钙调蛋白依赖性蛋白激酶Ⅰ(Calmodulin Dependent Protein Kinase Ⅰ,CAMK1)基因是一种蛋白质编码基因,参与细胞分化,可能会与繁殖相关。本研究在课题组前期关于阳原驴产仔数血液转录组分析的基础上,探索候选基因CAMK1在阳原驴群体中的...钙调蛋白依赖性蛋白激酶Ⅰ(Calmodulin Dependent Protein Kinase Ⅰ,CAMK1)基因是一种蛋白质编码基因,参与细胞分化,可能会与繁殖相关。本研究在课题组前期关于阳原驴产仔数血液转录组分析的基础上,探索候选基因CAMK1在阳原驴群体中的遗传多态性,并与产仔数进行关联分析,寻找与阳原驴产仔数显著相关的分子标记,为驴的专门化品种培育提供参考。试验以86头阳原驴母驴(其中10头为产双仔母驴)为研究对象,通过PCR-SSCP技术检测CAMK1基因SNP位点并将SNP位点与阳原驴产仔数进行关联分析。结果表明,在CAMK1基因中发现3个单碱基突变,分别为g.5309T>C、g.8164T>C、g.9059C>T,且3个突变位点均检测出CC、CT、TT 3种基因型。结果显示,g.5309T>C位点CT基因型个体的平均产仔数显著高于TT基因型;g.8164T>C位点CC和CT基因型个体的平均产仔数显著高于TT基因型;g.9059C>T位点CC基因型个体的平均产仔数显著高于TT基因型。综上表明,CAMK1基因3个位点与产仔数性状显著相关,推测CAMK1基因可作为阳原驴产仔数性状研究的候选基因和分子标记。展开更多
目的探讨经颅直流电刺激(transcranial direct current stimulation,tDCS)对单眼剥夺(Monocular deprivation,MD)大鼠初级视皮层(V1)中CAMKⅡ、CREB和SYN1蛋白表达及视觉能力的影响,为弱视治疗提供理论基础。方法55只SPF级新生SD大鼠,...目的探讨经颅直流电刺激(transcranial direct current stimulation,tDCS)对单眼剥夺(Monocular deprivation,MD)大鼠初级视皮层(V1)中CAMKⅡ、CREB和SYN1蛋白表达及视觉能力的影响,为弱视治疗提供理论基础。方法55只SPF级新生SD大鼠,从中选取43只于出生后14 d缝合右侧上下眼睑以建立单眼剥夺模型,单眼剥夺2周后选取36只缝合严密不漏光的大鼠随机分为模型组(MD)、阳极治疗组(anodaltDCS,a-tDCS)、假治疗组(shamtDCS,s-tDCS)。模型建立后进行为期14 d的tDCS治疗,对a-tDCS组每天施以1mA直流电刺激,每天1次,每次20 min;s-tDCS组只在刺激开始后1 min和结束前1 min给予电流刺激。视觉水迷宫用于测试视觉辨别能力和选择行为。图形视觉诱发电位(Pattern visual evoked potential,P-VEP)检测各组大鼠P100波振幅和潜伏期;HE染色观察V1中细胞形态和排列情况;Westernblot法检测V1中CaMKⅡ、CREB和SYN1蛋白表达水平;免疫组织化学染色观察SYN1阳性细胞数量。结果与Control组比较,MD组大鼠视觉辨别能力降低,选择正确率显著下降(P<0.0001);P100波振幅显著降低(P<0.001),潜伏期延长(P<0.001);V1中核仁皱缩,细胞排列不规整;CaMKⅡ、CREB和SYN1蛋白表达量下降(P<0.001);SYN1蛋白的平均阳性细胞数量减少(P<0.0001)。a-tDCS组与MD组相比,大鼠视觉辨别能力显著改善,选择正确率升高(P<0.0001);P100波振幅升高(P<0.001),潜伏期缩短(P<0.001);V1神经元排列规整,皱缩有所减少;CaMKⅡ、CREB和SYN1蛋白表达量显著升高(P<0.001);V1中SYN1蛋白的平均阳性细胞数量增加(P<0.05)。s-tDCS对MD组无治疗作用。结论a-tDCS可改善单眼剥夺大鼠的视觉能力,其机制可能与a-tDCS通过CaMKⅡ/CREB通路调节V1神经元突触可塑性有关。展开更多
Background:Glioma is the most common tumor of the central nervous system with a poor prognosis.This study aims to explore the role of calcium/calmodulin-dependent protein kinase IIβ(CAMK2B)in regulating the malignant...Background:Glioma is the most common tumor of the central nervous system with a poor prognosis.This study aims to explore the role of calcium/calmodulin-dependent protein kinase IIβ(CAMK2B)in regulating the malignant progression of glioma cells,as well as the molecular mechanisms underlying these malignant behaviors.Methods:The correlation between CAMK2B expression in gliomas and patient prognosis was analyzed using immunohistochemistry,quantitative reverse transcription polymerase chain reaction(qRT-PCR),and western blot.Furthermore,the study explored the role of CAMK2B in glioma cell proliferation,invasion,and migration