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Neuroinflammation is a fundamental contributor to progressive neuronal damage, which arouses a heightened interest in neurodegenerative disease therapy. Ubiquitin-specific protease 7 (USP7) has a crucial role in regulating protein stability in multiple biological processes; however, the potential role of USP7 in neurodegenerative progression is poorly understood. Here, we discover the natural small molecule eupalinolide B (EB), which targets USP7 to inhibit microglia activation. Cocrystal structure reveals a previously undisclosed covalent allosteric site, Cys 576 , in a unique noncatalytic HUBL domain. By selectively modifying Cys 576 , EB allosterically inhibits USP7 to cause a ubiquitination-dependent degradation of Keap1. Keap1 function loss further results in an Nrf2-dependent transcription activation of anti-neuroinflammation genes in microglia. In vivo, pharmacological USP7 inhibition attenuates microglia activation and resultant neuron injury, thereby notably improving behavioral deficits in dementia and Parkinson’s disease mouse models. Collectively, our findings provide an attractive future direction for neurodegenerative disease therapy by inhibiting microglia-mediated neuroinflammation by targeting USP7.

Alzheimer’s disease (AD) is a chronic neurodegenerative disease, which is characterized by cognitive and synaptic plasticity damage. Rapamycin is an activator of autophagy/mitophagy, which plays an important role in identifying and degrading damaged mitochondria. The aim of this study was to investigate the effect of rapamycin on cognitive and synaptic plasticity defects induced by AD, and further explore if the underlying mechanism was associated with mitophagy. The results show that rapamycin increases Parkin-mediated mitophagy and promotes fusion of mitophagosome and lysosome in the APP/PS1 mouse hippocampus. Rapamycin enhances learning and memory viability, synaptic plasticity, and the expression of synapse-related proteins, impedes cytochrome C-mediated apoptosis, decreases oxidative status, and recovers mitochondrial function in APP/PS1 mice. The data suggest that rapamycin effectively alleviates AD-like behaviors and synaptic plasticity deficits in APP/PS1 mice, which is associated with enhanced mitophagy. Our findings possibly uncover an important function of mitophagy in eliminating damaged mitochondria to attenuate AD-associated pathology.

This study aims to identify the anti-pneumonia targets of Xiaoer Xiaoji Zhike Oral Liquid(XXZL) withquot;target fishingquot; strategy and investigate the related signaling pathways, thereby clarifying the anti-pneumonia mechanism of XXZL. To be specific, the magnetic nanoparticles cross-linked with XXZL extract were prepared based on the photochemical activity of benzophenone, which were then used to capture the target proteins from the lysate of tissue with lipopolysaccharide(LPS)-induced pneumonia in mice. Then, the target proteins were identified by liquid chromatography-tandem mass spectrometry(LC-MS/MS). The signaling pathways and interactions of target proteins were explored with KEGG and STRING analysis on Cytoscape, and the possible biological functions of the target proteins were verified by immunohistochemistry(IHC) and RT-PCR. The result showed that LC-MS/MS identified 62 potential anti-pneumonia targets of XXZL in the lungs. The targets were involved in Ras signaling pathway, mitophagy, leukocyte transendothelial migration, mitogen-activated protein kinase(MAPK) signaling pathway, platelet activation, and actomyosin structure organization, which were closely related to inflammation, pulmonary microcirculation, pulmonary fibrosis, and energy metabolism. XXZL up-regulated the content of CD31, and heat shock protein 60(HSP60) and ATP5 b mRNA expression, down-regulated interleukin-6(IL-6), tumor necrosis factor-α(TNF-α), COL1 A1 content, and alleviated fibrosis, which suggested the obvious effects of XXZL such as anti-inflammation, pulmonary microcirculation improvement, pulmonary fibrosis inhibition, and energy metabolism regulation. This study explained the anti-pneumonia mechanism of XXZL from targets, which can serve as a reference for the clinical application of the prescription.

Carbon quantum dots (CQDs) offer huge potential due to their enzymatic properties as compared to natural enzymes. Thus, discovery of CQDs-based nanozymes with low toxicity from natural resources, especially daily food, implies a promising direction for exploring treatment strategies for human diseases. Here, we report a CQDs-based biocompatible nanozyme prepared from chlorogenic acid (ChA), a major bioactive natural product from coffee. We found that ChA CQDs exhibited obvious GSH oxidase-like activities and subsequently promoted cancer cell ferroptosis by perturbation of GPX4-catalyzed lipid repair systems. In vivo, ChA CQDs dramatically suppressed the tumor growth in HepG2-tumor-bearing mice with negligible side toxicity. Particularly, in hepatoma H22-bearing mice, ChA CQDs recruited massive tumor-infiltrating immune cells including T cells, NK cells, and macrophages, thereby converting "cold" to "hot" tumors for activating systemic antitumor immune responses. Taken together, our study suggests that natural product-derived CQDs from coffee can serve as biologically safe nanozymes for anticancer therapeutics and may aid the development of nanotechnology-based immunotherapeutic.

Follistatin like protein 1 (FSTL1) is a famous growth regulatory protein. FSTL1 has been noticed in many diseases, including heart and lung ischemia, cerebral ischemia, glioma, schizophrenia, and Autism. The role of FSTL1 has been declared in the genetics and development of the central nervous system. Therefore, we designed this study to investigate the function and the role of FSTL1 in Alzheimer's disease. Firstly, we noticed upregulated expression level of FSTL1 among four to six-month-old 5XFAD AD mice. Accordingly, we hypothesized that FSTL1-Knockdown improved AD model mice's cognitive function and recover from Alzheimer's disease. Thus, AD model mice were made by single intracerebroventricular injections of Aβ

Existing treatment methods encounter difficulties in effectively promoting skin wound healing, making this a serious challenge for clinical treatment. Extracellular vesicles (EVs) secreted by stem cells have been proven to contribute to the regeneration and repair of wound tissue, but they cannot be targeted and sustained, which seriously limits their current therapeutic potential. The recombinant human collagen III protein (rhCol III) has the advantages of good water solubility, an absence of hidden viral dangers, a low rejection rate and a stable production process. In order to achieve a site-specific sustained release of EVs, we prepared a rhCol III hydrogel by cross-linking with transglutaminase (TGase) from Streptomyces mobaraensis, which has a uniform pore size and good biocompatibility. The release profile of the rhCol III-EVs hydrogel confirmed that the rhCol III hydrogel could slowly release EVs into the external environment. Herein, the rhCol III-EVs hydrogel effectively promoted macrophage changing from type M1 to type M2, the migration ability of L929 cells and the angiogenesis of human umbilical vein endothelial cells (HUVECs). Furthermore, the rhCol III-EVs hydrogel is shown to promote wound healing by inhibiting the inflammatory response and promoting cell proliferation and angiogenesis in a diabetic rat skin injury model. The reported results indicate that the rhCol III-EVs hydrogel could be used as a new biological material for EV delivery, and has a significant application value in skin wound healing.

Alzheimer's disease (AD) is a complex pathological neurodegenerative disease that seriously threatens human health. Therefore, how to effectively improve and treat AD is an urgent problem. In this study, a novel multitarget derivative based on tacrine (named 9i), which could work simultaneously on more than one pathological target, was used to treat AD model APP/PS1 transgenic mice. After 4 weeks of intragastric administration, cognitive function and synaptic plasticity were significantly improved and β-amyloid (Aβ) plaques that are main pathological hallmarks of AD were decreased in the APP/PS1 mice. On the one hand, 9i inhibited the excessive activation of the Raf/MEK/ERK signaling pathway to alleviate the loss of neurons, which provides a foundation for structural integrity. On the other hand, synaptic associated proteins and the density of synaptic spines were increased in APP/PS1 mice treated with 9i, which provides the basis for the improvement of synaptic plasticity and cognitive impairment. Interestingly, 9i also reduced Aβ plaques in the DG region, which is consistent with previous in vitro experiments showing that 9i inhibited the self-assembly of Aβ fibers, thus protecting neurons from Aβ plaque neurotoxicity. Our results suggest that 9i as a novel compound can effectively improve the cognitive function and the pathological changes of AD in APP/PS1 transgenic mice.

Five undescribed sesquiterpenoids stellerasespenes A‒E and four reported congeners were isolated from the roots of Stellera chamaejasme. The structures were elucidated by comprehensive spectroscopic analyses together with X-ray single crystal diffraction and theoretical calculations. The structure of holosericin B was revised. All the isolated compounds were evaluated for NO production in murine microglial BV2 cells induced by LPS. Stellerasespene A showed better inhibitory activity than the positive control minocycline, inhibiting NO production and overexpression of pro-inflammatory cytokine IL-1β in LPS-activated BV2 cells.

