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Occurrence of cGMP/nitric oxide-sensitive store-operated calcium entry in fibroblasts and its effect on matrix metalloproteinase secretion

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World Journal of Gastroenterology 2006年第34期12卷 5483-5489页

作者： Yong Huang Min-Qiang Lu Hua Li Chi Xu Shu-Hong Yi Gui-Hua Chen Liver Transplantation Centre the Third Affiliated Hospital Sun Yat-Sen University Guangzhou 510630 Guangdong Province China

AIM： To examine the existence of Nitric oxide/ cGMP sensitive store-operated Ca^2＋ entry in mouse fibroblast NIH/3T3 cells and its influence on matrix metalloproteinase （MMP） production and adhesion ability of fibroblasts. METHODS： NIH/3T3 cells were cultured. Confocal laser scanning microscopy was used to examine the existence of thapsigargin-induced store-operated Ca^2＋ entry in fibroblasts. Gelatin zymography and semiquantitative reverse transcriptase-polymerase chain reaction （RTPCR） were employed to detect the involvement of [Ca^2＋]i and NO/cGMP in MMP secretion. The involvement of NO/ cGMP-sensitive Ca^2＋ entry in adhesion was determined using matrigel-coated culture plates. RESULTS： 8-bromo-cGMP inhibited the thapsigargin-induced Ca^2＋ entry in 3T3 cells. The cGMP-induced inhibition was abolished by an inhibitor of protein kinase G, KT5823 （1μmol/L）. A similar effect on the Ca^2＋ entry was observed in 3T3 cells in response to a NO donor, （±）-S-nitroso-N-acetylpenicillamine （SNAP）. The inhibitory effect of SNAP on the thapsigargin-induced Ca^2＋ entry was also observed, indicating NO/cGMP-regulated Ca^2＋ entry in 3T3 cells. Results of gelatin zymography assay showed that addition of extracellular Ca^2＋ concentration induced MMP release and activation in a dose-dependent manner. RT-PCR also showed that cGMP and SNAP reduced the production of MMP mRNA in 3T3 cells. Experiments investigating adhesion potentials demonstrated that cGMP and SNAP could upgrade 3T3 cell attachment rate to the matrigel-coated culture plates.CONCLUSION： NO/cGMP sensitive store-operated Ca^2＋ entry occurs in fibroblasts, and attenuates their adhesion potentials through its influence on MMP secretion.

关键词： cGMP Nitric oxide Protein kinase G Storeoperated Ca^2+ entry Matrix metalloproteinase fibroblast

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Bone marrow mesenchymal stem cell-derived exosomal microRNAs target PI3K/Akt signaling pathway to promote the activation of fibroblasts

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World Journal of Stem Cells 2023年第4期15卷 248-267页

作者： Fang-Qi Li Wen-Bo Chen Zhi-Wen Luo Yi-Sheng Chen Ya-Ying Sun Xiao-Ping Su Jun-Ming Sun Shi-Yi Chen Department of Sports Medicine Huashan HospitalFudan UniversityShanghai 200040China Guangxi Key Laboratory of Oral and Maxillofacial Rehabilitation and Reconstruction Guangxi Medical UniversityNanning 530021Guangxi Zhuang Autonomous RegionChina Laboratory Animal Center Guangxi Medical UniversityNanning 530021Guangxi Zhuang Autonomous RegionChina

BACKGROUND fibroblast plays a major role in tendon-bone *** derived from bone marrow mesenchymal stem cells(BMSCs)can activate fibroblasts and promote tendon-bone healing via the contained microRNAs(miRNAs).However,the underlying mechanism is not comprehensively ***,this study aimed to identify overlapped BMSC-derived exosomal miRNAs in three GSE datasets,and to verify their effects as well as mechanisms on *** To identify overlapped BMSC-derived exosomal miRNAs in three GSE datasets and verify their effects as well as mechanisms on *** BMSC-derived exosomal miRNAs data(GSE71241,GSE153752,and GSE85341)were downloaded from the Gene Expression Omnibus(GEO)*** candidate miRNAs were obtained by the intersection of three data *** was used to predict potential target genes for the candidate *** and pathway analyses were conducted using the Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)databases,respectively,by processing data with the *** interconnected genes in the protein-protein interaction(PPI)network were analyzed using Cytoscape ***,wound healing assay,collagen contraction assay and the expression of COL I andα-smooth muscle actin positive were applied to investigate the cell proliferation,migration and collagen *** real-time reverse transcription polymerase chain reaction was applied to determine the cell fibroblastic,tenogenic,and chondrogenic *** Bioinformatics analyses found two BMSC-derived exosomal miRNAs,has-miR-144-3p and hasmiR-23b-3p,were overlapped in three GSE *** network analysis and functional enrichment analyses in the GO and KEGG databases indicated that both miRNAs regulated the PI3K/Akt signaling pathway by targeting phosphatase and tensin homolog(PTEN).In vitro experiments confirmed that miR-144-3p and miR-23b-3p stimulated proliferation,migration and collagen sy

