Objective: To construct DNA vaccine against C region gene of SARS-CoV S protein, and observe the immune response induced by DNA vaccine injection and mucosal immunization. Methods: A recombinant DNA vaccine plasmid was constructed by cloning Sc gene fragment into pcDNA3. 1( - ) ,which could induce anti-SARS-CoV specific antibodies in mouse. After mice immunized with the recombinant DNA vaccine by injection and mucosal immunization, the SARS-CoV antibodies in the sera of mice were detected by ELISA,the lymphocyte phenotypes were detected by FCM,the specific antigen expressed in tissues was detected with immuno histochemical analysis. Detect spleen and MLN lymphocyte proliferation and CD8+ CTL response in vitro. Results: The titer of anti-Sc IgG was much higher in the mouse injected pcDNA3. 1-Sc than that of control group (P

Objective:To study the alveolar bone morphology in the anterior teeth area of the skeletal Class Ⅱ malocclusion subjects with different vertical skeletal types.Methods:64 patients with skeletal Class Ⅱ malocclusion and 15 subjects with normal occlusion were included.The alveolar bone structure of the anterior teeth were observed using CBCT.Results:The labial and lingual alveolar bone height and the alveolar bone thickness of the incisors of the patients were much lower than those of the normal controls.The height of labial and lingual alveolar bone and the alveolar bone thickness of anterior teeth in high-angle subgroup were lower than those in low-angle subgroup.Conclusion:The thickness of the anterior teeth alveolar bone of skeletal Class Ⅱ malocclusion is low,espe-cially in the high-angle group.

Objective To investigate the clinical efficacy of prucalopride in the treatment of severe chronic constipation.Methods The clinical data of 60 patients with severe chronic constipation [slow transit constipation (STC), functional defecation disorder (FDD) and constipation-predominant irritable bowel syndrome (IBS-C)] who were admitted to the Third Affiliated Hospital of Nanjing University of Chinese Medicine from February to August 2014 were prospectively analyzed.A prospective, clinical observational study was performed.Treatment plans included that patients withdrew the initial treatments of laxative and exema and took orally 2 mg prucalopride once daily for 2 weeks, and continued to be treated by oral prucalopride if frequency of the spontaneous complete bowel movement (SCBM) per week was satisfactory (or improvement of symptoms was more than 50％) till 4 weeks, and then were followed up after stopping prucalopride.If improvement of symptoms was less than 50％ after 2-week treatment, other treatment plans were performed according to symptoms of patients from week 3 to week 6 : (1) for patients with STC, prucalopride + two chain bacillus subtilis probiotic capsules were administered orally if patients were satisfied with frequency of SCBM per week and without improvement of abdominal distension;prucalopride + Chinese herb decoction were administered orally if patients had improvement of frequency of SCBM per week with abdominal distension or poor stool output;oral prucalopride + acupuncture were administered if patients were unsatisfied with frequency of SCBM per week or less bowel movements and without improvement of abdominal distension or poor stool output.(2) For patients with FDD, oral prucalopride + acupuncture + biofeed-back therapy were administered.(3) For patients with IBS-C, prucalopride + two chain bacillus subtilis probiotic capsules were administered orally if patients had abdominal distension;prucalopride Chinese herb decoction were administered orally if patients had improvement of frequency of SCBM per week and no improvement of abdominal distension or poor stool output.All patients used a diary for recording the frequency of SCBM per week, stool consistence, exertion in defecation and adverse reactions, which was submitted to doctors for inputting data at the return visit weekly.Results There was good overall medicine compliance in patients.Of 60 patients, 43 patients completed treatments (21 with STC, 11 with FDD and 11 with IBS-C).After 2-week treatment, there were 19 patients with satisfied therapeutic effects, 14 with improvement of constipation and 10 with poor therapeutic effects.After 4-week treatment, constipation in 17 patients was cured, constipation in 18 patients was improved,and constipation in 8 patients was not improved.Nineteen of 60 patients were complicated with adverse reactions within 1 week of the medication, including 6 patients dropping out of the trial due to medication withdrawal and others with improvement by symptomatic treatment or spontaneous remission.Conclusions Prucalopride is effective for the treatment of severe chronic constipation with a good toleration, and it can improve the overall satisfaction of patients combined with Chinese herb decoction and acupuncture.

