ObjectiveTo study the effect of Taikong Yangxin Pills on the metabolism of simulated weightless rats based on metabolomics and discuss the metabolism mechanism. MethodsIn the simulated space capsule environment on the ground， the rat model of simulated weightlessness was established by the tail suspension method. Rats were randomly grouped as follows： out-of-capsule control， in-capsule control， model， and high （3.0 g·kg^-1） and low （1.5 g·kg^-1） doses of Taikong Yangxin Pills， and they were administrated with corresponding drugs by gavage for 28 days. The serum levels of endogenous metabolites in rats were determined by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry （UPLC-Q-TOF/MS）. The obtained data were processed by principal component analysis （PCA） and orthogonal partial least squares-discriminant analysis （OPLS-DA） to screen for differential metabolites and potential biomarkers. MetaboAnalyst 5.0 was used for pathway enrichment analysis to explain the metabolic regulation mechanism of the drug. ResultsCompared with the out-of-capsule control group， the in-capsule control group showed elevated levels of thirteen metabolites， including 14-hydroxyhexadecanoic acid， linoleic acid， and α-linolenic acid （P<0.05）， which suggested that the space capsule environment mainly affected the metabolism of α-linolenic acid and linoleic acid in the rats. Compared with the in-capsule control group， the model group showed lowered levels of fourteen metabolites， including 4-imidazolone-5-propionic acid， isocitric acid/citric acid， and L-tyrosine （P<0.05）， which were recovered after the treatment with Taikong Yangxin pills （P<0.05）. The pathway enrichment analysis revealed that weightlessness induced by tail suspension and drug intervention mainly involved the phenylalanine， tyrosine， and tryptophan biosynthesis， tyrosine metabolism， histidine metabolism， and citric acid cycle. ConclusionThe simulated space capsule environment and simulated weightlessness induced by tail suspension can both affect the metabolism level of rats. Taikong Yangxin pills can ameliorate the metabolic abnormality in the rat model of weightlessness by regulating various amino acids and energy metabolism-related pathways.

Excavating the quantitative trait locus (QTL) associated with rice cooking quality, analyzing candidate genes, and improving cooking quality-associated traits of rice varieties by genetic breeding can effectively improve the taste of rice. In this study, we used the indica rice HZ, the japonica rice Nekken2 and 120 recombinant inbred lines (RILs) populations constructed from them as experimental materials to measure the gelatinization temperature (GT), gel consistency (GC) and amylose content (AC) of rice at the maturity stage. We combined the high-density genetic map for QTL mapping. A total of 26 QTLs associated with rice cooking quality (1 QTL associated with GT, 13 QTLs associated with GC, and 12 QTLs associated with AC) were detected, among which the highest likelihood of odd (LOD) value reached 30.24. The expression levels of candidate genes in the localization interval were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR), and it was found that the expression levels of six genes were significantly different from that in parents. It was speculated that the high expression of LOC_Os04g20270 and LOC_Os11g40100 may greatly increase the GC of rice, while the high expression of LOC_Os01g04920 and LOC_Os02g17500 and the low expression of LOC_Os03g02650 and LOC_Os05g25840 may reduce the AC. The results lay a molecular foundation for the cultivation of new high-quality rice varieties, and provide important genetic resources for revealing the molecular regulation mechanism of rice cooking quality.
