Collagen made a tremendous impact in the field of regenerative medicine as a bioactive material.For decades,collagen has been used not only as a scaffolding material but also as an active component in regulating cells...Collagen made a tremendous impact in the field of regenerative medicine as a bioactive material.For decades,collagen has been used not only as a scaffolding material but also as an active component in regulating cells'biological behavior and phenotype.However,animal-derived collagen as a major source suffered from problems of immunogenicity,risk of viral infection,and the unclear relationship between bioactive sequence and function.Recombinant humanized collagen(rhCol)provided alternatives for regenerative medicine with more controllable risks.However,the characterization of rhCol and the interaction between rhCol and cells still need further investigation,including cell behavior and phenotype.The current study preliminarily demonstrated that recombinant humanized collagen typeⅢ(rhColⅢ)conformed to the theoretical amino acid sequence and had an advanced structure resembling bovine collagen.Furthermore,rhColⅢcould facilitate basal biological behaviors of human skin fibroblasts,such as adhesion,proliferation and migration.rhColⅢwas beneficial for some extracellular matrix-expressing cell phenotypes.The study would shed light on the mechanism research of rhCol and cell interactions and further understanding of effectiveness in tissue regeneration.展开更多
AIM： To identify the role of herbal compound 861 （Cpd 861） in the regulation of mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells （HSCs）. METHODS： mRNA levels o...AIM： To identify the role of herbal compound 861 （Cpd 861） in the regulation of mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells （HSCs）. METHODS： mRNA levels of collagen types I and III, matrix metalloproteinase 1 （MMP-1）, matrix metalloproteinase 2 （MMP-2）, membrane type-1 matrix metalloproteinase （MT1-MMP）, tissue inhibitor of metalloproteinase 1 （TIMP-1）, and transforming growth factor β1 （TGF-β1） in cultured-activated HSCs treated with Cpd 861 or interferon-γ, （IFN-γ,） were determined by real-time PCR. RESULTS： Both Cpd 861 and IFN-γ reduced the mRNA levels of collagen type Ⅲ, MMP-2 and TGF-β1. Moreover, Cpd 861 significantly enhanced the MMP-1 mRNA levels while down-regulated the TIMP-1 mRNA expression, increasing the ratio of MMP-1 to TIMP-1 to （6.3 ＋ 0.3）- fold compared to the control group. CONCLUSION： The anti-fibrosis function of Cpd 861 may be mediated by both decreased interstitial collagen sythesis by inhibiting the transcription of collagen type Ⅲ and TGF-β1 and increased degradation of these collagens by up-regulating MMP-1 and down-regulating TIMP-1 mRNA levels.展开更多
Recently,evidence has suggested that chronic endometritis(CE)is a crucial factor associated with infertility and failure of assisted reproductive techniques,prompting concern in the reproductive field.Studies have sho...Recently,evidence has suggested that chronic endometritis(CE)is a crucial factor associated with infertility and failure of assisted reproductive techniques,prompting concern in the reproductive field.Studies have shown that persistent infiltered immune cells stimulation result in the disturbance of endometrial immune microenvironment could lead to the infertility of CE patients finally.Conventional treatments are limited because they lack immune regulation,so it is urgent to develop a novel approach to treat CE and promote embryo implantation in patients with CE.Herein,we prepared recombinant humanized type III collagen(rhCol III)with high cell adhesion activity to regulate macrophages and repair the endometrium.In this study,M1 macrophages and M1 macrophages cultured medium and lipopolysaccharide(LPS)co-stimulated inflammatory endometrium stromal cells(ESCs)were established in vitro to mimic CE condition.rhCol III promoted M1 macrophages toward M2 phenotype,improved cell migration,viability and collagen components of inflammatory ESCs.Also,the inflammatory response of inflammatory ESCs was downregulated after rhCol III treatment.Subsequently,LPS was used for CE rat model and a 28-day observation was performed;inflammatory cells’infiltration,endometrium repair,extracellular matrix(ECM)remodeling and pregnancy outcomes were promoted after rhCol III endometrial infusion.In conclusion,rhCol III promoted(i)macrophage polarization toward M2 macrophages,(ii)pro-inflammatory cytokine production and anti-inflammatory cytokine reduction,(iii)ECM remodeling and(iv)fertility restoration.Meanwhile,rhCol III enhanced cell biological functions by interacting with discoidin domain receptors,regulated cell metabolism and reduced the inflammatory response through the inhibition of the NF-κB/YAP signaling pathway.Overall,the results illustrated the potential therapeutic prospects of rhCol III for CE treatment.展开更多
