Animals ; Carcinoma, Hepatocellular ; blood supply ; virology ; Cell Transformation, Neoplastic ; Endothelial Growth Factors ; biosynthesis ; genetics ; Gene Expression Regulation, Viral ; Hepatitis B virus ; pathogenicity ; physiology ; Humans ; Liver Neoplasms ; blood supply ; virology ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Trans-Activators ; physiology

OBJECTIVETo investigate the effects of HBx gene on proliferation activity of hepatoma cells in vitro and in vivo.

METHODSThe plasmid pHA-HBx carrying HBx gene was transfected into HepG(2) cells, and the positive clones were screened and identified with G418 and RT-PCR, respectively. The growth curve and population doubling time were calculated, and the cell cycle was analyzed by flow cytometry (FCM). The proliferation activity of transformed cells was measured with (3)H-TdR incorporation rate and nude mice model in vitro and in vivo.

RESULTSThe result of RT-PCR indicated that HBx gene was integrated into the genome DNA of HepG(2) cells and transcripted. The growth curve and population doubling time showed a high proliferation activity of transformed cells. The amount of cells at stage S and G(2)/M were significantly higher, and cells at stage G(0)/G(1) were lower than those in control group. The tumors developed from transfected cells grew much quicker than those developed from HepG(2) cells in nude mice model.

CONCLUSIONHBx gene can facilitate the proliferation of hepatoma cells both in vitro and in vivo.

Animals ; Carcinoma, Hepatocellular ; genetics ; pathology ; Cell Cycle ; genetics ; Cell Division ; genetics ; Cell Line, Tumor ; Female ; Flow Cytometry ; Humans ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Reverse Transcriptase Polymerase Chain Reaction ; Trans-Activators ; genetics ; Transfection ; Transplantation, Heterologous

OBJECTIVETo study the inhibitory effects of insulin on nuclear factor-kappa B (NF-kappaB) nuclear translocation of vascular endothelial cells induced by burn serum and its correlative mechanism.

METHODSHuman umbilical vein endothelial cells (HUVECs) were cultured in vitro and divided into 5 groups: blank control group (BC, ordinary culture without any stimulation), normal serum control group (NS, cultured with nutrient solution containing 20% healthy human serum), burn serum stimulation group (BS, cultured with nutrient solution containing 20% burn human serum), burn serum+insulin treatment group (BI, cultured with nutrient solution containing 20% burn human serum and 1x10(-7) mol/L insulin), inhibitor pretreatment group [IP, pretreated with 50 micromol/L protein kinase B (Akt) specific inhibitor LY-294002, then cultured with the same medium as used in BI group 30 minutes later] according to the random number table. Six hours later, the injury and apoptosis of HUVECs was respectively observed by the scanning electron microscope and determined by the flow cytometry. Meanwhile, the phosphorylation of inhibitor kappa B-alpha (p-IkappaB-alpha) and Akt (p-Akt) in cytoplasm, and the content of NF-kappaB-p65 in nucleus were determined with Western blot.

RESULTS(1) Compared with those in BC group, HUVECs in BS group shrank obviously with irregular nuclear structure, and intercellular links jagged or vanished. Slight change was observed in HUVECs structure in NS and BI groups, with the cell ductility and nuclear structure much better than those in BS group. (2) The apoptosis rates of HUVECs in BS group [(28.5+/-2.3)%], BI group [(22.3+/-1.8)%], and IP group [(29.7+/-2.4)%] were all obviously higher than that in BC group [(15.7+/-2.2)%, F=14.288, P<0.05 or P<0.01]. There was no significant statistical difference between NS group [(17.0+/-2.5)%] and BC group in apoptosis rate (F=14.288, P>0.05). The apoptosis rate of HUVECs in BI group was obviously lower than that in BS group (F=14.288, P<0.05). (3) Compared with those in BC group, the protein expressions of p-IkappaB-alpha in cytoplasm and NF-kappaB-p65 in nucleus were up-regulated, and the protein expression of p-Akt in cytoplasm was down-regulated in BS and IP groups. The expression levels of the three proteins in NS and BI groups were close to those in BC group.

