The Differential Developmental Neurotoxicity of Valproic Acid on Anterior and Posterior Neural Induction of Human Pluripotent Stem Cells.

Valproic acid (VPA), widely used as an antiepileptic drug, exhibits developmental neurotoxicity when exposure occurs during early or late pregnancy, resulting in various conditions ranging from neural tube defects to autism spectrum disorders. However, toxicity during the very early stages of neural development has not been addressed. Therefore, we investigated the effects of VPA in a model where human pluripotent stem cells differentiate into anterior or posterior neural tissues. Exposure to VPA during the induction of neural stem cells induced different developmental toxic effects in a dose-dependent manner. For instance, VPA induced cell death more profoundly during anteriorly guided neural progenitor induction, while inhibition of cell proliferation and enhanced differentiation were observed during posteriorly guided neural induction. Furthermore, acute exposure to VPA during the posterior induction step also retarded the subsequent neurulation-like tube morphogenesis process in neural organoid culture. These results suggest that VPA exposure during very early embryonic development might exhibit cytotoxicity and subsequently disrupt neural differentiation and morphogenesis processes.

Guidelines for Manufacturing and Application of Organoids: Brain.

This study offers a comprehensive overview of brain organoids for researchers. It combines expert opinions with technical summaries on organoid definitions, characteristics, culture methods, and quality control. This approach aims to enhance the utilization of brain organoids in research. Brain organoids, as three-dimensional human cell models mimicking the nervous system, hold immense promise for studying the human brain. They offer advantages over traditional methods, replicating anatomical structures, physiological features, and complex neuronal networks. Additionally, brain organoids can model nervous system development and interactions between cell types and the microenvironment. By providing a foundation for utilizing the most human-relevant tissue models, this work empowers researchers to overcome limitations of two-dimensional cultures and conduct advanced disease modeling research.

Biological production of epicoccamide-aglycone and its cytotoxicity.

Epicoccamide (EPC) is an O-d-mannosylated acyltetramic acid of Epicoccum origin and is a bolaamphiphilic fungal polyketide. EPC displays weak toxicity against Staphylococcus aureus and HeLa cell lines. The EPC biosynthetic gene cluster was previously identified in Epicoccum nigrum and knockout of the glycosyltransferase gene (epcB) abolished EPC production. EPC-aglycone was expected in the epcB knockout but was not found. This study demonstrates that extractive culture using the hydrophobic resin Diaion HP-20 resulted in the production of EPC-aglycone, which was isolated using chromatographic separation techniques, and its structural identity was substantiated by chemical analyses. EPC-aglycone displayed strong antibacterial activity against Staphylococcus aureus, with the minimal inhibitory concentration of 1 µg/mL (64 µg/mL for EPC). EPC-aglycone displayed higher levels of growth inhibition against HeLa cell line (the half inhibitory concentration, 19 µM) and WI-38 (15 µM) cell line than EPC (76 µM and 38 µM vs. HeLa and WI-38, respectively). The dose-response curve fit of growth inhibition indicated that EPC-aglycone adopted a shallow curve (low slope factor), which was different from that of EPC, suggesting that their cellular targets are distinct from each other. This study substantiates that the d-mannose attachment is the final step in EPC biosynthesis, showcasing a glycosylation-mediated modulation of the biological activity of simple acyltetramic acid. This study also highlights the usefulness of extractive cultures in mining cryptic microbial natural products.

Transcriptional Signature of Valproic Acid-Induced Neural Tube Defects in Human Spinal Cord Organoids.

In vertebrates, the entire central nervous system is derived from the neural tube, which is formed through a conserved early developmental morphogenetic process called neurulation. Although the perturbations in neurulation caused by genetic or environmental factors lead to neural tube defects (NTDs), the most common congenital malformation and the precise molecular pathological cascades mediating NTDs are not well understood. Recently, we have developed human spinal cord organoids (hSCOs) that recapitulate some aspects of human neurulation and observed that valproic acid (VPA) could cause neurulation defects in an organoid model. In this study, we identified and verified the significant changes in cell-cell junctional genes/proteins in VPA-treated organoids using transcriptomic and immunostaining analysis. Furthermore, VPA-treated mouse embryos exhibited impaired gene expression and NTD phenotypes, similar to those observed in the hSCO model. Collectively, our data demonstrate that hSCOs provide a valuable biological resource for dissecting the molecular pathways underlying the currently unknown human neurulation process using destructive biological analysis tools.

