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The mitochondrial GTPase mitofusin-2 (MFN2) gene can suppress the cell cycle and regulate cell proliferation in a number of cell types. However, its function in hepatic fibrosis remains largely unexplored. We attempted to understand the mechanism of MFN2 in hepatic stellate cell (HSC) proliferation and the development of hepatic fibrosis. Rat HSC-T6 HSC were cultured and transfected by adenovirus- (Ad-) Mfn2 or its negative control (NC) vector (Ad-green fluorescent protein (GFP)); a rat liver cirrhosis model was established via subcutaneous injection with carbon tetrachloride (CCl4). Seventy-two rats were randomly divided into four groups: CCl4, Mfn2, GFP, and NC. Ad-Mfn2 or Ad-GFP was transfected into the circulation via intravenous injection at day 1, 14, 28, 42, or 56 after the first injection of CCl4 in the Mfn2/GFP groups. Biomarkers related to HSC proliferation and the development of hepatic fibrosis were detected using western blotting, hematoxylin-eosin and Masson staining, and immunohistochemistry. In vitro, Mfn2 interfered specifically with platelet-derived growth factor- (PDGF-) induced signaling pathway (phosphatidylinositol 3-kinase- (PI3K-) AKT), inhibiting HSC-T6 cell activation and proliferation. During the process of hepatic fibrosis in vivo, extracellular collagen deposition and the expression of fibrosis-related proteins increased progressively, while Mfn2 expression decreased gradually. Upregulating Mfn2 expression at the early stage of fibrosis impeded the process, triggered the downregulation of type I collagen, and antagonized the formation of factors associated with liver fibrosis. Mfn2 suppresses HSC proliferation and activation and exhibits antifibrotic potential in early-stage hepatic fibrosis. Therefore, it may represent a significant therapeutic target for eradicating hepatic fibrosis.

Given the threats to the regional river ecological flow, including the sharp increase in per capita water demand, the increase in the number of hydraulic engineering, and the strong seasonal runoff of rainfall recharge channels, we clarified the appropriate ecological flow thresholds for the middle reaches of Bailong River of northwest China. Based on the monthly runoff data in the Wudu Hydrological Station of Bailong River from 1990 to 2020, seven suitable hydrological methods were used to couple the estimation of the ecological flow in the middle reaches of Bailong River, which were tested by runoff satisfaction level analysis, using quadratic fit and 95% confidence interval to determine the monthly ecological flow and threshold of the river in the study area. The results by using the single hydrological methods showed that the calculation results from the monthly minimum ecological runoff calculation method, the improved RVA method, the Tennant method, and the DC method were four better estimation methods, which had a satisfaction level of more than 90% in the whole year. The improved RVA method and the Tennant method were more suitable for the diversion power station whose main task aimed at power generation. Although the economic benefits of hydropower stations were better when the improved RVA method was used to calculate ecological flow, the calculation results from the improved RVA method were tested and analyzed by the Tennant method, which were extremely poor from November to April in the following year, and could not meet the minimum ecological water requirements for the biological community in the river channel. While the other five methods all meet this requirement. We suggest that the minimum ecological flow in the middle reaches of the Bailong River maintained at 27.28 m3⁄s, and the maximum retained at 116.33 m3⁄s. The calculation results were in line with the dynamics of runoff in the study area, which could improve the accuracy of water ecological protection. The calculation results of this study could be used for the middle reaches of the Bailong River and may provide a reference for follow-up ecological restoration research and management in similar ecological zones.

In a fully mechanized top coal caving mining process, the mechanical properties of the top coal deteriorate constantly, and the stability of the support is determined by the boundary position of the top coal entering the limit equilibrium state. Top coal flaking occurs more easily along the advancing direction owing to an increase in the distance of the boundary from the wall. Moreover, the strength of the coal seam is an important factor that determines the boundary position. The equation of the top coal limit equilibrium zone boundary, used in fully mechanized top coal caving mining, was determined based on the limit equilibrium theory, continuum damage mechanics, and the Hoek–Brown rock empirical strength criterion. Using the RFPA3D and UDEC numerical analysis methods, the effect of top coal strength on the mechanical behavior during the mining process was determined; and the space–time relationship between the top coal deformation failure, damage evolution, and boundary of the limit equilibrium zone was established. The results revealed that when the coal mass entered the limit equilibrium zone, accelerated crack development, accelerated damage, and axial unloading occurred. Moreover, an increase in the coal seam strength caused the boundary of the top coal limit equilibrium zone to be closer to the wall. By comparing the numerical results with the theoretical results, the overall trend was found to be consistent, allowing the verification of the rationality and reliability of the determination equation of the boundary position of the limit equilibrium zone.

