MMP9、FeNO以及血清IgE与儿童哮喘急性发作严重程度的相关性分析

目的研究呼出气一氧化氮(FeNO)、血清免疫球蛋白E(IgE)和基质金属蛋白酶9(MMP9)的水平与儿童哮喘急性发作之间的关系,为儿童哮喘的预防及治疗提供依据。方法选取沈阳市妇婴医院于2020年11月至2022年11月收治的98例支气管哮喘急性发作期儿童作为急性组,按照病情程度分成轻度组(n=32)、中度组(n=38)和重度组(n=28),按照2∶1的比例选出49例同期在门诊治疗的处于支气管哮喘缓解期的儿童作为缓解组,随机选取健康体检儿童49例作为健康对照组,分别对他们进行FeNO、MMP9和血清IgE及肺功能[用力肺活量(FVC)、1秒用力呼气量(FEV_(1))、FEV_(1)/FVC%、最大呼气流量(PEF)]检测。应用Pearson相关分析探讨哮喘急性发作期FeNO、MMP9及血清IgE和肺功能之间的联系,并对三者在支气管哮喘急性发作中的预测价值进行分析。结果急性组、缓解组和对照组的年龄、性别、体重指数和病程的比较,差异无统计学意义(P>0.05)。急性组FeNO、MMP9、血清IgE分别为(59.95±12.65)ppb、(4.87±1.44)pg/ml、(330.63±74.88)IU/ml,缓解组分别为(25.23±8.23)ppb、(1.21±0.02)pg/ml、(152.23±32.12)IU/ml,均高于对照组的(12.43±4.09)ppb、(0.53±0.24)pg/ml、(126.34±57.33)IU/ml,差异具有统计学意义(P<0.05)。急性期和缓解期FVC、FEV1、FEV1/FVC%、PEF均低于对照组,差异具有统计学意义(P<0.05)。支气管哮喘急性发作中度组FeNO、MMP9、血清IgE水平分别为(49.23±6.23)ppb、(1.21±0.02)pg/ml、(282.61±59.83)IU/ml,重度组分别为(67.43±10.09)ppb、(0.53±0.24)pg/ml、(356.49±70.82)IU/ml,均高于轻度组的(34.62±10.65)ppb、(4.87±1.44)pg/ml,(189.21±14.33)IU/ml,差异具有统计学意义(P<0.05)。在轻度组中FeNO、MMP9和血清IgE水平均较低,而在中度组中这些指标均较高,其中FVC、FEV_(1)、FEV_(1)/FVC%和PEF均较低,差异具有统计学意义(P<0.05)。FeNO以及MMP9与血清IgE水平呈正相关(P<0.05),FeNO、MMP9以及血清IgE水平与FVC、FEV_(1)、FEV_(1)/FVC%、PEF均呈负相关(P<0.05)。MMP9在支气管哮喘的诊断中表现出了显著的优势,当达到最大约登指数时,对应的截断值为1.17,曲线下面积(Area under curve,AUC)为0.83,敏感度和特异性也分别达到了90.13%和86.5%。结论支气管哮喘急性发作的儿童血清中的FeNO、MMP9以及血清IgE水平显著增高,随肺部功能恶化程度加重而上升,可能与支气管哮喘急性发作患儿肺功能损害程度有关。

Knock-out of GhPDCT with the CRISPR/Cas9 system increases the oleic acid content in cottonseed oil

Cotton is a pivotal economic crop for natural textile fibers that also serves as an important source of edible oil(Long et al.2023).Cottonseed oil contains approximately14%oleic acid and 59%linoleic acid.An increase in monounsaturated fatty acids,particularly oleic acid,enhances the oxidative stability and nutritional value of edible oil(Chen et al.2021).

