Aged ; Humans ; Male ; Palatal Neoplasms ; pathology ; Uvula ; pathology

Objective To clone and express the alkaline protease AprA , one important virulence factor secreted by Pseudomonas aeruginosa(PAE)in Escherichia coli, to clone and express the inhibitor of AprA (AprI) and its substrate flagellin , and to detect the function of AprA and the inhibitory function of AprI .Methods The genes encoding AprA ,AprI and flagellin gene were amplified respectively by PCR using PAE PAO 1 genome DNA as the template .The expression vec-tors (pET-28a-AprA, pET-28a-AprI and pET-28a-Flagellin) were constructed and transformed into E.coli BL21(DE3) respectively.The recombinant AprA protein was expressed by IPTG induction and purified via denaturing and renaturation. The recombinant AprI and flagellin were expressed and purified by Ni 2+affinity chromatography .The cleavage activities of AprA on flagellin were detected by SDS-PAGE.Results Recombinant AprA , AprI and flagellin protein were expressed and purified .It was demonstrated that AprA cleaved flagellin , which was blocked by AprI .Conclusion Recombinant AprA could cleave its substrates as an alkaline protease , and its inhibitor AprI inhibits the activities of AprA .This study will contribute to further investigations on the role of AprA in the pathogenesis of PAE .

Objective To express the beta hemolysin ( Hlb), an important toxin secreted by Staphylococcus aureus ( S.aureus) and the mutant protein Hlb H-149-N , to detect the hemolytic activity of Hlb and Hlb H-149-N on sheep erythrocytes , and to prepare the specific antibodies against Hlb which can inhibit the hemolytic activity of Hlb .Methods Hlb gene was amplified by PCR using S.aureus NCTC-8325 genome DNA as template.The expression vector pET-28a-hlb was constructed and transformed into E.coli BL21(DE3).The expression vector pET28a-hlbH-149-Nwas constructed through point mutation.The recombinant Hlb and Hlb H-149-N protein were expressed and purified by Ni 2+affinity chromatography .The hemolytic activity of Hlb and Hlb H-149-N was measured by sheep erythrocyte lysis assay .Results Recombinant Hlb protein and the mutant were obtained .Further investigations showed that Hlb could significantly induce the lysis of SRBC while HlbH-149-N could not.The specific polyclonal antibodies against Hlb (anti-Hlb) were prepared.It was found that anti-Hlb recognized Hlb and Hlb H-149-N .Moreover , it was found that anti-Hlb blocked the hemolytic activity of Hlb .Conclusion The recombinant Hlb protein with high hemolytic activity and Hlb H-149-N without hemolytic activity are obtained while its neutralized antibody is pepared .Hlb from S.aureus has different hemolytic effects on erythrocytes from various species .Our findings will facilitate the investigation on the role of Hlb in the pathogenesis of S.aureus.

Objective To clone and express alpha hemolysin ( Hla ) , one important virulence factor secreted by Staphylococcus aureus in Escherichia coli to tdetect the hemolytic activity of Hla on erythrocytes from diffrerent species ,and to prepare specific antibodies against Hla that can inhibit the hemolytic activity of Hla .Methods Hla gene was amplified by PCR using S.aureus NCTC-8325 genome DNA as a template.The expression vector pET-28a-Hla was constructed and transformed into E.coli BL21(DE3).The recombinant Hla protein was expressed by IPTG induction ,and then purified by Ni2+affinity chromatography .The hemolytic activity of Hla was measured by erythrocyte lysis assays .Results Soluble recombinant Hla protein was expressed and purified .Further investigations showed that Hla could significantly induce the lysis of rabbit erythrocytes .However , erythrocytes from humans or sheep were more resistant to the lysis mediated by Hla . The specific polyclonal antibodies against Hla ( anti-Hla) were prepared and detected by ELISA .Moreover, it was found that anti-Hla could inhibit the hemolytic activity of Hla .Conclusion We found that Hla from S.aureus has different hemolytic effects on erythrocytes from various species .The prepared Hla-antibodies can specifically block the hemolytic activity of Hla.

