Combined deficiency of coagulation factor V (FV) and factor VIII (FVIII) is a rare bleeding disease caused by variants in either lectin mannose binding 1 (LMAN1) or multiple coagulation factor deficiency 2 (MCFD2) gene. Reducing the level of FVIII by inhibiting the LMAN1-MCFD2 complex may become a new anticoagulant approach. We aimed to find a new therapeutic option for anticoagulation by RNA interference (RNAi) targeting LMAN1 and MCFD2. siRNA sequences with cross-homology between mice and humans were designed based on LMAN1 or MCFD2 transcripts in NCBI and were screened with the Dual-Luciferase reporter assay. The optimal siRNAs were chemically modified and conjugated with three N-acetylgalactosamine molecules (GalNAc-siRNA), promoting their targeted delivery to the liver. The expression of LMAN1 and MCFD2 in cell lines or mice was examined by RT-qPCR and western blotting. For the mice administered with siRNA, we assessed their coagulation function by measuring APTT and the activity of FVIII factor. After administration, siRNAs GalNAc-LMAN1 and GalNAc-MCFD2 demonstrated effective and persistent LMAN1 and MCFD2 inhibition. 7 days after injection of 3mg/kg GalNAc-LMAN1, the LMAN1 mRNA levels reduced to 19.97

BACKGROUND:How concurrent TP53 mutations affect targeted therapy of advanced Epidermal Growth Factor Receptor (EGFR) mutant lung adenocarcinoma remains controversial, particularly the deep classification of TP53 mutations. METHODS:This study retrospectively analyzed the clinical data of advanced EGFR mutant lung adenocarcinoma patients treated with EGFR-tyrosine kinase inhibitors (TKIs) in the First Affiliated Hospital of Soochow University. The survival rates were compared using Log-rank tests. Potential prognostic factors were identified using multivariate Cox hazard regression models. RESULTS:Total 156 advanced lung adenocarcinoma patients treated with EGFR-TKIs were included in this study. Multivariate analysis showed that male [hazard rate (HR): 1.537, 95% confidence interval (CI): 1.055-2.240, P = 0.025], brain metastasis (HR: 1.707, 95%CI: 1.086-2.682, P = 0.020) and concurrent TP53 mutations (HR: 1.569, 95%CI: 1.051-2.341, P = 0.028) were independent negative predictors of progression-free survival (PFS). EGFR L858R mutations (HR: 2.475, 95%CI: 1.443-4.248, p = 0.001), smoking history (HR: 2.530, 95%CI: 1.352-4.733, P = 0.004) and concurrent TP53 mutations (HR: 2.326, 95%CI: 1.283-4.218, P = 0.005) were associated with worse survival. Further analysis revealed that mutations in TP53 exons 4, 5 and 8 (P<0.05), missense mutations (P = 0.006) and nondisruptive mutations (P<0.001) were associated with shorter PFS, whereas mutations in TP53 exons 5 and 7 (P<0.05), missense mutations and non-missense mutations (P = 0.006; P = 0.007), disruptive mutations and nondisruptive mutations (P = 0.013; P = 0.013) were all associated with poorer survival times. In addition, the PFS and overall survival (OS) of nondisruptive mutations in exon 7 were worse than those in other exons (P = 0.041; P<0.001). CONCLUSIONS:Concurrent TP53 mutations conferred worse EGFR-TKIs efficacy and prognosis in advanced EGFR mutant lung adenocarcinoma and the effects of different TP53 mutation types were heterogeneous.

