BACKGROUND: The relationship between cellular morphosis and function is undetachable, but there is little investigation about ultrastructure of human umbilical cord mesenchymal stem cells (hUCMSCs).OBJECTIVE: To study the relationship between the functions of hUCMSCs and their ultrastructure obtained by atomic force microscope (AFM). METHODS: hUCMSCs were isolated, cultured, expanded after enzyme digestion. P3 cells were observed under AFM. Immunophenotype and cell cycle were analyzed by flow cytometry, as well as induction of the adipogenic, osteogenic differentiation of hUCMSCs were identified using Oil red O staining and alkaline phosphatase staining. RESULTS AND CONCLUSION: hUCMSCs at passage 3 were strongly positive for CD44 and CD29, weakly positive for CD106, but negative for hematopoietic marker CD34. Cells in G_0/G_1 phase accounted for 80%. Proliferation index was 19.9%. Following adipogenic induction, alkaline phosphatase staining demonstrated brown cytoplasm in cube and polygonal cells. AFM showed hUCMSCs were spindle shape, obvious cytoskeletal filament that connected into nets, which fit for the strong capacities for proliferation, migration and differentiation.

[Objective]To investigate the value of 16-slice spiral CT(MSCT)postprocessing in styloid process in vivo and provide the vivisection data for diagnosing and curing styloid process syndrome.[Method]Totally 100 volunteers with normal styloid processes underwent 16-slice spiral CT(MSCT). Based on the initial datum,4 kinds of postprocessing including multiple planar reconstruction(MPR), curve planar reconstruction(CPR),surface shaded display(SSD) and volume rendering (VR) were performed.The length and inclination angle of stytoid processes were measured. The distance of between styloid processes and carotid canal external aperture,jugular foramen, stylomastoid foramen ,mastoid process, internal carotid artery and internal jugular vein were surveyed. [Result]The average length of styloid processes was 24.51 mm?3.05 mm, left was 24.33 mm?3.15 mm and right was 24.69 mm?2.95 mm. The anterior inclination angle was 19.35??2.92? and medial was 24.00??4.07?.The distance of between styloid processes and carotid canal external aperture, jugular foramen,stylomastoid foramen ,mastoid process , internal carotid artery and internal jugular vein were 8.53 mm?2.45 mm, 2.87 mm?0.88 mm, 2.34 mm?0.74 mm, 17.23 mm?1.85 mm,6.49 mm?1.58 mm and 3.70 mm?1.46 mm respectively. [Conclusion]Sixteen-slice Spiral CT postprocessing demonstrate the stytoid processes in vivo accurately and fully, among which multiple planar reconstruction and surface shaded display are more valuable, because they can evaluate the length and inclination of the styloid processes and the distance between the processes and their surrounding structures in vivo more accurately.

Objective:To investigate the quantification of CD4~+CD25~+ regulatory T cells and distribution of CD4~+CD8~+ T lymphocyte subgroup in peripheral blood of patients in chronic hepatitis B (CHB),and to reveal relationship between CD4~+CD25~+ regulatory T cells,CD4~+CD8~+ T lymphocyte subgroup and HBV infetion as well.Methods:CD4~+CD25~(high),CD4~+CD25~+Foxp3~+Treg and CD3~+CD4~+CD8~+T lymphocyte subgroup in peripheral blood from 50 patients with CHB and 20 healthy controls was analyzed using flow cytometry.HBV DNA was detected by fluorescence quantitative PCR.Results:The number of CD4~+CD25~(high)Tregs in patients with CHB was obviously higher than that in healthy controls(P＜0.01)and increased with copies of HBV DNA.The same with the change of CD4~+CD25~+Foxp3~+Tregs in patients with CHB and there was a positive correlation between CD4~+CD25~(high)Tregs and CD4~+CD25~+Foxp3~+Tregs(r=0.890,P＜0.001).Compared with healthy controls,the frequency of CD4~+T cells and the ratio of CD4~+/CD8~+ in patients with CHB was declined,but there was no significant difference in the frequency of CD3~+T cells and CD8~+T cells between them(P＞0.05).The variation in the number of CD4~+CD25~(high)Tregs was correlated positively with the copies of HBV DNA(r=0.782,P＜0.001)and glutamic-pyruvic transaminase(ALT)(r=0.432,P＜0.005)separately,but negatively with the frequency of CD3~+,CD4~+,CD8~+T cells and the ratio of CD4~+/CD8~+(P＞0.05).The variation in the frequency of CD3~+,CD4~+,CD8~+T cells and the ratio of CD4~+/CD8~+ was also correlated negatively with the copies of HBV DNA(P＞0.05).Conclusion:The number of CD4~+CD25~(high)Tregs increases in patients with CHB and is in accordance with the copies of HBV DNA and increased level of ALT.Further studies should be done to investigate weather CD4~+CD8~+ T lymphocyte subgroup could be used to monitor the state of community.