ObjectivesGastric cancer with liver metastasis (GCLM) is highly aggressive and has a poor prognosis. This study aims to evaluate the survival benefit of primary tumor resection (PTR) for gastric cancer with liver metastasis.MethodsData on patients with GCLM was extracted from the Surveillance, Epidemiology, and End Results (SEER) database from 2010 to 2015. A 1:1 propensity score matching (PSM) analysis was performed to minimize the heterogeneity between the PTR and no-PTR groups. The Kaplan–Meier method and Cox regression analysis were used to assess the impact of primary tumor resection (PTR) on overall survival (OS) and cause-specific survival (CSS).ResultsA total of 3,001 patients with GCLM were included, with 328 patients treated with primary tumor resection (PTR), whereas the other 2,673 patients were not. Patients with PTR had a significantly higher OS and CSS rate than those without PTR in unmatched and PSM cohorts. In an unmatched cohort, the median OS was 12.0 months (95% CI, 10 months to 14 months) for those who underwent PTR and 4 months (95% CI, 4 months to 5 months) for those without PTR; the median CSS for those who underwent PTR was 12.0 months (95% CI, 10 months to14 months) and 4 months (95% CI, 4 months to 5 months) for those without PTR, respectively. After PMS, the median OS was 12.0 months (95% CI, 10 months to 17 months) for those who underwent PTR and 7 months (95% CI, 5 months to 10 months) for those without PTR, respectively; the median CSS for those who underwent PTR was 12.0 months (95% CI, 11 months to 17 months) and 7 months (95% CI, 5 months to 8 months) for those without PTR, respectively. In addition, multivariate Cox analysis in the PSM cohort showed that PTR, age, degree of tumor differentiation, and chemotherapy were independent prognostic factors for OS and CSS in GCLM. Specifically, PTR was a significant protective factor for OS (HR: 0.427; 95% CI, 0.325 to 0.561, P ConclusionPrimary tumor resection improves the survival of gastric cancer patients with liver metastasis.

To establish a dynamic nomogram based on preoperative clinical data for prediction of lateral lymph node metastasis (LLNM) of papillary thyroid carcinoma.Retrospective study.The Sixth Affiliated Hospital of Sun Yat-Sen University.The data of 477 patients from 2 centers formed the training group and validation group and were retrospectively reviewed. Preoperative clinical factors influencing LLNM were identified by univariable and multivariable analysis and were to construct a predictive dynamic nomogram for LLNM. Receiver operating characteristic analysis and calibration curves were used to evaluate the predictive power of the nomogram.The following were identified as independent risk factors for LLNM: male sex (odds ratio [OR] = 4.6,The dynamic nomogram for preoperative prediction of LLNM in papillary thyroid carcinoma can help surgeons identify high-risk patients and develop individualized treatment plans.

PolarLight is a compact soft X-ray polarimeter onboard a CubeSat, which was launched into a low-Earth orbit on October 29, 2018. In March 2019, PolarLight started full operation, and since then, regular observations with the Crab nebula, Sco X-1, and background regions have been conducted. Here we report the operation, calibration, and performance of PolarLight in the orbit. Based on these, we discuss how one can run a low-cost, shared CubeSat for space astronomy, and how CubeSats can play a role in modern space astronomy for technical demonstration, science observations, and student training., Comment: accepted for publication in Advances in Space Research

Colorectal cancer (CRC) is the third and second most common type of cancer diagnosed in males and females, respectively, and is the fourth leading cause of cancer-associated mortality worldwide. Liver metastasis is the primary cause of mortality in patients with CRC, and therefore requires therapeutic focus. Regulatory T cells (Tregs) and hepatic stellate cells (HSCs) are potentially involved in regulating the immune response during liver metastasis. The aim of the present study was to evaluate the influence of CD4+ forkhead box p3 (Foxp3)+ Tregs and the HGF/c-Met signaling pathway in the liver metastasis of CRC. A model of the latter was established using Balb/c mice via splenic injection of human CRC cells (CT-26 line). The mice were monitored for 3 weeks after being injected, and the spleens and livers were removed on day 22 for further analysis. Moreover, the single-cell suspensions were labeled with CD4 and Foxp3 antibodies, and were analyzed using flow cytometry. Expression levels of α-smooth muscle actin (SMA), hepatocyte growth factor (HGF) and hepatocyte growth factor receptor (c-Met) were analyzed using immunohistochemistry. Mice injected with CT-26 cells exhibited signs of illness and significant weight loss, compared with the control mice (P=0.013), and they also developed liver metastases, at an average of 20.5 tumors per mouse. Pathological evaluation using hematoxylin and eosin staining confirmed the tumors as liver metastases of CRC. The numbers of CD4+ T cells were significantly decreased in the spleen (P

This paper selects the annual panel data of China and its provinces from 1997 to 2015 and explores the impacts of financial development on CO2 emissions based on the extended STIRPAT model. In order to analyze the impacts of financial development on CO2 emissions under different levels of financial development, full-sample panels and sub-panels were estimated. The estimation results show that financial development efficiency and stock trading volume have positive impacts on CO2 emissions, while financial development scale and the market value of listed companies have negative impacts on CO2 emissions. The elasticity coefficients of the financial development variables are different for different sub-panels with different financial development. In addition, we do robustness tests by estimating the panel data model of four different sub-panels, which estimation results are consistent with the previous regression results. Finally, we suggest China's policy makers the following policy implications: China's government should formulate and improve financial credit policies, which can encourage financial institutions to carry out green credit business. Laws and regulations of stock markets should match the environmental protection and carbon emission reduction. China's government should formulate and implement fiscal and taxation policies to encourage enterprises to prevent high energy consumption and carbon emission, especially non-stated owned enterprises and small size enterprises. Moreover, China should gradually develop the domestic carbon finance market and carbon trading market, which can promote carbon emission reduction and doesn't require additional subsidy from the government.

