Objective:To explore the in vitro anticancer effect of bleomycin (Ble) combined with cisplatin (DDP) on human gastric cancer cells.Methods:CCK-8 experiment was conducted to detect the effect of Ble and DDP alone or in combination on the proliferation of gastric cancer cells; Flow cytometry was used to detect changes in cell apoptosis and changes in reactive oxygen species. Cell scratch test was employed to detect cell migration; Western blot experiment was conducted to detect protein expression.Results:The results of CCK-8 showed that 20 μM DDP could inhibit the proliferation of six types of gastric cancer cells by 40%, while the effect of 20 μM Ble was only about 20%. When used in combination, the inhibitory rate on six types of gastric cancer could reach over 60%, with the strongest inhibitory effect on AGS cells. When 10 μM Ble and 10 μM DDP were combined, the proportion of AGS apoptotic cells reached 75.68%, higher than that of the two drugs treated with 20%, respectively 20 μM processing effect. When 10 μM Ble and 10 μM DDP were used in combination, the number of migrating and invading cells was significantly reduced, and the inhibition rate reached 50%. Compared with single use, the difference was statistically significant ( P<0.05). Flow cytometry results showed that the combination of Ble and DDP promoted cancer cell apoptosis by up-regulating the level of reactive oxygen species in cells; Western blot results showed that the expression of p-JAK2 and p-STAT3 decreased after Ble and DDP were combined, indicating that the JAK2/STAT3 signal pathway was inhibited. Conclusions:The combination of Ble and DDP can inhibit the proliferation and migration of gastric cancer cells, promote the apoptosis of gastric cancer cells, and play a synergistic anti-cancer effect. Its mechanism may be related to the up-regulation of ROS levels in cells, thereby inhibiting the activation of the JAK2/STAT3 pathway.

Objective:To investigate the biomechanical characteristics and clinical outcomes of arthroscopic superior capsular reconstruction for massive irreparable rotator cuff tears (MIRCT) using the long head of biceps tendon (LHBT) with tenotomizing its distally or not (the "Chinese way" ).Methods:Eight fresh-frozen cadaveric shoulders were used to create a MIRCT model by detaching the footprints of the supraspinatus and infraspinatus tendons on the greater tuberosity. LHBT autograft was transferred and securely fixed onto the footprint of supraspinatus tendon for superior capsular reconstruction. Further, all cadaveric specimens were assigned to the tenotomy group or reservation group (4 cadaveric specimens in each group) according to whether the distal part of LHBT was tentomized or not. Biomechanical tests were conducted to observe the stiffness, ultimate load of fixed LHBT and to measure the length between LHBT tear site and its insertion on the superior labrum. A total of 41 patients with MIRCT who underwent arthroscopic superior capsular reconstruction using LHBT autograft between July 2016 and December 2018 were enrolled in the study. There were 17 males and 24 females, aged from 46 to 76 years (62.6±7.3 years). All patients were assigned to the tenotomy group (23 cases) or reservation group (18 cases) according to whether the distal part of LHBT was tentomized or not. The visual analogue scale (VAS), University of California, Los Angeles (UCLA) score, American Shoulder and Elbow Surgeons (ASES) score, Constant-Murley score and Fudan University Shoulder Score (FUSS) were used to evaluate the clinical outcomes. The range of motion (ROM) of shoulder was recorded before and after operation. Magnetic resonance imaging was used to assess the structural integrity of reconstructed tissue at 12 months after operation (refers to the failure of the transposed LHBT, which may be accompanied by a retear of partial repaired supraspinatus tendon).Results:Biomechanical research showed that the stiffness, ultimate load of fixed LHBT and the length between LHBT tear site and its insertion on the superior labrum in the reservation group (54.0±6.6 N/mm, 141.8±15.9 N, 93.3±12.4 mm, respectively) were significantly higher than those in the tenotomy group (25.7±4.2 N/mm, 80.8±8.0 N, 47.4±2.0 mm, respectively) ( P<0.05). All patients were followed up for 12-18 months (14.5±1.8 months) without significant complications and adverse reactions. No matter the distal part of LHBT was tentomized or not, the ROM and clinical scores (VAS score, UCLA score, Constant-Murley score, ASES score and FUSS) of patients improved significantly at 1 year follow-up than that before operation ( P<0.05). However, there were no significant differences between the reservation group and tenotomy group in terms of postoperative ROM [flexion, abduction, external rotation at side, internal rotation (vertebral level) were 144.3°±15.5° vs. 148.0°±10.3°, 145.1°±14.1° vs. 142.3°±11.2°, 67.3°±14.4° vs. 62.7°±11.7°, 8.3±2.1 vs. 7.8±2.5, respectively], VAS scores (2.3±1.6 vs.1.5±1.2), functional scores (Constant-Murley score, UCLA score, ASES score and FUSS were 88.2±11.4 vs. 85.6±9.6, 29.3±2.8 vs. 31.4±3.5, 86.8±11.8 vs. 82.6±9.2, 92.1±10.1 vs. 88.3±8.2, respectively) and structural failures (35.2% vs. 30.0%, P>0.05). Conclusion:Arthroscopic superior capsular reconstruction using LHBT with reserving its distal part could achieve higher mechanics strength. However, the short-term follow-up showed that tenotomizing the distal part of LHBT exerted no obvious influence on postoperative function and structural integrity.