Objective To discusse the research status ofyanjing medical school experience method, and provide a reference for relative researchers.Methods Articles in TCM Database System, SinoMed and CNKI were retrieved refined and analyzed t from the year, author, journal, organization and so on after duplicate removing.Results There were 389 related articles published in journals from 1956 to 2012. With 41.90% articles published in 10 core journals, written by 14 main authors. There were 3 hot research areas including experience of famous doctor,syndrome differentiation and treatment and medical record.Conclusion There was lacking of combing on Yanjing medical school and inheritance system. The inheritance of Yanjing medical school was done far from good, and the study on how to inherit Yanjing medical school need strengthening.

Objective:To construct the plant expression vector of Der f1 allergen of Dermatophagoides pteronyssinu and expression in tobacco lamina.Methods:The Der f1 gene was amplified from the glycerin bacterium which contained pET28a(+)-Der f1 plasmid,cloned into the pMD 19-T plasmid,and then sequenced.The Der f1 gene was digested by ClaⅠand SalⅠ,and cloned into potato virus X (PVX) to construct plant expression vector PVX-Der f1,and then was transformed agrobacterium tumefaciens.The positive one was selected to infect tobacco lamina for expressing target protein.The protein was identified and analysed by SDS-PAGEand Western blot.Results:Digestion and sequence analysis confirmed that the plant expression vector was correct,and the SDS-PAGE and Western blot results showed that the molecular weight of the protein was about 34M_r and it could specific binding with positive serum.Conclusion:The plant expression vector of Der f1 is successfully constructed and the recombinant protein is also produced.

In order to scientifically classify ancient TCM literature and promote the development and use of the literature, this paper combs the classification methods used in the bibliography literature, the modern databases and the libraries of TCM, analyzes the features of the classification methods, summarizes the existing problems in the classify of ancient TCM literature, and proposes the solutions.

Objective To analyse distribution and antibacterial resistance status of pathogenic bacteria isolated from blood cul‐tures of hospitalized infants ,in order to provide references for rational use of antimicrobial agents in the treatment of bloodstream infection .Methods A total of 299 strains of pathogenic bacteria isolated from positive blood culture specimens from infants(3 or less than 3 months of age) suspected with bloodstream infections in this hospital from January 2011 to May 2015 were collected ,the bacteria identification and drug sensitivity test were carried out by using the VITEK 2 Compact automatic microorganism analyzer . The composition and antibacterial resistance of these isolates were analyzed .Results Among the 299 strains of pathogenic bacteria , there were 169 strains of gram‐positive cocci(accounted for 56 .5% ) ,including 95 strains of coagulase negative Staphylococcus (ac‐counted for 31 .8% ) which was the main isolates ,and followed by 28 strains of Staphylococcus aureus(accounted for 9 .4% );there were 120 strains of gram‐negative bacilli (accounted for 40 .1% ) and mainly were Escherichia coli (53 strains ,accounted for 17 .7% );otherwise ,there were 8 strains of fungi (accounted for 2 .7% ) and 2 strains of gram‐positive bacillus (accounted for 0 .7% ) .The results of drug susceptibility test indicated that the gram‐positive cocci had multiple drug resistance to antibacterial a‐gents except for vancomycin and linezolid;the gram‐negative bacilli shown multiple drug resistance except for amikacin ,imipenem and meropenem .The fungus ,however ,displayed high sensitivty to all antifungal drugs .Conclusion Gram‐positive and gram‐nega‐tive bacteria are the main pathogens of hospitalized infants with bloodstream infection ,and are severely resistant to antibacterial a‐gents .Rational use of antimicrobial agents should be recommend for improving clinical efficacy and prohibiting the emergence of drug‐resistant strains .

ObjectiveObjective To investigate the value of pancreatic elasticity based on shear wave elastography (SWE) in predicting the risk of pancreatic fistula, to measure pancreatic hardness by determining the SWE level of the pancreatic body, and to reduce the development of pancreatic fistula after pancreaticoduodenectomy. MethodsA retrospective analysis was performed for the clinical data of 53 patients who were admitted to The Affiliated Hospital of Binzhou Medical University from October 2017 to February 2019 and underwent pancreaticoduodenectomy (PD). The 53 patients were divided into pancreatic fistula group with 10 patients who developed pancreatic fistula after PD and non-pancreatic fistula group with 43 patients who did not develop this disease after PD. The elasticity value of the pancreatic body measured by SWE was used to reflect the tissue elasticity of the pancreas and evaluate the hardness of the pancreas. The t-test was used for comparison of continuous data between two groups, and the chi-square test was used for comparison of categorical data between groups. A logistic regression analysis was used for univariate analysis, and the Spearman’s correlation coefficient was used to investigate the correlation between SWE and other laboratory data. ResultsBody mass index (BMI) (t=1.321), preoperative total bilirubin (t=1.347), diameter of the main pancreatic duct (t=1.385), maximum SWE value (t=1.728), mean SWE value (t=1.634), and intraoperative pancreatic hardness (χ2=4.983) were risk factors for pancreatic fistula (all P＜0.05). Maximum SWE value and mean SWE value were negatively correlated with age and time of operation (maximum SWE value: r=-0.329 and -0.260, both P＜0.05; mean SWE value: r=-0.282, and -0.282, both P＜0.05) and positively correlated with BMI (r=0.275 and 0.350, both P＜0.05). ConclusionSWE level is an independent risk factor for pancreatic fistula after pancreaticoduodenectomy, and the SWE level of the pancreatic body has a high predictive value and guiding significance in surgical treatment.

