Background/Aims: Biliary tract cancer (BTC) frequently harbors KRAS mutations, which are associated with resistance to traditional treatment and a poor prognosis. Synthetic lethality (SL) strategy may provide other targets of KRAS. Therefore, we aim to identify and validate potential therapeutic targets of KRAS for the treatment of BTC via SL. Methods: The dependency (DepMap) projects were used to predict the synthetic lethal gene of KRAS. FDA-approved anticancer drug library was applied to screen potential drugs effective against KRAS-mutant BTC. Furthermore, the synthetic lethal effects or corresponding mechanisms of potential genes and drugs on BTC were investigated using KRAS-mutant and KRAS-wild type BTC cell lines, patient-derived xenografts (PDX), and KRAS oncogene-driven tumor models, as well as other KRAS-mutant cancer cell lines. Results: Initially, we discovered that the loss of GATA2 reduced the viability of KRAS-mutant but not KRAS-wild-type BTC. Subsequently, the drug library screened out disulfiram, which primarily exerts a synthetic lethal effect by inhibiting interleukin-1β (IL-1β) in KRAS-mutant BTC. Mechanistically, GATA2 specifically enhanced the transcription of IL-1β to promote NF-κB signaling in KRAS-mutant BTC. IL-1β inhibition phenocopied GATA2 deficiency, leading to reduced KRAS-mutant BTC viability. These synthetically lethal effects were confirmed using PDX, a KRAS oncogene-driven tumor model, as well as in other KRAS-mutant cancer cell lines. Conclusions In summary, these results indicate that inhibiting GATA2/IL1β could be a therapeutic strategy in KRAS-mutant BTC and potentially other cancers.

Objective To determine the effects of hypoxia pre-treatment combined with radiation damage on the hematopoietic cells in the bone marrow of mice.Methods A total of 165 male C57BL/6 mice(10～12 weeks old,weighing 20～25 g)were randomly divided into 7 groups:normal control(Control,n=33),6 Gy irradiation(6-Gy,n=43),7 d hypoxia-6 Gy irradiation(Hy-7 d+6 Gy,n=43),7 Gy irradiation(7 Gy,n=12),7 d hypoxia-7 Gy irradiation(Hy-7 d+7 Gy,n=12),7 Gy continuous hypoxia treatment(Hy-7 d+7 Gy+Hy,n=12),and 6 Gy continuous hypoxia treatment(Hy-7 d+6 Gy+Hy,n=10).The mice of the hypoxia treatment groups were given 7-day hypoxic pretreatment(12%oxygen)in a normobaric hypoxic chamber,while those of the other groups were housed in normoxic condition.After pretreatment,the mice of the irradiation groups were exposed to a single 6 or 7 Gy of whole-body 60Co γ-irradiation in normoxia.The mice of the hypoxia and irradiation groups were kept in hypoxic condition in 24 h post-irradiation followed by being resumed to normoxia,while those of the continuous hypoxia treatment groups were remained in hypoxia.After bone marrow cell suspensions were prepared from the Control,6 Gy,and Hy-7 d+6 Gy groups,bone marrow nucleated cells(BMNCs)were counted via automated cell counter.HE staining was employed to observe pathologic changes in medullary cavity,and flow cytometry was used to assess Lin-Sca1?c-Kit?(LSK)hematopoietic stem/progenitor cells,myeloid progenitors(MPs),and mature T/B/myeloid cells.The mice of the 7 Gy,Hy-7 d+7 Gy,and Hy-7 d+7 Gy+Hy groups were monitored for 30-day survival after hypoxic pretreatment.The dynamic changes in the counts of red blood cells(RBC),white blood cells(WBC)and platelets(PLT),and hemoglobin(HGB)level were observed in the 6 Gy,Hy-7 d+6 Gy,and Hy-7 d+6 Gy+Hy groups with aid of a fully automatic blood analyzer.Single-cell RNA sequencing was performed on bone marrow cell suspension derived from the mice euthanized in 17 d after irradiation from the Control,6 Gy,and Hy-7 d+6 Gy groups.Results ①Compared to the Control group,the 6 Gy group showed significantly reduced BMNCs(P<0.01),dilated bone marrow sinusoids,and erythrocyte extravasation.The Hy-7 d+6 Gy group exhibited higher cellular density and attenuated BMNC loss than the 6 Gy group(P<0.01).