OBJECTIVE： To investigate the effects of Danhong injection （DHI） on gene expression profile of acute myocardial infarction （AMI） model rats. METHODS： Male SD rats were randomly divided into sham operation group， model group and DHI group （0.76 mL/kg）， with 10 rats in each group. AMI model was established by ligation of left anterior descending coronary artery in model group and DHI group. After modeling， sham operation group and model group were given constant volume of normal saline intramuscularly， and DHI group was given relevant medicine intramuscularly， once a day， for consecutive 14 days. After last administration， myocardial tissue in the marginal zone of infarction was separated. The change of gene expression profile was detected by gene chip technique. Using fold-change of relative expression as index， differentially expressed microRNA （miRNA） were screened. On the basis of retrieving their corresponding genes， gene ontology （GO） and KEGG pathway enrichment analysis were carried out by using DAVID bioinformatics resource database and KEGG pathway database， respectively. TargetScan database was used to predict the target gene messenger RNA （mRNA） corresponding to differentially expressed miRNA. Cytoscape 3.6.1 software was used to construct and analyze the miRNA-mRNA network. Agilent GeneSpring GX v11.5 software was used to screen target genes and miRNA related to inflammation in the above networks. RESULTS： Compared with sham operation group， there were 22 differentially expressed miRNAs in model group， 5 up-regulated and 17 down-regulated. Compared with model group， there were 26 differentially expressed miRNAs in DHI group， and all of them were up-regulated. The differentially expressed miRNAs related to DHI therapy for AMI included rno-let-7a-5p， rno-let-7d-5p， rno-let-7f-5p， rno-miR-26b-5p， rno-miR-29b-3p， cel-miR-39-3p， cel-miR-39-5p， rno-miR-142-5p， rno-miR-191a-5p， rno-miR-409a-3p. Results of GO analysis and KEGG pathway enrichment analysis showed that differentially expressed miRNAs were mainly concentrated in membrane-bound organelles， cytoplasm， endometrial system and other cell components. The molecular functions such as protein binding and ion binding were exerted through biological processes such as anatomical structure development， multicellular tissue development and development process，which were mainly enriched in calcium signaling pathway， PPAR signaling pathway， VEGF signaling pathway， cell apoptosis， glycosylphosphatidylinositol anchored biosynthesis， valine， leucine and isoleucine degradation， etc. miRNA-mRNA network analysis showed that there were 25 target gene mRNAs corresponding to differentially expressed miRNA and 24 miRNAs related to it. There were 6 inflammation-related target genes （IL6， IL1b， TNF， TLR4， CRP， CXCL12） in this network， involving 19 differentially expressed miRNAs. CONCLUSIONS： The therapeutic effect of DHI on AMI may be related to regulating the expression of related miRNA， affecting signal transduction of calcium ion， PPAR and VEGF pathways， and regulating the secretion of inflammatory markers such as interleukin， chemokine and C-reactive protein.

