ABSTRACT
ß-Sitosterol is a natural compound with demonstrated anti-cancer properties against various cancers. However, its effects on hepatocellular carcinoma (HCC) and the underlying mechanisms are not well understood. This study aims to investigate the impact of ß-sitosterol on HCC. In this study, we investigated the effects of ß-sitosterol on HCC tumour growth and metastasis using a xenograft mouse model and a range of molecular analyses, including bioinformatics, real-time PCR, western blotting, lentivirus transfection, CCK8, scratch and transwell assays. The results found that ß-sitosterol significantly inhibits HepG2 cell proliferation, migration and invasion both in vitro and in vivo. Bioinformatics analysis identifies forkhead box M1 (FOXM1) as a potential target for ß-sitosterol in HCC treatment. FOXM1 is upregulated in HCC tissues and cell lines, correlating with poor prognosis in patients. ß-Sitosterol downregulates FOXM1 expression in vitro and in vivo. FOXM1 overexpression mitigates ß-sitosterol's inhibitory effects on HepG2 cells. Additionally, ß-sitosterol suppresses epithelial-mesenchymal transition (EMT) in HepG2 cells, while FOXM1 overexpression promotes EMT. Mechanistically, ß-sitosterol inhibits Wnt/ß-catenin signalling by downregulating FOXM1, regulating target gene transcription related to HepG2 cell proliferation and metastasis. ß-Sitosterol shows promising potential as a therapeutic candidate for inhibiting HCC growth and metastasis through FOXM1 downregulation and Wnt/ß-catenin signalling inhibition.
Subject(s)
Carcinoma, Hepatocellular , Liver Neoplasms , Sitosterols , Humans , Animals , Mice , Carcinoma, Hepatocellular/pathology , Liver Neoplasms/pathology , beta Catenin/metabolism , Cell Line, Tumor , Wnt Signaling Pathway , Cell Proliferation , Cell Movement , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Forkhead Box Protein M1/geneticsABSTRACT
Shape processing, whether by seeing or touching, is pivotal to object recognition and manipulation. Although the low-level signals are initially processed by different modality-specific neural circuits, multimodal responses to object shapes have been reported along both ventral and dorsal visual pathways. To understand this transitional process, we conducted visual and haptic shape perception fMRI experiments to test basic shape features (i.e. curvature and rectilinear) across the visual pathways. Using a combination of region-of-interest-based support vector machine decoding analysis and voxel selection method, we found that the top visual-discriminative voxels in the left occipital cortex (OC) could also classify haptic shape features, and the top haptic-discriminative voxels in the left posterior parietal cortex (PPC) could also classify visual shape features. Furthermore, these voxels could decode shape features in a cross-modal manner, suggesting shared neural computation across visual and haptic modalities. In the univariate analysis, the top haptic-discriminative voxels in the left PPC showed haptic rectilinear feature preference, whereas the top visual-discriminative voxels in the left OC showed no significant shape feature preference in either of the two modalities. Together, these results suggest that mid-level shape features are represented in a modality-independent manner in both the ventral and dorsal streams.
Subject(s)
Pattern Recognition, Visual , Visual Perception , Pattern Recognition, Visual/physiology , Visual Perception/physiology , Occipital Lobe/diagnostic imaging , Touch/physiology , Parietal Lobe , Magnetic Resonance Imaging/methods , Brain MappingABSTRACT
BACKGROUND: Blepharoptosis may result in an unattractive appearance and vision problems. According to the severity of ptosis, patients may undergo correction surgery using upper eyelid retractors. The conventional incision for surgical procedures was the double-eyelid incision, potentially resulting in an obvious and unnatural scar or long-lasting edema and prolonged recovery time. OBJECTIVES: The aim of this study was to introduce a supraciliary incision as an alternative to the double-eyelid incision for blepharoptosis correction that creates a scarless, natural appearance with a quick recovery time. METHODS: From June 2019 to June 2021, 32 patients (36 eyelids) underwent blepharoptosis correction through a supraciliary incision. MRD1, the height of the eyelid fissure, and the patient's satisfaction with the shape and scar as well as postoperative complications (eyelid insufficiency, conjunctival prolapse, inadequate correction of ptosis, and excessive correction of ptosis). RESULTS: All 32 patients (36 eyelids) were followed up for 6 to 18 months, with an average follow-up of 11.6 months. The postoperative satisfaction rate was 96.43%. There was no overcorrection, but one patient (1 eyelid, 2.8%) was under correction that required secondary correction. One patient (1 eyelid, 2.8%) experienced conjunctival prolapse. Sixteen patients showed lagophthalmos early after surgery, in which one patient experienced early-stage keratitis and completely recovered within two months. CONCLUSION: Blepharoptosis correction via supraciliary incision allows for broader indications and fewer surgical scars without disrupting eyelid integrity, resulting in quick recovery after surgery. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .
