Nonalcoholic fatty liver disease (NAFLD), along with metabolic syndrome and obesity, has seen a dramatic rise in frequency around the world. Arctigenin (ATG) has better efficacy in the treatment of non-alcoholic fatty liver in vitro, but its application in vivo models is limited due to its low oral bioavailability and poor water solubility. Compound ARC-18 is a derivative of ATG. As a prodrug, it improves the bioavailability and solubility of ATG, so that ARC-18 can also have better pharmacological activity in oral administration. The main purpose of this article is to explore the therapeutic effect of compound ARC-18 in nonalcoholic steatohepatitis and its possible mechanism. C57BL/6 J mice were fed with methionine and choline deficient (MCD) diet for 8 weeks to induce NAFLD and were given ARC-18 by gavage for 6 weeks (Proteomics Analysis and Histopathological analysis). The effect of ARC-18 was also investigated in AML12 cells exposed to palmitic acid. Our study showed that ARC-18 (33.0 mg/kg body weight) improved hepatic lipid accumulation, inflammatory damage and liver fibrosis in MCD diet feeding mice. Proteomic analyses indicated that ARC-18 exerts beneficial effects on inflammation, oxidative stress, and lipid metabolism. ARC-18 decreased lipid accumulation and promoted lipid oxidation by activating adiponectin receptor 1 (Adipo1 receptor) -mediated Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) signaling pathway in vivo and vitro. Our data implies that ARC-18 is a therapeutic medication for the treatment of NAFLD, and that this protective effect is achieved by activating the hepatic Adipo1 receptor-mediated AMPK signaling pathway.

Alzheimer’s disease (AD), the most prevalent form of dementia, is characterized as a slowly progressing neurodegenerative disease marked by senile plaques and neurofibrillary tangles due to the buildup of amyloid-beta peptide (Aβ) and phosphorylated tau in the brain. It is reported that arctigenin (ATG) reduces the level of the enzyme 1 that cleaves β-site amyloid precursor protein and increases Aβ clearance by enhancing autophagy. Compound ARC-18 is a derivative of ATG. The main objective of this study is to investigate whether ARC-18 could improve cognitive function and disease progression by promoting autophagy in Alzheimer-like animal models. Three-month-old 5 × FAD mice were orally treated with the drug for three consecutive months. Water maze and novel object recognition were used to assess cognitive abilities of 5 × FAD mice. In the hippocampus of the mice’ brain, APP processing-related proteins (sAPPβ, BACE1) and autophagy-related proteins (LC3B, P62, LAMP1) were detected. N2a/APPswe cells were used to do experiments to further identify the effect and mechanisms of the drug. Our study demonstrated that ARC-18 enhances the behavioral performance of 5 × FAD mice and mitigates Aβ aggregation in the hippocampus and cortex. This effect is achieved through the activation of adiponectin receptor 1 (AdipoR1)-mediated autophagy and the reduction of Aβ production by modulating amyloid precursor protein (APP) processing. Therefore, ARC-18 holds promise as a potential therapeutic agent for Alzheimer’s disease.

Huntington's disease (HD) is a rare, inherited neurodegenerative disorder caused by mutations in the huntingtin (HTT) gene. The classic concept is that HD is a degenerative disease that primarily affects the striatum, caused by a gain-of-function mutant mHTT that kills neurons. However, increasing evidence suggests that the effects of mHTT on development may be an alternative view of HD. Therefore, we describe the importance of HTT for neurodevelopment and then summarize the effects of mHTT on neurodevelopment that have been revealed so far in different models. Importantly, we provide new insights into the use of different models to study HD development, and propose new therapeutic strategies for intervening in HD early in development to improve disease progression. Furthermore, we explore potential connections between neurodevelopmental abnormalities and neurodegenerative processes in HD. This review provides a systematic synthesis of current knowledge regarding HD development and pathogenesis.

