Objective:To express and purify the TRIM5? chimaera[TRIM5? H(R328-332)] protein and to explore the interaction between the TRIM5? H(R328-332)and HIV-1gag. Methods:The plasmid pET28aTRIM5? H(R328-332) was transformed to E.coli BL21 (DE3) strain ,and the expression of TRIM5? H(R328-332) protein was induced by IPTG,purified with Ni2+ chromatography.The expression and purification of TRIM5? H(R328-332) were analyzed by SDS-PAGE and Western blot,and the interaction between TRIM5? H(R328-332) and HIV-1gag was detected by co-immunoprecipitation,His pull-down and ELISA. Results:The recombinant plasmid pET28aTRIM5? H(R328-332) was successfully expressed in E.coli. The results showed that the purified full length TRIM5? H(R328-332) interacted with HIV-1gag protein. Conclusion:The human TRIM5? chimaera was expressed successfully in vitro,and the study demonstrates that the human TRIM5? chimaera interacts with HIV-1 gag in vitro.

AlM:To observe the relationship between axial length and complications of phacoemulsification with intraocular lens ( lOL) implantation in high axial myopia eyes and normal axis eyes.
METHODS: A retrospective review of 843 consecutive patients ( 1 042 eyes ) of cataract extraction with phacoemulsification and lOL implantation in our hospital from February 2012 to February 2013 was performed. The patients were divided into two groups according to the axial length: 853 eyes were in normal axis group ( 21-24mm) and 189 eyes were in high axial myopia group (≥26mm). The two groups were compared regarding surgical complications, such as vitreous loss, posterior capsular rupture, nucleolus drop, and abnormal location of lOL.
RESULTS:Age was a risk factor in both groups. There was positive correlation between age and surgical complications, and between axial length and surgical complications, especially for complications with posterior capsular rupture and vitreous loss.
CONCLUSlON:As the results illustrate, in this survey, age and high axial lengthare statistically significant risk factors for incidence of complications of phacoemulsification. Anticipation of these complications and also preparation and prophylactic measures may decrease incidence of these complications.

Forty batches of Lonicerae Japonica Fse i collected extensively and prepared as the test solution. Their chromatographic fingerprints and anti-influenza virus IC50 value (half maximal inhibitory concentration) were determined respectively. Then Unscrambler software was used, and spectrum-efficient correlation analysis was done for chromatographic fingerprints data and IC50 data by partial least squares regression method, to establish spectrum-efficient correlation model for anti-influenza virus of Lonicerae Japonicae Flos. Then the other 10 batches of Lonicerae Japonicae Flos were used to verify the model and explore the adaptability of this spectrum-efficient correlation model based on partial least squares regression method. The mathematical model obtained R2 of 0.969489 and RM-SEC of 0.070691 for calibration set; R2 of 0.959042 and RMSECV of 0.084005 for cross validation set. The verification experiment results showed that the relative error between the predicted values and measured values was within 10% in all 10 hatches, and within 5% in 80% of them. The results showed that the established spectrum-efficient correlation model could be used to evaluate the biological activity of anti-influenza virus of Lonicerae Japonicae Flos by determining its HPLC fingerprints.
Antiviral Agents ; analysis ; pharmacology ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; pharmacology ; Flowers ; chemistry ; Least-Squares Analysis ; Lonicera ; chemistry ; Molecular Structure ; Orthomyxoviridae ; drug effects

OBJECTIVESTo investigate whether antisense Bmi-1 plasmid could inhibit the proliferation of Jurkat cells.

METHODSThe antisense plasmid was constructed by PCR amplification of a 171 bp segment spanning Bmi-1 start codon and zinc finger structure and the PCR product was subsequently inserted reversely to plasmid pLNCX2. The final construct was confirmed through restriction enzyme digestion. G418 was added into the medium after the plasmid was successfully introduced into Jurkat cells by using lipofectin-mediated DNA transfection. The proliferation of Jurkat cells were determined by MTT and colony formation assays. Cell cycle was determined by flow cytometry. The p16 expression of Jurkat cells was studied by immunofluorescent histochemistry.