Subject(s)
Blepharoplasty , Blepharoptosis , Humans , Asian People , Blepharoplasty/methods , Blepharoptosis/surgery , Cicatrix/surgery , Cohort Studies , Esthetics , Patient Satisfaction , Retrospective Studies , Treatment Outcome , Postoperative ComplicationsABSTRACT
Cumulative evidence indicates the important role of Nur77 in organ fibrogenesis. However, the role of Nur77 in hepatitis B virus (HBV)-related liver fibrosis (LF) remains unclear. Cells were transfected with the microRNA mimic miRNA-506-3p or inhibitor, and pcDNA3.1-Nur77 or Nur77 guide RNA. Exosomes were isolated from HBV-infected HepG2-sodium taurocholate cotransporting polypeptide cells. The levels of miR-506-3p, Nur77, and LF-related genes and proteins were detected by quantitative polymerase chain reaction (qPCR) and western blot analysis, respectively. The pathology of the liver from HBV-infected patients was examined using hematoxylin-eosin and Masson's staining. The expression of Nur77 in liver tissue was determined by immunohistochemistry, and the LF score was assessed using the METAVIR system. The relationship between miR-506-3p/Nur77 and LF score was analyzed by correlation analysis. HBV infection downregulated miR-506-3p expression and upregulated Nur77 levels in hepatocytes. Exosomes from HBV-infected hepatocytes also displayed decreased gene expression of miR-506-3p and increased expressions of Nur77- and LF-related genes in stellate cells compared with exosomes from hepatocytes with mock infection. These changes were reversed by Nur77 guide RNA. Nur77 expression in liver tissue was strongly correlated with LF, whereas serum miR-506-3p was strongly negatively correlated with LF. Exosomes from HBV-infected hepatocytes activate stellate cells and aggravate LF through the miR-506-3p/Nur77 pathway. These exosomes may be the basis of a promising therapeutic strategy.
ABSTRACT
INTRODUCTION: The increased migration of vascular smooth muscle cells (VSMCs) is an essential pathological factor in the early development of atherosclerosis. Beta-sitosterol (BS), a natural phytosterol abundant in plant seeds, exhibits various bioactivities, including cardioprotective effects. However, its effects on VSMC migration and underlying mechanisms remain to be explored. METHOD AND RESULT: BS inhibited the proliferation and migration of angiotensin II-induced A7r5 cells and reduced intracellular oxidative stress. Targets related to VSMC migration and the targets of BS were screened, cross-referenced, and analyzed by network pharmacology combined with molecular docking technology. The identified targets were verified at the protein and gene levels using Western blotting and quantitative PCR, respectively. BS was observed to activate peroxisome proliferator-activated receptor-γ (PPARG) and adenosine 5'-monophosphate-activated protein kinase (AMPK) and negatively regulate mammalian target of rapamycin (mTOR) expression. Furthermore, a PPARG inhibitor reversed the BS-induced activation of AMPK and mTOR. CONCLUSION: This study indicated that regulation of the PPARG/AMPK/mTOR signaling pathway could potentially contribute to the inhibitory effects of BS on angiotensin II-induced VSMC migration.
