Objective To induce directional differentiation from spinal cord-derived neural stem cells(SNSCs)into neurocytes in the simulated microenvironment of development of embryonic spinal cord of rat after isolation and culture of SNSCs and to investigate the proliferation and differentiation of SNSCs in vitro.Methods SNSCs were isolated mechanically from rat embryonic(E10 to 11d)spinal cord under microscope.SNSCs were cultured and maintained in serum-free medium.Directional differentiation from SNSCs to neurocytes was induced by adding N4 supplement.The morphologic features of the differentiated cells were noted.Cultured and differentiated cells were identified by immunochemistry stain and t he mean differentiating percentages of the positive cells were calculated.Resul ts SNSCs isolated by the mechanical method under microscope were vigorous and pr oliferative,and could differentiate into neurons,astrocytes and oligodendrocyt es.Conclusion The mechanical method under microscope of isolating SNSCs is simp le and efficient to obtain a high percentage of neurons.N4 supplement can induc e SNSCs to differentiate into neurocytes.

Acupuncture Therapy ; Adult ; Chronic Disease ; therapy ; Humans ; Male ; Moxibustion ; Pelvic Pain ; therapy ; Prostatitis ; therapy ; Treatment Outcome ; Young Adult

Objective To investigate the polymorphism at positions-137 and -607 in the upstream promoter 1 region of exon 2 of interleukin(IL)-18 gene in Han children with atopic dermatitis(AD)in Chongqing,China,as well as its correlation with the development of AD.Methods Blood samples were collected from 82 patients with atopic dermatitis and 100 healthy controls.DNA was extracted from the samples and subjected to test with PCR.The polymorphism of IL-18 gene at positions-137 and -607 in the upstream promoter 1 region was analyzed by polymerase chain reaction(PCR)-sequence specific primers(SSP)and gene sequencing.Genotype frequency was compared between the patients and controls.Resuits A G/C polymorphism(GG,GC and CC genotypes)was identified at position-137 in exon 2 of IL-18 gene,and a C/A polymorphism(CC.CA and AA genotypes)at position-607 of this gene.The frequency of genotype GC at position-137 was significantly higher in the patients than that in the controls(47% vs 27%,P＜0.05;odds ratio=2.33.95% confidence interval 1.26-4.33).Increased frequency of C allele was also noted at the position-137 in the patients compared with the controls(0.24 vs 0.15,P＜0.05;odds ratio=1.76,95% confidence interval 1.04.2.97).Patients with severe AD(SCORAD score＞50)were more likely to carry C allele at position-137 of IL-18 gene than those with mild AD(SCORAD＜20).There was no statistical difference in allele frequency at -607(C/A)among patients with mild,moderate,severe AD and the controls(P＞0.05).Conclusions There is a polymorphism at positions -137 and -607 in the upstream promoter 1 region of IL-18 exon 2.The GC genotype of IL-18 at position -137 may confer the susceptibility to AD in Han children in Chongqing.

Objective To investigate the expression changes of microRNA 146a (miRNA146a) in UVBdamaged mouse skin. Methods C57/BL6 mice were divided into dose groups to be irradiated on the back with UVB of 30, 60, 90, 180, 270 mJ/cm2 respectively for 24 hours, and time groups irradiated with UVB of 180 mJ/cm2 for 1, 12, 24 and 48 hours respectively. The mice received no irradiation served as the control. Skin samples were subsequently obtained from the irradiated sites of mice and subjected to real-time fluorescent PCR for the detection of miRNA146a expression as well as immunohistochemical staining (IHC) for the detection of STAT3. Results The expression levels of miRNA146a were 0.01158 ± 0.00098, 0.01083 ± 0.00104,0.00872 ± 0.00031, 0.00851 ± 0.00033, 0.00810 ± 0.00057 and 0.00770 ± 0.00031 in the unirradiated control mice and mice irradiated with UVB of 30, 60, 90, 180, 270 mJ/cm2 for 24 hours, respectively, 0.00730 ±0.00036, 0.00805 ± 0.00035, 0.00810 ± 0.00057 and 0.00837 ± 0.00039 in mice irradiated with UVB of 180mJ/cm2 for 1, 12, 24 and 48 hours, respectively. IHC suggested an intensive expression of phosphorylated STAT3 in mice irradiated with a high dose UVB. Conclusions miRNA146a may play an essential role in the mechanism of UVB-induced photodamge, which is mainly correlated with the negative regulation of inflammatory reaction likely mediated by the JAK/STAT3 signal transduction pathway.

