Cathepsin S-Mediated Negative Regulation of Wnt5a/SC35 Activation Contributes to Ischemia-Induced Neovascularization in Aged Mice

Abstract

stress in aged humans and animals remain largely unknown.The pathogenesis of human atherosclerosis and angiogenesis involves extensive vascular wall extracellular matrix (ECM) remodeling, which needs the participation of proteolytic enzymes. 7-9 We demonstrated that human hypoxic atherosclerotic plaques contained high levels of these proteases. 10-12 Biological hypoxia/ischemia and inflammatory stresses usually cause increases in the expression of several proteases including members of the cathepsin family (i.e., cathepsins K, L, and S), which then modulates vascular cellular events including migration, invasion, and proliferation and/or lumen maturation. 13-15 A ging impairs the body's ability to form new blood vessels under ischemic pathological conditions, and this impairment causes a decline in the body's tissue regeneration capacity. 1 In various mammals, the age-associated declines in angiogenesis and vascularization are characterized by endothelial cell (EC) and bone-marrow (BM)-derived endothelial progenitor cells (EPCs), a shift in the balance between vascular cell proliferation and apoptosis, and changes in the extracellular microenvironment (e.g., harmful changes in oxidative stress, inflammatory cytokines, and growth factors). 2-6 Indeed, the cellular and molecular mechanisms responsible for the defective angiogenic and vasculogenic reactions to chronic ischemic

Cathepsin K Deficiency Impaired Ischemia-Induced Neovascularization in Aged Mice

“…After the isolation of mononuclear cells, bone marrow-derived c-Kit + cells were isolated by using CD117 MicroBeads and magnetic-activated cell sorting (MACS) according to the manufacturer's instructions (Miltenyli Biotec, Bergisch-Gladbach, Germany). The c-Kit + bone marrow cells were >90% positive for CD31 + , as described [30]. The cultured cells were exhibited EC surface markers: CD31 and CD117.…”

“…The administration of bone marrow-derived or peripheral blood-derived EPCs has improved postischemic neovascularization in various experimental and clinical trials [22,30,32]. Impaired angiogenesis in individuals at advanced ages might be due to an interior decline in the regenerative capacity of vascular progenitors and/or a decline in a proregenerative niche.…”

“…CatK −/− and wild-type (CatK +/+ ) littermate controls were subjected to unilateral hindlimb ischemic surgery under ketamine (70 mg/kg, Sankyo Pharmaceutical, Tokyo) and xylazine (4.6 mg/kg, Bayer, Pharmaceutical, Tokyo) anesthesia [ 30 ]. At the indicated time points after surgery, mice were euthanized by an overdose of a ketamine and xylazine mixture, and then skeletal muscles were dissected out and immediately frozen at liquid nitrogen temperature before they were stored at −80°C until the western blot analysis.…”

“…At the indicated time points, the ratio of blood flow in the ischemia to that in the normal limb was measured with laser speckle blood flow imaging (LSBFI, Omegazone, OZ-1, OmEGA Wave, Tokyo) as described in [30]. With LSBFI, the blood flow is shown as the changes in the laser frequency in pixels of different colors.…”

“…We assayed the capillary density in cross-sections of adductor muscle at postoperative day 14 with anti-CD31 monoclonal antibody (mAb) (Santa Cruz Biotechnology, Santa Cruz, CA). The endothelial cells were quantified by measuring the number of cells that were positive for CD31 per high-power field (400×) [30]. We measured five randomly chosen microscopic fields from 5 to 7 different sections in each tissue block to determine the number of capillaries per muscle fiber.…”