Ligustilide inhibits Purkinje cell ferritinophagy via the ULK1/NCOA4 pathway to attenuate valproic acid-induced autistic features.

BACKGROUND: Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder in which social impairment is the core symptom. Presently, there are no definitive medications to cure core symptoms of ASD, and most therapeutic strategies ameliorate ASD symptoms. Treatments with proven efficacy in autism are imminent. Ligustilide (LIG), an herbal monomer extracted from Angelica Sinensis and Chuanxiong, is mainly distributed in the cerebellum and widely used in treating neurological disorders. However, there are no studies on its effect on autistic-like phenotypes and its mechanism of action. PURPOSE: Investigate the efficacy and mechanism of LIG in treating ASD using two Valproic acid(VPA)-exposed and BTBR T + Itpr3tf/J (BTBR) mouse models of autism. METHODS: VPA-exposed mice and BTBR mice were given LIG for treatment, and its effect on autistic-like phenotype was detected by behavioral experiments, which included a three-chamber social test. Subsequently, RNA-Sequence(RNA-Seq) of the cerebellum was performed to observe the biological changes to search target pathways. The autophagy and ferroptosis pathways screened were verified by WB(Western Blot) assay, and the cerebellum was stained by immunofluorescence and examined by electron microscopy. To further explore the therapeutic mechanism, ULK1 agonist BL-918 was used to block the therapeutic effect of LIG to verify its target effect. RESULTS: Our work demonstrates that LIG administration from P12-P14 improved autism-related behaviors and motor dysfunction in VPA-exposed mice. Similarly, BTBR mice showed the same improvement. RNA-Seq data identified ULK1 as the target of LIG in regulating ferritinophagy in the cerebellum of VPA-exposed mice, as evidenced by activated autophagy, increased ferritin degradation, iron overload, and lipid peroxidation. We found that VPA exposure-induced ferritinophagy occurred in the Purkinje cells, with enhanced NCOA4 and Lc3B expressions. Notably, the therapeutic effect of LIG disappeared when ULK1 was activated. CONCLUSION: LIG treatment inhibits ferritinophagy in Purkinje cells via the ULK1/NCOA4-dependent pathway. Our study reveals for the first time that LIG treatment ameliorates autism symptoms in VPA-exposed mice by reducing aberrant Purkinje ferritinophagy. At the same time, our study complements the pathogenic mechanisms of autism and introduces new possibilities for its therapeutic options.

Effects of six teaching strategies on medical students: protocol for a systematic review and network meta-analysis.

INTRODUCTION: Mounting evidence has suggested that novel teaching strategies have a positive impact on the quality and efficiency of medical education. However, the comprehensive evidence about the superiority among various strategies is not clear. To address this issue, we aim to conduct a systematic review and network meta-analysis (NMA) to evaluate the effects of six main strategies on medical education, including case-based learning, problem-based learning, team-based learning, flipped classrooms, simulation-based education and bridge-in, objective, preassessment, participatory learning, postassessment and summary. METHODS AND ANALYSIS: A systematic search will be conducted in PubMed, Embase, Web of Science and the Cochrane Library, covering studies published from database inception to November 2023. Randomised controlled trials which evaluated the different teaching methods and meet the eligibility criteria will be included. The effectiveness of medical students' learning, which is evaluated by theoretical test score, experimental or practical test score, will be analysed as the primary outcomes. Besides, the secondary outcomes consist of learning satisfaction of students and formative evaluation score. The study selection and data extraction will be independently performed by two authors. The risk of bias in each study will be assessed using V.2 of the Cochrane risk-of-bias tool for randomised controlled trials. To compare the effects of six teaching strategies, pairwise meta-analysis and NMA will be performed using Rev Man, STATA and R software. Statistical analyses including homogeneity tests, sensitivity analysis, consistency tests, subgroup analysis, Egger's test and publication bias will also be completed. ETHICS AND DISSEMINATION: No formal research ethics approval is required because this study is a meta-analysis based on published studies. The results will be disseminated to a peer-reviewed journal for publication. PROTOCOL REGISTRATION NUMBER: CRD42023456050.

Cell-free DNA testing for early hepatocellular carcinoma surveillance.