Quantitative Trait Loci ; Oryza/genetics* ; Plant Breeding ; Cooking ; Genetic Association Studies

Objective To investigate the effect and mechanism of ammonium pyrrolidine dithiocarbamate(PDTC)on neuroinflammatory injury in the penumbra of traumatic brain injury(TBI)in rats.Methods Sixty Sprague Dawley rats were divided into the PDTC group,TBI group,sham operation group and control group according to the random number table method,with 15 rats in each group.Rats in the PDTC group were intraperitoneally injected with PDTC(100 mg·kg-1)at 15 minutes before surgery;while the rats in the TBI group,sham operation group,and control group were intraperitoneally injected with the same volume of double distilled water.After the cranial window of rats in the TBI group and PDTC group was created,a 2.5 g steel rod with an inner diameter of 6.0 mm was dropped freely from a height of 75 cm through a transparent polyvinyl chloride tube with an inner diameter of 7.0 mm to impact the dura mater and induce right parietal lobe contusion and laceration to establish the TBI model;rats in the sham operation group were sealed with bone wax after the cranial window creation,without any impact applied;rats in the control group were raised under normal conditions.The modified neurological severity score(mNSS)was used to evaluate the degree of neurobehavioral damage in rats in each group at 1,4 and 7 days after modeling.At 2 days after modeling,5 rats in each group were decapitated,and brain tissues were taken for hematoxylin & eosin(HE)staining,and morphological changes of the brain tissues were observed under an optical microscope.The expressions of β-amyloid precursor protein(β-APP)and glial fibrillary acidic protein(GFAP)in the brain tissues of rats in each group were detected by immunohistochemical staining.At 24 hours after modeling,5 rats in each group were decapitated,and the right injured penumbra tissues were obtained;the expressions of nuclear transcription factor-κB(NF-κB)P65,phosphorylated NF-κB P65,inhibitor of NF-κB(IκB),phosphorylated IKB,NOD-like receptor protein 3(NLRP3)and caspase-1 protein in the right injured penumbra tissue of rats in each group were detected by Western blot,and the expressions of NF-κB P65,IκB,NLRP3 and caspase-1 mRNA in the right injured penumbra tissue of rats in each group were determined by real-time quantitative polymerase chain reaction.Results At 1,4,and 7 days after modeling,the mNSS scores of rats in the TBI group were signifi-cantly higher than those in the PDTC group,control group and sham operation group.The mNSS scores of rats in the PDTC group were significantly higher than those in the control group and sham operation group(P＜0.05);there was no statistically significant difference in mNSS scores between the sham operation group and the control group(P＞0.05).The neurons and neurogliocyte of rats in the control group and the sham operation group exhibited normal morphology,without swelling and wide-ning of intercellular space.Diffuse hemorrhagic changes were observed in the brain tissues of rats in the TBI group,with different morphologies of neuronal cell body,unclear cell membrane and cytoplasm,pyknosis of cell nuclei,often triangular shape,disappearance of normal structure and nucleoli,and diffuse white blood cells and red blood cells filling the field of vision.The lesion surrounding area of rats in the PDTC group showed ischemic changes,with mild shrinkage of neuronal volume,a uniform light red color,karyopyknosis,nuclear-cytoplasmic dissociation,disappearance of normal structure and nucleoli,and localization of neuroinflammation.There was no significant expression of β-APP and GFAP in the cerebral cortex of rats in the control group and the sham operation group,while the accumulation of β-APP and GFAP in neuronal serosae and/or axons was observed in the brain tissues of rats in the TBI group and the PDTC group.Compared with the TBI group,a decrease in the number and the expression intensity of β-APP and GFAP-positive stained neuronal cells in the cerebral cortex of rats was observed in the PDTC group.The relative expression of NF-κB P65 protein in the brain tissues of rats in the sham operation group,TBI group and PDTC group was significantly higher than that in the control group,and the relative expression of NF-κB P65 protein in the brain tissues of rats in the PDTC group was significantly higher than that in the TBI group(P＜0.05).The relative expression of phosphorylated NF-κB P65 protein in the brain tissues of rats in the PDTC