关键词： Exosome MicroRNA fibroblast Mesenchymal stem cell Tendon-bone healing

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Multitranscriptome analyses of keloid fibroblasts reveal the role of the HIF-1α/HOXC6/ERK axis in keloid development

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Burns & Trauma 2022年第1期10卷 529-545页

作者： Qi Wang Yixiu Zhong Zhijia Li Dingheng Zhu Hongyan Lu Pingjiao Chen Changxing Li Xuebiao Peng Qian Li Kang Zeng Department of Dermatology Nanfang HospitalSouthern Medical UniversityGuangzhou 510515China Molecular Diagnosis and Treatment Center for Infectious Diseases Dermatology HospitalSouthern Medical UniversityGuangzhou 510091China Department of Dermatologic Surgery Dermatology HospitalSouthern Medical UniversityGuangzhou 510091China

Background:A keloid is a disease of excessive fibrosis that is characterized by the aberrant prolifer-ation of ***,the molecular mechanisms of fibroblasts during the development of keloids remain *** study aims to identify new molecular targets that promote the proliferation and migration of keloid fibroblasts,providing new ideas for the prevention and treatment of ***:We utilized bioinformatics tools to analyze data from keloid fibroblasts(KFs)available in the Gene Expression Omnibus(GEO)database to identify the key genes involved in keloid *** C6(HOXC6)emerged as a hub gene in KFs from the GEO database was verified in keloid tissue samples and KFs using reverse transcription-quantitative polymerase chain reaction,western blot(WB)and ***,the effects of down-regulated HOXC6 expression on the cellular behaviors of KFs were examined by performing Cell Counting Kit-8,flow cytometry,transwell migration and WB ***,we performed transcriptome sequencing and gene set enrichment analysis(GSEA)to further explore HOXC6-related mechanisms and validated the signaling pathways by performing a series of ***:HOXC6 was the top-ranking hub gene of KFs in microarray datasets from GEO and was upregulated in keloid tissue samples and *** of HOXC6 inhibited proliferation,migration and extracellular matrix(ECM)accumulation and promoted KF *** pre-dicted that the hypoxia signaling pathway was associated with HOXC6 in *** sequencing suggested that the extracellular regulated protein kinase(ERK)pathway was one of the downstream pathways of HOXC6 in *** experiments confirmed that hypoxia-inducible factor-1α(HIF-1α)upregulates HOXC6,contributing to KFs proliferation,migration,apoptosis inhibition and collagen accumulation through the ERK signaling ***:Our findings first revealed that HOXC6 acts as an oncogenic driver in the

关键词： Keloid fibroblast HOXC6 HIF-1α ERK Transcriptome

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CD26 upregulates proliferation and invasion in keloid fibroblasts through an IGF-1-induced PI3K/AKT/mTOR pathway

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Burns & Trauma 2020年第1期8卷 41-54页

作者： Yu Xin Peiru Min Heng Xu Zheng Zhang Yan Zhang Yixin Zhang Department of Plastic and Reconstructive Surgery Shanghai 9th People’s HospitalShanghai Jiao Tong University School of Medicine639 Zhi Zao Ju RoadShanghai200011China

