MicroRNA-329-3p inhibits the Wnt/β-catenin pathway and proliferation of osteosarcoma cells by targeting transcription factor 7-like 1

An important factor in the emergence and progre sion of osteosarcoma(OS)is the dysregulated expression of microRNAs(miRNAs).Transcription factor 7-like 1(TCF7LI),a member of the T cell factor/lymphoid enhancer factor(TCF/LEF)transcription factor family,interacts with the Wnt signaling pathway regulator β-catenin and acts as a DNA-specific binding protein.This study sought to elucidate the impact of the interaction between miR 3293p and TCF7L1 on.the growth and apoptosis of OS and analyze the regulatory expression relationship between miRNA and mRNA in osteosarcoma cells using a variety of approaches.MiR329-3p was significantly downregulated,while TCF7L1 was considerably up-regulated in all examined OS cell lines.Additionally,a clinical comparison study was performed using the TCGA database.Subsequently,the regulatory relationship between miR-329-3p and TCF7L1 on the proliferation and apoptosis of OS cells was verified through in vitro and in vivo experiments.When miR 329-3p was transfected into the OS cell line,the expression of TCF7L1 decreased,the proliferation of OS cells was inhibited,the cytoskeleton disintegrated,and the nucleus condensed to fom apoptotic bodies.The expression of proteins that indicate apoptosis increased simultaneously.The cell cycle was arrested in the G0/G1 phase,and the G1/S transition was blocked.The introduction of miR 3293p also inhibited downstream Cyclin D1 of the Wnt pathway.Xenograf experiments indicated that the overexpression of miR-329-3p signi ficanly inhibited the growth of OS xenografts in nude mice,and the expression of TCF7L1 and C-Myc in tumor tssues decreased.MiR 329-3p was significantly reduced in OS cells and played a suppressive role in tumorigenesis and proliferation by targeting TCF7L1 both in vitro and in vivo.Osteosarcoma cell cycle arrest and pathway inhibition were observed upon the regulation of TCF7LI by miR 3293p.Summarizing these results,it can be inferred that miR.3293p exerts anticancer efects in osteosarcoma by inhibiting TCF7L1.

Acalypha australis L.extract inhibits B16 melanoma cell metastasis through PI3K/AKT signaling pathway

Background:Melanoma is a deadly skin tumor resulting from the malignant transformation of melanocytes.It is highly malignant and invasive,with the highest mortality rate among skin cancers.Acalypha australis L.(AAL),a plant with dual medicinal and culinary purposes,is commonly regarded as an edible wild vegetable in southern China.Additionally,AAL has a long history of medicinal use in China,often employed for its hemostatic,anti-diarrheal,and anti-inflammatory properties.Modern pharmacology has demonstrated that AAL possesses functions such as weight loss,antimicrobial activity,antiviral effects,and treatment for ulcerative colitis.However,there is currently no research available regarding its effectiveness and mechanisms of action on melanoma.Methods:In this investigation,we used methyl thiazolyl tetrazolium assay to detect cell viability,transwell assay to detect cell migration and invasion ability,and Western blot assay to detect relevant signaling pathways.Results:The present study reveals that 2 mg/mL AAL effectively suppresses the metastasis of B16 cells,while simultaneously triggering the expression of key apoptosis-related proteins,including Bcl-2,Bax,and cleaved caspased 3.Subsequent investigations demonstrate that AAL exerts this inhibitory effect via the PI3K/AKT signal transduction pathway,as evidenced by the observed deficits in Ras,AKT,p-AKT,and PI3K expression levels.Conclusion:These findings indicated that AAL could be a valuable therapeutic option for reducing the metastatic potential of B16 melanoma cells.