Time-efficient implantable catheters for draining malignant ascites in terminal cancer patients.

BACKGROUND: There is a need for an improved version of the implantable catheter for malignant ascites in the abdominal cavity. OBJECTIVE: New implantable catheters have been developed that drain ascites from the abdominal cavity to the bladder by applying pressure. Based on pigtail catheters, these newly designed catheters have silicone membranes and apertures. METHODS: Experimental instruments controlled flow rates and water level to observe changes of the activation pressure and its cycle time along flow rates and turns of catheters. Furthermore, various normality tests, difference tests and non-parametric tests were investigated to observe statistical validity. RESULTS: Cycle times were significantly affected by flow rate (3/4 cases of p< 0.05). The effects of flow rate on activation pressure, however, were not significant (1/4 case of p< 0.05). Cycle times were not significantly affected by the number of turns of the catheter (3/8 cases of p< 0.05). In contrast, the effects of the turns on activation pressure were significant (5/8 cases of p< 0.05). CONCLUSION: Overall, there was no significant difference between cycle times for 1.5 turns and 2.0 turns of catheters. In addition, catheters with 1.5 turns have a lower activation pressure than catheters with 2.0 turns. It is possible to customize catheters based on the ascites excretion and urination rates of various terminal patients.

Glymphatic system dysfunction in patients with cluster headache.

INTRODUCTION: The aim of this study was to investigate alterations of the glymphatic system function in patients with cluster headache. METHODS: We enrolled patients with cluster headache and healthy controls, and they underwent brain magnetic resonance imaging (MRI), including diffusion tensor imaging (DTI). We used the MRIcron and DSI studio programs for DTI preprocessing and DTI analysis with perivascular space (DTI-ALPS) index calculation. RESULTS: Fourteen patients with cluster headache and 23 healthy controls were enrolled. The DTI-ALPS indexes of the groups were significantly different. The DTI-ALPS index for the patients with cluster headache was lower than that for the healthy controls (1.586 vs. 1.786, p = 0.044). There was a significant negative correlation between the DTI-ALPS index and age in the patients with cluster headache (r = -0.549, p = 0.042). However, the DTI-ALPS index was not associated with other clinical characteristics, including disease duration and headache intensity (r = -0.405, p = 0.150; r = -0.048, p = 0.869, respectively). CONCLUSION: Patients with cluster headache had a lower DTI-ALPS index than the healthy controls; this might indicate glymphatic system dysfunction in the patients with cluster headache. Further research is required to determine whether glymphatic system dysfunction is related to the pathophysiology of cluster headache.

Cell-line dependency in cerebral organoid induction: cautionary observations in Alzheimer's disease patient-derived induced pluripotent stem cells.

The cerebral organoid (CO) model has been used in the study of various neurodegenerative diseases owing to its physiological implications. However, the CO model may only be representative of certain clinical findings in affected patients, while some features are not recapitulated. In this study, we found that neurons in the CO model from patients with Alzheimer's disease were less responsive to depolarization, in contrast to previous reports. This difference may be partly attributed to the variations in brain spatial identity depending on the genetic background of the induced pluripotent stem cells. Our current observation raises concerns that the phenotypes observed in the CO model need to be carefully evaluated for their clinical implications.

Microfluidic electrode array chip for electrical stimulation-mediated axonal regeneration.

The precise manipulation of the neural stem cell (NSC)-derived neural differentiation is still challenging, and there is a technological barrier to regulate the axonal regeneration in a controlled manner. Here, we developed a microfluidic chip integrated with a microelectrode array as an axonal guidance platform. The microfluidic electrode array chip consisted of two compartments and a bridge microchannel that could isolate and guide the axons. We demonstrated that the NSCs were largely differentiated into neural cells as the electric field was applied to the microfluidic electrode array chip. We also confirmed the synergistic effects of the electrical stimulation (ES) and neurotrophic factor (NF) on axonal outgrowth. This microfluidic electrode array chip can serve as a central nervous system (CNS) model for axonal injury and regeneration. Therefore, it could be a potentially powerful tool for an in vitro model of the axonal regeneration.

