Objective To investigate the construction of compound membrane with corneal stromal cells and collagen-chitosan by tissue engineering technique and its biocompatibility.Methods Rabbit and human corneal stromal cells were separated and seeded into collagen-chitosan membrane.The compound membrane was transplanted into rabbit corneal stroma.Then the growth condition of keratocytes,the effect on normal keratocytes and degradation of compound membrane were detected by corneal confocal microscope,anterior OCT and histological and immunohistochemical methods ex vivo 1,2,4 weeks after grafting.Results The rabbit and human corneal stromal cells grown well in collagen-chitosan scaffold.The compound membrane degradated gradually after grafting.There was no necrosis and dissolvation.Corneal epithelium,stroma and endothelial cells were all normal.Conclusion Collagen-chitosan can be used as a biological scaffold for construction of corneal stroma.Corneal confocal microscopy and anterior OCT are new methods to observe the biological activity of constructed corneal stroma.

OBJECTIVE:To study the effects and mechanism of rapamycin on invasion and metastasis of cervical cancer HeLa cell. METHODS:HeLa cells were divided into control group and rapamycin low-dose,medium-dose and high-dose groups (10, 30,100 nmol/L). After treated for 48 h,cell viability was measured by MTT assay,and inhibitory rate was calculated;migration and invasion of cell was tested by Transwell assay. The expression of matrix metalloproteinase 2(MMP-2),MMP-9,Vimentin and E-cadherin,and phosphorylation of protein kinase B (Akt),mammalian target of rapamycin (mTOR) were detected by Western blot. RESULTS:Compared with control group,the inhibition rate of cell viability was increased in rapamycin groups(P<0.01);the number of invasion and metastasis cells decreased(P<0.01);the expression of MMP-2,MMP-9 and Vimentin were decreased (P<0.01 or P<0.05);the expression of E-cadherin was enhanced(P<0.01 or P<0.05);the phosphorylation of Akt and mTOR were reduced (P<0.01). CONCLUSIONS:Rapamycin could inhibit invasion and metastasis of HeLa cell via Akt/mTOR signal pathway.

Objective To investigate effect of folic acid combined with xin funing on CRP, HGF, IL-2,TNF-αof patients with cervical cancer caused by human papillomavirus.Methods 80 cases of cervical cancer patients were randomly divided into control group, 40 cases in the control group were given conventional chemotherapy, 40 cases in the experimental group were on the base of the control with folic acid combined with xin funing.CRP, HGF, TNF-α, IL-2 and T lymphocyte subsets were compared before and after the treatment.Results Compared with the control group, the serum CRP, HGF and TNF-αof the experiment group were lower(P<0.05), IL-2 levels was higher (P<0.05), CD4 +and CD4 +/CD8 +level were higher(P<0.05), level of CD8 +was lower(P<0.05) and the clinical effective rate were higher(P<0.05).Conclusion Folic acid combined with Xin Funing has important significance for the treatment of patients with cervical cancer.It is speculated that the mechanism may be to reduce the level of serum CRP and HGF in patients with cervical cancer, and to increase the level of IL-2, and to regulate immune cells.

Objective To establish the fingerprints analysis of the methanol extracts of nutmeg,and study quality uniformity of nutmeg in different areas.Methods A ZORBAX EclipseXDB-C18 (4.6 mm?150 mm,5 ?m) column was used.The mobile phase consisted of methanol-acetonitrile-water (25∶35∶40),the flow rate was 1 mL/min,the column temperature was 30 ℃,the detective wavelength was 270 nm.Dehydrodiisoeugenol was used as reference compound.Results Fingerprint of nutmeg was established,consisted of l7 common peaks.The similarity of fingerprints was over 0.9.Conclusion The fingerprints of nutmeg in different areas have no differences.This method is accurate,reliable and provides a scientific basis for the quality control of nutmeg.