Subject(s)
Muscle, Smooth, Vascular , PPAR gamma , AMP-Activated Protein Kinases/metabolism , Angiotensin II/pharmacology , Cell Movement , Cell Proliferation , Molecular Docking Simulation , Myocytes, Smooth Muscle/metabolism , PPAR gamma/genetics , PPAR gamma/metabolism , Sitosterols , TOR Serine-Threonine Kinases/metabolismABSTRACT
Background: Porcine deltacoronavirus (PDCoV) is a newly discovered porcine intestinal pathogenic coronavirus with a single-stranded positive-sense RNA genome and an envelope. PDCoV infects pigs of different ages and causes acute diarrhea and vomiting in newborn piglets. In severe cases, infection leads to dehydration, exhaustion, and death in sick piglets, entailing great economic losses on pig farms. The clinical symptoms of PDCoV infection are very similar to those of other porcine enteroviruses. Although it is difficult to distinguish these viral infections without testing, monitoring PDCoV is very important because it can spread in populations. The most commonly used methods for the detection of PDCoV is qPCR, which is time-consuming and require skilled personnel and equipment. Many farms cannot meet the conditions required for detection. Therefore, it is necessary to establish a faster and more convenient method for detecting PDCoV. Aims: To establish a rapid and convenient detection method for PDCoV by combining RPA (Recombinase Polymerase Isothermal Amplification) with CRISPR/Cas13a. Methods: Specific RPA primers and crRNA for PDCoV were designed, and the nucleic acids in the samples were amplified with RPA. Fluorescent CRISPR/Cas13a detection was performed. We evaluated the sensitivity and specificity of the RPA-CRISPR/Cas13a assay using qPCR as the control method. Results: CRISPR/Cas13a-assisted detection was completed within 90 min. The minimum detection limit of PDCoV was 5.7 × 101 copies/µL. A specificity analysis showed that the assay did not cross-react with three other porcine enteroviruses. Conclusion: The RPA-CRISPR/Cas13a method has the advantages of high sensitivity, strong specificity, fast response, and readily accessible results, and can be used for the detection of PDCoV.
ABSTRACT
Background and aim: Transmissible gastroenteritis virus (TGEV) is a highly contagious gastrointestinal virus that causes diarrhea, vomiting, anorexia, dehydration, and weight loss in piglets. In clinical practice, it often occurs in mixed infections with other pathogens, and is therefore difficult to diagnose and prevent. It mainly harms piglets of about 2 weeks old, causing huge losses on farms. The clinical confirmation of TGEV usually requires a laboratory diagnosis, but traditional PCR and immunofluorescence assays have some limitations. Moreover, most farms in China are ill-equipped to accurately diagnose the disease. Therefore, a new detection method with high sensitivity and specificity and less dependence on instrumentation is required. Methods: We used recombinase polymerase amplification (RPA), combined with the nuclease characteristics of the activated Cas13a protein to establish a visual CRISPR-Cas13a-assisted detection method for TGEV by adding a reporter RNA with fluorescent and quenching moieties to the system. Result: We selected the optimal RPA primer and best CRISPR RNA (crRNA). The reaction system was optimized and its repeatability, specificity, and sensitivity verified. The TGEV detection system did not cross-react with other common diarrhea viruses, and its detection limit was 101 copies, which is similar with the sensitivity of qPCR. We successfully established an RPA-CRISPR-Cas13a-assisted detection method, and used this detection system to analyze 123 pig blood samples. qPCR was used as the gold standard method. The sensitivity, specificity, positive coincidence rate, and negative coincidence rate of the new method were 100, 98.93, 96.66, and 100%, respectively.
ABSTRACT
Hepatocellular carcinoma (HCC) is highly refractory. ß-Sitosterol has been reported to suppress proliferation and migration as well as interfere with cell metabolism in tumors. However, there is limited information on the effects of ß-sitosterol on HCC. Herein, we used a xenograft mouse model to investigate the effects of ß-sitosterol on HCC tumor growth. The molecular mechanism was elucidated using quantitative real-time PCR, western blotting, lentiviral transfection, CCK8, scratch, Transwell, and Ad-mCherry-GFP-LC3B assays. The results showed that HepG2 cells highly expressed complement C5a receptor 1. ß-Sitosterol antagonized complement component 5a and exerted inhibitory effects on the proliferation and migration of HepG2 cells. The inhibitory effect of ß-sitosterol was reversed by the knockdown of complement C5a receptor 1. Bioinformatics analysis suggested alpha fetoprotein (AFP) as a downstream factor of complement C5a receptor 1. ß-Sitosterol inhibited AFP expression, which was reversed by complement C5a receptor 1 knockdown. The inhibitory effects of ß-sitosterol on cell proliferation and migration were weakened by AFP overexpression. Furthermore, ß-sitosterol induced autophagy in HepG2 cells, which was reversed by complement C5a receptor 1 knockdown and AFP overexpression. Blockade of autophagy by 3-MA attenuated ß-sitosterol inhibition of proliferation and migration in HepG2 cells. Moreover, ß-sitosterol inhibited HCC progression in vivo. Our findings demonstrate that ß-sitosterol inhibits HCC advancement by activating autophagy through the complement C5a receptor 1/AFP axis. These findings recommend ß-sitosterol as a promising therapy for HCC.
