Loss-of-function variants of low-density lipoprotein receptor-related protein 5(LRP5)can lead to reduced bone formation,culminating in diminished bone mass.Our previous study reported transcription factor osterix(SP7)...Loss-of-function variants of low-density lipoprotein receptor-related protein 5(LRP5)can lead to reduced bone formation,culminating in diminished bone mass.Our previous study reported transcription factor osterix(SP7)-binding sites on the LRP5 promoter and its pivotal role in upregulating LRP5 expression during implant osseointegration.However,the potential role of SP7 in ameliorating LRP5-dependent osteoporosis remained unknown.In this study,we used mice with a conditional knockout(c KO)of LRP5 in mature osteoblasts,which presented decreased osteogenesis.The in vitro experimental results showed that SP7 could promote LRP5 expression,thereby upregulating the osteogenic markers such as alkaline phosphatase(ALP),Runt-related transcription factor 2(Runx2),andβ-catenin(P<0.05).For the in vivo experiment,the SP7 overexpression virus was injected into a bone defect model of LRP5 c KO mice,resulting in increased bone mineral density(BMD)(P<0.001)and volumetric density(bone volume(BV)/total volume(TV))(P<0.001),and decreased trabecular separation(Tb.Sp)(P<0.05).These data suggested that SP7 could ameliorate bone defect healing in LRP5 c KO mice.Our study provides new insights into potential therapeutic opportunities for ameliorating LRP5-dependent osteoporosis.展开更多
Sp7/Osterix as a zinc finger transcription factor is expressed specifically in osteoblasts. Embryonic lethality of Sp7 knockout mice, however, has prevented from examining the functions of Sp7 in osteoblast and bone f...Sp7/Osterix as a zinc finger transcription factor is expressed specifically in osteoblasts. Embryonic lethality of Sp7 knockout mice, however, has prevented from examining the functions of Sp7 in osteoblast and bone formation in live animals. Here we used TALEN, a versatile genome-editing tool, to generate one zebrafish sp7 mutant line. Homozygous sp7-/- mutant zebrafish are able to survive to adulthood. Alizarin Red staining and Micro-CT analysis showed that sp7-/- larvae and adult fish fail to develop normal opercula, and display curved tail fins and severe craniofacial malformation, while Alcian Blue staining showed no obvious cartilage defects in sp7-/- fish. Quantitative RT-PCR showed that a number of osteoblast markers including sppl, phex, collala, and collalb are significantly down-regulated in sp7-/- fish. Furthermore, col10a1a, whose ortholog is the cartilage marker in mice, was shown to be a novel downstream gene of Sp7 as an osteoblast marker in zebrafish. Together, these results suggest that Sp7 is required for zebrafish bone development and zebrafish sp7 mutants provide animal models for investigating novel aspects of bone development.展开更多
目的 对从混合培养的海洋软珊瑚共附生真菌中获得的结构新颖、活性良好的杂萜类成分进行研究。方法采用大米培养基对2株软珊瑚共附生曲霉属真菌EGF7-0-1和EGF15-0-3进行共培养;根据杂萜类化合物的结构特征,在基于质谱的分子网络(GNPS)...目的 对从混合培养的海洋软珊瑚共附生真菌中获得的结构新颖、活性良好的杂萜类成分进行研究。方法采用大米培养基对2株软珊瑚共附生曲霉属真菌EGF7-0-1和EGF15-0-3进行共培养;根据杂萜类化合物的结构特征,在基于质谱的分子网络(GNPS)和薄层色谱(TLC)的共同指导下,运用多种柱色谱及高效液相色谱(HPLC)等方法对其杂萜类成分进行目标导向分离;通过核磁共振(NMR)、高分辨质谱(HRMS)和旋光(ORD)等技术及物理常数对照等手段对单体化合物进行结构鉴定;采用液相色谱质谱联用(LC-MS^(n))和GNPS技术分析比较获得的杂萜类成分在2株真菌共培养及EGF7-0-1单培养中的含量。结果 从2株海洋真菌Aspergillus sp. EGF7-0-1和EGF15-0-3共培养的大米培养基中共分离得到7个杂萜类化合物,结构分别为terretonin (1)、terretonin A (2)、terretonin D1 (3)、terretonin H (4)、terreustoxin E (5)、aperterpene N (6)和asperterpene J (7)。LC-MSn和GNPS分析结果表明共培养条件下杂萜类化合物更为丰富。结论 化合物1~7均为具6/6/6骈并骨架的3,5-二甲基苔色酸(DMOA)途径衍生而成的杂萜。与菌株EGF7-0-1单培养相比,2株真菌共培养可以诱导产生丰富的杂萜,为进一步新颖杂萜类化合物的挖掘提供支持数据。展开更多
文摘Loss-of-function variants of low-density lipoprotein receptor-related protein 5(LRP5)can lead to reduced bone formation,culminating in diminished bone mass.Our previous study reported transcription factor osterix(SP7)-binding sites on the LRP5 promoter and its pivotal role in upregulating LRP5 expression during implant osseointegration.However,the potential role of SP7 in ameliorating LRP5-dependent osteoporosis remained unknown.In this study,we used mice with a conditional knockout(c KO)of LRP5 in mature osteoblasts,which presented decreased osteogenesis.The in vitro experimental results showed that SP7 could promote LRP5 expression,thereby upregulating the osteogenic markers such as alkaline phosphatase(ALP),Runt-related transcription factor 2(Runx2),andβ-catenin(P<0.05).For the in vivo experiment,the SP7 overexpression virus was injected into a bone defect model of LRP5 c KO mice,resulting in increased bone mineral density(BMD)(P<0.001)and volumetric density(bone volume(BV)/total volume(TV))(P<0.001),and decreased trabecular separation(Tb.Sp)(P<0.05).These data suggested that SP7 could ameliorate bone defect healing in LRP5 c KO mice.Our study provides new insights into potential therapeutic opportunities for ameliorating LRP5-dependent osteoporosis.