耐力训练和补充L-Arg对大鼠心肌、肝脏eNOS mRNA表达的影响

目的探讨耐力训练和补充L-Arg对大鼠心肌、肝脏eNOSmRNA表达的影响。采用RT-PCR法对大鼠心肌、肝脏eNOSmRNA的表达进行检测。实验结果心肌组织中,所有运动组eNOSmRNA的表达均较安静组显著下降(P<0.01);力竭组与其他各组相比较均有高度显著性差异(P<0.01),所有其他组相对表达量较力竭组相对表达量高。肝脏中,大小剂量力竭组和大剂量耐力训练组eNOSmRNA表达较安静对照组、耐力训练组、力竭组和小剂量耐力训练组均有显著下降(P<0.01)。结论耐力训练抑制心肌eNOSmRNA的表达;力竭运动提高其表达。耐力训练显著提高肝脏eNOSmRNA的表达(P<0.05);补充L-Arg抑制其表达,且抑制作用随L-Arg剂量的增加而相应增大。

IFN-γ、L-Arg及L-NNA对感染旋毛虫的沙鼠血液及腓肠肌NO合成和组织病理变化的影响

目的探讨IFN-γ、L-Arg和L-NNA对感染旋毛虫的沙鼠血液及腓肠肌NO合成和组织病理变化的影响。方法经腹腔给予感染旋毛虫后不同时期的沙鼠不同剂量的IFN-γ、L-Arg和L-NNA,检测沙鼠血清及腓肠肌NO的含量。观察腓肠肌的病理变化。结果IFN-γ和L-Arg组沙鼠血清及腓肠肌NO含量随药物浓度的增加而升高;L-NNA阴性对照组血清NO含量和感染25 d的L-NNA组腓肠肌NO含量升高,L-NNA实验组血清NO含量及阴性对照组、旋毛虫感染7 d组4、0 d组腓肠肌NO含量降低。实验组腓肠肌内囊包数较阳性对照组略有减少。结论IFN-γ和L-Arg能促进感染旋毛虫的沙鼠血液及腓肠肌NO合成,L-NNA则具有增强或抑制NO合成的双重作用。

L-Arg-NO途径对铝抑制大鼠海马CA3区诱发电位的影响

实验在70只乌拉坦麻醉的SD大鼠上进行。在海马CA3区记录诱发的群体锋电位（PS）,探讨了左旋精氨酸——氧化氮（L-Arg-NO）途径对铝抑制PS波幅的作用。结果:（1）0.5 mol/L AICI_3注入CA3区后,PS幅度较注药前显著减小（P<0.05）。（2）CA3区内注入0.1和 0.3mol/L硝基左旋精氨酸（ NLA）能分别加强0.25和0.5 mol/L AICl_3 的抑制效应。（3）CA3区注入0.3 mol/L左旋精氨酸（L-Arg）能拮抗0.5 mol/L AICI_3 对PS波幅的抑制作用,此拮抗作用并能为 NLA所翻转。（4）CA3区内注入 0.2mmol/L亚甲蓝（MB）能加强铝对PS的抑制作用。结果提示:AICI_3对CA3区诱发的PS幅度有抑制作用,此作用可能与L-Arg-NO途径受到损害有关。

体外培养中IFN-γ、L-Arg及L-NNA对NO合成的影响及NO抗旋毛虫的作用

目的 探讨体外培养中IFN γ、L Arg及L NNA对NO合成的影响及NO抗旋毛虫的作用。 方法 分离、纯化长爪沙鼠腹腔巨噬细胞 ,置RPMI16 4 0培养液中培养。设IFN γ组、L Arg组、L NNA组和对照组 ,每个实验组又分 5个不同的浓度组。分别向含有巨噬细胞的培养瓶中加入不同浓度的IFN γ、L Arg及L NNA进行体外培养。培养 2 4h后 ,用硝酸还原酶法分别测定培养液中的NO含量。将旋毛虫幼虫分别加入上述培养体系中进行体外培养 ,观察旋毛虫幼虫的活动及损伤。结果 ①体外培养中 ,激活的巨噬细胞能产生NO ,IFN γ和L Arg能促进NO的合成 ,L NNA则能抑制NO的合成 ,这种促进或抑制NO合成的作用均具有剂量依赖性 ,剂量越高作用越明显。②加入旋毛虫幼虫后 ,在IFN γ和L Arg培养体系中 ,随着NO浓度的升高及作用时间的延长 ,对虫体的抑制及杀伤作用越来越明显 ,导致其活动度减弱 ,虫体破裂 ,最终死亡 ;在L NNA培养体系中 ,L NNA浓度越高 ,对虫体的影响越小。结论 ①体外培养中 ,通过激活的巨噬细胞 ,IFN γ和L Arg能促进NO的合成 ,给予L NNA则能抑制NO的合成。