Background: Forkhead box L2 (FOXL2) has been recognized as a transcription factor in the progression of many malignancies, but its role in non-small cell lung cancer (NSCLC) remains unclear. This research clarified on the role of FOXL2 and the specific molecular mechanism in NSCLC.Methods: RNA and protein levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting assays. Cell proliferation was examined by cell counting kit-8 (CCK-8) and clonogenic assays. Transwell and wound healing assays were used to detect cell invasion and migration. Cell cycle alterations were assessed by flow cytometry. The relationship between FOXL2 and miR-133b was verified by dual-luciferase reporter assays. In vivo metastasis was monitored in the tail vein-injected mice.Results: FOXL2 was upregulated in NSCLC cells and tissues. Downregulation of FOXL2 restrained cell proliferation, migration, and invasion and arrested the cell cycle of NSCLC cells. Moreover, FOXL2 promoted the epithelial-mesenchymal transition (EMT) process of NSCLC cells by inducing the transforming growth factor-beta (TGF-beta)/Smad signaling pathway. miR-133b directly targeted the 3 & PRIME;-UTR of FOXL2 and negatively regulated FOXL2 expression. Knockdown of FOXL2 blocked metastasis in vivo.Conclusions: miR-133b downregulates FOXL2 by targeting the 3 & PRIME;-UTR of FOXL2, thereby inhibiting cell proliferation, EMT and metastasis induced by the TGF-beta/Smad signaling pathway in NSCLC. FOXL2 may be a potential molecular target for treating NSCLC.

Triggering receptor expressed on myeloid cells (TREM) like transcript-1 (TLT-1) is a membrane protein receptor found in α-granules of megakaryocytes and platelets. Upon platelet activation TLT-1 is rapidly relocated to the surface of platelets. In plasma, a soluble form of TLT-1 (sTLT-1) is present. Plasma levels of sTLT-1 are significantly elevated in thrombotic diseases. In the present study, we investigated to whether TLT-1 reflects platelet activation in pregnant women with preeclampsia. We studied 30 preeclamptic patients who were matched with 30 normotensive pregnant women and 30 non-pregnant controls. Basal TLT-1, P-selectin, and CD63 expressions on platelets were analyzed with the use of flow-cytometry (FCM). Platelet reactivity was induced by thrombin receptor activation peptide and determined by FCM. Plasma concentrations of sTLT-1 and soluble P-selectin (sP-selectin) were measured by an enzyme-linked immunosorbent assay. Results show that basal platelet expression of TLT-1, P-selectin and CD63 were increased in women with preeclampsia (PE) compared with normotensive pregnant women (NP). Platelets from PE women and NP women were more responsive compared to from nonpregnant women controls (NC), and which was demonstrated by increased expression of TLT-1, P-selectin, and CD63 upon stimulation in vitro. Plasma concentration of sTLT-1 was greater in PE women compared to NP women and NC women. Plasma sP-selectin level was higher in pregnant women than in nonpregnant women, but there were no significant differences between PE and NP women. In summary, our results revealed that platelet activation is prominent in preeclampsia, TLT-1 reflects platelet activation and may be a useful indicator for preeclampsia.

目的 分析冬春季佩戴口罩对血液透析患者呼吸道感染的预防作用.方法 回顾性分析2017-2020年每年2～4月因肾衰竭行规律血液透析患者的临床资料;其中,2020年患者治疗时严格佩戴口罩,2017-2019年患者治疗时未佩戴口罩.分析佩戴口罩对呼吸道感染率的影响.结果 与2017、2018、2019年相比,2020年患者佩戴口罩后呼吸道感染率降低(0.16％vs.0.36％、0.36％、0.47％)(P＜0.01).2020年年龄＜60岁和血液透析时间＞2年患者佩戴口罩后呼吸道感染率均较2017、2018、2019年降低(0.12％vs.0.44％,0.42％、0.49％和0.18％vs.0.42％、0.43％、0.50％)(P＜0.01或P＜0.05);而2017-2020年间年龄≥60岁和血液透析时间≤2年患者的呼吸道感染率比较差异均无统计学意义(P＞0.05).结论 在呼吸道感染高发季节佩戴口罩可以有效降低血液透析患者呼吸道感染率,尤其是年龄＜60岁和血液透析时间＞2年的患者.