BACKGROUND: Liver cirrhosis (LC) is the highest risk factor for hepatocellular carcinoma (HCC) development worldwide. The efficacy of the guideline-recommended surveillance methods for patients with LC remains unpromising. METHODS: A total of 4367 LCs not previously known to have HCC and 510 HCCs from 16 hospitals across 11 provinces of China were recruited in this multi-center, large-scale, cross-sectional study. Participants were divided into Stage Ⅰ cohort (510 HCCs and 2074 LCs) and Stage Ⅱ cohort (2293 LCs) according to their enrollment time and underwent Tri-phasic CT/enhanced MRI, US, AFP, and cell-free DNA (cfDNA). A screening model called PreCar Score was established based on five features of cfDNA using Stage Ⅰ cohort. Surveillance performance of PreCar Score alone or in combination with US/AFP was evaluated in Stage Ⅱ cohort. FINDINGS: PreCar Score showed a significantly higher sensitivity for the detection of early/very early HCC (Barcelona stage A/0) in contrast to US (sensitivity of 51.32% [95% CI: 39.66%-62.84%] at 95.53% [95% CI: 94.62%-96.38%] specificity for PreCar Score; sensitivity of 23.68% [95% CI: 14.99%-35.07%] at 99.37% [95% CI: 98.91%-99.64%] specificity for US) (P < 0.01, Fisher's exact test). PreCar Score plus US further achieved a higher sensitivity of 60.53% at 95.08% specificity for early/very early HCC screening. INTERPRETATION: Our study developed and validated a cfDNA-based screening tool (PreCar Score) for HCC in cohorts at high risk. The combination of PreCar Score and US can serve as a promising and practical strategy for routine HCC care. FUNDING: A full list of funding bodies that contributed to this study can be found in Acknowledgments section.

Puerarin attenuates valproate-induced features of ASD in male mice via regulating Slc7a11-dependent ferroptosis.

Autism spectrum disorder (ASD) is a complicated, neurodevelopmental disorder characterized by social deficits and stereotyped behaviors. Accumulating evidence suggests that ferroptosis is involved in the development of ASD, but the underlying mechanism remains elusive. Puerarin has an anti-ferroptosis function. Here, we found that the administration of puerarin from P12 to P15 ameliorated the autism-associated behaviors in the VPA-exposed male mouse model of autism by inhibiting ferroptosis in neural stem cells of the hippocampus. We highlight the role of ferroptosis in the hippocampus neurogenesis and confirm that puerarin treatment inhibited iron overload, lipid peroxidation accumulation, and mitochondrial dysfunction, as well as enhanced the expression of ferroptosis inhibitory proteins, including Nrf2, GPX4, Slc7a11, and FTH1 in the hippocampus of VPA mouse model of autism. In addition, we confirmed that inhibition of xCT/Slc7a11-mediated ferroptosis occurring in the hippocampus is closely related to puerarin-exerted therapeutic effects. In conclusion, our study suggests that puerarin targets core symptoms and hippocampal neurogenesis reduction through ferroptosis inhibition, which might be a potential drug for autism intervention.

Extracellular vesicles from organoid-derived human retinal progenitor cells prevent lipid overload-induced retinal pigment epithelium injury by regulating fatty acid metabolism.

Retinal degeneration (RD), a group of diseases leading to irreversible vision loss, is characterised by retinal pigment epithelium (RPE) or retinal neuron damage and loss. With fewer risks of immune rejection and tumorigenesis, stem cell-secreted extracellular vesicles (EVs) offer a new cell-free therapeutic paradigm for RD, which remains to be investigated. Human retinal organoid-derived retinal progenitor cells (hERO-RPCs) are an easily accessible and advanced cell source for RD treatment. However, hERO-RPCs-derived EVs require further characterisation. Here, we compared the characteristics of EVs from hERO-RPCs (hRPC-EVs) with those of human embryonic stem cell (hESC)-derived EVs (hESC-EVs) as controls. Based on in-depth proteomic analysis, we revealed remarkable differences between hRPC-EVs and hESC-EVs. A comparison between EVs and their respective cells of origin demonstrated that the protein loading of hRPC-EVs was more selective than that of hESC-EVs. In particular, hESC-EVs were enriched with proteins related to angiogenesis and cell cycle, whereas hRPC-EVs were enriched with proteins associated with immune modulation and retinal development. More importantly, compared with that of hESC-EVs, hRPC-EVs exhibited a lower correlation with cell proliferation and a unique capacity to regulate lipid metabolism. It was further confirmed that hRPC-EVs potentially eliminated lipid deposits, inhibited lipotoxicity and oxidative stress, and enhanced phagocytosis and survival of oleic acid-treated ARPE-19 cells. Mechanistically, hRPC-EVs are integrated into the mitochondrial network of oleic acid-treated ARPE-19 cells, and increased the level of mitochondrial fatty acid ß-oxidation-related proteins. Thus, organoid-derived hRPC-EVs represent a promising source of cell-free therapy for RD, especially for blinding diseases related to abnormal lipid metabolism in RPE cells.