Production of human spinal-cord organoids recapitulating neural-tube morphogenesis.

Human spinal-cord-like tissues induced from human pluripotent stem cells are typically insufficiently mature and do not mimic the morphological features of neurulation. Here, we report a three-dimensional culture system and protocol for the production of human spinal-cord-like organoids (hSCOs) recapitulating the neurulation-like tube-forming morphogenesis of the early spinal cord. The hSCOs exhibited neurulation-like tube-forming morphogenesis, cellular differentiation into the major types of spinal-cord neurons as well as glial cells, and mature synaptic functional activities, among other features of the development of the spinal cord. We used the hSCOs to screen for antiepileptic drugs that can cause neural-tube defects. hSCOs may also facilitate the study of the development of the human spinal cord and the modelling of diseases associated with neural-tube defects.

Glymphatic system dysfunction in temporal lobe epilepsy patients with hippocampal sclerosis.

OBJECTIVE: This study aimed to evaluate glymphatic system function in temporal lobe epilepsy (TLE) patients with hippocampal sclerosis (HS) in comparison to healthy controls, using diffusion tensor imaging (DTI)-analysis along the perivascular space (ALPS) method. We hypothesized that there is glymphatic system dysfunction in TLE patients with HS. METHODS: We retrospectively enrolled 25 TLE patients with HS and 26 age- and sex-matched healthy controls. All participants underwent DTI with the same 3T magnetic resonance imaging scanner, and the DTI-ALPS index was calculated. We evaluated the differences in the DTI-ALPS index between TLE patients with HS and healthy controls. Moreover, we evaluated the correlation between the DTI-ALPS index and clinical characteristics of epilepsy, including age, age at seizure onset, duration of epilepsy, and number of anti-seizure medications (ASMs). RESULTS: There was a difference in the DTI-ALPS index between TLE patients with HS and healthy controls. The DTI-ALPS index in TLE patients with HS was lower than that in healthy controls (1.497 vs. 1.668, P = .015). However, there was no difference in the DTI-ALPS index between the newly diagnosed TLE patients with HS and the chronic TLE patients with HS. The DTI-ALPS index was negatively correlated with age (r = -0.420, P = .036). However, the DTI-ALPS index was not correlated with other clinical characteristics, including age at seizure onset, duration of epilepsy, and number of ASMs. SIGNIFICANCE: Our findings showed that the DTI-ALPS index was significantly lower in TLE patients with HS than in healthy controls, indicating the presence of glymphatic system dysfunction in TLE patients with HS. Our study also suggests that the DTI-ALPS method may be useful for evaluating glymphatic system function in epilepsy.

Glymphatic system function in patients with newly diagnosed focal epilepsy.

INTRODUCTION: The aim of this study was to analyze the glymphatic system function and its relationship with clinical characteristics, global diffusion tensor imaging (DTI) parameters, and global structural connectivity in treatment-naïve patients with newly diagnosed focal epilepsy. METHODS: This retrospective single-center study investigated patients with focal epilepsy and healthy controls. All participants underwent routine brain magnetic resonance imaging and DTI. DTI analysis along the perivascular space (DTI-ALPS) was used to evaluate glymphatic system function. We also calculated the measures of global DTI parameters, including whole-brain fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (AD), and radial diffusivity (RD), and performed a graph theoretical network analysis to measure global structural connectivity. RESULTS: A total of 109 patients with focal epilepsy and 88 healthy controls were analyzed. There were no significant differences in the DTI-ALPS index (1.67 vs. 1.68, p = 0.861) between the groups. However, statistically significant associations were found between the DTI-ALPS index and age (r = -0.242, p = 0.01), FA (r = 0.257, p = 0.007), MD (r = -0.469, p < 0.001), AD (r = -0.303, p = 0.001), RD (r = -0.434, p < 0.001), and the assortative coefficient (r = 0.230, p = 0.016) in patients with focal epilepsy. CONCLUSION: The main finding of this study is that DTI-ALPS index is significantly correlated with global DTI parameters and structural connectivity measures of the brain in patients with focal epilepsy. In addition, DTI-ALPS index decreases with age in these patients. We conclude that the DTI-ALPS index can be used to investigate glymphatic system function in patients with focal epilepsy.

