目的研究乳铁蛋白对原代大鼠成骨细胞胰岛素生长因子结合蛋白-3(IGFBP-3)、胰岛素生长因子结合蛋白-4(IGFBP-4)、胰岛素生长因子结合蛋白-5(IGFBP-5)m RNA表达的影响。方法用混合酶消化法分离大鼠颅骨成骨细胞,进行原代培养。细胞以6...目的研究乳铁蛋白对原代大鼠成骨细胞胰岛素生长因子结合蛋白-3(IGFBP-3)、胰岛素生长因子结合蛋白-4(IGFBP-4)、胰岛素生长因子结合蛋白-5(IGFBP-5)m RNA表达的影响。方法用混合酶消化法分离大鼠颅骨成骨细胞,进行原代培养。细胞以6×103/cm2接种于六孔板内,用不同浓度乳铁蛋白即0μg/ml(对照组)、0.1μg/ml(乳铁蛋白1组)、1μg/ml(乳铁蛋白2组)、10μg/ml(乳铁蛋白3组)、100μg/ml(乳铁蛋白4组)、1000μg/ml(乳铁蛋白5组)干预原代大鼠成骨细胞1、3、5、7 d后提取总RNA,采用荧光定量PCR技术分析IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达。结果与对照组比较,各浓度组及时间段对IGFBP-3 m RNA表达的影响不稳定,时高时低。与对照组比较,除了乳铁蛋白1组外,其他浓度对IGFBP-4 m RNA的表达呈浓度梯度的抑制(P<0.01)。与对照组比较,实验干预第1天,呈浓度梯度抑制IGFBP-5 m RNA的表达(P<0.01),乳铁蛋白4、5组在第5、7天表现为继续抑制(P<0.01),而其他时间及浓度对IGFBP-5 m RNA的表达调控不稳定。结论乳铁蛋白影响原代大鼠成骨细胞IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达,其中乳铁蛋白对IGFBP-3、IGFBP-5 m RNA表达的调控不稳定,而乳铁蛋白抑制IGFBP-4 m RNA的表达呈浓度依赖性,浓度越高抑制作用越强。展开更多
Objective： To construct a recombinant adenovirus vector-carrying human growth and differentiation factor-5 （GDF-5） gene, investigate the biological effects of adenovirus-mediated GDF-5 （Ad-GDF-5） on extracellular...Objective： To construct a recombinant adenovirus vector-carrying human growth and differentiation factor-5 （GDF-5） gene, investigate the biological effects of adenovirus-mediated GDF-5 （Ad-GDF-5） on extracellular matrix （ECM） expression in human degenerative disc nucleus pulposus （NP） cells, and explore a candidate gene therapy method for intervertebral disc degeneration （IDD）. Methods： Human NP cells of a degenerative disc were isolated, cultured, and infected with Ad-GDF-5 using the AdEasy-1 adenovirus vector system. On Days 3, 7, 14, and 21, the contents of the sulfated glycosaminoglycan （sGAG）, deoxyribonucleic acid （DNA） and hydroxyproline （Hyp）, synthesis of proteoglycan and collagen II, gene expression of collagen II and aggrecan, and NP cell proliferation were assessed. Results： The adenovirus was an effective vehicle for gene delivery with prolonged expression of GDF-5. Biochemical analysis revealed increased sGAG and Hyp contents in human NP cells infected by Ad-GDF-5 whereas there was no conspicuous change in basal medium （BM） or Ad-green fluorescent protein （GFP） groups. Only cells in the Ad-GDF-5 group promoted the production of ECM, as demonstrated by the secretion of proteoglycan and up-regulation of collagen II and aggrecan at both protein and mRNA levels. The NP cell proliferation was significantly promoted. Conclusions： The data suggest that Ad-GDF-5 gene therapy is a potential treatment for IDD, which restores the functions of degenerative intervertebral disc through enhancing the ECM production of human NP ceils.展开更多
OBJECTIVE: To investigate the effect of Buyanghuanwu decoction(BYHWD) on gene expression in ventricular remodeling post-myocardial infarction in rats.METHODS: Animal models of myocardial infarction were established by...OBJECTIVE: To investigate the effect of Buyanghuanwu decoction(BYHWD) on gene expression in ventricular remodeling post-myocardial infarction in rats.METHODS: Animal models of myocardial infarction were established by permanent ligation of the left anterior descending coronary artery. Echocardiography measurements were performed after the treatment of BYHWD(18 g·kg-1 collagen was observ·d-1) for 90 days.Myocardialed by mallory trichrome staining. Capillary density was quantified by using Factor rentially expⅧre immunohistochemical staining.Diffessed genes were explored by a short-read sequencing technology combined with a tag-based digital gene expression profiling(DGE)system. Real-time quantitative polymerase chain reaction detecting system(q PCR) was used to validate the sequencing