BACKGROUND:Previous studies have found that estrogen deficiency causes a reduction in the activity of bone marrow mesenchymal stem cel s (BMSCs). OBJECTIVE:To explore the effect of endothelial progenitor cel s (EPCs) on the BMSCs proliferation and apoptosis ability of osteoporosis rats. METHODS:Healthy female Sprague-Dawley rats, 6 weeks old, were enrol ed and subjected to bilateral ovariectomy to make osteoporosis models. BMSCs and EPCs were isolated using density gradient centrifugation combined with adhesion method, and identified with surface markers, cel proliferation and immunocytochemistry in vitro. We used Transwel inserts to establish EPCs and OVX-BMSCs indirect co-culture system. Control groups were OVX-BMSCs group and sham-BMSCs group in which rats were only subjected to remove the equal amount of fat tissues around the ovary. Flow cytometry was applied to detect BMSCs proliferation and apoptosis ability. RESULTS AND CONCLUSION:Compared with the control groups, the results of flow cytometry test showed that the proportion of OVX-BMSCs at S phase was significantly increased at 3 days of indirect co-culture with EPCs and the apoptosis rate was significanty reduced at 10 days of indirect co-culture with EPCs (both P<0.05). These results suggest that EPCs can promote the proliferation but inhibit the apoptosis of OVX-BMSCs.

BACKGROUND:Numerous studies have demonstrated that estrogen can regulate the proliferation and migration of endothelial progenitor cel s (EPCs), while EPCs can also promote the function and activity of bone marrow mesenchymal stem cel s (BMSCs) in vitro. OBJECTIVE:To evaluate the ability of the BMSCs and EPCs which construct the composite cel sheet in the repair of alveolar bone defect in ovariectomized rats. METHODS:BMSCs/EPCs composite sheet, EPCs sheet and BMSCs sheet were respectively implanted into the defects of the alveolar bone in ovariectomized rats. Rats with no implantation served as control group. Repaired alveolar bone was assessed by gross examination, histological observation and micro-CT scan at 2, 4, 8 weeks after operation. RESULTS AND CONCLUSION:BMSCs/EPCs composite sheet has greater osteogensis activity and bone repair capacity than BMSCs or EPCs sheet alone.

BACKGROUND:Many studies have showed that enough blood supply is an essential condition of bone repair and regeneration. OBJECTIVE:To construct the endothelial progenitor cells/bone marrow mesenchymal stem cells (EPCs/BMSCs) composite sheet. METHODS:After isolation and culture, EPCs and BMSCs were co-cultured directly to form EPCs/BMSCs sheet by cellsheet-inducing medium. After 10 days of induction, the sheet was investigated by gross observation, inverted microscope and hematoxylin-eosin staining. The distribution and communication of EPCs and BMSCs during the process of cellsheet induction were observed after the fluorescence labeling separately. Alkaline phosphatase assay and alizarin red staining were applied to examine the ability of osteogenic differentiation of EPCs/BMSCs sheet.
RESULTS AND CONCLUSION:EPCs/BMSCs sheet was harvested after 10-day induction. Cel-cellcontact between EPCs and BMSCs could be observed during the process of the cellsheet preparation. The harvested sheet was composed of multiple layers of cells and cel-produced extracellular matrix. Alkaline phosphatase assay and alizarin red staining both demonstrated that EPCs/BMSCs sheet had good osteogenic differentiation ability. These results suggested that EPCs/BMSCs sheet can be constructed successful y, and the sheet has strong osteogenic differentiation capability in vitro, providing the foundation for the repair of bone defects.

Objective The adverse reactions of levonorgestrel releasing intrauterine system ( LNG-IUS) has been emphasized with the increase of its indications.The study was to investigate the feasibility of interventional treatment in treating uterine adenomyosis with LNG-IUS by observing its adverse reactions, trying to reduce its adverse reaction, increase its utilization ratio and service life and improve its efficacy. Methods Retrospective analysis was made on 67 patients with uterine adenomyosis who were willing to accept LNG-IUS treatment from January 2012 to December 2013 in our hospital.The patients were randomly divided into observation group ( n=34) and control group( n=34) according to different therapies.The observation group were given interventional treatment immedi-ately after the placement of LNG-IUS, that is to take oral non-steroidal anti-inflammatory drug ( indomethacin, 25 mg, 1 pill each time, three times a day) for 7 consecutive days and desogestrel-ethinylestradiol ( marvelon, 1 pill a day) for 21 consecutive 21 days, 3 con-secutive cycles;while the control group were only given communications without any special treatment.Observation and record were made on related adverse reactions at 1 month, 3 months and 6 months after the treatment. Results At 1 month, 3 months and 6 months after the interventional treatment, the incidence rates of abnormal uterine bleeding between two groups were of significant difference ( 23.5%vs 60.6%, 11.7%vs 36.4%, 0%vs 12.1%, P<0.05) in the control group. At 6 month after the treatment, statistical difference was found in the total incidence rates of amenorrhea, ovarian cysts, dislocation and out of place of IUD between these two groups ( 23.5%vs 6.1%, 2.9%vs 21.2%, 2.9%vs 18.2%, P<0.05). Conclusion The research has indicated that the adoption of interventional treatment after placing LNG-IUS can obviously reduce the occurrence of the main adverse reactions, so it is feasible in clinical application.