基于CRISPR/Cas9技术建立Lepr与eNos双基因敲除的糖尿病肾病小鼠模型

目的基于CRISPR/Cas9基因编辑技术建立瘦素受体基因(leptin receptor,Lepr)与血管内皮一氧化氮合酶基因(endothelial nitric oxide synthase,eNos)双基因敲除(double-knockout,DKO)小鼠模型,构建晚期糖尿病肾病小鼠模型。方法根据eNos基因制备对应的gRNA,将CRISPR-Cas9体系显微注射于C57BL/Ks(BKS)背景小鼠的受精卵内。将受精卵转移至有假孕状态雌性小鼠的输卵管内部。幼鼠出生后经聚合酶链反应(polymerase chain reaction,PCR)鉴定及测序分析分选出为eNos^(+/-)基因型的F0代阳性小鼠,获得BKS背景下的Lepr基因杂合小鼠,即基因型为Lepr^(db/m)的Lepr-F0代杂合子小鼠。将eNos-F0与Lepr-F0代小鼠杂交,获得eNos^(+/-)/Lepr^(db/m)双杂合F1代小鼠,将双杂合F1代小鼠进一步交配,筛选得到Lepr与eNos双基因敲除小鼠。采用PCR法鉴定小鼠基因型,按基因鉴定结果分为野生型(wild-type,WT)组与DKO组小鼠。监测各组小鼠体质量、血糖与饮水进食量;采用酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)检测小鼠尿白蛋白与尿肌酐水平,并计算尿白蛋白排泄率;苏木精-伊红(hematoxylin-eosin,HE)与过碘酸六胺银(periodic acid-silver metheramine,PASM)染色检查各组小鼠肾组织病理改变。结果PCR检测结果显示,成功构建lepr^(db/db)/eNos^(-/-)DKO小鼠。与同窝对照组相比,DKO小鼠的体质量、血糖水平与饮水进食量均显著高于同窝对照小鼠。DKO小鼠的尿白蛋白与尿白蛋白排泄率显著高于WT小鼠。病理学结果显示,DKO小鼠的肾小球体积明显增大,系膜基质增生明显。结论基于CRISPR/Cas9基因编辑技术可成功构建lepr^(db/db)/eNos^(-/-)DKO小鼠,DKO小鼠可反映糖尿病肾病的典型表现,为深入研究糖尿病肾病的作用机制提供动物模型。

A review of the literature on the use of CRISPR/Cas9 gene therapy to treat hepatocellular carcinoma

Noncoding RNAs instruct the Cas9 nuclease to site speifillyl cleave DNA in the CRISPR/Cas9 system.Despite the high incidence of hepatocellular carcinoma(HCC),the patient's outcome is poor.As a result of the emergence of therapeutic resistance in HCC patients,dlinicians have faced difficulties in treating such tumor.In addition,CRISPR/Cas9 screens were used to identify genes that improve the dlinical response of HCC patients.It is the objective of this article to summarize the current understanding of the use of the CRISPR/Cas9 system for the treatment of cancer,with a particular emphasis on HCC as part of the current state of knowledge.Thus,in order to locate recent developments in oncology research,we examined both the Scopus database and the PubMed database.The ability to selectively interfere with gene expression in combinatorial CRISPR/Cas9 screening can lead to the discovery of new effective HCC treatment regimens by combining clinically approved drugs.Drug resistance can be overcome with the help of the CRISPR/Cas9 system.HCC signature genes and resistance to treatment have been uncovered by genome-scale CRISPR activation screening although this method is not without limitations.It has been extensively examined whether CRISPR can be used as a tool for disease research and gene therapy.CRISPR and its applications to tumor research,particularly in HCC,are examined in this study through a review of the literature.

Generation of double knockout cattle via CRISPR-Cas9 ribonucleoprotein(RNP)electroporation

Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestational periods,single pregnancy,and high raising cost.Furthermore,technically demanding methods such as microinjection and somatic cell nuclear transfer(SCNT)are needed for gene editing in cattle.In this point of view,electroporation in embryos has been risen as an alternative.Results First,editing efficiency of our electroporation methods were tested for embryos.Presence of mutation on embryo was confirmed by T7E1 assay.With first combination,mutation rates for MSTN and PRNP were 57.6%±13.7%and 54.6%±13.5%,respectively.In case of MSTN/BLG,mutation rates were 83.9%±23.6%for MSTN,84.5%±18.0%for BLG.Afterwards,the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing.Thirteen recipients were transferred for MSTN/PRNP,4 calves were delivered,and one calf underwent an induction for double KO.Ten surrogates were given double-KO embryos for MSTN/BLG,and four of the six calves that were born had mutations in both genes.Conclusions These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied.Finally,MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.

