Objective : To explore the effect of Wilms′ tumor 1-associated protein(WTAP) on tissue collagen deposition in pulmonary fibrosis caused by bleomycin. Methods : 60 mice were randomly divided into four groups: control group(Control group), Bleomycin-induced pulmonary fibrosis group(BLM group), pulmonary fibrosis lentivirus empty vector control group(BLM+LV-NC group), pulmonary fibrosis WTAP lentivirus group virus group(BLM+LV-WTAP group). Experimental pulmonary fibrosis mouse model was established by subcutaneous injection of bleomycin(35 mg/kg) into the abdomen, twice a week for a total of 8 times. After modeling, Western Blot was used to detect the protein expression of fibrosis-related markers α-smooth muscle actin(α-SMA), type I collagen(Collagen Ⅰ), fibronectin(Fibronectin), and WTAP protein. Masson staining and Sirius Red staining were used to detect collagen deposition. RT-qPCR was used to detect WTAP mRNA expression, WTAP lentivirus infection effect, and Collagen Ⅰ mRNA expression. Results: Compared with the Control group, the expression of pulmonary fibrosis markers α-SMA(P<0.001), Collagen Ⅰ(P<0.001), and Fibronectin(P<0.01) protein in the BLM group all increased. Masson staining(P<0.001) and Sirius Red staining(P<0.001) confirmed that significant collagen deposition occurred in the lung tissue of the BLM group. In addition, the expression of WTAP protein in the lung tissue of the BLM group increased(P<0.01). Compared with the Control group, the expression of WTAP mRNA in the BLM group increased(P<0.001). Compared with the BLM+LV-NC group, the expression of WTAP mRNA in the tissues of the BLM+LV-WTAP group decreased(P<0.001), proving that virus infection is effective. After infection with WTAP lentivirus, collagen fiber deposition decreased(P<0.001), Collagen Ⅰ mRNA(P<0.001) level decreased, and protein(P<0.001) expression decreased in the BLM+LV-WTAP group. Conclusion Knocking down of WTAP can reduce collagen deposition in bleomycin-induced pulmonary fibrosis tissue in mice and improve experimental pulmonary fibrosis.

Objective: To investigate the effect of sirtuin 2(SIRT2) on the proliferation and migration of cardiac fibroblasts(CFs)in C57BL/6 mice under angiotensin II(Ang Ⅱ) stimulation. Methods : The hearts were taken from 1 to 2 days C57BL/6 milk mice. After cutting and digesting, CFs were extracted by different adhesion centrifugation. After CFs attachment, the cells were cultured under control medium and Ang Ⅱ(100 nmol/L) medium and treated using OE-SIRT2 plasmid to overexpression the SIRT2 gene. RT-qPCR was used to detect mRNA expression of SIRT2 proliferating cell nuclear antigen(PCNA), periostin(POSTN)and type Ⅰ collagen procollagen A1(Col1A1), Western blot assay was used to measure the protein expression levels of SIRT2, PCNA, POSTN and Col1A1, CCK-8 assay and EdU assay were used to evaluate CFs proliferation rate, Transwell experiment was used to assess CFs migration activity. Results: Compared with control group, Ang Ⅱ stimulation led to down-regulation of SIRT2 expression in CFs, increased collagen expression, and promoted CFs proliferation and migration. The expression of SIRT2 was up regulated in CFs treated with OE-SIRT2 plasmid under Ang Ⅱ stimulation, Col1A1, POSTN and PCNA expression was down regulated, and CFs proliferation and migration ability decreased. Conclusion Overexpression of SIRT2 can inhibit the proliferation and migration of CFs under Ang Ⅱ stimulation, indicating that SIRT2 may be a key regulatory point in the onset and progression of cardiac fibrosis.