基金supported by the grants from National High Technology Research and Development Program of China (2011AA100402-2)the National Natural Science Foundation of China (NSFC) (31271356, 31030062, 81070455)+5 种基金National Basic Research Program of China (2012CB947600)the Jiangsu Distinguished Professorship Program (SR13400111)the Natural Science Foundation of Jiangsu Province (BK2012052)the Priority Academic Program Development (PAPD) of Jiangsu Higher Education Institutions (YX13400214)the High-Level Innovative Team of Jiangsu Provincethe ‘‘333” project of Jiangsu Province (BRA2015328)
文摘Sp7/Osterix as a zinc finger transcription factor is expressed specifically in osteoblasts. Embryonic lethality of Sp7 knockout mice, however, has prevented from examining the functions of Sp7 in osteoblast and bone formation in live animals. Here we used TALEN, a versatile genome-editing tool, to generate one zebrafish sp7 mutant line. Homozygous sp7-/- mutant zebrafish are able to survive to adulthood. Alizarin Red staining and Micro-CT analysis showed that sp7-/- larvae and adult fish fail to develop normal opercula, and display curved tail fins and severe craniofacial malformation, while Alcian Blue staining showed no obvious cartilage defects in sp7-/- fish. Quantitative RT-PCR showed that a number of osteoblast markers including sppl, phex, collala, and collalb are significantly down-regulated in sp7-/- fish. Furthermore, col10a1a, whose ortholog is the cartilage marker in mice, was shown to be a novel downstream gene of Sp7 as an osteoblast marker in zebrafish. Together, these results suggest that Sp7 is required for zebrafish bone development and zebrafish sp7 mutants provide animal models for investigating novel aspects of bone development.
文摘目的 对从混合培养的海洋软珊瑚共附生真菌中获得的结构新颖、活性良好的杂萜类成分进行研究。方法采用大米培养基对2株软珊瑚共附生曲霉属真菌EGF7-0-1和EGF15-0-3进行共培养;根据杂萜类化合物的结构特征,在基于质谱的分子网络(GNPS)和薄层色谱(TLC)的共同指导下,运用多种柱色谱及高效液相色谱(HPLC)等方法对其杂萜类成分进行目标导向分离;通过核磁共振(NMR)、高分辨质谱(HRMS)和旋光(ORD)等技术及物理常数对照等手段对单体化合物进行结构鉴定;采用液相色谱质谱联用(LC-MS^(n))和GNPS技术分析比较获得的杂萜类成分在2株真菌共培养及EGF7-0-1单培养中的含量。结果 从2株海洋真菌Aspergillus sp. EGF7-0-1和EGF15-0-3共培养的大米培养基中共分离得到7个杂萜类化合物,结构分别为terretonin (1)、terretonin A (2)、terretonin D1 (3)、terretonin H (4)、terreustoxin E (5)、aperterpene N (6)和asperterpene J (7)。LC-MSn和GNPS分析结果表明共培养条件下杂萜类化合物更为丰富。结论 化合物1~7均为具6/6/6骈并骨架的3,5-二甲基苔色酸(DMOA)途径衍生而成的杂萜。与菌株EGF7-0-1单培养相比,2株真菌共培养可以诱导产生丰富的杂萜,为进一步新颖杂萜类化合物的挖掘提供支持数据。
