Obesity-induced chronic inflammation exacerbates multiple types of tissue/organ deterioration and stem cell dysfunction;however,the effects on skeletal tissue and the underlying mechanisms are still unclear.Here,we sh...Obesity-induced chronic inflammation exacerbates multiple types of tissue/organ deterioration and stem cell dysfunction;however,the effects on skeletal tissue and the underlying mechanisms are still unclear.Here,we show that obesity triggers changes in the microRNA profile of macrophage-secreted extracellular vesicles,leading to a switch in skeletal stem/progenitor cell(SSPC)differentiation between osteoblasts and adipocytes and bone deterioration.Bone marrow macrophage(BMM)-secreted extracellular vesicles(BMM-EVs)from obese mice induced bone deterioration(decreased bone volume,bone microstructural deterioration,and increased adipocyte numbers)when administered to lean mice.Conversely,BMM-EVs from lean mice rejuvenated bone deterioration in obese recipients.We further screened the differentially expressed microRNAs in obese BMM-EVs and found that among the candidates,miR-140(with the function of promoting adipogenesis)and miR-378a(with the function of enhancing osteogenesis)coordinately determine SSPC fate of osteogenic and adipogenic differentiation by targeting the Pparα-Abca1 axis.BMM miR-140 conditional knockout mice showed resistance to obesity-induced bone deterioration,while miR-140 overexpression in SSPCs led to low bone mass and marrow adiposity in lean mice.BMM miR-378a conditional depletion in mice led to obesity-like bone deterioration.More importantly,we used an SSPC-specific targeting aptamer to precisely deliver miR-378a-3p-overloaded BMM-EVs to SSPCs via an aptamer-engineered extracellular vesicle delivery system,and this approach rescued bone deterioration in obese mice.Thus,our study reveals the critical role of BMMs in mediating obesity-induced bone deterioration by transporting selective extracellular-vesicle microRNAs into SSPCs and controlling SSPC fate.展开更多
In order to improve the accuracy for quantitating the bovine foamy virus (BFV) in vitro,we developed a baby hamster kidney cell (BHK)-21-derived indicator cell line containing a plasmid that encodes the firefly lucife...In order to improve the accuracy for quantitating the bovine foamy virus (BFV) in vitro,we developed a baby hamster kidney cell (BHK)-21-derived indicator cell line containing a plasmid that encodes the firefly luciferase driven by the BFV long terminal repeat promoter (LTR,from-7 to 1012).The BFV titer could be determined by detecting the luciferase expression since the viral trans-activator BTas protein activates the promoter activity of the LTR.One clone,designated BFVL,was selected from ten neomycin-resistant clones.BFVL showed a specific and inducible dose-and time-dependent luciferase activity in response to BFV infection.Although the changes in luciferase activity of BFVL peaked at 84 h post infection,it was possible to differentiate infected and uninfected cells at 48 h post infection.A linear relationship was established between the multiplicity of infection (MOI) of BFV and the activated ratio of luciferase expression in BFVL.Moreover,the sensitivity of the BFVL-based assay for detecting infectious BFV was 10,000 times higher than the conventional CPE-based assay at 48 h post infection.These findings suggest that the BFVL-based assay is rapid,easy,sensitive,quantitative and specific for detection of BFV infection.展开更多
In order to quantitate the bovine immunodeficiency virus line （BIVL） was established by transfecting baby hamster kidney （BIV） cells infection in vitro, a BIV indicator cell with reporter plasmids containing the f...In order to quantitate the bovine immunodeficiency virus line （BIVL） was established by transfecting baby hamster kidney （BIV） cells infection in vitro, a BIV indicator cell with reporter plasmids containing the firefly luciferase gene driven by a BIV long terminal repeat promoter. The BIV activates promoter activity of the LTR to express luciferase upon infection. BIV infection could therefore by quantified by detection of luciferase activity. Compared to standard assays used to detect BIV infection, the BIVL-based assay is 10 times more sensitive than the the CPE-based assay, and has similar sensitivity with the viral capsid protein Western blot assay BIV indicator cell line could detect BIV infection specifically. Luciferase activity of BIV infected BIVL cells showed a time dependent manner, and 60 h post infection is the optimal time to detect BIV infection. Luciferase activity of BIVL cells correlates with the BIV capsid protein expression. Moreover, a linear relationship was found between MOI and the activated intensity of luciferase expression. In brief, the BIV indicator cell line is an easy, robust and quantitive method for monitoring BIV infection.展开更多
基金supported by the National Key R&D Program of China(Grant 2022YFC3601900 to G.H.L.,2022YFC3601903,and 2022YFC3601905 to C.J.L.,2019YFA0111900 to Y.J.)the National Natural Science Foundation of China(Grant Nos.82261160397,82272560,81922017 to C.J.L.)+4 种基金the Hunan Provincial Science and Technology Department(2023JJ30896 to C.J.L.,2023JJ40965 to L.L.)the Key Research and Development Program of Hunan Province(2022SK2023 to C.J.L.)Science and Technology Innovation Program of Hunan Province(2023RC1027 to C.J.L.,2022RC1009 to J.W.,and 2022RC3075 to C.Z.)The NSFC/RGC Joint Research Scheme,the Research Grants Council(UGC)of the Hong Kong Special Administrative Region and the National Natural Science Foundation of China(NSFC/RGC Project No.N_CUHK483/22 to Y.J.)the Center for Neuromusculoskeletal Restorative Medicine[CNRM at InnoHK,to Y.J.]by Innovation and Technology Commission(ITC)of Hong Kong SAR,China.