The Gamma-Ray Integrated Detectors (GRID) is a space mission concept dedicated to monitoring the transient gamma-ray sky in the energy range from 10 keV to 2 MeV using scintillation detectors onboard CubeSats in low Earth orbits. The primary targets of GRID are the gamma-ray bursts (GRBs) in the local universe. The scientific goal of GRID is, in synergy with ground-based gravitational wave (GW) detectors such as LIGO and VIRGO, to accumulate a sample of GRBs associated with the merger of two compact stars and study jets and related physics of those objects. It also involves observing and studying other gamma-ray transients such as long GRBs, soft gamma-ray repeaters, terrestrial gamma-ray flashes, and solar flares. With multiple CubeSats in various orbits, GRID is unaffected by the Earth occultation and serves as a full-time and all-sky monitor. Assuming a horizon of 200 Mpc for ground-based GW detectors, we expect to see a few associated GW-GRB events per year. With about 10 CubeSats in operation, GRID is capable of localizing a faint GRB like 170817A with a 90% error radius of about 10 degrees, through triangulation and flux modulation. GRID is proposed and developed by students, with considerable contribution from undergraduate students, and will remain operated as a student project in the future. The current GRID collaboration involves more than 20 institutes and keeps growing. On August 29th, the first GRID detector onboard a CubeSat was launched into a Sun-synchronous orbit and is currently under test., accepted for publication in Experimental Astronomy

Multi-drug resistance (MDR) in breast cancer poses a great threat to chemotherapy. The expression and function of the ATP binding cassette (ABC) transporter are the major cause of MDR. Herein, a linear polyethylene glycol (PEI) conjugated with dicyandiamide, which called polymeric metformin (PolyMet), was successfully synthesized as a simple and biocompatible polymer of metformin. PolyMet showed the potential to reverse MDR by inhibiting the efflux of the substrate of ATP-binding cassette (ABC) transporter from DOX resistant MCF-7 cells (MCF-7/DOX). To test its MDR reversing effect, PolyMet was combined with DOX to treat mice carrying MCF-7/DOX xenografts. In order to decrease the toxicities of DOX and delivery PolyMet and DOX to tumor at the same time, PolyMet was complexed with poly-γ-glutamic acid-doxorubicin (PGA-DOX) electrostatically at the optimal ratio of 2:3, which were further coated with lipid membrane to form lipid/PolyMet-(PGA-DOX) nanoparticles (LPPD). The particle size of LPPD was 165.8 nm, and the zeta potential was +36.5 mV. LPPD exhibited favorable cytotoxicity and cellular uptake in MCF-7/DOX. Meanwhile, the bioluminescence imaging and immunohistochemical analysis indicated that LPPD effectively conquered DOX-associated MDR by blocking ABC transporters (ABCB1 and ABCC1) via PolyMet. Remarkably, LPPD significantly inhibited the tumor growth and lowered the systemic toxicity in a murine MCF-7/DOX tumor model. This is the first time to reveal that PolyMet can enhance the anti-tumor efficacy of DOX by dampening ABC transporters and activating the AMPK/mTOR pathway, which is a promising strategy for drug-resistant breast cancer therapy.

To design potentially more effective therapies, we need to further understand the mechanisms underlying epilepsy. Here, we uncover the role of Rev-erbα in circadian regulation of epileptic seizures. We first show up-regulation of REV-ERBα/Rev-erbα in brain tissues from patients with epilepsy and a mouse model. Ablation or pharmacological modulation of Rev-erbα in mice decreases the susceptibility to acute and chronic seizures, and abolishes diurnal rhythmicity in seizure severity, whereas activation of Rev-erbα increases the animal susceptibility. Rev-erbα ablation or antagonism also leads to prolonged spontaneous inhibitory postsynaptic currents and elevated frequency in the mouse hippocampus, indicating enhanced GABAergic signaling. We also identify the transporters Slc6a1 and Slc6a11 as regulators of Rev-erbα-mediated clearance of GABA. Mechanistically, Rev-erbα promotes the expressions of Slc6a1 and Slc6a11 through transcriptional repression of E4bp4. Our findings propose Rev-erbα as a regulator of synaptic function at the crosstalk between pathways regulating the circadian clock and epilepsy., Rev-erbα is a known regulator of the circadian clock. Here, the authors show that Rev-erbα is also a regulator of synaptic dysfunction in preclinical models of epilepsy.