CONCLUSIONSInsulin could inhibit the IkappaB phosphorylation, and then restrict NF-kappaB nuclear translocation and improve the vascular endothelial cells function accordingly through regulating phosphatidylinositol 3 kinase/Akt pathway.

Apoptosis ; Burns ; blood ; Cells, Cultured ; Endothelial Cells ; metabolism ; Endothelium, Vascular ; cytology ; metabolism ; Humans ; I-kappa B Proteins ; metabolism ; Insulin ; pharmacology ; NF-kappa B ; metabolism ; Phosphorylation ; Serum ; metabolism ; Umbilical Veins ; cytology

OBJECTIVETo study the protective effect of intensive insulin treatment on the myocardium of severely scalded rats, and to primarily explore its mechanism.

METHODSEighteen SD rats were divided into three groups, with 6 rats in each group. The rats in burn and intensive insulin group were inflicted with 30% TBSA full-thickness injury on the back. Isotonic saline containing 0.12 U/ml insulin solution, and 100 g/L glucose solution were infused into the rats in the intensive insulin group to keep plasma glucose at the level of 4.0 - 6.6 mmol/L (the total fluid amount was 2 ml x kg(-1) x 8h(-1)). In sham burn group,fluid was given according to physiological demand. The same amount of isotonic saline was infused into the rats in burn group. The venous blood was obtained for the detection of plasma glucose contents, and the left ventricular systolic pressure (LVSP) and left ventricular end-diastolic pressure (LVEDP) were recorded via aortic ventricle cannula before scald and at 1, 2, 3, 4, 5, 6 post-scald hours (PSH). The tissue of the left ventricle was harvested at 6 PSH for the detection of troponin T expression in myocardiocytes.

RESULTSPlasma glucose level was increased to (7.6 +/- 1.7) mmol/L - (8.4 +/- 4.7) mmol/L in burn group during 1-6 PSH, which was significantly higher than that in intensive insulin group (4.5 +/- 0.9) mmol/L - (5.2 +/- 1.3) mmol/L, P < 0.01). Compared with the intensive insulin group, LVSP was markedly decreased in the burn group (60 +/- 11 mm Hg vs 72 +/- 8 mm Hg, P < 0.05) at 1 PSH,whereas LVEDP was increased significantly (21.3 +/- 11.3 mmHg vs 11.7 +/- 5.2 mmHg, P < 0.05). Intensive insulin treatment could significantly inhibit the loss of troponin T protein in myofilaments of myocardium.

CONCLUSIONIntensive insulin treatment possesses a protective effect on myocardia function after severe burns, and it may be related to its preventive effect on the loss of contractile protein in cardiocytes.

Animals ; Blood Glucose ; metabolism ; Burns ; drug therapy ; metabolism ; Insulin ; administration & dosage ; therapeutic use ; Male ; Myocardial Contraction ; Myocardium ; metabolism ; Rats ; Rats, Sprague-Dawley ; Troponin T ; metabolism

OBJECTIVETo investigate the anti-apoptosis effect of intensive insulin treatment on cardiac myocytes and its underlying mechanism in severe scald rats.

METHODSTwelve SD rats were suffered from 30% TBSA full thickness scald, and they were divided into: IT group [with intravenous injection of isotonic saline including insulin (15 mU x kg(-1) x min(-1)) and 100 g/L glucose], B group [with treatment of isotonic saline (2 mL x kg(-1) x %TBSA(-1) x 8 h(-1)]. Six SD rats received sham burn as controls[sham(S)group, with treatment of fluid at physiologic dose]. + dp/ dtmax (the rate of the rise of left ventricular pressure) and -dp/ dtmax (the rate of the fall of left ventricular pressure)at 6 post burn hour (PBH)were recorded. Apoptosis were determined by TUNEL staining and DNA ladder. The phosphorylation f Akt and protein expression of Bcl-2 in cardiomyocyte were assayed by Western blotting.