On the Impairment of Stress-Induced Changes in Triglyceride Levels via a Sub-Toxic Dose of Unmethylated Cytidine Phosphate Guanosine Oligodinucleotide (a Toll-Like Receptor 9 Ligand)

Changes in lipid metabolism have been implicated in protection against infectious diseases. In the first experiment of this study, we measured clinical lipid parameters in a murine model where the unmethylated cytidine phosphate guanosine (CpG) oligodinucleotide (ODN1826), a Toll-like receptor 9 (TLR9) agonist was administered in combination with D-galactosamine (GalN) that caused relatively liver-specific inflammation and toxicity. In the control mice group injected with phosphate-buffered saline (PBS) (acute psychological stress model associated with blood sampling), the serum triglyceride (TG) levels showed a rapid decrease followed by a rebound at 24 h as we have recently reported. However, such a TG rebound was impaired in the CpG/GalN- and solely CpG-treated groups of mice despite an absence of liver injury based on serum alanine aminotransferase levels in the latter group. Thus, the stress-associated serum TG rebound was abrogated by the injection of a sub-hepatotoxic CpG dose. In the second experiment, we simply measured the hepatic CD36 and SACRB1 (the gene for scavenger receptor B1 (SR-B1)) transcripts after the i.p. administration of PBS, CpG or CpG/GalN. There was a remarkable elevation of hepatic CD36 transcript expression in both the CpG- and CpG/GalN-treated mice at 8 h post-CpG injection whereas the increase in the PBS-treated mice was slower than the former two groups, suggesting that hepatic CD36 transcript expression is more pronounced in the combined stress models than under psychological stress alone. The individual mice data showed that the increase in CD36 expression was accompanied by a reduction in SCARB1 mRNA, showing reciprocal regulation between these two genes. Together with our previously reported findings, these data suggest that, in a murine model combining psychological stress with TLR-triggered hepatic inflammation, the psychological stress facilitates liver uptake of plasma TG (and its components fatty acids), but the subsequent re-esterification and/or release of TG-rich lipoproteins from the liver is impaired due to the concomitant TLR-signaling. We hypothesize that lipid metabolism during acute stress shifts toward an elevated hepatic uptake of lipids due to concomitant TLR signaling, facilitating the clearance of bacterial lipids by the liver.

Inactivated H9N2 vaccines developed with early strains do not protect against recent H9N2 viruses:Call for a change in H9N2 control policy

H9N2 virus has been widely distributed in wild birds and poultry around the world since its first emergence in the United States of America in 1966(Gu et al.2017;Carnaccini and Perez 2020).The virus appeared in chickens in China in the early 1990s,and over the last two decades has gradually become the dominant epidemic subtype(Sun and Liu 2015;Bi et al.2020).Although H9N2 virus infection alone cannot cause severe disease or death in poultry,H9N2 virus-infected birds experience a degree of egg production drop and can be easily infected by other pathogens,thus causing economic losses for poultry industry.

A simple and efficient CRISPR/Cas9 system permits ultra-multiplex genome editing in plants

The development and maturation of the CRISPR/Cas genome editing system provides a valuable tool for plant functional genomics and genetic improvement.Currently available genome-editing tools have a limited number of targets,restricting their application in genetic research.In this study,we developed a novel CRISPR/Cas9 plant ultra-multiplex genome editing system consisting of two template vectors,eight donor vectors,four destination vectors,and one primer-design software package.By combining the advantages of Golden Gate cloning to assemble multiple repetitive fragments and Gateway recombination to assemble large fragments and by changing the structure of the amplicons used to assemble sg RNA expression cassettes,the plant ultra-multiplex genome editing system can assemble a single binary vector targeting more than 40 genomic loci.A rice knockout vector containing 49 sg RNA expression cassettes was assembled and a high co-editing efficiency was observed.This plant ultra-multiplex genome editing system advances synthetic biology and plant genetic engineering.

