title: Type-2 diabetic Lepr(db/db) mice show a defective microvascular phenotype under basal conditions and an impaired response to angiogenesis gene therapy in the setting of limb ischemia.
abstract: Diabetes mellitus is associated with macro- and micro-angiopathy, leading to increased risk of peripheral ischemia. In the present study, we have characterized the microvascular phenotype at the level of limb muscles and the spontaneous angiogenesis response to surgically-induced unilateral limb ischemia in a murine model of type-2 diabetes, the obese C57BL/KsOlaHsd-Lepr(db/db) mice (Lepr(db/db)), and in non-diabetic heterozygous Lepr(db/+). Wild type C57BL mice (WT) were used as controls. The basal microvascular phenotype was determined in mice aged 3 or 5 months, while the response to limb ischemia was studied only in 5-month old mice. Moreover, in 5-month old ischemic Lepr(db/db) and Lepr(db/+), we have tested the therapeutic potential of local angiogenesis gene therapy with human tissue kallikrein (hTK) or constitutively-activated Akt kinase (Myr-Akt). We found that in the muscles of 3- or 5-month old Lepr(db/db), apoptosis of endothelial cells was enhanced and the densities of capillary and arteriole were reduced. Arterioles of Lepr(db/db) showed hypertrophic remodelling and, occasionally, lumen occlusion. Following ischemia, Lepr(db/db) showed a defective reparative angiogenesis in ischemic muscle, delayed blood flow recovery, and worsened clinical outcome as compared with controls. Five-month old Lepr(db/+) displayed an increase in endothelial cell apoptosis under basal conditions, while capillary and arteriole densities were normal. Lepr(db/+) mounted a proper reparative angiogenesis response to limb ischemia and regained blood flow to the ischemic limb, regularly. Local gene therapy with hTK or Myr-Akt induced angiogenesis in ischemic muscles of Lepr(db/+) and Lepr(db/db). However, in the Lepr(db/db) neither gene therapy approach improved the blood flow recovery and the clinical outcome from ischemia. In contrast, either hTK or Myr-Akt gene transfer improved the post-ischemic recovery of Lepr(db/+). Type-2 diabetes has a negative impact on the basal microvascular phenotype and severely impairs post-ischemic recovery of limb muscles. Gene therapy-induced stimulation of neovascularization might not suffice as a sole therapeutic strategy to combat type-2 diabetes-related vascular complications. In type-2 diabetic patients, therapeutic angiogenesis may need to be further optimized before being recommended for clinical applications.
authors.affiliation: Bristol Heart Institute, University of Bristol, Bristol, UK. mdxce@bristol.ac.uk
authors.forename: Costanza
authors.initials: C
authors.lastname: Emanueli
authors.forename: Andrea
authors.initials: A
authors.lastname: Caporali
authors.forename: Nicolle
authors.initials: N
authors.lastname: Krankel
authors.forename: Brunella
authors.initials: B
authors.lastname: Cristofaro
authors.forename: Sophie
authors.initials: S
authors.lastname: Van Linthout
authors.forename: Paolo
authors.initials: P
authors.lastname: Madeddu
mesh_terms: D000818:Animals; D002196:Capillaries / Q000473:pathology; D003924:Diabetes Mellitus, Type 2 / Q000150:complications*; D003925:Diabetic Angiopathies / Q000473:pathology / Q000503:physiopathology / Q000628:therapy*; D015316:Genetic Therapy; D006801:Humans; D007511:Ischemia / Q000473:pathology / Q000503:physiopathology / Q000628:therapy*; D035002:Lower Extremity / Q000098:blood supply; D051379:Mice; D008810:Mice, Inbred C57BL; D008820:Mice, Obese; D018482:Muscle, Skeletal / Q000098:blood supply / Q000378:metabolism; D018919:Neovascularization, Physiologic; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D010641:Phenotype; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics; D011956:Receptors, Cell Surface / Q000235:genetics; D054411:Receptors, Leptin; D020840:Tissue Kallikreins / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D011956:Receptors, Cell Surface; D054411:Receptors, Leptin; D042461:Vascular Endothelial Growth Factor A; D052250:Nitric Oxide Synthase Type III; D051057:Proto-Oncogene Proteins c-akt; D020840:Tissue Kallikreins