The balance between ubiquitination and deubiquitination is crucial for protein stability, function and location under physiological conditions. Dysregulation of E1/E2/E3 ligases or deubiquitinases (DUBs) results in malfunction of the ubiquitin system and is involved in many diseases. Increasing reports have indicated that ubiquitin-specific peptidases (USPs) play a part in the progression of many kinds of cancers and could be good targets for anticancer treatment. Glioma is the most common malignant tumor in the central nervous system. Clinical treatment for high-grade glioma is unsatisfactory thus far. Multiple USPs are dysregulated in glioma and have the potential to be therapeutic targets. In this review, we collected studies on the roles of USPs in glioma progression and summarized the mechanisms of USPs in glioma tumorigenesis, malignancy and chemoradiotherapy resistance.

Tacrine (Amino tetrahydroacridine hydrochloride hydrate) is a non-competitive and reversible inhibitor of acetylcholine esterase, and butylcholinesterase. Alzheimer's disease (AD) shows multiple types of pathological pathway in which cholinergic neuron deficiency is 95 % popular and the oldest pathological mechanism. However, the effect of tacrine on the hippocampal dependent memory is not yet known. In this study, we did verify that tacrine induced recovery of the specific pattern associated memory along with long-term memory through the improvement in the pattern of neural oscillation from deficits condition in the hippocampus of 6th month old AD mice. Our results showed that tacrine improved the performance of Morris water maze related spatial cognitive functions, and enhanced LTP in AD-TAC mice. Furthermore, our results implied that tacrine strongly improve the patterns of neural oscillations, and hippocampal synaptic plasticity in the 6th month old APP-PS1 double transgenic AD-TAC mice via changing the theta and alpha power spectra including with the improvement in theta, alpha and gamma synchronization. Moreover, tacrine generated the improvement in the theta cross spectra, theta-gamma phase-phase synchronization and theta-gamma phase-amplitude coupling. Besides, the data represented that tacrine accelerated the expression of NR2B, SYP and GAD65 while it caused deceleration on the expression of GAD67 neurotransmitter and Aβ. Thus, our results infer that tacrine works as a strong causative agent for improving the specific pattern-associated spatial long-term memory in the AD mice without showing any side effect.

Glioblastoma is a kind of malignant tumour and originates from the central nervous system. In the last century, some researchers and clinician have noticed that the psychosocial and neurocognitive functioning of patients with malignant gliomas can be impaired. Many clinical studies have demonstrated that part of patients, adults or children, diagnosed with glioblastoma will suffer from cognitive deficiency during their clinical course, especially in long-term survivors. Many nanoparticles (NPs) can inhibit the biological functions of tumours by modulating tumour-associated inflammation, which provokes angiogenesis and tumour growth. As one of the best antiviral nanoparticles (AVNPs), AVNP2 is the 2nd generation of AVNP2 that have been conjugated to graphite-graphene for improving physiochemical performance and reducing toxicity. AVNP2 inactivates viruses, such as the H1N1 and H5N1influenza viruses and even the SARS coronavirus, while it inhibits bacteria, such as MRSA and E. coli. As antimicrobials, nanoparticles are considered to be one of the vectors for the administration of therapeutic compounds. Yet, little is known about their potential functionalities and toxicities to the neurotoxic effects of cancer. Herein, we explored the functionality of AVNP2 on inhibiting C6 in glioma-bearing rats. The novel object-recognition test and open-field test showed that AVNP2 significantly improved the neuro-behaviour affected by C6 glioma. AVNP2 also alleviated the decline of long-term potentiation (LTP) and the decreased density of dendritic spines in the CA1 region induced by C6. Western blot assay and immunofluorescence staining showed that the expressions of synaptic-related proteins (PSD-95 and SYP) were increased, and these findings were in accordance with the results mentioned above. It revealed that the sizes of tumours in C6 glioma-bearing rats were smaller after treatment with AVNP2. The decreased expression of inflammatory factors (IL-1β, IL-6 and TNF-α) by Western blotting assay and ELISA, angiogenesis protein (VEGF) by Western blotting assay and other related proteins (BDNF, NF-ĸB, iNOS and COX-2) by Western blotting assay in peri-tumour tissue indicated that AVNP2 could control tumour-associated inflammation, thus efficiently ameliorating the local inflammatory condition and, to some extent, inhibiting angiogenesis in C6-bearing rats. In conclusion, our results suggested that AVNP2 could have an effect on the peri-tumor environment, obviously restraining the growth progress of gliomas, and eventually improving cognitive levels in C6-bearing rats.

Because of their widespread use and potential adverse effects in young developing organism, this study focused on the nephrotoxicity and genotoxicity of chronic low-dose exposure to silver nanoparticles (AgNPs) in 32 14-day-old male Wistar rats, randomly divided into three groups receiving AgNP solution (3 mg/kg body weight) intraperitoneally for one, two, or three weeks and the untreated control group (eight animals per group). When the rats were eight weeks old, blood creatinine and urine microalbumin were tested, followed by haematoxylin and eosin (H&E) staining. Proteinuria was found in the animals treated with AgNP for three weeks, and H&E staining revealed pathological changes in the kidney sections of this group. DNA damage was detected with the alkaline comet assay in the groups treated for two and three weeks. All results indicate that chronic exposure, even at a low dose, may affect animal health. The main culprit might be increased and time-dependent reactive oxygen species (ROS) production. Highly reactive ROS could cause a major structural damage to proteins and DNA, change the expression of ion channel proteins, and trigger inflammation. The findings of our in vivo experiment raise concern about nephrotoxic and genotoxic effects of silver nanoparticles in young organisms and call for further investigation of nanoparticle properties that can be modified to minimise the risks., Zbog široke primjene i rizika od štetnog djelovanja srebrnih nanočestica (AgNP) na mladi organizam u razvoju, u središtu pažnje ovog istraživanja bila je njihova nefrotoksičnost i genotoksičnost u 32 mužjaka Wistar štakora od 14. dana starosti koji su intraperitonealno primali otopinu AgNP-a (3 mg/kg tjelesne težine). Štakori su nasumce raspoređeni u tri izložene skupine (osam po skupini) – prva jedan tjedan, druga dva, a treća tri – i jednu kontrolnu skupinu, koja je primala fiziološku otopinu. Kad su štakori navršili osam tjedana, napravili smo pretrage krvi na kreatinin i mokraće na mikro albumin, a zatim i histološku pretragu s bojenjem hematoksilinom i eozinom (H&E). U životinja koje su tri tjedna bile izložene AgNP-u utvrđena je proteinurija, a H&E bojenjem utvrđene su patološke promjene na presjecima bubrežnog tkiva. Alkalnim komet testom otkriveno je oštećenje DNA u skupinama izloženima dva odnosno tri tjedna. Svi naši rezultati upućuju na to da kronična izloženost AgNP-u, makar i u niskim dozama, može utjecati na zdravlje životinja. Primarni je razlog za to vjerojatno povišena proizvodnja reaktivnih kisikovih spojeva (engl. reactive oxygen species, krat. ROS), koja je rasla s vremenom izloženosti. Visokoreaktivni kisikovi spojevi mogu uzrokovati velika strukturna oštećenja bjelančevina i DNA, izmijeniti ekspresiju ionskih kanala bjelančevina te potaknuti upalu. Rezultati našega in vivo istraživanja pozivaju ne samo na zabrinutost zbog nefrotoksičnih i genotoksičnih učinaka srebrnih nanočestica na mlade organizme nego i na potrebu daljnjeg istraživanja promjenjivih svojstava nanočestica ne bi li se rizik od njihova štetnog djelovanja sveo na najmanju moguću razinu.

The purpose of this study was to investigate whether soy isoflavone supplementation is effective in preventing periodontal destruction exacerbated by estrogen deficiency (ED) and its potential mechanism.The progression of periodontitis is affected by host factors, such as smoking, diabetes mellitus, and steroid use. Bone loss in periodontitis can be aggravated by ED.A rat model of experimental periodontitis (EP) with ED was established by silk ligature and inoculation with Porphyromonas gingivalis, and some EP rats were subjected to bilateral ovariectomy (OVX). The treatment groups received an intravenous injection of 17-β-estradiol (EEstrogen deficiency impaired the expression of genes encoding tight junction proteins in the gingiva, increased IL-17 level, and accelerated alveolar bone resorption. SI treatment alleviated tight junction protein expression, decreased IL-17 and ROS levels, and prevented the absorption of alveolar bone. Furthermore, GRHL2 expression was significantly induced by SI in human oral keratinocytes-1 (HOK-1) cells; GRHL2 knockdown impaired the expression of OCLN and ZO-1 induced by SI treatment.Soy isoflavones alleviates periodontitis in OVX rats, as observed by the increased expression of tight junction proteins, and reduced IL-17 level and alveolar bone loss. The in vitro studies suggested that the enhancement of oral epithelial barrier by SI treatment was partially dependent on GRHL2.