RESULTSThe + dp/ dtmax in the S group, IT group and B group at6 PBH were respectively (5.5 +/- 0.5) x 10(3) mm Hg/s, (3.4 +/- 0.4) x 10(3 mm Hg/s and (2.5 +/- 0.5) x 10(3) mm Hg/s (1 mm Hg = 0.133 kPa), the - dp/ dtmax were respectively (4.55 +/- 0.34) x 10(3) mmHg/s, (2.94 +/- 0.22) x 10(3) mm Hg/s and (2.05 +/- 0.19) x 10(3) mmHg/s.The +/- dp/dtmax in IT group was significantly higher than those in B group( P < 0.01). The apoptosis index in B group was (13.1 +/- 3.4)%, which was obviously higher than that in IT group (6.7 +/- 1.8)% and S group (0.6 +/- 0.4)% (P < 0.01). DNA ladder showed that no DNA fragmentation in S group, but obvious DNA fragmentation forming ladder pattern in B group, and no obvious ladder pattern in IT group. The phosphorylation of Akt and level of Bcl-2 protein in B group were markedly higher than those in IT group ( P < 0.05 or P < 0.01).

CONCLUSIONIntensive insulin treatment can upregulate the activity of Akt and enhance the expression of Bcl-2, and they might constitute the mechanisms for anti-apoptosis in cardiomyocyte and protection of cardiac function.

Animals ; Apoptosis ; drug effects ; Burns ; drug therapy ; pathology ; Insulin ; administration & dosage ; pharmacology ; Male ; Myocytes, Cardiac ; cytology ; metabolism ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the protective effect of insulin on oxygen-radical induced hepatic injury in severely scalded rats in early stage of severe scald.

METHODSEighty-four male Sprague-Dawley rats were randomly divided into three groups: i. e, normal group, saline group, and insulin group, with 28 rat in each group. The rats in the latter two groups were subjected to 30% TBSA full-thickness scald on the back, and received intra-peritoneal injection of 40ml/kg isotonic saline, and subcutaneous injection of 3 IU/kg insulin, respectively. The total anti-oxygen capability (T-AOC), the expression of superoxide dismutase (SOD), reactive oxygen species (ROS) and intercellular adhesion molecule (ICAM-1) in hepatic tissue, and serum alanine transaminase (ALT) were determined in each group at 6, 12, 24, 48 post-scald hours (PSH) with corresponding methods.

RESULTSThe hepatic T-AOC and SOD content were obviously decreased, while the ROS content were markedly increased at 6 PSH in saline group compared with that in normal group (P < 0.05 or P < 0.01). The expression of ICAM-1 and serum content of ALT were significantly higher than that in normal group at 12 PSH and 48 PSH (P < 0.01). At 24 PSH, the hepatic T-AOC (386 +/- 75) U/g and SOD content (210 +/- 39 ) U/g were obviously higher in insulin group than those in saline group [(124 +/- 18), (111 +/- 9) U/g, respectively, P < 0.01), but the ROS content (154 +/- 29 ) U/g was much lower than that in saline group [(351 +/- 41) U/g, respectively, P < 0.01]. At 48 PSH, the serum content of ALT and hepatic expression of ICAM-1 in insulin group exhibited obvious difference when compared with those in saline group (P < 0.01). Meanwhile, Pathological examination showed that hepatic injury was alleviated by insulin administration after scald.

CONCLUSIONInsulin administration early after severe scald exhibits protective effect on liver function by improving anti-oxygen radical ability of rat liver.

Alanine Transaminase ; blood ; Animals ; Burns ; metabolism ; pathology ; Insulin ; pharmacology ; Liver ; drug effects ; metabolism ; pathology ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species ; metabolism ; Superoxide Dismutase ; metabolism

Objective To identify potential microRNAs (miRNAs) in salivary adenoid cystic carcinoma and to construct a miRNA-mRNA regulatory network to better understand its potential molecular mechanisms. Methods Two microarray datasets of SACC were downloaded from the database Gene Expression Omnibus (GEO), and the differentially expressed miRNAs and mRNA were analyzed by the R language. FunRich 3. 1. 3 software was used to enrich and analyze the transcription factors of differential miRNAs and to predict the target genes of differentially expressed miRNAs. The target genes of differential miRNAs in SACC were utilized to perform Gene Onotology (GO) and Kyoto Encyclopedia of Gene Genomes (KEGG) pathway enrichment analyses, and protein-protein interaction. The miRNA-mRNA regulatory network was constructed in Cytoscape 3.7.0. Results A total of 144 differentially expressed miRNA (DEMs) and 1216 differentially expressed mRNA (DEGs) were screened. The enrichment analysis of KEGG signaling pathway revealed that target genes were mainly involved in the regulation of Rapi signaling pathway, mitogen active protein kinase (MAPK) signaling pathway, phosphatidylinositol 3-kinase/protein kinase B(PI3K/Akt) signaling pathway, and regulation of actin cytoskeleton. STRING protein interaction analysis shows that ACSL1, SCD, MGLL, FABP4 may be the key proteins in the protein interaction network. Conclusion Differentially expressed miRNA and mRNA between SACC tissues and normal tissues were screened out and the signaling pathways and functions of these differential molecules were found in our research.

