AIM:To ascertain the species status of the Anopheles dirus from Hainan Province,Chi- na.METHODS:The nucleotide sequence of the second internal transcribed spacer(ITS2 ) of PCR- amplified r DNA was determined for the An.dirusspecimens which included 2 individu- als of the species A colony (AFRIMS) from Thailand and 5 individuals from Hainan Province.RESUL TS:A841 bp fragment was amplified from single mosquito.The fragment included the ITS2 and small portions of flanking 5 .8S (96 bp) and 2 8S (2 9bp) genes.The ITS2 was71 6 bp in length,the sequence was identical for all7individuals from both Hainan Province and Thailand.No evidence of intraspecies or intrapopulation variation was detected in the ITS2 and flanking regions.CONCL USION:The result suggests the existence of An. dirus A in Hainan Province,which was in agreement with previous studies from cytogenetic analysis and egg characteristics determined by scanning electron microscopy.

Objective To investigate the protective effects of different ATP-sensitive potassium (K_ ATP) channel openers. Pinacidil and Diazoxide, on myocardium injury in isolated rabbit hearts caused by ischemia/reperfusion and the possible changes after application of ATP-sensitve potassium channel blocker, 5-HD. Methods Observation was made on rabbit hearts perfused with a Langendorff apparatus. Forty rabbits were randomly divided into five respective groups: 1. Pinacidil (Group P), 2. Diazoxide (Group D), 3. 5-HD+Pinacidil (Group HP), 4. 5-HD+Diazoxide (Group HD) and 5. the control (Group C). All groups were subjected to 40 minutes of occlusion, followed by 20 minutes of reperfusion as cardiac arrested by cold cardioplegia. Any one of Pinacidil, Diazoxide , Pinacidil or Diazoxide mixed with 5-HD was infused 15 minutes before cardioplegic heart rested in the experimental group. Cardiac tissue ultrastructure, hemodynamics variables, levels of adenine nucleotides and lipid peroxide of the myocardium were measured. Results (1) In Group P, Group D and Group HP, the recovery of myocardial contractility and heart rate after reperfusion was faster but MDA level and the amount of albumen released were lower than Group C. Moreover levels of myocardial adenosine triphosphate (ATP) were much higher compared with Group C. (2) In Group HP the recovery of myocardial contractility and heart rate was not as good as Group P, but its myocardium MDA level and amount of albumen released was higher than group P. Conclusion ATP-sensitive Potassium Channel openes may enhance myocardial protection against ischemia /reperfusion injury. The above effect of myocardial protection was only partially closed down by ATP-sensitive potassium channel blocker: 5-HD. The mitoK_ ATP channels on myocardium may be an important pathway of protection during ischemia/reperfusion process.

Objective To isolate and identify genes related to malaria parasite infection in vector mosquito, and to explore the mechanisms. Methods Anopheles stephensi infected with Plasmodium yoelii was used as tester (T) group, while uninfected but normal blood fed as driver (D) one. Engorged female mosquitoes of two groups were collected separately at 24 hours after biting. An enriched subtractive cDNA pool was generated through the course of suppression subtractive hybridization (SSH) and selective PCR amplification. The subtracted library was screened by hybridization using T and D cDNA mixture as probes, respectively. The positive clones, which produced stronger signal when probed with T than with D, were sequenced and their sequence homologues in GenBank database were searched with BLAST by internet. Results The analysis of subtraction efficiency showed that the differentially expressed genes in T comparing to in D were enriched significantly. In dot blot screening, 24 of 58 randomly selected clones (41.4%) were shown up regulation in malaria infected mosquitoes. The BLAST search of 23 genes revealed that 12 were homologous to functionally known genes, 4 were homologous to functionally unknown entries, and 7 were novel without any relatives. Nine of the 23 genes (39.1%) also hit homologous sequences in the An. gambiae EST database generated from an immune competent cell line treated with lipopolysaccharide (LPS). Conclusion An enriched cDNA pool of the mosquito genes which up regulated responsively at the early stage of malaria parasite infection was obtained. Expression screening against the pool indicated that various biochemical processes and mechanisms might be involved in the response of mosquito to parasite infection, especially those related with the innate immune system and energy metabolism.