Cognitive dysfunction is a high incidence of diabetes mellitus (DM). However, the relationship between DM-induced cognitive defect and neuronal network oscillations is still unknown. In this study, adult male C57BL/6 J mice were intraperitoneally injected with streptozotocin (STZ) to duplicate DM. After 12 weeks, local field potentials were recorded in the perforant fiber pathway (PP) and dentate gyrus (DG) regions. Data showed that mice in the STZ group exhibited impairment of spatial learning and memory by the Morris Water Maze test. The low gamma (LG) and high gamma (HG) power were increased in the PP and DG areas of the STZ group. Moreover, the phase synchronization and the information flow at theta and LG rhythms between the PP and DG areas were decreased, and the theta-LG phase-amplitude coupling strength was markedly reduced in the PP region, DG region, and the PP-DG pathway in the STZ group. Additionally, the concentration of glutamate was increased by the high-performance liquid chromatography. Moreover, the NR2B and PSD95 expressions were markedly reduced, and the Akt/GSK-3β pathway was inhibited. Interestingly, the expressions of mGluRIIs (mGluR2 and mGluR3) were significantly decreased. The reduction of mGluRIIs may limit their function, such as restricting presynaptic glutamate release and reversing the dysfunction of NR2B via Akt/GSK-3β signaling pathway. In conclusion, our data suggest that DM alters the hippocampal neural network partly related to the dysfunction of mGluRIIs.

Inflammatory processes play a pivotal role in the development and progression of depression. Since Follistatin-like protein 1 (FSTL1) has been identified as a novel inflammatory protein, a variety of studies suggest that targeting FSTL1 may be useful in the treatment of diseases in which inflammation plays a central role. In the study, we aimed to investigate the causal relationship between FSTL1 signaling and the development of depression. To explore the effect and mechanism of FSTL1 on chronic stress-induced depression, the chronic unpredictable mild stress (CUMS) paradigm was used. Animals subjected to CUMS for 4 weeks exhibited depressive-like symptoms, including decreased sucrose preference and obvious behavioral despair, concomitantly with increased FSTL1 level in the hippocampus. In contrast, mice with FSTL1 knockdown abolished CUMS induced depression-like and anxiety-like behaviors. Moreover, FSTL1 knockdown reversed CUMS induced synaptic plasticity deficits in the PP-DG pathway of the hippocampus and increased the expression of synaptic associated proteins in the hippocampus of CUMS exposed mice. Microglia activation induced by CUMS paradigm could be significantly inhibited by FSTL1 knockdown. Furthermore, Western blot revealed that FSTL1 knockdown considerably decreased the expression of indicated molecules TLR4/MyD88/NF-κB signaling pathway in CUMS exposed mice. In conclusion, our data implies that FSTL1 may modulate the microglial activation through TLR4/MyD88/NF-κB signaling, which affects depression-like behaviors and synaptic function deficits induced by CUMS in mice. These results suggested that the role of FSTL1 in mediating microglia-related mechanisms in depression may shed light on developing new therapeutic strategies to treat this prevalent disease.

Alzheimer's disease (AD) is the most prevalent form of dementia in the old adult and characterized by progressive cognitive decline and neuronal damage. The mammalian Ste20-like kinase1/2 (MST1/2) is a core component in Hippo signaling, which regulates neural stem cell proliferation, neuronal death and neuroinflammation. However, whether MST1/2 is involved in the occurrence and development of AD remains unknown. In this study we reported that the activity of MST1 was increased with Aβ accumulation in the hippocampus of 5xFAD mice. Overexpression of MST1 induced AD-like phenotype in normal mice and accelerated cognitive decline, synaptic plasticity damage and neuronal apoptosis in 2-month-old 5xFAD mice, but did not significantly affect Aβ levels. Mechanistically, MST1 associated with p53 and promoted neuronal apoptosis by phosphorylation and activation of p53, while p53 knockout largely reversed MST1-induced AD-like cognitive deficits. Importantly, either genetic knockdown or chemical inactivation of MST1 could significantly improve cognitive deficits and neuronal apoptosis in 7-month-old 5xFAD mice. Our results support the idea that MST1-mediated neuronal apoptosis is an essential mechanism of cognitive deficits and neuronal loss for AD, and manipulating the MST1 activity as a potential strategy will shed light on clinical treatment for AD or other diseases caused by neuronal injury.

Diabetic neuropathy is one of the most common complications in patients with diabetes and leads to cognitive impairment. It is suggested that protracted hyperglycemia is the main trigger of cognitive deficits in diabetes and causes hippocampal abnormalities. Rapamycin, an inhibitor of mammalian target of rapamycin complex 1 (mTORC1), can significantly ameliorate cognitive deficits in neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease and Huntington disease. We employed whole-cell patch clamping to examine the effects of rapamycin on neuronal electrophysiological characteristics of the hippocampal dentate gyrus in mice under the condition of high glucose. We recorded the action potentials (APs) and the miniature excitatory postsynaptic currents (mEPSCs) of dentate gyrus neurons. We found that high glucose increased the half-width, the duration and decreased the peak amplitude of Aps as well as the inter-event interval (IEI) of mEPSCs in hippocampal dentate gyrus neurons. However, rapamycin pre-treatment reversed the changes induced by high glucose. Moreover, we demonstrated that rapamycin pre-treatment reversed the down-regulation of postsynaptic density protein 95 (PSD-95) expression caused by high glucose. Therefore, pre-treatment with rapamycin could ameliorate high glucose-induced alteration of synaptic transmission in the hippocampal dentate gyrus.

Frequent outbreaks of coronaviruses underscore the need for antivirals and vaccines that can counter a broad range of coronavirus types. We isolated a human antibody named 76E1 from a COVID-19 convalescent patient, and report that it has broad-range neutralizing activity against multiple α- and β-coronaviruses, including the SARS-CoV-2 variants. 76E1 also binds its epitope in peptides from γ- and δ-coronaviruses. 76E1 cross-protects against SARS-CoV-2 and HCoV-OC43 infection in both prophylactic and therapeutic murine animal models. Structural and functional studies revealed that 76E1 targets a unique epitope within the spike protein that comprises the highly conserved S2' site and the fusion peptide. The epitope that 76E1 binds is partially buried in the structure of the SARS-CoV-2 spike trimer in the prefusion state, but is exposed when the spike protein binds to ACE2. This observation suggests that 76E1 binds to the epitope at an intermediate state of the spike trimer during the transition from the prefusion to the postfusion state, thereby blocking membrane fusion and viral entry. We hope that the identification of this crucial epitope, which can be recognized by 76E1, will guide epitope-based design of next-generation pan-coronavirus vaccines and antivirals.

Inhibition of asparagine endopeptidase (AEP) has been implied to be effective for treating tau- and amyloid-beta-mediated neurodegenerative diseases, although a method for targeted intracerebral delivery of AEP inhibitors has not yet been achieved. Here, we fabricated ultrasound-responsive nanobubbles (NBs) to load AEP inhibitor RR-11a, and modified the NB surface with either AEP recognizable peptide AAN or pro-transendothelial transversal motif RGD, i.e. NB(11a)-A and NB(11a)-R, for AEP-targeted treatment of Alzheimer's disease (AD). The developed NBs were uniform, small in size (50.1 ± 1.5 nm), with strong echogenicity and high drug loading efficiency (∼91.97%). When intravenously co-injected in the APP/PS1 mouse model, NB(11a)-R could adhere to endothelial cells and enhance transient opening of the blood-brain barrier (BBB) upon focused ultrasound oscillations, allowing the rest NBs/localized released RR-11a molecules to enter the brain, and then NB(11a)-A could selectively bind with the impaired neurons and deposit RR-11a molecules at the AD lesion. As a result, co-administration of NB(11a)-A and NB(11a)-R significantly promoted accumulation of RR-11a in the mouse brain, and substantially alleviated both tau cleavage and amyloid plaques deposition in the hippocampus. Most strikingly, the cognitive ability of the AD model mice was dramatically improved, achieving a level close to the normal mice. Overall, this unique AEP-targeted nanobubble design provides an efficient intracerebral drug delivery strategy and significantly enhances treatment efficacy of AD. STATEMENT OF SIGNIFICANCE: Asparagine endopeptidase (AEP) is an innovative therapeutic target simultaneously involved in Aβ and tau-mediated Alzheimer's disease (AD) pathology, but targeted delivery of AEP inhibitors has not been achieved yet. Here we developed an efficient strategy to deliver AEP inhibitor RR-11a towards impaired neurons. We fabricated RR-11a-loaded ultrasound-responsive nanobubbles (NBs) and modified the NB surface with RGD peptide to promote BBB crossing upon focused ultrasound oscillations, or with AAN peptide to increase binding of NBs on the neurons. Our results indicated that, co-administration of the NB(11a)-A and NB(11a)-R significantly enhanced accumulation of RR-11a molecules at the AD lesion, alleviated both tau cleavage and amyloid plaques deposition in the hippocampus, and consequently restored cognitive function of the AD model mice.