We report follow-up observations of the Crab nebula with the PolarLight X-ray polarimeter, which revealed a possible variation in polarization associated with a pulsar glitch in 2019. The new observations confirm that the polarization has recovered roughly 100 days after the glitch. With the new observations, we find that the polarization angle (PA) measured with PolarLight from the total nebular emission has a difference of 18.0 +- 4.6 (deg) from that measured 42 years ago with OSO-8, indicating a secular evolution of polarization with either the Crab nebula or pulsar. The long-term variation in PA could be a result of multiple glitches in the history, magnetic reconnection or movement of synchrotron emitting structures in the nebula, or secular evolution of the pulsar magnetic geometry., Comment: ApJ Letters to appear

Temporal lobe epilepsy (TLE) is a common drug-resistant epilepsy associated with abundant cell death in the hippocampus. Here, we develop a novel gene therapy-mediated cell therapy that regenerates GABAergic neurons using internal hippocampal astrocytes to suppress seizure activity in a rat TLE model. We discovered that TLE-induced reactive astrocytes in the hippocampal CA1 region can be efficiently converted into GABAergic neurons after overexpressing a neural transcription factor NeuroD1. The astrocyte-converted neurons showed typical markers of GABAergic interneurons, fired action potentials, and formed functional synaptic connections with other neurons. Following NeuroD1-mediated astrocyte-to-neuron conversion, the number of hippocampal interneurons was significantly increased, and the spontaneous recurrent seizure (SRS) activity was significantly decreased. Moreover, NeuroD1 gene therapy treatment rescued total neuronal loss in the CA1 region and ameliorated the cognitive and mood dysfunctions in the TLE rat model. These results suggest that regeneration of GABAergic interneurons through gene therapy approach may provide a novel therapeutic intervention to treat drug-resistant TLE.

We report the detection of X-ray polarization in the neutron star low mass X-ray binary Scorpius (Sco) X-1 with PolarLight. The result is energy dependent, with a non-detection in 3-4 keV but a 4$\sigma$ detection in 4-8 keV; it is also flux dependent in the 4-8 keV band, with a non-detection when the source displays low fluxes but a 5$\sigma$ detection during high fluxes, in which case we obtain a polarization fraction of $0.043 \pm 0.008$ and a polarization angle of $52.6^\circ \pm 5.4^\circ$. This confirms a previous marginal detection with OSO-8 in the 1970s, and marks Sco X-1 the second astrophysical source with a significant polarization measurement in the keV band. The measured polarization angle is in line with the jet orientation of the source on the sky plane ($54^\circ$), which is supposedly the symmetric axis of the system. Combining previous spectral analysis, our measurements suggest that an optically thin corona is located in the transition layer under the highest accretion rates, and disfavor the extended accretion disk corona model., Comment: Submitted to AAS journals. We are grateful to some experts for useful comments

Lentinan is widely used as a therapeutic agent for treatment of malignant tumors in clinical practice. The chemical structure of lentinan is highly associated with its biological activity. In this study, the correlation between the structure of lentinan and its immune activity was investigated to assess the function of key parameters that can influence quality control of lentinan. The results showed that the batch-to-batch consistency of two lentinan samples was satisfactory, indicating the stability of production process of lentinan. However, although the chemical composition and triple-helical conformation (THC) of the tested samples were relatively similar, their Mw, polydispersity index (PDI), and Rgz remarkably varied due to different production processes. In vitro immunomodulatory assay reflects that lentinan could stimulate the macrophages phagocytic capacity. Meanwhile, lentinan samples could improve the spleen and thymus indices, promote the proliferation of lymphocytes and adjust for the percentages of CD4+ and CD8+ T cells in vivo. Furthermore, the immunomodulatory effect of lentinan sample B (Mw: 650,700 g/mol) was superior than that of the sample A (Mw: 4,818,700 g/mol). It was noted that the Mw should be detected as a necessary index for quality control of lentinan to ensure stability and effectiveness of the production process.

Metabolomics has been a potential tool for strain improvement through analyzing metabolite changes in the context of different conditions. However, the availability of a universal metabolite profiling analysis is still a big challenge. In this study, we presented an optimized liquid chromatography-tandem mass spectrometry-based metabolomics methodology for Corynebacterium glutamicum, an important industrial workhorse. It was found that quenching the cellular metabolism with 5-fold volume of - 20 °C 40% methanol was highly recommended due to its lower cell damage rate and higher intracellular metabolite recovery rate. For extracting intracellular metabolites, ethanol/water (3:1, v/v) at 100 °C combined with acidic acetonitrile/water (1:1, v/v, with 0.1% formic acid) at - 20 °C achieved the unbiased metabolite profiling of C. glutamicum. The established methodology was then applied to investigate the intracellular metabolite differences between C. glutamicum ATCC 13032 and an mscCG-deleted mutant under biotin limitation condition. It was observed that in the presence of the functional L-glutamate exporter MscCG, biotin limitation led to accumulation of intracellular 2-oxoglutarate but not L-glutamate. Deletion of mscCG severely inhibited L-glutamate excretion and resulted in a dramatical increase of intracellular L-glutamate, which in turn affected the metabolite profile. The optimized metabolomics methodology holds promise for promoting studies on metabolic mechanism of C. glutamicum.