②Flow cytometry revealed less LSK,MP,and mature T/B/myeloid cells in the 6 Gy group than the Control group(P<0.05),and the reduced counts of LSK and MP were mitigated in the Hy-7 d+6 Gy group(P<0.01).③The Hy-7 d+7 Gy group demonstrated improved 30-day survival than the 7 Gy group(P<0.01),while continuous hypoxia(Hy-7 d+7 Gy+Hy)failed to enhance the survival.No statistical difference was seen in the survival rate between the 2 groups(P=0.12),though the Hy-7 d+7 Gy group showing higher survival rate.④Routine blood test revealed that the Hy-7 d+6 Gy group showed faster WBC recovery(vs the 6 Gy and Hy-7 d+6 Gy+Hy groups,P<0.05),higher pre-irradiation RBC/HGB levels,and accelerated PLT restoration(P<0.05).⑤Single-cell RNA sequencing indicated that hypoxia pretreatment suppressed the numbers of long-term hematopoietic stem cells/short-term hematopoietic stem cells(LT-HSC/ST-HSC)depletion in the Hy-7 d+6 Gy group when compared with the 6 Gy group,which was consistent with the results of flow cytometry.Pseudotime trajectory aligned the Hy-7 d+6 Gy group,as the Control group,showed enriched undifferentiated LSKs.Differential gene analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis revealed that oxidative phosphorylation pathway was strongly activated in the 6 Gy group,while the Hy-7 d+6 Gy group had enriched in chromatin remodeling and mRNA surveillance pathways.Conclusion Hypoxic preconditioning alleviates radiation-induced bone marrow injury,and post-irradiation normoxia restoration promotes hematopoietic recovery in acute radiation-exposed mice.

Objective:To investigate the predictive value of serum hypersensitive C-reactive protein (hs-CRP) for stroke-associated pneumonia (SAP) in patients with acute ischemic stroke (AIS) who received intravenous thrombolysis.Methods:From May 2015 to April 2017, the clinical data of the patients with AIS treated with intravenous thrombolysis in Nanjing First Hospital were collected retrospectively. Multivariate logistic regression analysis was used to determine the independent risk factors for SAP in patients with AIS after intravenous thrombolysis. Receiver operating characteristic (ROC) curve and nomogram-based methods were used to analyze the predictive value of hs-CRP for SAP. Results:A total of 243 patients with AIS who received intravenous thrombolysis were included, and 63 (34.6%) of them had SAP. There were significant differences in age ( P=0.006), leukocyte count ( P=0.044), fasting blood glucose level ( P=0.003), serum hs-CRP level ( P=0.001), hs-CRP classification ( P=0.001) and dysphagia rate ( P=0.035) between the SAP group and non-SAP group. Multivariate logistic regression analysis showed that after adjusting for the confounding factors, taking the first quartile of serum hs-CRP level as a reference, the third quantile (odds ratio [ OR] 18.790, 95% confidence interval [ CI] 4.771-74.007; P=0.001) and the fourth quantile ( OR 54.054, 95% CI 12.248-324.088; P=0.001) of hs-CRP were the independent predictors of SAP. The area under the ROC curve of the baseline serum hs-CRP level for predicting SAP was 0.805 (95% CI 0.742-0.868; P<0.001). When the optimal cut-off value of hs-CRP was 5.54 mg/L, the sensitivity and specificity of predicting SAP were 76.11% and 76.19%, respectively. The analysis of nomogram also showed that hs-CRP was an independent predictor of SAP (consistency index 0.862, 95% CI 0.738-0.986; P<0.001). Conclusions:The increased serum hs-CRP was an independent predictor of SAP in patients with AIS receiving intravenous thrombolysis, and had a higher predictive value.