In addition to motor dysfunction, cognitive impairments have been reported to occur in patients with early-stage Parkinson's disease (PD). In this study, we examined a PD mouse model induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). This treatment led to the degeneration of nigrostriatal dopaminergic neurons in mice, a phenomenon that is consistent with previous studies. Besides, spatial memory and object recognition of MPTP-treated mice were impaired, as denoted by the Morris water maze (MWM) and novel object recognition (NOR) tests, respectively. Moreover, hippocampal synaptic plasticity (long-term potentiation and depotentiation) and the levels of synaptic proteins in hippocampus were decreased after MPTP treatment. We also found that MPTP resulted in the microglial activation and an inflammatory response in the striatum and hippocampus. Mammalian asparagine endopeptidase (AEP), a cysteine lysosomal protease, is involved in the cleavage and activation of Toll-like receptors (TLRs). The deletion of AEP can inhibit TLR4 in a mouse model of Alzheimer's disease, and TLR4 is upregulated in PD, inducing microglial activation and inflammation. We found that AEP deletion provided greater resistance to the toxic effects of MPTP. AEP knockout ameliorated the cognition and the synaptic plasticity defects in the hippocampus. Furthermore, AEP deletion decreased the expression of TLR4 and reduced microglial activation and the levels of several proinflammatory cytokines. Thus, we suggest that AEP plays a role in the inflammation induced by MPTP, and TLR4 might also involve in this process. AEP deletion could be a possible treatment strategy for the cognitive deficits of PD.

Antimicrobial peptides are a fundamental component of mollusks’ defense systems, though they remain a thinly investigated subject. Here, infection by Vibrio parahemolyticus triggered a significant increase in antimicrobial activity in oyster plasma. By using PBS-challenged oysters as a control, plasma peptides from immunologically challenged oysters were subjected to peptidomic profiling and in silico data mining to identify bioactive peptides. Thirty-five identified plasma peptides were up-regulated post infection, among which, six up-regulated peptides (URPs) showed a relatively high positive charge. URP20 was validated with significant antibacterial activity. Virtually, URP20 triggered aggregation of bacterial cells, accompanied by their membrane permeabilization. Interestingly, URP20 was found to be active against Gram-positive and Gram-negative foodborne pathogens as well as Candida albicans, with no cytotoxicity to mammalian cells and mice. Our study provides the first large-scale plasma peptidomic dataset that identifies novel bioactive peptides in marine mollusks. Further exploration of peptide diversity in marine invertebrates should prove a fruitful pursuit for designing novel AMPs with broad applications.

This study identifies the endolysosomal two-pore channel TPC2 as a potential novel drug target for age-related macular degeneration (AMD). Inhibition of TPC2 reduces two key clinical hallmarks of the AMD, retinal inflammation and formation of leaky choroi., Age-related macular degeneration (AMD) is the most common cause of blindness among the elderly and can be classified either as dry or as neovascular (or wet). Neovascular AMD is characterized by a strong immune response and the inadequate release of cytokines triggering angiogenesis and induction of photoreceptor death. The pathomechanisms of AMD are only partly understood. Here, we identify the endolysosomal two-pore cation channel TPC2 as a key factor of neovascularization and immune activation in the laser-induced choroidal neovascularization (CNV) mouse model of AMD. Block of TPC2 reduced retinal VEGFA and IL-1β levels and diminished neovascularization and immune activation. Mechanistically, TPC2 mediates cationic currents in endolysosomal organelles of immune cells and lack of TPC2 leads to reduced IL-1β levels in areas of choroidal neovascularization due to endolysosomal trapping. Taken together, our study identifies TPC2 as a promising novel therapeutic target for the treatment of AMD.

Generating and improving antibodies and peptides that bind specifically to membrane protein targets such as ion channels and G protein-coupled receptors (GPCRs) can be challenging using established selection methods. Current strategies are often limited by difficulties in the presentation of the antigen or the efficiency of the selection process. Here, we report a method for obtaining antibodies specific for whole cell membrane-associated antigens which combines a cell–cell interaction format based on yeast display technology with fluorescence-activated cell sorting of dual fluorescent complexes. Using this method, we were able to direct the affinity maturation of an antagonist antibody specific for the proton-gated ion channel ASIC1a and showed that both the affinity and potency were improved. We were also able to use this method to do kinetic selections to generate clones with better dissociation profiles. In addition, this method was employed successfully to handle the difficult problem of selecting antibodies specific to a GPCR target, the mu-opioid receptor.

The transient receptor potential vanilloid 4 (TRPV4) channel may be opened by mechanical stimuli to mediate Ca2+ and Na+ influxes, and it has been suggested to mediate glaucoma retinopathy. However, it has been mostly unclear how TRPV4 activities affect the function of primate retinal ganglion cells (RGCs). We studied RGCs and bipolar cells (BCs) in the peripheral retina of the old-world primate using whole-cell current-clamp and voltage-clamp recordings, immunomarkers and confocal microscopy. RGCs were distinguished from displaced amacrine cells (ACs) by the absence of GABA and glycine immunoreactivity and possession of an axon and a large soma in the RGC layer. Strong TRPV4 signal was concentrated in medium to large somas of RGCs, and some TRPV4 signal was found in BCs (including PKCα-positive rod BCs), as well as the end feet, soma and outer processes of Mȕller cells. TRPV4 immunoreactivity quantified by the pixel intensity histogram revealed a high-intensity component for the plexiform layers, a low-intensity component for the soma layers of ACs and Mȕller cells, and both components in the soma layers of RGCs and BCs. In large RGCs, TRPV4 agonists 4α-phorbol 12,13 didecanoate (4αPDD) and GSK1016790A reversibly enhanced the spontaneous firing and shortened the delay of voltage-gated Na+ (Nav) currents under current-clamp conditions, and under voltage-clamp conditions, 4αPDD largely reversibly increased the amplitude and frequency of spontaneous excitatory postsynaptic currents. In BCs, changes in the membrane tension induced by either applying pressure or releasing the pressure both activated a transient cation current, which reversed at ~ −10 mV and was enhanced by heating from 24 °C to 30 °C. The pressure for the half-maximal effect was ~18 mmHg. These data indicate that functional TRPV4 channels are variably expressed in primate RGCs and BCs, possibly contributing to pressure-related changes in RGCs in glaucoma.

In this study, we have, for the first time, demonstrated the beneficial effects of transcranial magneto-acoustic stimulation (TMAS), a technique based on focused ultrasound stimulation within static magnetic field, on the learning and memory abilities and neuroplasticity of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson’s disease (PD). Our results showed that chronic TMAS treatment (2 weeks) improved the outcome of Morris water maze, long-term potentiation (LTP), and dendritic spine densities in the dentate gyrus (DG) region of the hippocampus of PD model mice. To further investigate into the underlying mechanisms of these beneficial effects by TMAS, we quantified the proteins in the hippocampus that regulated neuroplasticity. Results showed that the level of postsynaptic density protein 95 was elevated in the brain of TMAS-treated PD model mice while the level of synaptophysin (SYP) did not show any change. We further quantified proteins that mediated neuroplasticity mechanisms, such as brain-derived neurotrophic factor (BDNF) and other important proteins that mediated neuroplasticity. Results showed that TMAS treatment elevated the levels of BDNF, cAMP response element–binding protein (CREB), and protein kinase B (p-Akt) in the PD model mouse hippocampus, but not in the non-PD mouse hippocampus. These results suggest that the beneficial effects on the neuroplasticity of PD model mice treated with TMAS could possibly be conducted through postsynaptic regulations and mediated by BDNF. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13311-019-00732-5) contains supplementary material, which is available to authorized users.