Subject(s)
Carcinoma, Hepatocellular , Liver Neoplasms , Humans , Animals , Mice , Carcinoma, Hepatocellular/drug therapy , alpha-Fetoproteins , Liver Neoplasms/drug therapy , Autophagy , Complement C5a , Disease Models, AnimalABSTRACT
Objective: To explore the effects of ß-sitosterol on VSMC proliferation. Materials and Methods: A7r5 cells were pretreated with 2 µM angiotensin II (Ang II) for 24 hr to establish an excessive VSMC proliferation model, followed by treatment with ß-sitosterol for 24 hr. Cells were divided into five groups: control, Ang II, and Ang II + ß-sitosterol (2, 4, 8 µM). CCK-8 assay, flow cytometry, and Ad-mCherry-GFP-LC3B assay analyzed cell proliferation, cell cycle, cell apoptosis, and autophagic flux. Additionally, the expression of proteins was detected by the western blotting. Results: ß-Sitosterol effectively inhibited Ang II-induced A7r5 cell proliferation (IC50 : 6.841 µM at 24 hr). It achieved this by arresting cell cycle progression, promoting apoptosis, inhibiting autophagy, and suppressing the contractile-synthetic phenotypic switch. Mechanistically, ß-sitosterol downregulated PCNA, Cyclin D1, and Bcl-2, while upregulating pro-caspase 3, cleaved-caspase 3, and Bax to induce cell cycle arrest and apoptosis. Additionally, it suppressed the contractile-synthetic phenotypic transformation by downregulating OPN and upregulating α-SMA. The Ad-mCherry-GFP-LC3B Assay and western blotting revealed ß-sitosterol's autophagy inhibitory effects by downregulating LC3, ULK1, and Beclin-1 while upregulating P62 expression. Discussion and Conclusion. This study found for the first time that ß-sitosterol could inhibit the proliferation of A7r5 cells induced by Ang II. ß-Sitosterol treatment may be recommended as a therapeutic strategy to prevent the cardiovascular diseases.
Subject(s)
Angiotensin II , Myocytes, Smooth Muscle , Angiotensin II/pharmacology , Angiotensin II/metabolism , Caspase 3/metabolism , Caspase 3/pharmacology , Cell Proliferation , Cells, CulturedABSTRACT
Sepsis-induced acute kidney injury (SI-AKI) causes renal dysfunction and has a high mortality rate. Protein arginine methyltransferase-1 (PRMT1) is a key regulator of renal insufficiency. In the present study, we explored the potential involvement of PRMT1 in SI-AKI. A murine model of SI-AKI was induced by cecal ligation and perforation. The expression and localization of PRMT1 and molecules involved in the transforming growth factor (TGF)-ß1/Smad3 and interleukin (IL)-6/signal transducer and activator of transcription 3 (STAT3) signaling pathways were detected in mouse kidney tissues by western blot analysis, immunofluorescence, and immunohistochemistry. The association of PRMT1 with downstream molecules of the TGF-ß1/Smad3 and IL-6/STAT3 signaling pathways was further verified in vitro in mouse renal tubular epithelial cells. Cecal ligation and perforation caused epithelial-mesenchymal transition, apoptosis, and inflammation in renal tissues, and this was alleviated by inhibition of PRMT1. Inhibition of PRMT1 in SI-AKI mice decreased the expression of TGF-ß1 and phosphorylation of Smad3 in the renal cortex, and downregulated the expression of soluble IL-6R and phosphorylation of STAT3 in the medulla. Knockdown of PRMT1 in mouse renal tubular epithelial cells restricted the expression of Cox-2, E-cadherin, Pro-caspase3, and phosphorylated Smad3 (involved in the TGF-ß1-mediated signaling pathway), and also blocked IL-6/soluble IL-6R, inducing the expression of Cox-2 and phosphorylated-STAT3. In conclusion, our findings suggest that inhibition of PRMT1 mitigates SI-AKI by inactivating the TGF-ß1/Smad3 pathway in the cortex and the IL-6/STAT3 pathway in the medulla. Our findings may aid in the identification of potential therapeutic target molecules for SI-AKI.