group and the sham operation group was significantly lower than that in the control group,the relative expression of phosphorylated NF-κB P65 protein in the brain tissues of rats in the TBI group was significantly higher than that in the control group and the sham operation group,and the relative expression of phosphorylated NF-κB P65 protein in the brain tissues of rats in the PDTC group was significantly lower than that in both TBI group and sham operation group(P＜0.05).There was no significant difference in the relative expression of IκB protein in the brain tissues of rats between the sham operation group and the control group(P＞0.05);the relative expression of IκB protein in the brain tissues of rats in the TBI group was significantly higher than that in the control group,and the relative expression of IκB protein in the brain tissues of rats in the PDTC group was significantly lower than that in the control group,sham operation group,and TBI group(P＜0.05).The relative expression of phosphorylated IκB protein in the brain tissues of rats in the TBI group was significantly lower than that in the control group and the sham operation group,and the relative expression of phosphorylated IκB protein in the brain tissues of rats in the PDTC group was significantly higher than that in the TBI group(P＜0.05).The relative expression of NLRP3 protein in the brain tissues of rats in the sham opera-tion group was significantly higher than that in the control group,the relative expression of NLRP3 protein in the brain tissues of rats in the TBI group and the PDTC group was significantly lower than that in the sham operation group and the control group,and the relative expression of NLRP3 protein in the brain tissues of rats in the TBI group was significantly lower than that in the PDTC group(P＜0.05).The relative expression of caspase-1 protein in the brain tissues of rats in the sham opera-tion group,PDTC group,and TBI group was significantly higher than that in the control group,and the relative expression of caspase-1 protein in the brain tissues of rats in the PDTC group was significantly lower than that in the TBI group(P＜0.05).The relative expression of NF-κB P65 mRNA in the brain tissues of rats in the PDTC group,TBI group,and sham operation group was significantly higher than that in the control group,the relative expression of NF-κB P65 mRNA in the brain tissues of rats in the PDTC group and TBI group was significantly higher than that in the sham operation group,and the relative expres-sion of NF-κB P65 mRNA in the brain tissues of rats in the PDTC group was significantly lower than that in the TBI group(P＜0.05).The relative expression of IκB mRNA in the brain tissues of rats in the PDTC group and TBI group were signifi-cantly higher than that in the control group,and the expression of IκB mRNA in the brain tissues of rats in the sham operation group was significantly lower than that in the control group(P＜0.05).The relative expression of IκB mRNA in the brain tis-sues of rats in the PDTC group and TBI group was significantly higher than that in the sham operation group,and the relative expression of IκB mRNA in the brain tissues of rats in the PDTC group was significantly lower than that in the TBI group(P＜0.05).The relative expression of NLRP3 mRNA in the brain tissues of rats in the PDTC group and TBI group was significantly higher than that in the control group,the relative expression of NLRP3 mRNA in the brain tissues of rats in the sham operation group was significantly lower than that in the control group,the relative expression of NLRP3 mRNA in the brain tissues of rats in the PDTC group and TBI group was significantly higher than that in the sham operation group,and the relative expression of NLRP3 mRNA in the brain tissues of rats in the PDTC group was significantly lower than that in the TBI group(P＜0.05).The relative expression of caspase-1 mRNA in the brain tissues of rats in the sham operation group,TBI group,and PDTC group was significantly lower than that in the control group,the relative expression of caspase-1 mRNA in the brain tissues of rats in the TBI group and PDTC group was significantly higher than that in the sham operation group(P＜0.05).Conclusion PDTC can effectively improve neural functional deficit score and reduce neuroinflammatory injury in TBI rats,the mechanism of which may be related to regulating mRNA and protein expression of NF-κB/NLRP3 axis-related inflammatory injury indicators and regulating downstream inflammatory factors.