using cell counting kit-8(CCK-8),5-Ethynyl-2′-deoxyuridine(EdU),wound healing,transwell,and in vivo tumor xenograft assays.Result:Patients with high CAMK2B expression exhibited significantly better prognostic outcomes compared to those with low expression levels.Furthermore,CAMK2B expression was significantly lower in glioma tissues and cells compared to both normal brain tissue and human astrocyte cell lines.Notably,overexpression of CAMK2B in glioma cells led to an approximate 40%reduction in proliferative capacity and a 60–70%decrease in invasive and migratory abilities,compared to control glioma cells.These differences were statistically significant at p<0.05.Conversely,knockdown of CAMK2B using siRNA-CAMK2B significantly enhanced the proliferative,invasive,and migratory capabilities of glioma cells in both in vitro and in vivo settings,enhancing these abilities by 1.5 to 3 times.Notably,these effects were reversed through the application of the Rat Sarcoma viral oncogene homolog(Ras)pathway inhibitor,Salirasib.Western blot analysis revealed that knockdown of CAMK2B led to activation of the Ras/Rapidly Accelerated Fibrosarcoma(Raf)/Mitogen-activated protein kinase kinase(MEK)/Extracellular signal-regulated kinase(ERK)signaling pathway in glioma cell lines,whereas overexpression of CAMK2B resulted in the suppression of this pathway.Conclusion:CAMK2B inhibits glioma proliferation,invasion,andmigration through the Ras/Raf/MEK/ERK signaling pathway.展开更多
目的对“远志-酸枣仁”中潜在作用于钙/钙调蛋白依赖性蛋白激酶Ⅱ(calcium/calmodulin-dependent protein kinaseⅡ,CaMKⅡ)靶点的活性物质进行筛选,阐明其药效物质基础。方法采用以药效团为基础的虚拟筛选手段,通过收集已出版文献中报...目的对“远志-酸枣仁”中潜在作用于钙/钙调蛋白依赖性蛋白激酶Ⅱ(calcium/calmodulin-dependent protein kinaseⅡ,CaMKⅡ)靶点的活性物质进行筛选,阐明其药效物质基础。方法采用以药效团为基础的虚拟筛选手段,通过收集已出版文献中报道的具有调控CaMKⅡ作用的化合物信息,建立基于CaMKⅡ配体的HipHop药效团模型,收集文献中报道的“远志-酸枣仁”中的成分并建立化合物库,对“远志-酸枣仁”中的成分与药效团进行匹配,随后使用分子模拟对接手段对匹配到的小分子化合物与CaMKⅡ靶点(PDB ID:2V7O)进行对接,研究潜在活性单体与CaMKⅡ的作用。用戊四唑(pentylenetetrazol,PTZ)点燃模型对活性单体细叶远志苷A(39)的抗癫痫作用进行测定,并对CaMKⅡ水平的调控作用进行评价。结论通过文献检索建立包含“远志-酸枣仁”中109种化学成分的化合物库,通过测试集验证确定将药效团03用于对“远志-酸枣仁”化合物库的虚拟筛选,对匹配度排前6位的单体通过分子模拟对接分析其与CaMKⅡ的作用情况,并用抗癫痫动物模型对单体的活性进行评价,化合物39能够在中高剂量下延长PTZ致癫痫发作的潜伏时间,改善PTZ所致的焦虑抑郁样行为,改善PTZ诱导的癫痫发作后脑内尼氏体丢失,提升PTZ所致的CaMKⅡ表达水平降低,提升海马CA1、CA3区及颞叶皮层内CaMKⅡ免疫阳性细胞数量并进一步对nAChR4受体水平进行调控。表明基于虚拟筛选与活性评价手段探讨“远志-酸枣仁”中CaMKⅡ调控作用的成分具有一定的准确性。展开更多
基金supported by the Natural Science Foundation of Hebei Province(H2021206037)the Government-funded Project on Training of Outstanding Clinical Medical Personnel of Hebei Province in the year 2021(303-16-20-06)the Medical Research Project of Hebei Provincial Health Commission(20230031).
