Objective Clarify the effect of neutrophil extracellular traps (NETs) on endothelial cell injury, and investigate whether the heparin can exert a protective effect on endothelial cells by reducing the endothelial cell injury induced by NETs.Methods Neutrophils (PMN) were obtained from healthy human peripheral blood by Percoll-Histopaque density gradient centrifugation, and was stimulated with phorbol ester (PMA) to induce NETs. The qualitative and quantitative analysis of NETs was detected by immunofluorescence staining combined with fluorescence detector. The NETs were used to induce human umbilical vein endothelial cells (HUVEC)in vitro. Recombinant DNA hydrolytic enzymes (rhDNase) and heparin intervention were added respectively. The activity of HUVEC was measured by methyl thiazolyl tetrazolium (MTT) method after 6 hours.Results PMA can stimulate PMN to produce NETs. Immunofluorescence staining showed the formation of reticular formation around the PMN. The concentration of cell-free DNA in the supernatant of PMN stimulated by PMA was significant increased compared with the control group through the detection of PicoGreen fluorescent labeling instrument (2 hours: 119.62±14.83 vs. 24.27±0.67, 4 hours: 146.67±21.24 vs. 28.35±2.98, bothP < 0.05). Application of NETs to stimulate the HUVEC, cell damage was dose dependent and inhibition rate increased gradually. The endothelial cell inhibition induced by NETs can be antagonized after adding rhDNase [10μg/L NETs: (8.65±0.51)% vs. (10.99±0.35)%, 20μg/L NETs:(14.85±0.43)% vs. (16.85±0.49)%, 30μg/L NETs: (26.06±3.51)% vs. (27.54±0.62)%, allP < 0.05]. Heparin with different concentrations were added into the experimental group (0.01, 0.1, 1, 10 kU/L). We found that the endothelial cell inhibition rate decreased compared with control group [10μg/L NETs: (8.96±0.70)%, (5.32±1.36)%, (0.70±0.30)%, (0.75±0.20)% vs. (10.99±0.35)%; 20μg/L NETs: (15.57±0.62)%, (13.28±0.65)%, (6.91±0.15)%, (5.86±0.17)% vs. (16.85±0.49)%; 30μg/L NETs: (30.49±0.74)%, (29.41±1.41)%, (23.45±0.75)%, (21.72±1.52)% vs. (27.54±0.62)%, allP < 0.05].Conclusions NETs can induce endothelial cell injury, and the injury degree was increased with the concentration of NETs. Heparin can reduce endothelial cell injury induced by NETs, which may be a potential mechanism for the protective effect of heparin on sepsis.

Objective To investigate the influence of heparin pretreatment on serum and lung tissue level of neutrophil extracellular traps (NETs) in septic mice model and its molecular mechanism.Methods Ninety male C57BL/6J mice were randomly divided into control group (n = 30), lipopolysaccharides (LPS) group (n = 30, 30 mg/kg LPS in 100μL normal saline was intraperitoneally injected) and LPS+heparin group (n = 30, 8 U of heparin in 20μL normal saline was subcutaneously injected 30 minutes before the injection of LPS). Six hours later of LPS injection, blood was collected and lung tissue was harvested. Enzyme linked immunosorbent assay (ELISA) was used to assess the concentration of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and histones 2AX (H2AX), neutrophil elastase (NE), which reflected NETs concentration. PicoGreen fluorescent dyes was used to detect serum circulating free DNA (cf-DNA/NETs) concentration. The protein expression levels of H2AX and NE in lung tissue were examined by Western Blot.Results The serum concentrations of TNF-α, IL-6, H2AX, NE, cf-DNA/NETs, and the protein expression levels of H2AX and NE in lung tissue of septic mice were significantly higher than those of control group [TNF-α (ng/L): 133.0±14.1 vs. 2.7±1.0, IL-6 (ng/L): 3911.2±189.2 vs. 298.9±52.5, H2AX (ng/L): 545.5±40.0 vs. 21.9±8.3, NE (μg/L): 6.48±0.12 vs. 0.47±0.15, cf-DNA/NETs (μg/L): 846.3±137.5 vs. 152.7±36.4, H2AX protein (gray value): 1.14±0.09 vs. 0.68±0.04, NE protein (gray value): 0.56±0.03 vs. 0.32±0.04, allP < 0.05]. After heparin pretreatment, levels of serum TNF-α, H2AX, NE, cf-DNA/NETs, and protein expression levels of H2AX and NE in lung tissue were significantly reduced [TNF-α (ng/L): 83.2±7.6 vs. 133.0±14.1, H2AX (ng/L): 435.0±39.0 vs. 545.5±40.0, NE (μg/L): 4.26±0.17 vs. 6.48±0.12, cf-DNA/NETs (μg/L): 606.5±73.9 vs. 846.3±137.5, H2AX protein (gray value): 0.91±0.03 vs. 1.14±0.09, NE protein (gray value): 0.42±0.03 vs. 0.56±0.03, allP < 0.05], but no significant change was found in IL-6 (ng/L: 3919.9±166.6 vs. 3911.2±189.2,P > 0.05).Conclusion Heparin pretreatment could significantly decrease the level of NETs in serum and lung tissue, and can be the potential mechanism of its organ protection in sepsis.