results. After assembling the gene information from Sham, model and BYHWD groups, we constructed three DGE libraries based on each group. The sequencing of three libraries generated 66 000-73 000 unique tags, which were mapped to reference sequences for annotation of expressed genes.RESULTS: Among them, 511 and 352 differentially expressed genes were found in comparison with sham/model and model/BYHWD, respectively. Fifty-five genes exhibited reversed direction of gene expression differences between Sham/Model and Model/BYHWD groups. We found that transforming growth factor beta receptor-1, junctophilin-2,monocyte chemotactic protein 1, neuropeptide Y,arachidonate 5-Lipoxygenase, arachidonate 15-Lipoxygenase were significantly modulated, which suggested the involvement of these genes in BYHWD treatment.CONCLUSION: The DGE profiling data provide comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the pharmacological mechanisms of BYHWD in ventricular remodeling post-myocardial infarction.展开更多
文摘目的研究乳铁蛋白对原代大鼠成骨细胞胰岛素生长因子结合蛋白-3(IGFBP-3)、胰岛素生长因子结合蛋白-4(IGFBP-4)、胰岛素生长因子结合蛋白-5(IGFBP-5)m RNA表达的影响。方法用混合酶消化法分离大鼠颅骨成骨细胞,进行原代培养。细胞以6×103/cm2接种于六孔板内,用不同浓度乳铁蛋白即0μg/ml(对照组)、0.1μg/ml(乳铁蛋白1组)、1μg/ml(乳铁蛋白2组)、10μg/ml(乳铁蛋白3组)、100μg/ml(乳铁蛋白4组)、1000μg/ml(乳铁蛋白5组)干预原代大鼠成骨细胞1、3、5、7 d后提取总RNA,采用荧光定量PCR技术分析IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达。结果与对照组比较,各浓度组及时间段对IGFBP-3 m RNA表达的影响不稳定,时高时低。与对照组比较,除了乳铁蛋白1组外,其他浓度对IGFBP-4 m RNA的表达呈浓度梯度的抑制(P<0.01)。与对照组比较,实验干预第1天,呈浓度梯度抑制IGFBP-5 m RNA的表达(P<0.01),乳铁蛋白4、5组在第5、7天表现为继续抑制(P<0.01),而其他时间及浓度对IGFBP-5 m RNA的表达调控不稳定。结论乳铁蛋白影响原代大鼠成骨细胞IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达,其中乳铁蛋白对IGFBP-3、IGFBP-5 m RNA表达的调控不稳定,而乳铁蛋白抑制IGFBP-4 m RNA的表达呈浓度依赖性,浓度越高抑制作用越强。
基金Project supported by the National Natural Science Foundation of China(Nos.81171472 and 81201407)the Innovation Team Project of Sichuan Provincial Education Department(No.13TD0030)+1 种基金the Major Transformation Cultivation Project of Sichuan Provincial Education Department(No.15CZ0021)the Science and Technology Project of Nanchong City(No.14A0021),China
文摘Objective： To construct a recombinant adenovirus vector-carrying human growth and differentiation factor-5 （GDF-5） gene, investigate the biological effects of adenovirus-mediated GDF-5 （Ad-GDF-5） on extracellular matrix （ECM） expression in human degenerative disc nucleus pulposus （NP） cells, and explore a candidate gene therapy method for intervertebral disc degeneration （IDD）. Methods： Human NP cells of a degenerative disc were isolated, cultured, and infected with Ad-GDF-5 using the AdEasy-1 adenovirus vector system. On Days 3, 7, 14, and 21, the contents of the sulfated glycosaminoglycan （sGAG）, deoxyribonucleic acid （DNA） and hydroxyproline （Hyp）, synthesis of proteoglycan and collagen II, gene expression of collagen II and aggrecan, and NP cell proliferation were assessed. Results： The adenovirus was an effective vehicle for gene delivery with prolonged expression of GDF-5. Biochemical analysis revealed increased sGAG and Hyp contents in human NP cells infected by Ad-GDF-5 whereas there was no conspicuous change in basal medium （BM） or Ad-green fluorescent protein （GFP） groups. Only cells in the Ad-GDF-5 group promoted the production of ECM, as demonstrated by the secretion of proteoglycan and up-regulation of collagen II and aggrecan at both protein and mRNA levels. The NP cell proliferation was significantly promoted. Conclusions： The data suggest that Ad-GDF-5 gene therapy is a potential treatment for IDD, which restores the functions of degenerative intervertebral disc through enhancing the ECM production of human NP ceils.