BACKGROUND:Extracellular matrix can simulate microenvironment and make the stem cells proliferate maintaining the characteristics of stem cells wel in vitro. OBJECTIVE:To prepare the extracellular matrix from human bone marrow cells and to analyze its microstructure and composition preliminarily. METHODS:Human bone marrow cells of passage 4 were cultured for 14 days, and the induction medium was used during the last 8 days. After decellularization, cells were removed to prepare human bone marrow cells-derived extracellular matrix. The surface morphology of human bone marrow cells-derived extracellular matrix was observed by inverted microscope and scanning electron microscope. Changes of col agen I and biglycan before and after decellularization were observed by immunofluorescence staining. Human periodontal ligament stem cells were seeded onto human bone marrow cells-derived extracellular matrix, fibronectin coated 6-wel plate and normal culture plate to compare the influence of different matrix on cellmorphology and adhesion. RESULTS AND CONCLUSION:We obtained intact human bone marrow cells-derived extracellular matrix by chemical combined with physical decellularization. The structure and amount of col agen I and biglycan had not been compromised dramatical y after decellularization. Human periodontal ligament stem cells growing on the human bone marrow cells-derived extracellular matrix developed in accordance with the orbit of the extracellular matrix, differing from the original cellmorphology. There were more human periodontal ligament stem cells adhering to the extracellular matrix during the same time. These findings indicate that effective decellularization can produce intact the extracellular matrix membrane without destroying its microstructure. Extracellular matrix protein is not compromised due to decellularization. The extracellular matrix affects cellmorphology and promotes celladhesion. We can use the extracellular matrix model to simulate stem cellmicroenvironment and thereafter, acquire a large number of adult stem cells with high quality in vitro.

BACKGROUND:Human bone marrow cells-derived extracellular matrix can promote proliferation of human periodontal ligament stem cells and maintain stem cellproperties.
OBJECTIVE:To preliminarily investigate the effect of human bone marrow cells-derived extracellular matrix on the proliferation of human periodontal ligament stem cells.
METHODS:Human periodontal ligament stem cells and bone marrow cells were separately derived from human periodontal tissue and jaw bone marrow, and human bone marrow cells-derived extracellular matrix was prepared. Human periodontal ligament stem cells were cultured and purified using limited dilution cloning method, and transmission electron microscope was used for ultrastructure observation. Human periodontal ligament stem cells at passage 2 were cultured with human bone marrow cells-derived extracellular matrix and normal culture medium (control group). The cellcounting kit-8 and flow cytometry were used to determine the proliferation potential of human periodontal ligament stem cells cultured on human bone marrow cells-derived extracellular matrix.
RESULTS AND CONCLUSION:Compared with the control group, human periodontal ligament stem cells cultured on human bone marrow cells-derived extracellular matrix had a superior capacity of proliferation (P<0.05), and the cells met their morphological and biological characteristics, and grew in good conditions. Human bone marrow cells-derived extracellular matrix is a promising matrix for large-scale expanding human periodontal ligament stem cells for future use in stem cel-based therapy.

This study was aimed to establish determination method of content and related substances of piperaquine in A rtemisinin and Piperaquine Tablets, and to set the limit of related substance. HPLC was adopted on a SHISEIDO CAPCELL PAK C18 (4.6 mm í 250 mm, 5 μm) using an isocratic mobile phase consisted of acetonitrile: 0.1%trichloroaceticacid:triethylamine (18:82:0.2, V:V:V, pH 2.5) with a flow rate of 1.0 mL·min-1. The column tempera-ture was kept at 30oC and the detection wavelength was set at 216 and 237 nm, separately for the determination of related substance and content. The results showed that piperaquine and its related impurity can be separated effec-tively. The concentration-response relationship was linear over the range of 0.01-0.2 mg·mL-1 (R2=0.999 9). The av-erage recovery rate was 98.14% (RSD=0.77%, n=9). The minimum detection limit was 0.06 μg·mL-1. The solution was stable for 12 h. It was concluded that the method was specific, accurate, sensitive and suitable for the determi-nation of content and related substances of piperaquine in A rtemisinin and Piperaquine Tablets.