Additive color mixing modulated by digital pulses enables universal applications in lights and displays. Conventional methods map binarized signals to ordinary red–green–blue color spaces, loosely connected with the color perception of human eyes, causing the complexity of gamut mapping and inaccuracy of chromatic manipulations. Here we developed a complete theory that encodes and decodes digital signals directly to a perceptually nonuniform color space of the commission internationale de l’éclairage, featuring a strict bijection between the duty cycles of each binary pulse and color components of the mixed light and an analytic nonlinear gamut volume. Exemplary applications in biophotonic lighting, active full-color displaying, and metaphotonic sensing confirmed our theory. The method applies to the fields with accurate manipulations of chromaticity and luminosity of a light, opening an avenue toward the next generation of perceptual displays, cameras, and sensors.

AMAP1 was a GTPase-activating protein that regulates cytoskeletal structures in focal adhesions, circular dorsal ruffles, and promote cell differentiation in tumor cells. But the activation and function of AMAP1 in breast cancer remain largely unexplored. Here we show that AMAP1 was phosphorylated and translocated to plasma membrane and formed a stable complex with Pyk2 in response to CCL18. Moreover, CCL18-dependent AMAP1 translocation interfered the AMAP1-IKK-β interaction, resulting in nuclear factor-kappaB (NF-κB) activation. Depletion of AMAP1 expression by RNAi efficiently reversed the CCL18-induced epithelial to mesenchymal transition (EMT) of breast cancer cells and as well as CCL18-induced adhesion, migration and invasion. Strikingly, AMAP1 overexpression was found in breast cancers that had undergone metastasis and was strongly predictive of poor prognosis in breast cancers. Given that AMAP1 mediated CCL18-induce activation of NF-κB and promoted breast cancer metastasis, AMAP1 may represent a therapeutic target for the eradication of breast cancer metastasis.

Colorectal cancer (CRC) is the third and second most common type of cancer diagnosed in males and females, respectively, and is the fourth leading cause of cancer-associated mortality worldwide. Liver metastasis is the primary cause of mortality in patients with CRC, and therefore requires therapeutic focus. Regulatory T cells (Tregs) and hepatic stellate cells (HSCs) are potentially involved in regulating the immune response during liver metastasis. The aim of the present study was to evaluate the influence of CD4+ forkhead box p3 (Foxp3)+ Tregs and the HGF/c-Met signaling pathway in the liver metastasis of CRC. A model of the latter was established using Balb/c mice via splenic injection of human CRC cells (CT-26 line). The mice were monitored for 3 weeks after being injected, and the spleens and livers were removed on day 22 for further analysis. Moreover, the single-cell suspensions were labeled with CD4 and Foxp3 antibodies, and were analyzed using flow cytometry. Expression levels of α-smooth muscle actin (SMA), hepatocyte growth factor (HGF) and hepatocyte growth factor receptor (c-Met) were analyzed using immunohistochemistry. Mice injected with CT-26 cells exhibited signs of illness and significant weight loss, compared with the control mice (P=0.013), and they also developed liver metastases, at an average of 20.5 tumors per mouse. Pathological evaluation using hematoxylin and eosin staining confirmed the tumors as liver metastases of CRC. The numbers of CD4+ T cells were significantly decreased in the spleen (P

SHANK3 mutations, including de novo deletions, have been associated with autism spectrum disorders (ASD). However, the effects of SHANK3 loss of function on neurodevelopment remain poorly understood. Here we generated human induced pluripotent stem cells (iPSC) in vitro, followed by neuro-differentiation and lentivirus-mediated shRNA expression to evaluate how SHANK3 knockdown affects the in vitro neurodevelopmental process at multiple time points (up to 4 weeks). We found that SHANK3 knockdown impaired both early stage of neuronal development and mature neuronal function, as demonstrated by a reduction in neuronal soma size, growth cone area, neurite length and branch numbers. Notably, electrophysiology analyses showed defects in excitatory and inhibitory synaptic transmission. Furthermore, transcriptome analyses revealed that multiple biological pathways related to neuron projection, motility and regulation of neurogenesis were disrupted in cells with SHANK3 knockdown. In conclusion, utilizing a human iPSC-based neural induction model, this study presented combined morphological, electrophysiological and transcription evidence that support that SHANK3 as an intrinsic, cell autonomous factor that controls cellular function development in human neurons.

Autistic spectral disorder (ASD) is a prevalent neurodevelopmental disease that affects multiple brain regions. Both clinical and animal studies have revealed the possible involvement of the cerebellum in ASD pathology. In this study, we generated a rodent ASD model through a single prenatal administration of valproic acid (VPA) into pregnant mice, followed by cerebellar morphological and functional studies of the offspring. Behavioral studies showed that VPA exposure led to retardation of critical motor reflexes in juveniles and impaired learning in a tone-conditioned complex motor task in adults. These behavioral phenotypes were associated with premature migration and excess apoptosis of the granular cell (GC) precursor in the cerebellar cortex during the early postnatal period, and the decreased cell density and impaired dendritic arborization of the Purkinje neurons. On acute cerebellar slices, suppressed synaptic transmission of the Purkinje cells were reported in the VPA-treated mice. In summary, converging evidence from anatomical, electrophysiological and behavioral abnormalities in the VPA-treated mice suggest cerebellar pathology in ASD and indicate the potential values of motor dysfunction in the early diagnosis of ASD.

