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 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 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 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.

BACKGROUND:Compared with the conventional composite resin, the 3M Z350 nano-resin has good wear
resistance, physical mechanical properties, and polishing, and exerts a lower irritation to the dental pulp. Besides fil ing materials, a reliable tooth-prosthesis bonding interface is necessary for resin bonded repairs.
OBJECTIVE:To compare the clinical effects of self-etch bonding Adper Easy One and total-etch bonding Single Bond 2 on nano-resin bonding restoration of the anterior teeth.
METHODS:120 anterior teeth with vital pulp, which had defects at the incisal ends and were to be restored with nano resins, were divided into two groups randomly. Two kinds of adhesives, self-etch adhesive and total-etch adhesive, combined with nano-resin were used to restore the teeth. The patients were re-examined immediately, 6 months, 1 year and 2 years after the treatment. The fil ings, teeth and pulps of patients were examined,
including whether the prosthesis and tooth color were coordinated, whether the gap between the prosthesis and the teeth were sealed, whether the surface of the prosthesis was intact with no loose, whether the prosthesis and teeth had no staining and secondary caries, whether the condition of the tooth pulp had hot or cold
stimulation-induced pain.
RESULTS AND CONCLUSION:No significant difference in the fil ing effects was found between the two groups when the patients were re-examined immediately, 6 months and 1 year after the treatment (P>0.05). The pulp
lesions of the self-etching group were fewer than those of the total-etch group 2 years after the treatment (P<0.05). Self-etching group had 1, 6, 0, 2 cases and total-etch group had 0, 2, 1, 2 cases in uncoordinated color, edge seal,
incomplete restoration and secondary caries, respectively. No statistical y significant differences were found in these four aspects between the two groups (P>0.05). The 2-year fol ow up showed a low incidence of pulp lesions and satisfactory clinical performance after 3M Z350 nano-resin working with self-etching bonding system in the nano-resin fil ing of
anterior teeth with vital pulp.

Coronary heart disease (CHD) is a common kind of cardiovascular disease which does serious harm to human health. It has become the first cause of people's death in many countries and regions. This article analyzed CHD-related animal experiments in recent ten years. It made a review on progress in the establishment of chronic and acute animal models of CHD from the animal selection and model establishment. It can provide guidance for the further research of CHD.

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.

Background The pathogenic mechanism of thyroid associated ophthalmopathy (TAO) is still unclear,which is considered to be an autoimmune disease.It is confirmed that interleukin-17A (IL-17A) plays an important role in the occurrence and development of many autoimmune diseases.It is unclear that whether IL-17A participates in the pathogenesis of TAO.Objective This study was to explore whether IL-17A secreted by coculture system of peripheral blood mononuclear cells (PBMCs) and orbital fibroblasts (OFs) participates in the pathogenesis of TAO and its possible mechanism.Methods Periphery blood and orbital connective tissue were obtained from 12 patients with TAO and 8 patients who received prosthesis implantation for eyeball atrophy in Xiangya Hospital during April to December 2014.PBMCs were isolated by density gradient centrifugation,and OFs were cultured by explant culture method.The purity of T leukomonocyte in PBMCs was tested by flow cytometry,and OFs were identified by Giemsa staining and immunochemistry.OFs and PMBCs were incubated into 96-well plate in a 1:20 proportion to establish co-culture system.Different concentrations of phytagglutinin (PHA) (0,1.0,2.5,5.0,10.0 μg/ml) was added for 72 hours,and IL-6,IL-17A levels in the co-culture system supernatant and IL-17A receptor (IL-17RA) of the total cell membranes in the co-culture system were assayed by ELISA.The differences of IL-6,IL-17A,IL-17RA levels in co-culture system were compared between the TAO group and control group.Results The mean purity of T leukomonocyte in PBMCs was (81.10±0.21)％ in the TAO group and (80.05 ±0.38)％ in the control group respectively,with no significant difference between them(t =0.923,P＞0.05).Cultured OFs showed the positive response for Vimentin expression and Giemsa staining.After stimulated by 1.0 μg/ml PHA,the proliferation of both PBMCs and OFs were increased in the co-culture system.Apoptosis exist in PBMCs and the number of OFs decreased when PHA was higher than 1.0 μg/ml.The growth of PBMCs and OFs was faster in the TAO group than that in the control group in the same concentration of PHA.The contents of IL-6,IL-17A and IL-17RA in co-culture system were significantly different among various concentrations of PHA subgroups (IL-6:Fgroup =12.561,P=0.000;F ion =23.356,P =0.001.IL-17A:Fgroup =12.037,P =0.000;Fconcentration =19.206,P=0.000.IL-17RA:Fgroup =16.216,P=0.000;Fconcentraction =4.627,P=0.018).The production of IL-6,IL-17A and IL-17RA reached peak in both TAO group and the control group after 1.0 μg/ml PHA stimulated.However,the concentrations of IL-6,IL-17A and IL-17RA reduced with the increase of PHA concentration.The concentrations of IL-6,IL-17A and IL-17RA in co-culture system were significantly higher in the TAO group than those in the control group under the stimulation of the same concentration of PHA (all at P＜0.05).Conclusions The co-culture system of PBMCs and OFs stimulated with PHA can be the imitation of TAO pathogenesis in vitro,and PHA can amplify its immune reaction to imitate TAO pathogenic processes intuitively.The IL-6,IL-17A and IL-17RA secreted by PBMCs and induced by PHA are increased in TAO patients,implying that IL-17A participates in the pathogenesis of TAO through magnifying cellular immune response and inflammatory reaction.

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.