The application of medical devices to repair skin damage is clinically accepted and natural polymer enjoys an important role in this field,such as collagen or hyaluronic acid,etc.However,the biosafety and efficacy of ...The application of medical devices to repair skin damage is clinically accepted and natural polymer enjoys an important role in this field,such as collagen or hyaluronic acid,etc.However,the biosafety and efficacy of these implants are still challenged.In this study,a skin damage animal model was prepared by UV-photoaging and recombinant humanized type Ⅲ collagen(rhCol Ⅲ)was applied as a bioactive material to implant in vivo to study its biological effect,comparing with saline and uncrosslinked hyaluronic acid(HA).Animal skin conditions were non-invasively and dynamically monitored during the 8 weeks experiment.Histological observation,specific gene expression and other molecular biological methods were applied by the end of the animal experiment.The results indicated that rhCol Ⅲ could alleviate the skin photoaging caused by UV radiation,including reduce the thickening of epidermis and dermis,increase the secretion of Collagen Ⅰ(Col Ⅰ)and Collagen Ⅲ(Col Ⅲ)and remodel of extracellular matrix(ECM).Although the cell-material interaction and mechanism need more investigation,the effect of rhCol Ⅲ on damaged skin was discussed from influence on cells,reconstruction of ECM,and stimulus of small biological molecules based on current results.In conclusion,our findings provided rigorous biosafety information of rhCol Ⅲ and approved its potential in skin repair and regeneration.Although enormous efforts still need to be made to achieve successful translation from bench to clinic,the recombinant humanized collagen showed superiorities from both safety and efficacy aspects.展开更多
通过基因工程法优化人胶原蛋白的密码子序列,以人Ⅲ型胶原蛋白α1链为模板,(Gly-X-Y)为最小研究单位,构建重组质粒pET30a(+)-3CH,利用Escherichia coli发酵和蛋白纯化技术制备重组人Ⅲ型胶原蛋白(Recombinant human type Ⅲ collagen,RH...通过基因工程法优化人胶原蛋白的密码子序列,以人Ⅲ型胶原蛋白α1链为模板,(Gly-X-Y)为最小研究单位,构建重组质粒pET30a(+)-3CH,利用Escherichia coli发酵和蛋白纯化技术制备重组人Ⅲ型胶原蛋白(Recombinant human type Ⅲ collagen,RHCⅢ).研究结果证实,RHCⅢ的表达量为3.5 g/L,利用亲和/离子交换层析工艺纯化后的RHCⅢ纯度为95.44%,RHCⅢ蛋白溶液在4℃下可以保存12个月,稳定性良好,无细胞毒性,为重组人胶原蛋白产业化奠定了良好基础.展开更多
基金supported by the National Key Research and Development Program of China(2018YFC1106200 and 2018YFC1106203)the National Natural Science Foundation of China(32071330).
文摘Collagen made a tremendous impact in the field of regenerative medicine as a bioactive material.For decades,collagen has been used not only as a scaffolding material but also as an active component in regulating cells'biological behavior and phenotype.However,animal-derived collagen as a major source suffered from problems of immunogenicity,risk of viral infection,and the unclear relationship between bioactive sequence and function.Recombinant humanized collagen(rhCol)provided alternatives for regenerative medicine with more controllable risks.However,the characterization of rhCol and the interaction between rhCol and cells still need further investigation,including cell behavior and phenotype.The current study preliminarily demonstrated that recombinant humanized collagen typeⅢ(rhColⅢ)conformed to the theoretical amino acid sequence and had an advanced structure resembling bovine collagen.Furthermore,rhColⅢcould facilitate basal biological behaviors of human skin fibroblasts,such as adhesion,proliferation and migration.rhColⅢwas beneficial for some extracellular matrix-expressing cell phenotypes.The study would shed light on the mechanism research of rhCol and cell interactions and further understanding of effectiveness in tissue regeneration.
文摘AIM： To identify the role of herbal compound 861 （Cpd 861） in the regulation of mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells （HSCs）. METHODS： mRNA levels of collagen types I and III, matrix metalloproteinase 1 （MMP-1）, matrix metalloproteinase 2 （MMP-2）, membrane type-1 matrix metalloproteinase （MT1-MMP）, tissue inhibitor of metalloproteinase 1 （TIMP-1）, and transforming growth factor β1 （TGF-β1） in cultured-activated HSCs treated with Cpd 861 or interferon-γ, （IFN-γ,） were determined by real-time PCR. RESULTS： Both Cpd 861 and IFN-γ reduced the mRNA levels of collagen type Ⅲ, MMP-2 and TGF-β1. Moreover, Cpd 861 significantly enhanced the MMP-1 mRNA levels while down-regulated the TIMP-1 mRNA expression, increasing the ratio of MMP-1 to TIMP-1 to （6.3 ＋ 0.3）- fold compared to the control group. CONCLUSION： The anti-fibrosis function of Cpd 861 may be mediated by both decreased interstitial collagen sythesis by inhibiting the transcription of collagen type Ⅲ and TGF-β1 and increased degradation of these collagens by up-regulating MMP-1 and down-regulating TIMP-1 mRNA levels.