The effects of direct hemoperfusion with polymyxin B-immobilized fiber in patients with acute exacerbation of interstitial lung disease.

BACKGROUND: Acute exacerbation of interstitial lung disease (AE-ILD) causes clinically significant deterioration and has an extremely poor prognosis with high mortality. Recently, several studies reported the effectiveness of direct hemoperfusion with a polymyxin B-immobilized fiber column (PMX-DHP) in patients with AE-ILD as a potential therapy. This study describes the clinical effectiveness and safety of PMX-DHP in patients with AE-ILD. METHODS: We retrospectively reviewed the medical records of 10 patients (11 episodes) with AE-ILD treated with PMX-DHP from January 2018 to June 2019. We compared laboratory and physiologic data of the ratio of partial pressure arterial oxygen to fraction of inspired oxygen (P/F ratio) and level of inflammatory markers before and after implementation of PMX-DHP. RESULTS: Ten patients were included according to the 2016 revised definition of acute exacerbation of idiopathic pulmonary fibrosis (IPF). Nine patients had IPF and one patient had fibrotic nonspecific interstitial pneumonia. Most patients (90.9%) were treated with a steroid pulse, and four patients (36.4%) were treated with an immunosuppressant. The median number of PMX-DHP cycles was 2, and the median duration of each cycle was 6 hours. After PMX-DHP, the mean P/F ratio improved (86 [range, 63-106] vs. 145 [86-260], P=0.030) and interleukin-6 and c-reactive protein decreased (79 [35-640] vs. 10 [5-25], P=0.018 and 14 [4-21] vs. 5 [2-6], P=0.019, respectively). The 30-day mortality rate was 27.3% and the 90-day mortality rate was 72.7%. CONCLUSIONS: PMX-DHP treatment improved P/F ratio and reduced inflammatory markers in AE-ILD patients.

Anteroposterior Wnt-RA Gradient Defines Adhesion and Migration Properties of Neural Progenitors in Developing Spinal Cord.

Mammalian embryos exhibit a transition from head morphogenesis to trunk elongation to meet the demand of axial elongation. The caudal neural tube (NT) is formed with neural progenitors (NPCs) derived from neuromesodermal progenitors localized at the tail tip. However, the molecular and cellular basis of elongating NT morphogenesis is yet elusive. Here, we provide evidence that caudal NPCs exhibit strong adhesion affinity that is gradually decreased along the anteroposterior (AP) axis in mouse embryonic spinal cord and human cellular models. Strong cell-cell adhesion causes collective migration, allowing AP alignment of NPCs depending on their birthdate. We further validated that this axial adhesion gradient is associated with the extracellular matrix and is under the control of graded Wnt signaling emanating from tail buds and antagonistic retinoic acid (RA) signaling. These results suggest that progressive reduction of NPC adhesion along the AP axis is under the control of Wnt-RA molecular networks, which is essential for a proper elongation of the spinal cord.

Alterations in Structural and Functional Connectivities in Patients with End-Stage Renal Disease.

BACKGROUND AND PURPOSE: The aim of this study was to evaluate the structural and functional connectivities of brain network using graph theoretical analysis in neurologically asymptomatic patients with end-stage renal disease (ESRD). We further investigated the prevalence of cognitive impairment (CI) in ESRD patients and analyzed the association between network measures of brain connectivity and cognitive function. METHODS: We prospectively enrolled 40 neurologically asymptomatic ESRD patients, 40 healthy controls, and 20 disease controls. All of the subjects underwent diffusion-tensor imaging (DTI) and resting-state functional magnetic resonance imaging (rs-fMRI). We calculated measures of structural and functional connectivities based on DTI and rs-fMRI, respectively, and investigated differences therein between the ESRD patients and the healthy controls. We assessed cognitive function in the ESRD patients using the Korean version of the Consortium to Establish a Registry for Alzheimer's Disease neuropsychological battery. RESULTS: The ESRD patients exhibited decreased global structural and functional brain connectivities, as well as alterations of network hubs compared to the healthy controls and disease controls. About 70% of the ESRD patients had CI. Moreover, ESRD patients without CI exhibited decreased global connectivity and alterations of network hubs. Furthermore, there was a significant positive association between measures of brain connectivity and cognitive function. CONCLUSIONS: We found that ESRD patients exhibited decreased structural and functional brain connectivities, and that there was a significant association between brain connectivity and cognitive function. These alterations in the brain network may contribute to the pathophysiological mechanism of CI in ESRD patients.

