Objective:To analyze the expression of E-cadherin in the epithelial-mesenchymal transition (EMT) induced by transforming growth factor-β1 (TGF-β1) in neuroblastoma.Methods:TGF-β1(1 μg/L, 5 μg/L, 10 μg/L), was applied to SK-N-SH cells in vitro compared with the blank control group.EMT-related genes mRNA and protein expression were detected by carrying out real-time PCR assays and Western blot.A scratch test and migration assay were performed to verify the alteration of SK-N-SH cell migration capacity.Data collected from 18 cases of neuroblastoma patients were selected from the Department of Hematology Oncology, Shanghai Children′s Hospital from January 2008 to December 2012.The expression of E-cadherin in the tumor tissue of the neuroblastoma patients after operation was detected by immunohistochemistry.The clinical features and survival prognosis of these patients were analyzed. Results:Compared with the control group, after SK-N-SH cells were treated with TGF-β1(1 μg/L, 5 μg/L, 10 μg/L), real-time PCR assays and Western blot revealed that the mRNA(0.603±0.081, 0.606±0.008, 0.716±0.166 vs.1.000) and protein expression levels(0.855±0.026, 0.600±0.017, 0.495±0.011 vs.1.000) of E-cadherin were significantly decreased ( F=8.144, P=0.040; F=74.810, P<0.001), while the mRNA(2.132±0.167, 3.494±0.017, 4.184±0.021 vs.1.000) and protein expression levels (1.175±0.053, 1.189±0.058, 1.225±0.106 vs.1.000)of α - smooth muscle actin were significantly increased ( F=547.300, P<0.001; F=68.810, P=0.007), suggesting that EMT changes occur in cells.Scratch test and Transwell migration assay revealed that the number of migrating cells increased obvious with the treatment of TGF-β1 (5 μg/L) ( t=16.070, P=0.040). The 10-year overall survival(OS) rates of neuroblastoma patients with E-cadherin strong positive expression, positive expression, weak positive expression and negative expression in the pathology were (77.78±13.86)%, (75.00±21.66)%, (25.00±21.65)% and 0, respectively ( F=8.160, P=0.040). Conclusions:TGF-β1 can induce the EMT in SK-N-SH cells and increase cell migration.The decrease expression of E-cadherin in neuroblastoma patients is closely associated with clinical progress and recurrence or metastasis of the disease.

Objective To explore the function of fluorescent probe JC-1 in detecting the changes of mitochondrial membrane potential(△?m)in early apoptotic cells.Methods After 2-ME was used to induce MUTZ-1 cell apoptosis,cells were dyed with fluorescent probe JC-1,and then the changes of △?m in the early stage of apoptotic cells were analyzed by flow cytometry or detected under fluorescent microscope. Results The control cells with high △?m are those forming JC-1 aggregates in the inner membrane of mitochondria,thus showing orange-red fluorescence.2-ME caused decrease of △?m in MUTZ-1 cells,in which JC-1 maintains monomeric form,thus showing only green fluorescence.The decreases of △?m were in a time-dependent manner,which were significantly higher than those in control group(P

· AIM: This study was designed to evaluate the effectiveness of anterior lens capsule inclusion in combined trabeculectomy and cataract surgery in preventing scar formation of the filtering bleb in rabbit model.· METHODS: Twerty-Four eyes (12 rabbits) with glaucoma model were studied, anterior lens capsule inclusion in trabeculectomy with the small-incision cataract surgery were performed on all right eyes (experimental group) and no implants were applicator in trabeculectomy with the small-incision cataract surgery on all left eyes (control group). These operated eyes were followed up from day 1 to 6 months postoperatively. Intraocular pressure (IOP) was measured and filtering blebs were observed after surgery. Other main outcome measures: cornea、conjunctiva、formation of the anterior chamber, anterior chamber depth、inflammatory reaction、achievement ratio of operation and complications were analyzed. On week 1, 2, 4, 12 and 24 after surgery the animal were killed in batch. Tissue was harvested from the bleb area and was made pathological section. HE staining、light microscope and micro photo analysis technique were applied to observe the cytological and histopathologic characteristics of the filtering tunnels.· RESULTS: There were significant differences between the two groups on IOP (1, 2, 4 weeks)、filtration bleb, achievement ratio of operation and complications. In experimental group, at the first month postoperatively, anterior lens capsule absorption started with inflammatory characteristics. The peak of inflammatory reaction occurred 1 week after operation and all the cells in the filtrating tunnel disappeared 6month after surgery. The fibroblast proliferation in control group occurred at I week and the filtrating tunnel closed with angiogenesis at 1 month after surgery. Fibroblast proliferation started 1week after surgery with no statistical difference during the time course (P ＞0.05). Significant statistical differences were observed by comparing the fibro blasts numbers per unit area in the filtrating tunnel in experimental group and those in control groups (P＜0.05).· CONCLUSION: Anterior lens capsule was totally absorbed at 6 months postoperatively. Anterior lens capsule inclusion in trabeculectomy with cataract surgery can possibly control intraocular pressure effectively, long-term sustainability of functional filtration bleb, inhibition of the proliferation of fibroblasts and opening of the filtrating pathway in the experimental animal models were satisfied. Compared to the control group, anterior lens capsule application has less complication.