Ethnopharmacological relevance Curcuma Longa Radix (Huangsiyujin in Chinese, HSYJ) has been used in clinical for thousands of years as a traditional Chinese medicine (TCM) and has the effects of invigorating the blood circulation, relieving pain, promoting qi circulation and relieving depression, besides, it is able to clear heart heat and cool blood and cure jaundice. The study aims to determine the active fractions of HSYJ for promotion of blood circulation and relief of pain, and to identify their chemical constituents. Material and methods HSYJ was extracted by the systematic solvent method. The effects of promoting blood circulation and relieving pain were determined by measuring pain threshold (hot-plate test), latency of twisting and writhing times (acetic acid induced writhing test), and hemorheology of mice. Chemical constituents of the extractive fractions were analyzed by Gas Chromatography-Mass Spectrometer (GC-MS) or High Performance Liquid Chromatography (HPLC). Results The results showed that five fractions (volatile oil fraction, petroleum ether fraction, ethyl acetate fraction, 1-butanol fraction and water fraction) all could increase pain threshold, prolong latency of twisting, decrease the writhing times and influence hemorheology. However, the effects of the volatile oil fraction and ethyl acetate fraction were better than other fractions. Three constituents, namely bisdesmethoxycurum, desmethoxycurumin and curcumin were identified from the active fractions by comparing with standard preparation through HPLC analysis. Forty-five compounds including aR-Turmerone, Curlone, Tumerone, Cyclohexene,1-methyl-4-(1-methylethylidene)-, trans-Sesquisabinene hydrate, (E)-Atlantone, α-curcumene and Zingiberene were identified from the volatile oil fraction by GC-MS. Conclusion HSYJ possessed the effect of promoting blood circulation and relieving pain. Curcumins and their derivatives were the major constituents and might be the main bioactive ingredients for the promotion of blood circulation and relief of pain. The study provided evidence and formed a basis for the establishment of HSYJ comprehensive quality evaluation system related to traditional efficacy. It is beneficial to the quality control of HSYJ and its proprietary Chinese medicine.

6-Phosphogluconate dehydrogenase (6PGD) is a key enzyme that converts 6-phosphogluconate into ribulose-5-phosphate with NADP+ as cofactor in the pentose phosphate pathway (PPP). 6PGD is commonly upregulated and plays important roles in many human cancers, while the mechanism underlying such roles of 6PGD remains elusive. Here we show that upon EGFR activation, 6PGD is phosphorylated at tyrosine (Y) 481 by Src family kinase Fyn. This phosphorylation enhances 6PGD activity by increasing its binding affinity to NADP+ and therefore activates the PPP for NADPH and ribose-5-phosphate, which consequently detoxifies intracellular reactive oxygen species (ROS) and accelerates DNA synthesis. Abrogating 6PGD Y481 phosphorylation (pY481) dramatically attenuates EGF-promoted glioma cell proliferation, tumor growth and resistance to ionizing radiation. In addition, 6PGD pY481 is associated with Fyn expression, the malignancy and prognosis of human glioblastoma. These findings establish a critical role of Fyn-dependent 6PGD phosphorylation in EGF-promoted tumor growth and radiation resistance., 6-phosphogluconate dehydrogenase is commonly upregulated in cancers. Here, the authors show that activation of EGFR induces phosphorylation of this enzyme at Y481 to activate the pentose phosphate pathway, which consequently reduces ROS and accelerates DNA synthesis to promote tumor growth and radioresistance.

The current cytological evaluation technique of health food raw materials does not entirely meet the needs of evaluating health food. Our study adopted the microfluidic chip technique for the first time to construct a hepatocyte model of evaluating emodin, which was composed of a human hepatocellular carcinoma cell (HepG2) and microfluidic chip. The mixed glue of a model with rat tail collagen type I (1.3 mg/mL) + gelatin (7.5%) was used to simulate the microenvironment of a cell. The validity of this model was evaluated by cell proliferation activity and cell staining, and the toxicity of emodin was evaluated by a series of metabolic indicators on this model. The results indicated that the repeatability of the constructed hepatocyte model was favorable, with a coefficient of variation (CV) of 2.8%. After emodin continuously was exposed for 48 h, the cell inhibition was obvious at 100 and 200 μM, and the number of dead cells gradually increased with the increasing of emodin concentration, and the difference of BUN was significant between the emodin group and blank group (p < 0.05). The constructed model has a favorable applicability in evaluating emodin. This study provides an important platform and a potential in vitro alternative model for assessing and predicting the health effects of health food.

Antibiotic residues contained in poultry eggs pose threat to human health. However, the classes and concentrations of antibiotics in poultry egg in southwestern China is unknown due to insufficient monitoring and research. A total of 513 egg samples were collected from supermarkets and farm markets in Kunming city in 2020 and the levels of 7 antibiotics were analyzed using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method. The linear correlation coefficients were above 0.990 for all antibiotics tested. The limits of detection and limits of quantification in poultry eggs were 0.002 to 0.010 μg/g and 0.007 to 0.033 μg/g, respectively. The average recoveries of the 7 analytes from poultry egg samples were 80.00 to 128.01%, with relative standard deviations of less than 13.97%. A total of 93 (18.13%) samples tested positive for antibiotics, with the highest concentration being 2.48 μg/g. The concentration range of ofloxacin, danofloxacin, difloxacin, sulfadimethoxine, sulfamonomethoxine, sulfamethoxypyridazine, and sulfamethoxazole in poultry eggs was 0.01 to 0.37 μg/g, 0.06 to 0.48 μg/g, 0.05 to 0.29 μg/g, 0.03 to 0.16 μg/g, 0.06 to 1.00 μg/g, 0.05 to 0.37, and 0.07 to 2.48 μg/g, respectively. Sulfamonomethoxine was detected from hen eggs with the highest concentration level at 1.00 μg/g. Sulfamethoxazole was detected with the highest concentration level from both duck and quail eggs, at 1.87 and 2.48 μg/g, respectively. The antibiotic with the highest residue level in pheasant eggs was danofloxacin, which was 0.37 μg/g. Sulfamethoxypyridazine was identified in 30 samples with the highest positive rate of 5.85%, sulfadimethoxine was identified in 3 samples with the lowest positive rate of 0.58%. We observed that 7 targeted antibiotic residues in quail eggs and 3 targeted antibiotic residues in pheasant eggs. We also found that there were antibiotic residues in free-range hen eggs and the concentration was not low. The antibiotic with the highest residue level in free-range eggs was sulfamonomethoxine, which was 1.00 μg/g. These findings suggest that continual antibiotic residue monitoring of poultry eggs is essential in China.

Recently, we reported that presenilin 1 considerably increased the expression level of U1 small nuclear RNA (snRNA) accompanied with the adverse change of amyloid precursor protein (APP) expression, β-amyloid (Aβ) production and cell apoptosis. In the present study, it was found that U1 snRNA overexpression significantly elevated the expression level of autophagy. Moreover, rapamycin further enhanced the Aβ production and cell apoptosis, whereas these processes were effectively inhibited by 3-MA. Acridine orange staining images showed that U1 snRNA overexpression not only activated autophagy pathway, but also led to the autophagic-lysosomal system dysfunction in cells. Immunofluorescence assay showed autophagic vacuoles localization with APP, which was the precursor protein of main component of toxic protein in AD. Meanwhile, the superoxide dismutase activity was remarkably decreased and MDA level was significantly increased by U1 snRNA overexpression in cells, suggesting that there was a possible pathway to elucidate how the U1 snRNA overexpression induced cell damage. We further found that U1 snRNA overexpression altered lysosomal biogenesis and autophagic-lysosomal fusion. In combination with our previous results, it suggests that the malfunction of autophagy pathway provides important insight into molecular mechanisms of augment the aggregation of Aβ and induction of cell apoptosis contributed to AD.

Hypertension is the most common chronic disease accompanied by cognitive decline and anxiety-like behavior. Angiotensin II (Ang II) induces hypertension by activating angiotensin II receptor subtype 1 (AT1R). The purpose of the study was to examine the potential underlying mechanism of alterations in cognition and anxiety-like behavior induced by Ang II. Adult C57 mice were intraperitoneal injected with either 1 mg/kg/d Ang II or saline individually for 14 consecutive days. Ang II resulted in cognitive decline and anxious like behavior in C57 mice. Moreover, Ang II disturbed bidirectional synaptic plasticity and neural oscillation coupling between high theta and gamma on PP (perforant pathway)-DG (dentate gyrus) pathway. In addition, Ang II decreased the expression of N-methyl-d-aspartate receptor (NR) 2A and NR 2B and increased the expression of GABAAR α1. The data suggest that Ang II disturb neural oscillations via altering excitatory and inhibitory (E/I) balance and eventually damage cognition and anxiety-like behavior in mice.