Objective:To clarify the potential correlation between biological changes of meninges in periodontitis mice and cognitive impairment by analyzing the biological changes of meninges in periodontitis mice using single-cell RNA sequencing.Methods:Thirty C57BL/6 mice were divided into two groups by using random number table method (15 mice in each group). Mice in the control group were locally administered 2% carboxyl methyl cellulose (CMC) without Porphyromonas gingivalis (Pg) on both buccal sides. A mixture of Pg W83 and 2% CMC was applied on both buccal sides in the experimental group mice three times a week, lasting for 16 weeks in total. The absorption of alveolar bone, locomotor activity and cognitive function, the activation of microglia and astrocytes in the cortex were observed and assessed. The mRNA expression levels of Occludin in meninges and brain were detected in two groups. Single-cell RNA sequencing data of meninges were processed by uniform manifold approximation and projection (UMAP). Differential genes expressions of endothelial cells were processed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. In addition, real-time fluorescence quantitative PCR (RT-qPCR) was used to verify the expressions of transcription activating factor 3 (Atf3) and apolpoprotein L domain-containing 1 (Apold 1). Results:Methylene blue staining found the distances of buccal and palatal cement-enamel junction-alveolar bone crest in experimental mice [(185.60±17.60), (206.90±13.37) μm] increased significantly compared with the control group [(135.33±9.57), (163.05±14.98) μm] ( t=5.02, P=0.002; t=4.37, P=0.005). Open field experiment showed the total distance and average speed of mice in the experimental group [(971.88±164.57) cm, (3.25±0.55) cm/s] were not statistically significant compared with the control group [(914.24±278.81) cm, (3.05±0.93) cm/s] ( t=0.65, P=0.525; t=0.65, P=0.520). The recognition index of the experimental group [(48.02±16.92) %] was lower than the control group [(66.27±17.90) %] ( t=2.40, P=0.027) by novel object recognition tests. Compared with the control group [(63.56±11.88) %], the alternation of experimental group [(50.99±14.17) %] was significantly decreased in Y maze tests ( t=2.33, P=0.030). Immunohistochemistry results showed microglia and astrocytes were activated in the cortex of experimental mice. Compared with the control group (1.02±0.25, 1.04±0.31), the relative mRNA expressions of Occludin decreased significantly in the meninges and brain of periodontitis mice, respectively (0.61±0.10, 0.64±0.20) ( t=3.47, P=0.010; t=2.66, P=0.024). By single-cell RNA sequencing, meninges cells were divided into 11 types, such as endothelial cells, fibroblasts, immune cells and so on. Endothelial cells were the main cell types in meninges [the control group: 26.47% (1 589/6 004), the experimental group: 26.26% (807/3 073)]. Compared with the control group [5.56% (334/6 004)], the percentage of granulocytes increased in the periodontitis mice [11.65% (358/3 073)]. Using clustering analysis to further focus on endothelial cells, GO enrichment analysis revealed differential genes were mainly related to angiogenesis, cell adhesion, apoptosis and so on. KEGG enrichment analysis revealed that differential genes were related to signaling pathways of interleukin-17, relaxin and so on. The relative mRNA expressions of Atf3 and Apold1 in meninges of periodontitis mice (0.42±0.24, 0.54±0.27) were significantly lower than the control group (1.03±0.26, 1.02±0.23) ( t=3.88, P=0.005; t=3.02, P=0.017). Conclusions:The mice chronically infected with Pg W83 occurred memory impairment, neuroinflammation and changes of barrier function. In the meninges of periodontitis mice, there were infiltration of immune cells and down-regulation expressions of Atf3 and Apold1 by single-cell RNA sequencing. Meningeal immunity and changes of barrier function may play an important role in the cognitive impairment caused by periodontitis.