CRISPR/Cas9-mediated knockout of E4 gene promotes maturation in soybean

Soybean is a broadly popular and extensively cultivated crop,however,many high-yield and high-quality varieties require specific growth conditions,restricting their widespread adoption.The appropriate light conditions and photoperiod must be attained for these varieties to thrive in new environments.In this study,we employed CRISPR/Cas9 to design two sgRNAs aimed at knocking out the maturity-related gene E4 in a major American soybean variety called''Jack'',which belongs to maturity group MGII.E4 gene is primarily involved in the photoperiodic flowering and maturity in soybean,making it an ideal candidate for genetic manipulation.We successfully obtained 1 homozygous E4-SG1 mutant type with 1-bp insertion,and 4 homozygous E4-SG2 mutants type with 2-bp deletion,7-bp deletion,61-bp deletion,and 1-bp insertion,respectively.The homozygous e4 mutant plants contained early termination codons devoid of transgenic elements.Additionally,no potential offtarget sites of the E4 gene were detected.A comparative analysis revealed that,unlike the wild-type,the maturity time of homozygous e4 mutants was early under both short-day and long-day conditions.These mutants offer novel germplasm resources that may be used to modify the photoperiod sensitivity and maturity of soybean,enhancing its adaptability to high-latitude regions.

Knock-in of exogenous sequences based on CRISPR/Cas9 targeting autosomal genes and sex chromosomes in the diamondback moth,Plutella xylostella

Genetic pest control strategies based on precise sex separation and only releasing sterile males can be accomplished by site-specific genome editing.In the current study,we showed that the mutation of single-allele Pxfl(2)d can significantly impair the normal mating behavior and testis development in male adults of the notorious cruciferous insect pest Plutella xylostella,in addition to its known functions in the ovarian development in female adults and egg hatching.Subsequent CRISPR/Cas9-based knock-in experiments revealed that site-specific integration of an exogenous green fluorescent protein(GFP)gene into autosomal Pxfl(2)d for labelling mutants could be achieved.However,this gene is not a suitable target for GFP insertion to establish a genetically stable knock-in strain because of the severe decline in reproductive capacity.We further screened for the W-chromosome-linked and Z-chromosome-linked regions to test the knock-in efficiency mediated by CRISPR/Cas9.The results verified that both types of chromosomes can be targeted for the site-specific insertion of exogenous sequences.We ultimately obtained a homozygous knock-in strain with the integration of both Cas9 and cyan fluorescent protein(CFP)expression cassettes on a Z-linked region in P.xylostella,which can also be used for early sex detection.By injecting the sgRNA targeting Pxfl(2)d alone into the eggs laid by female adults of the Z-Cas9-CFP strain,the gene editing efficiency reached 29.73%,confirming the success of expressing a functional Cas9 gene.Taken together,we demonstrated the feasibility of the knock-in of an exogenous gene to different genomic regions in P.xylostella,while the establishment of a heritable strain required the positioning of appropriate sites.This study provides an important working basis and technical support for further developing genetic strategies for insect pest control.

Combining prognostic value of serum carbohydrate antigen 19-9 and tumor size reduction ratio in pancreatic ductal adenocarcinoma

BACKGROUND Pancreatic ductal adenocarcinoma(PDAC)is a common cancer with increasing morbidity and mortality due to changes of social environment.AIM To evaluate the significance of serum carbohydrate antigen 19-9(CA19-9)and tumor size changes pre-and post-neoadjuvant therapy(NAT).METHODS This retrospective study was conducted at the Chongqing Key Laboratory of Translational Research for Cancer Metastasis and Individualized Treatment,Chongqing University Cancer Hospital.This study specifically assessed CA19-9 levels and tumor size before and after NAT.RESULTS A total of 156 patients who completed NAT and subsequently underwent tumor resection were included in this study.The average age was 65.4±10.6 years and 72(46.2%)patients were female.Before survival analysis,we defined the post-NAT serum CA19-9 level/pre-NAT serum CA19-9 level as the CA19-9 ratio(CR).The patients were divided into three groups:CR<0.5,CR>0.5 and<1 and CR>1.With regard to tumor size measured by both computed tomography and magnetic resonance imaging,we defined the post-NAT tumor size/pre-NAT tumor size as the tumor size ratio(TR).The patients were then divided into three groups:TR<0.5,TR>0.5 and<1 and TR>1.Based on these groups divided according to CR and TR,we performed both overall survival(OS)and disease-free survival(DFS)analyses.Log-rank tests showed that both OS and DFS were significantly different among the groups according to CR and TR(P<0.05).CR and TR after NAT were associated with increased odds of achieving a complete or near-complete pathologic response.Moreover,CR(hazard ratio:1.721,95%CI:1.373-3.762;P=0.006),and TR(hazard ratio:1.435,95%CI:1.275-4.363;P=0.014)were identified as independent factors associated with OS.CONCLUSION This study demonstrated that post-NAT serum CA19-9 level/pre-NAT serum CA19-9 level and post-NAT tumor size/pre-NAT tumor size were independent factors associated with OS in patients with PDAC who received NAT and subsequent surgical resection.