Amyotrophic lateral sclerosis (ALS) is an devastating neurodegenerative disorder with a very fast course and a very high fatality rate. The review discusses the intricate pathophysiology of ALS, such as the alterations caused by the genetic mutations of the C9orf72 and SOD1 genes, the misfolding and aggregation of proteins, oxidative stress, the excitotoxicity of glutamate, neuroinflammation, malfunctions in mitochondria, and axonal transport. Heterogeneity of the disease makes the development of biomarkers in ALS challenging; however, some promising candidates have been identified. Protein aggregation markers, including TDP-43 and SOD1, oxidative stress markers, such as 8-oxodG, neuroinflammatory markers, such as CRP and MCP-1, and neurological injury markers, such as NfL and pNfH, have potential in diagnosis, monitoring, and prediction. The miRNAs and particular metabolites can also provide clues to the molecular basis of ALS. The creation of biomarkers is challenged by the presence of a significant amount of disease heterogeneity and the lack of animal model reliability. The review highlights the importance of further research on biomarkers aimed at improving the diagnosis, treatment, and development of drugs for ALS. It supports the concept of a systematic biomarker development process, including genetic testing and molecular subgroup analysis, to enhance diagnostic accuracy and prognostic prediction capabilities. Exploring the interrelationship between the pathological process of ALS and the treatment based on multi-biomarker strategies is crucial for achieving effective management of this disease. As our understanding of ALS deepens, we expect to discover more new biomarkers in the future. This will significantly improve the diagnosis, treatment, and overall management of this devastating diseas.

BACKGROUND AND PURPOSE:Cullin 4B (Cul4B), a scaffold protein of CUL4B-RING E3 ligase complex, functions primarily as a negative regulator of inflammation. This study aims to investigate its role in acute lung injury (ALI) and NLRP3 ubiquitination. EXPERIMENTAL APPROACH:Cul4B expression and distribution were examined in two sepsis-induced ALI mouse models. Lung-specific overexpression and myeloid-specific Cul4B deletion were applied to evaluate its effects on lung histopathology, pulmonary inflammation, and alveolar capillary barrier dysfunction. The role of Cul4B in NLRP3 inflammasome activation was evaluated in THP-1 cells and Cul4B-deficient BMDMs. Cul4B-NLRP3 interaction and Cul4B-mediated NLRP3 ubiquitination were determined. The involvement of Cul4B in MN-08-mediated ALI protection and inflammasome suppression was also investigated. KEY RESULTS:Cul4B was significantly downregulated in lung tissues of sepsis-induced ALI mice. Pulmonary overexpression of Cul4B attenuated lung injury and inflammation, whereas myeloid-specific deletion of Cul4B exacerbated epithelial barrier damage and inflammatory responses, along with enhanced Nigericin-induced NLRP3 inflammasome activation. Cul4B was shown to interact with both endogenous and exogenous NLRP3 and suppressed NLRP3 inflammasome activation by promoting NLRP3 ubiquitination. MN-08, a small molecule previously reported to alleviate lipopolysaccharide (LPS)-induced ALI, was found to upregulate Cul4B expression and inhibit NLRP3 inflammasome activation by promoting NLRP3 ubiquitination. CONCLUSIONS AND IMPLICATIONS:Upregulating Cul4B promotes NLRP3 ubiquitination, thereby inhibiting inflammasome activation and mitigating pulmonary epithelial barrier dysfunction and inflammation in ALI. Cul4B emerges as a potential therapeutic target, and these findings provide new insights into the mechanism by which MN-08 protects against sepsis-induced ALI.

Small interfering RNA (siRNA) therapeutics offer considerable promise due to their gene-specific mechanism of action via Watson–Crick base pairing. While GalNAc–siRNA conjugation has demonstrated success in liver-targeted therapies, addressing complex diseases such as cancer and cardiovascular disorders often requires simultaneous silencing of multiple genes. Here, we report a dual-siRNA delivery platform composed of two modular building blocks: DL05 and B13. DL05 features a GalNAc moiety and an azide group for conjugation, while B13 contains an alkyne group for click chemistry with DL05. Both modules were synthesized efficiently and linked to solid supports for oligonucleotide elongation. A proof-of-concept experiment demonstrated the conjugation of two siRNA strands bearing unnatural nucleotides via click chemistry, followed by annealing with complementary antisense strands to form a dual-target siRNA construct. Structural validation via HPLC and mass spectrometry confirmed the success of this approach, highlighting the potential of this delivery platform for multigene-targeting siRNA therapies.