Objective To study the changes of immune function of endotoxin tolerant dendritic cell (ETDC) and to observe its effect on sepsis in mouse model.Methods ETDC were prepared by pretreated bone marrow dendritic cells derived from BALB/c mice with lipopolysaccharide stimulation.The cells were collected and the expressions of surface markers including major histocompatibility complex ( MHC)Ⅱ, CD86 and CD11c were detected by flow cytometry.The proliferation rate of T lymphocytes was evaluated by cell counting kit-8 and the concentrations of cytokines in the supernatant were detected by enzyme linked immuno sorbent assay.Afterwards, 36 mice were randomly assigned into 4 groups.The blank control group did not receive any treatment, the sham-operated group underwent simple incision suture, the sepsis group and ETDC reinfusion group underwent cecal ligation and puncture to establish sepsis.Before sepsis model establishment, 0.9% sodium chloride solution or suspension of ETDC and 0.9%sodium chloride were reinfused by tail vein.The serum levels of alanine aminotransferase (ALT) and aspartate transaminase (AST), tumor necrosis factor (TNF)-α, interleukin(IL)-6 and IL-10, and the proportion of help T cell ( Th) 17/regulatory T cell ( Treg) in spleen of each group were detected.The pathological manifestations of liver, kidney and ileum in each group were observed.T test and χ2 test were used for comparisons between groups.Results The results of flow cytometry showed that MHCⅡ, CD86 and CD11c of ETDC were 70.4%, 43.1%, and 73.1%, respectively, which were significantly lower than those of mature dendritic cell (mDC) (96.1%, 89.5%, and 84.6%, respectively) (χ2 ＝56.47, 83.78, and 23.29, respectively, all P＜0.01).The concentrations of IL-10, TNF-αand IL-6 in the supernatant of ETDC were (978.04 ±56.70), (980.34 ±111.96) and (12 743.03 ±865.81) ng/L, respectively, and those of mDC were (741.35 ±99.23), (1 703.11 ±117.00) and (19 052.28 ±1 145.84) ng/L, respectively.The differences were all statistically significant (t＝5.073, 10.93, and 10.76, respectively, all P＜0.01).The proliferation rates of T lymphocytes co-cultured with ETDC in 1∶5 and 1∶10 ratio group were (676.95 ±85.99)%and (514.00 ±106.39)%, respectively, which were lower than those of the mDC group (956.25 ±127.12)%and (772.07 ±214.08)%, respectively.The pathological injuries of liver, kidney and ileum in ETDC treatment group were significantly lighter than those in sepsis group.Serum ALT and AST levels in the ETDC reinfusion group were (299.71 ±36.91) and (690.39 ±154.92) U/L, respectively, and TNF-α, IL-6 and IL-10 were (0.067 ±0.005), (0.428 ±0.051) and (0.058 ±0.005) ng/L, respectively.Serum ALT and AST levels in the sepsis group were (620.67 ±69.27) and (1 430.17 ±134.05) U/L, respectively, and TNF-α, IL-6 and IL-10 were (0.085 ±0.007), (0.774 ±0.088) and (0.036 ±0.005) ng/L, respectively.The differences were all statistically significant (t＝11.60, 10.96, 5.991, 8.657, and 8.04, respectively, all P ＜0.01).The proportion of Treg/Th17 in the ETDC reinfusion group was 23.4%, and that in the sepsis group was 60.8%(χ2 ＝28.69, P＜0.01).Conclusion The reinfusion of ETDC has a protective effect on sepsis in mouse model, which may play a negative immune regulatory role by regulating the differentiation of T cells.