Objective To evaluate the diagnostic value of the common test parameters in acute fever without obvious infection focus and sick appearance in children under 5 years.Methods The hospitalized children with fever duration less than 7 days, anal temperature higher than or equal to 38°C, age younger than or equal to 5 years, and without obvious infection focus and sick appearance were recruited, we investigated the diagnosis value of common test parameters including C-reactive protein (CRP), procalcitonin (PCT), the white blood cell count (WBC), and neutrophil percentage (N%) , according to the ifnal diagnostic.Results Of 228 children, 42 children (18.42%) had serious diseases, the difference of CRP, PCT between serious diseases group and non-serious diseases group were statistically signiifcant (P<0.001). The diagnostic cut-off point of CRP was 67.1 mg/L by speciifcity of 0.810 and sensitivity of 0.715, that of PCT was 0.505 ng/L by speciifcity 0.762 and sensitivity 0.672. The speciifcity and sensitivity combining CRP with PCT was respectively 0.918 and 0.617. Of 228 children, 32 children had viral infections, 40 children had bacterial infections, 15 children had mycoplasma infections. The difference of CRP, PCT, WBC, and N% among three groups were statistically signiifcant (P<0.01).The cut-off point of CRP was 38 mg/L by sensitivity 0.900 and spec-iifcity 0.813, that of PCT was 0.450 ng/L by sensitivity 0.700 and speciifcity 0.812, and the speciifcity and sensitivity combining CRP with PCT was respectively 0.965 and 0.630, to distinguish bacterial infections from viral infections. The diagnostic cut-off point of CRP was 80.75 mg/L by sensitivity 0.700 and speciifcity 0.933 distinguishing bacterial infections from mycoplasma infections.Conclusions The parameters CRP and PCT have the diagnostic value for the children with the acute fever and age younger than or equal to 5 years and without obvious infection focus and sick appearance in etiology and serious diseases, espe-cially the value of combining CRP with PCT is better.

Objective To observe the effects of hawthorn leaf procyanidins (HLP) on over expressions of ICAM-1 and E-selectin in human umbilical vein endothelial cells (HUVEC) induced by TNF-α,and clarify the mechanism of HLP's anti-inflammation effect. Methods HUVEC were cultured in vitro. MTT assay was used to detect cell viabilities. The expressions of ICAM-1 and E-selectin in HUVEC were detected by flowcytometry. Results Up to 200 mg/L, HLP showed no significant decrease in cell viabilities; the expression levels of ICAM-1 and E-selectin in the model group significantly increased, compared with that in the normal group; 10, 20, 30, 40, 50 mg/L HLP inhibited the expression elevations of ICAM-1 and E-selectin in concentration-dependent manner; and there were statistical significances in 40, 50 mg/L HLP groups, compared with the model group. Conclusion HLP can inhibit the over expressions of ICAM-1 and E-selectin of vascular endothelial cells induced by TNF-α, which possibly underlies HLP's anti-inflammation effect.

Objective To investigate the differential expression of miR-155 in newly diagnosed pediatric acute myeloid leukemia(AML) and its clinical significances.Methods Fifty-two AML children and 30 non-malignant disease matched children were recruited as the controls.The preliminary AML children were divided into favorable group,moderate group and poor group according to the National Comprehensive Cancer Network(NCCN) 2013.Real-time quantitative polymerase chain reaction was applied to validate the expressions of miR-155 in bone marrow samples (the data presented by 2-△△Ct).Results By comparing expressions of miR-155 between AML patients and controls,the miR-155 expressions were significantly higher in the AML children than those in the controls (Z =-5.391,P ＜ 0.001).There were significant differences among different prognostic groups,with a significantly lower level in the favorable group compared with others (x2 =12.586,P =0.002).It was also found that differential expressions existed not only in kinds of mutation cohort,with the highest level in FLT3-ITD and the lowest one in FLT3-TKD mutation group (x2 =11.216,P =0.024),but also among fusion gene subgroups (x2 =12.254,P =0.016),with the highest level in AML-ETO group and the lowest level in PML-RARa group:meanwhile,the expressions of miR-155 were statistic different according to French-America-British (FAB) subtypes (x2 =17.814,P =0.013),which was lower in M3 patients than non-M3 patients (Z =-3.291,P =0.001).Conclusions It indicates that the expressions of miR-155 may increase sharply in preliminary AML children,and the lower expression of miR-155 is closely related to favorable prognosis.

Objective To investigate the inhibitory effects of RNA interference on expression of matrix metalloproteinase-2(MMP-2)gene and invasiveness of human pancreatic cancer cell line PANC-1 in vitro.Methods Small interference RNA targeting MMP-2 gene was designed and constructed to plasmid pGCsi-U6.Recombinant plasmids were transfected to pancreatic carcinoma PANC1 cells with Lipofectamine 2000.The efficiency of transfection was evaluated by flow cytometry.RQPCR was used to detect the expression of MMP-2 mRNA.The expression of MMP-2 protein was determined by ELISA.The invasiveness of PANC-1 cells was measured by transwell chamber experiment.MTT assay was used to detect the proliferation and growth of PANC-1 cells.Results Sequencing confirmed that the MMP-2 siRNA plasmid was successfully constructed.The best efficiency of transfecting recombinant plasmid was 82.1%.After transfection of the MMP-2 siRNA plasmid, the MMP-2 gene expression of PANC-1 cells was suppressed to 71.74 %(P＜0.05),and protein expression of MMP-2 fell to 49.82%(P＜0.05).The corresponding inhibition ratio of invasiveness was 33.0%(P＜0.05).There was no marked difference in proliferation rate measured by MTT assay among different groups(P＞0.05).Conclusions RNAi targeting MMP-2 can suppress invasiveness of PANC-1 cells in vitro.This suggests that MMP-2 could be a target for gene therapy of pancreatic carcinoma.RNAi is expected to open up a new prospect for tumor therapy.