Subject(s)
Acute Kidney Injury , Sepsis , Mice , Animals , Transforming Growth Factor beta1/metabolism , Interleukin-6/metabolism , Cyclooxygenase 2/metabolism , Signal Transduction , Sepsis/complicationsABSTRACT
Introduction: Chimeric antigen receptors (CARs) can redirect T cells against antigen-expressing tumors, and each component plays an important role in the function and anti-tumor efficacy. It has been reported that using human sequences or a low affinity of CAR single-chain variable fragments (scFvs) in the CAR binding domains is a potential way to enhance the function of CAR-T cells. However, it remains largely unknown how a lower affinity of CARs using humanized scFvs affects the function of CAR-T cells until recently. Methods: We used different humanized anti-HER2 antibodies as the extracellular domain of CARs and further constructed a series of the CAR-T cells with different affinity. Results: We have observed that moderately reducing the affinity of CARs (light chain variable domain (VL)-based CAR-T) could maintain the anti-tumor efficacy, and improved the safety of CAR therapy both in vitro and in vivo compared with high-affinity CAR-T cells. Moreover, T cells expressing the VL domain only antibody exhibited long-lasting tumor elimination capability after multiple challenges in vitro, longer persistence and lower cytokine levels in vivo. Discussion: Our findings provide an alternative option for CAR-T optimization with the potential to widen the use of CAR T cells.
Subject(s)
Neoplasms , Receptors, Chimeric Antigen , Single-Chain Antibodies , Single-Domain Antibodies , Humans , Receptors, Chimeric Antigen/genetics , Receptors, Chimeric Antigen/metabolism , Single-Domain Antibodies/genetics , Single-Domain Antibodies/metabolism , T-LymphocytesABSTRACT
Blastocystis sp. is an important gastrointestinal parasite with global distribution, prevalent in humans, farmed animals, and wildlife. Therefore, this study aimed to investigate the prevalence and genetic diversity of Blastocystis sp. in Asiatic brush-tailed porcupines (Atherurus macrourus), bamboo rats (Rhizomys pruinosus), and masked palm civets (Paguma larvata) in Hainan Province, China. A total of 900 fecal samples were collected from three farmed animal species including 257 porcupines, 360 rats, and 283 civets. Genomic DNA was extracted from each fecal sample and Blastocystis sp. was detected by PCR at the small subunit ribosomal RNA (SSU rRNA) gene. A phylogenetic tree was constructed using the maximum likelihood method. Blastocystis sp. was detected in 47 (5.2%) fecal samples: 12 (4.7%) Asiatic brush-tailed porcupines, 8 (2.2%) bamboo rats, and 27 (9.5%) masked palm civets. Three known Blastocystis sp. subtypes, including ST1, ST4, ST5, and one unnamed subtype (unST), were found in one, 19, 26, and one animal, respectively. Subtypes ST4 and unST were detected in porcupines, ST4 in rats, and ST1 and ST5 in civets. Our results suggest that the three farmed animal species reported in this study could serve as reservoirs for potentially zoonotic Blastocystis sp. subtypes and transmit this parasite to humans, other farmed animals, and wildlife.
Title: Prévalence et répartition des sous-types de Blastocystis chez les athérures à longue queue (Atherurus macrourus), les rats des bambous (Rhizomys pruinosus) et les civettes masquées (Paguma larvata) élevés en Chine dans le Hainan. Abstract: Blastocystis sp. est un parasite gastro-intestinal important avec une distribution mondiale, répandu chez les humains, les animaux d'élevage et la faune. Par conséquent, cette étude visait à étudier la prévalence et la diversité génétique de Blastocystis sp. chez les athérures à longue queue (Atherurus macrourus), les rats des bambous (Rhizomys pruinosus) et les civettes masquées (Paguma larvata) dans la province de Hainan, en Chine. Au total, 900 échantillons fécaux ont été collectés sur ces trois espèces animales d'élevage dont 257 athérures, 360 rats et 283 civettes. L'ADN génomique a été extrait de chaque échantillon fécal et Blastocystis sp. a été détecté par PCR au niveau du gène de la petite sous-unité de l'ARN ribosomal. Un arbre phylogénétique a été construit en utilisant la méthode du maximum de vraisemblance. Blastocystis sp. a été détecté dans 47 (5,2 %) échantillons fécaux : 12 (4,7 %) athérures, 8 (2,2 %) rats et 27 (9,5 %) civettes. Trois sous-types de Blastocystis sp., dont ST1, ST4, ST5 et un sous-type sans nom (unST), ont été trouvés respectivement chez 1, 19, 26 et 1 animal. Les sous-types ST4 et unST ont été détectés chez les athérures, ST4 chez les rats et ST1 et ST5 chez les civettes. Nos résultats suggèrent que les trois espèces animales d'élevage concernées par cette étude pourraient servir de réservoirs à des sous-types potentiellement zoonotiques de Blastocystis sp. et transmettre ce parasite aux humains, à d'autres animaux d'élevage et à la faune.
