ObjectiveTo observe the effect of bee acupuncture therapy on clinical symptoms and signs， as well as the level of inflammatory cytokine interleukin-6 （IL-6） in synovial fluid of patients with knee osteoarthritis. MethodsThe 94 patients with knee osteoarthritis were divided into the control group and the trial group by the random number table method， with 47 cases in each group. Both groups were given one tablet （60 mg） of etoricoxib orally every morning for 2 weeks. The control group also received microneedle shallow acupuncture therapy， once a day for 5 consecutive times followed 2-day pause， and continued 5 consecutive times， as a course of treatment； the trial group was treated with bee acupuncture therapy once every 2 days， 2 times a week， and 4 times as a course of treatment. Both groups have a course of treatment for 2 weeks. The changes in clinical symptoms and signs of patients in the two groups were observed and evaluated before treatment， after 1- and 2-week treatment， and 12-week follow-up and the differences in Lequesne index scores， HSS scores， Visual Analogue Scale （VAS） scores and IL-6 levels in knee synovial fluid between the two groups of patients were also compared. ResultsNo patients lost to follow up in either group. The Lequesne index scores and VAS scores were lower， and the HSS scores were higher in both groups at all time points after treatment compared with those before treatment （P＜0.05）. Compared at the same time after treatment， the Lequesne index scores and VAS scores of the trial group were lower than those of the control group， and the HSS scores were higher than those of the control group （P＜0.05）. IL-6 in synovial fluid was lower in the trial group at the 12-week follow-up than before treatment （P＜0.05）， but there was no significant difference between two groups at each time point（P＞0.05）. ConclusionBee acupuncture therapy for knee osteoarthritis can significantly improve clinical signs and symptoms， but has no significant effect on the level of IL-6 in knee synovial fluid.

Objective:To discuss the effect of Aspergillus fumigatus(Af)on DNA damage and interleukin(IL)-33 expression in the human bronchial epithelial cells,and to clarify its related mechanism.Methods:Different concentrations(1,5,and 10 mg·L-1)of Af were used to stimulate the bronchial epithelial BEAS-2B cells to select the appropriate stimulation concentration.When the BEAS-2B cells were treated with N-acetylcysteine(NAC)and Af,the cells were divided into control group,Af group,NAC group,and Af+NAC group.When the BEAS-2B cells were treated with DNA double-strand break repair inhibitor NU7441 and Af,the cells were divided into control group,Af group,NU7441 group,and Af+NU7441 group.The comet assay was used to detect the percentages of comet tail DNA of cells in various groups;immunofluorescence method was used to detect the expression levels of DNA damage-related protein phosphorylated H2AX(yH2AX)in the cells in various groups;2,7-dichlorofluorescein diacetate(DCFH-DA)fluorescence probe was used to detect the levels of reactive oxygen species(ROS)in the cells in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of interleukih-33(IL-33),thymic stromal lymphopoietin(TSLP),and interleukih-25(IL-25)mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of phosphorylated nuclear factor κB(p-NF-κB),phosphorylated ataxia telangiectasia mutated(p-ATM),and γH2AX proteins in the cells in various groups.Results:Compared with control group,the percentage of comet tail DNA and the expression level of γH2AX in the cells in 1 mg·L-1 Af group showed no significant difference(P＞0.05),while the percentage of comet tail DNA and the expression level of γH2AX in the cells in 5 mg·L-1 Af group were significantly increased(P＜0.01);compared with 5 mg·L-1 Af group,the percentage of comet tail DNA and the expression level of γH2AX in the cells in 10 mg·L-1 Af group were significantly increased(P＜0.01).Compared with control group,the ROS levels in the bronchial epithelial cells in 1 mg·L-1 Af group was significantly increased(P＜0.05);compared with 1 mg·L-1 Af group,the ROS level in the cells in 5 mg·L-1 Af group was significantly increased(P＜0.01);compared with 5 mg·L-1 Af group,the ROS level in the cells in 10 mg·L-1 Af group was significantly increased(P＜0.05).After treatment of NAC,compared with Af group,the percentage of comet tail DNA(P＜0.01),the expression level of γH2AX(P＜0.05),and the ROS level(P＜0.01)in the cells in Af+NAC group were significantly decreased;after treatment of NU7441,compared with Af group,the percentage of comet tail DNA and the expression level of yH2AX in the cells in Af+NU7441 group were significantly increased(P＜0.01).The RT-qPCR results showed that after treatment of NAC,compared with control group,the expression level of IL-33 mRNA in the cells in Af group was significantly increased(P＜0.05);compared with Af group,the expression level of IL-33 mRNA in the cells in Af+NAC group was significantly decreased(P＜0.05);after treatment of NU7441,compared with Af group,the expression level of IL-33 mRNA in the cells in Af+NU7441 group was significantly increased(P＜0.05).The Western blotting results showed that after treatment of NAC,compared with control group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af group were significantly increased(P＜0.05);after treatment of NU7441,compared with Af group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af+NAC group were significantly decreased(P＜0.05);After treat ment of NU7441,compared with Af group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af+NU7441 group were significantly increased(P＜0.05).Conclusion:Af promotes the IL-33 expression in the human bronchial epithelial cells by causing DNA damage,and its mechanism may be related to the activation of ATM/NF-κB signaling pathway.