ObjectiveTo investigate the serum prostate-specific antigen (tPSA),serum free PSA to total PSA ratio (f/tPSA),prostate volume (PV) and prostate-specific antigen density (PSAD) in early prostate cancer (PCa) diagnosis.MethodsRetrospective analysis was performed on serum PSA values and related test results from 252 cases of BPH patients and 49 patients with PCa.Prostate volume (PV) was measured by transrectal ultrasound (TRUS),and the f/tPSAand PSAD values were calculated.The differences of serum tPSA,f/tPSA,PV,and PSAD between BPH and PCa group were compared,the area under the ROC curve was used to evaluate these four indicators for its diagnostic sensitivity and diagnostic specificity.ResultsThe values of tPSA,PSAD in PCa group were significantly higher than BPH group ( P ＜0.05),while the values of f/tPSA,PV in PCa group were significantly lower than BPH group ( P ＜0.01orP ＜0.05).The ROC area showed that serum tPSA(0.8013),f/tPSA(0.7390),PV(0.5613) had lower diagnosis value than PSAD(0.9214) in early prostate cancer ( PSAD ＞ tPSA ＞ f/tPSA ＞ PV).When the upper limit of normal PSA was set to take 4ng/ml,the sensitivity was 91.49％,diagnostic specificity was 51.05％.When the f/tPSA threshold set to 0.16,the diagnostic sensitivity was 57.78％,diagnostic specificity was 78.72％.When PSAD threshold was set to 0.15,diagnostic sensitivity was 88.24％,diagnostic specificity was 81.52％.ConclusionsPSA,f/tPSA and PSAD are indicators for biopsy or followup in early diagnosis of prostate cancer.In particular,the diagnostic value of PSAD has higher sensitivity and specificity than PSA and f/tPSA in the diagnosis of prostate cancer.

Objective To compare the mRNA level of macrophage inflammatory protein-3α(MIP-3α) in 3 different mucosal epithelial cell lines. Methods RNA standards were prepared by in vitro transcription. One-step real-time RT-PCR was established and optimized using TaqMan EZ RT-PCR Core Reagents with TaqMan probes and primers specific to human MIP-3α mRNA sequence. The specificity of one-step real-time RT-PCR method was confirmed by sequencing the PCR products. The sensitivity and reproducibility of the method was examined by repeating the test 8 times with the same sample. Results The one-step real-time RT-PCR with a wide detection range is sensitive, reproducible. It was found that MIP-3α mRNA level in Caco-2 and T-84 cells was much higher than that in the HeLa cells. Conclusion High level of MIP-3α mRNA could be found in mucosal epithelial cells and difference in transcription level of MIP-3α may exist in epithelial cells from different mucosa.

Though all the marketed drugs of dipeptidyl peptidase IV inhibitors are structurally different, their inherent correlation is worthy of further investigation. Herein we rapidly discovered a novel DPP-IV inhibitor 8g (IC50 = 4.9 nmol.L-1) which exhibits as good activity and selectivity as the market drugs through scaffold hopping and drug splicing strategies based on alogliptin and linagliptin. This study demonstrated that the employment of classic medicinal chemistry strategy to the marketed drugs with specific target is an efficient approach to discover novel bioactive molecules.
Dipeptidyl-Peptidase IV Inhibitors ; chemical synthesis ; chemistry ; Drug Design ; Drug Discovery ; methods ; Humans ; Hypoglycemic Agents ; chemical synthesis ; chemistry ; Linagliptin ; chemical synthesis ; chemistry ; Molecular Structure ; Piperidines ; chemical synthesis ; chemistry ; Structure-Activity Relationship ; Uracil ; analogs & derivatives ; chemical synthesis ; chemistry

Nucleic acid sequence-based amplification(NASBA) is a sensitive,isothermal,transcription-based amplification system specifically designed for the detection of RNA targets,which could amplify templete RNA in 2h under isothermal condition at about 42?C and without any special equipment.NASBA is now widely applicated in diagnosis of many pathogenic microorganism.It is mainly about principles and applications of NASBA in viral diagnosis.