Objective To investigate the characteristics of adenovirus infection in hospitalized children with respiratory tract infection in Shenzhen.Methods Nasopharyngeal swabs obtained from 25 602 children hospitalized with respiratory tract infections in Shenzhen Children's Hospital during 2014 to 2016,were tested for adenovirus with direct immunofluorescence assay.The detection rate of adenovirus and diagnosis in hospitalized children with respiratory tract infection were analyzed.Results The total adenovirus detection rate was 2.97 ％ in 25 602 samples,with a male to female ratio of 2.04:1,no significantly difference in detection rate in male (3.12％) and female (2.70％).Accounted for 724 (95.14％) of the total adenovirus positive detection children below six years old,and 409(53.75％) children were detected below two years old.There was a distinct seasonality;the detection rate was higher in summer and winter (x2 =36.631,P＜0.01).In 761 hospitalized patients of ADV positive,431 were pneumonia,109 were bronchitis,74 were tonsillitis,14 were conjunctivitis pharynx and 133 were acute upper respiratory infection.Conclusion Our study demonstrates that respiratory adenovirus infection is an important cause of hospitalization in children below the age of 6 years in Shenzhen,China.The detection rate was higher in summer and winter than spring and autumn.Most adenovirus positive children were diagnosed by pneumonia,bronchitis and acute upper respiratory tract infection.

Objective To investigate protective activity against Aβ25-35-induced cytotoxicity in PC12 cells of different extracts and ursolic acid, which were isolated from pyrola decorata. Methods Aβ25-35-induced cytotoxicity in PC12 cells was established as the model in vitro. The cultured PC12 cells were divided into blank control group, DMSO control group, model control group, different extract groups of pyrola decorate and ursolic acid(UA) group. The different extract groups included ether extract (PE), acetidin extract (AE), n-butanol extract (BE), the water extract (WE), 50% ethanol extract (HEE). MTT assay was used to test the optimum concentration, and the number of viable cells in culture medium was measured by ELISA at 490 nm wavelength. Results The cell viabilities in different extracts groups(PE, AE, BE, WE, HEE) were respectively 89.3%, 77.2%, 79. 2%, 75. 1% ,74. 0% at the concentration of 5. 0 mg ? mL-1 . Moreover, ursolic acid showed the best neuroprotective activity (88.9%) at the concentration of 500 μg?L-1 . Compared with model control group, the survival rate of each group was remarkably increased, and the protective activities of PE and UA were more significant among them. Conclusion Different polar extracts of pyrola decorata and isolated ursolic acid have neuroprotective effects on Aβ25-35-induced cytotoxicity in PC12 cells in certain degrees.

Objective To investigate the role of osteopontin (OPN) in the pathogenesis ot cholesterol gallstone formation in bile.Methods The nucleation time of OPN in model bile and human gallbladder bile was studied by the nucleation time assay,the effect of OPN on cholesterol crystal growth in model bile was examined by the cholesterol crystal growth assay.The effect of OPN on vesicle was detected by the transmission electron microscopy in model bile and gallbladder bile; then the content of OPN and calcium were detected via the commercial kits in human bile.Results Osteopontin prolonged nucleation time in a dose dependent manner in model bile and human bile,and this effect was correlated with calcium.Compared with control group,the nucleation times were prolonged by 1.50and 1.93 times in lithogenic bile at the concentration of osteopontin 50 μg/ml and 100 μg/ml (P＜0.01),respectively. Nucleation time were prolonged by 1.17 and 1.33 times in normal bile (P＜0.01) and by 1.29 and 1.48 times in model bile (P＜0.01),respectively.The rate of cholesterol crystals growth was not influenced by calcium ions,but inhibited by osteopontin in a dose dependent manner in the model bile.Furthermore,the formation,aggregation and fusion of vesicles were delayed by osteopontin in bile samples as indicated by the transmission electron microscopy.The concentration of osteopontin [(0.53± 0.08) mg/ml vs. (0.65 ± 0.14) mg/ml,P＜0.05] and the calcium ions [ (0.71 ± 0.17) mmol/L vs. ( 0.84 ± 0.08 ) mmol/L,P ＜ 0.05 ] were lower in lithogenic bile than in control.Conclusions Osteopontin can inhibit the cholesterol gallstone formation in model and human gallbladder bile as the anti nucleating factor.

Objective To investigate the role of osteopontin (OPN) in cholesterol gallstone formation.MethodsGallbladder bile was obtained from patients with cholelithiasis (n=36,the experimental group) and from donors of liver transplantation (n=19,the control group).OPN,calcium ion and lipid were analysed quantitively.The nucleating role of OPN in bile was evaluated using nucleating time (NT) approach.ResultsOPN inhibited cholesterol nucleation in a dose dependent manner.OPN (50 μg/ml and 100 μg/ml) prolonged NT by 48.90％ (91.51％) and 17.07％ (32.93％) in lithogenic and control bile,respectively.OPN (100 μg/ml) also inhibited the nucleating effect induced by calcium ion.Furthermore,a combination of OPN (50 μg/ml) and calcium prolonged NT by 75.78％ and 33.96％ in lithogenic and control bile,respectively.A combination of OPN (100 μg/ml) and calcium prolonged NT by 125.9％ and 62.26％ in the 2 groups.The contents of osteopontin and calcium were significantly lower in lithogenic bile than control bile (P＜0.05).On the other hand,the cholesterol saturation index and the contents of cholesterol,phospholipid and bile acid were significantly higher (P＜0.05).ConclusionsOPN inhibited cholesterol gallstone formation.It may be involved in the pathogenesis of cholelithiasis.