Background:Keloid is a fibrotic dermal disease characterized by an abnormal increase in fibroblast proliferation and *** pathological behaviours may be related to the heterogeneity of keloid fibroblasts(KFs);however,because of a lack of effective biomarkers for KFs it is difficult to study the underlying *** previous studies revealed that the expansion of CD26+KFs was responsible for increased keloid proliferation and invasion capabilities;the intrinsic relationship and mechanism between CD26 and keloid is therefore worthy of further *** aim of this studywas to explore molecular mechanisms in the process of CD26 upregulated KFs proliferation and invasion abilities,and provide more evidence for CD26 as an effective biomarker of keloid and a new clinical therapeutic ***:Flow cytometry was performed to isolate CD26+/CD26−fibroblasts from KFs and normal *** generate stably silenced KFs for CD26 and insulin-like growth factor-1 receptor(IGF-1R),lentiviral particles encoding shRNA targeting CD26 and IGF-1R were used for *** proliferations were analysed by cell counting kit-8 assay and 5-ethynyl-2-deoxyuridine(EdU)incorporation *** assay and transwell assay were used to assess cell migration and invasion *** further quantify the regulatory role of CD26 expression in the relevant signalling pathway,RT-qPCR,western blot,ELISA,PI3K activity assay and immunofluorescence were ***:Aberrant expression of CD26 in KFs was proven to be associated with increased proliferation and invasion of ***,the role of the IGF-1/IGF-1 receptor axis was also studied in CD26 and was found to upregulate KF proliferation and *** PI3K/protein kinase B(AKT)/mammalian target of rapamycin(mTOR)pathway was shown to affect CD26-regulated KF proliferation and invasion by increasing phosphorylation levels of S6 kinase and 4E-binding ***:CD26 can be the effective biomark

关键词： CD26 IGF-1 Invasion Keloids PI3K/AKT/mTOR signalling pathway Proliferation fibroblast

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Injecting skin fibroblasts into muscles to express human clotting factor IX cDNA

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Science China(Life Sciences) 1998年第2期41卷 146-150页

作者：王宏伟杨晓青邢永娜卢大儒邱信芳薛京伦 Institute of Genetics Fudan University Shanghai China

Primary rabbit skin fibroblasts (RSF) containing lacZ reporter gene or human clotting factor IX (hFIX) cDNA could long termly exist and be expressed after being implanted in rabbit muscles. Among them, the beta glactosidase expression lasted at least 3 months, and hFIX expression lasted 1 month. These results will be helpful to organically combine the advantages of both skin fibroblasts and muscle, and provide a new way for somatic gene therapy of hemophilia B and other genetic diseases.

关键词： muscle injection of fibroblast expression of human clotting factor IX.

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Regenerative material for aneurysm embolizationA 3-dimensional culture system of fibroblasts and calcium alginate gel

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Neural Regeneration Research 2011年第10期6卷 762-766页

作者： Jingdong Zhang Kan Xu Jinlu Yu Jun Wang Qi Luo Department of Neurosurgery First Hospital of Jilin University Changchun 130021 Jilin Province China

Calcium alginate gel（CAG） has been shown to successfully model aneurysm embolization within a short period of ***,gradually degrading CAG potentially results in aneurysm *** the present study,a regenerative embolic material was designed by seeding rat fibroblasts in a *** study investigated the feasibility of constructing a 3-dimensional culture *** fibroblasts grew well and firmly attached to the *** was conducive for fibroblast growth,and resulted in a 3-dimensional culture *** show that CAG can be used theoretically as a vascular,regenerative,embolic material.

关键词： intracranial aneurysm embolic material detachable coil fibroblast alginate neural regeneration

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Reprogrammed astrocytes display higher neurogenic competence, migration ability and cell death resistance than reprogrammed fibroblasts

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Translational Neurodegeneration 2020年第1期9卷 55-65页

作者： Xiaohuan Xia Chunhong Li Yi Wang Xiaobei Deng Yizhao Ma Lu Ding Jialin Zheng Center for Translational Neurodegeneration and Regenerative Therapy Shanghai Tenth People’s Hospital affiliated to Tongji University School of MedicineShanghai 200072China Collaborative Innovation Center for Brain Science Tongji UniversityShanghai 200092China Departments of Pharmacology and Experimental Neuroscience University of Nebraska Medical CenterOmahaNE 68198-5930USA Department of Pathology and Microbiology University of Nebraska Medical CenterOmahaNE 68198-5930USA

The direct reprogramming of somatic cells into induced neural progenitor cells(iNPCs)has been envisioned as a promising approach to overcome ethical and clinical issues of pluripotent stem cell *** previously reported that astrocyte-derived induced pluripotent stem cells(iPSCs)have more tendencies for neuronal differentiation than fibroblast-derived ***,the differences of neurogenic potential between astrocytederived iNPCs(AiNPCs)and iNPCs from non-neural origins,such as fibroblast-derived iNPCs(FiNPCs),and the underlying mechanisms remain *** results suggested that AiNPCs exhibited higher differentiation efficiency,mobility and survival capacities,compared to *** whole transcriptome analysis revealed higher activities of TGFβsignaling in AiNPCs,versus FiNPCs,following a similar trend between astrocytes and *** higher neurogenic competence,migration ability,and cell death resistance of AiNPCs could be abrogated using TGFβ signaling inhibitor ***,our study demonstrates the difference between iNPCs generated from neural and non-neural cells,together with the underlying mechanisms,which,provides valuable information for donor cell selection in the reprogramming approach.