?AlM: To investigate the outcome and safety of Ahmed glaucoma valve implantation treatment in uncontrolled primary congenital glaucoma ( PCG) .
? METHODS: Twenty - two eyes in 22 children with uncontrolled PCG were reviewed retrospectively and underwent Ahmed glaucoma valve implantation treatment from January 2011 to December 2014. Main checking index included intraocular pressure ( lOP ) before and after operation, corneal diameter and complications.
?RESULTS: Preoperative mean age was 3. 74±2. 24y, and 2. 59 ± 1. 78y apart from the last operation. Postoperative average lOP was 35. 22 ± 6. 36mmHg. Average corneal diameter was 12. 79 ± 0. 75mm. Mitomycin C ( 0. 3 - 0. 5mg/mL ) was used in all operations for 3-5min. Glaucoma valves were implanted in the temporal or nose above the equator sclera. Postoperative lOP was 11. 4±4. 45mmHg at 1wk, and 16. 73± 7. 23mmHg after 12mo. As lOP< 21mmHg for success criteria, lOP of 16 eyes ( 73%) were controlled after 12mo. Preoperative 6 cases had shallow anterior chamber, recovered spontaneously. No serious complication was recorded, such as rejection of glaucoma valve, endoophthalmitis and corneal decompensation.
?CONCLUSlON:Ahmed glaucoma valve implantation in uncontrolled PCG is a safe and viable treatment.

Objective To evaluate the relationship between CD4+ T lymphocyte count and results of enzyme-linked immunospot (ELISPOT) assay in human immunodeficiency virus (HIV)-Mycobacterium tuberculosis (M.tb) coinfected patients.Methods A total of 193 HIV-infected individuals in Yunnan Province and Shanghai were enrolled.T-SPOT.TB assay was employed to detect M.tb specific T lymphocyte in the peripheral blood mononuclear cells (PBMC).CD4+ T lymphocyte in PBMC from the enrolled subjects was detected by flow cytometry.Data were analyzed using t test.ResultsThe incidence of latent tuberculosis in HIV-infected individuals was 30.6％.The CD4+ T lymphocyte counts in HIV-infected individuals with active tuberculosis were 190×106/L,which were significantly lower than those in HIV-infected individuals with latent tuberculosis (484×106/L; t=6.665,P＜0.01).The HIV-infected individuals were stratified according to CD4+ T lymphocyte counts of ＞500×106/L,200×106-500×106/L,and ＜200×106/L and the constituent ratios of active tuberculosis/latent tuberculosis were 1∶16.2,1∶1.3 and 5.6∶1,respectively.Among 79 subjects with positive T-SPOT.TB results,20 were coinfected with active tuberculosis,in which 14 had CD4+ T lymphocyte counts of ＜200 ×106/L,5 had 200×105-500×106/L and 1 had ＞500×106/L.Fifty-two in 59 HIV/latent tuberculosis patients individuals had CD4+ T lymphocyte counts of ＞200×106/L.ConclusionsThe prevalence of latent tuberculosis in HIV-infected individuals is high in China.Cellular immunity in HIV-infected individuals with active tuberculosis is severely impaired.With the decrease of CD4 ′ T lymphocyte counts,patients with latent tuberculosis are prone to develop active tuberculosis in HIV-infected individuals.The negative predictive value of T-SPOT.TB is significantly diminished in patient with low CD4+ T lymphocyte counts,especially less than 200×106/L.

