Effect of rapamycin-eluting stents on in-stent restenosis and early inflammatory response in coronary artery narrowing animal models

Abstract

Objective

it was to evaluate the efficacy and safety of rapamycin-eluting stents at different doses in the treatment of coronary artery narrowing in miniature pigs.

Methods

a total of 20 miniature pigs were randomly assigned into four groups: S1 group (low-dose rapamycin-coated stent, 55 µg/mm²), S2 group (medium-dose rapamycin-coated stent, 120 µg/mm²), S3 group (high-dose rapamycin-coated stent, 415 µg/mm²), and D0 group (bare metal stent). The stent size was 3.0 mm × 18 mm, with an over-expansion ratio of 1.1. Each group consisted of five pigs. Stent implantation was followed by euthanasia and tissue collection after 1 month. Vascular measurements, inflammatory response scores, cardiovascular injury scores, endothelialization scores, liver and kidney function indices, and myocardial injury markers were compared among the groups.

Results

the neointimal thickness in the S2 and S3 groups was significantly lower than that in the S1 and D0 groups (S1 group: 24.08 ± 3.95, S2 group: 1.86 ± 0.28, S3 group: 2.72 ± 0.74, D0 group: 22.85 ± 3.15, P < 0.05). The residual lumen area in the S2 and S3 groups was significantly larger than that in the S1 and D0 groups (S1 group: 2.73 ± 0.51, S2 group: 4.25 ± 0.78, S3 group: 3.91 ± 0.73, D0 group: 2.91 ± 0.44, P < 0.05). The neointimal area in the S2 and S3 groups was significantly smaller than that in the S1 and D0 groups (S1 group: 3.44 ± 0.84, S2 group: 1.78 ± 0.25, S3 group: 2.07 ± 0.41, D0 group: 3.43 ± 0.72, P < 0.05). The degree of lumen narrowing in the S2 and S3 groups was significantly lower than that in the S1 and D0 groups (S1 group: 44.25 ± 3.66%, S2 group: 14.19 ± 2.01%, S3 group: 15.29 ± 2.45%, D0 group: 21.79 ± 3.51%, P < 0.05). The inflammation scores of coronary artery walls in the S2 and S3 groups of miniature pigs were markedly lower than those in the S1 and D0 groups (P < 0.05). The cardiovascular injury scores (P = 0.072) and endothelialization scores (P = 0.085) differed slightly among the four groups (P > 0.05). Post-operative liver function indicators (alanine transaminase, aspartate transaminase), kidney function indicators (blood urea nitrogen, serum creatinine), and myocardial injury markers (creatine kinase, creatine kinase-MB) also showed neglectable differences among the four groups (P > 0.05).

Conclusion

medium and high doses of rapamycin-eluting stents effectively inhibit neointimal hyperplasia and local vascular inflammatory response in miniature pigs without causing damage to liver and kidney functions or myocardial cells. These stents demonstrate high efficacy and safety. Rapamycin-coated coronary stents, as an effective treatment for coronary artery stenosis, may achieve further improvement in therapeutic efficacy through optimization of drug dosage and stent design.

Background

Coronary artery disease (CAD) is a leading cause of mortality globally in both men and women, accounting for approximately one-third of all deaths [1,2,3]. CAD, particularly atherosclerosis of the coronary arteries, can progress to coronary artery stenosis and even arterial occlusion [4]. Coronary artery stenosis is a common condition that primarily affects individuals over the age of 40, with a higher incidence observed in men and those engaged in sedentary or intellectually demanding occupations, reflecting an epidemic in industrialized nations. The disease is primarily caused by coronary atherosclerosis and is more prevalent in individuals with a family history of cardiovascular disease, as well as those with obesity or diabetes mellitus. Additionally, long-term smoking, psychological stress, and unbalanced diet may contribute to the development of coronary artery stenosis [5, 6]. Initially, patients may be asymptomatic or experience symptoms only during intense physical exertion. As coronary arteries continue to narrow, blood flow to the heart progressively diminishes, leading to more severe or frequent symptoms, including chest pain, palpitations, and heart failure [7]. Some patients may exhibit systemic symptoms, primarily fever, accompanied by tachycardia, elevated white blood cell count, and accelerated erythrocyte sedimentation rate [8].