Objective:To investigate the effects of persistent isolated hypothyroxinemia in the first and second trimester of pregnancy on complications and adverse outcomes of pregnancy.Methods:A retrospective analysis was conducted in 784 pregnant women including 111 cases of persistent isolated hypothyroxinemia in the first and second trimester of pregnancy and 673 pregnant women with normal thyroid function as control group. All women were registered and delivered in the Department of Obstetrics of our hospital from April 2016 to April 2017. The complications and adverse outcomes of pregnancy in the two groups were analyzed.Results:Age, body weight before pregnancy, body mass index(BMI), 1 h plasma glucose and 2 h plasma glucose during oral glucose tolerance test in persistent isolated hypothyroxinemia group were higher than those in control group( P<0.05), with increased incidence of anemia during pregnancy( P<0.05). However, there were no significant differences in the incidences of gestational diabetes mellitus and gestational hypertension between the two groups( P>0.05). No significant statistical differences were found in macrosomia, stillbirth, neonatal malformation, postpartum hemorrhage, acute delivery, premature delivery, fetal intrauterine development delay, and small full-term infants between the two groups( P>0.05). Logistic regression analysis showed that age( OR=1.1, 95% CI 1.0-1.1, P=0.002) and pre-pregnancy body weight( OR=1.0, 95% CI 1.0-1.1, P=0.046) were risk factors for the occurrence of persistent isolated hypothyroxinemia in the first and second trimesters of pregnancy. Persistent isolated hypothyroxinemia in the first and second trimesters was associated with anemia during pregnancy( OR=1.9, 95% CI 1.1-3.2, P=0.024). Conclusions:Pregnant women who are older and heavier before pregnancy should pay more attention to their thyroid function. Pregnant women with persistent isolated hypothyroxinemia in the first and second trimesters should be concerned for anemia.

Objective Through the study of color Doppler ultrasound imaging, to explore the mechanism of portal vein thrombosis for patients with liver cirrhotic portal hypertension splenectomy and its influencing factors. Methods From January 2013 to December 2013, a total of 284 cases of patients with cirrhosis and portal hypertension plenectomy plus pericardial devascularization in 302 Hospital of PLA were reviewed. Color Doppler ultrasound technique was employed to record the thrombosis of portal vein, splenic vein and superior mesenteric vein before and after the surgical procedure. According to the occurrence of portal vein thrombosis after splenectomy, patients were divided into portal vein thrombosis group and non portal vein thrombosis group; the factors that may affect the formation of portal vein thrombus were analyzed by using Logistic regression. Results One hundred and twenty-nine cases of portal vein thrombosis occurred among the 284 patients with splenectomy, the incidence rate was 45.42% (129/284). Logistic regression analysis shows that inner diameters of splenic vein for the group with portal vein thrombosis are significantly different from that of the group without portal vein thrombosis (Z=2.034, P < 0.05), postoperative inner diameter of portal vein (Z=2.037, P<0.05), and prothrombin time (Z=-2.171, P<0.05) are significantly higher in the group with portal vein thrombosis, while preoperative platelet count is significantly lower in the group with portal vein thrombosis (Z=-2.146, P < 0.05); gender, preoperative hepatic artery flow velocity, and blood coagulation time are also the influencing factors of portal vein thrombosis (all P>0.05). Conclusion Color Doppler ultrasound technology can not only monitor, the thrombus formation in portal venous system post splenectomy, it can assess the risk factors of portal vein thrombosis post splenectomy.