Differences of connectivity between ESRD patients with PD and HD.

OBJECTIVES: The aim of this study was to investigate alterations in structural and functional brain connectivity between patients with end-stage renal disease (ESRD) who were undergoing peritoneal dialysis (PD) and hemodialysis (HD). METHODS: We enrolled 40 patients with ESRD who were undergoing PD (20 patients) and HD (20 patients). We also enrolled healthy participants as a control group. All of the subjects underwent diffusion tensor imaging (DTI) and resting-state functional magnetic resonance imaging (rs-fMRI). Using data from the structural and functional connectivity matrix based on DTI and rs-fMRI, we calculated several network measures using graph theoretical analysis. RESULTS: The measures of global structural connectivity were significantly different between the patients with ESRD who were undergoing PD and healthy subjects. The global efficiency and local efficiency in the patients with PD were significantly decreased compared with those in healthy participants. However, all of the measures of global structural connectivity in the patients with HD were not different from those in healthy participants. Conversely, in the global functional connectivity, the characteristic path length was significantly increased and the small-worldness index was decreased in patients with HD. However, the measures of the global functional connectivity in the patients with PD were not different from those in healthy subjects. CONCLUSION: This study revealed that alterations in structural and functional connectivity in patients who were undergoing PD and HD were different than those in healthy controls. These findings suggest that brain networks may be affected by different types of renal replacement therapy.

Genetic localization of the orevactaene/epipyrone biosynthetic gene cluster in Epicoccum nigrum.

Epipyrone (EPN)-A (syn. orevactaene) is a polyketide compound of 3-d-galactosyl-4-hydroxy-2-pyrone with a modified heptaene acyl moiety, produced from Epicoccum nigrum and was reported to have various biological activities. Genome analysis identified a hypothetical EPN biosynthetic gene cluster (BGC) composed of the four genes epnABCD, which encode a highly-reducing fungal polyketide synthase, a glycosyltransferase, a cytochrome P450, and a transporter. The individual gene inactivation of epnABC resulted in the total loss of EPN production, while the inactivation of a nearby transcription factor-encoding gene had no effect on the production of EPN, substantiating that epnABCD is the EPN BGC. mRNA expression indicated no epnA transcription in the epnB knockout mutant and the occurrence of the bicistronic transcription of epnAB. This study defined an EPN BGC, which is the first blueprint reported for glycosylated 2-pyrone polyketide biosynthesis.

Alterations of gray matter volumes and connectivity in patients with tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is an inherited genetic disorder caused by mutations in the TSC1 or TSC2 genes, encoding hamartin and tuberin. We aimed to evaluate structural volumes and connectivity of patients with TSC compared to those of healthy subjects. We consecutively enrolled 13 patients with a diagnosis of TSC and 15 age- and sex-matched healthy control subjects. Subjects underwent three-dimensional volumetric T1-weighted imaging, suitable for a quantitative analysis. Structural volumes were investigated using FreeSurfer image analysis software, and structural connectivity was calculated from a connectivity matrix, which was estimated from the correlation analysis of structural volumes using the Brain Analysis using Graph Theory software package. Differences in structural volumes and connectivity were analyzed between patients with TSC and healthy subjects. There were no differences of cortical volumes between the patients with TSC and healthy controls. However, we found decreased gray matter volumes in several subcortical regions in the patients with TSC compared to those in healthy controls, specifically in the putamen (0.3212 vs. 0.3841%, p = 0.001), even after multiple corrections. Regarding global structural connectivity, the small-worldness index was significantly decreased in patients with TSC compared to that in healthy controls (0.907 vs. 0.977, p = 0.049). This study revealed structural volumes and connectivity in patients with TSC that are significantly different from those in healthy controls. These alterations have implications for the pathogenesis of TSC.