The treatment of coronary artery stenosis should be tailored according to the underlying cause and the severity of the stenosis [9,10,11,12]. The treatment should be timely and effective, aimed at rapidly alleviating myocardial ischemia symptoms caused by coronary artery stenosis. For patients with mild disease, pharmacological therapy is the first choice, while for those with severe coronary stenosis complicated by myocardial ischemia, surgical or interventional thrombolytic therapy may be considered. Commonly used medications include morphine, pethidine, nitroglycerin, β-blockers, perindopril, heparin, and enalapril, among others [13]. Venetsanos et al. (2018) [14] investigated the use of bivalirudin, with or without the addition of glycoprotein IIb/IIIa inhibitors, in combination with standard heparin in coronary interventions for 23,800 patients with ST-segment elevation myocardial infarction. The study found that bivalirudin combined with glycoprotein IIb/IIIa inhibitors was significantly associated with a reduced risk of in-hospital major bleeding, thereby improving the safety of the treatment. Stent implantation is a mechanical interventional procedure, wherein the principle involves percutaneous access through the femoral artery, followed by the use of specialized catheters and guidewires to deliver the stent to the site of coronary artery stenosis or occlusion under fluoroscopic guidance. The stent expands the narrowed vessel wall, restoring blood flow and improving patency [15]. This procedure is an important method for addressing severe coronary artery stenosis that leads to inadequate blood supply, offering advantages such as no need for open chest surgery, local anesthesia, and quick recovery. However, for long or multiple lesions, coronary artery bypass grafting (CABG) remains necessary [16]. Chaszczewski et al. (2022) [17] analyzed the use of percutaneous coronary stent angioplasty for the treatment of post-operative coronary occlusion in critically ill infants, and found that stent angioplasty is a feasible and effective treatment for post-operative coronary occlusion in these infants. This technique allows for short-term reperfusion, promoting recovery of ventricular function, and may facilitate collateral circulation development when long-term stent patency can’t be achieved. Kim et al. (2014) [18] suggested that the strategy of local drug-eluting stent (DES) implantation is more effective than covering the entire lesion, significantly reducing the incidence of target vessel failure at 1 year in patients with coronary artery stenosis. This indicates that while stent implantation effectively addresses short-segment coronary artery stenosis, in-stent restenosis (ISR) and stent-associated inflammatory responses remain significant clinical challenges. In recent years, the application of DES has provided new solutions to mitigate ISR, particularly rapamycin-coated stents. As an immunosuppressant, rapamycin inhibits smooth muscle cell proliferation, reducing the occurrence of ISR, and has been widely used in coronary interventional therapies.

Although rapamycin-coated stents have shown significant efficacy in reducing in-stent restenosis, most existing studies have primarily focused on the effects of drug release rates and dosages, with limited in-depth research on how the changes in local drug concentrations after release affect the mechanisms of early inflammatory responses. Therefore, this study utilized a miniature pig animal model to evaluate the interventional treatment effects of rapamycin-eluting stents on coronary artery stenosis, with a particular focus on their role in inflammatory responses and immune mechanisms. The aim of the study was to explore the potential of rapamycin-coated stents at different dosages in the treatment of coronary artery stenosis, addressing gaps in the current literature regarding drug dosage and biological responses, and providing new theoretical foundations and experimental data for the prevention and treatment of coronary artery stenosis.

Study hypothesis

Hypothesis 1

There were significant differences in the therapeutic effects of rapamycin-coated stents at different dosages in the miniature pig model.

This hypothesis was based on previous studies showing that rapamycin, as an antiproliferative agent, effectively inhibits neointimal hyperplasia in coronary arteries. Therefore, it was hypothesized that medium- and high-dose rapamycin-coated stents may significantly reduce neointimal thickness, elastic lamina coverage area, lumen narrowing, and suppress local vascular inflammatory responses.

Hypothesis 2

Rapamycin-coated stents exhibited good biocompatibility and did not cause damage to the liver, kidney function, or myocardial cells in the pig model.

It was hypothesized that the use of rapamycin-coated stents won’t significantly impact the physiological parameters of the miniature pigs and that the stents would exhibit a high level of safety. This hypothesis was based on previous research on rapamycin-eluting stents, which has demonstrated that these stents typically do not cause significant systemic toxicity or organ damage.

Hypothesis 3

Low-dose rapamycin-coated stents may be insufficient to achieve the desired therapeutic effects.

It was hypothesized that low-dose rapamycin-coated stents may fail to effectively inhibit neointimal hyperplasia and local inflammatory responses due to the lower drug concentration, thus not achieving the therapeutic outcomes observed with medium- and high-dose stents.

Materials and methods

Research animals

A total of 20 Bama miniature pigs, of both sexes, weighing 30 ± 5 kg and aged between 5 and 7 months, were purchased from Chengdu Dashuo Experimental Animal Co., Ltd. Animal samples underwent standard immunization procedures following experimental animal protocols, including rigorous deworming and professional bathing procedures to maintain cleanliness of the body surface. This study adhered to the principles of the 3Rs (Replacement, Reduction, Refinement) and received ethical approval from the Beidahuang Industry Group General Hospital Institutional Animal Ethics Committee.