L-Arg在心肺联合移植中对心肺保护作用的实验研究

目的研究L-Arg在心肺联合移植中对心肺的保护作用及可能机制。方法将健康成年犬20只随机分为两组,对照组以4℃LPD液灌注及保存供肺,实验组以4℃含L-Arg(500mg/kg)的LPD液灌注及保存供肺。心脏灌注液采用4℃St.thomas液,实验组中加入L-Arg300mg/500mL。分别监测受体麻醉后和心肺移植后主动脉开放5min和主动脉开放30min的血气分析,测定心肌和肺组织中一氧化氮(NO)、超氧化物歧化酶(SOD)、丙二醛(MDA)的含量及肺组织的湿/干重比(W/D),并观察移植肺组织的超微结构变化。结果实验组血氧分压(PaO2)高于对照组,二氧化碳分压(PaCO2)明显低于对照组。实验组心肌和肺组织中的NO及SOD的含量较对照组增高(P<0.01),而MDA的含量较对照组降低(P<0.05)。实验组较对照组的W/D低(P<0.05)。电镜检查实验组的肺组织损伤轻于对照组。结论L-Arg可增强保存液对心脏的保护作用,改善移植肺的肺功能,能减轻供肺损伤,增加心肺联合移植的成功率。

L-Arg及L-NNA对旋毛虫病沙鼠小肠及肝NO合成和组织病理变化的影响

目的探讨L-Arg和L-NNA对感染旋毛虫的沙鼠小肠及肝NO合成和组织病理变化的影响。方法在感染旋毛虫后的早、中、晚期经腹腔给予沙鼠不同剂量的L-Arg和L-NNA,检测沙鼠小肠及肝组织NO的含量;观察小肠组织的病理变化。结果L-Arg组沙鼠小肠及肝NO含量均明显升高,且随药物浓度的增加而升高。给予L-NNA后,小肠组织实验7d组、25d组NO量明显高于阳性对照组,且随L-NNA剂量的增加而升高,40d组及阴性对照组NO量则明显低于阳性对照组(P<0.05),且随药物浓度的增加而降低;肝组织实验组NO量普遍明显低于阳性对照组(P<0.01),且随L-NNA剂量的增加及感染时间的延长而降低。高剂量组小肠组织的病理损伤加重。结论L-Arg能促进感染旋毛虫的沙鼠小肠及肝NO的合成,L-NNA则表现出不同的增强或抑制NO合成的双重作用。

血小板L-arg/NO系统在妊高征中的表达及意义

目的:观察重度妊高征患者血小板L-arg/NO系统的变化。方法:选择正常妊娠孕晚期患者19例(对照组),重度妊高征患者21例(观察组),采用放射性核素3H-L-arg标记法测定血小板内NOS的活性,血小板L-arg转运,比色法测定血小板内NO含量。结果:重度妊高征患者血小板L-arg转运为3.28±0.71pmol/108pt·min-1,较正常降低43.15%(P<0.01);血小板NOS活性为5.33±1.75pmol/108pt·min-1,较正常降低31.84%(P<0.01);NO水平较正常降低,有显著性差异(P<0.05)。结论:重度妊高征时血小板L-arg转运降低,NOS的活性降低,血小板内NO水平降低,血小板L-arg/NOS/NO系统明显损伤。

RGDS肽对胶元活化的大鼠血小板L-Arginine/NO途径的影响

本工作在胶原活化的大鼠血小板上，观察RGDS肽对血小板聚集、L一精氨酸（L－Arg）转运、一氧化氮合酶（NOS）活性和亚硝酸盐（NO2－）含量的影响。结果发现，4μg／mL胶原引起血小板聚集时，L－Arg转运增强、NOS活性增高和NO2-含量增加。血小板L－Arg转运的Vmax与NOS活性和NO2-生成呈明显正相关（r=0.8100和0.8343，p＜0．01）；血小板聚集与NO2-生成亦呈明显正相关（r=0.7660，P＜0.05）。RGDS肽200μmol／L凡与胶原共同孵育，可明显抑制胶原引起的血小板聚集、L－Arg转运增强、NOS活性增高和NO2-含量增加。提示，胶原激活的血小板L－Arg／NO途径增加是通过Ⅱb／Ⅲa复合物所介导，RGDS肽可括抗其增加作用。