Objective To explore the clinical application value and feasibility of contrast-enhanced ultrasonography (CEUS) in rapid diagnosis for bleeding of hepatocellular carcinoma. Methods From January 2009 to December 2012, 58 patients from the 302nd Hospital of the People′s Liberation Army underwent CEUS and conventional ultrasound, who were suspicioused primary bleeding of liver cancer or secondary bleeding of liver cancer after percutaneous radiofrequency ablation (RFA). The change of effusion volume in front of liver and in abdominal cavity, the perfusion time, range and characteristics of contrast agent, tumor vessel and the characteristic manifestation of active bleeding within the tumor and subcapsular were evaluated. Theχ2 test was used to compare the show rates of lesion boundary and the bleeding point by conventional ultrasound and CEUS. The t test was used to compare the change of effusion in front of liver and in abdominal cavity from 0 min to 30 min after bleeding were diagnosis by CEUS. Results For 58 cases of suspicious rupture and bleeding of liver cancer, 34 cases were conifrmed by clinical and imaging ifndings. The bleeding points were detected by conventional ultrasound in 5 cases. The bleeding points in 30 cases (including 4 cases of rapid bleeding, 7 cases of medium bleeding and 19 cases of slow bleeding) were detected by CEUS. The detection rate of lesion boundary, tumor vessel and bleeding point of CEUS were higher than those of conventional ultrasound, and the differences were statistically significant (χ2=10.350, P=0.001; χ2=4.300, P=0.034;χ2=36.790, P=0.0007). CEUS showed that contrast agent continuously concentrated at the bleeding point of lesions, and different degrees of bleeding were shown as′jet-like′,′linear′or′intermittent′spillover. The volume of effusion in front of the liver and in abdominal cavity at 30 min was more than those at 0 min after bleeding were diagnosis by CEUS, and the differences were statistically significant (the volume of effusion in abdominal cavity:t=-3.467, P=0.026;t=-12.895, P=0.000;t=-3.055, P=0.007;the volume of effusion in front of the liver:t=-8.110, P=0.001;t=-5.642, P=0.002;t=-5.981, P=0.000). Conclusions CEUS can show direct signs of rupture and bleeding of liver cancer, and the degree of bleeding can be evaluated according to the extravasation characteristics of contrast medium and the changes of lfuid volume. Therefore, CEUS can provid an objective basis for rapid diagnosis of ruptured hepatocellular carcinoma in the emergency scene and bedside.

Objective To study the detection rate of contrast-enhanced ultrasound (CEUS) with different perfusion methods on rabbit VX2 small hepatocellular carcinoma less than 10.0 mm. Methods VX2 tumor cells were inoculated into the subcutaneous tissue of New Zealand rabbit′s thigh. Then the plant tumor were cut into small blocks under sterile conditions and transplanted into hepatic parenchyma in 30 New Zealand rabbits. The contrast media was injected through peripheral vein by single and double perfusion methods. The detection rate of two methods were compared. Results There were 41 hepatocellular carcinoma lesions in the 30 rabbits. There were 15 lesions with size between 3.0 mm and 5.0 mm, and 26 lesions between 5.0 mm and 10.0 mm in diameter. On CEUS, the VX2 tumor presented fast-in and fast-out pattern. In arterial phase, the lesion was enhanced rapidly. In portal venous phase, contrast began to wash out from the carcinoma. In delay phase, the enhancement of lesion was signiifcantly lower than the surrounding normal liver parenchyma. A total of 32 lesions were detected by single perfusion method, including 7 lesions ranging 3.0-5.0 mm and 25 lesions ranging 5.0-10.0 mm. A total of 39 lesions were detected by double perfusion method, including 13 lesions ranging 3.0-5.0 mm and 26 lesions ranging 5.0-10.0 mm. The detection rate of micro-hepatocellular carcinoma by single and double perfusion method was 78% and 95% respectively. The difference was statistically signiifcant (χ2=5.150, P=0.023). The detection rate of 3.0-5.0 mm lesions by single and double perfusion method was 47%and 87%, respectively. The difference was statistically signiifcant ( χ2=5.400, P=0.025). The detection rate of 5.0-10.0 mm lesions by single and double perfusion method was 96% and 100%, respectively. There was no statistically signiifcant difference (χ2=1.020, P=0.500). Conclusion The double perfusion method greatly promotes the detection of micro hepatocellular carcinoma, especially for the lesions less than 5.0 mm in diameter.