Two-dimensional layered In_(2)P_(3)S_(9): A novel superior anode material for sodium-ion batteries

Developing reliable and efficient anode materials is essential for the successfully practical application of sodium-ion batteries.Herein,employing a straightforward and rapid chemical vapor deposition technique,two-dimensional layered ternary indium phosphorus sulfide(In_(2)P_(3)S_(9)) nanosheets are prepared.The layered structure and ternary composition of the In_(2)P_(3)S_(9) electrode result in impressive electrochemical performance,including a high reversible capacity of 704 mA h g^(-1) at 0.1 A g^(-1),an outstanding rate capability with 425 mA h g^(-1) at 5 A g^(-1),and an exceptional cycling stability with a capacity retention of88% after 350 cycles at 1 A g^(-1).Furthermore,sodium-ion full cell also affords a high capacity of 308 and114 mA h g^(-1) at 0.1 and 5 A g^(-1).Ex-situ X-ray diffraction and ex-situ high-resolution transmission electron microscopy tests are conducted to investigate the underlying Na-storage mechanism of In_(2)P_(3)S_(9).The results reveal that during the first cycle,the P-S bond is broken to form the elemental P and In_(2)S_(3),collectively contributing to a remarkably high reversible specific capacity.The excellent electrochemical energy storage results corroborate the practical application potential of In_(2)P_(3)S_(9) for sodium-ion batteries.

MAP3K9通过上调Notch-1促进胰腺癌细胞的增殖与迁移

目的:探讨MAP3K9通过调控Notch-1促进胰腺癌细胞增殖、迁移的机制。方法:在胰腺癌细胞中分别转染MAP3K9过表达质粒和siRNA,RT-qPCR检测MAP3K9 mRNA表达水平,MTT实验检测转染后细胞增殖能力的改变,流式细胞术检测细胞凋亡能力的变化,划痕试验检测细胞迁移能力的变化,Transwell实验检测细胞的迁移与侵袭能力,Western Blot检测MAP3K9、Notch-1及EMT相关蛋白的表达情况。结果:RT-qPCR和Western Blot显示,PaTu-8988和PANC-1细胞的MAP3K9表达水平低于BXPC-3细胞;RT-qPCR结果表明,转染MAP3K9过表达质粒可以显著提高PaTu-8988和PANC-1细胞MAP3K9 mRNA表达水平,而转染siRNA MAP3K9可明显下调BXPC-3细胞MAP3K9 mRNA表达水平;上调胰腺癌细胞MAP3K9表达,可提高胰腺癌细胞增殖、迁移与侵袭能力,并抑制细胞凋亡;在胰腺癌细胞中过表达MAP3K9,可上调Notch-1、Vimentin、N-cadherin以及slug蛋白表达水平,并降低E-cadherin表达水平。结论:MAP3K9能够激活Notch-1,进而促进胰腺癌细胞的上皮—间充质转化(EMT)过程,增强胰腺癌细胞增殖及迁移能力。

The combined detection of carcinoembryonic antigen,carcinogenic antigen 125,and carcinogenic antigen 19-9 in colorectal cancer patients

