Objective To investigate the morphologic characteristics of neuron-like cells induced from the rat mesenchymal stem cells(MSCs). Methods The rat MSCs from bone marrow were cultured by being adhered to the culture dish.The thirdgeneration of the MSCs were induced by the neonate rat brain homogenate supernatant for 48 hours.The morphological characteristics and the ultrastructures of both non-induced and induced MSCs were observed under the inverted microscope and the electron microscope.The property of the induced cells was identified by using immunocytochemical method. Results Under the inverted microscope MSCs showed spindle or polygon-shaped cell bodies and the nuclei with one or two nucleoli were located in the middle of the cells.After the inducement the cells appeared neuron-like with axonlike and dendrite-like processes.The neuron-like cells were neuron special endolase(NSE),neurofilament protein(NF) positive and glial filbrillary acidic protein(GFAP) negative by irnmunocytochemical staining.Under the electron microscope the MSCs had plentiful cytoplasm and organelles and had an obvious nuclei containing a nucleoli.There were a lot of microvilli on the surfaces of the MSCs.The neuron-like cells had plentiful cytoplasm and irregular nuclei with one to three nucleoli.There were also plenty of microvilli on the cell surfaces.Conclusion MSCs are one kind of multipotent stem cells and can differentiate neurons with matured organelles induced by the neonate rat brain homogenate supernatant.

Abstract: In order to investigate immune protection against swine-origin influenza virus (S-OIV) A H1N1, the helper-dependent adenovirus vector (HDAd) system was exploited to construct recombinant HDAd encoding hemagglutinin (HA). The HA gene was synthesized and cloned to the HDAd backbone. Then, the HDAd/HA DNA molecules were transfected into 293Cre4 cells with calcium phosphate. The cells were infected by helper virus 16 hours after the transfection. The 293Cre4 cells were coinfected with HDAd/HA and the helper virus for large-scale preparation of HDAd/HA. The HDAd/HA was obtained and purified twice with CsCI density ultracentrifugation and observed morphologically under transmission electron microscope, and the expression of HA protein was analyzed with RTPCR. Recombinant HDAd/HA expressing HA protein was successfully constructed which could pave the way for in vivo investigation on immunogenicity and efficacy against S-OIV A H1N1 infection.
Adenoviridae ; Cell Line ; Cloning, Molecular ; Genetic Vectors ; Helper Viruses ; Hemagglutinin Glycoproteins, Influenza Virus ; biosynthesis ; Humans ; Influenza A Virus, H1N1 Subtype

Objective To investigate the pathogenic mechanism of Campylobacter jejuni(C.jejuni) associated with Guillain-Barré syndrome(GBS) and provide strategy for gene modification, the cstⅡ gene from 8 GBS-associated C.jejuni strains were compared with that from 3 GBS-unrelated C.jejuni strains, getting the base and amino acid mutations, the changes of secondary structures and finding the region which may be responsible for the pathogenicity of C.jejuni inducing GBS. Methods Three GBS-associated C.jejuni strains isolated from stools of GBS patients in north China were selected and cultured, which has been confirmed as GBS-associated by animal model. After sequencing the genome of them, the nucleotide sequences of cstⅡ gene were got through sequence alignment. The nucleotide sequences and deduced amini acid sequences of 3 GBS-associated cstⅡ genes were compared with that from 3 GBS-unrelated C.jejuni strains through bioinformatics software, getting the base and amino acid mutations, the changes of secondary structures. Other 5 GBS-associated cstⅡ genes were also aligned to know whether the differences we got above makes sense. In this way the genetic differences between two kinds of C.jejuni strains may be found and speculating the gene region related to the pathogenicity of GBS became possible. Results The cstⅡgene of 3 GBS-associated C.jejuni strains were all composed of 876 base pairs. Compared with GBS-unrelated C.jejuni strains, there were 9 consistent mutation sites in cstⅡ gene of LL and QYT stains, leading to 3 consistent amino acid mutation. The amino acid mutation of 114 and 182 sites in LL and QYT stains existed in other 5 GBS-associated C.jejuni strains. The sole amino acid mutation of ZHX strain -169 site, located near the 182 site. The seventh α-helix(165-180 region)of the secondary structure of the amino acid sequence from GBS-associated strains were shorter than that from GBS-unrelated strains, and the shorter regions were opened to form β-sheet or coli, which also existed in other GBS-related strains in this study.75% of the GBS-associated cstⅡ genes were Asn-51, while 25% of the GBS-associated and all of the GBS-unrelated cstⅡ genes were Thr-51.LL strain showed highly identity to other GBS-unrelated strains in this study. Conclusion The 165-180 segment of secondary structures in cstⅡ gene from local 3 GBS-associated C.jejuni strains are probably the responsible region involved in inducing GBS. The senior structure changes in this region may affect the activity of sialyltransferase and the structures of ganglioside epitope, so that the C.jejuni can acquire the pathogenicity of GBS. This finding may give a clue to genetic modified site. The bi-functional cstⅡ of C.jejuni may be related to the pathogenicity of GBS. The cstⅡ of LL strain to some extent represents the characteristics of Asian strains, which may directs strains monitoring.