The role of specific isoforms of CaV2 and the common C-terminal of CaV2 in calcium channel function in sensory neurons of Aplysia.

The presynaptic release apparatus can be specialized to enable specific synaptic functions. Habituation is the diminishing of a physiological response to a frequently repeated stimulus and in Aplysia, habituation to touch is mediated by a decrease in transmitter release from the sensory neurons that respond to touch even after modest rates of action potential firing. This synaptic depression is not common among Aplysia synaptic connections suggesting the presence of a release apparatus specialized for this depression. We found that specific splice forms of ApCaV2, the calcium channel required for transmitter release, are preferentially used in sensory neurons, consistent with a specialized release apparatus. However, we were not able to find a specific ApCaV2 splice uniquely required for synaptic depression. The C-terminus of ApCaV2 alpha1 subunit retains conserved binding to Aplysia rab-3 interacting molecule (ApRIM) and ApRIM-binding protein (ApRBP) and the C-terminus is required for full synaptic expression of ApCaV2. We also identified a splice form of ApRIM that did not interact with the ApCav2 alpha 1 subunit, but it was not preferentially used in sensory neurons.

Assessing the effectiveness of massive open online courses on improving clinical skills in medical education in China: A meta-analysis.

Massive Open Online Courses (MOOCs) are a new phenomenon in education worldwide. In China, MOOCs have been widely used in medical courses. However, the effects of MOOCs on improving clinical skills are controversial. Therefore, we conducted the study to verify whether the application of MOOCs in medical courses can improve participants' clinical skills in China. A systematic literature search was carried out using the PubMed, Embase, Web of Science, CNKI and Wanfang databases according to the predetermined criteria. The Hedges' g and its corresponding 95% confidence interval were selected to assess the effects of MOOCs on participants' clinical skills. Subgroup analyses, sensitivity analysis and publication bias test were performed in the study. A total of thirty-two records (thirty-two studies) with 3422 participants were identified in our study. There was a significant improvement in clinical skill scores of participants in the MOOC group compared with the control group. Subgroup analyses showed similar results in different student groups. Our study supported the notion that the MOOC-based teaching method appeared to be a more effective method than the conventional teaching technique for the improvement of participants' clinical skills in China.

A controllable perfusion microfluidic chip for facilitating the development of retinal ganglion cells in human retinal organoids.

Retinal organoids (ROs) derived from human pluripotent stem cells (hPSCs) have become a promising model in vitro to recapitulate human retinal development, which can be further employed to explore the mechanisms of retinal diseases. However, the current culture systems for ROs lack physiologically relevant microenvironments, such as controllable mechano-physiological cues and dynamic feedback between cells and the extracellular matrix (ECM), which limits the accurate control of RO development. Therefore, we designed a controllable perfusion microfluidic chip (CPMC) with the advantages of precisely controlling fluidic shear stress (FSS) and oxygen concentration distribution in a human embryonic stem cell (hESC)-derived RO culture system. We found that ROs cultured under this system allow for expanding the retinal progenitor cell (RPC) pool, orchestrating the retinal ganglion cell (RGC) specification, and axon growth without disturbing the spatial and temporal patterning events at the early stage of RO development. Furthermore, RNA sequencing data revealed that the activation of voltage-gated ion channels and the increased expression of ECM components synergistically improve the growth of ROs and facilitate the differentiation of RGCs. This study elaborates on the advantages of the designed CPMC to promote RO growth and provide a controllable and reliable platform for the efficient maturity of RGCs in the ROs, promising applications in modeling RGC-related disorders, drug screening, and cell transplantation.

Minocycline improves autism-related behaviors by modulating microglia polarization in a mouse model of autism.

