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Here, a liver\u2010specific knockdown mouse model was developed based on a recombinant AAV (rAAV) tool, which is rapid and economical, with optimized delivery methods and systematically evaluated knockdown efficiency. Hepatic ischemia\/reperfusion (IR) injury\u2010related genes (\n                    <jats:italic>Btg2<\/jats:italic>\n                    ,\n                    <jats:italic>Flrt3<\/jats:italic>\n                    ,\n                    <jats:italic>Klf4<\/jats:italic>\n                    ) were screened through GEO data mining. To knock down these genes, rAAV vectors featuring a liver\u2010specific promoter TBG669, Firefly luciferase (Fluc) for real\u2010time tracking, and miR30\u2010based regulatory elements were engineered. Delivery protocols were optimized through comparative analysis of reporting systems (GFP vs. Fluc), administration routes (tail vein vs. dorsal penile vein), and doses (half [2.5\u2009\u00d7\u200910\n                    <jats:sup>11<\/jats:sup>\n                    GC] vs. full dose [5.0\u2009\u00d7\u200910\n                    <jats:sup>11<\/jats:sup>\n                    GC]). Safety and knockdown efficiency were systematically assessed using histopathology, serum biochemistry, qRT\u2010PCR, and western blot analysis both in bulk liver tissue and single\u2010cell subsets. The optimized rAAV system achieved 100% hepatic infection rates with minimal off\u2010target expression. Dorsal penile vein injection simplified delivery without compromising efficacy. Full\u2010dose administration produced stronger bioluminescence by Week 4, with no adverse effects on organ function or histology. For knockdown efficiencies,\n                    <jats:italic>Btg2<\/jats:italic>\n                    and\n                    <jats:italic>Flrt3<\/jats:italic>\n                    mRNA and protein levels were reduced by &gt;\u200950%, whereas\n                    <jats:italic>Klf4<\/jats:italic>\n                    exhibited moderate suppression in bulk liver tissue. In single\u2010cell subsets, the three genes were all effectively knocked down in Kupffer cells, followed by hepatocytes and LSECs. A time\u2010 and cost\u2010efficient rAAV\u2010based hepatic gene knockdown murine model was established, enabling precise investigation of liver disease mechanisms and therapeutic development.\n                  <\/jats:p>","DOI":"10.1096\/fj.202502602r","type":"journal-article","created":{"date-parts":[[2025,10,30]],"date-time":"2025-10-30T08:41:51Z","timestamp":1761813711000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["In\u2010Depth Optimization and Systematic Evaluation of a Recombinant\n                    <scp>AAV<\/scp>\n                    \u2010Based Hepatic\u2010Targeted Gene Knockdown Murine Model"],"prefix":"10.1096","volume":"39","author":[{"given":"Ying","family":"Zhu","sequence":"first","affiliation":[{"name":"Department of Hepatobiliary and Pancreatic Surgery the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"},{"name":"Henan Key Laboratory for Hepatopathy and 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Zhengzhou Henan China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaodan","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Hepatobiliary and Pancreatic Surgery the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"},{"name":"Henan Key Laboratory for Hepatopathy and Transplantation Medicine  Zhengzhou Henan China"},{"name":"Henan Key Laboratory of Digestive Organ Transplantation the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhiping","family":"Yan","sequence":"additional","affiliation":[{"name":"Department of Hepatobiliary and Pancreatic Surgery the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"},{"name":"Henan Key Laboratory for Hepatopathy and Transplantation Medicine  Zhengzhou Henan China"},{"name":"Henan Key Laboratory of Digestive Organ Transplantation the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mingjie","family":"Ding","sequence":"additional","affiliation":[{"name":"Department of Hepatobiliary and Pancreatic Surgery the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"},{"name":"Henan Key Laboratory for Hepatopathy and Transplantation Medicine  Zhengzhou Henan China"},{"name":"Henan Key Laboratory of Digestive Organ Transplantation the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0821-1318","authenticated-orcid":false,"given":"Wenzhi","family":"Guo","sequence":"additional","affiliation":[{"name":"Department of Hepatobiliary and Pancreatic Surgery the First Affiliated Hospital of Zhengzhou University  Zhengzhou Henan China"},{"name":"Henan Key Laboratory for Hepatopathy and Transplantation Medicine  Zhengzhou Henan 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