基金supported by Scientific and Technological Research Program of Chongqing Municipal Education Commission(Grant No.KJCXZD2020017).
文摘Recently,evidence has suggested that chronic endometritis(CE)is a crucial factor associated with infertility and failure of assisted reproductive techniques,prompting concern in the reproductive field.Studies have shown that persistent infiltered immune cells stimulation result in the disturbance of endometrial immune microenvironment could lead to the infertility of CE patients finally.Conventional treatments are limited because they lack immune regulation,so it is urgent to develop a novel approach to treat CE and promote embryo implantation in patients with CE.Herein,we prepared recombinant humanized type III collagen(rhCol III)with high cell adhesion activity to regulate macrophages and repair the endometrium.In this study,M1 macrophages and M1 macrophages cultured medium and lipopolysaccharide(LPS)co-stimulated inflammatory endometrium stromal cells(ESCs)were established in vitro to mimic CE condition.rhCol III promoted M1 macrophages toward M2 phenotype,improved cell migration,viability and collagen components of inflammatory ESCs.Also,the inflammatory response of inflammatory ESCs was downregulated after rhCol III treatment.Subsequently,LPS was used for CE rat model and a 28-day observation was performed;inflammatory cells’infiltration,endometrium repair,extracellular matrix(ECM)remodeling and pregnancy outcomes were promoted after rhCol III endometrial infusion.In conclusion,rhCol III promoted(i)macrophage polarization toward M2 macrophages,(ii)pro-inflammatory cytokine production and anti-inflammatory cytokine reduction,(iii)ECM remodeling and(iv)fertility restoration.Meanwhile,rhCol III enhanced cell biological functions by interacting with discoidin domain receptors,regulated cell metabolism and reduced the inflammatory response through the inhibition of the NF-κB/YAP signaling pathway.Overall,the results illustrated the potential therapeutic prospects of rhCol III for CE treatment.
基金financially supported by the National Key Research and Development Program of China(2018YFC1106200 and 2018YFC1106203)the National Natural Science Foundation of China(32071330).
文摘The application of medical devices to repair skin damage is clinically accepted and natural polymer enjoys an important role in this field,such as collagen or hyaluronic acid,etc.However,the biosafety and efficacy of these implants are still challenged.In this study,a skin damage animal model was prepared by UV-photoaging and recombinant humanized type Ⅲ collagen(rhCol Ⅲ)was applied as a bioactive material to implant in vivo to study its biological effect,comparing with saline and uncrosslinked hyaluronic acid(HA).Animal skin conditions were non-invasively and dynamically monitored during the 8 weeks experiment.Histological observation,specific gene expression and other molecular biological methods were applied by the end of the animal experiment.The results indicated that rhCol Ⅲ could alleviate the skin photoaging caused by UV radiation,including reduce the thickening of epidermis and dermis,increase the secretion of Collagen Ⅰ(Col Ⅰ)and Collagen Ⅲ(Col Ⅲ)and remodel of extracellular matrix(ECM).Although the cell-material interaction and mechanism need more investigation,the effect of rhCol Ⅲ on damaged skin was discussed from influence on cells,reconstruction of ECM,and stimulus of small biological molecules based on current results.In conclusion,our findings provided rigorous biosafety information of rhCol Ⅲ and approved its potential in skin repair and regeneration.Although enormous efforts still need to be made to achieve successful translation from bench to clinic,the recombinant humanized collagen showed superiorities from both safety and efficacy aspects.
文摘通过基因工程法优化人胶原蛋白的密码子序列,以人Ⅲ型胶原蛋白α1链为模板,(Gly-X-Y)为最小研究单位,构建重组质粒pET30a(+)-3CH,利用Escherichia coli发酵和蛋白纯化技术制备重组人Ⅲ型胶原蛋白(Recombinant human type Ⅲ collagen,RHCⅢ).研究结果证实,RHCⅢ的表达量为3.5 g/L,利用亲和/离子交换层析工艺纯化后的RHCⅢ纯度为95.44%,RHCⅢ蛋白溶液在4℃下可以保存12个月,稳定性良好,无细胞毒性,为重组人胶原蛋白产业化奠定了良好基础.
