目的探究肌醇缺乏环境下骨形成蛋白(bone morphogenetic proteins,BMP)/果蝇与秀丽隐杆线虫蛋白同源物(homologues of the drosophila protein,mothers against decapentaplegic and the caenorhabditis elegans protein,Smad)1/5/8通...目的探究肌醇缺乏环境下骨形成蛋白(bone morphogenetic proteins,BMP)/果蝇与秀丽隐杆线虫蛋白同源物(homologues of the drosophila protein,mothers against decapentaplegic and the caenorhabditis elegans protein,Smad)1/5/8通路对小鼠神经干细胞增殖的影响,进一步阐明胚胎神经发育异常的分子机制。方法选取NE-4C细胞系,分为对照组和0.005、0.01、0.05、0.1、0.5、1、5、10、50、100、150和200 mmol/L肌醇处理组;应用肌醇合成酶抑制剂碳酸锂处理细胞,分为对照组和0.75、1、1.25、1.5、1.75、2和2.5 mmol/L碳酸锂处理组;用BMP信号通路抑制剂LDN-193189进行挽回实验,分别将细胞分为对照组和0.05、0.1、0.5、1、2、4和8μmol/L LDN-193189处理组;以及对照组、碳酸锂组、无肌醇组、碳酸锂+LDN-193189组和LDN-193189组,培养24 h,3-(4,5-二甲基-2-噻唑)-2,5-二苯基四氮唑溴盐[3-(4,5-dimethylthiazol-2-yl)-2,5-di-phenyltetrazolium bromide,MTT]法检测各组细胞活性;逆转录-聚合酶链式反应(reverse transcription-polymerase chain reaction,RT-PCR)检测BMP信号通路关键基因Bmp2、Bmp4、Smad1、Smad5、Smad8以及下游靶基因Runx家族转录因子2(Runx family transcription factor 2,Runx2)的表达情况;免疫印迹法及免疫荧光检测BMP/Smad1/5/8通路关键基因的表达及定位。统计学方法采用方差分析和Dunnett's t检验。结果①0.005、0.01、0.05、0.1、0.5、1、5、10、50、100、150和200 mmol/L肌醇处理组与对照组细胞活性分别为(119.1±1.5)%、(101.7±2.6)%、(98.9±1.4)%、(101.5±1.2)%、(97.9±1.5)%、(97.5±1.0)%、(96.5±1.2)%、(97.8±1.1)%、(94.4±1.2)%、(92.9±1.9)%、(82.9±1.5)%、(63.6±1.7)%与(147.8±2.4)%,随着培养基中肌醇浓度增加,细胞增殖能力逐渐降低(P值均<0.01)。1、1.25、1.5、1.75、2和2.5 mmol/L碳酸锂处理组与对照组细胞活性分别为(119.8±1.6)%、(128.1±1.8)%、(138.5±2.2)%、(114.4±2.3)%、(111.0±1.9)%、(106.3±1.7)%与(99.5±1.6)%,碳酸锂显著促进细胞增殖(P值均<0.05),且浓度为1.5 mmol/L时细胞增殖作用最明显。②碳酸锂组、无肌醇组与对照组Bmp2(2.17±0.26、19.81±0.92与1.00±0.13)、Bmp4(1.97±0.18、4.23±0.31与1.00±0.27)、Smad1(1.91±0.15、4.10±0.25与1.00±0.37)、Smad5(1.94±0.20、2.88±0.27与1.00±0.28)、Smad8(1.62±0.14、3.20±0.18与1.00±0.13)及Runx2(1.79±0.13、5.31±0.49与1.00±0.21)的mRNA水平比较,碳酸锂组和无肌醇组基因表达增强(P值均<0.01)。③0.05、0.1、0.5、1、2、4和8μmol/L LDN-193189处理组与对照组细胞活性分别为(96.5±0.7)%、(80.4±1.6)%、(63.8±1.1)%、(56.9±1.8)%、(29.3±2.4)%、(4.6±0.2)%、(2.6±0.2)%与(99.4±1.0)%,LDN-193189抑制细胞增殖且具有剂量依赖性(P值均<0.001)。碳酸锂组、无肌醇组、碳酸锂+LDN-193189组、无肌醇+LDN-193189组、LDN-193189组与对照组细胞活性分别为(122.1±2.0)%、(144.7±2.7)%、(82.6±2.3)%、(66.6±2.0)%、(57.3±2.1)%与(101.2±2.2)%,LDN-193189能逆转碳酸锂或无肌醇诱导的细胞增殖(P值均<0.001)。