Objective:To explore the influence factors on amplification of single cell duplex-nested PCR.Methods:The mutational loci region CD41-42 and IVS-Ⅱ654 of ?-globin gene were amplified by duplex-nested PCR with different combination of primers concentration, different Taq DNA polymerases, different neutralization buffers and with or without predenaturation at 98 ℃ before the PCR amplification in single lymphocyte or single blastomere, thus, to investigate the influence of these factors on the amplification efficiency of PCR.Results:TaKaRa EX Taq was the most efficient Taq DNA ploymerase among different Taq DNA ploymerases; primer pair R1+F1 at final concertration of 0.25 ?mol/L and R2+F2 at 0.3 ?mol/L were the most efficient ones in amplification among different combinations of primers concentrations; the amplification efficiency in neutralization buffer-1 (200 mmol/L Tricine) was obviously higher than that of neutralization buffer-2 (900 mmol/L Tris-HCl, pH 8.3/300 mmol/L KCl/200 mmol/L HCl)(P0.05). Conclusion:There were remarkable differences of the amplification efficiency of single cell duplex-nested PCR while using different combination of primers concentrations, different Taq DNA polymerases, different neutralization buffers. However, predenaturation at 98 ℃ before the single cell PCR amplification could not improve the PCR amplification efficiency

Objective: To study the distribution of flavonoids among the plants for medicinal use of Selaginella . Methods: HPCE analysis was carried out on the ethyl acetate extracts of 18 Selaginella plants (from 11 species), using MECC separation model. Results: HPCE fingerprints of 18 Selaginella plants were established with quercetin as interanl standard and relative migration time and relative peak area as parameters. Conclusion: The contributions of flavonoids has not only generity, but also difference among the tested samples.

AIM: To isolate targeted the peptides that binding and internalizing into endometrial carcinoma cell line EAC. METHODS: The tumor cells were screened five rounds of whole cell screen through the Ph.D.-12 phage display library. Analysis of the displayed peptides specific binding efficiency to the tumor cells was proceeded. The DNA of phages was extracted, sequenced and translated to the sequences of amino acid and analysis using computer software. RESULTS: After five biopannings, the isolated phages showed high specificity and strong affinity for their cognate cell types relative to different cell lines. Through sequencing, the sequences of displayed peptides were obtained. CONCLUSIONS: Whole cell screening against EAC cells through random phage peptide library can obtain phage peptides with a highly tissue specific binding and internalizing ability. The peptides do provide a basis for tumor's targeted delivery as therapy vector.

Objective To study the effect of schistosomiasis transmission interruption model based on intensive agriculture in hilly endemic areas,so as to provide the reference for the similar endemic areas. Methods Based on the development of in-tensive agriculture in Guanghan City,a comprehensive demonstration area of schistosomiasis control with measures such as new rural construction,hardening ditches,the adjustment of industrial structure and water remediation measures was constructed. Ji-nhua,Shiguan and Hongyan villages were chosen as the evaluation sites to comparatively analyze the indexes of intensive agri-culture and schistosomiasis control effects. Results Compared with the demonstration area before construction,in 2014,the harden rates of ditches and village roads were increased by 49.57％and 39.33％respectively;and the proportion of agricultural machinery increased by 25％. The positive rate of serological tests of schistosomiasis was decreased by 81.74％. The Oncomela-nia hupensis snail area was decreased from 2.44 hm2(2007)to 0(2014). The awareness rate of schistosomiasis control knowl-edge and correct behavior rate of the residents were increased from 51.28％ and 90.85％ to 91.29％ and 97.69％ respectively. The experience of the demonstration area ensured the entire Guanghan District achieved the schistosomiasis transmission inter-ruption criterion at the end of 2014. Conclusions The schistosomiasis control model of intensive agriculture combined with oth-er comprehensive measures has a good effect on interrupting the endemic of schistosomiasis,and it can realize the sustainable development of the agricultural economy and schistosomiasis control.

