PMID- 36497083 OWN - NLM STAT- MEDLINE DCOM- 20221216 LR - 20221221 IS - 2073-4409 (Electronic) IS - 2073-4409 (Linking) VI - 11 IP - 23 DP - 2022 Nov 29 TI - The Efficacy of HGF/VEGF Gene Therapy for Limb Ischemia in Mice with Impaired Glucose Tolerance: Shift from Angiogenesis to Axonal Growth and Oxidative Potential in Skeletal Muscle. LID - 10.3390/cells11233824 [doi] LID - 3824 AB - BACKGROUND: Combined non-viral gene therapy (GT) of ischemia and cardiovascular disease is a promising tool for potential clinical translation. In previous studies our group has developed combined gene therapy by vascular endothelial growth factor 165 (VEGF165) + hepatocyte growth factor (HGF). Our recent works have demonstrated that a bicistronic pDNA that carries both human HGF and VEGF165 coding sequences has a potential for clinical application in peripheral artery disease (PAD). The present study aimed to test HGF/VEGF combined plasmid efficacy in ischemic skeletal muscle comorbid with predominant complications of PAD-impaired glucose tolerance and type 2 diabetes mellitus (T2DM). METHODS: Male C57BL mice were housed on low-fat (LFD) or high-fat diet (HFD) for 10 weeks and metabolic parameters including FBG level, ITT, and GTT were evaluated. Hindlimb ischemia induction and plasmid administration were performed at 10 weeks with 3 weeks for post-surgical follow-up. Limb blood flow was assessed by laser Doppler scanning at 7, 14, and 21 days after ischemia induction. The necrotic area of m.tibialis anterior, macrophage infiltration, angio- and neuritogenesis were evaluated in tissue sections. The mitochondrial status of skeletal muscle (total mitochondria content, ETC proteins content) was assessed by Western blotting of muscle lysates. RESULTS: At 10 weeks, the HFD group demonstrated impaired glucose tolerance in comparison with the LFD group. HGF/VEGF plasmid injection aggravated glucose intolerance in HFD conditions. Blood flow recovery was not changed by HGF/VEGF plasmid injection either in LFD or HFD conditions. GT in LFD, but not in HFD conditions, enlarged the necrotic area and CD68+ cells infiltration. However, HGF/VEGF plasmid enhanced neuritogenesis and enlarged NF200+ area on muscle sections. In HFD conditions, HGF/VEGF plasmid injection significantly increased mitochondria content and ETC proteins content. CONCLUSIONS: The current study demonstrated a significant role of dietary conditions in pre-clinical testing of non-viral GT drugs. HGF/VEGF combined plasmid demonstrated a novel aspect of potential participation in ischemic skeletal muscle regeneration, through regulation of innervation and bioenergetics of muscle. The obtained results made HGF/VEGF combined plasmid a very promising tool for PAD therapy in impaired glucose tolerance conditions. FAU - Stafeev I, Iurii S AU - Stafeev I IS AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. FAU - Boldyreva, Maria A AU - Boldyreva MA AUID- ORCID: 0000-0002-8309-0018 AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. AD - Cell and Molecular Biotechnology Unit, Faculty of Biology and Biotechnology, National Research University Higher School of Economics, 101000 Moscow, Russia. FAU - Michurina, Svetlana S AU - Michurina SS AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. AD - Faculty of Biology, Lomonosov Moscow State University, 117192 Moscow, Russia. FAU - Agareva, Margarita Yu AU - Agareva MY AUID- ORCID: 0000-0002-9603-9780 AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. AD - Institute of Fine Chemical Technologies Named after M.V. Lomonosov, 119571 Moscow, Russia. FAU - Radnaeva, Arina V AU - Radnaeva AV AD - Faculty of Medicine, Lomonosov Moscow State University, 119192 Moscow, Russia. AD - Institute for Regenerative Medicine, Medical Research and Education Centre, Lomonosov Moscow State University, 119192 Moscow, Russia. FAU - Menshikov, Mikhail Yu AU - Menshikov MY AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. FAU - Hu, Yu-Chen AU - Hu YC AD - Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300044, Taiwan. AD - Frontier Research Center on Fundamental and Applied Sciences of Matters, National Tsing Hua University, Hsinchu 300044, Taiwan. FAU - Makarevich, Pavel I AU - Makarevich PI AUID- ORCID: 0000-0001-8869-5190 AD - Faculty of Medicine, Lomonosov Moscow State University, 119192 Moscow, Russia. AD - Institute for Regenerative Medicine, Medical Research and Education Centre, Lomonosov Moscow State University, 119192 Moscow, Russia. FAU - Parfyonova, Yelena V AU - Parfyonova YV AD - National Medical Research Centre for Cardiology Named after Academician E.I.Chazov, 121552 Moscow, Russia. AD - Institute for Regenerative Medicine, Medical Research and Education Centre, Lomonosov Moscow State University, 119192 Moscow, Russia. LA - eng GR - 20-45-08003/Russian Science Foundation/ GR - 20-015-00181/Russian Foundation of Basic Research/ PT - Journal Article DEP - 20221129 PL - Switzerland TA - Cells JT - Cells JID - 101600052 RN - 67256-21-7 (Hepatocyte Growth Factor) RN - 0 (Vascular Endothelial Growth Factor A) RN - 0 (HGF protein, human) SB - IM MH - Mice MH - Male MH - Humans MH - Animals MH - Hepatocyte Growth Factor/genetics/metabolism MH - Vascular Endothelial Growth Factor A/genetics/metabolism MH - *Glucose Intolerance/complications/genetics/therapy MH - *Diabetes Mellitus, Type 2/complications/genetics/therapy MH - Mice, Inbred C57BL MH - Ischemia/metabolism MH - Genetic Therapy/methods MH - Muscle, Skeletal/metabolism PMC - PMC9737863 OTO - NOTNLM OT - HGF OT - VEGF OT - gene therapy OT - high-fat diet OT - limb ischemia OT - obesity OT - plasmid COIS- The authors declare no conflict of interest. EDAT- 2022/12/12 06:00 MHDA- 2022/12/15 06:00 PMCR- 2022/11/29 CRDT- 2022/12/11 01:03 PHST- 2022/11/03 00:00 [received] PHST- 2022/11/23 00:00 [revised] PHST- 2022/11/27 00:00 [accepted] PHST- 2022/12/11 01:03 [entrez] PHST- 2022/12/12 06:00 [pubmed] PHST- 2022/12/15 06:00 [medline] PHST- 2022/11/29 00:00 [pmc-release] AID - cells11233824 [pii] AID - cells-11-03824 [pii] AID - 10.3390/cells11233824 [doi] PST - epublish SO - Cells. 2022 Nov 29;11(23):3824. doi: 10.3390/cells11233824.