Brain accumulation of toxic soluble α-synuclein (α-syn) oligomers represents a prodromal marker of synucleinopathies in Parkinson’s disease (PD), contributing to progressive nigrostriatal neurodegeneration. Dysfunction in beta-glucocerebrosidase (GCase) and leucine-rich repeat kinase 2 (LRRK2) mutation are genetic risks for developing synucleinopathies. However, whether pharmacological GCase activation ameliorated synucleinopathies in LRRK2-PD was unexplored. Here, we showed that long-term treatment of ambroxol (ABX), a brain-penetrant GCase activator, reduced α-syn oligomer accumulation in aged mutant LRRK2R1441G mouse striatum. Acute ABX treatment (50 µM) increased cellular GCase enzymatic activity and reduced Ser129-α-syn phosphorylation in human SH-SY5Y cells and mutant LRRK2 mouse fibroblasts, independent to LRRK2 kinase activity. Real-time DQ-BSA assay revealed lysosomal dysfunction in mutant MEFs, which was partially attenuated by ABX treatment. Lysosomal stress by bafilomycin-A1 induced endogenous GCase activity in wildtype (WT) MEFs, which was not observed in the LRRK2 mutant. Single gavage of ABX (400 mg/kg) in aged mice achieved peak drug level in serum and brain within 6 h post-administration. Ad libitum feeding of ABX (in food pellets) over 18 weeks (average dose: 45.9 mg/kg/day) elevated brain GCase activity in both WT and mutant striatum without affecting body weight. This regimen significantly reduced α-syn oligomer level in mutant striatum to a comparable physiological level in age-matched WT without altering total α-syn and Ser129-phosphorylation levels. This is the first study demonstrating reduced α-syn oligomer accumulation by chronic treatment of GCase activator in aged mouse brains vulnerable to PD, suggesting early intervention to alter progression of synucleinopathies as a key determinant of clinical outcomes of PD.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease that lacks ideal models to comprehensively recapitulate its pathological features. TDP-43 pathology, a hallmark of neurodegenerative diseases, plays a critical role in disease progression. Given the anatomical and physiological similarities between pig and human brains, large animal models offer a unique advantage in more accurately simulating patient-specific disease characteristics. In this study, we rapidly established a TDP-43-induced neurodegenerative disease model in pigs through ear vein injection of the TDP-43M337V virus. Disease progression was systematically evaluated using behavioral assessments and pathological analyses. This porcine model produced extremely severe motor dysfunction accompanied by significant muscle atrophy and fibrosis. Additionally, characteristic TDP-43 pathological phenotypes were observed, including degeneration of spinal motor neurons and proliferation of glial cells in both the brain and spinal cord. Notably, TDP-43M337V induction led to a significant upregulation of TMEM106B, SOD1, and APOE4 levels. This TDP-43 porcine model recapitulates multiple key features of ALS and serves as a valuable complement to existing animal models, providing a robust platform for investigating TDP-43-related pathogenic mechanisms of TDP-43 and developing effective therapeutics.

Alzheimer's disease (AD) and vascular dementia (VaD) are two prevalent forms of dementia. VaD is linked to cerebrovascular lesions, such as those from white matter ischemia and chronic cerebral hypoperfusion, which can also occur in AD. Nitric oxide (NO) regulates cerebral blood flow (CBF) in the central nervous system. Memantine is an NMDA receptor antagonist approved for AD treatment. This study investigated the efficacy and molecular mechanism of MN-08, a novel memantine nitrate, in one VaD model (2VO) and two AD models (APP/PS1 mice and Aβ1-42-induced mice). MN-08 increased CBF, ameliorated cognitive and memory functions in VaD and AD, and was more effective than memantine. MN-08 increased the survival rate of CA1 neurons and mitigated white matter lesions and axonal damage. Moreover, MN-08 protected neurons from OGD-induced loss and promoted axonal outgrowth in the hippocampus by upregulating phosphorylated Akt (p-Akt), glycogen synthase kinase-3β (p-GSK3β), and high-molecular-weight neurofilaments (p-NFH). The beneficial effects of MN-08 were attenuated by carboxy-PTIO, a potent NO scavenger, suggesting that MN-08-derived NO may alleviate cognitive impairment from cerebral hypoperfusion. Taken together, our studies demonstrate that MN-08 is a promising therapeutic agent for the treatment of dementia including VaD and AD.

