Beneficial effects of vascular endothelial growth factor B gene transfer in aged heart
Beneficial effects of vascular endothelial growth factor B gene transfer in aged heart
Aged male C57Bl/6J wildtype mice were purchased from Janvier (Le Genest SaintIsle, France) and from Charles River (Sulzfeld, Germany). Homozygosity of these inbred mice was controlled by Janvier and Charles River using exome sequencing.  To overexpress VEGFB186 in aged mouse hearts, AAV9 vectors carrying the VEGFB186 gene under the control of the CAG promotor were used. Non targeting AAV9 particles served as negative control. Both viruses were provided by K.A and I.S. The viral particles were injected intraperitoneally, at a concentration of 2.2 x 1011 viral particles per animal to 18-month-old mice. Before AAV9-treatment (baseline) and 4 and 8 weeks after the treatment, echocardiography was used to monitor the systolic and diastolic function of each mouse.  To isolate total RNA from cardiac tissue, hearts combined with 700µl of Qiazol and ¼” ceramic spheres and were homogenized three times for 20 seconds (4 min/s). The miRNeasy Mini kit (217004, Qiagen) was used for further steps of RNA isolation. The RNA concentration was determined by measuring absorption at 260 nm and 280 nm with the NanoDrop ND 2000-spectrophotometer (PeqLab). Library quantification and quality assessment was performed using Bioanalyzer Agilent 2100 using a High Sensitivity DNA chip (5067-4627 Agilent Genomics).