Polyaromatic Resin HP-20 Induced Accumulation of Intermediate Azaphilones in Monascus purpureus ∆mppC and ∆mpp7 Strains.

Monascus purpureus recombinant mppC and mpp7 knockout strains were subjected to extractive fermentation in the context of azaphilone pigment production. Inclusion of Diaion HP-20 resin resulted in the selective production of unreduced azaphilone congeners, in addition to the early intermediate FK17-P2a, from ∆mppC and ∆mpp7 strains that would otherwise mainly produce reduced congeners. Structural determination of two novel unreduced azaphilones from the ∆mpp7 strain was accomplished. The unreduced azaphilone compound was converted into the cognate reduced congener in recombinant M. purpureus strains, demonstrating its intermediate role in azaphilone biosynthesis. This study demonstrates the possibility that extractive fermentation with Diaion HP-20 resin can be used to obtain cryptic azaphilone metabolites.

Alterations of brain network hubs in reflex syncope: Evidence from a graph theoretical analysis based on DTI.

OBJECTIVE: We evaluated global topology and organization of regional hubs in the brain networks and microstructural abnormalities in the white matter of patients with reflex syncope. METHODS: Twenty patients with reflex syncope and thirty healthy subjects were recruited, and they underwent diffusion tensor imaging (DTI) scans. Graph theory was applied to obtain network measures based on extracted DTI data, using DSI Studio. We then investigated differences in the network measures between the patients with reflex syncope and the healthy subjects. We also analyzed microstructural abnormalities of white matter using tract-based spatial statistics analysis (TBSS). RESULTS: Measures of global topology were not different between patients with reflex syncope and healthy subjects. However, in reflex syncope patients, the strength measures of the right angular, left inferior frontal, left middle orbitofrontal, left superior medial frontal, and left middle temporal gyrus were lower than in healthy subjects. The betweenness centrality measures of the left middle orbitofrontal, left fusiform, and left lingual gyrus in patients were lower than those in healthy subjects. The PageRank centrality measures of the right angular, left middle orbitofrontal, and left superior medial frontal gyrus in patients were lower than those in healthy subjects. Regarding the analysis of the white matter microstructure, there were no differences in the fractional anisotropy and mean diffusivity values between the two groups. CONCLUSIONS: We have identified a reorganization of network hubs in the brain network of patients with reflex syncope. These alterations in brain network may play a role in the pathophysiologic mechanism underlying reflex syncope.

Procalcitonin as a potential predicting factor for prognosis in bacterial meningitis.

We investigated the potential role of serum procalcitonin in differentiating bacterial meningitis from viral meningitis, and in predicting the prognosis in patients with bacterial meningitis. This was a retrospective study of 80 patients with bacterial meningitis (13 patients died). In addition, 58 patients with viral meningitis were included as the disease control groups for comparison. The serum procalcitonin level was measured in all patients at admission. Differences in demographic and laboratory data, including the procalcitonin level, were analyzed between the groups. We used the mortality rate during hospitalization as a marker of prognosis in patients with bacterial meningitis. Multiple logistic regression analysis showed that high serum levels of procalcitonin (>0.12ng/mL) were an independently significant variable for differentiating bacterial meningitis from viral meningitis. The risk of having bacterial meningitis with high serum levels of procalcitonin was at least 6 times higher than the risk of having viral meningitis (OR=6.76, 95% CI: 1.84-24.90, p=0.004). In addition, we found that high levels of procalcitonin (>7.26ng/mL) in the blood were an independently significant predictor for death in patients with bacterial meningitis. The risk of death in patients with bacterial meningitis with high serum levels of procalcitonin may be at least 9 times higher than those without death (OR=9.09, 95% CI: 1.74-47.12, p=0.016). We found that serum procalcitonin is a useful marker for differentiating bacterial meningitis from viral meningitis, and it is also a potential predicting factor for prognosis in patients with bacterial meningitis.