A cold-adapted (ca), temperature sensitive (ts), live attenuated influenza B virus strain B/Ann Arbor/1/66 was chosen for influenza virus rescue research, in which six internal gene segments, PB1, PB2, PA, NP, M, NS, were fully synthesized and nine amino acid substitutions were artificially alter by human intervention. The resultant B/Ann Arbor/1/66 plasmids were named as pAB121-PB1, pAB122-PB2, pAB123-PA, pAB124-HA, pAB125-NP, pAB126-NA, pAB127-M and pAB128-NS, respectively. A recombinant influenza A virus was previously generated entirely from cloned cDNA. An infectious recombinant influenza B virus was generated here, and designated as rMDV-B, by plasmid-based reverse genetics. The rMDV-B virus contained HA and NA genes from an epidemic influenza B vires strain B/Malaysia/2506/2004 in the background of internal genes derived from influenza B virus strain B/Ann Arbor/1/66. HA titer of rMDV-B in MDCK cells and embryonated chicken eggs ranged from 1 : 64 to 1 : 512. The results may allow an effective live influenza B vaccine to be produced from a single master strain, providing a model for the design of future live human influenza vaccines.

Objective To set up a technical platform of reverse genetics based on the 8 plasmid.virus rescue system of cold-adapted influenza virus strain. Methods The cold-adapted, temperature sensitive, live attenuated influenza virus strain A/AnnArbor/6/60(H2N2)was chosen as the master donor virus(MDV)for rescue research,and its six internal gene fragments PB2,PB1,PA,NP,M and NS were artificially synthesized. Meanwhile, five amino acid mutations have been introduced as tags. Six fragments were ligated with modified pAD3000 for the construction of rescue plasmid. Six transcription/expression plasmids(pMDV-A-PB2,pMDV-A-PB1,pMDV-A-PA,pMDV-A-NP,pMDV-A-M,and pMDV-A-NS)were obtained, and their sequences were accurate. Results The reassorted virus named as rMDV-A contains HA and NA gene segments derived from PR8 strain along with six gene segments,PB2,PB1,PA,NP,M and NS,from MDV. The COS-1 cells were co-transfected with eight recombinant plasmids. The results showed that a cold-adapted, attenuated reassortant influenza A virus with hemagglutination activity was rescued successfullv bv"6+2" combination of MDV and PR8, and the allanotoic fluid of the injected eggs gave a posigenes of A/AA/6/60 used as backbone has provided experimental materials for further research on the gene function and novel vaccine candidate of cold-adapted, attenuated human influenza virus.

Six gene segments,PB1,PB2,PA,NP,M and NS,were fully synthesized which derived from the master donor virus (MDV),cold-adapted(ca),temperature sensitive(ts),live attenuated influenza virus strain A/Ann Arbor/6/60(MDV-A).Meanwhile,five amino acid substitutions (PB1-391E,58lG,661T,PB2-265S,NP-34G) were artificially altered by human intervention.HA and NA fragments derived from the 2006-2007 circulating strain A/New Caledonia/20/99 (H1N1).Eight fragments were ligated with modified pAD3000 for rescue plasmid construction.Eiight transcription/expression plasmids were named as pMDV-A-PB2,pMDV-A-PB1,pMDV-A-PA,pMDV-A-NP,pMDV-A-M,pMDV-A-NS,pMDV-A-HA,pMDV-A-NA,respectively.The COS-l cells were co-transfected with eight plasmids representing 6 internal viral backbone of the strain A/AA/6/60 and two plasmids containing the CDNA of the HA and NA segments of the strain A/New Caledonia/20/99 (H1N1),the results showed that cold-adapted,attenuated reassortant influenza A virus Was rescued successfully.Titers of a reassorted influenza A virus in embryonated chicken eggs mnged from 1:29to l:210.The rescue system of six intemal genes used as backbone opens the way for further research on gene function and neotype vaccine candidate of cold-adapted,live attenuated human influenza virus.