Autism spectrum disorder (ASD) is a heterogeneous neurodevelopmental disorder with few pharmacological treatments. Minocycline, a tetracycline derivative that inhibits microglial activation, has been well-identified with anti-inflammatory properties and neuroprotective effects. A growing body of research suggests that ASD is associated with neuroinflammation, abnormal neurotransmitter levels, and neurogenesis. Thus, we hypothesized that minocycline could improve autism-related behaviors by inhibiting microglia activation and altering neuroinflammation. To verify our hypothesis, we used a mouse model of autism, BTBR T + Itpr3tf/J (BTBR). As expected, minocycline administration rescued the sociability and repetitive, stereotyped behaviors of BTBR mice while having no effect in C57BL/6J mice. We also found that minocycline improved neurogenesis and inhibited microglia activation in the hippocampus of BTBR mice. In addition, minocycline treatment inhibited Erk1/2 phosphorylation in the hippocampus of BTBR mice. Our findings show that minocycline administration alleviates ASD-like behaviors in BTBR mice and improves neurogenesis, suggesting that minocycline supplementation might be a potential strategy for improving ASD symptoms.

Early-life stress contributes to depression-like behaviors in a two-hit mouse model.

BACKGROUND: Depression is a common psychological disorder with pathogenesis involving genetic and environmental interactions. Early life stress can adversely affect physical and emotional development and dramatically increase the risk for the development of depression and anxiety disorders. METHODS: To examine potential early life stress driving risk for anxiety and depression, we used a two-hit developmental stress model，injecting poly(I: C) into neonatal mice on P2-P6 followed by peripubertal unpredictable stress in adolescence. RESULTS: Our study shows that early-life and adolescent stress leads to anxiety and depression-related behavioral phenotypes in male mice. Early-life stress exacerbated depression-like behavior in mice following peripubertal unpredictable stress. We confirmed that early life stress might be involved in the decreased neuronal activity in the medial prefrontal cortex (mPFC) and might be involved in shaping behavioral phenotypes of animals. We found that increased microglia and neuroinflammation in the mPFC of two-hit mice and early life stress further boost microglia activation and inflammatory factors in the mPFC region of mice following adolescent stress. LIMITATIONS: The specific neural circuits and mechanisms by which microglia regulate depression-like behaviors require further investigation. CONCLUSIONS: Our findings provide a novel insight into developmental risk factors and biological mechanisms in depression and anxiety disorders.

Novel, high accuracy models for hepatocellular carcinoma prediction based on longitudinal data and cell-free DNA signatures.

BACKGROUND & AIMS: Current hepatocellular carcinoma (HCC) risk scores do not reflect changes in HCC risk resulting from liver disease progression/regression over time. We aimed to develop and validate two novel prediction models using multivariate longitudinal data, with or without cell-free DNA (cfDNA) signatures. METHODS: A total of 13,728 patients from two nationwide multicenter prospective observational cohorts, the majority of whom had chronic hepatitis B, were enrolled. aMAP score, as one of the most promising HCC prediction models, was evaluated for each patient. Low-pass whole-genome sequencing was used to derive multi-modal cfDNA fragmentomics features. A longitudinal discriminant analysis algorithm was used to model longitudinal profiles of patient biomarkers and estimate the risk of HCC development. RESULTS: We developed and externally validated two novel HCC prediction models with a greater accuracy, termed aMAP-2 and aMAP-2 Plus scores. The aMAP-2 score, calculated with longitudinal data on the aMAP score and alpha-fetoprotein values during an up to 8-year follow-up, performed superbly in the training and external validation cohorts (AUC 0.83-0.84). The aMAP-2 score showed further improvement and accurately divided aMAP-defined high-risk patients into two groups with 5-year cumulative HCC incidences of 23.4% and 4.1%, respectively (p = 0.0065). The aMAP-2 Plus score, which incorporates cfDNA signatures (nucleosome, fragment and motif scores), optimized the prediction of HCC development, especially for patients with cirrhosis (AUC 0.85-0.89). Importantly, the stepwise approach (aMAP -> aMAP-2 -> aMAP-2 Plus) stratified patients with cirrhosis into two groups, comprising 90% and 10% of the cohort, with an annual HCC incidence of 0.8% and 12.5%, respectively (p <0.0001). CONCLUSIONS: aMAP-2 and aMAP-2 Plus scores are highly accurate in predicting HCC. The stepwise application of aMAP scores provides an improved enrichment strategy, identifying patients at a high risk of HCC, which could effectively guide individualized HCC surveillance. IMPACT AND IMPLICATIONS: In this multicenter nationwide cohort study, we developed and externally validated two novel hepatocellular carcinoma (HCC) risk prediction models (called aMAP-2 and aMAP-2 Plus scores), using longitudinal discriminant analysis algorithm and longitudinal data (i.e., aMAP and alpha-fetoprotein) with or without the addition of cell-free DNA signatures, based on 13,728 patients from 61 centers across mainland China. Our findings demonstrated that the performance of aMAP-2 and aMAP-2 Plus scores was markedly better than the original aMAP score, and any other existing HCC risk scores across all subsets, especially for patients with cirrhosis. More importantly, the stepwise application of aMAP scores (aMAP -> aMAP-2 -> aMAP-2 Plus) provides an improved enrichment strategy, identifying patients at high risk of HCC, which could effectively guide individualized HCC surveillance.

