This paper tries to analyse the linguistic feafures of AAVE with a case study of a typical and popular AAVE hip-pop"Live your life"song sang by T.I.and Rihanna.And it also briefly analyses the potential soci...This paper tries to analyse the linguistic feafures of AAVE with a case study of a typical and popular AAVE hip-pop"Live your life"song sang by T.I.and Rihanna.And it also briefly analyses the potential social reasons that may contribute to the particular linguistic features of AAVE which is different from those of Standard American English.展开更多
Retinitis pigmentosa is a group of inherited diseases that lead to retinal degeneration and photoreceptor cell death.However,there is no effective treatment for retinitis pigmentosa caused by PDE6B mutation.Adeno-asso...Retinitis pigmentosa is a group of inherited diseases that lead to retinal degeneration and photoreceptor cell death.However,there is no effective treatment for retinitis pigmentosa caused by PDE6B mutation.Adeno-associated virus(AAV)-mediated gene therapy is a promising strategy for treating retinitis pigmentosa.The aim of this study was to explore the molecular mechanisms by which AAV2-PDE6B rescues retinal function.To do this,we injected retinal degeneration 10(rd10)mice subretinally with AAV2-PDE6B and assessed the therapeutic effects on retinal function and structure using dark-and light-adapted electroretinogram,optical coherence tomography,and immunofluorescence.Data-independent acquisition-mass spectrometry-based proteomic analysis was conducted to investigate protein expression levels and pathway enrichment,and the results from this analysis were verified by real-time polymerase chain reaction and western blotting.AAV2-PDE6B injection significantly upregulated PDE6βexpression,preserved electroretinogram responses,and preserved outer nuclear layer thickness in rd10 mice.Differentially expressed proteins between wild-type and rd10 mice were closely related to visual perception,and treating rd10 mice with AAV2-PDE6B restored differentially expressed protein expression to levels similar to those seen in wild-type mice.Kyoto Encyclopedia of Genes and Genome analysis showed that the differentially expressed proteins whose expression was most significantly altered by AAV2-PDE6B injection were enriched in phototransduction pathways.Furthermore,the phototransductionrelated proteins Pde6α,Rom1,Rho,Aldh1a1,and Rbp1 exhibited opposite expression patterns in rd10 mice with or without AAV2-PDE6B treatment.Finally,Bax/Bcl-2,p-ERK/ERK,and p-c-Fos/c-Fos expression levels decreased in rd10 mice following AAV2-PDE6B treatment.Our data suggest that AAV2-PDE6B-mediated gene therapy promotes phototransduction and inhibits apoptosis by inhibiting the ERK signaling pathway and upregulating Bcl-2/Bax expression in retinitis pigmentosa.展开更多
Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images...Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images in shRNA group in Fig.3 were accidentally used rather than the final,formal experiments.To retain consistency,the entire Fig.3 is replaced here with original images of the experiments.The authors declare that this correction will not affect the conclusion of the study.展开更多