左旋精氨酸通过激活L-arg-NO通路对异丙肾上腺素致大鼠心力衰竭的作用

目的探究左旋精氨酸(L-arg)通过激活L-arg-NO通路对异丙肾上腺素(ISO)导致的大鼠心力衰竭的保护作用。方法雄性SD大鼠随机分为对照组、ISO组、L-arg干预组。腹腔注射ISO复制大鼠心力衰竭模型,灌胃给予L-arg。30 d后监测血流动力学参数:心率(HR)、左室收缩压力(LVSP)、左室舒张末期压力(LVEDP)、左室压力最大上升或下降速率(±dp/dtmax),计算心肺系数,检测血清超氧化物歧化酶(SOD)、肿瘤坏死因子-α(TNF-α)、一氧化氮(NO)含量以及一氧化氮合酶(NOS)活性;HE染色观察心肌组细胞排列;Masson染色观察心肌胶原纤维并计算心肌胶原纤维容积分数(CVF)。结果与对照组比较,ISO组大鼠血流动力学参数:HR、LVSP、+dp/dtmax降低,LVEDP、-dp/dtmax升高,心肺系数增加,血清TNF-α、SOD升高,NO、NOS降低,差异均有统计学意义(P<0.05)。L-arg干预组HR、LVSP、+dp/dtmax均高于ISO组,LVEDP、-dp/dtmax、心肺系数低于ISO组,血清SOD和TNF-α降低,NO和NOS增加,差异均有统计学意义(P<0.05)。HE染色显示对照组心肌细胞形态正常,间质无扩张充血,心肌细胞排列整齐;ISO组心肌细胞排列紊乱,炎性细胞浸润,而L-arg干预组心肌细胞接近对照组。ISO组Masson染色心肌细胞间可见大量蓝染的胶原纤维,而对照组观察不到蓝染区域,L-arg干预组在对照组与ISO组之间;ISO组CVF明显高于L-arg干预组(P<0.05),L-arg干预组高于对照组(P<0.05)。结论 L-arg可以通过激活Larg-NO通路,增加机体内NO含量,改善血流,减轻心肌重构,产生对心力衰竭心肌的保护作用。

外源L-Arg处理对蜜橘果实贮藏品质的影响

用100、200 μmol/L外源L-精氨酸（L-arginine,L-Arg）溶液浸泡处理蜜橘果实10 min,探讨该处理对采后蜜橘果实贮藏品质的影响.结果表明,适宜浓度（200 μmol/L）的外源L-Arg处理可以有效抑制蜜橘果实黄化,在一定程度上抑制果皮细胞膜透性的增加;在贮藏的前28 d内,L-Arg浸泡处理能够维持果实可溶性固形物的含量;各处理组之间的呼吸速率没有显著性差异,适宜浓度（200 μmol/L L-Arg）的L-Arg浸泡处理能够在一定程度上抑制蜜橘果实质量损失率的增加;延缓果实总酚、抗坏血酸含量的减少,贮藏前期（0～21 d）处理组果实果肉类黄酮含量与对照组之间无显著差异,贮藏后期（28～35 d）处理组果实类黄酮含量显著低于对照组果实.即200 μmol/L外源L-Arg处理能够在一定程度上有效延缓采后蜜橘果实的衰老和品质下降.

