascade.

Induction of Parkinsonian-Like Changes via Targeted Downregulation of Astrocytic Glutamate Transporter GLT-1 in the Striatum.

BACKGROUND: Previous investigations have suggested that decreased expression of glutamate transporter-1 (GLT-1) is involved in glutamate excitotoxicity and contribute to the development of Parkinson's disease (PD), GLT-1 is decreased in animal models of PD. GLT-1 is mainly expressed in astrocytes, and the striatum is a GLT-1-rich brain area. OBJECTIVE: The aim was to explore the function and mechanism of astrocytic GLT-1 in PD-like changes. METHODS: In the study, PD-like changes and their molecular mechanism in rodents were tested by a behavioral assessment, micro-positron emission tomography/computed tomography (PET/CT), western blotting, immunohistochemical and immunofluorescence staining, and high performance liquid chromatography pre-column derivatization with O-pthaldialdehida after downregulating astrocytic GLT-1 in vivo and in vitro. RESULTS: In vivo, after 6 weeks of brain stereotactic injection of adeno-associated virus into the striatum, rats in the astrocytic GLT-1 knockdown group showed poorer motor performance, abnormal gait, and depression-like feature; but no olfactory disorders. The results of micro-PET/CT and western blotting indicated that the dopaminergic system was impaired in astrocytic GLT-1 knockdown rats. Similarly, tyrosine hydroxylase (TH) positive immune-staining in neurons of astrocytic GLT-1 knockdown rats showed deficit in cell count. In vitro, knockdown of astrocytic GLT-1 via RNA interference led to morphological injury of TH-positive neurons, which may be related to the abnormal calcium signal induced by glutamate accumulation after GLT-1 knockdown. Furthermore, the GLT-1 agonist ceftriaxone showed a protective effect on TH-positive neuron impairment. CONCLUSION: The present findings may shed new light in the future prevention and treatment of PD based on blocking glutamate excitotoxicity.

Role of G Protein-Coupled Receptors in Microglial Activation: Implication in Parkinson's Disease.

Parkinson's disease (PD) is one of the prevalent neurodegenerative diseases associated with preferential loss of dopaminergic (DA) neurons in the substantia nigra compacta (SNc) and accumulation of α-synuclein in DA neurons. Even though the precise pathogenesis of PD is not clear, a large number of studies have shown that microglia-mediated neuroinflammation plays a vital role in the process of PD development. G protein-coupled receptors (GPCRs) are widely expressed in microglia and several of them act as regulators of microglial activation upon corresponding ligands stimulations. Upon α-synuclein insults, microglia would become excessively activated through some innate immune receptors. Presently, as lack of ideal drugs for treating PD, certain GPCR which is highly expressed in microglia of PD brain and mediates neuroinflammation effectively could be a prospective source for PD therapeutic intervention. Here, six kinds of GPCRs and two types of innate immune receptors were introduced, containing adenosine receptors, purinergic receptors, metabotropic glutamate receptors, adrenergic receptors, cannabinoid receptors, and melatonin receptors and their roles in neuroinflammation; we highlighted the relationship between these six GPCRs and microglial activation in PD. Based on the existing findings, we tried to expound the implication of microglial GPCRs-regulated neuroinflammation to the pathophysiology of PD and their potential to become a new expectation for clinical therapeutics.

α-synuclein suppresses microglial autophagy and promotes neurodegeneration in a mouse model of Parkinson's disease.

The cell-to-cell transfer of α-synuclein (α-Syn) greatly contributes to Parkinson's disease (PD) pathogenesis and underlies the spread of α-Syn pathology. During this process, extracellular α-Syn can activate microglia and neuroinflammation, which plays an important role in PD. However, the effect of extracellular α-Syn on microglia autophagy is poorly understood. In the present study, we reported that extracellular α-Syn inhibited the autophagy initiation, as indicated by LC3-II reduction and p62 protein elevation in BV2 and cultured primary microglia. The in vitro findings were verified in microglia-enriched population isolated from α-Syn-overexpressing mice induced by adeno-associated virus (AAV2/9)-encoded wildtype human α-Syn injection into the substantia nigra (SN). Mechanistically, α-Syn led to microglial autophagic impairment through activating toll-like receptor 4 (Tlr4) and its downstream p38 and Akt-mTOR signaling because Tlr4 knockout and inhibition of p38, Akt as well as mTOR prevented α-Syn-induced autophagy inhibition. Moreover, inhibition of Akt reversed the mTOR activation but failed to affect p38 phosphorylation triggered by α-Syn. Functionally, the in vivo evidence showed that lysozyme 2 Cre (Lyz2cre )-mediated depletion of autophagy-related gene 5 (Atg5) in microglia aggravated the neuroinflammation and dopaminergic neuron losses in the SN and exacerbated the locomotor deficit in α-Syn-overexpressing mice. Taken together, the results suggest that extracellular α-Syn, via Tlr4-dependent p38 and Akt-mTOR signaling cascades, disrupts microglial autophagy activity which synergistically contributes to neuroinflammation and PD development.