Regular Transient Limb Ischemia Prevents Atherosclerosis Progression in Hypercholesterolemic Rabbits
Atherosclerosis, a leading cause of death worldwide, is characterized by the formation of plaque within arterial walls. The pathogenesis of atherosclerosis is complex, with endothelial dysfunction being a critical initial factor. Endothelial injury triggers an inflammatory-fibroproliferative response, leading to plaque formation. Therefore, interventions aimed at restoring endothelial function are essential in reducing cardiovascular risk. Transient limb ischemia (TLI), a technique involving brief episodes of limb blood flow occlusion followed by reperfusion, has shown promise in protecting endothelial function. This study explores the potential of regular TLI (RTLI) in preventing atherosclerosis progression in hypercholesterolemic rabbits.
The study was conducted on 28 male New Zealand white rabbits, randomly assigned to four groups: control, cholesterol, sham, and ischemia groups, each consisting of seven rabbits. The control group was fed a normal diet, while the other three groups were fed a hypercholesterolemic diet for 12 weeks. The ischemia group underwent six cycles of RTLI daily, while the sham group had a deflated cuff on the left hind limb for 60 minutes daily. Serum samples were collected at weeks 0, 4, 8, and 12 to measure total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C). At week 12, the entire aorta was harvested, stained with Sudan IV to identify plaque, and analyzed using Image J software.
The results showed that serum concentrations of TC, HDL-C, and LDL-C were significantly higher in the cholesterol group compared to the control group at weeks 4, 8, and 12. For instance, TC levels in the cholesterol group at week 4 were 29.60 mmol/L compared to 1.00 mmol/L in the control group. Similarly, HDL-C and LDL-C levels were significantly elevated in the cholesterol group. However, there were no significant differences in lipid profiles among the hypercholesterolemic groups (cholesterol, sham, and ischemia groups) at any time point.
Plaque area analysis revealed that the cholesterol group had a significantly higher percentage of plaque area (47.22%) compared to the control group (0%). The ischemia group showed a smaller plaque area (20.45%) compared to the cholesterol group (47.22%) and the sham group (37.88%). After transforming the data to square root values, the ischemia group had a significantly smaller plaque area (0.44) compared to both the cholesterol (0.67) and sham groups (0.61).
The study demonstrated that RTLI could significantly reduce the progression of atherosclerosis in hypercholesterolemic rabbits by decreasing the percentage of plaque area in the aorta. This finding suggests that RTLI might be an effective intervention in preventing atherosclerosis, potentially offering a novel therapeutic approach for atherosclerotic diseases.
The experimental protocol involved feeding the rabbits a hypercholesterolemic diet consisting of 1.5 g cholesterol, 9 g corn oil, and 69.5 g normal chow in the evening, while the control group received normal chow. The RTLI procedure involved inflating a blood pressure cuff to 200 mmHg for 5 minutes, followed by deflation for 5 minutes, repeated six times daily. This protocol was chosen based on previous studies demonstrating its safety and efficacy in inducing ischemic preconditioning.
Serum lipid analysis was performed using an automatic biochemical analyzer, and plaque area quantification was conducted using Sudan IV staining and image analysis software. Statistical analysis included ANOVA and Mann-Whitney U tests, with significance set at P < 0.05. The study also noted that there were no significant differences in weight gain among the groups over the 12-week period, indicating that the hypercholesterolemic diet and RTLI did not adversely affect the rabbits' overall health.
The findings of this study align with previous research indicating that TLI can protect endothelial function and reduce systemic inflammation. For example, Kimura et al. found that RTLI improved endothelial function in healthy humans by increasing nitric oxide production and endothelial progenitor cells. Similarly, Shimizu et al. reported that RTLI modulated systemic inflammatory responses by altering neutrophil function and cytokine levels.
The study’s limitations include the lack of investigation into the underlying mechanisms of RTLI’s protective effects, the absence of histological analysis of aortic tissue, and the need to explore different intensities and durations of RTLI. Future research should address these limitations to further elucidate the potential of RTLI in preventing atherosclerosis.
In conclusion, this study provides evidence that RTLI can significantly reduce plaque area in hypercholesterolemic rabbits, suggesting a potential therapeutic strategy for preventing atherosclerosis. The findings underscore the importance of further research into the mechanisms and optimal protocols of RTLI to translate these benefits into clinical practice.
doi.org/10.1097/CM9.0000000000000204
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