Transferrin receptor 1 (TfR1) is a ubiquitously expressed receptor characterized by rapid internalization kinetics and efficient receptor recycling, making it an attractive target for drug delivery. Herein, we investigated the potential of TfR1-binding peptide-siRNA conjugates for central nervous system (CNS)-specific gene silencing. A panel of TfR1-binding peptides and conjugation linkers were synthesized to enable siRNA attachment and evaluate their gene-silencing effects. Conjugation with the hTfR No. 894 peptide achieved effective siRNA delivery both in vitro and in vivo. Compared to ribose 2'-O-hexadecyl (C16)-siRNA conjugates, the hTfR No. 894-siRNA conjugation (POC2) elicited favorable pharmacokinetic characteristics and robust and durable silencing of the target gene across CNS regions following local administration, with minimal impact on peripheral tissues. These findings support TfR1-binding peptide conjugation as a promising strategy for CNS-targeted siRNA delivery.

Importance Tetramethylpyrazine nitrone has exhibited promising results in improving motor dysfunction in neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). Objective To evaluate the safety and efficacy of orally administered tetramethylpyrazine nitrone in patients with ALS. Design, Setting, and Participants This phase 2, multicenter, double-masked, placebo-controlled, randomized clinical trial was conducted from December 24, 2020, through July 14, 2023, in 11 centers in China, with a 180-day follow-up. Patients aged 45 to 70 years, with ALS onset within 2 years, ALS Functional Rating Scale–Revised (ALSFRS-R) scores of at least 2 points on each item, and forced vital capacity (FVC) of at least 80% were included. Patients experienced a 1- to 4-point decrease in ALSFRS-R score during a 3-month screening period. Interventions Patients were randomly assigned 1:1:1 to receive low-dose tetramethylpyrazine nitrone (600 mg twice daily), high-dose tetramethylpyrazine nitrone (1200 mg twice daily), or placebo (twice daily) for 180 days. Main Outcomes and Measures The primary outcome was change in ALSFRS-R score (range of 0-48, with lower scores indicating worse function) from baseline to 180 days. The secondary outcomes were changes in FVC, grip strength, ALS Assessment Questionnaire-40 (ALSAQ-40) score, and end point events. Safety outcomes included adverse events. Results A total of 155 patients (mean [SD] age, 55.0 [6.5] years; 115 men [74.2%]) were randomized (51 [32.9%] to the low-dose tetramethylpyrazine nitrone group, 52 [33.6%] to the high-dose tetramethylpyrazine nitrone group, and 52 [33.6%] to the placebo group). No significant differences were observed in ALSFRS-R score changes between low-dose tetramethylpyrazine nitrone (least squares [LS] mean difference, −0.89 points; 95% CI −3.25 to 1.48 points) and high-dose tetramethylpyrazine nitrone (LS mean difference, −0.20 points; 95% CI −2.48 to 2.07 points) compared with placebo. High-dose tetramethylpyrazine nitrone showed a significantly slower decline in grip strength at day 180 (LS mean difference, 2.46 kg; 95% CI, 0.15-4.76 kg). In a subgroup of patients younger than 65 years with slower disease progression, tetramethylpyrazine nitrone significantly attenuated the decline in grip strength (LS mean difference, 3.63 kg; 95% CI, 0.84-6.41 kg), bulbar scores (LS mean difference, 0.66 points; 95% CI, 0.03-1.29 points), and respiratory scores (LS mean difference, 0.54 points; 95% CI, 0.03-1.06 points). Adverse events were mostly mild or moderate, with no severe treatment-related adverse events or deaths. Conclusions and Relevance This randomized clinical trial demonstrates that tetramethylpyrazine nitrone is safe and well-tolerated in patients with ALS. There was no difference in the primary end point across the low-dose, high-dose, and placebo groups, with significant benefits in a subgroup of younger patients with slower disease progression. Trial Registration ChiCTR Identifier: ChiCTR2000039689