Objective To construct the knowledge map of Shen Nong Ben Cao Jing;To analyze basic knowledge of materia medica,explore implicit knowledge,and conduct visualization display;To provide methodological references for the study of ancient books.Methods The types of knowledge entities and relationships between entities involved in the Shen Nong Ben Cao Jing were organized and expressed.A training corpus dataset was produced using the BIO sequence labeling method;a self-developed CNLP text labeling system was used for text labeling;the BERT model was used to recognize named entities;the relationships between entities were set based on rules and semantic associations;the data were imported into the Neo4j-community 4.4.9 graph database using Cypher language for storage and visualization display after knowledge fusion;finally a knowledge graph was constructed.Results The knowledge map of Shen Nong Ben Cao Jing included 5 273 nodes and 11 064 relationships.The pattern layer contained 14 entity classes and 16 relationship types.Through Cypher language query,knowledge was visualized from the aspects of TCM classification,medicinal property theory,compatibility of seven emotions and application of TCM.Conclusion The knowledge graph constructed in this study intuitively reflects the knowledge recorded in Shen Nong Ben Cao Jing and the recessive relationship,which is suitable for knowledge mining and intuitive multi-dimensional display of ancient TCM books.

Objective:To reduce the immunogenicity of vaccinia virus vector by replacing the D8L region, which is a neutralizing antibody epitope in vaccinia virus, with an exogenous gene.Methods:A gene fragment encoding influenza virus hemagglutinin (HA) was inserted into the D8L region to replace it using homologous recombination technique. Then, a recombinant vaccinia virus influenza vaccine was constricted. A recombinant vaccinia virus vaccine with the TK region expressing HA was used as a control. The expression of HA was validated by Western blot. BALB/c mice were immunized with the vaccines and the serum antibody titers two weeks after each immunization were evaluated by ELISA and hemagglutination inhibition assay. The protective efficacy of the recombinant vaccinia virus was assessed through a challenge experiment.Results:Western blot confirmed the successful expression of HAD8L protein in the constructed recombinant vaccines. ELISA and hemagglutination inhibition assay showed that after the primary immunization, the anti-HA antibody titer induced by the recombinant vaccinia virus with D8L region mutation was slightly higher than that induced by the vaccine with TK region mutation, and the difference was statistically significant with the increase of immunization times ( P<0.05). The recombinant vaccinia virus with D8L region mutation showed significantly lower immunogenicity than the recombinant virus with TK region mutation after the primary immunization, but there was no significant difference between them with the increase of immunization times ( P>0.05). After H1N1pdm challenge, no virus was detected in the mice immunized with the recombinant vaccinia virus with D8L region mutation and the mice showed mild lung inflammation and less tissue damage. Conclusions:This study indicated that inserting exogenous genes into the D8L region of the neutralizing antibody epitope in the vaccinia virus vector could help to reduce the immunogenicity of the vector itself and enhance the immunogenicity of the exogenous genes. This provided a reference for the use of the vaccinia virus vector as a delivery tool in the field of vaccines or gene therapy.

Duchene muscular dystrophy (DMD) is a serious progressive muscular dystrophy.Reports in recent years about abnormal lipid in DMD patients have increased, yet little attention has been paid to liver lipid.This study aimed to explore the effect of dystrophin gene defect on liver lipid synthesis.7-week-old mdx male mice were used as DMD model.The conditions of liver function, liver lipid accumulation and liver lipid synthesis were determined through liver tissue morphological examination, blood biochemical examination, and detection of hepatic gene and protein expression.The results showed that lipid droplets in liver of mdx mice increased significantly.The contents of total cholesterol and triglyceride in liver, aspartate aminotransferase and alanine aminotransferase in serum increased.The gene and protein expression of hepatic lipid synthesis-related enzymes such as fatty acid synthase, acetyl CoA carboxylase, and sterol regulatory element binding protein 1-c were up-regulated.These results showed accumulation of liver lipid in 7-week-old mdx male mice.