The cardiovascular disease, accompanied by psychological disorder frequently, often re-quiresphysical and mental (double heart) treatment. The clinical teaching of cardiovascular graduate stu-dents is very important for cultivating qualified cardiovascular doctors. According to present situation, we improved the diagnosis rate and the doctor-patient communication skills by the strengthening of learning thedouble heart theory, the analysis ofdouble heart typical cases, and the clinical practice of double heart medical model. Accordingly, we enhanced the consciousness of double heart, which is helpful to reducing the conflicts between doctors and patients, promoted the rational use of medical resources, and fi-nally promoted the application ofdouble heartmedical model in clinical practice.

Objective To observe the changes of microRNA141 expression in photodamaged skin of mice irradiated with ultraviolet B (UVB).Methods Eighteen C57/BL6 mice were equally divided into six groups to receive single irradiation on the hair-removed back with UVB of 0,30,60,90,180 and 270 mJ/cm2 respectively.Skin specimens were obtained from the irradiated sites 24 hours after the irradiation and subjected to paraffin embedding followed by hematoxylin-eosin (HE) staining for the observation of histopathological changes.Real-time quantitative PCR was performed to determine the miRNA141 expression in skin specimens,immunohistochemical staining (IHC) to quantify the expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN) protein.The TargetScan database was employed for the prediction of miRNA141 targets,and Gostat analysis for functional clustering.Data were processed using SPSS version 13.0 software,and intergroup comparisons were done by analysis of variance.Results The relative expression level of miRNA141 in skin tissue was 2.1354 ± 0.4289,2.4333 ± 0.2517,2.9328 ± 0.3126,3.4125 ± 0.3606,4.5667 ± 0.4014 and 6.7428 ± 0.5158 in mice irradiated with UVB of 0,30,60,90,180 and 270 mJ/cm2 respectively.There was a positive correlation between the expression level of miRNA141 and radiation dose (r =0.992,P ＜ 0.01).Gostat analysis showed that the targets of miRNA141 were mainly related to transcription regulation and signaling transduction.Moreover,PTEN expression gradually decreased with the increase in radiation dose.Conclusions miRNA141 may be involved in UVB-induced photodamage and inflammatory response,and PTEN may play a regulatory role in this process.

Objectives:To detect telomerase activity in various tumor tissues by PCR-PAGE and PCR-MPH.Methods:We used telomerase PCR-MPH，an assay in which a digoxigenin-labeled telomerase repeat-specific probe was hybridized to the PCR product.After revelation with an anti-digoxin-AKP conjugates,the amount of hybridized probe was determined by optical reading.Telomerase activity was analyzed by PAGE.Results:Telomerase activity was determined by PCR-MPH accurately and specifically in various tumor tissues,its sensitivity was one hundred times higher than that of PCR-PAGE.Conclusion:PCR-MPH is ideal for analyzing telomerase activity in various tumor tissues.It is simple,easy and suitable for clinical application.

Objective To evaluate the feasibility of bladder extracellular matrix (BACM) seeding with autologous cultured vaginal smooth muscle cells (VSMC) repaired rabbit vaginal defect.Methods This study included 24 female rabbits.BACM and vaginal acellular matrix (VACM) were obtained from 8 rabbits by decellularization process.The other 16 rabbits were randomly divided into experimental and control groups.Vaginal tissue biopsies ( ～ 1 cm2) were harvested from female New Zealand rabbits.VSMC were cultured and stained with a-smooth muscle actin antibodies. Cultured VSMC cells were seeded on BACM ( experimental group) or VACM ( control group) at a cell density of 1 × 107 cells/cm2.The cell-seeded matrixes were cultured for 5days.In experimental group of 8 rabbits,a 2 cm segment of vagina was resected and replaced with BACM seeding with VSMC.Then the regenerative segment was studied with histological technique by hematoxylin-eosin staining after 3,6 and 12 weeks postoperative aud vaginography was performed at 12 weeks postoperative.The 8 rabbits in control group underwent the exact same procedure as above but the vaginal defect was repaired with VACM seeding with VSMC,Results The prepared BACM and VACM were transparent,HE staining and scanning electron microscopy showed the two acellular matrix was both consisted of abundant network of fibers with the regular arrangement,without cellular debris.Primary culture of VSMC was successfully established and passaged,and was uniformly spindle-shaped in the confluent state and showed a characteristic hill and valley'formation.Immunohistochemical staining showed VSMC in culture stained positively with a-smooth muscle actin antibodies.VSMC began to adhere to the BACM 5 hour after implanting and the number of the adhered cells increased with time.Cells gradually expanded and showed the typical morphology of smooth muscle cells.Three weeks after implantation in vivo,the luminal surface of matrix was completely covered by vaginal epithelial tissue,a layer of smooth muscle cells was formed in the outer surface of the matrix.Multilayered vaginal epithelial and improved development of organized muscle bundles was observed after 6 weeks.The regenerative tissue was equivalent to the normal vaginal tissue at 12 weeks postoperatively.Vaginography demonstrated the maintenance of full patency and a wide vaginal caliber without fibrosis and graft rejection.There was no significant difference in all evaluated items between experimental and control groups.Conclusions BACM has the same regenerative process as VACM in the replacement of vaginal defect.Moreover,BACM has wider source and appears to be an suitable material for vaginal replacement.