Objective To study the susceptible factors of atherosclerotic stenosis before the coronary artery myocardial bridges.Methods The data from 88 myocardial bridge cases which received coronary angiography were statistically analyzed.Sixty-seven cages which suffered from atherosclerofic stenesis in proximal of the myocardial bridges were recruited into group A,and the other 21 cases which suffered no atherosclerotic stenosis or from atherosclerotic stenosis in distal of the myocardial bridges were mcmited into group B.Difference of the age,gender,length of myocardial bridge,systolic blood pressure (SBP),diastolic blood pressure (DBP),pulse pressure (PP),the oppression degree of myocardial bridge (Nobel classification),fasting plasma glucose (FPG),and blood fat,and so on,in two groups,were observed and statistically analyzed.Results The difference of the Nobel classification,SBP and PP in two groups showed a statistical significance (P＜0.05).While the difference of the age,gender,length of myocardial bridge,DBP,FPG,total cholesterol,low-density hpoprotein in two groups showed no statistical significance (P＞0.05).A further regression analysis suggested that Nobel classification and PP had a correlation with the comphcation of stonosis before the myocardial bridge (r=3.0569,0.9740,P＜0.05).Conclusions High blood pressure cases are liable to suffer from myocardial bridge.Myocardial bridges themselves trend to promote or accelerate the atherosclerotic stenosis of the coronary arteries before.them.The oppression degree of myocardial bridge and PP has a correlation with the complication of stenosis before the myocardial bridge,while has no correlations with age,gender,bloodfat,SBP,DBP,FPG,length of myocardial bridge,and so on.

Objective To investigate the effects of cryopreservation on the growth and osteogenesis capa- bility of human bone marrow stromal cells(BMSCs)on demineralized bene matrix(DBM).Methods Bone marrow aspirates were obtained from the lilac crests of three donors.The BMSCs were isolated from the bone marrow by density gradient centrifugation.Cells of passage 3 were cryopreserved in liquid nitrogen for 24 hours,and then re- covered.The non-cryopreserved BMSCs were used as the control,The cryopreserved and control BMSCs were cul- tured in osteogenic media,collected and labeled with Dil to be seeded onto the DBM when cells were confluent.The percentage of BMSCs adhered to the DMB was detected.The cell morphology and matrices secreted by BMSCs on the DBM were observed by the inverted phase-contrasted microscope,fluorescence microscope and scanning electron microscope(SEM).The growth and viability of BMSCs on the DBM were determined using the modified MTT ashy. The osteogenesis ability of BMSCs on the DBM was determined by assessment of the alkaline phosphatase(ALP) activity and osteocalcin(OCN)content.Results The percentages of the cryopreserved and control cells adhered to DBM were(97.25?1.17)% and(97.00?1.09)% respectively.The cells adhered well to the DBM and grew rapidly.Large amounts of matrices on the DBM were observed by the light microscope and SEM.The cells embedded in the matrices could be observed by fluorescence microscope.There were no significant differences in the assay values of MTT,ALP and OCN between the cryopreserved and control BMSCs on the DBM.Conclusion Since cryopreservation does not affect the growth and osteogenesis capability of BMSCs on DBM,the cryopreserved BMSCs can be used as a cell source in bone tissue engineering.

Sulfate-reducing bacteria (SRB) are widespread in the environment. SRB are obligate anaerobes and capable of dissimilatory reduction of sulfate. SRB have application prospects in the control of environmental pollution due to that many pollutants can be removed by SRB. The biological characteristics and metabolic mechanisms of SRB are introduced, and the application of SRB in the treatment of environmental pollution is described in this paper. The research progress of Cr(Ⅵ ) reduction and Cr(Ⅵ ) removal from wastewater by SRB is reviewed, and future direction of research on the control of Cr(Ⅵ ) pollution by SRB is also analysed.