BACKGROUND Hepatic metastases are common and difficult to treat after colorectal cancer(CRC)surgery.The predictive value of carcinoembryonic antigen(CEA),cancer antigen(CA)125 and CA19-9 combined tests for liver metastasis is unclear.AIM To evaluate predictive value of combined tests for CEA,CA125,and CA19-9 levels in patients with liver metastases of CRC.METHODS The retrospective study included patients with CRC alone(50 cases)and patients with CRC combined with liver metastases(50 cases)who were hospitalized between January 2021 and January 2023.Serum CEA,CA125 and CA19-9 levels were compared between the two groups,and binary logistic regression was used to analyze the predictive value of the combination of these tumor markers in liver metastasis.In addition,we performed receiver operating characteristic(ROC)curve analysis to assess its diagnostic accuracy.RESULTS The results showed that the serum CEA,CA125 and CA19-9 levels in the CRC with liver metastasis group were significantly higher than those in the CRC alone group.Specifically,the average serum CEA level in the CRC with liver metastasis group was 162.03±810.01 ng/mL,while that in the CRC alone group was 5.71±9.76 ng/mL;the average serum CA125 levels were 43.47±83.52 U/mL respectively.and 13.5±19.68 U/mL;the average serum CA19-9 levels were 184.46±473.13 U/mL and 26.55±43.96 U/mL respectively.In addition,binary logistic regression analysis showed that CA125 was significant in predicting CRC liver metastasis(P<0.05).ROC curve analysis results showed that the areas under the ROC curves of CEA,CA125 and CA19-9 were 0.607,0.692 and 0.586.CONCLUSION These results suggest that combined detection of these tumor markers may help early detection and intervention of CRC liver metastasis,thereby improving patient prognosis.

Serum tumor markers (carcinoembryonic antigen, carbohydrate antigen 19-9, carbohydrate antigen 72-4, carbohydrate antigen 24-2, ferritin) and gastric cancer prognosis correlation

BACKGROUND Gastric cancer is a kind of malignant tumor which is prevalent all over the world.Although some progress has been made in the treatment of gastric cancer,its prognosis is still not optimistic,so it is of great significance to find reliable prog-nostic indicators to guide the treatment and management of patients with gastric cancer.AIM To explore the relationship between serum levels of five biomarkers[carcinoem-bryonic antigen(CEA),carbohydrate antigen(CA)19-9,CA72-4,CA24-2,and ferritin]and prognosis in patients with gastric cancer.METHODS This study included 200 patients with gastric adenocarcinoma,and conducted an in-depth analysis of their baseline characteristics,relationship between tumor markers and staging,and prognosis.The study found that CA19-9 has a signi-ficant correlation with tumor stage,the average levels of CA24-2,CEA,CA72-4 and ferritin were slightly increased disregarding the stage of tumor.Survival analysis showed that increases in CEA,CA19-9,CA24-2,and ferritin were all associated with shortened overall survival of patients.Further multivariate ana-lysis revealed that elevated serum CA72-4 levels were an inde-pendent adverse prognostic factor.RESULTS This study reveals that there is a significant correlation between the expression levels of serum tumor markers CEA,CA19-9,CA72-4,CA24-2 and ferritin in patients with gastric cancer and prognosis,and can be used as important indicators for prognostic evaluation of gastric cancer.In particular,markers that appear abnormally elevated initially may help identify gastric cancer patients with poor prognosis.CONCLUSION Serum CEA and CA19-9 play an important role in the prognosis assessment of gastric cancer,and are effective tools to guide clinical practice and optimize individualized treatment strategies for gastric cancer patients.

猎鹰9号火箭发射及箭体复用的分析

猎鹰9号火箭截至2023年年底已完成286次发射。对猎鹰9号火箭特别是Block5版本火箭的发射情况进行了完整梳理统计,重点分析了一级箭体的复用情况以及发射、回收的相关能力保障,对火箭的经济性和未来前景作了预测。