AlM:To compare the effect of different incision in corneal astigmatism after phacoemulsification.
METHODS: Totally 88 cases ( 122 eyes ) with pure cataract were randomly divided into two groups. Forty cases (60 eyes) were clarity corneal incision in group A, and 48 cases ( 62 eyes ) were sclera tunnel incision in group B. Mean corneal astigmatism, surgically induced astigmatism ( SlA ) , uncorrected visual acuity ( UCVA ) and best correct vision acuity ( BCVA ) were observed in pre- and post-operation at 1d;1wk;1mo.
RESULTS: The mean astigmatism had statistically significant difference between two groups at 1d; 1wk;1mo after operation(P<0. 05). The SlA had statistically significant difference at 1d ( P<0. 05 ); The SlA had no statistically significant difference between two groups at 1mo after operation (P>0. 05). UCVA≥0. 5 and BCVA≥0. 8 had statistically significant difference at 1d; 1wk ( P<0. 05) . There had no statistically significant difference at 1mo after operation (P>0. 05).
CONCLUSlON: Phacoemulsification with scleral tunnel incision remove combined intraocular lens ( lOL ) implantation has small changes to corneal astigmatism. By selecting personalized corneal incision according to the corneal topography might be more beneficial.

Objective To investigate the current status of lead(Pb) poisoning in children and correlations between Pb levels and calcium(Ca), iron(Fe), zinc(Zn) and environmental factors. Methods ①Questionnaire survey was conducted among 3160 parents in Qingdao area, regarding factors that might have effects on the levels of Pb. ②Levels of blood Pb, Ca, Fe and Zn were determined by graphite furnace atomic absorption (GFAA) spectrometry for 3160 children, who were divided into three groups: mildly increased Pb levels, moderately increased Pb levels and severely increased Pb levels, and the effects of Ca, Fe and Zn on children's blood Pb levels were analyzed by multiple linear regressions.③Children having mildly Pb increase were divided into four groups and received Ca, Fe or Zn preparations or placebo. Results ① Children living with smokers, in new houses, in houses or kindergartens located in industrial areas or main traffic roads, consuming preserved eggs or canned drinks regularly were more easily poisoned.②Percentages of children with increased levels who had insufficient blood Ca, Fe and Zn : 53.31% (306/574), 46.17% (265/574) and 68.47% (393/574), respectively, in the group of children with mildly increased Pb levels; 57.68% (263/456), 49.34% (225/456) and 77.41% (353/456), respectively, in the group of children with moderately increased Pb levels; 64.71% (11/17), 58.82% (10/17) and 82.35%(14/17), respectively, in the group of children with severely increased Pb levels. In contrast, the percentages of children in the control group who had low blood Ca, Fe and Zn were 19.78%(418/2113), 5.21% (110/2113) and 14.62%(309/2113), respectively. ③The levels of Ca, Fe and Zn had a negative correlations with blood levels of Pb(coefficient of regression were-0.870,-0.430,-0.565, respectively). ④When given Ca, Fe and Zn, the children had the blood Pb levels decreased by 78.32% (112/143), 66.43% (95/143) and 67.83% (97/143), respectively. Conclusions ①Children's increased levels of Pb have adverse effects on the absorption of Ca, Fe and Zn. ②Intaking Ca, Fe and Zn preparations can reduce blood Pb level.

?AIM: To discuss the application of anterior segment optic coherence tomography ( AS-OCT ) in the diagnosis of corneal ulcer.?METHODS: The cross linear scan was used in 88 patients ( 88 eyes ) with corneal lesion by AS-OCT to gather the image data, observe the pathological changed tissue by measuring all layers for patients with initial inspection, providing important visual images and data for treatments. All the patients were followed up for 2mo.?RESULTS:Clear images with structure of all layers were obtained. It can provide the intuitive image data and scan the same position and show the changes during the treatment.?CONCLUSION: AS-OCT can discover the important condition immediately. It also can monitor course of disease dynamically, provide the intuitive image data for clinical treatment.

To study the effect of miR-490 on Coxsackievirus B3 (CVB3) replication, HeLa cells were trans- fected with miR-490 in vitro, and infected with a Renilla luciferase (RLuc)-expressing CVB3 variant (RLuc-CVB3). The activities of RLuc in these cells were measured at 8h intervals from 0 to 40 h post-infection (p.i.), and the effects of miR-490 on RLuc-CVB3 replication were observed. In a further study, HeLa cells were transfected with either miR-490 or antisense miR-490 (AMO-miR-490), and were then infected with an enhanced green fluorescence protein (EGFP)-expressing CVB3 variant (EGFP-CVB3). The replication of EGFP-CVB3 was then determined by detecting the expression of EGFP. We observed that miR-490 could significantly inhibit the expression of RLuc in infected cells at 32 h p. i. Furthermore, in HeLa cells infected with EGFP-CVB3 at 32 h p.i., EGFP expression was also significantly inhibited by the presence of mniR-490. The inhibitory effect of miR-490 on EGFP expression in EGFP-CVB3-infected cells could be reversed by tranfection with AMQ-miR-490. These results indicated that miR-490 significantly inhibits the replication and expression of QVB3.
Enterovirus B, Human ; genetics ; physiology ; Enterovirus Infections ; genetics ; metabolism ; virology ; HeLa Cells ; Humans ; MicroRNAs ; genetics ; metabolism ; Virus Replication