Mylabris is the dried body of the Chinese blister beetle (Mylabris sp.), which has been used in traditional Chinese medicine and achieved significant positive effects in the treatment of cancer including liver cancer, lung cancer, and rectal cancer. However, heavy metal pollution and accumulation of Mylabris insects could pose threat to human health. This study was carried out to assess levels of different heavy metals like Cu, As, Cd, Hg, and Pb, along with soil–plant-insect system and health risks using two representative Mylabris insects from the Hasi Mountains of Gansu Province, China. The results showed that the heavy metal concentration of plants and insects followed the order Cu > Pb > As > Hg > Cd. Compared with soil and plants, the content of Cu in insects was the highest, reaching 45.65 mg/kg. Cu was the main element that caused insects to absorb and accumulate. The quantitative risk analysis implied the two Mylabris insects had carcinogenic risks, with the contribution of As providing 63% and 60.7%, respectively. This kind of carcinogenic risk that the human body could bear was not easy to cause side effects to normal people, but it was difficult and dangerous for cancer patients. Thus, the evaluation of health risk lays the foundation for pollutant risk monitoring.

F-box and WD repeat domain containing 7 (FBXW7) is a substrate receptor of the ubiquitin ligase SKP1-Cullin1-F-box complex and a potent tumor suppressor that prevents unregulated cell growth and tumorigenesis. However, little is known about FBXW7-mediated control of cell metabolism and related functions in cancer therapy. Here, we report that FBXW7 expression inversely correlates with the expression levels of the key metabolic enzyme isocitrate dehydrogenase 1 (IDH1) in patients with glioma and public glioma datasets. Deletion of FBXW7 significantly increased both wild-type (WT) and mutant IDH1 expression, which was mediated by blocking degradation of sterol regulatory element binding protein 1 (SREBP1). The upregulation of neomorphic mutant IDH1 by FBXW7 deletion stimulated production of the oncometabolite 2-hydroxyglutarate at the expense of increasing pentose phosphate pathway activity and NADPH consumption, limiting the buffering ability against radiation-induced oxidative stress. In addition, FBXW7 knockout and IDH1 mutations induced nonhomologous end joining and homologous recombination defects, respectively. In vitro and in vivo, loss of FBXW7 dramatically enhanced the efficacy of radiation treatment in IDH1-mutant cancer cells. Taken together, this work identifies FBXW7 deficiency as a potential biomarker representing both DNA repair and metabolic vulnerabilities that sensitizes IDH1-mutant cancers to radiotherapy. Significance: Deficiency of FBXW7 causes defects in DNA repair and disrupts NADPH homeostasis in IDH1-mutant glioma cells, conferring high sensitivity to radiotherapy.

Notch1 not only plays a key role in the development of the nervous system but also modulates synaptic plasticity and memory. However, the role of Notch1 in the brain of diabetes is still unclear. We hypothesize that Notch1 is involved in type I diabetes-induced cognitive dysfunction. In this study, adult male C57BL/6J mice carrying a heterozygous null mutation in the Notch1 gene (Notch1+/-) and wild-type littermate controls were used in this experiment. They were subjected to streptozocin (55 mg/kg, i.p.) for consecutive five days. After 12 weeks, the cognitive function of all mice was detected by novel object recognition (NOR) test and electrophysiological recording. Our results demonstrated that the levels of Notch1 mRNA and Notch1 receptor were increased in the hippocampus of the wild-type diabetic mice at 12 weeks. It suggested that the Notch1 signal pathway was activated. Compared with the wild-type diabetic mice, the discrimination index and the long-term potentiation was further decreased in the Notch1+/- diabetic group, the impairment of neuronal ultrastructure was exacerbated in the hippocampus of the Notch1+/- diabetic mice, and the number of synapses and autophagic vacuoles were significantly reduced in the Notch1+/- diabetic group. Moreover, some postsynaptic associated protein expressions were down-regulated, as well as the Beclin1 expression and the ratio of LC3II/LC3I were reduced in the hippocampus of the Notch1+/- diabetic mice. Interestingly, the phosphorylation of mTOR, Akt, and ERK1/2 were all inhibited in the Notch1+/- diabetic group. Taken together, these results suggest that Notch1 deficiency deteriorates the synaptic plasticity and inhibits the activation of autophagy partly via the mTOR-independent signal pathway in the hippocampus of type I diabetic mice.

MDM2 (mouse double-minute) and p53 form a negative feedback loop and play a prominent role in preventing the induction of uncontrolled apoptosis. To better understand their potential roles in oyster Crassostrea hongkongensis, MDM2 and p53 homologues were first isolated and cloned in C. hongkongensis (named ChMDM2 and Chp53), and their mRNA expression patterns in tissues and developmental stages were analyzed. Multiple sequence alignment analysis and phylogenetic analysis of ChMDM2 and Chp53 displayed a high degree of homology and conservation. In addition, exposure to Vibrio coralliilyticus resulted in DNA damage and apoptosis in the hemocytes of C. hongkongensis, and found that the mRNA expression level of ChMDM2 was decreased, while the relative expression of Chp53 was significantly increased in the hemocytes and gills. Furthermore, fluorescence from ChMDM2-EGFP and Chp53-Red were found to be distributed in the nucleus of HEK293T cells. Besides, dual-luciferase reporter assays showed that ChMDM2 antagonized with Chp53 and participates in p53 signaling pathway. In addition, the interaction between ChMDM2 and Chp53 was confirmed strongly by Co-immunoprecipitation assays. Furthermore, the results of RNAi showed that ChMDM2 and Chp53 participated in apoptosis which induced infection of V. coralliilyticus. Taken together, our results characterized the features of ChMDM2 and Chp53, which played a critical role in apoptosis of C. hongkongensis.

Hepatocellular carcinoma (HCC), the most prevalent liver cancer, is considered one of the most lethal malignancies with a dismal outcome. There is an urgent need to find novel therapeutic approaches to treat HCC. At present, natural products have served as a valuable source for drug discovery. Here, we obtained five known biflavones from the root of Stellera chamaejasme and evaluated their activities against HCC Hep3B cells in vitro. Chamaejasmenin E (CE) exhibited the strongest inhibitory effect among these biflavones. Furthermore, we found that CE could suppress the cell proliferation and colony formation, as well as the migration ability of HCC cells, but there was no significant toxicity on normal liver cells. Additionally, CE induced mitochondrial dysfunction and oxidative stress, eventually leading to cellular apoptosis. Mechanistically, the potential target of CE was predicted by database screening, showing that the compound might exert an inhibitory effect by targeting at c-Met. Next, this result was confirmed by molecular docking, cellular thermal shift assay (CETSA), as well as RT-PCR and Western blot analysis. Meanwhile, CE also reduced the downstream proteins of c-Met in HCC cells. In concordance with above results, CE is efficacious and non-toxic in tumor xenograft model. Taken together, our findings revealed an underlying tumor-suppressive mechanism of CE, which provided a foundation for identifying the target of biflavones.

Ischemic stroke remains one of the leading causes of death worldwide, thereby highlighting the urgent necessary to identify new therapeutic targets. Deoxyhypusine hydroxylase (DOHH) is a fundamental enzyme catalyzing a unique posttranslational hypusination modification of eukaryotic translation initiation factor 5A (eIF5A) and is highly involved in the progression of several human diseases, including HIV-1 infection, cancer, malaria, and diabetes. However, the potential therapeutic role of pharmacological regulation of DOHH in ischemic stroke is still poorly understood. Our study first discovered a natural small-molecule brazilin (BZ) with an obvious neuroprotective effect against oxygen-glucose deprivation/reperfusion insult. Then, DOHH was identified as a crucial cellular target of BZ using HuProt™ human proteome microarray. By selectively binding to the Cys232 residue, BZ induced a previously undisclosed allosteric effect to significantly increase DOHH catalytic activity. Furthermore, BZ-mediated DOHH activation amplified mitophagy for mitochondrial function and morphology maintenance via DOHH/eIF5A hypusination signaling pathway, thereby protecting against ischemic neuronal injury in vitro and in vivo. Collectively, our study first identified DOHH as a previously unreported therapeutic target for ischemic stroke, and provided a future drug design direction for DOHH allosteric activators using BZ as a novel molecular template.

