title: The stimulatory effects of eNOS/F92A-Cav1 on NO production and angiogenesis in BMSCs.
abstract: BACKGROUND
  Nitric oxide (NO) is generated in endothelial cells by endothelial nitric oxide synthase (eNOS). Caveolin-1 (Cav1) inhibits eNOS function and NO production. Modifying Cav1 scaffold domain, in particular Phenylalanine at position 92 (F92) is critical for the inhibitory actions of Cav1 toward eNOS. The aims of this study were to investigate the effect of enhanced NO production in term of in vitro angiogenesis on rat bone marrow derived mesenchymal stem cells (BMSCs) transduced with a novel bicistronic lentiviral vector co-expressing eNOS and mutant Cav1 (F92A).
  
  
  METHODS
  A bicistronic eNOS/F92-Cav1 lentiviral vector was constructed, and used to transduce rat BMSCs. The expression of eNOS and VEGF protein were confirmed by western-blot. NO production was detected by the greiss assay and in vitro angiogenesis was assessed by matrigel assisted capillary tube formation. The cell viability was evaluated using a Cell Counting Kit (CCK)-8.
  
  
  RESULTS
  The bicistronic eNOS/F92A-Cav1 lentiviral vector increased eNOS and VEGF protein expression, NO production compared to controls. In vitro capillary formation was increased in eNOS-F92A transduced cells and cell viability was not affected by transduction.
  
  
  CONCLUSIONS
  Transduction of rat BMSCs with an eNOS-F92A-Cav1 lentiviral vector can increase NO production by enhancing eNOS protein expression. The increased NO production did not reduce cell viability. This study demonstrates that genetic modification of BMSCs to enhance NO producton could be applied in stem cell based therapeutic approaches to treat diseases such as pulmonary arterial hypertension (PAH) which is characterized by decreased endothelial NO release.
authors.affiliation: Department of Cardiology, Liaocheng People's Hospital and Affiliated Liaocheng People's Hospital of Shandong University, Liaocheng, Shandong 252000, China; Taishan Medical University, Taian, Shandong 271016, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Xia
authors.affiliation: Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China.
authors.forename: Hai-Ying
authors.initials: HY
authors.lastname: Chen
authors.affiliation: Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China.
authors.forename: Shuang-Feng
authors.initials: SF
authors.lastname: Chen
authors.affiliation: Department of Cardiology, Liaocheng People's Hospital and Affiliated Liaocheng People's Hospital of Shandong University, Liaocheng, Shandong 252000, China; Taishan Medical University, Taian, Shandong 271016, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.affiliation: School of Biomedical Sciences, Charles Sturt University, Wagga Wagga, NSW 2650, Australia.
authors.forename: Pádraig Michael
authors.initials: PM
authors.lastname: Strappe
authors.affiliation: Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China.
authors.forename: Hong-Li
authors.initials: HL
authors.lastname: Yang
authors.affiliation: Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China.
authors.forename: Chang-Hui
authors.initials: CH
authors.lastname: Zhou
authors.affiliation: Department of Cardiology, Liaocheng People's Hospital and Affiliated Liaocheng People's Hospital of Shandong University, Liaocheng, Shandong 252000, China; Taishan Medical University, Taian, Shandong 271016, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China.
authors.forename: Ying-Xin
authors.initials: YX
authors.lastname: Zhang
authors.affiliation: Department of Cardiology, Liaocheng People's Hospital and Affiliated Liaocheng People's Hospital of Shandong University, Liaocheng, Shandong 252000, China; Taishan Medical University, Taian, Shandong 271016, China; Central Laboratory, and Key Laboratory of Oral and Maxillofacial-Head and Neck Medical Biology, Liaocheng People's Hospital, Liaocheng, Shandong 252000, China; School of Biomedical Sciences, Charles Sturt University, Wagga Wagga, NSW 2650, Australia. Electronic address: maldongchang@yahoo.com.cn.
authors.forename: Long-le
authors.initials: LL
authors.lastname: Ma
authors.affiliation: Department of Cardiology, Liaocheng People's Hospital and Affiliated Liaocheng People's Hospital of Shandong University, Liaocheng, Shandong 252000, China; School of Biomedical Sciences, Charles Sturt University, Wagga Wagga, NSW 2650, Australia. Electronic address: lwang@csu.edu.au.
authors.forename: Lexin
authors.initials: L
authors.lastname: Wang
mesh_terms: D000818:Animals; D051242:Caveolin 1 / Q000235:genetics*; D002470:Cell Survival; D002478:Cells, Cultured; D005434:Flow Cytometry; D005822:Genetic Vectors; D019276:Glycocalyx; D016086:Lentivirus; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D009389:Neovascularization, Pathologic / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D052250:Nitric Oxide Synthase Type III / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D051381:Rats; D014161:Transduction, Genetic
chemical_list: D051242:Caveolin 1; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III
keywords: Bone marrow derived mesenchymal stem cells; Caveolin-1; Endothelial nitric oxide synthase; Nitric oxide