④碳酸锂组、无肌醇组、碳酸锂+LDN-193189组、LDN-193189组与对照组磷酸化Smad1/5/8相对表达量分别为1.30±0.10、1.62±0.10、0.59±0.15、0.11±0.08与1.00±0.18,表明LDN-193189能逆转肌醇缺乏环境下磷酸化Smad1/5/8蛋白表达上调(P<0.01)。⑤免疫荧光结果显示,红色荧光标记的Runx2蛋白定位于细胞核,与对照组比较,碳酸锂组和无肌醇组Runx2表达增高,LDN-193189组几乎无表达,碳酸锂+LDN-193189组表达降低,LDN-193189能逆转肌醇缺乏环境下诱导的Runx2表达上调。碳酸锂组、无肌醇组、碳酸锂+LDN-193189组、LDN-193189组与对照组Runx2蛋白相对表达量分别为2.00±0.08、2.09±0.11、1.75±0.05、1.76±0.13与1.04±0.10,表明肌醇缺乏促进Runx2表达上调(P<0.01),LDN-193189逆转碳酸锂诱导的Runx2表达上调的作用不明显,但P值接近0.05(P=0.051)。结论肌醇缺乏环境会导致BMP/Smad1/5/8通路被异常激活,调控下游靶基因Runx2,可促进神经干细胞异常增殖。展开更多
目的:观察益气温经方在老年骨质疏松性肱骨近端骨折内固定术后抗骨质疏松治疗中的应用价值。方法:2018年1—12月,采用益气温经方对49例老年骨质疏松性肱骨近端骨折内固定术后患者进行抗骨质疏松治疗。男18例,女31例。年龄62~78岁,中位...目的:观察益气温经方在老年骨质疏松性肱骨近端骨折内固定术后抗骨质疏松治疗中的应用价值。方法:2018年1—12月,采用益气温经方对49例老年骨质疏松性肱骨近端骨折内固定术后患者进行抗骨质疏松治疗。男18例,女31例。年龄62~78岁,中位数70岁。左侧26例,右侧23例。中医辨证为肾阳虚证。骨折AO分型,A型18例、B型31例。骨质疏松症病程1~2年,中位数2年。采用双能X线吸收法测量L 1~L 4椎体骨密度,采用酶联免疫吸附法测定血清Ⅰ型前胶原氨基端前肽(N-terminal propeptide of typeⅠprecollagen,PⅠNP)及Ⅰ型胶原羧基端交联端肽(C-terminal cross-linked telopeptide of typeⅠcollagen,CTX-Ⅰ)含量,采用Neer肩关节评分标准评价综合疗效,观察不良反应发生情况。结果:所有患者均获随访,随访时间12~14个月,中位数13个月。骨密度,术前(0.67±0.04)g·cm^-2、术后6个月(0.70±0.12)g·cm^-2、术后12个月(0.71±0.04)g·cm^-2。血清PⅠNP,术前(38.98±24.17)ng·mL-1、术后6个月(45.20±20.26)ng·mL-1、术后12个月(49.47±30.48)ng·mL-1。血清CTX-Ⅰ,术前(12.35±0.96)ng·mL-1、术后6个月(10.76±0.16)ng·mL-1、术后12个月(9.47±0.48)ng·mL-1。术后12个月Neer肩关节评分(83.3±12.1)分,优20例、良20例、可9例。治疗期间均未出现明显不良反应。结论:益气温经方能够提高老年骨质疏松性肱骨近端骨折内固定术后患者的骨密度、改善骨代谢指标,综合疗效好、安全性高,值得临床推广应用。展开更多
N-phosphoryl-α-amino acids can react to form esters, peptides, phosphoryl ester-exchanged products and phosphoryl group migration products under mild conditions, which are very important in researches on arohebiont. ...N-phosphoryl-α-amino acids can react to form esters, peptides, phosphoryl ester-exchanged products and phosphoryl group migration products under mild conditions, which are very important in researches on arohebiont. It has been proposed that intramolecular mixed carboxylic-phosphoric anhydrides (IMCPAs) might be the common intermediates of the reactions, which are difficult to be experimentally observed because of their high