Objective To predict the potential mechanism of Yifei Sanjie prescription in the treatment of lung adenocarcinoma based on network pharmacology,and to verify one of the key signal pathways,Janus protein tyrosine kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3),by cell experiments in vitro.Methods To screen the main active components and potential action targets of Yifei Sanjie prescription,with traditional Chinese medicine system pharmacological database(TCMSP).To search and retrieve the main targets of lung adenocarcinoma,with human genetic database(GeneCards)and online human Mendelian genetic database(OMIM).To obtain the intersection targets by screening and apply Wayne diagram,then analysis the topology and establish the traditional Chinese medicine-active compound-target network diagram by using of Cytoscape 3.7.2 software.To construct the protein-protein interaction(PPI)network,with the protein-protein interaction platform(STRING)and Cytoscape3.7.2 software.To analyze the functional enrichment of gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG),with the Metascape database.To carry out the molecular docking verification by using of Vina1.2.3 software.Using CCK-8 method to detect the effect of Yifei Sanjie prescription on cell activity.Using the cell scratch test to observe the effect on cell migration.And using Western blot method to test the expression of p-STAT3,STAT3,p-JAK2,JAK2 and VEGF-A.Results 94 active components,329 related drug targets and 1358 lung adenocarcinoma targets were obtained from Yifei Sanjie prescription,among which,150 of them intersected.PPI network visualization analysis shows that the potential key targets of Yifei Sanjie prescription in the treatment of lung adenocarcinoma are protein kinase B1(AKT1),β-actin(ACTB),tumor suppressor gene p53(TP53),serum albumin(ALB),caspase-3(CASP3)and vascular endothelial growth factor A(VEGFA).KEGG enrichment analysis screened 138 related signal pathways,indicating that JAK/STAT signaling pathway may play a key role in the treatment of lung adenocarcinoma with Yifei Sanjie prescription.Molecular docking results showed that quercetin,luteolin,and ursolic acid had good binding activities with JAK2 and STAT3.The cell experiment showed that compared with the blank group,Yifei Sanjie prescription could significantly inhibit the activity of A549 cells,inhibit the migration of A549 cells,and decrease the expression of p-JAK2/JAK2,p-STAT3/STAT3 and VEGF-A protein.In addition,Colivelin,an activator of JAK2/STAT3 pathway,could reverse the effect of Yifei Sanjie prescription on the expression of A549 related proteins.Conclusion Yifei Sanjie prescription has the characteristics of multi-component,multi target and multi pathway in the treatment of lung adenocarcinoma,and its mechanism may be related to the down-regulation of p-JAK2,p-STAT3 and VEGF-A protein expression,thereby inhibiting cell proliferation and migration.

Objective:To investigate the effect of sequential suture and adhesion on craniomaxillofacial skin contusion and laceration.Methods:A total of 189 patients with craniomaxillofacial skin contusion and laceration (CMFSCL) were randomly divided into three groups: 66 cases in SSA group, 63 cases in CS group and 60 cases in TS group. Operation time, visual analogue scale (VAS), Vancouver scar scale (VSS) and adverse reactions incidence were compared and analyzed between the three groups. Effect and satisfactory scale were evaluated.Results:Operation time in SSA group (10.67±1.26) min was significantly less than that in CS (18.91±1.38) min and TS group (17.96±1.43) min ( P<0.05). VAS in SSA group 24 h post-operation (3.11±1.01) was significantly lower than that in CS and TS group ( P<0.05). VSS in SSA group 6 months post-operation (1.18±0.21) was significantly lower than that in CS (3.78±1.01) ( P<0.05) and TS group (5.98±1.06) ( P<0.01). Total effective rate of SSA group (96.5%) was significantly higher than that in CS (85.7%) ( P<0.05) and TS group (56.1%) ( P<0.01); total effective rate in CS group was significantly higher than that in TS group ( P<0.05). Infection and dehiscence rates in SSA group were lower than those in CS and TS group ( P<0.01). Satisfactory rate of SSA group (99%) was significantly higher than that of CS (89.1%) and TS group (71.3%) ( P<0.05); the satisfactory rate of CS group was significantly higher than that of TS group ( P<0.05). Conclusions:Sequential suture and adhesion technique is simple and effective for craniomaxillofacial skin contusion and laceration, which is worthy of clinical promotion.

Objective:To investigate the pathological change mechanism of patients with chronic active Epstein-Barr virus infection (CAEBV).Methods:The clinical manifestations, laboratory examinations, diagnosis and treatment of one CAEBV patient in Suzhou Hongci Hematology Hospital of Jiangsu Province were retrospectively analyzed with review of related literature.Results:Combined with the results of laboratory, lymph node biopsy and bone marrow tests in different periods, the patient initially showed lymphoproliferative disease, and gradually transformed into lymphoma with the diagnosis results of EB virus-related lymphocyte clones in different periods. The patient received the corresponding treatment plan in different periods, but eventually died due to hemophagocytic syndrome and rapid progress of the disease.Conclusion:CAEBV may cause lymphocytes to gradually evolve from polyclonal to monoclonal state.