Objective To investigate the influence of the purging fire and removing toxin method on chemokines and adhesion factors related to vascular endothelialitis injury induced by toxin of trimeresurus stejnegeri bite.Methods ① Animal experiment:50 healthy New Zealand white rabbits were chosen.According to random numbers generated by statistical software,they were divided into normal control group,model group,low,middle and high dose Sheshang capsule groups,10 in each group.Trimeresurus stejnegeri bite model was replicated by injecting 0.75 mL/kg snake venom into subcutaneous tissues of rabbits' right hind legs.And the same volume of normal saline was injected into the rabbit in the normal control group.After the model was established for 6 hours,the rabbits in low,middle and high dose Sheshang capsule groups received 174,348 and 522 mg· kg-1 · d-1 of Sheshang capsule solution respectively (the content of capsules was dissolved in normal saline to make liquid with 17.4,34.8 and 52.2 g/L Sheshang solution respectively,so the volume of gavage of each group was 10 mL· kg-1 · d-1);in the model and normal control groups,the same amount of normal saline was given by gavage,once daily for consecutive one week.24 hours after the last gavage,the blood of the rabbits was collected through an auricular border vein and the serum was separated by centrifuge ready for use.Meanwhile,the whole abdominal aorta segment of the rabbit was harvested and kept them in liquid nitrogen ready for use.② Cell experiment:human umbilical vascular endothelial cell (HUVEC) was cultured with MEM for 24 hours.The solution was replaced and according to the random number generated by statistical software,the cells were divided into blank control group,model group and low,middle,high dose Sheshang capsule medicinal serum groups,10 wells in each group.Trimeresurus stejnegeri toxin cell model was reproduced by addition of 5 mg/L snake venom into the cell culture medium.After 6-hour culture,the cells of model group and blank control group received 10％ normal rabbit serum,and the cells of low,middle and high dose Sheshang medicinal serum capsule groups received serum containing 5％,10％ and 15％ drug,respectively.After culture for 72 hours,the cells were collected and the total RNA was extracted.The real-time fluorescent quantitative polymerase chain reaction (qPCR) was used to detect the levels of mRNA of interleukin-8 (IL-8),monocyte chemoattractant protein-1 (MCP-1),intercellular adhesion molecule-1 (ICAM-1) and vascular endothelial cell adhesion molecule-1 (VCAM-1) in the vascular endothelial cells of rabbit aorta abdominalis and human umbilical vein,and the content of serum E-select element (CD62E) was measured by enzyme linked immunosorbent assay (ELISA).Results In model group,the expression levels of mRNA in IL-8,MCP-1,ICAM-1,VCAM-1 and the content of CD62E were all increased significantly in the endothelial cells of rabbit aorta abdominalis and HUVEC compared with those in control group [when the mRNA expression levels of IL-8,MCP-1,ICAM-1 and VCAM-1 in normal and blank control group were all being 1,the mRNA expression levels (2-△ △Ct) of the above mentioned inflammatory factors and adhesion molecule in animal model group were 3.96 ± 0.39,3.07 ± 0.27,3.71 ± 0.26,3.94 ± 0.26,and the mRNA expression levels (2-△ △Ct) of the above mentioned inflammatory factors and adhesion molecule in HUVEC model group were 3.53±0.70,2.24±0.48,3.13±0.44,2.80±0.13,respectively,all P ＜ 0.01].The content of CD62E in serum was increased significantly in model group compared with that in normal control group (μg/L:1.31 ± 0.22 vs.0.82 ± 0.13,P ＜ 0.01),the mRNA expression levels of IL-8,MCP-1,ICAM-1 and VCAM-1 were decreased significantly in low,middle,high dose Sheshang capsule groups compared with those in model group in endothelial cells of aorta abdominalis of rabbits and HUVEC [abdominal aorta:IL-8 mRNA (2-△ △Ct) were 1.13 ± 0.19,1.26 ± 0.16,1.27 ± 0.17 vs.3.96 ± 0.39,MCP-1 mRNA (2-△ △ Ct) were 1.79 ± 0.24,2.22 ± 0.38,1.76±0.19 vs.3.07±0.27,ICAM-1 mRNA (2 △△Ct) were 2.05±0.11,1.68±0.09,2.37±0.48 vs.3.71±0.26,VCAM-1 mRNA (2-△△Ct) were 1.59±0.08,1.40±0.11,1.84±0.11 vs.3.94±0.26;HUVEC:IL-8 mRNA (2-△△Ct) were 2.33±0.59,2.82±0.82,2.51±0.77 vs.3.53±0.70,MCP-1 mRNA (2-△△Ct) were 1.59±0.35,1.48±0.36,1.54±0.29 vs.2.24±0.48,ICAM-1 mRNA (2-△△Ct) were 1.46±0.38,1.77±0.65,1.73±0.50 vs.3.13±0.44,VCAM-1 mRNA (2-△△Ct) were 2.49±0.24,2.18±0.19,2.45±0.24 vs.2.80±0.13,all P ＜ 0.05].The contents of CD62E were decreased significantly in middle,high dose Sheshang capsule groups compared with the content in model group (μg/L:1.01 ±0.14,1.04±0.13 vs.1.31 ±0.22,all P ＜ 0.01),but there were no statistical significant differences among the three drug group (all P ＞ 0.05).Conclusion The therapy of purging fire and removing toxin can treat vascular endothelial injury by inhibiting the inflammatory response induced by Trimeresurus stejnegeri bites.