ObjectiveTo observe β cell function in newly diagnosed diabetic patients with ketosis before and in remission period and evaluate its classification and predictive value.MethodsA total of 206 patients newly diagnosed as diabetic ketosis who had been treated with intensive insulin therapy in our hospital and entered in the honeymoon after the withdraw of insulin therapy were followed for 36 months from onset of diabetes.They were divided into two groups of type 1 and type2 diabetes ( group A and B),according to the dependence or independence on insulin treatment. The β cell function of the two groups before and in remission period was compared by oral glucose tolerance test (OGTF).β cell function was measured with the AUC of insulin and C-peptide and homeostatic model assessment β-cell function (HOMA-β),while homeostatic model assessment insulin resistant (HOMA-IR) for insulin resistant.The duration of the honeymoon and the change of insulin and C-peptide curve before and in honeymoon were also observed.ResultsThe AUC of insulin and C-peptide,the HOMA-β and the HOMA-IR before and after the intensive insulin treatment were lower in group A than that in group B [ before the insulin treatment:(10.18 ±2.36)mIU · h · L-1 vs (20.28 ±6.89)mIU · h · L-1,(1.56 ±0.53) μg · h · L-1 vs (3.75 ±0.67) μg · h · L-1,3.68 ± 1.08 vs 18.20 ±6.59,1.22 ±0.49 vs 3.06 ± 1.54,respectively;after the insulin treatment:(29.86 ± 8.65 ) mIU · h · L-1 vs (93.35 ± 19.42 ) mIU · h · L-1,( 3.99 ± 0.79 )μg · h · L-1 vs ( 12.54 ±3.83) μg · h · L-1,8.50 ±2.46 vs 56.17 ± 19.42,0.63 ±0.56 vs 1.42 ±0.78,respectively ].The duration of the honeymoon in group A was significantly shorter than in group B [ (7.9 ±5.2) months vs (20.9 ± 9.9 ) months ].In oral glucose insulin and C-peptide release test,the peak of insulin and C-peptide releasing curve in group A was brought forward by a half to 1 hour after intensive treatment while delayed in group B by 1 or 2 hours.The releasing peak of insulin and C-peptide in group A was less than two folds of the basic value,while four to ten fold of the basic value in group B.The positive ratio of glutamic acid decarboxylase antibody,insulin autoantibody and insular cellular antibody in group A and group B were 21.2％ vs 4.8％,18.1％ vs 3.3％,9.2％ vs 10.6％,respectively.ConclusionsOf all the patients newly diagnosed as diabetes ketosis who had entered into the honeymoon after intensive insulin therapy,91％ were type 2 diabetes.Inferior β cell function before insulin therapy,weaker remission after insulin therapy and shorter duration of remission period suggest the classification of type 1 diabetes.

OBJECTIVE To investigate the bacterial distribution and antibiotic resistance situation with urinary tract infection(UTI) for the guidance of rational use of antibiotics.METHODS The antibiotic resistance of clinical isolates from urinary tract infection from Mar 2005 to Jul 2006 was analyzed. RESULTS The most common pathogens in urinary tract infection were Escherichia coli(50.2%),Enterococcus(14.4%),Staphyloccus aureus(8.7%),Klebsiella pneumoniae(7.3%),and Proteus mirabilis(3.9%).E.coli,K.pneumoniae,and P.mirabilis were found to be highly resistant to ampicillin,quinolones and SMZ(70.6-100.0%).Enterococcus were highly resistant to penicillin and quinolones(81.0-96.8%).41.4% of E.coli and 31.3% of K.pneumoniae isolates produced ESBLs.HLGR-Enterococcus were 79.4%.78.9% S.aureus isolates were resistant to oxacillin.CONCLUSIONS The high antibiotic resistance of commonly encountered pathogens is a serious problem and much attention should be paid to detect pathogens and their antibiotic resistance.

Objective To determine the content of sophocarpine, matrine, oxysophocarpine, sophoridine, oxymatrine in Sophora Flavescentis Radix from different areas. Methods Agilent ZORBAX NH2 column (4.6 mm×150 mm, 5 μm) was used with mobile phase of acetonitrile-ethanol-3% phosphate (84∶10∶6), at a flow rate of 1 mL/min. The wavelength of detection was 210 nm. Results The linear range of sophocarpine, matrine, oxysophocarpine, sophoridine and oxymatrine were 0.022 88-0.114 4 μg (r=0.999 7), 0.083 2-0.416 0 μg (r=0.999 7), 0.376 2-1.836 0 μg (r=0.999 8), 0.104 4-0.522 μg (r=0.999 2), 0.491 2-2.456 μg (r=0.999 9), respectively. The average recovery were 101.63% (RSD=2.08%), 98.29%(RSD=1.87%), 101.89% (RSD=1.97%), 99.87% (RSD=2.06%), 102.66% (RSD=1.34%), respectively. Conclusion The method is simple, rapid and accurate, and suitable for the quality control of Sophorae Flavescentis Radix.