activity. In this note, it has been revealed, by means of MNDO, that the IMCPAs are situated at the relatively minimal points of the potential surface and can form in a non-synchronous concerted reaction pathway that passes through a transition state with a hydrogen-bond-bridge structure. The results give a successful explanation to the structure and formation of IMCPAs and their ability to promote the various reactions mentioned above.展开更多
文摘目的:观察益气温经方在老年骨质疏松性肱骨近端骨折内固定术后抗骨质疏松治疗中的应用价值。方法:2018年1—12月,采用益气温经方对49例老年骨质疏松性肱骨近端骨折内固定术后患者进行抗骨质疏松治疗。男18例,女31例。年龄62~78岁,中位数70岁。左侧26例,右侧23例。中医辨证为肾阳虚证。骨折AO分型,A型18例、B型31例。骨质疏松症病程1~2年,中位数2年。采用双能X线吸收法测量L 1~L 4椎体骨密度,采用酶联免疫吸附法测定血清Ⅰ型前胶原氨基端前肽(N-terminal propeptide of typeⅠprecollagen,PⅠNP)及Ⅰ型胶原羧基端交联端肽(C-terminal cross-linked telopeptide of typeⅠcollagen,CTX-Ⅰ)含量,采用Neer肩关节评分标准评价综合疗效,观察不良反应发生情况。结果:所有患者均获随访,随访时间12~14个月,中位数13个月。骨密度,术前(0.67±0.04)g·cm^-2、术后6个月(0.70±0.12)g·cm^-2、术后12个月(0.71±0.04)g·cm^-2。血清PⅠNP,术前(38.98±24.17)ng·mL-1、术后6个月(45.20±20.26)ng·mL-1、术后12个月(49.47±30.48)ng·mL-1。血清CTX-Ⅰ,术前(12.35±0.96)ng·mL-1、术后6个月(10.76±0.16)ng·mL-1、术后12个月(9.47±0.48)ng·mL-1。术后12个月Neer肩关节评分(83.3±12.1)分,优20例、良20例、可9例。治疗期间均未出现明显不良反应。结论:益气温经方能够提高老年骨质疏松性肱骨近端骨折内固定术后患者的骨密度、改善骨代谢指标,综合疗效好、安全性高,值得临床推广应用。
文摘N-phosphoryl-α-amino acids can react to form esters, peptides, phosphoryl ester-exchanged products and phosphoryl group migration products under mild conditions, which are very important in researches on arohebiont. It has been proposed that intramolecular mixed carboxylic-phosphoric anhydrides (IMCPAs) might be the common intermediates of the reactions, which are difficult to be experimentally observed because of their high activity. In this note, it has been revealed, by means of MNDO, that the IMCPAs are situated at the relatively minimal points of the potential surface and can form in a non-synchronous concerted reaction pathway that passes through a transition state with a hydrogen-bond-bridge structure. The results give a successful explanation to the structure and formation of IMCPAs and their ability to promote the various reactions mentioned above.