Biological scaffold materials

The Xylo^® drug-eluting stent system (rapamycin) was purchased from GW Medical Products Ltd., USA. The stent size was 3.0 mm × 18 mm, with an over-expansion ratio of 1.1. This system features a cobalt-chromium alloy stent platform loaded with the classic anti-proliferative drug rapamycin, including low-dose rapamycin-eluting stents (55 µg/mm²), medium-dose rapamycin-eluting stents (120 µg/mm²), and high-dose rapamycin-eluting stents (415 µg/mm²). Cobalt-chromium alloy bare-metal stents were used as a blank control. All coated stents were sterilized and stored after ethylene oxide sterilization.

Coronary artery stenosis model in the Bama miniature pig

The animals were fasted for twelve hours prior to surgery and deprived of water for six hours. Anesthesia was induced using ketamine and pentobarbital sodium, and continuous electrocardiographic monitoring was performed. A guiding sheath was introduced via femoral artery puncture, and heparinization was applied to prevent thrombus formation. A contrast catheter was advanced to the coronary artery for coronary angiography to confirm the location and morphology of the target vessel (e.g., left anterior descending artery or right coronary artery). High-pressure balloon angioplasty was then performed, with the appropriate balloon catheter selected based on the vessel diameter, typically 1.2 to 1.5 times the diameter of the target vessel. The balloon catheter was advanced to the target coronary artery, and high-pressure inflation (typically 8–12 atm) was applied at the stenotic site, with each inflation lasting 10–20 s until the stenosis was fully dilated. After balloon dilation, coronary angiography was repeated to confirm vessel patency. Postoperatively, the animals were administered antibiotics (e.g., penicillin) and antiplatelet drugs (e.g., aspirin, clopidogrel) to prevent thrombus formation.

Experimental grouping

A total of 20 miniature pigs were randomly rolled into four groups: S1 (low-dose rapamycin-eluting stent), S2 (medium-dose rapamycin-eluting stent), S3 (high-dose rapamycin-eluting stent), and D0 (bare-metal stent), with 5 pigs in each group.

Stent implantation surgery

Three days prior to surgery, each group of miniature pigs received aspirin (Huainan China Hengcheng Pharmaceutical Group Co., Ltd., National Drug Approval No. H34022418, 100 mg/day) and clopidogrel bisulfate tablets (National Drug Approval No. J20180029, 50 mg/day). Initially, the pigs were anesthetized by subcutaneous injection of 15 mg/kg ketamine hydrochloride (Jiangsu Hengrui Medicine Co., Ltd.) and 0.05 g/L pentobarbital sodium (Shanghai Xinya Pharmaceutical Co., Ltd.). Subsequently, the pigs were positioned supine and securely fixed on the operating table. Small animal electrocardiogram monitoring equipment (Shanghai Yuyan Scientific Instruments Co., Ltd.) was employed to record heart rate, systolic blood pressure, diastolic blood pressure, and other parameters throughout the procedure.

Subsequently, the left femoral artery was punctured utilizing the Seldinger technique, and a 7 F arterial sheath (Antai Guorui Medical Devices Co., Ltd.) was inserted. A bolus of 200 IU/kg heparin was administered through the sheath. Next, a Johnson JR3.5 angiographic catheter (National Medical Device Registration No. 20183032563, Beijing Antai Jiahua Technology Co., Ltd.) was advanced into the coronary arteries for angiography. Based on the angiographic findings, guidewires were guided into the left anterior descending artery and right coronary artery using a ratio of balloon to vessel diameter (1.2:1.0). The balloon was inflated for approximately nine seconds at an appropriate pressure to fully appose the stent against the vessel wall. Completion of the stent implantation procedure was confirmed with contrast imaging to ensure patency of the lumen. Subsequently, the catheter, sheath, and other instruments were removed, and the incision was sutured. The pigs were then returned to their housing facility. Postoperatively, each pig received daily injections of penicillin (3 million units) for three consecutive days. Within 28 days post-surgery, they were administered aspirin (100 mg/day) and clopidogrel bisulfate tablets (50 mg/day).

HE staining

After euthanasia by exsanguination, the hearts of the pigs were carefully extracted. The implanted segments of the vessels were isolated and fixed in 10% formaldehyde solution for 1 day. After one month following stent implantation, the animals were euthanized, and tissue samples were collected for analysis. Subsequently, the specimens were dehydrated through a series of alcohol gradients (70%, 80%, 90%, 95%, 100%) and embedded in Technovit 7100 resin (Shanghai Nuoning Biotechnology Co., Ltd.). Once embedded, tissue blocks were sectioned using a McIlwain tissue slicer (Shanghai Yuyan Scientific Instruments Co., Ltd.). For each stented vessel segment, at least three tissue sections were obtained, corresponding to the proximal, middle, and distal regions.