Aluminum hydroxide exposure induces neurodevelopmental impairment in hESC-derived cerebral organoids.

Aluminum (Al) has been classified as a cumulative environmental pollutant that endangers human health. There is increasing evidence to suggest the toxic effects of Al, but the specific action on human brain development remains unclear. Al hydroxide (Al(OH)3), the most common vaccine adjuvant, is the major source of Al and poses risks to the environment and early childhood neurodevelopment. In this study, we explored the neurotoxic effect of 5 µg/ml or 25 µg/ml Al(OH)3 for six days on neurogenesis by utilizing human cerebral organoids from human embryonic stem cells (hESCs). We found that early Al(OH)3 exposure in organoids caused a reduction in the size, deficits in basal neural progenitor cell (NPC) proliferation, and premature neuron differentiation in a time and dose-dependent manner. Transcriptomes analysis revealed a markedly altered Hippo-YAP1 signaling pathway in Al(OH)3 exposed cerebral organoid, uncovering a novel mechanism for Al(OH)3-induced detrimental to neurogenesis during human cortical development. We further identified that Al(OH)3 exposure at day 90 mainly decreased the production of outer radial glia-like cells(oRGs) but promoted NPC toward astrocyte differentiation. Taken together, we established a tractable experimental model to facilitate a better understanding of the impact and mechanism of Al(OH)3 exposure on human brain development.

Chronic exposure to (2 R,6 R)-hydroxynorketamine induces developmental neurotoxicity in hESC-derived cerebral organoids.

(R,S)-ketamine (ketamine) has been increasingly used recreationally and medicinally worldwide; however, it cannot be removed by conventional wastewater treatment plants. Both ketamine and its metabolite norketamine have been frequently detected to a significant degree in effluents, aquatic, and even atmospheric environments, which may pose risks to organisms and humans via drinking water and aerosols. Ketamine has been shown to affect the brain development of unborn babies, while it is still elusive whether (2 R,6 R)-hydroxynorketamine (HNK) induces similar neurotoxicity. Here, we investigated the neurotoxic effect of (2 R,6 R)-HNK exposure at the early stages of gestation by applying human cerebral organoids derived from human embryonic stem cells (hESCs). Short-term (2 R,6 R)-HNK exposure did not significantly affect the development of cerebral organoids, but chronic high-concentration (2 R,6 R)-HNK exposure at day 16 inhibited the expansion of organoids by suppressing the proliferation and augmentation of neural precursor cells (NPCs). Notably, the division mode of apical radial glia was unexpectedly switched from vertical to horizontal division planes following chronic (2 R,6 R)-HNK exposure in cerebral organoids. Chronic (2 R,6 R)-HNK exposure at day 44 mainly inhibited the differentiation but not the proliferation of NPCs. Overall, our findings indicate that (2 R,6 R)-HNK administration leads to the abnormal development of cortical organoids, which may be mediated by inhibiting HDAC2. Future clinical studies are needed to explore the neurotoxic effects of (2 R,6 R)-HNK on the early development of the human brain.

Icariin alleviates autistic-like behavior, hippocampal inflammation and vGlut1 expression in adult BTBR mice.