OBJECTIVE: To investigate the expression of HPA, CK2beta and HIF-1alpha gene in nasopharyngeal carcinoma (NPC) tissues, and the correlation between their expression with the clinical characteristics of NPC and the relativity of HPA, CK2beta and HIF-1alpha gene in NPC tissues. METHOD: HPA, CK2beta and HIF-1alpha were detected with Super-Vision immunohistochemical method using antibody in 49 NPC specimens and 30 specimens with chronic nasopharyngitis tissue (CNT). RESULT: The expression of HPA, CK2beta and HIF-1alpha in NPC tissue were significantly higher than those in CNT tissue (P<0.05, separately). The expression of HPA, CK2beta and HIF-1alpha were significantly related to the TNM stage and whether recurrence or metastasis occur after treatment (P<0.05, separate ly), but there was no obvious correlation between its expression and the sex of NPC patient (P>0.05). The expression of HIF-1alpha was significantly related to the age of NPC patient (P<0.05), while HPA, CK2beta were not. The expression of HPA, CK2beta and HIF-1alpha in NPC tissue was positively correlated with each other (P<0.05, separately). CONCLUSION HPA, CK2beta and HIF-1alpha play synergetic role in development of NPC, which plays an important role in invasiveness,recurrence and metastasis of NPC. There could be a positive cooperation among HPA, CK2beta and HIF-1alpha in the carcinogenesis and development of NPC.
Carcinoma ; Casein Kinase II ; metabolism ; Female ; Heparin Lyase ; metabolism ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Male ; Middle Aged ; Nasopharyngeal Carcinoma ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Neoplasm Staging

BACKGROUND: The incidence of ankylosing spondylitis (AS) presents a trend of rising year by year, accompanied by a higher disability. Therefore, fully understanding the related factors to the development of AS is of great significance to develop a rational treatment scheme.OBJECTIVE: To evaluate the expression levels of Dickkopf-1 and sclerostin in Wnt signaling pathway as well as vascular endothelial growth factor A (VEGF-A) in the AS patients, and to evaluate their correlation with AS progression.METHODS: AS patients and healthy volunteers were recruited, and their baseline data and medical history were collected. The expression levels of Dickkopf-1, sclerostin, VEGF-A and C-reactive protein in the peripheral blood were detected; the AS progression was evaluated by Bath AS Disease Activity Index and Bath AS Functional Index; the imaging performance was assessed by modified Stroke AS Spine Score; the correlation of Dickkopf-1, sclerostin and VEGF-A with AS progression, imaging performance and inflammatory reaction was analyzed by Spearman's rank correlation analysis and multiple linear regression analysis.RESULTS AND CONCLUSION: No significant differences were found in the expression levels of Dickkopf-1, sclerostin and VEGF-A between two groups (P > 0.05). The AS patients without syndesmophyte and with higher erythrocyte sedimentation rate and C-reactive protein level appeared with an significant increase in the Dickkopf-1 level (P < 0.05),which was significantly related to sclerostin level (r=0.592, P=0.000). The As patients with the history of smoking,increase in erythrocyte sedimentation rate and C-reactive protein, as well as higher Bath AS Disease Activity Index and Bath AS Functional Index presented with a higher level of VEGF-A. Multiple linear regression analysis showed that erythrocyte sedimentation rate, C-reactive protein, syndesmophyte and sclerostin level were the independent factors affecting the Dickkopf-1 level (P ≤ 0.001); the history of smoking, erythrocyte sedimentation rate, and C-reactive protein were the independent factors affecting VEGF-A (P < 0.005). These results suggest that in AS patients, the Dickkopf-1 level is related to syndesmophyte and systemic inflammatory response, while the history of smoking affects VEGF-A level. Therefore, all above indicators can be used to evaluate osteophyte formation and bone mass loss.