Subject(s)
Blastocystis Infections , Blastocystis , Porcupines , Animals , Humans , Blastocystis/genetics , Blastocystis Infections/epidemiology , Viverridae , Prevalence , Phylogeny , Genetic Variation , Animals, Wild , China/epidemiology , Feces/parasitologyABSTRACT
Background: Atherosclerosis (AS) is a common chronic vascular inflammatory disease and one of the main causes of cardiovascular/cerebrovascular diseases (CVDs). Autophagy-related genes (ARGs) play a crucial part in pathophysiological processes of AS. However, the expression profile of ARGs has rarely been adopted to explore the relationship between autophagy and AS. Therefore, using the expression profile of ARGs to explore the relationship between autophagy and AS may provide new insights for the treatment of CVDs. Methods: The differentially expressed ARGs of the GSE57691 dataset were obtained from the Human Autophagy Database (HADb) and the Gene Expression Omnibus (GEO) database, and the GSE57691 dataset contains 9 aortic atheroma tissues and 10 normal aortic tissues. The differentially expressed ARGs of the GSE57691 dataset were analyzed by protein-protein interaction (PPI), gene ontology analysis (GO), and Kyoto Encyclopedia of Genes and Genomes analysis (KEGG) and were chosen to explore related miRNAs/transcriptional factors. Results: The GSE57691 dataset had a total of 41 differentially expressed ARGs. The GO analysis results revealed that ARGs were mainly enriched in autophagy, autophagosome, and protein serine/threonine kinase activity. KEGG analysis results showed that ARGs were mainly enriched in autophagy-animal and longevity regulating signaling pathways. Expressions of ATG5, MAP1LC3B, MAPK3, MAPK8, and RB1CC1 were regarded as focus in the PPI regulatory networks. Furthermore, 11 related miRNAs and 6 related transcription factors were obtained by miRNAs/transcription factor target network analysis. Conclusions: Autophagy and ARGs may play a vital role in regulating the pathophysiology of AS.
Subject(s)
Atherosclerosis , MicroRNAs , Animals , Atherosclerosis/genetics , Autophagy/genetics , Gene Expression Profiling , Humans , MicroRNAs/genetics , Protein Interaction Maps/geneticsABSTRACT
Autophagy is essential to vessel homeostasis and function in the cardiovascular system. Ligustilide (LIG) is one of the main active ingredients extracted from traditional Chinese medicines, such as Ligusticum chuanxiong, Angelica, and other umbelliferous plants, and reported to have cardiovascular protective effects. In this study, we explore the effects and the potential mechanism of ligustilide on the Ang II-induced autophagy in A7r5 cells. Our results showed that ligustilide inhibited the Ang II-induced autophagy in A7r5 cells and down regulated the expression of autophagy-related proteins LC3, ULK1, and Beclin-1. Ligustilide exerted a protective effect on the reduction of the concentrations of reactive oxygen species and Ca2+ and upregulated the nitric oxide concentration in A7r5 cells with Ang II-induced autophagy. Additionally, the analyses of network pharmacological targets and potential signal pathways indicated that the target of ligustilide to regulate autophagy was related to the Akt/mTOR signaling pathway. Furthermore, ligustilide could upregulate the expression of p-Akt and p-mTOR and inhibit the expression of LC3II in A7r5 cells with Ang II-induced autophagy. These findings showed that ligustilide inhibited the autophagic flux in A7r5 cells induced by Ang II via the activation of the Akt/mTOR signaling pathway.
Subject(s)
4-Butyrolactone/analogs & derivatives , Angiotensin II/metabolism , Autophagy/drug effects , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction/drug effects , TOR Serine-Threonine Kinases/metabolism , 4-Butyrolactone/pharmacology , Angiotensin II/toxicity , Animals , Autophagy/genetics , Autophagy-Related Protein-1 Homolog/metabolism , Beclin-1/metabolism , Calcium/metabolism , Cell Line , Cell Survival/drug effects , Down-Regulation/drug effects , Microtubule-Associated Proteins/metabolism , Nitric Oxide/genetics , Nitric Oxide/metabolism , Protein Interaction Maps , Rats , Reactive Oxygen Species/metabolism , Up-Regulation/drug effectsABSTRACT
Antibacterial textiles, which effectively inhibit bacterial breeding and resist pathogenic diseases, have wide applications in medicine, hygiene, and related fields. However, traditional antibacterial textiles exhibit significant limitations, such as poor antibacterial durability and contamination during preparation. In this work, nanofiber yarn loaded with a high-efficiency antibacterial agent was prepared using electrospinning technology. Polyethyleneimine (PEI) was introduced as a solubilizing material to functionalize graphene oxide (GO) to form GO-PEI composites. A facile microwave heating method was used to synthesize GO-PEI and silver nanoparticles (AgNPs). A multi-needle conjugated electrospinning device was used to blend the nanofibers with the GO-PEI-Ag composite to form a nanofiber core-spun yarn. The antibacterial agent was firmly fixed on the fiber to prevent easy removal. A uniformly oriented yarn structure and internal morphology were observed, and the antibacterial activity of the fabric was measured. The antibacterial rate of the fabric was over 99.99%for both Escherichia coli and Staphylococcus aureus. After ten washes, the antibacterial rate remained above 99.99%. Thus, nanofiber fabric from electrospinning displays high antibacterial activity and excellent durability, thereby providing a feasible methodology for future production of antibacterial textiles.