急性脑梗死rt-PA溶栓时NO/ET-1、L-Arg、PCT与神经功能关系及预测血管再通效能

目的 探讨急性脑梗死重组人组织型纤溶酶原激活物(rt-PA)溶栓时一氧化氮与内皮素-1比值(NO/ET-1)、L-精氨酸(L-Arg)、血小板压积(PCT)与神经功能关系及预测血管再通效能。方法 选取2018年3月至2021年1月河南中医药大学第一附属医院收治的182例具有静脉溶栓指征急性脑梗死患者,根据rt-PA溶栓后血管再通情况分为非再通组(n=31)、再通组(n=151),比较两组基线资料、NO/ET-1、L-Arg、PCT水平,对所获得数据进行统计分析。结果 非再通组糖尿病心房颤动、NIHSS评分、NO/ET-1、L-Arg、PCT与再通组比较,差异有统计学意义(P<0.05);NO/ET-1(r=-0.875,P<0.001)、L-Arg(r=-0.901,P<0.001)与NIHSS评分呈负相关,PCT(r=0.804,P<0.001)与NIHSS评分呈正相关;将糖尿病、心房颤动、NIHSS评分控制后,NO/ET-1、L-Arg、PCT仍是血管再通的相关影响因素(P<0.05);NO/ET-1、L-Arg联合PCT预测血管再通的受试者工作特征曲线下面积(AUC)(0.896)大于单一的NO/ET-1(0.816)、L-Arg(0.778)、PCT(0.812);NO/ET-1、L-Arg高水平者血管再通率高于低水平者,PCT高水平者血管再通率低于低水平者(χ^(2)=32.475、28.344、35.495,P<0.05)。结论 急性脑梗死rt-PA溶栓前NO/ET-1、L-Arg降低及PCT升高与神经功能恶化和血管非再通风险增加有关,联合检测三者能为临床预测血管再通提供参考,从而为临床决策、治疗提供依据。

L-精氨酸通过L-Arg-NO途径促进大鼠背部跨区皮瓣的成活

背景:有研究表明一氧化氮可有效改善皮瓣术后血供,促进皮瓣成活,但具体机制尚不清楚。目的:通过实验验证L-Arg-NO途径在L-精氨酸促进大鼠背部跨区皮瓣成活中的作用。方法:成功建立大鼠背部三穿支体跨区皮瓣模型的雄性SD大鼠81只,随机被分为3组,分别于术后即刻、术后1-7 d腹腔注射不同药物,L-Arg组注射L-精氨酸400 mg/(kg·d);L-NAME组注射一氧化氮合酶抑制剂40 mg/(kg·d);空白组注射等体积等渗氯化钠溶液。术后7 d观察皮瓣成活情况,计算皮瓣成活率;颈总动脉灌注明胶-氧化铅行血管造影,观察血管走形和分布;硝酸还原酶法测定术后即刻、1,3,5,7 d chokeⅡ区一氧化氮水平;苏木精-伊红染色观察chokeⅡ区新生血管密度和管径;蛋白印记法检测术后3,7 d chokeⅡ区血管内皮生长因子和基质金属蛋白酶2蛋白表达。实验方案经解放军联勤保障部队第九四〇医院动物实验伦理委员会批准(批准号为2015KYLL046)。结果与结论:①术后7 d,L-Arg组大鼠皮瓣成活率最高(89.47±3.17)%,3组比较差异有显著性意义(F=49.908,P<0.001);②术后7 d,L-Arg组chokeⅡ区血管结构较完整,走形清楚,扩张达到真性吻合的血管沿着皮瓣纵轴方向一直延伸到皮瓣末端,空白组和L-NAME组chokeⅡ区血管结构和走形杂乱,L-NAME组皮瓣末端血管结构消失;③L-Arg组一氧化氮水平术后开始升高,术后3 d达到高峰,之后逐渐下降;④术后7 d,L-Arg组新生血管密度和管径最高,3组比较差异有显著性意义(均P<0.001);⑤术后3 d,L-Arg组血管内皮生长因子和基质金属蛋白酶2蛋白表达量最高,3组比较差异有显著性意义(均P<0.05);术后7 d,L-Arg组血管内皮生长因子和基质金属蛋白酶2蛋白表达量最高,3组之间比较血管内皮生长因子差异有显著性意义(P<0.05),基质金属蛋白酶2蛋白差异无显著性意义;⑥结果说明,一氧化氮可有效改善大鼠背部三穿支体皮瓣chokeⅡ区血供,促进chokeⅡ区微血管扩张和增生,而L-精氨酸可通过L-Arg-NO途径提高皮瓣术后组织内一氧化氮水平,促进皮瓣术后成活。