关键词： Reprogramming Astrocyte fibroblast Induced neural progenitor cells TGFβsignaling Neurogenesis Proliferation Migration Survival

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Neutrophil extracellular traps contribute to myofibroblast differentiation and scar hyperplasia through the Toll-like receptor 9/nuclear factor Kappa-B/interleukin-6 pathway

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Burns & Trauma 2022年第1期10卷 116-128页

作者： Yiming Shao Zaiwen Guo Yunxi Yang Lu Liu Jiamin Huang Yi Chen Linbin Li Bingwei Sun Department of Burns and Plastic Surgery Affiliated Suzhou Hospital of Nanjing Medical UniversitySuzhou 215002Jiangsu ProvinceChina

Background:Inflammation is an important factor in pathological *** role of neutrophils,one of the most important inflammatory cells,in scar hyperplasia remains *** purpose of this article is to study the correlation between neutrophil extracellular traps(NETs)and scar hyperplasia and identify a new target for inhibiting scar ***:Neutrophils were isolated from human peripheral blood by magnetic-bead *** in plasma and scars were detected by enzyme-linked immunosorbent assays(ELISAs),immunofluorescence and flow *** was used to assess neutrophil(CD66B)infiltration in hypertrophic *** observe the entry of NETs into fibroblasts we used immunofluorescence and flow ***:We found that peripheral blood neutrophils in patients with hypertrophic scars were more likely to form NETs(pfibroblasts in vitro to promote their differentiation and *** of NETs with cytochalasin in wounds reduced the hyperplasia of scars in *** induced neutrophils to generate NETs with different stimuli in vitro and detected the proteins carried by *** did not find an increase in the expression of common scarring factors[interleukin(IL)-17 and transforming growth factor-β(TGF-β),p>0.05].However,inhibiting the production of NETs or degrading DNA reduced the differentiation of fibroblasts *** vitro,NETs were found to be mediated by Toll-like receptor 9(TLR-9)in fibroblasts and further phosphorylated nuclear factor Kappa-B(NF-κB).We found that IL-6,which is downstream of NF-κB,was increased in ***,IL-6 uses autocrine and paracrine signaling to promote differentiation and ***:Our experiments found that NETs activate fibroblasts through the TLR-9/NF-κB/IL-6 pathway,thereby providing a new target for regulating hypertrophic scars.

关键词： Neutrophil extracellular traps Hypertrophic scar Toll-like receptor 9 fibroblast Inflammation Differentiation Nuclear factor Kappa-B Interleukin-6

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P-MSC-derived extracellular vesicles facilitate diabetic wound healing via miR-145-5p/CDKN1A-mediated functional improvements of high glucose-induced senescent fibroblasts

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Burns & Trauma 2023年第1期11卷 325-348页

作者： Jianlong Su Qian Wei Kui Ma Yaxi Wang Wenzhi Hu Hao Meng Qiankun Li Yuehou Zhang Wenhua Zhang Haihong Li Xiaobing Fu Cuiping Zhang Research Center for Tissue Repair and Regeneration Affiliated to the Medical Innovation Research Division and the 4th Medical Center of Chinese PLA General Hospital 51 Fucheng RoadHaidian DistrictBeijing 100048China School of Medicine NanKai University94Weijin RoadNankai DistrictTianjin 300071China Research Unit of Trauma Care Tissue Repair and RegenerationChinese Academy of Medical Sciences2019RU05151 Fucheng RoadHaidian DistrictBeijing 100048China Burn and Plastic Surgery Zhongda Hospital Affiliated Southeast UniversityDingjiaqiao 87Gulou DistrictNanjing 210009China Department of Wound Repair Institute of Wound Repair and Regeneration MedicineSouthern University of Science and Technology HospitalSouthern University of Science and Technology School of Medicine6019 Xililiuxian RoadNanshan DistrictShenzhen 518055China PLA Key Laboratory of Tissue Repair and Regenerative Medicine and Beijing Key Research Laboratory of Skin Injury Repair and Regeneration51 Fucheng RoadHaidian DistrictBeijing 100048China