Objective To investigate the correlation between reproductive hormone concentration and the amplitude and latency of speech-evoked auditory brainstem response (speech-ABR) in young adults, and to explore the effects of reproductive hormone on the speech processing ability of young people.Methods Speech-ABR of thirty five normal hearing young adults, including seventeen females (27.29±1.83 years old) and eighteen males (28.17±2.50 years old) were recorded.The speech syllable /da/ was transmitted as a stimulus sound to the right ears through insert earphones in speech-ABR test.All participants had air conduction hearing thresholds of 20 dB HL or better across the standard audiometric frequencies (250~8 000 Hz) in both ears, and click-ABRs were also within normal limits.At the same time, the concentrations of estradiol and testosterone in the serum were examined.Results ① Females had a shorter latency than males in transient responses (waves V, A and O) and sustained responses (waves D, E and F) of speech-ABR (P<0.05, respectively).The amplitude of transient response (waves V and A) and sustained response (waves D, E and F) in females was also significantly larger than that in males (P<0.05, respectively), except for amplitude of peak O (P>0.05).The V/A slope in females was significantly steeper than that in males (P<0.05).② Estradiol levels in females (118.77±102.66 pg/ml) were significantly higher than that in males (52.91±14.77 pg/ml) (P<0.05), and the total testosterone concentration in females (457.65±140.82 pg/ml) was significantly lower than that in males (3 677.37±1 155.80 pg/ml) (P<0.05).③ A correlation analysis between speech-ABR and estradiol or total testosterone showed that all peak latencies of speech-ABR in transient responses (waves V, A and O) and sustained responses (waves D, E and F) were negatively correlated with the estradiol concentration (P<0.05 respectively), in which the correlation coefficient was between 0.2~0.4.All peak latencies of speech-ABR were positively correlated with the total testosterone concentration (P<0.05 respectively), in which the correlation coefficient was between 0.4~0.7.④ The amplitudes of speech-ABR increased with estradiol concentration growing, the wave V and estradiol concentrations were positively correlated (P<0.05).The estradiol concentrations showed a significant negative correlation with wave A, D, E, F and O wave (P<0.05 respectively), with a correlation coefficient between 0.2~0.7.On the contrary, the amplitudes of speech-ABR decreased with the increasing of total testosterone concentration, and the wave V, wave A, V/A slope and total testosterone concentration were moderately correlated (P<0.05),with a correlation coefficient between 0.4-0.6.The correlation between the amplitudes of D wave and total testosterone concentration was not statistically significant (P>0.05), and the correlation between wave E and wave F and total testosterone concentration was weakly correlated (P<0.05).In addition, the amplitudes of the wave O were also independent with testosterone levels (r=0.133, P>0.05).Conclusion There are correlations between the level of reproductive hormone and the amplitude and latency of speech-ABR.It is one of the reasons for the gender difference in the brainstem speech coding ability of normal young adult.

Objective: To evaluate the impact of intracoronary administration of eptifibatide oncoronary no-reflow and myocardium perfusion in patients with ST-elevation myocardial infarction (STEMI) at percutaneous coronary intervention (PCI). Methods: A total of 80 STEMI patients with emergent PCI were randomly divided into 2 groups: Eptifibatide group, the patients received intracoronary administration of eptiifbatide and Control group, the patients received the same volume of normal saline.n=40 in each group. The baseline condition, post-operative vascular recanalization, changes of platelet aggression at pre- and post-medication were compared between 2 groups. Echocardiography was examined at immediately and 24 weeks after operation;myocardial infusion imaging was examined at l week after operation. All patients were followed-up for 24 weeks to observe the incidence of major adverse cardiovascular events (MACE). Results: Compared with Control group, Eptifibatide group showed increased ratios of post-operative TIMI grade 3 (72.5%vs 92.5%) and myocardium perfusion (70.0% vs 90.0%), bothP<0.05; decreased post-operative and 2h post-medicinal platelet aggression and they were both lower than Control group at the same period, allP<0.05. Eptiifbatide group had obviously improved LVEDD and LVEF at 24-week than 1-week after PCI and they were both superior to Control group, allP<0.05. There were 7 (17.5%) patients in Eptiifbatide group and 7 (7.5%) in Control group suffering from small bleeding events, P>0.05; no severe bleeding eventand no in-hospital thrombocytopeniaoccurred. MACE occurrence rates during 24-week follow-up period were 12.5% vs 22.5%, P>0.05. Conclusion: Intracoronary administration of eptiifbatide in STEMI patients at emergent PCI could effectively improve coronary blood lfow,increase myocardium perfusion and enhance cardiac function without severe bleeding events.