Essential Oil from Carpesium abrotanoides L.Induces Apoptosis via Activating Mitochondrial Pathway in Hepatocellular Carcinoma Cells

The effects of essential oil from Carpesium abrotanoides L.(CAEO)on the proliferation and apoptosis of human hepatic cancer cells were investigated in this study.MTT assays indicated that CAEO inhibited the proliferation of HCC cells with the IC50 values ranging from 41.28±3.06 to 130.36±20.79μg/mL.Moreover,many obviously nuclear morphological changes of apoptotic cells in CAEO-treated HepG2 cells were detected by Hoechst 33258staining and fluorescence microscopy.Flow cytometry was used to detect cell apoptosis and cell cycle,and noticeable findings showed that CAEO arrested cell-cycle at S and G2/M phases.The decreased Bcl-2/Bax protein ratio and the activation of caspase-3,caspase-9 were also detected by Western blotting.All results suggested that CAEO is a potential agent to fight against liver cancer,and the mitochondria-mediated intrinsic apoptotic pathway could be involved in CAEO-mediated apoptosis of human liver carcinoma cells.

The NAC-like transcription factor Si NAC110 in foxtail millet(Setaria italica L.) confers tolerance to drought and high salt stress through an ABA independent signaling pathway

Foxtail millet（Setaria italica（L.）P.Beauv）is a naturally stress tolerant crop.Compared to other gramineous crops,it has relatively stronger drought and lower nutrition stress tolerance traits.To date,the scope of functional genomics research in foxtail millet（S.italic L.）has been quite limited.NAC（NAM,ATAF1/2 and CUC2）-like transcription factors are known to be involved in various biological processes,including abiotic stress responses.In our previous foxtail millet（S.italic L.）RNA seq analysis,we found that the expression of a NAC-like transcription factor,SiNAC110,could be induced by drought stress;additionally,other references have reported that SiNAC110 expression could be induced by abiotic stress.So,we here selected SiNAC110 for further characterization and functional analysis.First,the predicted SiNAC110 protein encoded indicated SiNAC110 has a conserved NAM（no apical meristem）domain between the 11–139 amino acid positions.Phylogenetic analysis then indicated that SiNAC110 belongs to subfamily III of the NAC gene family.Subcellular localization analysis revealed that the SiNAC110-GFP fusion protein was localized to the nucleus in Arabidopsis protoplasts.Gene expression profiling analysis indicated that expression of SiNAC110 was induced by dehydration,high salinity and other abiotic stresses.Gene functional analysis using SiNAC110 overexpressed Arabidopsis plants indicated that,under drought and high salt stress conditions,the seed germination rate,root length,root surface area,fresh weight,and dry weight of the SiNAC110 overexpressed lines were significantly higher than the wild type（WT）,suggesting that the SiNAC110 overexpressed lines had enhanced tolerance to drought and high salt stresses.However,overexpression of SiN AC110 did not affect the sensitivity of SiNAC110 overexpressed lines to abscisic acid（ABA）treatment.Expression analysis of genes involved in proline synthesis,Na＋/K＋transport,drought responses,and aqueous transport proteins were higher in the SiNAC110overexpressed lines than in the WT,whereas expression of ABA-dependent pathway genes did not change.These results indicated that overexpression of SiNAC110 conferred tolerance to drought and high salt stresses,likely through influencing the regulation of proline biosynthesis,ion homeostasis and osmotic balance.Therefore,SiNAC110 appears to function in the ABA-independent abiotic stress response pathway in plants.

