Variability in Endogenous Perfusion Recovery of Immunocompromised Mouse Models of Limb Ischemia

Abstract

Immunocompromised hind limb ischemia (HLI) murine models are essential for preclinical evaluation of human cell-based therapy or biomaterial-based interventions. These models are used to generate proof of principle that the approach is effective and also regulatory preclinical data required for translation to the clinic. However, surgical variations in creation of HLI models reported in the literature introduce variability in the pathological manifestation of the model, in consequence affecting therapeutic endpoints. This study aims to compare the extent of vascular regeneration in HLI-induced immunocompromised murine models to obtain a stable and more reproducible injury model for testing. Athymic and Balb/C nude mice underwent HLI surgery with single and double ligation of femoral artery (FA). The recovery from surgery was observed over a period of 2 weeks with respect to ischemia reperfusion using laser Doppler and clinical signs of necrosis and ambulatory impairment. Double ligation of the FA results in a more severe response to ischemia in Balb/C with endogenous perfusion recovery up to 50% ± 10% compared with 75% ± 20% in athymic nude mice. Single iliac artery (IA) and FA lead to creation of mild ischemia compared with femoral artery-vein (FAV) pair ligation in Balb/C. Microcirculatory parameters indicate significantly lower capillary numbers (26 ± 3/mm(2)) and functional capillary density (203 ± 5 cm/cm(2)) in the FAV group. In this study, we demonstrate a reproducible, arterial double ligation in an immunocompromised Balb/C nude mouse model that exhibits characteristic pathological signs of ischemia with impaired endogenous recovery.

Acidic preconditioning of endothelial colony-forming cells (ECFC) promote vasculogenesis under proinflammatory and high glucose conditions in vitro and in vivo

“…In the present study, we further extended those findings by demonstrating that preconditioned ECFC not only augmented blood flow but also increased vascular density and reduced the inflammation score as revealed by histological analysis of gastrocnemius muscle sections. Although endogenous recovery from ischemia in the PBS-treated animals at day 14 was elevated, this is expected for immunocompromised mice in general and this mouse strain in particular [ 24 ]. Obesity-related disorders, including diabetes and hypertension, are associated with impaired endothelial progenitor cell functions [ 25 ].…”

Fat extract promotes angiogenesis in a murine model of limb ischemia: a novel cell-free therapeutic strategy

“…A unilateral hindlimb ischemia model was generated in nude mice, aged 10–12 weeks, via ligation of the left femoral artery and its branches, as previously described [ 33 , 34 ]. In brief, the mice were anesthetized via isoflurane (2–3%) inhalation.…”

“…A ratio of 1 before surgery indicated equal blood perfusion of both legs. Tissue necrosis was scored as previously described [ 34 ]: 0, normal/no necrosis; 1, mild necrosis and/or deep cyanosis of toes; 2, necrosis/amputation of toes (two or more); 3, foot amputation; and 4, severe limb loss.…”

Consensus guidelines for the use and interpretation of angiogenesis assays

“…In addition to these two commonly used methods to monitor the neovascularization in the ischemic limb of the mouse, sometime a functional test is used in which the use of the foot and limb is analyzed for instance by assessment of the plantar/dorsiflexion [ 594 ]. In the more severe models, necrosis of toes or even feet ensues, leading to auto-amputation.…”