BACKGROUND Voltage-gated sodium channels(VGSCs,or Navs)are highly expressed in various tumors and play a critical role in tumor metastasis and invasion.AIM To identify Nav1.6-associated cancer genes through bioinforma...BACKGROUND Voltage-gated sodium channels(VGSCs,or Navs)are highly expressed in various tumors and play a critical role in tumor metastasis and invasion.AIM To identify Nav1.6-associated cancer genes through bioinformatics analysis and experimental validation,with the goal of determining the role of Nav1.6 in colorectal cancer(CRC)metastasis.METHODS The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)data were analyzed using weighted correlation network analysis(WGCNA)and Venn analysis to identify Nav1.6-associated genes in CRC.siRNA,real-time PCR,and western blotting were employed to validate the Nav1.6-associated cancer genes and signaling pathways identified in CRC.Cell counting kit-8 and Transwell migration assays were used to assess the proliferation and migration of CRC cells.RESULTS The analysis of TCGA and GEO datasets,along with WGCNA,identified 575 differentially expressed genes associated with SCN8A(Nav1.6)in CRC,which were particularly enriched in MAPK signaling pathways.Tissue microarray analysis of surgical samples revealed elevated Nav1.6 levels in CRC tissues,which were predominantly in the cytoplasm and nucleus rather than in the membrane.Cytoplasmic Nav1.6 expression increased with T stage increases,consistent with the TCGA findings.SCN8A knockdown in colon tumor cells significantly reduced cell proliferation and invasion and downregulated key proteins in the RAF-MAPK pathway.CONCLUSION These findings suggest that Nav1.6 promotes CRC cell proliferation and invasion which is related to the MAPK signaling pathway.展开更多
目的:基于电压门控钠离子通道NAV1.7探究运气方苁蓉牛膝汤对膝骨关节炎(knee osteoarthritis,KOA)慢性疼痛的影响及作用机制。方法:将18只8周龄C57BL/6J雄性小鼠随机分为3组,每组6只。模型组和苁蓉牛膝汤组采用内侧半月板失稳术于小鼠...目的:基于电压门控钠离子通道NAV1.7探究运气方苁蓉牛膝汤对膝骨关节炎(knee osteoarthritis,KOA)慢性疼痛的影响及作用机制。方法:将18只8周龄C57BL/6J雄性小鼠随机分为3组,每组6只。模型组和苁蓉牛膝汤组采用内侧半月板失稳术于小鼠右后肢构建KOA模型,假手术组暴露关节腔后缝合皮肤。造模手术后第2天起,苁蓉牛膝汤组按0.02 mL·g^(-1)以苁蓉牛膝汤浓缩液(生药浓度1 g·mL^(-1))灌胃,模型组和假手术组以等量生理盐水灌胃,每天1次,连续灌胃12周。造模后6周、8周、10周、12周分别采用Von Frey测痛纤维丝和智能热板仪测定小鼠机械性疼痛阈值和热敏性疼痛阈值。造模后12周药物干预结束后,取小鼠右侧膝关节和L_(4)~L_(6)右侧背根神经节,采用Micro-CT进行膝关节骨微结构检测;采用阿尔辛蓝-苏木素-橙黄G染色进行膝关节软骨组织病理学观察,并采用国际骨关节炎研究学会(Osteoarthritis Research Society International,OARSI)评分系统对膝关节软骨病理变化进行评价;采用免疫组织化学染色检测膝关节软骨中Ⅱ型胶原、基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)、白细胞介素(interleukin,IL)-1β、IL^(-1)8表达情况;采用免疫荧光技术检测L_(4)~L_(6)背根神经节中NAV1.7和c-fos表达情况及膝关节软骨中NAV1.7表达情况。结果:①机械性疼痛阈值测定结果。模型组机械性疼痛阈值随时间变化不明显(F=1.764,P=0.186),假手术组和苁蓉牛膝汤组机械性疼痛阈值随时间变化均呈先升高后降低的趋势(F=6.764,P=0.002;F=4.188,P=0.019)。