SHANK3 mutations, including de novo deletions, have been associated with autism spectrum disorders (ASD). However, the effects of SHANK3 loss of function on neurodevelopment remain poorly understood. Here we generated human induced pluripotent stem cells (iPSC) in vitro, followed by neuro-differentiation and lentivirus-mediated shRNA expression to evaluate how SHANK3 knockdown affects the in vitro neurodevelopmental process at multiple time points (up to 4 weeks). We found that SHANK3 knockdown impaired both early stage of neuronal development and mature neuronal function, as demonstrated by a reduction in neuronal soma size, growth cone area, neurite length and branch numbers. Notably, electrophysiology analyses showed defects in excitatory and inhibitory synaptic transmission. Furthermore, transcriptome analyses revealed that multiple biological pathways related to neuron projection, motility and regulation of neurogenesis were disrupted in cells with SHANK3 knockdown. In conclusion, utilizing a human iPSC-based neural induction model, this study presented combined morphological, electrophysiological and transcription evidence that support that SHANK3 as an intrinsic, cell autonomous factor that controls cellular function development in human neurons.

Carbon nanotubes (CNTs) have shown potential applications in neuroscience as growth substrates owing to their numerous unique properties. However, a key concern in the fabrication of homogeneous composites is the serious aggregation of CNTs during incorporation into the biomaterial matrix. Moreover, the regulation mechanism of CNT-based substrates on neural differentiation remains unclear. Here, a novel strategy was introduced for the construction of CNT nanocomposites via layer-by-layer assembly of negatively charged multi-walled CNTs and positively charged poly(dimethyldiallylammonium chloride). Results demonstrated that the CNT-multilayered nanocomposites provided a potent regulatory signal over neural stem cells (NSCs), including cell adhesion, viability, differentiation, neurite outgrowth, and electrophysiological maturation of NSC-derived neurons. Importantly, the dynamic molecular mechanisms in the NSC differentiation involved the integrin-mediated interactions between NSCs and CNT multilayers, thereby activating focal adhesion kinase, subsequently triggering downstream signaling events to regulate neuronal differentiation and synapse formation. This study provided insights for future applications of CNT-multilayered nanomaterials in neural fields as potent modulators of stem cell behavior.

Activity of the dentate gyrus, which gates information flow to the hippocampus, is under tight inhibitory regulation by interneurons with distinctive axonal projections, intrinsic and synaptic characteristics and neurochemical identities. Total molecular layer cells (TML-Cs), a class of morphologically distinct GABAergic neurons with axonal projections across the molecular layer, are among the most frequent interneuronal type in the dentate subgranular region. However, little is known about their synaptic and neurochemical properties. We demonstrate that synapses from morphologically identified TML-Cs to dentate interneurons are characterized by low release probability, facilitating short-term dynamics and asynchronous release. TML-Cs consistently show somatic and axonal labeling for the cannabinoid receptor type 1 (CB1R) yet fail to express cholecystokinin (CCK) indicating their distinctive neurochemical identity. In paired recordings, the release probability at synapses between TML-Cs was increased by the CB1R antagonist AM251, demonstrating baseline endocannabinoid regulation of TML-C synapses. Apart from defining the synaptic and neurochemical features of TML-Cs, our findings reveal the morphological identity of a class of dentate CB1R-positive neurons that do not express CCK. Our findings indicate that TML-Cs can mediate cannabinoid sensitive feed-forward and feedback inhibition of dentate perforant path inputs. © 2015 The Authors Hippocampus Published by Wiley Periodicals, Inc.

Strong perisomatic inhibition by fast-spiking basket cells (FS-BCs) regulates dentate throughput. Homotypic FS-BC interconnections that support gamma oscillations, and heterotypic inputs from diverse groups of interneurons that receive extensive neurochemical regulation, together, shape FS-BC activity patterns. However, whether seizures precipitate functional changes in inhibitory networks and contribute to abnormal network activity in epilepsy is not known. In the first recordings from dentate interneuronal pairs in a model of temporal lobe epilepsy, we demonstrate that status epilepticus (SE) selectively compromises GABA release at synapses from dentate accommodating interneurons (AC-INs) to FS-BCs, while efficacy of homotypic FS-BC synapses is unaltered. The functional decrease in heterotypic cannabinoid receptor type 1 (CB1R)-sensitive inhibition of FS-BCs resulted from enhanced baseline GABAB-mediated suppression of synaptic release after SE. The frequency of CB1R-sensitive inhibitory synaptic events in FS-BCs was depressed early after SE induction and remained reduced in epileptic rats. In biologically based simulations of heterogeneous inhibitory networks and excitatory-inhibitory cell networks, experimentally identified decrease in reliability of AC-IN to FS-BCs synaptic release reduced theta power and theta-gamma coupling and enhanced gamma coherence. Thus, the experimentally identified functional reduction in heterotypic inhibition of FS-BCs can contribute to compromised network oscillations in epilepsy and could precipitate memory and cognitive co-morbidities.