RESULTSThe growth rate of antisense Bmi-1 transfected Jurkat cells was significantly lower than that of the controls, and the colony forming capacity of the transfected cells decreased significantly (P < 0.01), the colony numbers being (90.7 +/- 9.07)/10(3) cells, (83.3 +/- 6.11)/10(3) cells and (56.0 +/- 5.56)/10(3) cells for control cells, empty plasmid transfected Jurkat cells and antisense Bmi-1 transfected Jurkat cells, respectively. The percentage of G, phase cells was increased and the p16 expression of antisense Bmi-1 transfected cells was significantly upregulated than that of control cells.

CONCLUSIONAntisense Bmi-1 can inhibit the growth and upregulate the expression of p16 of Jurkat cells in vitro.

Cell Cycle ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Humans ; Jurkat Cells ; Nuclear Proteins ; genetics ; Oligonucleotides, Antisense ; genetics ; Plasmids ; genetics ; Polycomb Repressive Complex 1 ; Proto-Oncogene Proteins ; genetics ; Repressor Proteins ; genetics ; Transfection

OBJECTIVESTo investigate whether antisense human telomerase reverse transcriptase (hTERT) could inhibit the activity of telomerase and the proliferation of K562 cells.

METHODSThe antisense plasmid was constructed by reverse insertion of hTERT PCR product into plasmid pLNCX-neo. Then the constructed plasmid was introduced into K562 cells by liposomes-mediated DNA transfection. The inhibition effects of telomerase on the proliferation of K562 cells were analyzed by MTT and colony formation assay, the telomerase activity of K562 cells by TRAP-PCR ELISA methods.

RESULTSThe growth rate of antisense hTERT transfected K562 cells was significantly lower than those of the controls, and the colony formation capacity of the transfected cells decreased significantly (P < 0.01), the colony number is (100.33 +/- 7.57)/10(3) cells, (92.67 +/- 5.86)/10(3) cells and (50.33 +/- 6.11)/10(3) cells for control K562 cells, K562 neo cells and antisense hTERT transfected HL60 cells, respectively. The telomerase activity of antisense hTERT transfected K562 cells was significantly inhibited.

CONCLUSIONThe expression of an antisense sequence to the mRNA sequence of telomerase protein subunit can inhibit the activity of telomerase, slow the cell growth and inhibit the capacity of colony formation of K562 cells.

Cell Division ; drug effects ; Humans ; K562 Cells ; Plasmids ; genetics ; RNA, Antisense ; genetics ; pharmacology ; RNA, Messenger ; genetics ; Telomerase ; drug effects ; genetics ; metabolism ; Transfection

Serum pharmacochemistry of traditional Chinese medicine (TCM) is designed to screen the efficacy material base of TCMs from the constituents absorbed into the blood after oral administration. The theory and method is in accordance with the effect characteristics of TCMs, and reflects the interaction between the body and the drugs, has become an effective pathway for researching the efficacy material base of TCMs which has been recognized and used widely. In the paper, the previous research contents and methods of the serum pharmacochemistry of TCM were reviewed, and on the basis of the further validity of the special administration form of the TCM formula and the corresponding property to TCM syndrome, the new strategy of serum pharmacochemistry of TCM integrating the metabonomics technologies was put forward. According to the strategy, we take the biological characters of TCM syndrome as a research starting point, taking TCM formula as object, using the metabolic biomarkers of syndromes or disease to evaluate the therapeutic effect of formula and screen the compounds of TCMs in serum which are highly correlated with the metabolic biomarkers through the correlation analysis, and by further biological validation to finally confirm the efficacy material basis of TCMs. Integrating with the systems biology technologies, the theory and method of serum pharmacochemistry of TCM will further develop, and open a new chapter in the interpretation of the theory of TCM.
Animals ; Drug Therapy ; trends ; Drugs, Chinese Herbal ; chemistry ; pharmacokinetics ; Humans ; Metabolomics ; Serum ; chemistry

OBJECTIVETo study the effects of antisense Bmi-1 (B cell-specific moloney murine leukemia virus insertion site 1) RNA on the growth, cell cycle and apoptosis of lung cancer cell line A549.