We previously demonstrated the immunostimulatory efficacy of Pseudomonas aeruginosa flagellar hook protein FlgE on epithelial cells, presumably via ectopic ATP synthases or subunits ATP5B on cell membranes. Here, by using recombinant wild-type FlgE, mutant FlgE (FlgEM; bearing mutations on two postulated critical epitopes B and F), and a FlgE analog in pull-down assay, Western blotting, flow cytometry, and ELISA, actual bindings of FlgE proteins or epitope B/F peptides with ATP5B were all confirmed. Upon treatment with FlgE proteins, human umbilical vein endothelial cells (HUVECs) and SV40-immortalized murine vascular endothelial cells manifested decreased proliferation, migration, tube formation, and surface ATP production and increased apoptosis. FlgE proteins increased the permeability of HUVEC monolayers to soluble large molecules like dextran as well as to neutrophils. Immunofluorescence showed that FlgE induced clustering and conjugation of F-actin in HUVECs. In Balb/c-nude mice bearing transplanted solid tumors, FlgE proteins induced a microvascular hyperpermeability in pinna, lungs, tumor mass, and abdominal cavity. All effects observed in FlgE proteins were partially or completely impaired in FlgEM proteins or blocked by pretreatment with anti-ATP5B antibodies. Upon coculture of bacteria with HUVECs, FlgE was detectable in the membrane and cytosol of HUVECs. It was concluded that FlgE posed a pathogenic ligand of ectopic ATP5B that, upon FlgE–ATP5B coupling on endothelial cells, modulated properties and increased permeability of endothelial layers both in vitro and in vivo. The FlgE-ectopic ATP5B duo might contribute to the pathogenesis of disorders associated with bacterial infection or ectopic ATP5B-positive cells.

血栓形成即血液的局部凝结,可发生在动脉或静脉循环中,它是缺血性心脏病、缺血性脑卒中和静脉血栓栓塞症( venous thromboembolism,VTE)的共同病理基础.缺血性心脏病和中风占全球死亡人数的四分之一[1] ,已成为人类死亡的首要病因.深静脉血栓和肺栓塞统称VTE,是继心梗和卒中后第3位的心血管相关性疾病的死亡原因[2].

Background: There have been many studies on the effectiveness and complications of airway stent, but few had focused on factors that affect survival after stent placement. This study intended to assess the factors associated with the survival in patients with malignant central airway obstruction (MCAO) after airway metallic stent placement. Methods: The clinical data of adult MCAO patients who underwent stent placement form February 2003 to June 2017 in the First Affiliated Hospital of Soochow University in China were retrospectively analyzed. The survival rates were compared using Log-rank tests. Potential prognostic factors were identified using multivariate Cox hazard regression models. Results: Total 102 MCAO patients were included in this study. The median survival time of these patients after airway metallic stent placement was 4.1 months. Multivariate analysis showed that MCAO patients receiving radiotherapy [hazard ratio (HR) 0.554; 95% confidence interval (CI): 0.308-0.999] or chemoradiotherapy (HR 0.251; 95% CI: 0.126-0.499) after stenting had better prognosis. However, ECOG PS >= 3 score prior to the stenting (HR 2.193; 95% CI: 1.364-3.526) and stents placed in both trachea and main bronchus (HR 2.458; 95% CI: 1.384-4.366) were associated with worse survival. Conclusions: In our results, survival of MCAO patients after airway metallic stenting was related to ECOG PS score prior to the stenting, the site of stent placement and we have hereby proposed for the first time that having opportunity to receive radiotherapy or chemoradiotherapy after stenting contribute to better prognosis.

目的 探讨血液透析中心应对新型冠状病毒疫情的院内感染防控策略优化及效果.方法 回顾性分析苏州大学附属第一医院血液透析中心通过预约、预检、宣教培训、佩戴口罩、对疑似感染患者单间专人透析、加强重点区域消毒等优化策略,应对新型冠状病毒疫情,识别、隔离潜在感染者,保护易感人群的院内感染防控策略的效果.结果 截止2020年2月15日的疫情期间,苏州大学附属第一医院共首诊新型冠状病毒肺炎(以下简称COVID-19)患者10例,其中急诊3例,发热门诊7例.共血液透析310例患者,其中急诊室内单间血透13例.通过优化院内感染防控策略,未发生一例COVID-19院内感染.结论 通过预检分流、口罩防护、对疑似感染患者单间专人透析、加强重点区域消毒等优化策略,可有效防止COVID-19在血液透析患者中的院内传播.