Objective To investigate the value of CysC-based CFR in comparison with creatinine-based GFR (CG-GFR and MDRD-GFR) as an accurate serum marker in the prediction of early diabetic nephropathy. Methods A tatal of 133 type 2 diabetic patients (74 males and 59 females, aged 58.1 ±12.3) were enrolled. The level of diabetic nephropathy (normoalbuminuric, microalbuminuric and macroalbuminuric) was staged and estimated GFR based on serum creatinine and cystatin C(CysC) was calculated. The plasma clearance of Tc-DTPA, serum CysC, creatinine, BMI, HbA1c, serum lipid and blood pressure were measured. Results 99mTc-DTPA-GFR was used as golden standard. At 90 and 75 ml· min-1· (1.73 m2)-1 cut-points, diagnostic efficiencies of CysC-GFR (89% and 92%) were better than those of CG-GFR (79%=86%, P=0.004, 0.04) and MDRD-GFR (80%-86%, P=0.02, 0.04). At 60 ml · min-1 · (1.73 m2)-1 cut-point, diagnostic efficiencies of CysC-GFR,CG-GFR and MDRD-GFR were 92%, 90% and 92% respectively (P= 0.49, P=0.71). The Logistic regression analyses showed that retinopathy, HbA1c, CysC, diabetic duration, and CysC-GFR were indicators to predict the development of microalbuminuria. Conclusion CysC-GFR is more valuable than CG-GFR and MDRD-GFR in the prediction of early diabetic nephropathy and should be applied clinically.

Objective To obtain the specific antigens of the expressed major capsid proteins from Noroviruses with different sub-genotypes in Beijing area. Methods The full-length genes of the major capsid proteins (VP1)were obtained through the amplification of the VP1 encoding gene in the recombinant plasmids pBST-CR2987(G Ⅱ-3)and pBST-CR2932(GⅡ-4),which represented different Norovirus geno-types. The full-length genes were sub-cloned into the expression vector pET-30a(+),resulting in a recombinant plasmid, with which the BL21 competent cells were transformed, and the expression of the gene was induced by adding IPTG to the growth culture. The expression of the major capsid proteins were analyzed with Coomassie blue staining after SDS-PAGE, and assayed by Western blot with serum from human. Results (1)The major capsid protein genes of CR2987 and CR2932 were sub-cloned into expression vector pET-30a(+). The VP1 encoding genes were 1647 bp in length for CR2987 and 1623 bp for CR2932. The open reading frames(ORF)coded for 549 and 541 amino acids for these two proteins, respectively. (2)The expressed VP1s were present primarily as inclusion bodies,and the maximal amount of the expressed proteins occurred at 4-6 h after IPTG induction.(3)These VP1s could be recognized by specific immune serum against VP1 of Norovirus as well as His-tag antibody. Conclusion The VP1s of CR2987 and CR2932 are expressed in BL21 E.coli cells.The expressed VP1s could react with specific immune serum against VP1 of Norovirus, indicating that the expressed VP1s are of antigenicity.

ObjectiveTo investigate the correlation between glucose excursions and oxidative stress in the elderly type 2 diabetic patients. MethodsFifty five elderly type 2 diabetic patients (32 males and 23 females) were recruited. All the patients were submitted to continuous glucose monitoring system (CGMS) for three days. According to the result of CGMS monitoring ,the patients were divided into two groups: high glucose excursion group (30 cases) and low glucose excursion group (25 cases). In high glucose excursion group, the therapy was adjusted for 4 weeks based on the CGMS monitoring. After the treatments, the specimens of blood were collected again to detect the related indexes. Results(1) There were no differences in age, blood pressure, fasting blood glucose and glycosylated hemoglobin between the two groups (all P>0.05). The stardard deviation of blood glucose was higher, and the maximun and average amplitude of glycemic excursion were significantly lower in high glucose excursion group than in low glucose excursion group (t= 5. 4620,5. 9416,3. 8281, all P<0.05) ;(2) Compared with low glucose excursion group, plasma concentration of 8-iso-PGF2a was obviously higher in high glucose excursion group[(57.56 ± 3.86)ng/L vs.(34. 21±3. 82) ng/L, t= 18. 221, P=0. 0000)] . (3) Stepwise regression analysis showed that the standard deviation of blood glucose was involved in regression model (β= 1. 959, P= 0. 013).ConclusionsIn elderly patients with type 2 diabetes mellitus, glucose excursion is related with oxidation stress, which suggests that the glucose excursion may be the risk factor for oxidation stress.