METHODSRecombinant plasmids carrying antisense Bmi-1 RNA were transfected into A549 cells, which expressed a high level of endogenous Bmi-1. The mRNA level of A549 cell was analyzed by real time quantitative RT-PCR and the protein level was determined using Western blot. MTT growth curve and plate colony forming assay were used to measure the effect of antisense Bmi-1 RNA expression on the growth of A549. Flow cytometry was used to analyze cell cycle and apoptosis.

RESULTSAntisense Bmi-1 RNA reduced the Bmi-1 expression at the protein level, but did not alter the mRNA level in A549 cells. Compared with the control cells, A549 cells transfected with antisense Bmi-1 RNA showed a strong inhibition of the cell growth. The number of plate colony formation of the antisense Bmi-1 transfected cells (0.67 +/- 0.50) was less than those of the control (73.0 +/- 4.1) and cells transfected with empty vector (67.0 +/- 4.0, P < 0.01). Transfection of antisense Bmi-1 RNA arrested the A549 cells at G₀/G₁ phase of the cell cycle and did not increase the apoptosis.

CONCLUSIONAntisense Bmi-1 RNA expression inhibits A549 cells proliferation, likely through the interference of Bmi-1 leading to an arrest of the proliferating cells at the G₀/G₁ phase.

Apoptosis ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Down-Regulation ; Gene Expression Regulation, Neoplastic ; Humans ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Nuclear Proteins ; biosynthesis ; genetics ; Polycomb Repressive Complex 1 ; Proto-Oncogene Proteins ; biosynthesis ; genetics ; RNA, Antisense ; pharmacology ; RNA, Messenger ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; Repressor Proteins ; biosynthesis ; genetics ; Transfection

In order to characterize the molecular epidemiology of HFMD-associated Coxsackievirus A6 (CVA6) in Fujian Province, a total of 1340 specimens from non-EV71 non-CVA16 HFMD patients were collected during 2011-2013. Isolated virus strains were identified and subtyped. Full-length coding regions for the VP1 gene of the predominant serotype CVA6 isolates were amplified and sequenced. Among the 375 non-EV71 non-CVA16 HFMD cases confirmed by virus isolation and molecular subtyping, 182 (48.5%) were found to be caused by CVA6, accounting for 7.9%, 16.2% and 39.6% HFMD-associated enteroviruses in FujianProvince during 2011, 2012, and 2013, respectively. Compared with general features observed in the HFMD epidemic, no difference in CVA6-specificity or severity rates was observed between geographical origins, gender, or age groups. Nucleotide sequence analyses of VP1 genes revealed high diversity levels of 16.2%-18.6% among CVA6 strains from Fujian Province, in contrast to the prototype CVA6 strain, and showed low levels of diversity in the amino acid sequences (4.3%-6.2%). Phylogenetic analysis also indicated that CVA6 isolates from Fujian Province were distinct from the prototype strain and other isolates from abroad; however, it was homologous to domestic strains, although the Fujian isolates clustered into multiple branches. These results suggested that significant changes in the pathogenic spectrum of HFMD in Fujian Province occurred during 2011-2013, as CVA6 was one of the predominant serotypes of HFMD. CVA6 isolates from Fujian Province were co-circulating and co-evolving with other domestic strains as multiple closely related CVA6 transmission chains were observed in Fujian Province overall and within each prefecture.
Child ; Child, Preschool ; China ; epidemiology ; Enterovirus A, Human ; classification ; genetics ; isolation & purification ; Evolution, Molecular ; Female ; Hand, Foot and Mouth Disease ; epidemiology ; virology ; Humans ; Infant ; Male ; Molecular Epidemiology ; Molecular Sequence Data ; Phylogeny

OBJECTIVETo investigate the potential genetic mode of aggressive periodontitis (AgP) in Chinese Han nationality.