目的 探讨多肿瘤标志物蛋白芯片(C12芯片)连续3年筛查对肿瘤检出率的影响.方法 收集某高校40周岁及以上身体键康、无肿瘤病史教师2013-2015年连续3年的健康体检C12芯片检测结果 ,分析C12芯片连续筛查对肿瘤检出率的影响.按照年龄分为中年组(40～59岁)、老年组(60～79岁)和高龄组(≥80岁),比较不同性别和年龄组肿瘤标志物的阳性率和检测值的特点.结果共3634名教师完成了连续3年的C12芯片检测及相应的随访.汇总11017例的C12芯片检测结果分析表明,其中1027例有1项或多项标志物阳性,阳性率为9.32％.中年组、老年组和高龄组体检者肿瘤标志物总阳性率分别为4.45％、10.84％和26.97％,随年龄的增大而升高(P<0.01).再次检测后仍超过正常值2倍以上的受检者行进一步检查,肿瘤指标单次,连续2次和连续3次阳性的体检者,肿瘤检出率分别为1.46％,6.07％和7.08％,肿瘤检出率随肿瘤标志物持续阳性次数的增多而增大(P<0.01).结论 C12芯片检测的肿瘤标志物总阳性率和肿瘤检出率随年龄增大而增高,肿瘤检出率随肿瘤标志物持续阳性次数的增多而增大,C12芯片连续检测有助于提高肿瘤检出率.

Since its inception, anti-platelet antibodies have been an important tool for studying the interaction of platelets with blood components and blood vessels. At the same time, anti-platelet antibodies also play an important role in the detection and diagnosis of hemorrhagic and thrombotic diseases, and become a kind of powerful anti-thrombotic drugs. With the further understanding of the role of platelets in physiological hemostasis and pathological thrombosis, anti-platelet and anti-thrombotic antibodies also seek a balance between better anti-thrombotic effects and less bleeding side effects.

To develop an effective long-acting antidiabetic agent, we designed a novel Exendin-4 derivative (termed LEx4) containing an albumin-binding domain (ABD), a protease-cleavable linker and a native Exendin-4. Here, we present the LEx4 with balanced glucoregulatory activity and prolonged in vivo activity. As a first step, the LEx4 with purity more than 99% was prepared. Microscale thermophoresis (MST) results demonstrated that LEx4 associates with rat and monkey serum albumin with high-affinity (K-a = 1.26 x 10(6) M-1 and 1.52 x 10(6) M-1, respectively). Then the stability test in vitro showed the enhanced antiproteolytic ability of LEx4 in rat and human plasma compared to native Exendin-4. Oral glucose tolerance test (OGTT) in type 2 diabetic mice showed the glucose-lowering efficacy of LEx4 was clearly dosage-dependent within 25-250 nmol/kg. In addition, the protracted antidiabetic effects of LEx4 were further confirmed by both multiple OGITs and hypoglycemic efficacies test in type 2 diabetic mice. In Sprague Dawley (SD) rats, LEx4 also showed 3.3-fold longer elimination half-life (t(1/2)) than native Exendin-4. Furthermore, once daily injection of LEx4 to db/db mice achieved long-term beneficial effects on body weight, blood biochemical values, glucose tolerance and pancreatic tissue. We believe LEx4 has superior pharmaceutical potential as a therapeutic drug to against type-2 diabetes mellitus (T2DM) based on these results. This strategy of albumin binding is also applicable to other bioactive peptides for development of long-acting therapeutic drugs. (C) 2018 Elsevier Masson SAS. All rights reserved.