Objective To evaluate the expression of miR-196b in newly diagnosed pediatric acute myeloid leukemia (AML) and its clinical signiifcance. Methods Fifty-two AML children were enrolled in this study and 30 non-leukemia com-pared children were selected as controls. The expressions of miR-196b were detected in bone marrow samples by real-time quan-titative PCR (q-RT-PCR) and the results were expressed in 2-??Ct. Results miR-196b expressions were signiifcantly higher in M4-5 and lower in non-M4-5 of AML children than those in control (P<0.01), with a lowest level in t (15;17) and a highest level in MLL subtypes (P<0.01). The miR-196b expressions were signiifcantly different among different prognosis groups (P<0.01) and the level in the favorable prognostic group was lower than in poor prognosis group. It was also found that miR-196b expres-sion was lower in remission group than that in no-remission group after the ifrst induction remission therapy (P<0.05). Mean-while, the expression of miR-196b in the children with WBC≥100×109/L were statistically higher than that in the children with WBC<100×109/L (P<0.01), and miR-196b level was positively correlated with the platelet counts (r=0.302, P=0.030). Conclu-sions miR-196b expression is increased in poor prognosis group of AML children, and high expression of miR-196b is related with low response rate and poor prognosis. miR-1966 is expected to become a new target for the treatment of AML.

Objective To evaluate the accuracy of sentinel lymph node biopsy (SLNB) in predicting the axillary lymph node status in breast cancer patients undergoing a breastectomy. Methods From Mar 2002 to Jun 2003,patent blue dye and/or 99m Tc-Sc were used to detect sentinel lymph nodes,samples were sent for fast pathology.Three patients in which the SLN were judged as negative by both the two tests were freed of futher axillary lymph node dissection (ALND). Results SLN were successfully identified in 78 of 81 (96.3%) patients. The accuracy rate of SLNB to predict axillary lymph node status was 97.5%,with 9.7% false negative rate. The success rate,accuracy rate and false negative rate by blue dye only and by a combination of two above methods were 92.5%,94.2%,15.8%,and 100%,100%,0, respectively . Conclusions SLNB accurately predict the axillary status in clinically node negative breast cancer patients .

Objective: To observe the correlation between Nε-carboxymethyl-Lysine (CML), the main component of advanced glycation end products and the calcification of the anterior tibial artery plaque in patients with diabetic foot post foot amputation. Methods: Sixty patients hospitalized for foot amputation operation due to diabetic foot from June 2012 to June 2016 in the Department of Orthopedics, Affiliated Hospital of Jiangsu University were prospectively recruited.The patients were categorized into mild stenosis (0n=20), moderate stenosis (50%≤stenosis<70%, n=20) and severe stenosis (70%≤stenosis≤100%, n=20) based on the color Doppler ultrasound assessed severity of anterior tibial artery stenosis.The baseline clinical data of patients were collected and anterior tibial artery was isolated.Then, HE staining, O-Cresolphthalein Complexone method, enzymic method and ELASA analysis were then performed to detect the evolution of calcification, arterial calcium content, alkaline phosphatase activity and serum CML concentration, respectively. Results: The results from both color Doppler ultrasound scan before amputation and HE staining after amputation showed that echo intensity as well as spotty blue calcium particles of anterior tibial artery plaque increased significantly in proportion to degree of stenosis and destructed elastic plate of the arterial wall was evidenced in patients with severest stenosis.The content of calcium ((2.3±0.9), (3.9±1.3), (6.6±1.7) μmol/mg, respectively, P<0.001), ALP activity ((102.4±39.4), (202.3±73.4), (483.7±117.9) U/mg, respectively, P<0.001) and serum CML level ((28.9±4.4), (37.9±5.3), (57.3±7.1)μg/L, respectively, P<0.001) increased significantly in proportion to stenosis severity.Pearson correlation analysis showed that serum CML level was positively correlated with the content of calcium (r=0.749, P<0.001) and ALP activity (r=0.923, P<0.001), respectively. Conclusions Serum CML level is positively correlated with the calcification of anterior tibial arterial plaque in patients with diabetic foot and could be used to evaluate the calcification of anterior tibial arterial plaque and stenosis degree of anterior tibial arterial in these patients.