To obtain chicken CD40L protein, the cDNA was prepared from chicken splenic cells and used as a template to clone and amplify CD40L by PCR. The target gene was cloned into pFastBac vector to construct a pFastBac-chCD40L donor plasmid. Recombinant plasmid was transformed into DH10Bac and recombinant Bacmid-chCD40L was obtained. The Bacmid-chCD40L plasmid was transfected into sf9 insect cells to obtain His-chCD40L protein. In addition, the target gene was cloned into pQM01 vector to construct a pQM01-chCD40L plasmid, recombinant plasmid was transfected into HEK 293T cells to obtain Strep-chCD40L protein. The chCD40L protein was purified by affinity chromatography, and the concentration of purified chCD40L protein was determined to be 0.01 mg/mL. Primary cells were isolated from the bursal tissue of 3-week old SPF chickens, and the chCD40L protein was added to the culture medium to stimulate cells. The chCD40L could bind to CD40 on B cells as examined by Western blotting, indirect immunofluorescence assay and flow cytometry, suggesting that chCD40L protein is biologically active. We successfully obtained chicken CD40L protein of biological activity, which laid the foundation in the in vitro culture of primary B lymphocytes for the isolation and diagnosis of virulent IBDV.
Animals ; Baculoviridae/genetics* ; CD40 Ligand/genetics* ; Chickens ; Cloning, Molecular ; Genetic Vectors/genetics* ; Recombinant Proteins/genetics*

Objective:To identify important prognostic molecular markers of triple negative breast cancer (TNBC) using high throughput sequencing technology and to explore the correlation of spindle checkpoint protein BUB1B and clinicopathological features with patients′ prognosis.Methods:The clinicopathological data and prognostic information of TNBC diagnosed at the West China Hospital of Sichuan University from 2009 to 2017 were collected. Forty-seven fresh tumor samples and 139 formalin fixed paraffin-embedded samples were selected. The fresh tumor samples were subject to RNA sequencing (RNA-seq). The enrichment analysis and protein-protein interaction (PPI) analysis were performed after intersection of difference analysis between RNAseq and GEO (Gene Expression Omnibus) datasets GSE38959 and GSE65194. Kaplan-Meier plotter database was used to analyze the relationship between expression of BUB1B and prognosis. Immunohistochemical staining was used to verify its expression in TNBC and correlation with clinicopathological features and prognosis.Results:Using edgeR to perform differential expression analysis between 47 TNBC tumor tissues and 12 normal tissues, 1 559 up-regulated genes and 1 376 down-regulated genes were identified, while only 131 differentially expressed genes were overlapping with those in GSE38959 and GSE65194. Enrichment analysis was mainly enriched in cell cycle, JAK-STAT signaling pathway and p53 signaling pathway. The top 10 genes ranked by degree of association were TOP2A, BUB1B, MKI67, PLK1, RRM2, PCNA, KPNA2, SMC4, PBK and IGF1. Kaplan-Meier plotter database analysis showed that the expression of BUB1B was significantly correlated with the prognosis of TNBC [overall survival, hazard ratio (HR)=0.52, 95% CI (0.35-0.77), P=0.001; distant metastasis-free, HR=0.72, 95% CI (0.52-0.98), P=0.038]. The immunohistochemical analyses of 139 formalin fixed paraffin-embedded samples showed that the low expression of BUB1B was correlated with poor prognosis in TNBC [HR=0.41, 95% CI (0.18-0.95), P=0.024]. Conclusions:The low expression of BUB1B protein is associated with poor prognosis in TNBC patients, and the molecular mechanism related with prognosis and potential therapeutic targets need to be further studied.