Autism spectrum disorder (ASD) is a complicated, heterogeneous disorder characterized by social interaction deficits and repetitive stereotypical behaviors. Neuroinflammation and synaptic protein dysregulation have been implicated in ASD pathogenesis. Icariin (ICA) has proven to exert neuroprotective function through anti-inflammatory function. Therefore, this study aimed to clarify the effects of ICA treatment on autism-like behavioral deficits in BTBR mice and whether these changes were related to modifications in the hippocampal inflammation and the balance of excitatory/inhibitory synapses. ICA supplementation (80 mg/kg, once daily for ten days, i.g.) ameliorated social deficits, repetitive stereotypical behaviors, and short-term memory deficit without affecting locomotor activity or anxiety-like behaviors of BTBR mice. Furthermore, ICA treatment inhibited neuroinflammation via decreasing microglia number and the soma size in the CA1 region of the hippocampus, as well as the protein levels of proinflammatory cytokines in the hippocampus of BTBR mice. In addition, ICA treatment also rescued excitatory-inhibitory synaptic protein imbalance by inhibiting the increased vGlut1 level without affecting the vGAT level in the BTBR mouse hippocampus. Collectively, the observed results indicate that ICA treatment alleviates ASD-like features, mitigates disturbed balance of excitatory-inhibitory synaptic protein, and inhibits hippocampal inflammation in BTBR mice, and may represent a novel promising drug for ASD treatment.

Novel antidepressant-like properties of the fullerenol in an LPS-induced depressive mouse model.

Depression is a common mental disease and is highly prevalent in populations. Dysregulated neuroinflammation and concomitant-activated microglia are involved in the pathogenesis of depression. Experimental evidence has indicated that fullerenol exerts anti-neuroinflammation and protective effects against neurological diseases. Here, we evaluated fullerenol's effects against lipopolysaccharide (LPS)-induced mouse depressive-like behaviors. Fullerenol treatment produced an antidepressant-like effect, as indicated by preventing the LPS-induced reduction in the sucrose preference and shortening the immobility durations in both the tail suspension test and the forced swim test. We found that fullerenol treatment mitigated LPS-induced hippocampal microglia activation and released proinflammatory cytokines. Meanwhile, fullerenol promoted hippocampus neurogenesis, evidenced by increased DCX-positive cells in LPS-treated mice. Hippocampal RNA-Seq analysis revealed proinflammatory cytokine and neurogenesis involved in fullerenol's antidepressant-like effects. Our data indicate that fullerenol exerts antidepressant effects, which might be due to beneficial functions in reducing neuroinflammatory processes and promoting neurogenesis in the hippocampus.

Mitochondrial-targeting fluorescent small molecule IR-780 alleviates radiation-induced brain injury.

Radiation-induced brain injury (RIBI) is a common complication of radiation therapy for brain tumors. Vascular damage is one of the key factors closely related to the severity of the RIBI. However, effective vascular target treatment strategies are lacking. Previously, we have identified a fluorescent small molecule dye, IR-780, which shows the properties of injury tissue targeting and provided protection against various injuries by modulating oxidative stress. This study aims to validate the therapeutic effect of IR-780 on RIBI. The effectiveness of IR-780 against RIBI has been comprehensively evaluated through techniques such as behavior, immunofluorescence staining, quantitative real-time polymerase chain reaction, Evans Blue leakage experiments, electron microscopy, and flow cytometry. Results show that IR-780 improves cognitive dysfunction, reduces neuroinflammation, restores the expression of tight junction proteins in the blood-brain barrier (BBB), and promotes the recovery of BBB function after whole brain irradiation. IR-780 also accumulates in injured cerebral microvascular endothelial cells, and its subcellular location is in the mitochondria. More importantly, IR-780 can reduce the levels of cellular reactive oxygen species and apoptosis. Moreover, IR-780 has no significant toxic side effects. IR-780 alleviates RIBI by protecting vascular endothelial cells from oxidative stress, reducing neuroinflammation, and restoring BBB function, suggesting IR-780 as a promising treatment candidate for RIBI therapy.

The development of a cSMART-based integrated model for hepatocellular carcinoma diagnosis.