造模后6周、8周、10周、12周,模型组的机械性疼痛阈值均低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.042;P=0.002,P=0.009;P=0.000,P=0.000;P=0.000,P=0.001)。②热敏性疼痛阈值测定结果。热敏性疼痛阈值随时间变化总体呈先升高后降低的趋势(F=14.383,P=0.000)。模型组的热敏性疼痛阈值低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.000)。③膝关节骨微结构检测结果。Micro-CT检查结果显示,假手术组膝关节结构基本正常,模型组可见明显骨赘和软骨下骨钙化,苁蓉牛膝汤组可见轻微骨赘和软骨下骨钙化。模型组的骨体积分数、骨小梁厚度均高于假手术组和苁蓉牛膝汤组(P=0.000,P=0.012;P=0.000,P=0.000),骨小梁分离度低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.027)。④膝关节软骨组织病理学观察结果。模型组的膝关节软骨OARSI评分高于假手术组和苁蓉牛膝汤组(P=0.000,P=0.004)。⑤膝关节软骨中Ⅱ型胶原、MMP-9、IL-1β、IL-18表达情况检测结果。模型组的Ⅱ型胶原阳性表达面积小于假手术组和苁蓉牛膝汤组(P=0.000,P=0.000),MMP-9、IL-1β、IL-18阳性表达面积均大于假手术组和苁蓉牛膝汤组(P=0.000,P=0.001;P=0.000,P=0.002;P=0.000,P=0.000)。⑥L_(4)~L_(6)背根神经节中NAV1.7和c-fos表达情况检测结果。模型组L_(4)~L_(6)背根神经节中NAV1.7和c-fos阳性表达面积均大于假手术组(NAV1.7:P=0.001,P=0.000,P=0.000;c-fos:P=0.001,P=0.000,P=0.000)和苁蓉牛膝汤组(NAV1.7:P=0.006,P=0.013,P=0.003;c-fos:P=0.019,P=0.018,P=0.006)。⑦膝关节软骨中NAV1.7表达情况检测结果。模型组膝关节软骨中NAV1.7阳性表达面积大于假手术组和苁蓉牛膝汤组(P=0.005,P=0.006)。结论:运气方苁蓉牛膝汤可改善KOA模型小鼠的慢性疼痛症状,其机制可能与下调NAV1.7表达水平,以及减轻炎症反应和延缓软骨退变有关。展开更多
基金Supported by Science and Technology Project of Quzhou of China,No.2021Y011Beijing Science and Technology Innovation Medical Development Foundation,No.KC2021-JX-0186-81.
文摘BACKGROUND Voltage-gated sodium channels(VGSCs,or Navs)are highly expressed in various tumors and play a critical role in tumor metastasis and invasion.AIM To identify Nav1.6-associated cancer genes through bioinformatics analysis and experimental validation,with the goal of determining the role of Nav1.6 in colorectal cancer(CRC)metastasis.METHODS The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)data were analyzed using weighted correlation network analysis(WGCNA)and Venn analysis to identify Nav1.6-associated genes in CRC.siRNA,real-time PCR,and western blotting were employed to validate the Nav1.6-associated cancer genes and signaling pathways identified in CRC.Cell counting kit-8 and Transwell migration assays were used to assess the proliferation and migration of CRC cells.RESULTS The analysis of TCGA and GEO datasets,along with WGCNA,identified 575 differentially expressed genes associated with SCN8A(Nav1.6)in CRC,which were particularly enriched in MAPK signaling pathways.Tissue microarray analysis of surgical samples revealed elevated Nav1.6 levels in CRC tissues,which were predominantly in the cytoplasm and nucleus rather than in the membrane.Cytoplasmic Nav1.6 expression increased with T stage increases,consistent with the TCGA findings.SCN8A knockdown in colon tumor cells significantly reduced cell proliferation and invasion and downregulated key proteins in the RAF-MAPK pathway.CONCLUSION These findings suggest that Nav1.6 promotes CRC cell proliferation and invasion which is related to the MAPK signaling pathway.