文摘Obesity-induced chronic inflammation exacerbates multiple types of tissue/organ deterioration and stem cell dysfunction;however,the effects on skeletal tissue and the underlying mechanisms are still unclear.Here,we show that obesity triggers changes in the microRNA profile of macrophage-secreted extracellular vesicles,leading to a switch in skeletal stem/progenitor cell(SSPC)differentiation between osteoblasts and adipocytes and bone deterioration.Bone marrow macrophage(BMM)-secreted extracellular vesicles(BMM-EVs)from obese mice induced bone deterioration(decreased bone volume,bone microstructural deterioration,and increased adipocyte numbers)when administered to lean mice.Conversely,BMM-EVs from lean mice rejuvenated bone deterioration in obese recipients.We further screened the differentially expressed microRNAs in obese BMM-EVs and found that among the candidates,miR-140(with the function of promoting adipogenesis)and miR-378a(with the function of enhancing osteogenesis)coordinately determine SSPC fate of osteogenic and adipogenic differentiation by targeting the Pparα-Abca1 axis.BMM miR-140 conditional knockout mice showed resistance to obesity-induced bone deterioration,while miR-140 overexpression in SSPCs led to low bone mass and marrow adiposity in lean mice.BMM miR-378a conditional depletion in mice led to obesity-like bone deterioration.More importantly,we used an SSPC-specific targeting aptamer to precisely deliver miR-378a-3p-overloaded BMM-EVs to SSPCs via an aptamer-engineered extracellular vesicle delivery system,and this approach rescued bone deterioration in obese mice.Thus,our study reveals the critical role of BMMs in mediating obesity-induced bone deterioration by transporting selective extracellular-vesicle microRNAs into SSPCs and controlling SSPC fate.
基金National Natural Science Foundation of China(31070135,81071343)
文摘In order to improve the accuracy for quantitating the bovine foamy virus (BFV) in vitro,we developed a baby hamster kidney cell (BHK)-21-derived indicator cell line containing a plasmid that encodes the firefly luciferase driven by the BFV long terminal repeat promoter (LTR,from-7 to 1012).The BFV titer could be determined by detecting the luciferase expression since the viral trans-activator BTas protein activates the promoter activity of the LTR.One clone,designated BFVL,was selected from ten neomycin-resistant clones.BFVL showed a specific and inducible dose-and time-dependent luciferase activity in response to BFV infection.Although the changes in luciferase activity of BFVL peaked at 84 h post infection,it was possible to differentiate infected and uninfected cells at 48 h post infection.A linear relationship was established between the multiplicity of infection (MOI) of BFV and the activated ratio of luciferase expression in BFVL.Moreover,the sensitivity of the BFVL-based assay for detecting infectious BFV was 10,000 times higher than the conventional CPE-based assay at 48 h post infection.These findings suggest that the BFVL-based assay is rapid,easy,sensitive,quantitative and specific for detection of BFV infection.
基金The General Foundation of Tianjin Science Committee for Applied Basic Research (08JCZDJC21000)Chinese Ministry of Education (30770081)
文摘In order to quantitate the bovine immunodeficiency virus line （BIVL） was established by transfecting baby hamster kidney （BIV） cells infection in vitro, a BIV indicator cell with reporter plasmids containing the firefly luciferase gene driven by a BIV long terminal repeat promoter. The BIV activates promoter activity of the LTR to express luciferase upon infection. BIV infection could therefore by quantified by detection of luciferase activity. Compared to standard assays used to detect BIV infection, the BIVL-based assay is 10 times more sensitive than the the CPE-based assay, and has similar sensitivity with the viral capsid protein Western blot assay BIV indicator cell line could detect BIV infection specifically. Luciferase activity of BIV infected BIVL cells showed a time dependent manner, and 60 h post infection is the optimal time to detect BIV infection. Luciferase activity of BIVL cells correlates with the BIV capsid protein expression. Moreover, a linear relationship was found between MOI and the activated intensity of luciferase expression. In brief, the BIV indicator cell line is an easy, robust and quantitive method for monitoring BIV infection.