To investigate the transgenic expressing efficacy of helper-dependent adenoviral vector (HDAd) in vitro, we constructed a HDAd encoding enhanced green fluorescent protein (EGFP), denominated as HDAd/EGFP, performed large scale preparation and purification, and then identified the purified HDAd/EGFP under fluorescent microscope and electron microscope. After the concentration of HDAd/EGFP was determined by spectrophotometer, the transgenic expression efficiency of HDAd/EGFP was compared with first generation adenoviral vector encoding EGFP (FGAd/EGFP) in vitro. Therefore, we infected A549 cells with 2000 virus particles (vp) per cell by HDAd/EGFP and FGAd/EGFP respectively and analyzed EGFP expressing level by flow cytometry. Consequently, the fluorescent expression rate and fluorescent intensity of EGFP were higher in early infected A549 cells by HDAd/EGFP than by FGAd/EGFP. HDAd, capable of expressing transgene instantly and efficiently in vitro, is a potential vaccine vector.
Adenoviridae ; genetics ; metabolism ; Cell Line, Tumor ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; Helper Viruses ; genetics ; metabolism ; Humans ; Transgenes ; Viral Fusion Proteins ; genetics ; metabolism

OBJECTIVE: To investigate the expression of fibroblast growth factor-2 (FGF-2) and osteopontin (OPN) in non-small cell lung cancer (NSCLC) tissues and analyze the correlation between FGF-2 and OPN. METHODS: Immunohistochemical SP method was used to detect the expression of FGF-2 and OPN in 76 patients with NSCLC and 15 normal lung tissues. The effect of FGF-2 on OPN expression at mRNA and protein level in A549 cell was examined by RT-PCR and Western blot. RESULTS: The positive expression of FGF-2 (65.8%) and OPN (60.5%) in the NSCLC tissues was significantly higher than that in the normal lung tissues (13.3% and 0, respectively ) (P<0.01). The expression of FGF-2 and OPN was closely related to TNM stages and the lymph node metastasis (all Ps<0.01), but not to histological types, sex, and age of NSCLC patients (all Ps>0.05).A positive correlation was found between the expression of FGF-2 and OPN in NSCLC (r=0.552,P<0.01). The expression of OPN protein and mRNA was up-regulated by FGF-2 in A549 cells. CONCLUSION The overexpression of FGF-2 and OPN is related to the metastasis and invasion of NSCLC.FGF-2 may promote the metastasis and invasion of NSCLC depending on the upregulation of OPN expression.
Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; metabolism ; pathology ; Female ; Fibroblast Growth Factor 2 ; genetics ; metabolism ; Humans ; Lung Neoplasms ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Metastasis ; Osteopontin ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism

Objective:To find clues potentially valuable for fighting against infection with human respiratory syncytial virus (HRSV), the differentially expressed genes in HEp-2 cells infected with HRSV were analyzed.Methods:Gene expression profiles of HEp-2 cells infected with HRSV were collected from the public gene expression omnibus (GEO) database. The differentially expressed genes following HRSV infection at each time point of 4, 8, 12, and 15 hours were found using R language. The differentially expressed genes were analyzed by gene ontology (GO), KEGG pathway and protein-protein interaction network (PPI). Genes with relatively high protein interaction in PPI were randomly selected for quantitative reverse transcription-polymerase chain reaction (qRT-PCR) verification at the transcription level from HEp-2 cells after HRSV infection at 4 hours.Results:A total of 101 differentially expressed genes were determined, including 92 upregulated genes and 9 downregulated genes. Function enrichment analysis revealed that HRSV infection could cause significant changes in multiple signaling pathways such as immune response in HEp-2 cells. The results of qRT-PCR were consistent with the trend of transcriptome data.Conclusions:The differentially expressed genes and the change of signaling pathways in HRSV-infected HEp-2 cells is of great significance to the studies on pathogenic mechanism and prevention of HRSV infection.

Objective To evaluate the application of three deep learning algorithms in automatic segmentation of clinical target volumes (CTVs) in high-dose-rate brachytherapy after surgery for endometrial carcinoma. Methods A dataset comprising computed tomography scans from 306 post-surgery patients with endometrial carcinoma was divided into three subsets: 246 cases for training, 30 cases for validation, and 30 cases for testing. Three deep convolutional neural network models, 3D U-Net, 3D Res U-Net, and V-Net, were compared for CTV segmentation. Several commonly used quantitative metrics were employed, i.e., Dice similarity coefficient, Hausdorff distance, 95th percentile of Hausdorff distance, and Intersection over Union. Results During the testing phase, CTV segmentation with 3D U-Net, 3D Res U-Net, and V-Net showed a mean Dice similarity coefficient of 0.90 ± 0.07, 0.95 ± 0.06, and 0.95 ± 0.06, a mean Hausdorff distance of 2.51 ± 1.70, 0.96 ± 1.01, and 0.98 ± 0.95 mm, a mean 95th percentile of Hausdorff distance of 1.33 ± 1.02, 0.65 ± 0.91, and 0.40 ± 0.72 mm, and a mean Intersection over Union of 0.85 ± 0.11, 0.91 ± 0.09, and 0.92 ± 0.09, respectively. Segmentation based on V-Net was similarly to that performed by experienced radiation oncologists. The CTV segmentation time was < 3.2 s, which could save the work time of clinicians. Conclusion V-Net is better than other models in CTV segmentation as indicated by quantitative metrics and clinician assessment. Additionally, the method is highly consistent with the ground truth, reducing inter-doctor variability and treatment time.