Objective:To investigate the effects of tuina, treadmill running or both on the expression of factors related to gastrocnemius muscle proteins after acute muscle injury and to explore the mechanisms involved.Methods:Thirty adult male Sprague-Dawley rats were randomly divided into a control group, a natural recovery group, a tuina group, a treadmill running group and a combined treatment group, each of 6. An impactor was used to induce an acute skeletal muscle injury in the right hind legs of all of the rats except those of the control group. One day after the successful modelling, the tuina, treadmill running and combined groups were given interventions as their name implied, 5 times a week for 3 weeks. The gait of rats in each group was analyzed and the number of times the rats fell into and striken by the electrical grid was counted. The injured muscles′ cross-sectional areas (CSAs) and the diameters of muscle fibers were observed using hematoxylin and eosin staining. The expression of mTOR, p-mTOR, p70S6K, p-p70S6K and smad2/3 protein were tested using western blotting. The relative expression of myostatin (MSTN) mRNA was measured using real-time quantitative polymerase chain reactions.Results:Compared with the natural recovery group, all the other groups fell into the electrical grid significantly less often. The average CSA and wet weight of the affected gastrocnemius had increased significantly in the tuina, treadmill running and combined treatment groups, with the average CSA increases in the treadmill running and combined treatment groups significantly greater compared with the tuina group. The average relative expression of mTOR, p-mTOR, p70S6K, and p-p70S6K in the other four groups increased significantly compared with the control group, while that of Smad2/3 and MSTN decreased significantly. Compared with the natural recovery group, the average increases in the other groups were significantly greater. Compared with the tuina group, the treadmill running and combined treatment groups showed significantly better improvements, on average.Conclusions:Tuina, treadmill running and their combination all can improve recovery from skeletal muscle trauma, at least in rats. However, treadmill running and combined treatment are more effect than tuina alone.

Objective: To investigate the effect of Kamistad gel on oral ulcer healing and the expression of tumor necrosis factor α (TNF-α), interleukin 6 (IL-6) and epidermal growth factor (EGF) after oral administration in ulcer tissue of rats and to provide animal experimental data for the clinical application of Kamistad gel. Methods: The oral ulcer rat model was established by chemical cauterization. The rats were randomly divided into four groups: Kamistad group (Kamistad gel), befuxin group (befuxin gel), lidocaine group (lidocaine cream), blank control group (normal saline), with 12 rats in each group. The ulcer area of the rats in each group was measured before and 1, 3 and 5 days after treatment; at 1 day after treatment, the duration of swabbing behavior within 3 minutes of intraoral capsaicin infusion was recorded to evaluate the degree of pain; the ulcer tissue was collected at 5 days after treatment, and the histopathological changes were observed by HE staining, the expression of TNF-α, IL-6 and EGF in the ulcer tissue was detected by immunohistochemistry and ELISA. Results: At 1 day after treatment, the duration of mouth wiping induced by capsaicin was significantly shorter in the Kamistad group than in the blank control and befuxin groups (P < 0.05), but there was no significant difference between the Kamistad and lidocaine groups (P >0.05). At 5 days after treatment, the ulcer area was significantly smaller in the Kamistad group than in the blank control and lidocaine groups (P < 0.05), and there was no significant difference between the Kamistad and befuxin groups (P >0.05). At 5 days after treatment, H&E staining of the oral ulcer tissue sections showed significantly reduced levels of inflammatory cells and significantly proliferated fibroblasts and better epithelial hyperplasia in the Kamistad group compared with those in the lidocaine and blank control groups, and there were no differences between the Kamistad and befuxin groups. At 5 days after treatment, the expression levels of TNF-α, IL-6 and EGF in the ulcer tissue of rats in each group were significantly different (P < 0.05). Compared with the blank control and lidocaine groups, the expression of TNF-α and IL-6 was significantly decreased and the expression of EGF was significantly increased in the Kamistad group (P < 0.05); there were no significant differences in the expression of the above three factors between the Kamistad and befuxin groups (P > 0.05). Conclusion Kamistad gel exhibited anti-inflammatory, analgesic and healing effects on experimental oral ulcers.