Objective To evaluate the effects of small interfering RNA (siRNA) against peptidylarginine deiminase 4 (PADI4) gene on apoptosis of fibroblast-like synoviocytes (FLS) from synovium of rheumatoid arthritis (RA).Methods The siRNA targeting PADI4 was constructed and transfected into FLS cells in RA via LipofectamineTM 2000.The expression level of PDAI4 mRNA was detected by using real-time quantitative polymerase chain reaction (real-time PCR).The protein expression of PADI4,CyclinB1 and P21 was detected by Western blotting.The apoptosis of FLS cells in RA was examined by flow cytometry.The levels of IL-1β were detected by ELISA.T-test was used for statistical analysis.Results siRNA-PADI4 efficiently down-regulated the PADI4 expression compared with control group,1.00±0.20 vs 0.38±0.20 (t=9.607,P＜0.01),0.39±0.23(t=8.394,P＜0.01).FCM analysis showed that the percentage of apoptosis cells in PADI4 siRNA group in FLS was (5.4±0.6)％ (t=-19.223,P＜0.01) and (6.1±0.6)％ respectively (t=-24.229,P＜0.01),which was significantly higher than that in the control group in FLS (1.6±0.3)％.The expression of CyclinB1 protein was decreased,and P21 increased.The concentrations of IL-1β in culture medium of the transfected group were (26.8±0.7) ng/ml (t=-10.747,P＜0.01) and (27.7±0.7) ng/ml (t=-10.967,P＜0.01),higher than the control group [(23.9±0.7) ng/ml].Conclusion After being transfected with PADI4 siRNA,the apoptosis of FLS cells in RA is increased.Our results have demonstrated the potential role of CyclinB1 and P21 in PADI4 signaling.

Objective To compare the clinical efficacies of vasiculation moxibustion with pastes prepared in different ways and at different acupoint groups in treating allergic rhinitis due to deficient cold of lung qi. Method Eighty subjects were randomized into 4 groups, 20 in each group. Raw Bai Jie Zi (Semen Brassicae) was used in the vesiculation paste for group 1 and 2;stir-baked Bai Jie Zi (Semen Brassicae) was used for group 3 and 4. In group 1 and group 3, bilateral Feishu (BL 13), Fengmen (BL12), and Gaohuang (BL 43) were selected; in group 2 and group 4, bilateral Dazhu (BL11), Geshu (BL17), and Shenshu (BL23) were selected. The symptom and sign scores were compared before and after intervention. Result There were significant differences in comparing the symptoms and signs among the four groups (P＜0.05), and paired comparisons showed that the therapeutic efficacies of group 1 and group 3 were significantly higher than that of group 2 and group 4 (P＜0.05), while the differences in the therapeutic efficacy between group 1 and group 3, group 2 and group 4 were statistically insignificant (P>0.05). Conclusion Vesiculation moxibustion at the point group of Fengmen (BL12), Feishu (BL13), and Gaohuang (BL43) can markedly improve the symptoms and signs of allergic rhinitis due to deficient cold of lung qi;raw and stir-baked Bai Jie Zi (Semen Brassicae) can work equally in vesiculation paste for allergic rhinitis.

Objective:To study clinical effectiveness of sustained inflation (SI)on extrapulmonary acute respiratory disstress syndrome(ARDS).Methods:Twenty-four patients with extrapulmonary ARDS were included from January 2011 to September 2012 .Patients were randomly divided into control group (n=10)and SI group (n=14).Both groups were given lung protec-tive ventilation (LPV),and SI (45 cmH2 O,20 s)was conducted for alveolar recruitment in SI group.Heart rate (HR),mean arterial pressure (MAP),transcutaneous oxygen saturation (SpO2 ),PaO2/FiO2 ,mechanical ventilation time,ICU hospitaliza-tion time were compared between the two groups.Results:SpO2 ,PaO2/FiO2 were increased significantly at 2 h after applying SI in SI group (P <0.05),while there were no significant differences in HR,MAP,mechanical ventilation time,ICU hospitali-zation time between the two groups (P >0.05).No other serious complications were observed.Conclusions:SI can effectively improve oxygenation in patients with extrapulmonary ARDS.It is safe and effective to hemodynamics alveolar recruitment.