METHODSA total of 233 subjects from 73 nuclear families were recruited. All probands were diagnosed according to the criteria of AgP in 1999 classification of periodontal diseases. Ninety parents, 35 siblings and three grandparents and two offspring were examined based on full-mouth periodontal chartings (including parameter of probing depths, attachment loss, bleeding on probing at six sites per tooth) and full-mouth periapical radiographs. The genetic ratio was calculated and analyzed by the methods of Edwards and simple segregation.

RESULTSThe prevalence of AgP in probands' siblings was close to the square root of the prevalence of general population. The segregation ratio was 0.2419, which was close to the theoretical ratio for autosomal recessive inheritance. However, autosomal dominant inheritance could not be rejected in families whose parent(s) suffered from severe chronic periodontitis.

CONCLUSIONSThe genetic heterogeneity of AgP existed in Chinese Han nationality. The genetic mode was autosomal recessive inheritance in general, and autosomal dominant inheritance could not be excluded in families whose parent(s) suffered from severe chronical periodontitis. The results imply the genetic heterogeneity of AgP, and further demonstrate that AgP was a multifactorial disease with major genetic component in the disease etiology.

Aggressive Periodontitis ; epidemiology ; genetics ; Asian Continental Ancestry Group ; genetics ; Chronic Periodontitis ; epidemiology ; genetics ; Female ; Genes, Dominant ; Genes, Recessive ; Genetic Heterogeneity ; Humans ; Male ; Pedigree ; Prevalence ; Surveys and Questionnaires

Perioperative stroke is cerebral infarction occurring in the perioperative period.The incidence of perioperative stroke in non-cardiac,and non-neurologic surgery is about 0.7％,but the mortality can be as high as 26％ to 40％.The outcome of the patients with perioperative stroke can be disastrous.Here we report a case of perioperative ischemic stroke that occurred after surgery of lumbar decompression and pedical screw fixation.A 76-year-old female admitted to our hospital because of lumbar spinal stenosis.Her medical history included hypertension and diabetes for ten years.Her personal history included a smoking history of 60 years by 2 cigarettes per day,not quitting.Her carotid artery ultrasound showed multiple low echo plaques on the right side and multiple high echo plaques on the left side of the carotid artery,but without distinct stenosis.Other examinations and tests showed no distinct abnormality.She went on a lumbar decompression and pedical screw fixation uneventfully.The blood loss was 400 mL and autologous blood transfusion 150 mL.The arterial blood pressure (ABP) maintained during 100-130 mmHg/60-80 mmHg (1 mmHg =0.133 kPa).Sixty minutes after she recovered from general anesthesia,the patient developed symptoms of slurred speech and right limbs weakness.The anesthesiologist evaluated the patient immediately with National Institute of Health Stroke Scale (NIHSS).The NIHSS score was 11 and a stroke was highly suspected.The acute stroke team was therefore initiated and fast responded.Within 4 h,digital subtraction angiography (DSA) was proceeded,which showed the M1 segment of the left middle cerebral artery was occluded and the local stenosis of her right middle cerebral artery was up to 80％.After the successful embolectomy by Solitaire stent,the left middle cerebral artery reflowed and the forward blood flow was thrombolysis in myocardial infarction (TIMI) grade 3.The patient was discharged after 33 days after the surgery with a NIHSS of 9.Our case provides an example that an acute stroke team that included the department of anesthesiology can be beneficial to the patients' perioperative strokes.During the perioperative period,anesthesiologists should be included into the acute stroke team,because anesthesiologists and anesthesia nurses might be first observers of those early onset strokes.Our case also put forward this thought that a standard peri-operative stroke evaluation tool,like NIHSS,should be discussed and applied to facilitate and accelerate the initiation of perioperative acute stroke team.