We observed that miR‐22 can promote hepatic steatosis. Conversely, steatosis was alleviated after inhibition of miR‐22, indicating that miR‐22 is involved in fat deposition. Furthermore, miR‐22 was validated to have effects on downstream lipid metabolism‐related genes, revealing the pathways through which miR‐22 regulates lipid metabolism. Therefore, miR‐22 may be a promising target for the treatment of NAFLD and obesity., Nonalcoholic fatty liver disease (NAFLD) is highly correlated with obesity, and lifestyle changes to reduce weight remain the main therapeutic approach. The noncoding RNA miR‐22 has previously been reported to be highly abundant in the sera of NAFLD patients. In addition, miR‐22 directly targets peroxisome proliferative‐activated receptor, Pgc‐1α, peroxisome proliferator‐activated receptor α, and sirtuin 1 (Sirt1), which are important factors involved in fatty acid metabolism. Given that miR‐22 directly targets genes involved in the control of metabolism and obesity, we investigated whether miR‐22 contributes to metabolic alterations induced by obesity. We observed increased expression of miR‐22, decreased expression of Sirt1, and alterations in the expression of adipogenesis‐related genes in a mouse model of obesity and a human hepatocyte cell line. We identified that miR‐22 and the 3′‐UTR of Sirt1 are complementary. Mutation of the complementary fragment abolishes the ability of miR‐22 to regulate the Sirt1 gene. Furthermore, treatment of hepatic steatosis cells with miR‐22 mimics or inhibitors showed that miR‐22 can promote hepatic steatosis, and miR‐22 inhibitors effectively reduced triglyceride levels without affecting cell activity. Finally, we validated that miR‐22 has similar effects on downstream lipid metabolism‐related genes. Our data reveal the pathways and mechanisms through which miR‐22 regulates lipid metabolism and suggest that miR‐22 inhibitors may have potential as candidate drugs for NAFLD and obesity.

Alzheimer’s disease (AD) is a neurodegenerative disease of the central nervous system characterised by cognitive impairment. Its major pathological feature is the deposition of β-amyloid (Aβ) peptide, which triggers a series of pathological cascades. Autophagy is a main pathway to eliminate abnormal aggregated proteins, and increasing autophagy represents a plausible treatment strategy against relative overproduction of neurotoxic Aβ. Graphene oxide (GO) is an emerging carbon-based nanomaterial. As a derivative of graphene with neuroprotective effects, it can effectively increase the clearance of abnormally aggregated protein. In this article, we investigated the protective function of GO in an AD mouse model. GO (30 mg/kg, intraperitoneal) was administered for 2 weeks. The results of the Morris water maze test and the novel object recognition test suggested that GO ameliorated learning and memory impairments in 5xFAD mice. The long-term potentiation and depotentiation from the perforant path to the dentate gyrus in the hippocampus were increased with GO treatment in 5xFAD mice. Furthermore, GO upregulated the expression of synapse-related proteins and increased the cell density in the hippocampus. Our results showed that GO up-regulated LC3II/LC3I and Beclin-1 and decreased p62 protein levels in 5xFAD mice. In addition, GO downregulated the PI3K/Akt/mTOR signalling pathway to induce autophagy. These results have revealed the protective potential of GO in AD.

Background As a novel type of non-coding RNA, circular RNAs (circRNAs) play a critical role in the initiation and development of various diseases, including cancer. However, the exact function of circRNAs in human cervical cancer remains largely unknown. Methods We identified the circRNA signature of upregulated circRNAs between cervical cancer and paired adjacent normal tissues. Using two different cohorts and GEO database, a total of six upregulated circRNAs were identified with a fold change > 2, and P CircRNA pull-down, circRNA immunoprecipitation (circRIP) and Co-immunoprecipitation (Co-IP) assays were executed to investigate the molecular mechanism underlying the function of circZFR. Results Functionally, circZFR knockdown represses the proliferation, invasion, and tumor growth. Furthermore, circRNA pull-down experiments combined with mass spectrometry unveil the interactions of circZFR with Single-Stranded DNA Binding Protein 1 (SSBP1). Mechanistically, circZFR bound with SSBP1, thereby promoting the assembly of CDK2/cyclin E1 complexes. The activation of CDK2/cyclin E1 complexes induced p-Rb phosphorylation, thus releasing activated E2F1 leading to cell cycle progression and cell proliferation. Conclusion Our findings provide the first evidence that circZFR is a novel onco-circRNA and might be a potential biomarker and therapeutic target for cervical cancer patients.

The deficiency or insufficiency of androgen can trigger a range of reproductive diseases as well as other symptoms. Stem Leydig cells (SLCs) are critical for the formation and maintenance of a functional androgen-producing cell (Leydig cell, LC) population throughout adult male life. However, to date, our knowledge about SLCs is poor. Here we report the derivation and characterisation of a clonal stem LC line (designated as TSL) capable of 11- ketotestosterone (11-KT) production from a 3-month-old Nile tilapia (Oreochromis niloticus) testis. The cells retained stable proliferation after 77 generations with normal karyotype and growth factor dependency. They expressed platelet-derived growth factor receptor-α (pdgfrα), nestin and chicken ovalbumin upstream promoter transcription factor II (coup-tfIIa), which are characteristic of SLCs. Upon induction in defined medium, TSLs could undergo differentiation into steroidogenically active LCs and produce 11-KT. When implanted into recipient Nile tilapia testes from which endogenous LCs had been eliminated by ethane dimethanesulphonate (EDS) treatment, the PKH26-labelled TSLs could colonise the interstitium, subsequently express steroidogenic genes and restore 11-KT production. Taken together, our data suggest that TSLs possess the ability of continuous proliferation and potential of differentiation into functional LCs invitro and invivo. To the best of our knowledge TSL might represent the first stem LC line capable of 11-KT production to date. Our study may offer new opportunities for investigating the self-renewal of SLCs and steroidogenesis invitro, and provide an invaluable invitro model for investigating endocrine disruptors.

Emerging evidence indicates that circRNAs play essential roles in tumorigenesis and development. However, the role of circRNAs in cervical cancer (CC) remains unclear. CircRNA microarrays performed on the immortal cervical cell line H8 and the cervical cancer cell line SiHa were used to identify a circRNA, termed circNRIP1 (hsa_circ_0004771), which was upregulated in SiHa. QRT-PCR confirmed that circNRIP1 was upregulated in CC tissues, where its expression was correlated with lymphovascular space invasion. Besides, both in vitro and in vivo experiments demonstrated that circNRIP1 promotes cell proliferation, migration, and invasion. Additionally, we found that miR-629-3p induced tumor suppression by regulating PTP4A1 and the ERK1/2 pathway. Finally, we confirmed that circNRIP1 exerts its effect, at least partially, by sponging miR-629-3p and thereby regulating the PTP4A1/ERK1/2 pathway. Therefore, circNRIP1 may be useful as a potential prognostic biomarker and therapeutic target in CC patients.

The cysteine proteases of parasites are vital contributors that induce parasite migration to and invasion of host tissue. In this study, we analysed the cysteine protease ATG4B of Trichinella spiralis (TsATG4B) isolated from the soluble proteins of Trichinella spiralis (T. spiralis) adult worms to ascertain its biochemical properties and functions during invasion into the intestine of the host. The 43 kDa recombinant cysteine protease ATG4B protein (rTsATG4B) consists of a conserved peptidase_C54 domain and was expressed in Escherichia coli. Gelatine zymography showed that rTsATG4B could hydrolyse gelatine and that the hydrolytic activity was prevented by the cysteine protease inhibitor E-64 (pH 5.2). Immunofluorescence assays showed that TsATG4B is expressed at different stages and is localized at the cuticles and stichosomes of worms. Far-Western blotting and confocal microscopy revealed that rTsATG4B interacts with intestinal epithelial cells (IECs) and that it was subcellularly localized to the membrane and cytoplasm in IECs. Real‑time quantitative PCR (qPCR) results indicated that the transcription level of the TsATG4B gene was the higher in 6-day-old adult worms (6 days AW) than in any other stage. An in vitro larval invasion assay verified that rTsATG4B promoted larval invasion and that invasion was inhibited when rTsATG4B was pre-incubated with E-64, whereas anti-rTsATG4B serum inhibited larval invasion in a dose-dependent manner. Collectively, these results suggested that the enzymatic activity of TsATG4B significantly influences the hydrolysis process, which is necessary for larval invasion of the host intestinal epithelium.