Volatile Oil of Platycladus Orientalis(L.)Franco Leaves Exerts Strong Anti-inflammatory Effects via Inhibiting the IκB/NF-κB Pathway

This study was designed to investigate the anti-inflammatory effects of volatile oil of Platycladus orientalis(L.)Franco leaves(VOPF)and the underlying molecular mechanisms by using the non-infectious inflammation rat models and infectious inflammation mouse models.Ear swelling and intraperitoneal capillary permeability in mice,and carrageenan-induced toe swelling and cotton ball-induced granuloma in rats were used to reveal anti-inflammatory effects of VOPF.Moreover,the lipopolysaccharide(LPS)-induced mouse model of acute lung injury was used to explore the anti-inflammatory mechanism of VOPF.The results showed that VOPF could significantly inhibit auricular swelling,intraperitoneal capillary permeability in mice,and reduce granuloma swelling and paw swelling in rats.Furthermore,it significantly alleviated the pathological damage of the lung tissue.In addition,VOPF could reduce the contents of IL-1β and TNF-αand increase the content of IL-10 in the serum.It had little effect on the expression of p65 but reduced the phosphorylation level of p65 and IκB in NF-κB pathway.In conclusion,VOPF has anti-inflammatory effects and the mechanisms involve the down-regulation of the phosphorylation levels of p65 and IκB and blockage of the NF-κB signaling pathway.

Effect of arginine on stability of GST-ZNF191(243-368)

For better understanding the chemical or biological information of ZNF191 （243-368）, we expressed the fusion protein of GST and ZNF191 （243-368）, and used it to obtain the binding DNA sequence of this zinc finger protein. But in the process of expression and purification, we found this fusion protein slowly degradated. For resolving this problem, we simultaneously added charged amino acids L-Arg and L-Glu to the solution of fusion protein, and demonstrated that this method can dramatically increase the stability of this fusion protein. This method can make the fusion protein suitable for the continuous works, especially for situations where high protein concentration and long-term stability without precipitate and degradation of protein are required.

Immunoregulatory polysaccharides from Apocynum venetum L.flowers stimulate phagocytosis and cytokine expression via activating the NF-κB/MAPK signaling pathways in RAW264.7 cells

Two immunomodulatory polysaccharides(Vp2a-Ⅱ and Vp3) were isolated and identified from Apocynum venetum L. flowers, and their innate immune-stimulating functions and working mechanisms were evaluated in RAW264.7 cells. Both the level of released nitric oxide(NO) and expression of inducible nitric oxide synthase(iNOS) m RNA were significantly enhanced in the RAW264.7 macrophages cells treated by Vp2a-Ⅱ and Vp3. Vp2a-Ⅱ(100–800 μg/m L) and Vp3(400 μg/mL) could significantly increase the phagocytic activity of RAW264.7 cells and the secretion and m RNA expression of TNF-α and IL-6 in a concentrationdependent manner through affecting mitogen-activated protein kinase(MAPK) activity and nuclear factor κB(NF-κB) nuclear translocation. Vp2a-Ⅱ might activate the MAPK signaling pathways and induce the nuclear translocation of NF-κB p65, whilst Vp3 likely activated the NF-κB and MAPK signaling pathways without influencing the p38 MAPK route.

Genome-wide association study and metabolic pathway prediction of barrenness in maize as a response to high planting density

Increasing the planting density is one way to enhance grain production in maize.However,high planting density brings about growth and developmental defects such as barrenness,which is the major factor limiting grain yield.In this study,the barrenness was characterized in an association panel comprising 280 inbred lines under normal(67500 plants ha–1,ND)and high(120000 plants ha–1,HD)planting densities in 2017 and 2018.The population was genotyped using 776254 single nucleotide polymorphism(SNP)markers with criteria of minor allele frequency>5%and<20%missing data.A genome-wide association study(GWAS)was conducted for barrenness under ND and HD,as well as the barrenness ratio(HD/ND),by applying a Mixed Linear Model that controls both population structure and relative kinship(Q+K).In total,20 SNPs located in nine genes were significantly(P<6.44×10–8)associated with barrenness under the different planting densities.Among them,seven SNPs for barrenness at ND and HD were located in two genes,four of which were common under both ND and HD.In addition,13 SNPs for the barrenness ratio were located in seven genes.A complementary pathway analysis indicated that the metabolic pathways of amino acids,such as glutamate and arginine,and the mitogen-activated protein kinase(MAPK)signaling pathway might play important roles in tolerance to high planting density.These results provide insights into the genetic basis of high planting density tolerance and will facilitate high yield maize breeding.