Objective:To investigate whether human umbilical vein endothelial cell(HUVEC) vaccine combined with low dose docetaxel (DOC) could play a synergistic role in anti-breast cancer.Methods:BALB/c mice were randomly divided into normal saline group,HUVEC vaccine group,DOC group,and HUVEC vaccine combined with DOC treatment group (HUVEC-DOC) group.An experimental metastasis model by tail vein injection of EMT-6 breast cancer cells was employed to evaluate the anti-metastatic efficiency of the HUVEC-DOC combination treatment regime.Lymphocyte proliferation assay,cytotoxic T lymphocytes and an indirect enzyme-linked immunosorbent assay (ELISA) for detecting IFN-γ were used to investigate cellular immune responses elicited by the combination treatment regime.Results:Compared with HUVEC and DOC single drug group,the number of lung metastasis in HUVEC-DOC combination treatment group was significantly decreased(P<0.05).In vitro analysis of splenocytes isolated from immunized mice revealed an induction of cytotoxic T lymphocytes(CTLs) with a lytic activity against activated endothelium.IFN-γ in the serum of im-munized mice of the HUVEC-DOC combination treatment group was significantly higher than that in the other three groups(P<0.05). Conclusion:HUVEC vaccine with low dose of DOC could display synergistic anti-breast cancer effect.

We investigated epidemiological characteristic of scrub typhus in Yongshan County,Yunnan Province,China. The serum samples were collected from the patients with fever for detecting the antibody against Orientia tsutsugamsushi (Ot) by colloidal gold immunoassay assay.Rat traps were used to capture rodents.The spleen tissues of the captured rodents were detected by nested-polymerase chain reaction for rickettsia groEL segment.The groEL segments were sequenced and analyzed the homology with the other known sequences.Thirty-four scrub typhus cases were found in Yongshan County,Yunnan Prov-ince from May 2015 to October 2015.Among them,21 cases were confirmed by laboratory tests and 13 cases were clinical di-agnosis diseases.Of these patients,32.35% of the cases occurred in June.The 32.35% were in the group of the 40-49 year-old,and 79.41% were farmers,94.12% exhibited eschar or skin ulcer(31.25% were observed in groin of these cases),and rash developed in 50%.In 39 spleen tissue samples of Rattus flavipectus,9 samples showed positive for groEL gene Ot,but gro-EL gene of Typhus group rickettsia and spotted fever group rickettsia were negative.Sequence analysis showed that YSP30 was closely related to some Saitama related strains of Ot,such as HSB1,FAR1 and UAP4,while the other 8 strains were closely related to some Karp related strains of Ot,such as UT213,UT221 and SH205.It was confirmed that the Yongshan County was the natural foci of scrub typhus by the serological and molecular biological detections.There are Karp and Saitama genotype related Ots in the natural foci.

Objective To provide theoretical reference for the research on lateral postural control in older people by using the electromyography （EMG）technique to compare the neuromuscular reaction between the young and older people during lateral perturbation. Methods Postural reactions of 14 young people and 14 older people as subjects were evoked by sudden unpredictable medio lateral translation platform. The surface EMG data were collected from peroneus longus, anterior tibialis, gluteus medius and erector spinae of the left side of the body. Results The latencies in anterior tibialis, gluteus medius and erector spinae were obviously delayed in older people, and the duration of time to peak in peroneus longus, anterior tibialis and gluteus medius of older people was much longer than that of young people. Conclusions The delayed response of ankle, hip and trunk muscle and the low efficacy of contraction rate in ankle and hip muscles to the unexpected perturbation might be the primary reason for the decrease of lateral postural control in older people.