基金Supported by National Natural Science Foundation of China Project:Studies of Qi-Supplementing Therapy Promoting Angiogenesis after Myocardial Infarction by Simulation of Angptl6 Pathway in NK Cells(No.81302892)Effects of Polyamine-derived Aldehyde Load Injury in Long-term Prognosis after Myocardial Infarction Ventricular Remodeling,and the Modulation Mechanism of Buyanghuanwu Decoction(No.81173459)+4 种基金Studies of the Role of 5-LOX/Cys LT2R in Left Ventricular Remodeling after Myocardial Infarction and the Pharmacological Mechanisms of BYHWD(No.81373575)Effects of Hsp20 on Ventricular Remodeling after Myocardial Infarction,and the Modulation Mechanism of Buyanghuanwu decoction(No.81202841)Guangdong Natural Science Foundation Project:Studies of Qi-Supplementing Therapy Promoting Angiogenesis after Myocardial Infarction by Simulation of Angptl6 Pathway in NK Cells(No.S2013040016226)Studies of the role of 5-LOX/Cys LT2R in Left Ventricular Remodeling after Myocardial Infarction and the Pharmacological Mechanisms of BYHWD(No.S2013010014777)Specialized Research Fund for the Doctoral Program of Higher Education Project:Effects of Polyamine-derived Aldehyde Load Injury in Long-term Prognosis after Myocardial Infarction Ventricular Remodeling,and the Modulation Mechanism of Buyanghuanwu decoction(No.20124433110019)
文摘OBJECTIVE: To investigate the effect of Buyanghuanwu decoction(BYHWD) on gene expression in ventricular remodeling post-myocardial infarction in rats.METHODS: Animal models of myocardial infarction were established by permanent ligation of the left anterior descending coronary artery. Echocardiography measurements were performed after the treatment of BYHWD(18 g·kg-1 collagen was observ·d-1) for 90 days.Myocardialed by mallory trichrome staining. Capillary density was quantified by using Factor rentially expⅧre immunohistochemical staining.Diffessed genes were explored by a short-read sequencing technology combined with a tag-based digital gene expression profiling(DGE)system. Real-time quantitative polymerase chain reaction detecting system(q PCR) was used to validate the sequencing results. After assembling the gene information from Sham, model and BYHWD groups, we constructed three DGE libraries based on each group. The sequencing of three libraries generated 66 000-73 000 unique tags, which were mapped to reference sequences for annotation of expressed genes.RESULTS: Among them, 511 and 352 differentially expressed genes were found in comparison with sham/model and model/BYHWD, respectively. Fifty-five genes exhibited reversed direction of gene expression differences between Sham/Model and Model/BYHWD groups. We found that transforming growth factor beta receptor-1, junctophilin-2,monocyte chemotactic protein 1, neuropeptide Y,arachidonate 5-Lipoxygenase, arachidonate 15-Lipoxygenase were significantly modulated, which suggested the involvement of these genes in BYHWD treatment.CONCLUSION: The DGE profiling data provide comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the pharmacological mechanisms of BYHWD in ventricular remodeling post-myocardial infarction.