Objective:To explore the influencing factors of fatigue after stent implantation in patients with airway stenosis.Methods:Totally 104 patients with airway stenosis hospitalized in a hospital from July 2016 to June 2019 were selected, and all the patients were planned to received stent implantation. The patients were evaluated with Identity-Consequence Fatigue Scale (ICFS) postoperatively, and divided into the fatigue group and the non-fatigue group according to the evaluate results. The data of the two groups were consulted, and their gender, age, primary tumor, combined underlying disease, postoperative physical activity, pathological staging, history of radical surgery before stenting, preoperative radiotherapy and chemotherapy, degree of stenosis, type of stenosis, infiltration juga, stent placement site, and re-intervention after stenting were analyzed. Univariate and Logistic regression analysis were performed over the above influencing factors.Results:All the 104 patients with airway stenosis completed postoperative fatigue evaluation. The total score of ICFS was (82.16±8.07) . The scores of postoperative fatigue, attention, vitality, energy, daily activities and the total score were higher than those before operation, and the differences were statistically significant ( P<0.01) . Among the 104 patients with airway stenosis, 28 developed postoperative fatigue, with an incidence of 26.92%. Univariate and multivariate Logistic analysis revealed that postoperative physical activity, pathological staging, infiltrating and re-intervention after stenting were the main influencing factors to fatigue after stent implantation in patients with airway stenosis ( P<0.05) . Conclusions:Patients with airway stenosis have a high incidence of fatigue after stent implantation, which is affected by many factors. Effective intervention measures should be formulated according to the risk factors in order to reduce the incidence of postoperative fatigue.

Objective The effect of different doses of ethylnitrosourea(ENU)and cyclophosphamide(CP)on the loss rate of CD59 on peripheral blood erythrocytes was explored to optimize the detection method of Pig-a gene mutation. Methods According to the weight and loss rate of CD59 on peripheral blood erythrocytes,rats were divided into 4 groups:the control group,CP 40 mg/kg group,ENU 10 mg/kg group and ENU 40 mg/kg group(n=6). The control group was injected i.p. with PBS,other groups were injected i.p. with corresponding solutions. The body weight of rats on days 0,7,14,21, 28, 42 and 56 were recorded. At the same time, blood samples were collected and incubated with antibodies,and the loss rate of RBCCD59-was detected by flow cytometry. Results Compared with the control group, at different time points, the body weight and weight gain of ENU 10 mg/kg group and ENU 40 mg/kg group had no statistically significant difference(P > 0.05),while those in the CP 40 mg/kg group were significantly decreased(P <0.05). The loss rate of RBCCD59-was significantly increased in the CP 40 mg/kg group at 28,42 and 56 days, ENU 10 mg/kg group at 42 and 56 days,and ENU 40 mg/kg group at 7,14,21,28,42 and 56 days,(P < 0.05). The results showed a dose-response relationship. Conclusions Under the conditions of this Pig-a mutation detection method,ENU is superior to CP on raising loss rate of RBCCD59-,ENU 40 mg/kg is better than 10 mg/kg,and 28 days is suitable as the test period.