目的 构建 、鉴定自身互补双链DNA的腺相关病毒（scAAV）重组载体，使其分泌表达Exendin-4，检测其转导效率并观察在糖尿病大鼠模型中的治疗作用。 方法 应用基因工程方法改建穿梭质粒pSSHG-CMV，插入外源性基因Exendin-...目的 构建 、鉴定自身互补双链DNA的腺相关病毒（scAAV）重组载体，使其分泌表达Exendin-4，检测其转导效率并观察在糖尿病大鼠模型中的治疗作用。 方法 应用基因工程方法改建穿梭质粒pSSHG-CMV，插入外源性基因Exendin-4，构建重组scAAV载体，感染HEK293细胞，ELISA检测转染NIH3T3细胞上清Exendin-4滴度，链佐霉素诱导20只6周龄体质量180～220 g SD成年雄性大鼠为糖尿病鼠模型，逆向注射重组scAAV于糖尿病大鼠颌下腺，检测其血糖及胰岛素分泌水平。 结果 重组scpSSHG/exn4可有效包装和复制，病毒滴度为2.5×1011pfu/mL，转染细胞上清分泌Exendin-4浓度可达到4.53 ng/mL，scAAV治疗组血糖浓度在2、4周及8周均低于对照组[分别为（639.17±27.89）vs （396.00±34.00），（657.02±39.87） vs （315.62±42.56），（215.6±24.7） vs （458.6±19.7） mg/dL]，胰岛素浓度均高于对照组[分别为（156.8±24.5） vs （535.9±35.6），（236.5±12.3） vs （495.3±18.6），（620.43±46.90） vs （381.56±21.78） pg/mL]，二者比较有显著统计学差异（P〈0.05）。 结论 成功构建重组双链腺伴病毒scAAV-Ex-4，具有高效转导能力，对糖尿病大鼠模型具有控制血糖及增加胰岛素分泌作用。展开更多
常用的基因治疗表达载体有病毒表达载体和质粒表达载体,这两类传统基因治疗载体含有大量病毒或细菌DNA序列,会引发人体较严重的免疫反应、细胞炎症、细胞毒性副作用、以及基因表达沉默化,是基因治疗应用于人类疾病治疗的一大障碍。项目...常用的基因治疗表达载体有病毒表达载体和质粒表达载体,这两类传统基因治疗载体含有大量病毒或细菌DNA序列,会引发人体较严重的免疫反应、细胞炎症、细胞毒性副作用、以及基因表达沉默化,是基因治疗应用于人类疾病治疗的一大障碍。项目构建了一种新型的、基于腺相关病毒(AAV)倒置末端重复序列(ITR)的基因表达单链微载体(AAV-ITR mini vector),并用GFP基因作为报告基因。通过热变性的方法制备单链DNA,然后将带有GFP的质粒、双链DNA载体、AAV-ITR基因表达微载体转入真核表达细胞,采用荧光显微镜观察和流式细胞仪检测等较为简单的方法来检测其表达效率。实验结果显示,AAV-ITR基因表达微载体在293T细胞中具有较高的转染、表达效率,并且具有类似AAV病毒载体的特性。该研究结果将有助于进一步研发类似于AAV病毒载体的安全、无免疫原性的人造基因治疗载体。展开更多
文摘This paper tries to analyse the linguistic feafures of AAVE with a case study of a typical and popular AAVE hip-pop"Live your life"song sang by T.I.and Rihanna.And it also briefly analyses the potential social reasons that may contribute to the particular linguistic features of AAVE which is different from those of Standard American English.
基金supported by the National Natural Science Foundation of China,Nos.82071008(to BL)and 82004001(to XJ)Medical Science and Technology Program of Health Commission of Henan Province,No.LHGJ20210072(to RQ)Science and Technology Department of Henan Province,No.212102310307(to XJ)。
文摘Retinitis pigmentosa is a group of inherited diseases that lead to retinal degeneration and photoreceptor cell death.However,there is no effective treatment for retinitis pigmentosa caused by PDE6B mutation.Adeno-associated virus(AAV)-mediated gene therapy is a promising strategy for treating retinitis pigmentosa.The aim of this study was to explore the molecular mechanisms by which AAV2-PDE6B rescues retinal function.To do this,we injected retinal degeneration 10(rd10)mice subretinally with AAV2-PDE6B and assessed the therapeutic effects on retinal function and structure using dark-and light-adapted electroretinogram,optical coherence tomography,and immunofluorescence.Data-independent acquisition-mass spectrometry-based proteomic analysis was conducted to investigate protein expression levels and pathway enrichment,and the results from this analysis were verified by real-time polymerase chain reaction and western blotting.AAV2-PDE6B injection significantly upregulated PDE6βexpression,preserved electroretinogram responses,and preserved outer nuclear layer thickness in rd10 mice.Differentially expressed proteins between wild-type and rd10 mice were closely related to visual perception,and treating rd10 mice with AAV2-PDE6B restored differentially expressed protein expression to levels similar to those seen in wild-type mice.Kyoto Encyclopedia of Genes and Genome analysis showed that the differentially expressed proteins whose expression was most significantly altered by AAV2-PDE6B injection were enriched in phototransduction pathways.Furthermore,the phototransductionrelated proteins Pde6α,Rom1,Rho,Aldh1a1,and Rbp1 exhibited opposite expression patterns in rd10 mice with or without AAV2-PDE6B treatment.Finally,Bax/Bcl-2,p-ERK/ERK,and p-c-Fos/c-Fos expression levels decreased in rd10 mice following AAV2-PDE6B treatment.Our data suggest that AAV2-PDE6B-mediated gene therapy promotes phototransduction and inhibits apoptosis by inhibiting the ERK signaling pathway and upregulating Bcl-2/Bax expression in retinitis pigmentosa.