Objective To understand the status and drug resistant patterns of strains of extended spectrum ?-lactamase(ESBLs) in children with lower respiratory tract infection,and to give clinical suggestions for rational treatment.Methods Escherichia coli and klebsiella pneumoniae were isolated from the 2 969 nasopharyngeal secretions which collected from lower respiratory tract of children in our hospital from Jan.2006 to Dec. 2007.Dual-sheets and sheets-diffusing method (K-B method) were used to determine the ESBLs and antibiotic susceptibility was tested by K-B method which included 18 kinds of antibiotics,the results were marked by resistant,intermedial and sensitive.Chi-square test was used to analyze the data.Results Total 135 strains were detected,73 strains were escherichia coli,of which 54 strains(74.0%)produced ESBLs,62 strains were klebsiella pneumoniae,of which 33 strains(53.2%)produced ESBLs.The 2 bacterias were found more in children with 1-6 months old than those in other age groups,the ratio of which were 50 strains and 41 strains,respectively (Pa0.05).The resistant rate of ESBLs-producing strains to penicillins,cephalosporins,quinolones,aminoglycosides and sulfamido was higher than that of non ESBLs-producing strains respectively.And the resistant rates to beta-lactam antibiotics of ESBLs strains were located on a high level.Whether producing ESBLs or not,the 2 bacterias were still sensitive to amikacin,cefoxitin,cefoperazone/sulbactam and imipenem.Conclusions The prevalences of ESBLs-producing escherichia and klebsiella pneumonia were high.There was a multi-drug resistance to the varied antibiotics.It is very important to make sputum culture and use sensitive antibiotics in treatment according to drug sensitivity test to control the occurrence and conveying of the ESBLs.

Epithelial-to-mesenchymal transition (EMT) plays a critical role in cancer metastasis,and is relevant to the inflammatory microenvironment.Lipopolysaccharide (LPS),a cell wall constituent of gram-negative bacteria,has been reported to induce EMT of cancer cells through TLR4 signal.We previously reported that LPS promoted metastasis of mesenchymallike breast cancer cells with high expression of cyclin D 1 b.However,the role of cyclin D1b in LPS-induced EMT has not been fully elucidated.In the present study,we described that cyclin D1b augmented EMT induced by LPS in MCF-7 breast cancer cells.Cyclin D1b markedly amplified integrin αvβ3 expression,which was further up-regulated under LPS stimulation.Our results showed ectopic expression of cyclin D1b promoted invasiveness of epithelial-like MCF-7 cells under LPS stimulation.Additionally,LPS-induced metastasis and EMT in MCF-7-D1b cells might depend on αvβ3 expression.Further exploration indicated that cyclin D1b cooperated with HoxD3,a transcription factor promoting αvβ3 expression,to promote LPS-induced EMT.Knockout of HoxD3 repressed LPS-induced EMT and αvβ3 over-expression in MCF-7 cells with high expression of cyclin D1b.Specifically,all these effects were in a cyclin D1a independent manner.Taken all together,LPS up-regulated integrin αvβ3 expression in MCF-7 cells with high expression of cyclin D 1b and induced EMT in breast cancer cells,which highlights that cyclin D1b may act as an endogenous pathway participating in exogenous signal inducing EMT in breast cancer cells.