Subject(s)
Metal Nanoparticles , Nanocomposites , Nanofibers , Anti-Bacterial Agents/pharmacology , Graphite , Silver/pharmacology , TextilesABSTRACT
Objective To make a site-directed mutation of streptavidin (SA) to reduce its affinity to biotin, to prepare a streptavidin mutant agarose gel column, and to find the optimal elution conditions to purify the biotin-modified protein. Methods The streptavidin mutant protein expressed in prokaryotes was purified using Ni-NTA resin, and the purified SA mutant protein was coupled with CNBr-activated BestaroseTM 4FF agarose gel to prepare the column. The biotin-modified protein was incubated with the gel column, and was eluted and recovered using NaOH at various concentrations. The purity of protein was detected by Western blotting and Coomassie brilliant blue staining. Results Five SA mutants were expressed and purified, and gel columns were successfully prepared. The results of ELISA showed that the affinity of SA-mutant to biotin-modified protein was lower than that of wild type SA. By purifying the biotin-modified protein with different gel columns, we screened out the SA mutant M2 gel column that could purify the biotin-modified protein with 10 mmol/L NaOH. And the M2 gel column could be used to re-purify biotin-modified protein. Conclusion The SA mutant agarose gel column has been successfully prepared, which provides a feasible and reliable option for the purification of biotin-modified protein.
Subject(s)
Streptavidin/chemistry , Biotin , Blotting, Western , ProteinsABSTRACT
Objective To express the humanized immunoapoptotic protein with a modified cleavage site in mammalian cell expression system and verify its killing effect on human epidermal growth factor receptor 2 (HER2)-positve tumor cells. Methods Kozak sequence, the humanized anti-HER2 single chain antibody P1h3, Fc tag, cathepsin D cleavage site D1 and pro-apoptotic effector tBid gene were linked in sequence and cloned into the eukaryotic expression vector pLVX-EGFP. Cell culture supernatant was harvested and subjected to Western blot analysis to detect the secretory expression of the recombinant plasmid after its transfection into HEK293F cells. Killing effect of the recombinant plasmid on HER2-positive PC9 tumor cells was evaluated by cell counting and luciferase activity assay. Results The eukaryotic expression vector pLVX-P1h3-Fc-D1-tBid-EGFP was successfully constructed and its expression was confirmed by Western blot analysis. The humanized immunoapoptotic proteins significantly induced the death of HER2-positive PC9 cells. Conclusion The modified humanized immunoapoptotic proteins obtained by mammalian cell expression system could specifically kill HER2-positive tumor cells.
Subject(s)
Apoptosis , Receptor, ErbB-2/immunology , Single-Chain Antibodies/pharmacology , Cell Line, Tumor , HEK293 Cells , Humans , Plasmids , TransfectionABSTRACT
Vascular smooth muscle cells (VSMCs) excessive migration, a basic change of pathological intimal thickening, can lead to serious cardiovascular diseases such as atherosclerosis, myocardial infarction, and stroke. Ligustilide (LIG), the main active ingredient of angelica volatile oil, has been demonstrated to exert protective effects on the cardiovascular and cerebrovascular, circulatory system, and immune function. However, whether it protects against intimal thickening and VSMCs excessive migration and its underlying mechanism remains largely unknown. The aim of this study is to investigate the effect of LIG on VSMCs migration and its underlying mechanism. The protective effect of LIG on VSMCs excessive migration was assessed using an atherosclerotic spontaneously hypertensive rat model and an angiotensin II (AngII)-induced VSMCs migration model. The results showed that LIG exerted a protective effect against pathological intimal thickening as demonstrated by decreasing VSMCs migration in vivo and in vitro. In vivo, intimal thickening and VSMCs migration were inhibited and LIG performed a suppressive effect on the expression of c-Myc protein while enhanced phenotypic transformation related proteins α-SMA expression. Meanwhile, the administration of LIG significantly lowered the blood pressure and blood lipids level in atherosclerotic spontaneously hypertensive rats. In vitro, LIG suppressed AngII-induced VSMCs migration and downregulated the expression of migration related protein c-Myc, MMP2, ROCK1, ROCK2, p-JNK, and JNK. These findings suggested the protective effect of LIG on VSMCs migration was associated with the decrement of c-Myc/MMP2 signaling pathway and ROCK-JNK signaling pathway. Thus, LIG may serve as a novel therapeutic agent for preventing cardiovascular disease.