文摘Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images in shRNA group in Fig.3 were accidentally used rather than the final,formal experiments.To retain consistency,the entire Fig.3 is replaced here with original images of the experiments.The authors declare that this correction will not affect the conclusion of the study.
文摘为了建立一种快速、特异、灵敏的检测I群禽腺病毒的液相基因芯片方法。根据Hexon基因序列设计特异引物,上游引物5'端加TAG序列,下游引物5'端加生物素,进行PCR扩增,将扩增产物与链霉素亲和蛋白、磁珠37℃孵育30 min,通过Luminex200仪器分析。用所建立的液相基因芯片方法进行特异性、灵敏度、重复性及病料样品的检测。结果该方法的特异性强;检测的敏感性可达1×102copies/μL;批内批间的变异系数都在5%以下;检测结果与SYBR Green I荧光PCR方法 100%相符。表明建立的液相基因芯片方法特异性好、灵敏高、重复性好,可用于I群禽腺病毒的检测。
文摘目的 构建 、鉴定自身互补双链DNA的腺相关病毒（scAAV）重组载体，使其分泌表达Exendin-4，检测其转导效率并观察在糖尿病大鼠模型中的治疗作用。 方法 应用基因工程方法改建穿梭质粒pSSHG-CMV，插入外源性基因Exendin-4，构建重组scAAV载体，感染HEK293细胞，ELISA检测转染NIH3T3细胞上清Exendin-4滴度，链佐霉素诱导20只6周龄体质量180～220 g SD成年雄性大鼠为糖尿病鼠模型，逆向注射重组scAAV于糖尿病大鼠颌下腺，检测其血糖及胰岛素分泌水平。 结果 重组scpSSHG/exn4可有效包装和复制，病毒滴度为2.5×1011pfu/mL，转染细胞上清分泌Exendin-4浓度可达到4.53 ng/mL，scAAV治疗组血糖浓度在2、4周及8周均低于对照组[分别为（639.17±27.89）vs （396.00±34.00），（657.02±39.87） vs （315.62±42.56），（215.6±24.7） vs （458.6±19.7） mg/dL]，胰岛素浓度均高于对照组[分别为（156.8±24.5） vs （535.9±35.6），（236.5±12.3） vs （495.3±18.6），（620.43±46.90） vs （381.56±21.78） pg/mL]，二者比较有显著统计学差异（P〈0.05）。 结论 成功构建重组双链腺伴病毒scAAV-Ex-4，具有高效转导能力，对糖尿病大鼠模型具有控制血糖及增加胰岛素分泌作用。
文摘常用的基因治疗表达载体有病毒表达载体和质粒表达载体,这两类传统基因治疗载体含有大量病毒或细菌DNA序列,会引发人体较严重的免疫反应、细胞炎症、细胞毒性副作用、以及基因表达沉默化,是基因治疗应用于人类疾病治疗的一大障碍。项目构建了一种新型的、基于腺相关病毒(AAV)倒置末端重复序列(ITR)的基因表达单链微载体(AAV-ITR mini vector),并用GFP基因作为报告基因。通过热变性的方法制备单链DNA,然后将带有GFP的质粒、双链DNA载体、AAV-ITR基因表达微载体转入真核表达细胞,采用荧光显微镜观察和流式细胞仪检测等较为简单的方法来检测其表达效率。实验结果显示,AAV-ITR基因表达微载体在293T细胞中具有较高的转染、表达效率,并且具有类似AAV病毒载体的特性。该研究结果将有助于进一步研发类似于AAV病毒载体的安全、无免疫原性的人造基因治疗载体。