Objective To analyze the CT features of renal cell carcinoma,so as to improve the diagnostic accuracy of renal cell carcinoma.Methods Three hundred cases of renal cell carcinoma proved by pathology were examined by means of CT.There were 214 male and 86 female in this group.Their age ranged from 9 to 81 years,with a mean of 53.7 years.Their CT features were retrospectively reviewed. Results The masses were 1.5—16.0 cm(mean,4.8 cm)in greatest dimension,125 masses on left kidney and 175 masses on right kidney.According to WHO histological classification of tumours of the kidney in 2004,there were 238 cases of clear cell renal cell carcinoma,6 cases of multilocular clear cell renal cell carcinomas,23 cases of papillary renal cell carcinoma,14 cases of chromophobe renal cell carcinoma and 19 cases of renal cell carcinoma,unclassified.The above subtype of renal cell carcinoma demonstrated characteristic features.Clear cell renal cell carcinoma exhibited inhomogenous(due to hemorrhage,necrosis or cystic degeneration)and hypervaseular.Multilocular clear cell renal cell carcinoma presented as a multilocular cystic mass lacking an,expansile nodule,and with regular thin cyst wall and septa.Papillary renal cell carcinoma exhibited inhomogenous and hypovascular.Chromophobe renal cell carcinoma was relatively homogenous and hypovascular.Renal cell carcinoma,unclassified showed inhomogenous and hypervascular,and was more invading growth compared to clear cell renal cell carcinoma. Conclusion Common subtype of renal cell carcinoma demonstrated characteristic features in CT and it is helpful for differentiation.

Objective To obtain the fusion genes of several human orphan G protein coupled receptors (oGPCRs) with Gi1α subtype of G protein and their expression system. Methods The whole open reading frames of GPR45, GPR85, GPR174 and Gilα were cloned by RT-PCR from HepG2 cDNA separately,and the corresponding fusion genes were amplified by overlap extension PCR. Then, the fusion genes-containing pBacmids were successfully constructed with the Bac-to-Bac baculovirus expression system indicated by specific transposition and virus recombination. The insect Sf9 cells were transfected with pBacmid-oGPCRs-Gi1α, and the supernatant containing recombinant virus was harvested. With the supernatant, insect Sf9 cells were infected under an optimized condition (MOI=5, infection time=72 h) and the fusion proteins were prepared and detected by Western blotting.Results The three fusion genes of GPCR45, GPR85 or GPR174 with Gi1α were obtained. The corresponding fusion proteins could be properly prepared in Sf9 cells.Conclusion Human oGPCRs could be fused with Gilα, and the fusion genes could be expressed using the Bac-to-Bac baculovirus expression system in insect Sf9 cells.

Objective To investigate the diagnosis value of immunophenotype and karyotypes in newly diagnosed chronic lymphocytic leukemia (CLL).Methods To retrospect the flow cytometry (FCM) immunophenotype and karyotypes characteristics in newly diagnosed 70 CLL cases.Results In all cases,the positive rates of CD19,CD20,CD5,CD23,CD22 were 100 ％,88.5 ％ (54/61),77.1％ (54/70),67.6 ％ (23/34)and 51.9 ％ (27/52),respectively.And 6 were misdiagnosed,2 was CD+5CD+19(+),but CD20,CD22 were strongly positive,final diagnosed as mantle cell lymphoma (MCL) by FISH t(11;14) examination and CyclinDl; CD+5CD+19(-) CLL were 16 cases (22.9 ％),but 4 were misdiagnosed,the misdiagnosis rate was 25 ％,significantly higher than that of CD+5CD+19(-) CLL (P =0.030).59 cases were examined by conventional cytogcnetic (CC),and 13 cases were with abnormal karyotypes,positive rate was 22.0 ％,with complex karyotypes in 5 cases (8.5 ％); 10 cases combined with FISH abnormalities karyotype examination rate was 60 ％.Conclusion Typical CLL immunophenotypic characteristics were CD5,CD19,CD23 co-expression,and CD－5 CLL with higher misdiagnosis rate,combined with CD20 (,) CD22 expression and karyotype analysis helps to CLL and other B lymphoid proliferative diseases (B-LPD) identification.Conventional cytogenetic detection combined with FISH scan can improve the recognition ability of abnormal chromosome.