文摘目的:基于电压门控钠离子通道NAV1.7探究运气方苁蓉牛膝汤对膝骨关节炎(knee osteoarthritis,KOA)慢性疼痛的影响及作用机制。方法:将18只8周龄C57BL/6J雄性小鼠随机分为3组,每组6只。模型组和苁蓉牛膝汤组采用内侧半月板失稳术于小鼠右后肢构建KOA模型,假手术组暴露关节腔后缝合皮肤。造模手术后第2天起,苁蓉牛膝汤组按0.02 mL·g^(-1)以苁蓉牛膝汤浓缩液(生药浓度1 g·mL^(-1))灌胃,模型组和假手术组以等量生理盐水灌胃,每天1次,连续灌胃12周。造模后6周、8周、10周、12周分别采用Von Frey测痛纤维丝和智能热板仪测定小鼠机械性疼痛阈值和热敏性疼痛阈值。造模后12周药物干预结束后,取小鼠右侧膝关节和L_(4)~L_(6)右侧背根神经节,采用Micro-CT进行膝关节骨微结构检测;采用阿尔辛蓝-苏木素-橙黄G染色进行膝关节软骨组织病理学观察,并采用国际骨关节炎研究学会(Osteoarthritis Research Society International,OARSI)评分系统对膝关节软骨病理变化进行评价;采用免疫组织化学染色检测膝关节软骨中Ⅱ型胶原、基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)、白细胞介素(interleukin,IL)-1β、IL^(-1)8表达情况;采用免疫荧光技术检测L_(4)~L_(6)背根神经节中NAV1.7和c-fos表达情况及膝关节软骨中NAV1.7表达情况。结果:①机械性疼痛阈值测定结果。模型组机械性疼痛阈值随时间变化不明显(F=1.764,P=0.186),假手术组和苁蓉牛膝汤组机械性疼痛阈值随时间变化均呈先升高后降低的趋势(F=6.764,P=0.002;F=4.188,P=0.019)。造模后6周、8周、10周、12周,模型组的机械性疼痛阈值均低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.042;P=0.002,P=0.009;P=0.000,P=0.000;P=0.000,P=0.001)。②热敏性疼痛阈值测定结果。热敏性疼痛阈值随时间变化总体呈先升高后降低的趋势(F=14.383,P=0.000)。模型组的热敏性疼痛阈值低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.000)。③膝关节骨微结构检测结果。Micro-CT检查结果显示,假手术组膝关节结构基本正常,模型组可见明显骨赘和软骨下骨钙化,苁蓉牛膝汤组可见轻微骨赘和软骨下骨钙化。模型组的骨体积分数、骨小梁厚度均高于假手术组和苁蓉牛膝汤组(P=0.000,P=0.012;P=0.000,P=0.000),骨小梁分离度低于假手术组和苁蓉牛膝汤组(P=0.000,P=0.027)。④膝关节软骨组织病理学观察结果。模型组的膝关节软骨OARSI评分高于假手术组和苁蓉牛膝汤组(P=0.000,P=0.004)。⑤膝关节软骨中Ⅱ型胶原、MMP-9、IL-1β、IL-18表达情况检测结果。模型组的Ⅱ型胶原阳性表达面积小于假手术组和苁蓉牛膝汤组(P=0.000,P=0.000),MMP-9、IL-1β、IL-18阳性表达面积均大于假手术组和苁蓉牛膝汤组(P=0.000,P=0.001;P=0.000,P=0.002;P=0.000,P=0.000)。⑥L_(4)~L_(6)背根神经节中NAV1.7和c-fos表达情况检测结果。模型组L_(4)~L_(6)背根神经节中NAV1.7和c-fos阳性表达面积均大于假手术组(NAV1.7:P=0.001,P=0.000,P=0.000;c-fos:P=0.001,P=0.000,P=0.000)和苁蓉牛膝汤组(NAV1.7:P=0.006,P=0.013,P=0.003;c-fos:P=0.019,P=0.018,P=0.006)。⑦膝关节软骨中NAV1.7表达情况检测结果。模型组膝关节软骨中NAV1.7阳性表达面积大于假手术组和苁蓉牛膝汤组(P=0.005,P=0.006)。结论:运气方苁蓉牛膝汤可改善KOA模型小鼠的慢性疼痛症状,其机制可能与下调NAV1.7表达水平,以及减轻炎症反应和延缓软骨退变有关。