Subject(s)
4-Butyrolactone/analogs & derivatives , Atherosclerosis/drug therapy , Atherosclerosis/physiopathology , MAP Kinase Signaling System/drug effects , Muscle, Smooth, Vascular/drug effects , 4-Butyrolactone/administration & dosage , Animals , Atherosclerosis/genetics , Atherosclerosis/metabolism , Blood Pressure/drug effects , Cell Movement/drug effects , Cell Proliferation/drug effects , Cells, Cultured , Humans , Male , Matrix Metalloproteinase 2/genetics , Muscle, Smooth, Vascular/metabolism , Myocytes, Smooth Muscle/cytology , Myocytes, Smooth Muscle/metabolism , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Rats , Rats, Inbred SHR , Rats, Sprague-Dawley , rho-Associated Kinases/genetics , rho-Associated Kinases/metabolismABSTRACT
Pollution by atmospheric particulate matter (PM) is a serious threat to human health, and traditional window screens are unable to intercept fine particles. Therefore, it is urgent to develop anti-haze window screens to effectively protect the public. We prepared a polyurethane/silicon nitride (PU/Si3N4) electret nanofiber membrane with a minimum diameter (350â¯nm) and narrower diameter distribution than those of PU-Boehmite, PU-SiO2, and PU-TiO2 membranes by electrospinning. The PU-Si3N4 fiber membrane has good mechanical properties with stress of 18.05â¯MPa and strain of 170%. Moreover, PU/Si3N4 exhibits a maximum current peak (2.63â¯pA) at 90⯰C. Importantly, PU/Si3N4 membranes applied to window screens still maintain high filtration efficiency (79.36%) and low resistance (25â¯Pa) to air flow at a low areal density of 1.22â¯g/m2. Furthermore, they show good light transmittance (40%) and air permeability (46.42â¯mmâ¯s-1). Therefore, PU/Si3N4 electret nanofiber window screens have broad application prospects.
ABSTRACT
PURPOSE: To evaluate the effect of dexmedetomidine on emergence agitation after oral and maxillofacial surgery. METHODS: Fifty five patients who went into recovery room after oral and maxillofacial surgery were randomly divided into 2 groups: dexmedetomidine group (n=28) and control group (n=27). Patients in dexmedetomidine group were assigned to receive intravenous dexmedetomidine at a dose of 0.3 µg/kg when they came into recovery room. Patients in control group were assigned to receive intravenous normal saline. Emergence agitation was assessed and extubation time after operation was recorded. Mean arterial pressure, heart rate, arterial oxygen saturation, Ramsay scale were recorded at the time point of entering the recovery room instantly(T0) and 5 minutes(T1), 15 minutes(T2), 30 minutes(T3), 60 minutes(T4), 120 minutes(T5) after the patient came into recovery room. Statistical analysis was performed using SAS 9.1 software package. RESULTS: The incidence of emergence agitation was significantly lower in the dexmedetomidine group (18%) than in the control group (70%) (P<0.05). The Ramsay scale was significantly higher in dexmedetomidine group than in the control group at the time point of T1, T2, T3, T4 (P<0.05). The heart rate was significantly lower in dexmedetomidine group than in the control group at the time point of T1, T2, T3, T4, T5 (P<0.05). Mean arterial pressure was significantly lower in dexmedetomidine group than in the control group at the time point of T2, T3 (P<0.05). There was no significant difference on extubation time between 2 groups. There was no postoperative respiratory depression in 2 groups. CONCLUSIONS: Intravenous dexmedetomidine at a dose of 0.3 µg/kg can reduce emergence agitation after oral and maxillofacial surgery with safety and efficacy.