Metabolomics is an essential discipline in industrial biotechnology. Sample preparation approaches dramatically influence data quality and, ultimately, interpretation and conclusions from metabolomic experiments. However, standardized protocols for highly reproducible metabolic datasets are limited, especially for the fungal cell factory Aspergillus niger. Here, an improved liquid chromatography-tandem mass spectrometry-based pipeline for A. niger metabolomics is developed. It is found that fast filtration with liquid nitrogen is more suitable for cell quenching, causing minimal disruption to cell integrity, and improved intracellular metabolite recovery when compared to cold methanol quenching approaches. Seven solutions are evaluated for intracellular metabolite extraction, and found acetonitrile/water (1:1, v/v) at -20 °C, combined with boiling ethanol extraction protocols, showed unbiased metabolite profiling. This improved methodology is applied to unveil the dynamic metabolite profile of one citrate over-producing A. niger isolate under citrate fermentation. Citrate precursors, especially pyruvate, oxaloacetate, and malate, are maintained at a relatively high intracellular level, which can be necessary for high citrate synthesis flux. Glutamine shows a similar trend compared to citrate production, suggesting glutamine may be involved in intracellular pH homeostasis. Taken together, this study delivers a highly standardized and improved metabolomics methodology and paves the way for systems metabolic engineering in biotechnologically important fungi.

Neuronal spike encoding and synaptic transmission in the brain need be precise and reliable for well-organized behavior and cognition. Little is known about how a unitary synapse reliably transmits presynaptic sequential spikes and how multiple unitary synapses precisely drive their postsynaptic neurons to encode spikes. To address these questions, we investigated the dynamics of glutamatergic unitary synapses as well as their role in driving the encoding of cortical fast-spiking neurons. Synaptic transmission patterns randomly fluctuate among facilitation, depression and parallel over time. The postsynaptic calmodulin-signaling pathway enhances initial responses and converts this fluctuation to a synaptic depression. We integrated current pulses mathematically based on synaptic plasticity and found that they improve spike capacity and timing precision by shortening the spike refractory period at postsynaptic neurons. Our results indicate that the gain and fidelity of synaptic patterns enable reliable transmission of presynaptic signals by the synapse and precise encoding of spikes by postsynaptic neurons. These reproducible neural codes may be involved in controlling well-organized behavior.

Altered inhibition is a salient feature of hippocampal network reorganization in epilepsy. Hippocampal pyramidal cells and dentate granule cells show specific reduction in cannabinoid receptor type 1 (CB1R)-sensitive GABAergic inputs in experimental epilepsy. In the dentate gyrus, CB1Rs regulate synaptic release from accommodating interneurons (AC-INs) with adapting firing characteristics and axonal projections in the molecular layer, but not from fast-spiking basket cells (FS-BCs). However, it is not known whether the intrinsic physiology and synaptic inhibition of AC-INs responsible for CB1R-sensitive inhibition is altered in epilepsy. Using the pilocarpine-induced status epilepticus (SE) model of epilepsy, we find that the basic physiological characteristics of AC-INs in epileptic rats are not different from age-matched controls. In paired interneuronal recordings, the amplitude of unitary inhibitory synaptic currents (uIPSCs) between AC-INs doubled after SE. Non-stationary noise analysis revealed that the post-SE strengthening of synapses between AC-INs resulted from an increase in postsynaptic receptors. Baseline synaptic release and CB1R antagonist enhancement of release at synapses between AC-INs were not different between control and post-SE rats. Additionally, uIPSC amplitude in FS-BCs to AC-INs pairs was unchanged after SE indicating input-specific microcircuit alterations in inhibitory inputs to AC-INs. At the network level, AC-INs showed no reduction in spontaneous and miniature inhibitory synaptic current (sIPSC or mIPSC) frequency or amplitude after SE. However, AC-IN mIPSC amplitude was persistently enhanced in post-SE and epileptic rats. CB1R agonist reduced the amplitude and suppressed a greater proportion of sIPSCs in AC-INs from post-SE and epileptic rats demonstrating a novel, cell-type specific increase in CB1R-sensitive inhibition of AC-INs after SE. This unique post-SE strengthening of inhibition between AC-INs could lead to activity-dependent suppression of AC-IN firing and compromise dentate CB1R-sensitive inhibition in epilepsy.

Neurons program various patterns of sequential spikes as neural codes to guide animal behavior. Studies show that spike programming (capacity and timing precision) is influenced by inhibitory synaptic inputs and membrane afterhyperpolarization (AHP). Less is clear about how these inhibitory components regulate spike programming, which we investigated at the cortical neurons. Whole-cell current-clamp recording for action potentials and single channel recording for voltage-gated sodium channels (VGSC) were conducted at regular-spiking and fast-spiking neurons in the cortical slices. With quantifying the threshold potentials and refractory periods of sequential spikes, we found that fast-spiking neurons expressing AHP possess lower threshold potentials and shorter refractory periods, and the hyperpolarization pulse immediately after each of spikes lowers threshold potentials and shortens refractory periods at regular-spiking neurons. Moreover, the hyperpolarization pulses shorten the refractory periods for VGSC reactivation and threshold potentials for its sequential activation. Our data indicate that inhibitory components immediately after spikes, such as AHP and recurrent inhibition, improve spike capacity and timing precision via lowering the refractory periods and threshold potentials mediated by voltage-gated sodium channels.

