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Function and Mechanism Research of Recombinant Adenovirus Expressed CCN1 in Healing Process of Combined Radiation and Wound Injury

Author: DuYeJun
Tutor: SiWeiKe
School: Third Military Medical University
Course: Clinical Laboratory Science
Keywords: CCN1 Radiation Wound healing Adenovirus
CLC: R641
Type: Master's thesis
Year: 2008
Downloads: 10
Quote: 0
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Abstract


The purpose put record combined injury is one of the main injury type wartime nuclear explosions and peacetime nuclear accident occurred, delayed wound healing mainly healing injury, or even long-term healing. CCN1 (Cysteine ??rich 61, Cyr61/CCN1) is a member of the CCN family of immediate early genes in vitro studies can promote cell adhesion, proliferation, migration, regulating angiogenesis impact matrix metabolism involved in wound healing, about CCN1 combined injury put record The healing has not been reported. Put this topic detection CCN1 protein and mRNA in rat record composite wounds granulation tissue expression; build the wound tissue after the recombinant adenovirus AdCCN1 infection, promote wound healing, as well as the expression and regulation of the relevant factors of wound repair. Through research, to explore the role and mechanism of CCN1 put record composite injury together. Provides a new perspective for an in-depth understanding of the mechanism of the process of wound healing, and to explore new ways to promote better treatment of new drugs. Second, the methods and results. Put invasive rat model of combined injury and CCN1 expression in wound tissue and its significance: the rats were randomly divided into simple trauma group and put the record combined injury group. Put record the combined injury group a 60Coγ-ray body irradiation within 1h after irradiation system back full-thickness skin excision wound. Drawn CCN1 mRNA and protein expression levels were detected by RT-PCR and immunohistochemistry, found that approximately 15d after injury before CCN1 protein and mRNA than mere 0,3,6,9,15,21 d after injury trauma decreased expression, elevated expression levels of about 21d. CCN1 expression than simple trauma group delay. 2. CCN1 Adenovirus Preparation and activity detection: connection by digestion manner pCMV-CCN1 mice CCN1 expression frame incoming pAdTrace-TO4 shuttle vector to obtain pAdTrace-CCN1 linearized AdEasy -1 E. coli homologous recombination obtain pAdCCN1 adenoviral plasmid transfected into 293 cells, subsequently linearized virus particle packing. The collected viral lysate and repeatedly amplified viral infection of 293 cells infected wounds CHO cells and the wound tissue cells, using Western Blot and IHC identification CCN1 correct secretion in CHO cells, and high expression in the wound tissue cells. Put record combined injury repair 3.CCN1 recombinant adenovirus experimental study of its mechanism of action: after injury, the wound edge immediately were injected subcutaneously equal amount of AdCCN1 the virus and AdRFP virus, drawn 7 days after fixed paraffin sections, HE staining that AdCCN1 group wound fibroblasts increased microvessel density increased, IHC detection MMP1, TIMP1, bFGF wound healing associated factor expression are elevated. Conclusion 1. Established rat model of combined injury put record, the amount increased to about 15d before granulation tissue after injury compared with trauma CCN1 protein and mRNA expression decreased approximately 21d expression detected. CCN1 expression decline may be one of the reasons put record combined injury refractory. 2. The application AdEasy system building, prepared AdCCN1 recombinant adenovirus enzyme digestion results in line with expectations; discovered that can infect CHO cells correctly secretion CCN1; effective wound tissue infection, and can make high expression CCN1. 3.AdCCN1 in put record the composite injury cooperative mechanism may be that: infected wound tissue cells AdCCN1 high expression secretion CCN1 to increase MMP1, TIMP1, bFGF, trauma healing relevant factor expression play a role.

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CLC: > Medicine, health > Surgery > Traumatology > Trauma
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