Objective: Vascular endothelial growth factor A (VEGFA) was investigated as the key protein which might promote the specific metastasis progress of nasopharyngeal carcinoma. Methods: Sixteen specimens of pulmonary metastasis carcinoma and counterparts in primary nasopharyngeal carcinoma tissue were collected from patients. The expression of VEGFA through immunohistochemistry was investigated.VEGFA was knocked down by siRNA in two cell lines of nasopharyngeal carcinoma (CNE-1 and 5-8F), MTT and Transwell test were used to explore the role of VEGFA in praxiology. Then shRNA was used to cultivate the stable CNE-1 cell line with down-regulated-expression of VEGFA. The nude mice models were built through tail vein injection of specific nasopharyngeal carcinoma cells, and lungs were collected to perform further metastasis analysis. Results: Previous genetic studies showed that VEGFA had higher expression in metastasis tissue, and the result was validated in the present study using immunohistochemistry. The percentage of positive cells was 84.8% in pulmonary metastasis group, 51.5% in primary tissue group (t=8.639, P<0.05), average optical density was 0.154 in pulmonary metastasis group, 0.061 in primary tissue group (t=18.791, P<0.05). Low expression of VEGFA inhibited cell viability of optical density value of CNE-1 in siRNA gourp was 0.715, 0.902 in control group (t=7.274, P<0.05); 5-8F in siRNA group was 0.715, 0.935 in control group (t=7.751, P<0.05). Number counting suppressed migration of CNE-1 in siRNA group was 52 per high-power lens, 124 per high-power lens in control group (t=29.380, P<0.05), 5-8F in siRNA group was 65 per high-power lens, 155 per high-power lens in control group (t=18.181, P<0.05). Number counting invasion of CNE-1 in siRNA gourp was 38 per high-power lens, 86 per high-power lens in control group (t=27.665, P<0.05), 5-8F in siRNA group was 52 per high-power lens, 116 per high-power lens in control group (t=40.972, P<0.05) in vitro. Furthermore, knock-down of VEGFA in nasopharyngeal carcinoma reduced the pulmonary metastasis in vivo. Number counting of tumor volumes in shRNA group was 2.4, and 11.0 in control group (t=6.143, P<0.05); average optical density of immunohistochemistry in shRNA group was 0.033, and 0.176 in control group (t=15.734, P<0.05). Conclusions Results above reveal the overexpression of VEGFA in nasopharyngeal carcinoma can facilitate the pulmonary metastasis. Targeting VEGFA may provide a new biomarker of clinical study.

OBJECTIVETo investigate the correlation between CMV reactivation and obliterative bronchiolitis (BO) after allogeneic hematopoietic stem cell transplantation (allo-HSCT).

METHODSFrom January 2011 to December 2013, 769 patients underwent allo-HSCT. The CMV infection was diagnosed by fluorescence quantitative PCR method for detecting the level of CMV-DNA and immunofluorescence staining of PP65 antigen in white blood cell. The frequency of BO in patients with and without CMV infection was compared, and the correlation between CMV infection and BO was analyzed. The clinical data of CMV infection patients with and without BO were analyzed and compared.

RESULTSOf 259 diagnosed CMV infection patients, BO occurred in 32 cases, the incidence rate was 12.35%, while in 510 cases without CMV infection, BO occurred in 8 cases, the incidence was 1.56%. The incidence rate of BO is significantly higher in patients with CMV infection than that in patients without CMV infection (P<0.001). The CMV related clinical data between the 32 cases with BO and 227 cases without BO were analyzed among the 259 cases of diagnosed CMV infection patients. BO incidence is higher in patients with more than 10⁵ copies/ml CMV-DNA than that in patients with less than 10² copies/ml CMV-DNA.

CONCLUSIONAmong the risk factors related to BO post allo-HSCT, CMV infection is one of them to be worthy of attention. CMV reactivation with high virus titer, multiple CMV reactivations and CMV pneumonia are the risk factors.

Allografts ; Bronchiolitis ; Bronchiolitis Obliterans ; Cytomegalovirus ; Cytomegalovirus Infections ; Hematopoietic Stem Cell Transplantation ; Humans ; Real-Time Polymerase Chain Reaction ; Risk Factors ; Viral Load ; Virus Activation

Objective To investigate the effect of clotrimazole on apoptosis of hepatic cells after ischemia-reperfusion injury and its mechanism. Methods Hepatic ischemia-reperfusion rat model was established. Thirty-two male Sprague-Dawley rats were randomly allocated into sham-operated group, model control group, low dose clotrimazole group and high dose clotrimazole group. Apoptosis in hepatic tissue was assessed by TUNEL method. Protein expression levels of CYP3A1,Bcl-2,Bax and PARP were measured by Western blotting. Results As compared with model control group, the apoptosis rate, tissue injury,activity of plasma enzymes and the Bax/Bcl-2 expression ratio were reduced in low and high dose clotrimazole groups. The apoptotic index in both clotrimazole-treated groups was lower than that of model control group with statistically significant difference. CYP3A1 expression was significantly induced by clotrimazole compared to the sham-operated group. Conclusion Clotrimazole may inhibit apoptosis of hepatic cells by up-regulating Bcl-2 and down-regulating Bax, thus produce a protective effect on hepatic ischemia-reperfusion injury and it is also related to the inhibition of PARP shear.