OBJECTIVE：To study the effects of limonin on immune function and apo ptosis-related factors expression in MFC gastric cancer bearing model mice. METHODS ：MFC gastric cancer bearing model was established by inoculating MFC gastric cancer cells into the right armpit of mice. After modeling ，model mice were divided into model group ，cyclophosphamide group （positive control ，25 mg/kg）and limonin high-dose ，medium-dose and low-dose groups （100，50 and 25 mg/kg），with 10 mice in each group. Other groups were given relevant medicine intragastrically ，once a day ，for consecutive 14 days，except that model group was given 0.5％ sodium carboxymethyl cellulose intragastrically. Before administration and after last administration ，the body weight of mice was measured ；spleen，thymus and tumor tissue were taken after the last administration to calculate the spleen index，thymus index and tumor inhibition rate. The percentage of CD 4+ and CD 8+ T lymphocytes ，CD4+/CD8+ ratio were detected by flow cytometry. The expression of immune function related indexes （IL-2，IL-10，IFN-γ）in serum were detected by ELISA. RT-PCR and Western blot assay were adopted to detect relative mRNA and protein expression of apoptosis-related factors [cytochrome C （Cyt-C），Bcl-2，Bax] in tumor tissue of mice. RESULTS ：There was no significant difference in body weight among the other groups except that of cyclophosphamide group was decreased significantly ，compared with model group （P＜0.05）. Inhibitory rate of tumor were （58.16 ± 7.07）％ ，（37.09 ± 4.26）％ ，（27.30 ± 3.64）％ ，（15.13 ± 2.95）％ in cyclophosphamide group ，limonin high-dose ，medium-dose and low-dose groups. Compared with model group ，spleen index ， thymus index ，the percentages of CD 4+ and CD 8+T lymphocyte cells ，CD4+/CD8+ ratio，serum levels of IL- 2 and IL- 10，relative mRNA and protein expression of Bcl- 2 in tumor of mice in cyclophosphamide group as well as the expression of IL- 10，relative mRNA and protein expression of Bcl- 2 in limonin groups were decreased significantly （P＜0.05）. The expression of IFN-γ，relative mRNA and protein expression of Cyt-C and Bax of cyclophosphamide group as well as spleen index （except for low-dose group ）， thymus index ， the percentage of CD 4 + and CD 8 + T lymphocytes，CD4+/CD8+ ratio，the expression of IL- 2 and IFN-γ，and relative mRNA and protein expression of Cyt-C and Bax in limonin groups were increased significantly （P＜0.05）. CONCLUSIONS ：Limonin can inhibit tumor growth in MFC gastric cancer bearing model mice ，and the side effects are relatively weak. Its mechanism is related to the improvement of immune function and the induction of apoptosis.

ObjectiveTo investigate the effects of thyme herbal tea （BLX） on the proliferation of human coronavirus OC43 （HCoV-OC43） in vitro and in vivo. MethodThe chemical composition of BLX was analyzed by UPLC-MS. The cytotoxicity of BLX in HRT-18 cells and the effect of BLX treatment on the proliferation of HCoV-OC43 in cells were analyzed. Copies of viral gene were detected by real-time PCR. The effect of BLX treatment on the life cycle of HCoV-OC43 was detected by time-of-addition assay. The maximum tolerated dose of BLX and the influences of BLX on the body weight and survival time of suckling mice infected with HCoV-OC43 were determined. The expression of viral protein in the brain and lung tissue was analyzed by immunohistochemistry. ResultThere were 11 chemical components identified in BLX by UPLC-MS. BLX showed the 50% cytotoxic concentration （CC₅₀） of （13 859.56±319） mg·L^-1， the median inhibitory concentration （IC₅₀） of （1 439.09±200） mg·L^-1， and the selection index of 8.26-11.44 for HCoV-OC43 in HRT-18 cells. Compared with the cells infected with HCoV-OC43， BLX at the concentrations of 1 500， 1 000， 500 mg·L^-1 inhibited the proliferation of this virus （P<0.05， P<0.01）. BLX exhibited antiviral effect in the early stage of virus infection， and the inhibition role in the attachment stage was more significant than that in the entry stage （P<0.05）. In the suckling mice infected with HCoV-OC43， BLX at 1200 and 600 mg·kg^-1·d^-1 alleviated the symptoms， prolonged the survival period， reduced the death rate， and down-regulated the mRNA level of nucleocapsid protein in the mice. Moreover， BLX at 1 200 mg·kg^-1·d^-1 down-regulated the expression of nucleocapsid protein in the brain （P<0.01） and the lung （P<0.01）. ConclusionBLX contained multiple antiviral ingredients. It inhibited the proliferation of HCoV-OC43 both in vitro and in vivo by interference with viral attachment. This study provides theoretical reference for the treatment of acute respiratory tract infection with HCoV-OC43 and for further development and application of BLX.