More than 90% of thyroid cancer belongs to the papillary and follicular thyroid carcinomas based on pathological subtypes. Papillary and follicular thyroid carcinoma are generally associated with a good prognosis. In contrast, other pathological subtypes such as poorly-differentiated and anaplastic thyroid carcinoma (PDTC and ATC) have a poor clinical outcome with a short life expectancy. To identify the genetic variations and biomarkers that may potentially distinguish the aggressive form of thyroid cancer, we performed a retrospective analysis of the formalin-fixed paraffin-embedded tumor samples from 50 patients who mainly displayed aggressive thyroid cancer using next-generation sequencing of 416 solid tumor-related genes. We adopted extensive bioinformatic analysis to vigorously remove germline single-nucleotide polymorphism and systematic sequencing errors, and report here that mutation in DNMT3A gene was significantly enriched in patients with PDTC or ATC.

SZ-123, a murine monoclonal antibody that targets the human von Willebrand factor (VWF) A3 domain and blocks the binding of collagen, is a powerful antithrombotic. In a Rhesus monkey model of thrombosis, SZ-123 had no side effects, such as bleeding or thrombocytopenia.

Aspirin (ASA) is an anti-inflammatory and anti-platelet agent that irreversibly inhibits the cyclooxygenase (COX) and, thus thromboxane (TX) generation. Insufficient response on antiplatelet medication has become an intensively discussed issue because of the risk factor of recurrent adverse cardiovascular events. However, the monitoring of antiplatelet therapy requires appropriate, robust and reliable test methods. For the measurement of thienopyridine effects, the manufacturer of the PFA-200 System provides the INNOVANCE PFA P2Y cartridge.

The aim of the study was to investigate ITGA2B and ITGB3 genetic polymorphisms and to evaluate the variability in the platelet function in healthy Chinese subjects. The genetic sequence of the entire coding region of the ITGA2B and ITGB3 genes was investigated. Adenosine diphosphate-induced platelet aggregation, glycoprotein IIb/IIIa content, bleeding time, and coagulation indexes were detected. Thirteen variants in the ITGA2B locus and 29 variants in the ITGB3 locus were identified in the Chinese population. The rs1009312 and rs2015049 were associated with the mean platelet volume. The rs70940817 was significantly correlated with the prothrombin time. The rs70940817 and rs112188890 were related with the activated partial thromboplastin time, and ITGB3 rs4642 was correlated with the thrombin time and fibrinogen. The minor alleles of rs56197296 and rs5919 were associated with decreased ADP-induced platelet aggregation, and rs55827077 was related with decreased GPIIb/IIIa per platelet. The rs1009312, rs2015049, rs3760364, rs567581451, rs7208170, and rs117052258 were related with bleeding time. Further studies are needed to explore the clinical importance of ITGA2B and ITGB3 SNPs in the platelet function.

目的:探讨多肿瘤标志物蛋白芯片技术(C12芯片法)在老年人健康体检中的应用,并量化分析不同年龄和性别体检者肿瘤标志物检测值和阳性率的差别,阐明其临床意义.方法:选择健康体检者9654人,按年龄分为对照组(40～59岁)、老年组(60～79岁)和高龄组(80岁及以上).采用多肿瘤标志物芯片检测系统检测各组体检者甲胎蛋白(AFP)、糖链抗原15-3 (CA15-3)、糖链抗原19-9 (CA19-9)、糖链抗原125(CA125)、糖链抗原242 (CA242)、癌胚抗原(CEA)、神经元特异性烯醇化酶(NSE)、铁蛋白(ferritin)、β-人绒毛膜促性腺激素(β-HCG)、人生长激素(HGH)、前列腺特异性抗原(PSA)和游离PSA (f-PSA)共12项肿瘤标志物水平,以超出正常范围为阳性结果,分析3组体检者样本的肿瘤标志物检测值和阳性率.结果:①体检者9 654人中共检出阳性结果665人,阳性检出率为6.89％,其中32人经内窥镜、病理学和影像学检查确诊为早期或中期肿瘤,肿瘤检出率为0.33％.②男女体检者肿瘤标志物阳性率不同,男性前5位依次为f-PSA、PSA、CEA、CA19-9和NSE;女性前5位依次为CA19-9、CA125、CEA、NSE和CA242.③CA19-9、CA242、CEA、NSE、PSA和f-PSA阳性率分布呈随年龄增长而升高的趋势(P＜0.01).④各组体检者AFP、CA125、CA19-9、CA242、CEA、HGH、NSE和PSA检测值随着年龄增长而升高,高龄体检者上述指标高于老年组和对照组(P＜0.01),老年组上述指标高于对照组(P＜0.01).结论:C12芯片中CA19-9、CEA、NSE、PSA和f-PSA等肿瘤标志物在老年人、尤其是高龄老年人的健康筛查中阳性率高,有助于发现早期肿瘤.