BMP9调控Notch、TLR4信号通路诱导主动脉瓣间质细胞成骨样分化

目的:探究骨形态发生蛋白9(BMP9)调控Notch、Toll样受体-4(TLR4)信号通路诱导主动脉瓣膜间质细胞(AVICs)成骨样分化的作用机制。方法:将患者分为非钙化性主动脉瓣疾病(non-CAVD)组和CAVD组,免疫组化和Western blot检测患者BMP9、Runx2蛋白表达水平。取健康家猪主动脉瓣叶并分离AVICs,免疫荧光染色对其进行表型鉴定;采用BMP9干扰载体及BMP9腺病毒处理AVICs,分为对照组(control组)、干扰对照组(si-control)组、si-BMP9组、携带绿色荧光蛋白(GFP)基因的重组腺病毒(Ad)组(Ad-GFP组)和Ad-BMP9组,并建立体外AVICs钙化模型;采用碱性磷酸酶(ALP)染色、雄激素受体(AR)染色检测细胞早期和晚期的成骨分化能力;qRT-PCR检测Runx2、骨桥蛋白(OPN)、骨钙蛋白(OCN) mRNA表达;Western blot检测Runx2、OPN、OCN、BMP9、Notch2、Notch胞内域(NICD)、发状分裂相关增强子1(Hes1)、TLR4、NF-κB p65蛋白表达。结果:与non-CAVD组相比,CAVD组中BMP9和Runx2蛋白明显升高(P<0.05)。原代猪AVICs分离成功,其中α-平滑肌肌动蛋白(α-SMA)、Vimentin呈阳性表达,血小板内皮细胞黏附分子-1(PECAM-1,CD31)呈阴性表达。敲减BMP9后,AVICs中BMP9、Runx2、OCN、OPN mRNA水平、早期及晚期成骨分化能力及BMP9、Runx2、OCN、OPN、Notch2、NICD、Hes1、TLR4、NF-κB p65蛋白水平均下降(P<0.05);过表达BMP9后,AVICs中BMP9、Runx2、OCN、OPN mRNA水平,早期及晚期成骨分化能力,BMP9、Runx2、OCN、OPN、Notch2、NICD、Hes1、TLR4、NF-κB p65蛋白水平均升高(P<0.05)。结论:BMP9能够促进AVICs的成骨样分化,这一作用机制可能与Notch和TLR4信号通路的激活有关。

铸造增压压力对汽车用AlSi9Mg铝合金组织和力学性能的影响

通过改变低压铸造过程中不同增压压力来研究其对AlSi9Mg合金显微组织与力学性能的影响。结果表明,在高增压压力制备的AlSi9Mg合金样品具有更加致密的显微组织及更接近球形的共晶Si颗粒。同时,由于具有更好的填充能力,高增压压力下合金的气孔缺陷大大减少,合金具有更优异的力学性能。在90、150、210 kPa3种不同级别的增压压力下,晶粒尺寸呈逐渐减小的趋势,对应的合金晶粒尺寸和二次枝晶臂间距分别为246、197、149μm,44.9、37.3、31.1μm;3种增压压力对应的密度和孔隙率分别为2.656、2.659、2.674 g/cm^(3),0.96%、0.79%、0.38%;AlSi9Mg合金的抗拉强度、屈服强度和伸长率分别为238.76、174.68 MPa和3.14%;252.43、177.47 MPa和4.61%;256.10、178.81 MPa和6.85%,分别提高了7.26%、2.36%、118.15%。