Coronavirus disease 2019 (COVID-19), caused by the novel human coronavirus SARS-CoV-2, is currently a major threat to public health worldwide. The viral spike protein binds the host receptor angiotensin-converting enzyme 2 (ACE2) via the receptor-binding domain (RBD), and thus is believed to be a major target to block viral entry. Both SARS-CoV-2 and SARS-CoV share this mechanism. Here we functionally analyzed the key amino acid residues located within receptor binding motif of RBD that may interact with human ACE2 and available neutralizing antibodies. The in vivo experiments showed that immunization with either the SARS-CoV RBD or SARS-CoV-2 RBD was able to induce strong clade-specific neutralizing antibodies in mice; however, the cross-neutralizing activity was much weaker, indicating that there are distinct antigenic features in the RBDs of the two viruses. This finding was confirmed with the available neutralizing monoclonal antibodies against SARS-CoV or SARS-CoV-2. It is worth noting that a newly developed SARS-CoV-2 human antibody, HA001, was able to neutralize SARS-CoV-2, but failed to recognize SARS-CoV. Moreover, the potential epitope residues of HA001 were identified as A475 and F486 in the SARS-CoV-2 RBD, representing new binding sites for neutralizing antibodies. Overall, our study has revealed the presence of different key epitopes between SARS-CoV and SARS-CoV-2, which indicates the necessity to develop new prophylactic vaccine and antibody drugs for specific control of the COVID-19 pandemic although the available agents obtained from the SARS-CoV study are unneglectable.

Alzheimer's disease (AD) is a common neurodegenerative disease, and its pathogenesis is closely related to β-amyloid (Aβ) peptide. The deposition of Aβ in the brain due to impaired Aβ clearance is considered as an important cause of AD. The decrease in Aβ clearance is closely related to the autophagy dysfunction in brains of AD patients. It is feasible to treat AD by increasing the autophagy level of cells such as microglia and neurons to accelerate Aβ clearance. In this article we explored the ability of graphene oxide (GO) to clear Aβ through activating autophagy. Our work demonstrated that GO could inhibit the mTOR signaling pathway by activating AMPK to induce the autophagy of microglial and neurons. As expected, with the improvement of autophagy ability of microglia, GO promoted microglia-mediated Aβ phagocytosis. Under the conditions of co-culture of microglia and neurons, GO induced the autophagy of microglia and neurons, especially the autophagy of microglia, thereby promoting the clearance of Aβ, and ultimately achieved the effect of protecting neurons. Moreover, GO was not only non-cytotoxic to microglia and neurons but also able to reduce the toxicity of Aβ to neurons through its clearance. These results have shown the potential of GO in treating Alzheimer's disease.

Follistatin-like 1 (FSTL1), also named transforming growth factor (TGF)-β1-inducible gene, is a secreted extracellular glycoprotein expressing widely in nervous system. Several recent studies have revealed that FSTL1 plays an essential role in neurological diseases including neuropathic pain and ischemic stroke. It proves that FSTL1 suppresses synaptic transmission by activating Na/K-ATPase in DRG neurons and inhibits neuronal apoptosis by phosphorylation AKT signaling. However, it is not clear whether FSTL1 can play a role in other type of neuron or neurodegenerative diseases. In this study, we found that the mice with Fstl1 genetic knockdown showed not only the impairments of learning and memory abilities, but also abnormal neural oscillations and synaptic plasticity in the hippocampus. Subsequently, we identified broad transcriptional changes including 55 up-regulated and 184 down-regulated genes in Fstl1 knockdown mice by RNA-Seq analysis, as well as neurotransmitter transport, synaptic transmission and disease-related genes. The expression changes of some DEGs were further validated via quantitative Realtime PCR (qRT-PCR). Further patch-clamp whole cell recording showed that Fstl1+/- mice displayed a significant decrease in glutamatergic synaptic transmission and increase in GABAergic synaptic transmission, which were consistent with the RNA-Seq analysis. Taken together, our results provide an evidence and a possibly underlying mechanism for the critical role of FSTL1 in the hippocampus on learning and memory and normal neural oscillations, suggesting that FSTL1 may plays an important role in neurodegenerative diseases related to cognitive impairments.

Corticosterone (CORT) is a kind of corticosteroid produced by cortex of adrenal glands. Hypothalamic-pituitary-adrenal (HPA) axis hyperfunction leads to excessive CORT, which is associated with depression. Few studies have investigated the role of CORT in voltage-gated ion channels and its upstream signaling pathway in central nervous system. In this study, we investigated the mechanism of excessive CORT resulting in brain impairment on voltage-gated ion channels, and its upstream signaling effectors in hippocampal CA1 neurons. The action potential (AP) and voltage-gated potassium currents were determined by using whole-cell patch-clamp. Insulin and CORT improved the neuronal excitability. Independent effects existed in transient potassium channel (IA) and delay rectifier potassium channel (IK). The inhibition of potassium currents, IA in our experiment, could increase neuronal excitability. CORT led to the excitotoxicity of hippocampal neurons via phosphatidylinositol 3 kinase (PI3K)-mediated insulin-signaling pathway. Therefore, the stimulation of excessive CORT induces excitotoxicity of hippocampal neurons and sensitivity of potassium channels via PI3K-mediated insulin-signaling pathway, which indicates one possible way of depression treatment.

Herein we present a concept in cancer where an immune response is detrimental rather than helpful. In the cancer setting, the immune system is generally considered to be helpful in curtailing the initiation and progression of tumors. In this work we show that a patient’s immune response to their tumor can, in fact, either enhance or inhibit tumor cell growth. Two closely related autoantibodies to the growth factor receptor TrkB were isolated from cancer patients’ B cells. Although highly similar in sequence, one antibody was an agonist while the other was an antagonist. The agonist antibody was shown to increase breast cancer cell growth both in vitro and in vivo, whereas the antagonist antibody inhibited growth. From a mechanistic point of view, we showed that binding of the agonist antibody to the TrkB receptor was functional in that it initiated downstream signaling identical to its natural growth factor ligand, brain-derived neurotrophic factor (BDNF). Our study shows that individual autoantibodies may play a role in cancer patients.

Aims The activation of hepatic stellate cells (HSCs) plays a central role in liver fibrosis progression. Phospholipase D (PLD) enzymes participate in multiple cellular activities. However, whether and how PLD regulates HSCs activation remain elusive. Main methods The expression of intrahepatic PLD1 and PLD2 was determined in CCl4-induced mouse liver fibrosis models by western blot and immunohistochemistry. Cell model of liver fibrogenesis was constructed using rat HSCs line (HSC-T6) treated with recombinant transforming growth factor β1 (TGFβ1). Fibrogenesis was evaluated on the aspects of proliferation, expression of pro-fibrogenic markers and migration. The effects mediated by PLD1-mTOR axis on TGFβ1-induced fibrogenesis were evaluated using HSC-T6 treated with small-molecular PLD1 inhibitors, PLD1-SiRNA, rapamycin (mTOR inhibitor) and MHY1485 (mTOR activator). Key findings Significant increase of PLD1, not PLD2 was documented in CCl4-induced cirrhotic compared to normal liver tissues. Suppression of PLD1 activities by PLD inhibitors or down-regulation of PLD1 expression in HSC-T6 could significantly restrain TGFβ1-induced fibrogenesis, as reflected by decreased cell proliferation and reduced expression of pro-fibrogenic markers. Besides, either PLD1 inhibitor or PLD1-SiRNA significantly inhibited mTOR activity of HSC-T6. Moreover, PLD1 inhibitors not only exhibited similar effects with rapamycin in TGFβ1-induced fibrogenesis, but also blunted MHY1485 enhanced cell proliferation of HSC-T6. Significance The PLD1-mTOR axis of HSCs could be therapeutically targeted in advanced liver fibrosis.

Fatty acid binding protein 4 (FABP4) plays a critical role in metabolism and inflammatory processes and therefore is a potential therapeutic target for immunometabolic diseases such as diabetes and atherosclerosis. Herein, we reported the identification of naphthalene-1-sulfonamide derivatives as novel, potent and selective FABP4 inhibitors by applying a structure-based design strategy. The binding affinities of compounds 16dk, 16do and 16du to FABP4, at the molecular level, are equivalent to or even better than that of BMS309403. The X-ray crystallography complemented by the isothermal titration calorimetry studies revealed the binding mode of this series of inhibitors and the pivotal network of ordered water molecules in the binding pocket of FABP4. Moreover, compounds 16dk and 16do showed good metabolic stabilities in liver microsomes. Further extensive in vivo study demonstrated that 16dk and 16do exhibited a dramatic improvement in glucose and lipid metabolism, by decreasing fasting blood glucose and serum lipid levels, enhancing insulin sensitivity, and ameliorating hepatic steatosis in obese diabetic (db/db) mice.