Objective To study the role of Ⅲb isoform of human fibroblast growth factor receptor 1 (FGFR1-Ⅲb) in PANC-1 pancreatic cancer cells. Methods The plasmid of human full-length FGFR1-Ⅲb isoform，pSVK4/FGFR1-Ⅲb, was stable transfected into cultured PNAC-1 pancreatic cancer cell lines facilitated by lipofectamine. The function of FGFR1-Ⅲb in transfected pancreatic cancer cells were examined by MTT assay, soft agar assay, cell migration assay, single cell movement assay, In vivo tumorigenicity assay. Results The basal anchorage-dependent and -independent cell growth was significantly inhibited. Additionally, FGFR1-Ⅲb expression inhibited single cell movement and in vitro invasion as determined by time-lapse microscopy and boyden chamber assay as well as in vivo tumor formation and growth in nude mice. Microscopic analysis of the xenograft tumors revealed a reduced Ki-67 labelling, lower amount of tumor necrosis and higher grade of differentiation in FGFR1-Ⅲb expressing tumors. Conclusion We identified a functional human FGFR mRNA splice variant that inhibits the transforming potential of pancreatic cancer cells.

Objective:To construct a clinical-radiomics model based on MRI, and to explore its predictive value for biochemical recurrence (BCR) after radical prostatectomy in prostate cancer patients.Methods:A total of 212 patients with prostate cancer who underwent radical prostatectomy in the First Affiliated Hospital of Soochow University from January 2015 to December 2018 and had complete follow-up data were retrospectively analyzed. The random toolkit of Python language was used to randomly sample the patients at a ratio of 7∶3 without replacement, and they were divided into a training set (149 cases) and a test set (63 cases). The endpoint of follow-up was BCR or at least 3 years. BCR occurred in 50 patients in the training group and 21 patients in the test group. The imaging features of the main lesion area in the preoperative T 2WI, diffusion-weighted imaging and apparent diffusion coefficient map of patients in the training set were extracted, and the unsupervised K means clustering algorithm was used to screen the features. The selected features were fitted by a multivariate Cox regression model, and the radiomics model was constructed. Univariate Cox regression analyses were used to screen the main clinical risk factors associated with BCR, and the clinical-radiomics model was constructed combined with RadScore. In the test set, the time-dependent receiver operating characteristic (ROC) curve was constructed, and the area under the curve (AUC) was calculated to evaluate the predictive efficacy of the radiomics model, clinical-radiomics model and prostate cancer risk assessment after radical resection (CAPRA-S) score for the occurrence of BCR. Harrell consistency index (C-index) was used to evaluate the model to predict BCR consistency. The calibration curve was used to evaluate the degree of variation of the model. The decision curve was used to evaluate the clinical application value of the prediction model. Results:A total of 26 radiomics features were screened to establish the radiomics model. The univariate Cox showed that the preoperative clinical features included preoperative prostate-specific antigen level (HR=1.006, 95%CI 1.002-1.009, P=0.001), Gleason score of biopsy (HR=1.422, 95%CI 1.153-1.753, P=0.001), clinical T stage (HR=1.501, 95%CI 1.238-1.822, P<0.001). The multivariate Cox showed that the RadScore was an independent predictor of BCR after radical prostatectomy (HR=51.214, 95%CI 18.226-143.908, P<0.001). The selected preoperative clinical features were combined with RadScore to construct a clinical-radiomics model. In the test set, the AUCs of the time (3 years)-dependent ROC curves of the radiomics model, the clinical-radiomics model, and the CAPRA-S score were 0.824 (95%CI 0.701-0.948), 0.841 (95%CI 0.714-0.968), and 0.662 (95%CI 0.518-0.806), respectively. The C-index of the radiomics model, clinical-radiomics model and CAPRA-S score were 0.784 (95%CI 0.660-0.891), 0.802 (95%CI 0.637-0.912) and 0.650 (95%CI 0.601-0.821), respectively. The calibration curve showed that the predicted probability and actual probability of BCR by radiomics model, clinical-radiomics model and CAPRA-S score were in good agreement (χ 2=7.64, 10.61, 6.37, P=0.465, 0.225, 0.498). The decision curve showed that the clinical net benefit of the clinical-radiomics model and the radiomics model was significantly higher than the CAPRA-S score. When the threshold probability was 0.20-0.30, 0.40-0.50, and >0.55, the clinical net benefit of the clinical radiomics model was higher than that of the radiomics model. Conclusions:The clinical-radiomics model can effectively predict the occurrence of BCR in patients with prostate cancer after radical prostate ctomy, and the prediction efficacy is better than the radiomics model and CAPRA-S score.