INTRODUCTION:Neuroinflammation derived from the activation of the microglia is considered a vital pathogenic factor of Alzheimer's Disease (AD). T-006, a tetramethylpyrazine derivative, has been found to alleviate cognitive deficits via inhibiting tau expression and phosphorylation in AD transgenic mouse models. Recently, T-006 has been proven to dramatically decrease the levels of total Amyloid β (Aβ) peptide and Glial Fibrillary Acidic Protein (GFAP) and suppress the expression of ionized calcium binding adaptor molecule-1 (Iba-1) in APP/PS1 mice. Therefore, we have further investigated the effects of T-006 on neuroinflammation in AD-like pathology. METHODS:The anti-inflammatory effects of T-006 and its underlying mechanisms were evaluated in Lipopolysaccharide (LPS)-induced AD rats. The potential protective effects against LPS-activated microglia-mediated neurotoxicity were also measured. RESULTS:T-006 significantly improved the cognitive impairment in LPS-induced AD rats by inhibiting the microglia/astrocyte activation. Further cellular assays found that T-006 significantly reserved the anomalous elevation of inflammatory cytokines in LPS-induced BV2 microglial cells in a concentration-dependent manner, while T-006 treatment alone showed no effects on the normal cultured cells. T-006 also reduced the levels of Toll-like Receptor 4 (TLR4)/Myeloid Differentiation protein-88 (MyD88)/NF-κB signaling-related proteins in BV2 cells exposed to LPS stimulation. TAK242, which selectively inhibits TLR4, slightly lessened the effects of T-006 in LPS-treatment BV2 cells without significance. Importantly, T-006 protected neurons against LPS-induced neuroinflammation by inhibiting the Reactive Oxygen Species (ROS) production and maintaining mitochondrial function. CONCLUSION:T-006 inhibited TLR4-mediated MyD88/NF-κB signaling pathways to suppress neuroinflammation in the LPS-induced AD rat model.

Glaucoma stands out as the primary cause of permanent blindness worldwide, marked by elevated intraocular pressure (IOP) and the deterioration of retinal ganglion cells (RGCs) within the optic nerve. This study aimed to investigate the therapeutic efficacy of MN-08, a novel memantine nitrate derivative, in experimental models of glaucoma. In the rat model of retinal ischemia-reperfusion injury, MN-08 prevented the reduction in retinal ganglion cell complex (GCC) thickness and RGC loss. Meanwhile, MN-08 reduced the protein levels of cleaved-caspase-3, caspase-3, and Bax, while increasing the expression of Bcl-2 compared to the model group, indicating that MN-08 exerts an inhibitory effect on the apoptosis of RGCs. Furthermore, MN-08 lowered IOP in the transient ocular hypertension and glaucoma rabbit models. MN-08 significantly increased cGMP levels in human trabecular meshwork cells (HTMCs). Elevation of cGMP induced by MN-08 was eliminated by ODQ. Moreover, MN-08 attenuated the protein levels of MLCK and p-MLC-2 in HTMCs induced by endothelin-1. Additionally, MN-08 exhibited favorable distribution profiles and pharmacokinetic parameters in normal rabbit ocular tissues following topical instillation. These findings indicate that MN-08 could antagonize NMDA receptors to protect RGCs and release NO to relax TM for the treatment of glaucoma.