Although intradiscal cement leakage is a common complication of spine augmentation, the effects of cement leakage into the intervertebral disc (IVD) have not been well investigated. This study aimed to determine the effects of cement leakage on IVD degeneration in rabbits. Poly(methyl methacrylate) (PMMA) particles were injected into lumbar discs of rabbits using 26 G needles. Tissue effects were assessed using disc height, sagittal T2-weighted images, histology and immunohistochemistry. The results showed that stimulation with PMMA particles significantly reduced disc height compared with that in the sham-operation group at 3 weeks after injection. The mean signal intensity of the operated discs showed little to no changes among all groups at 3 weeks post-operation. After 6 weeks, the signal intensity of the PMMA-injected group decreased by 22% compared with that in the sham-operation group. Histological and quantitative immunohistochemical examination indicated phenotypic tissue changes from cartilaginous tissue into fibrotic tissue, with apparent degeneration in the PMMA group. Additionally, more collagen type II-containing tissues, but fewer matrix metalloproteinase-7-positive cells or apoptotic cells, were detected in the sham-operation group. The PMMA particle-induced degeneration rate was slower than that of the degeneration group, whereas the histologic data showed no difference in the progression of degeneration between the two groups. These data suggest that PMMA particles can moderately accelerate disc degeneration compared with the 18 G needle puncture model. In conclusion, intradiscal injection of PMMA particles induced significant IVD degeneration in vivo. Therefore, further study of the adverse effects of PMMA leakage on IVD degeneration is required.

The interactions between collagen, von Willebrand factor (VWF), and glycoprotein Ib (GPIb) are crucial for hemostasis and thrombosis. This axis represents a promising target for the development of new antithrombotic agents. In this study, we investigate the in vivo antithrombotic efficacy of an anti-VWF monoclonal antibody SZ-123 and its potential underlying mechanisms. Cyclic flow reductions (CFRs), an indicator of arterial thrombosis, were measured in the femoral artery of anesthetized Rhesus monkeys before and after intravenous administration of SZ-123. Ex vivo VWF binding to collagen, platelet agglutination, platelet count, and template bleeding time were used as measurements of antithrombotic activity. In addition, plasma VWF and SZ-123 levels, and VWF occupancy were measured by ELISA. Administration of 0.1, 0.3, and 0.6 mg/kg SZ-123 resulted in 45.3%, 78.2%, and 100% reductions in CFRs, respectively. When 0.3 and 0.6 mg/kg SZ-123 were administered, 100% of VWF was occupied by the antibody. Moreover, 100% ex vivo inhibition of VWF-collagen binding and 60-95% inhibition of platelet agglutination were observed from 15 min to 1 h. None of the doses resulted in significant prolongation of bleeding time. In vitro experiments revealed that SZ-123 not only blocks the collagen-VWF A3 interaction but also indirectly inhibits VWF A1 binding to GPIb alpha induced by ristocetin. Thus, we demonstrate that SZ-123 prevents in vivo arterial thrombus formation under high shear conditions by inhibiting VWF A3-collagen and VWF A1-platelet interactions and does not significantly prolong bleeding time. (c) 2013 Elsevier Inc. All rights reserved.