文摘Background:Glioma is the most common tumor of the central nervous system with a poor prognosis.This study aims to explore the role of calcium/calmodulin-dependent protein kinase IIβ(CAMK2B)in regulating the malignant progression of glioma cells,as well as the molecular mechanisms underlying these malignant behaviors.Methods:The correlation between CAMK2B expression in gliomas and patient prognosis was analyzed using immunohistochemistry,quantitative reverse transcription polymerase chain reaction(qRT-PCR),and western blot.Furthermore,the study explored the role of CAMK2B in glioma cell proliferation,invasion,and migration using cell counting kit-8(CCK-8),5-Ethynyl-2′-deoxyuridine(EdU),wound healing,transwell,and in vivo tumor xenograft assays.Result:Patients with high CAMK2B expression exhibited significantly better prognostic outcomes compared to those with low expression levels.Furthermore,CAMK2B expression was significantly lower in glioma tissues and cells compared to both normal brain tissue and human astrocyte cell lines.Notably,overexpression of CAMK2B in glioma cells led to an approximate 40%reduction in proliferative capacity and a 60–70%decrease in invasive and migratory abilities,compared to control glioma cells.These differences were statistically significant at p<0.05.Conversely,knockdown of CAMK2B using siRNA-CAMK2B significantly enhanced the proliferative,invasive,and migratory capabilities of glioma cells in both in vitro and in vivo settings,enhancing these abilities by 1.5 to 3 times.Notably,these effects were reversed through the application of the Rat Sarcoma viral oncogene homolog(Ras)pathway inhibitor,Salirasib.Western blot analysis revealed that knockdown of CAMK2B led to activation of the Ras/Rapidly Accelerated Fibrosarcoma(Raf)/Mitogen-activated protein kinase kinase(MEK)/Extracellular signal-regulated kinase(ERK)signaling pathway in glioma cell lines,whereas overexpression of CAMK2B resulted in the suppression of this pathway.Conclusion:CAMK2B inhibits glioma proliferation,invasion,andmigration through the Ras/Raf/MEK/ERK signaling pathway.
文摘目的对“远志-酸枣仁”中潜在作用于钙/钙调蛋白依赖性蛋白激酶Ⅱ(calcium/calmodulin-dependent protein kinaseⅡ,CaMKⅡ)靶点的活性物质进行筛选,阐明其药效物质基础。方法采用以药效团为基础的虚拟筛选手段,通过收集已出版文献中报道的具有调控CaMKⅡ作用的化合物信息,建立基于CaMKⅡ配体的HipHop药效团模型,收集文献中报道的“远志-酸枣仁”中的成分并建立化合物库,对“远志-酸枣仁”中的成分与药效团进行匹配,随后使用分子模拟对接手段对匹配到的小分子化合物与CaMKⅡ靶点(PDB ID:2V7O)进行对接,研究潜在活性单体与CaMKⅡ的作用。用戊四唑(pentylenetetrazol,PTZ)点燃模型对活性单体细叶远志苷A(39)的抗癫痫作用进行测定,并对CaMKⅡ水平的调控作用进行评价。结论通过文献检索建立包含“远志-酸枣仁”中109种化学成分的化合物库,通过测试集验证确定将药效团03用于对“远志-酸枣仁”化合物库的虚拟筛选,对匹配度排前6位的单体通过分子模拟对接分析其与CaMKⅡ的作用情况,并用抗癫痫动物模型对单体的活性进行评价,化合物39能够在中高剂量下延长PTZ致癫痫发作的潜伏时间,改善PTZ所致的焦虑抑郁样行为,改善PTZ诱导的癫痫发作后脑内尼氏体丢失,提升PTZ所致的CaMKⅡ表达水平降低,提升海马CA1、CA3区及颞叶皮层内CaMKⅡ免疫阳性细胞数量并进一步对nAChR4受体水平进行调控。表明基于虚拟筛选与活性评价手段探讨“远志-酸枣仁”中CaMKⅡ调控作用的成分具有一定的准确性。