To study the chemical constituents of Viola tianshanica.Compounds were isolated by silica column, pharmadex LH-20 column and polyamide column, and their structures were elucidated by UV, IR, ESI-MS and NMR.Six compounds were isolated and identified as daucosterol (1), kaempferol-7-O-beta-D-glucopyranoside (2), kaempferol- 3-O-beta-D-glucopyranoside (3), isorhamnetin-3-O-beta-glucoside (4), kaempferol (5) and quercetin (6).Compounds 2-5 were isolated from this plant for the first time.

A scheme of integrated guidance/autopilot design for missiles with impact angle constraints is studied in this paper. When the integrated guidance/autopilot model yaw plane is formulated, the guidance/control law with impact angel constraints is designed based on the backstepping idea and VSC theory. For the unavailable information of the maneuvering target, an estimating method is given. Finally, to verify the effectiveness and rightness of the integrated design scheme, the simulation of some missile has been made, the simulation results have shown that high accuracy of hitting target with impact angle constraints can be got.

Gamma frequency oscillations have been proposed to contribute to memory formation and retrieval. Fast-spiking basket cells (FS-BCs) are known to underlie development of gamma oscillations. Fast, high amplitude GABA synapses and gap junctions have been suggested to contribute to gamma oscillations in FS-BC networks. Recently, we identified that, apart from GABAergic synapses, FS-BCs in the hippocampal dentate gyrus have GABAergic currents mediated by extrasynaptic receptors. Our experimental studies demonstrated two specific changes in FS-BC GABA currents following experimental seizures [Yu et al., J. Neurophysiol. 109, 1746 (2013)]: increase in the magnitude of extrasynaptic (tonic) GABA currents and a depolarizing shift in GABA reversal potential (E(GABA)). Here, we use homogeneous networks of a biophysically based model of FS-BCs to examine how the presence of extrasynaptic GABA conductance (g(GABA-extra)) and experimentally identified, seizure-induced changes in g(GABA-extra) and E(GABA) influence network activity. Networks of FS-BCs interconnected by fast GABAergic synapses developed synchronous firing in the dentate gamma frequency range (40-100 Hz). Systematic investigation revealed that the biologically realistic range of 30 to 40 connections between FS-BCs resulted in greater coherence in the gamma frequency range when networks were activated by Poisson-distributed dendritic synaptic inputs rather than by homogeneous somatic current injections, which were balanced for FS-BC firing frequency in unconnected networks. Distance-dependent conduction delay enhanced coherence in networks with 30-40 FS-BC interconnections while inclusion of gap junctional conductance had a modest effect on coherence. In networks activated by somatic current injections resulting in heterogeneous FS-BC firing, increasing g(GABA-extra) reduced the frequency and coherence of FS-BC firing when E(GABA) was shunting (-74 mV), but failed to alter average FS-BC frequency when E(GABA) was depolarizing (-54 mV). When FS-BCs were activated by biologically based dendritic synaptic inputs, enhancing g(GABA-extra) reduced the frequency and coherence of FS-BC firing when E(GABA) was shunting and increased average FS-BC firing when E(GABA) was depolarizing. Shifting E(GABA) from shunting to depolarizing potentials consistently increased network frequency to and above high gamma frequencies (>80 Hz). Since gamma oscillations may contribute to learning and memory processing [Fell et al., Nat. Neurosci. 4, 1259 (2001); Jutras et al., J. Neurosci. 29, 12521 (2009); Wang, Physiol. Rev. 90, 1195 (2010)], our demonstration that network oscillations are modulated by extrasynaptic inhibition in FS-BCs suggests that neuroactive compounds that act on extrasynaptic GABA receptors could impact memory formation by modulating hippocampal gamma oscillations. The simulation results indicate that the depolarized FS-BC GABA reversal, observed after experimental seizures, together with enhanced spillover extrasynaptic GABA currents are likely to promote generation of focal high frequency activity associated with epileptic networks.

BackgroundAction potentials are the essential unit of neuronal encoding. Somatic sequential spikes in the central nervous system appear various in amplitudes. To be effective neuronal codes, these spikes should be propagated to axonal terminals where they activate the synapses and drive postsynaptic neurons. It remains unclear whether these effective neuronal codes are based on spike timing orders and/or amplitudes.Methodology/principal findingsWe investigated this fundamental issue by simultaneously recording the axon versus soma of identical neurons and presynaptic vs. postsynaptic neurons in the cortical slices. The axons enable somatic spikes in low amplitude be enlarged, which activate synaptic transmission in consistent patterns. This facilitation in the propagation of sequential spikes through the axons is mechanistically founded by the short refractory periods, large currents and high opening probability of axonal voltage-gated sodium channels.Conclusion/significanceAn amplification of somatic incomplete spikes into axonal complete ones makes sequential spikes to activate consistent synaptic transmission. Therefore, neuronal encoding is likely based on spike timing order, instead of graded analogues.