Background:Persistent hyperglycaemia in diabetes causes functional abnormalities of human dermal fibroblasts(HDFs),partially leading to delayed skin wound *** vesicles(EVs)containing multiple pro-healing microRNAs(miRNAs)have been shown to exert therapeutic effects on diabetic wound *** present study aimed to observe the effects of EVs derived from placental mesenchymal stem cells(P-MSC-EVs)on diabetic wound healing and high glucose(HG)-induced senescent fibroblasts and to explore the underlying ***:P-MSC-EVs were isolated by differential ultracentrifugation and locally injected into the full-thickness skin wounds of diabetic mice,to observe the beneficial effects on wound healing in vivo by measuring wound closure rates and histological ***,a series of assays were conducted to evaluate the effects of low(2.28 x 1010 particles/ml)and high(4.56 x 1010 particles/ml)concentrations of P-MSC-EVs on the senescence,proliferation,migration,and apoptosis of HG-induced senescent HDFs in ***,miRNA microarrays and real-time quantitative PCR(RT-qPCR)were carried out to detect the differentially expressed miRNAs in HDFs after EVs *** RNA inhibitors,miRNA mimics,and small interfering RNA(siRNA)were used to evaluate the role of a candidate miRNA and its target genes in P-MSC-EV-induced improvements in the function of HG-induced senescent ***:Local injection of P-MSC-EVs into diabetic wounds accelerated wound closure and reduced scar widths,with better-organized collagen deposition and decreased p16INK4a *** vitro,P-MSC-EVs enhanced the antisenescence,proliferation,migration,and antiapoptotic abilities of HG-induced senescent fibroblasts in a dose-dependent ***-145-5p was found to be highly enriched in ***-145-5p inhibitors effectively attenuated the P-MSC-EV-induced functional improvements of senescent ***-145-5p mimics simulated the effects of P-MSC-EVs on funct

关键词： Diabetic wounds Mesenchymal stem cells Extracellular vesicles miR-145-5p CDKN1A Senescence fibroblast Glucose

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Substrate Stiffness and Topography Affect the Morphology of Human fibroblasts in Mechanical Microenvironment

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Journal of Shanghai Jiaotong university(Science) 2023年第4期28卷 495-506页

作者：刘阳王雅靖温大渭张全有王立安美文刘勇 Institute of Biomedical Engineering Shanxi Key Laboratory of Material Strength&Structural ImpactResearch Center for Nano-Biomaterials&Regenerative MedicineCollege of Biomedical EngineeringTaiyuan University of TechnologyTaiyuan 030024China Department of Nuclear Medicine First Hospital of Shanxi Medical UniversityTaiyuan 030001China Dermatology Department Shanxi Bethune HospitalShanxi Academy of Medical SciencesTaiyuan 030032China

Hyperplastic scar is a common fibrotic disease that may ultimately lead to severe dysfunction anddeformity,causing physical and psychological ***,we aim to evaluate the effect of the mechanicalmicroenvironment of scar substrates on the morphology of human fibroblasts(HFbs).The micro-modular fabrication technique was used to design a new cross-groove topology and to construct four elastic substrates withthe stiffness of 19.3 kPa and 90.1 kPa coupled with parallel groove and cross groove,respectively,to simulate themechanical microenvironment of skin wounds and scar *** morphological changes in HFbs in differentsubstrates were observed,and the changes in the cell-long axis length,area,and the angle between cell-long axisand grooves were *** staining was performed to observe the distribution of microfila*** results indicated that substrate stiffness and topography affected the morphology of *** cellswere elongated in parallel grooves as well as in the area where cross grooves restricted groove length,the celllength was restricted,and the angle between the long axis and the groove was *** topography exertedno significant effect on the cell area,but the cell area increased with increasing the *** parallel groovepromoted the expression of the F-actin to a certain extent,and the fluorescence intensity of F-actin decreased withincreasing the *** the effect of the mechanical microenvironment of substrates on HFb morphologyis of great importance for understanding the mechanisms of scar formation and prevention.

关键词： mechanical micro-environment matrix stiffness topological structure fibroblast

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