【Objective】 To understand the frequency and significance of anti-" Mi^a" (anti-" Mi^a" mixtures of antibodies) in local population in Zhongshan, and the influence of different experimental conditions on the activity of human anti-" Mi^a" . 【Methods】 The microplate-based agglutination assay and polybrene method were used to screen anti-" Mi^a" in 3 587 blood samples from voluntary blood donors and patients using O type red blood cells with positive Mi^a antigen, then.rechecked by tube method and microcolumn gel card method. 【Results】 The frequency of anti-" Mi^a" was 1.06% (38/3 587), among which 60.5% (23/38) were IgM and 39.5% (15/38) were mixture of IgM and IgG; 0.61% (13/2 135) in local blood donors and 1.72% (25/1 452) in patients(P<0.01). 65.8% (25/38) of the population carrying anti-" Mi^a" had a history of immunity. 57.9% (22/38) were identified to be anti-" Mur" and 42.1% (16/38) anti-" Mi^a" using GP.Vw erythrocyte. The appropriate incubation time for anti-" Mi^a" test was 10 min. 【Conclusion】 The frequency of anti-" Mi^a" was relatively high among blood donors and patients in Zhongshan, and most of the anti-" Mi^a" carriers had a history of immunity. Most anti-Mi^a antibodies were active in saline, and some of them were mixture of IgM and IgG. It may be helpful to include Mi^a positive red blood cells in the irregular antibody screening cell panel to improve the safety of blood transfusion.

【Objective】 To investigate the frequency of CD36 deletion and gene mutation in voluntary blood donors of Zhongshan city, and to explore the possibility of establishing local CD36 negative platelet donor bank. 【Methods】 Platelet CD36 antigen was detected by ELISA in 1 654 voluntary blood donors.Some of the negative samples were confirmed by flow cytometry, and genotyping was also performed. 【Results】 Platelet CD36 antigen was negative in 27 cases, accounting for 1.6% (27/1654), among which 1.6% (18/1149) were males and 1.8% (9/505) were females.No significant difference was noticed between males and females in CD36 antigen deletion cases (P>0.05). Fifteen CD36 negative samples were randomly selected, genotyped and sequenced, with type I deletion in 1 case[ 6.7% (1/15)], type Ⅱ deletion in 14 cases[ 93.3% (14/15)], and gene mutation in exon 3-14 detected in 8 cases. 【Conclusion】 The frequency of platelet CD36 antigen deletion in Zhongshan is comparable to that in other southern regions of China.The establishment of CD36 negative platelet donor bank is conductive to improve the effectiveness of platelet transfusion.