BACKGROUND: Hepatocellular carcinoma (HCC) generally arises from a background of liver cirrhosis (LC). Patients with cirrhosis and suspected HCC are recommended to undergo serum biomarker tests and imaging diagnostic evaluation. However, the performance of routine diagnostic methods in detecting early HCC remains unpromising. METHODS: Here, we conducted a large-scale, multicenter study of 1675 participants including 490 healthy controls, 577 LC patients, and 608 HCC patients from nine clinical centers across nine provinces of China, profiled gene mutation signatures of cell-free DNA (cfDNA) using Circulating Single-Molecule Amplification and Resequencing Technology (cSMART) through detecting 931 mutation sites across 21 genes. RESULTS: An integrated diagnostic model called "Combined method" was developed by combining three mutation sites and three serum biomarkers. Combined method outperformed AFP in the diagnosis of HCC, especially early HCC, with sensitivities of 81.25% for all stages and 66.67% for early HCC, respectively. Importantly, the integrated model exhibited high accuracy in differentiating AFP-negative, AFP-L3-negative, and PIVKA-II-negative HCCs from LCs.

Di-(2-ethylhexyl) phthalate exposure impairs cortical development in hESC-derived cerebral organoids.

Di-(2-ethylhexyl) phthalate (DEHP), a ubiquitous environmental endocrine disruptor, is widely used in consumer products. Increasing evidence implies that DEHP influences the early development of the human brain. However, it lacks a suitable model to evaluate the neurotoxicity of DEHP. Using an established human cerebral organoid model, which reproduces the morphogenesis of the human cerebral cortex at the early stage, we demonstrated that DEHP exposure markedly suppressed cell proliferation and increased apoptosis, thus impairing the morphogenesis of the human cerebral cortex. It showed that DEHP exposure disrupted neurogenesis and neural progenitor migration, confirmed by scratch assay and cell migration assay in vitro. These effects might result from DEHP-induced dysplasia of the radial glia cells (RGs), the fibers of which provide the scaffolds for cell migration. RNA sequencing (RNA-seq) analysis of human cerebral organoids showed that DEHP-induced disorder in cell-extracellular matrix (ECM) interactions might play a pivotal role in the neurogenesis of human cerebral organoids. The present study provides direct evidence of the neurodevelopmental toxicity of DEHP after prenatal exposure.

Implication of Hippocampal Neurogenesis in Autism Spectrum Disorder: Pathogenesis and Therapeutic Implications.

Autism spectrum disorder (ASD) is a cluster of heterogeneous neurodevelopmental conditions with atypical social communication and repetitive sensory-motor behaviors. The formation of new neurons from neural precursors in the hippocampus has been unequivocally demonstrated in the dentate gyrus of rodents and non-human primates. Accumulating evidence sheds light on how the deficits in the hippocampal neurogenesis may underlie some of the abnormal behavioral phenotypes in ASD. In this review, we describe the current evidence concerning pre-clinical and clinical studies supporting the significant role of hippocampal neurogenesis in ASD pathogenesis, discuss the possibility of improving hippocampal neurogenesis as a new strategy for treating ASD, and highlight the prospect of emerging pro-neurogenic therapies for ASD.

Valproic acid exposure decreases neurogenic potential of outer radial glia in human brain organoids.

Valproic acid (VPA) exposure during pregnancy leads to a higher risk of autism spectrum disorder (ASD) susceptibility in offspring. Human dorsal forebrain organoids were used to recapitulate course of cortical neurogenesis in the developing human brain. Combining morphological characterization with massive parallel RNA sequencing (RNA-seq) on organoids to analyze the pathogenic effects caused by VPA exposure and critical signaling pathway. We found that VPA exposure in organoids caused a reduction in the size and impairment in the proliferation and expansion of neural progenitor cells (NPCs) in a dose-dependent manner. VPA exposure typically decreased the production of outer radial glia-like cells (oRGs), a subtype of NPCs contributing to mammalian neocortical expansion and delayed their fate toward upper-layer neurons. Transcriptomics analysis revealed that VPA exposure influenced ASD risk gene expression in organoids, which markedly overlapped with irregulated genes in brains or organoids originating from ASD patients. We also identified that VPA-mediated Wnt/ß-catenin signaling pathway activation is essential for sustaining cortical neurogenesis and oRGs output. Taken together, our study establishes the use of dorsal forebrain organoids as an effective platform for modeling VPA-induced teratogenic pathways involved in the cortical neurogenesis and oRGs output, which might contribute to ASD pathogenesis in the developing brain.