Oxygen shortage, known as hypoxia, occurs commonly in both physiological and pathological conditions. Transcriptional regulation by hypoxia-inducible factors is a dominant regulatory mechanism controlling hypoxia-responsive genes during acute hypoxia; however, recent studies suggest that post-transcriptional regulation, including RNA degradation, also involves hypoxia-induced gene expression during the chronic hypoxia. In this study, we developed a method to quantify the contributions of RNA synthesis and degradation to differential gene expression, and identified 102 genes mainly regulated via RNA degradation under chronic hypoxia in HCT116 cells. Bioinformatics analysis showed that the genes mainly regulated by RNA degradation were involved in glycolysis. We examined changes in the RNA-binding ability of RNA-binding proteins by RNA interactome capture and statistical analysis using public databases. We identified fragile X messenger ribonucleoprotein 1 (FMRP) as an RNA-binding protein involved in the chronic hypoxia-induced increase in mRNAs encoding rate-limiting enzymes. This study emphasizes the importance of post-transcriptional gene regulation under chronic hypoxia in HCT116 cells.

Huntington’s disease (HD) is a progressive neurodegenerative disorder characterized by motor dysfunction and cognitive decline. While retinal abnormalities have been documented in some HD patients and animal models, the nature of these abnormalities—specifically whether they originate in the inner or outer retina—remains unclear, particularly regarding their progression with age. This study investigates the retinal structure and function in HD transgenic mice (R6/1) compared to C57BL/6 J control mice at 2, 4, and 6 months of age, encompassing both pre-symptomatic and symptomatic stages of HD. Pathological assessments of the striatum and evaluations of motor function confirmed significant HD-related alterations in R6/1 mice at 6 months. Visual function was subsequently analyzed, accompanied by immunofluorescent staining of retinal and optic nerve tissues over time. Our findings revealed that R6/1 mice exhibited pronounced HD symptoms at 6 months, characterized by neuronal loss in the striatum and impaired locomotor abilities. Functionally, visual acuity declined at 6 months, while retinal light responses began to deteriorate by 4 months. Structurally, R6/1 mice demonstrated a global reduction in cone opsin expression as early as 2 months, with a decrease in rhodopsin levels at 4 months, alongside a thinner retinal structure compared to controls. Notably, rod bipolar cell populations were decreased at 6 months, exhibiting shorter dendritic branches and reduced synaptic connections with photoreceptors in the outer retina. Additionally, ganglion cell numbers in the inner retina decreased at 6 months, accompanied by aberrant neural fibers in the optic nerve. Microglial activation was evident at 4 months, while astrocytic activation was observed at 6 months. Aggregates of mutant huntingtin (mHTT) were first detected in the ganglion cell layer and optic nerve at 2 months, subsequently disseminating throughout all retinal layers with advancing age. These results indicate that retinal pathology in R6/1 mice manifests earlier in the outer retina than in the inner retina, which does not align with the progression of mHTT aggregation. Consequently, the R6/1 mouse retina may serve as a more effective model for elucidating the mechanisms underlying HD and evaluating potential therapeutic strategies, rather than functioning as an early diagnostic tool for the disease.

Recent research has shown the presence of blood-brain barrier (BBB) breakdown in Alzheimer's disease (AD). BBB is a dynamic interface consisting of a continuous monolayer of brain endothelial cells (BECs) enveloped by pericytes and astrocytes. The restricted permeability of BBB strictly controls the exchange of substances between blood and brain parenchyma, which is crucial for brain homeostasis by excluding blood-derived detrimental factors and pumping out brain-derived toxic molecules. BBB breakdown in AD is featured as a series of BEC pathologies such as increased paracellular permeability, abnormal levels and functions of transporters, and inflammatory or oxidative profile, which may disturb the substance transportation across BBB, thereafter induce CNS disorders such as hypometabolism, Aβ accumulation, and neuroinflammation, eventually aggravate cognitive decline. Therefore, it seems important to protect BEC properties for BBB maintenance and neuroprotection. In this review, we thoroughly summarized the pathological alterations of BEC properties reported in AD patients and numerous AD models, including paracellular permeability, influx and efflux transporters, and inflammatory and oxidative profiles, and probably associated underlying mechanisms. Then we reviewed current therapeutic agents that are effective in ameliorating a series of BEC pathologies, and ultimately protecting BBB integrity and cognitive functions. Regarding the current drug development for AD proceeds extremely hard, this review aims to discuss the therapeutic potentials of targeting BEC pathologies and BBB maintenance for AD treatment, therefore expecting to shed a light on the future AD drug development by targeting BEC pathologies and BBB protection.