miR-126过表达和ADAM9基因沉默对胃癌SGC-7901细胞生物学行为的影响及其机制

目的:探讨微小RNA-126(miR-126)过表达和解整联蛋白金属蛋白酶9(ADAM9)基因沉默对胃癌细胞生物学行为的影响,并阐明其作用机制。方法:体外培养人胃低分化腺癌SGC-7901细胞和人正常胃黏膜上皮NGEC细胞,提取细胞中总RNA,采用实时荧光定量PCR(RT-qPCR)法检测2种细胞中miR-126和ADAM9 mRNA表达水平。将处于对数生长期的SGC-7901细胞分为miR-126过表达组(miR-126-OE组)和ADAM9基因沉默组(ADAM9 siRNA组),以LipofectamineTM 2000为载体分别转染miR-126模拟物(miR-126 mimics)和敲低ADAM9的RNA寡核苷酸,并设置相应的阴性对照组。采用噻唑蓝(MTT)法检测各组细胞增殖活性,细胞划痕实验检测各组细胞迁移率,Transwell小室实验检测各组细胞的迁移细胞数和侵袭细胞数,Western blotting法检测各组细胞中E-钙黏蛋白、N-钙黏蛋白和波形蛋白表达水平。TargerScan网站预测miR-126的靶基因并通过双荧光素酶报告基因实验验证miR-126和ADAM9的靶向调控关系,采用RT-qPCR法和Western blotting法检测转染miR-126 mimics后SGC-7901细胞中ADAM9 mRNA和蛋白表达水平。结果:RT-qPCR法检测,与人正常胃黏膜上皮NGEC细胞比较,胃癌SGC-7901细胞中miR-126表达水平明显降低(P<0.05),而ADAM9mRNA表达水平明显升高(P<0.05)。MTT法检测,SGC-7901细胞过表达miR-126或沉默ADAM9基因48和72 h后,与相应阴性对照组比较,miR-126 OE组和ADAM9 siRNA组细胞增殖活性均明显降低(P<0.05或P<0.01)。细胞划痕实验检测,与相应阴性对照组比较,48 h后miR-126 OE组和ADAM9 siRNA组细胞迁移率明显降低(P<0.05)。Transwell小室实验检测,与相应阴性对照组比较,miR-126 OE组和ADAM9 siRNA组迁移细胞数和侵袭细胞数明显减少(P<0.05或P<0.01)。Western blotting法检测,与相应阴性对照组比较,miR-126-OE组和ADAM9 siRNA组细胞中E-钙黏蛋白表达水平明显升高(P<0.05或P<0.01),而N-钙黏蛋白和波形蛋白表达水平明显降低(P<0.05或P<0.01)。靶基因预测,ADAM9的3´-UTR含有与miR-126-3p互补的核苷酸序列。双荧光素酶报告基因实验,ADAM9是miR-126靶向负调控的下游基因。SGC-7901细胞转染miR-126 mimics 48 h后,与mimics NC组比较,miR-126 OE组细胞中ADAM9 mRNA和蛋白表达水平均明显降低(P<0.05或P<0.01)。结论:胃癌SGC-7901细胞中miR-126低表达而ADAM9基因高表达。miR-126过表达可抑制胃癌SGC-7901细胞增殖活性、迁移和侵袭能力,其机制可能与miR-126靶向负调控ADAM9并抑制胃癌细胞的上皮-间质转化(EMT)进程有关。

组织中MMP-9、E-cadherin与子宫内膜样癌伴MELF浸润方式及肿瘤侵袭程度的关系

目的分析子宫内膜样癌组织中基质金属蛋白酶(matrix metalloproteinase,MMP)-9、E-钙黏蛋白(E-cadherin)与微囊性、伸长及碎片状(microcystic,elongation,and fragmentation,MELF)浸润方式及肿瘤侵袭程度之间的关系,探讨能够识别子宫内膜样癌伴MELF浸润的指标,抑制肿瘤侵袭能力。方法回顾性分析在河北北方学院附属第一医院接受手术治疗的112例子宫内膜样癌患者临床资料,全部患者均顺利完成改良根治性子宫切除术,并获得肿瘤标准,根据病理检查结果对112例患者进行分组,分为MELF组和N-MELF组。统计112例患者一般临床资料,血液检查项目包括肿瘤标志物,免疫组织化学检查项目包括组织中MMP-9、E-cadherin,根据检查结果,构建Logistic回归模型,分析组织中MMP-9、E-cadherin及相关指标与子宫内膜样癌MELF浸润方式及肿瘤侵袭程度的关系。结果研究中112例子宫内膜样癌患者,评估为MELF浸润患者21例(18.75%);MELF浸润患者病理等级低级别,国际妇产科联盟(International Federation of Gynecology and Obstetrics,FIGO)ⅣA期,淋巴脉管间隙浸润(lymphatic vascular space,LVSI)、组织中MMP-9阳性患者占比多于N-MELF浸润患者,E-cadherin阳性患者占比少于N-MELF浸润患者,人附睾蛋白(humanepididymisprotein 4,HE4),甲壳质酶蛋白(human cartilageglycopmtein,YKL-40)水平高于N-MELF患者,差异有统计学意义(P<0.05)。多元Logistic回归分析提示,组织中E-cadherin表达率低、MMP-9表达率高,MELF浸润与子宫内膜样癌FIGO分期进展有关(P<0.05)。结论子宫内膜样癌发生MELF浸润与组织中MMP-9、E-cadherin有关,MMP-9表达率高和E-cadherin表达率低能够通过导致MELF浸润发生,从而增强肿瘤侵袭能力。