RANBP17 Overexpression Restores Nucleocytoplasmic Transport and Ameliorates Neurodevelopment in Induced DYT1 Dystonia Motor Neurons.

DYT1 dystonia is a debilitating neurological movement disorder, and it represents the most frequent and severe form of hereditary primary dystonia. There is currently no cure for this disease due to its unclear pathogenesis. In our previous study utilizing patient-specific motor neurons (MNs), we identified distinct cellular deficits associated with the disease, including a deformed nucleus, disrupted neurodevelopment, and compromised nucleocytoplasmic transport (NCT) functions. However, the precise molecular mechanisms underlying these cellular impairments have remained elusive. In this study, we revealed the genome-wide changes in gene expression in DYT1 MNs through transcriptomic analysis. We found that those dysregulated genes are intricately involved in neurodevelopment and various biological processes. Interestingly, we identified that the expression level of RANBP17, a RAN-binding protein crucial for NCT regulation, exhibited a significant reduction in DYT1 MNs. By manipulating RANBP17 expression, we further demonstrated that RANBP17 plays an important role in facilitating the nuclear transport of both protein and transcript cargos in induced human neurons. Excitingly, the overexpression of RANBP17 emerged as a substantial mitigating factor, effectively restoring impaired NCT activity and rescuing neurodevelopmental deficits observed in DYT1 MNs. These findings shed light on the intricate molecular underpinnings of impaired NCT in DYT1 neurons and provide novel insights into the pathophysiology of DYT1 dystonia, potentially leading to the development of innovative treatment strategies.

Assembling a Coculture System to Prepare Highly Pure Induced Pluripotent Stem Cell-Derived Neurons at Late Maturation Stages.

Generation of human induced pluripotent stem cell (hiPSC)-derived motor neurons (MNs) offers an unprecedented approach to modeling movement disorders such as dystonia and amyotrophic lateral sclerosis. However, achieving survival poses a significant challenge when culturing induced MNs, especially when aiming to reach late maturation stages. Utilizing hiPSC-derived motor neurons and primary mouse astrocytes, we assembled two types of coculture systems: direct coculturing of neurons with astrocytes and indirect coculture using culture inserts that physically separate neurons and astrocytes. Both systems significantly enhance neuron survival. Compared with these two systems, no significant differences in neurodevelopment, maturation, and survival within 3 weeks, allowing to prepare neurons at maturation stages. Using the indirect coculture system, we obtained highly pure MNs at the late mature stage from hiPSCs. Transcriptomic studies of hiPSC-derived MNs showed a typical neurodevelopmental switch in gene expression from the early immature stage to late maturation stages. Mature genes associated with neurodevelopment and synaptogenesis are highly enriched in MNs at late stages, demonstrating that these neurons achieve maturation. This study introduces a novel tool for the preparation of highly pure hiPSC-derived neurons, enabling the determination of neurological disease pathogenesis in neurons at late disease onset stages through biochemical approaches, which typically necessitate highly pure neurons. This advancement is particularly significant in modeling age-related neurodegeneration.

Unveiling the therapeutic potential: Evaluation of anti-inflammatory and antineoplastic activity of Magnolia champaca Linn's stem bark isolate through molecular docking insights.

Magnolia champaca Linn. has traditionally been used for medicinal activity in Asia for treating various chronic diseases as well as a source of food, medicines, and other commodities. Due to the long-used history of this plant, the present study was designed to explore the in vitro, in vivo and in silico anti-inflammatory and antineoplastic properties of the methanolic extract and fractions and the pure compound isolated from the most active chloroform fraction (CHF) of the stem bark of the plant. The isolated compound from the most active CHF was characterized and identified as a glycoside, trans-syringin, through chromatographic and spectroscopic (1H-NMR and 13C-NMR) analyses. In the in vitro anti-inflammatory assay, CHF was most effective in inhibiting inflammation and hemolysis of RBCs by 73.91 ± 1.70% and 75.92 ± 0.14%, respectively, induced by heat and hypotonicity compared to standard acetylsalicylic acid. In the egg albumin denaturation assay, CME and CHF showed the highest inhibition by 56.25 ± 0.82% and 65.82 ± 3.52%, respectively, contrasted with acetylsalicylic acid by 80.14 ± 2.44%. In an in vivo anti-inflammatory assay, statistically significant (p < 0.05) decreases in the parameters of inflammation, such as paw edema, leukocyte migration and vascular permeability, were recorded in a dose-dependent manner in the treated groups. In the antineoplastic assay, 45.26 ± 2.24% and 68.31 ± 3.26% inhibition of tumor cell growth for pure compound were observed compared to 73.26 ± 3.41% for standard vincristine. Apoptotic morphologic alterations, such as membrane and nuclear condensation and fragmentation, were also found in EAC cells after treatment with the isolated bioactive pure compound. Such treatment also reversed the increased WBC count and decreased RBC count to normal values compared to the untreated EAC cell-bearing mice and the standard vincristine-treated mice. Subsequently, in silico molecular docking studies substantiated the current findings, and the isolated pure compound and standard vincristine exhibited -6.4 kcal/mol and -7.3 kcal/mol binding affinities with topoisomerase-II. Additionally, isolated pure compound and standard diclofenac showed -8.2 kcal/mol and -7.6 kcal/mol binding affinities with the COX-2 enzyme, respectively. The analysis of this research suggests that the isolated bioactive pure compound possesses moderate to potent anti-inflammatory and antineoplastic activity and justifies the traditional uses of the stem bark of M. champaca. However, further investigations are necessary to analyze its bioactivity, proper mechanism of action and clinical trials for the revelation of new drug formulations.