Objective To investigate the effect and safety of intravenous thrombolytic therapy in the endovascular treatment of acute anterior circulation vascular occlusive stroke.Methods The clinical data of 226 patients with acute anterior circulation vascular occlusive stroke who underwent endovascular treatment in Nanjing First Hospital,Nanjing Medical University from May 2015 to May 2018 were retrospectively collected.According to whether or not intravenous thrombolysis was performed,the patients were classified into simple thrombectomy group (n=112) and bridging treatment group (n=114).The modified Thrombolysis in Cerebral Infarction Score (mTICI) was used to evaluate the vascular opening effect,and the blood vessel recanalization time,mTICI,the symptomatic intracranial hemorrhage rate,and the modified Rankin Scale (mRS) score at 90 days after surgery were evaluated.Results There were no statistically significant differences in gender,age,past history and National Institute of Health Stroke Scale score between the two groups (P＞0.05).There was no statistically significant difference in door-to-recanalization time between the two groups (P＞0.05).Excluding the patients with post-wake stroke and unexplained onset time,the simple thrombectomy group (n=63) and the bridging treatment group (n=1 11) showed statistically significant differences in onset-to-door time ((235.04± 182.64) min vs (102.48±60.51) min,t=7.01,P＜0.01)and onset-to-recanalization time ((405.31 ± 148.89) min vs (337.31 ± 117.65) min,t=3.32,P=0.01).The difference in number of thrombectomy between the simple thrombolysis group (2.55± 1.52) and the bridging treatment group (2.11± 1.48) was statistically significant (t=2.246,P=0.026).The total reperfusion (mTICI 2b/3) rate was 89.8％ (203/226),88.4％ (99/112) in the simple thrombectomy group and 91.2％ (104/114) in the bridging treatment group,with no statistically significant difference between the two groups (P＞0.05).The differences in symptomatic intracranial hemorrhage rate (8.93％ (10/112) vs 11.4％ (13/114)),mortality rate (12.5％ (12/112) vs 16.7％ (19/114)) and 90-day good functional outcome (mRS score 0-2;54.5％ (61/112) vs 55.8％ (63/114)) between the two groups were not statistically significant (P＞0.05).Conclusions In patients with acute anterior circulation vascular occlusive stroke undergoing endovascular treatment,intravenous thrombolysis can reduce the number of thrombectomy,not increase the door-to-recanalization time,the risk of symptomatic intracranial hemorrhage and mortality,and has similar good functional outcome as the simple thrombeetomy group.Therefore,intravenous thrombolysis is safe and effective for endovascular treatment of acute anterior circulation large vessel occlusive stroke.

Objective To study the effect of age-related white matter changes (ARWMC) on first symptomatic ischemic stroke events in the oldsters. Methods For the prospective study, a total of 368 eligible oldsters were enrolled in the study from January 2010 to August 2012. The degrees of ARWMC were assessed by ARWMC scale;according to the scores, they were divided into non ARWMC group, mild-moderate ARWMC group and severe ARWMC group. The patients were followed up once every 3 months. The clinical endpoint events and time (first symptomatic ischemic stroke, myocardial infarction and all-cause death) were recorded. Analyses of variance and Chi-square test were used to compare the differences of clinical data among the 3 groups. COX regression was used to assess the risk differences of first symptomatic ischemic stroke in the oldsters of three groups. Results After an average of follow-up for 48.7 months, 50 participants (13.6％) had first symptomatic ischemic stroke;25 (25.8％) were categorized as the severe ARWMC group, 22 (10.9％) were as the mild-medium group, and 3 (4.4％) were as the non ARWMC group. Among the three groups, the differences in age, history of hypertension, systolic blood pressure, incidence of clinical endpoint events and first symptomatic ischemic stroke, and follow-up time of endpoint events were statistically significant (P<0.05); patients from the severe ARWMC group were the oldest, and had the longest history of hypertension, the highest systolic blood pressure, the highest incidence of clinical end events and first symptomatic ischemic stroke, and the shortest follow-up period for clinical end events. COX regression analysis showed that the risk of first symptomatic ischemic stroke in the severe ARWMC group was about 8 times higher than that in the non ARWMC group (hazard ratio=9.012, 95％CI: 2.310-35.154, P=0.002). Conclusion In oldsters, severe ARWMC often accompany hypertension history and poor blood pressure controll, and it is an independent and serious risk factor for long-term first symptomatic ischemic stroke.

Objective To research effect of esomeprazole,mosapride,hydrotalcid treatment of gastroesophageal reflux disease(GERD),and find effective treatment methods.Methods 90 cases diagnosed as GERD patients were divided into the three groups:group A(30 cases),B group(30 cases),C group(30 cases).A group was treated with esomeprazole 40mg qd,B group was treated with esomeprazole 40mg qd,mosapride 5mg tid,C group was treated with esomeprazole 40mg qd,mosapride 5mg tid,three scenarios Hydrotalcid 1.0g tid treatment.The treatment methods remained for 4 weeks and 8 weeks and symptom remission were evaluated.Results The clinical remission rates were 66.7％,83.3％ and 90.0％ after four weeks treatment in A group,B group,C group; the clinical remission rates were 73.3％,90.0％ and 96.7％ after eight weeks treatment in A group,B group,C group.The total effective rate of group A and group C in eight weeks were significantly difference (P ＜ 0.05),A group and B group,B group and C group showed no significant difference(P ＞ 0.05).Conclusion Esomeprazole combined with mosapride,Hydrotalcid treated with GERD can get better efficacy,which can improve symptom relief rates.