The obtained tissue sections were immersed in alcohol gradients of varying concentrations (100%, 95%, 90%, 80%, 70%) for 3 min each, followed by rinsing with distilled water. They were then stained with hematoxylin for 5 min, rinsed again with distilled water for 30 s, and differentiated in 75% hydrochloric acid alcohol for 5 min. Subsequently, the sections were stained with eosin for 1 min, dehydrated in alcohol, cleared with xylene, and mounted with neutral resin. High-resolution microscope images were captured, and image analysis software was utilized for measurements. Parameters recorded included neointimal thickness, area surrounded by internal elastic lamina, residual lumen area, and area surrounded by external elastic lamina. Additionally, calculations were performed for neointimal area, medial area, and degree of luminal stenosis.

$$\begin{aligned}&\text{N}\text{e}\text{o}\text{i}\text{n}\text{t}\text{i}\text{m}\text{a}\text{l}\:\text{a}\text{r}\text{e}\text{a}\cr&\quad =Surrounding\:area\:of\:internal\:elastic\:lamina\cr&\qquad-Residual\:lumen\:area\end{aligned}$$

(1)

$$\begin{aligned}&Medial\:\:area\cr&\quad =Surrounding\:area\:of\:outer\:elastic\:lamina\cr&\qquad-Surrounding\:area\:of\:internal\:elastic\:lamina\end{aligned}$$

(2)

$$\begin{aligned}&\text{D}\text{e}\text{g}\text{r}\text{e}\text{e}\:\text{o}\text{f}\:\text{l}\text{u}\text{m}\text{e}\text{n}\:\text{s}\text{t}\text{e}\text{n}\text{o}\text{s}\text{i}\text{s}\cr&\quad=\frac{Pre-stenting\:lumen\:area-Post-stenting\:lumen\:area}{Pre-stenting\:lumen\:area}\cr&\qquad\times\:1\end{aligned}$$

(3)

Degree of vascular injury

The Schwartz scoring system was employed to assess the extent of vascular wall injury in each group of miniature pigs. The scoring was based on the sum of injury scores at each stent strut location and the number of stent struts, calculating the average injury score per vessel. Score 0: intact internal elastic lamina, endothelium loss, mild compression of the media; Score 1: clearly fractured internal elastic lamina, mild compression of the media; Score 2: obvious cracks in the internal elastic lamina, tearing of the media, relatively intact external elastic lamina; Score 3: severe cracks in the internal elastic lamina, sometimes visibility of stent struts within the outer membrane.

$$\begin{aligned}&\text{A}\text{v}\text{e}\text{r}\text{a}\text{g}\text{e}\:\text{i}\text{n}\text{t}\text{e}\text{g}\text{r}\text{a}\text{l}\:\text{o}\text{f}\:\text{v}\text{a}\text{s}\text{c}\text{u}\text{l}\text{a}\text{r}\:\text{i}\text{n}\text{j}\text{u}\text{r}\text{y}\cr&\quad=\frac{Damage\:integral\:at\:strut\:of\:each\:support}{Number\:of\:support\:poles}\cr&\qquad\times\:1\end{aligned}$$

(4)

Inflammatory response score

The degree of inflammatory response in each group of miniature pigs was evaluated using the coronary artery wall inflammation scoring criteria [19]. The scoring system was as follows: 0 points indicated an intact internal elastic lamina with endothelial denudation and compression of the media; 1 point represented significant fracturing of the internal elastic lamina with media compression; 2 points indicated prominent fissures in the internal elastic lamina, tearing of the media, and relatively intact external elastic lamina; and 3 points denoted severe fracturing of the internal elastic lamina, with visible metal stent struts (stent supports) within the adventitia, causing disruption of the vessel wall.

Notably, stent struts are the metal supporting components of the stent, typically comprising bars or rod-like structures that maintain vessel patency by preventing collapse.

Vascular endothelialization score

The endothelialization score of each group of miniature pigs was evaluated based on the area surrounded by endothelial cells around the vessel circumference. Score 0: endothelial cells surround less than one-quarter of the vessel circumference; Score 1: endothelial cells surround the vessel circumference between one-quarter and three-quarters; Score 3: endothelial cells surround more than three-quarters of the vessel circumference.

Biochemical indicators

Blood samples were collected from each group of miniature pigs to measure biochemical indicators including alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), serum creatinine (Scr), creatine kinase (CK), and creatine kinase-MB isoenzyme (CK-MB).

Statistical analysis

The data analysis utilized SPSS 22.0. Normally distributed quantitative data were denoted as mean ± standard deviation (𝑥̄±𝑠), while categorical data as frequencies and percentages (%). Non-normally distributed quantitative data between groups were compared utilizing the Mann-Whitney U test, and normally distributed quantitative data were assessed employing one-way analysis of variance. Group comparisons for categorical data were conducted adopting χ² test. Due to the multiple significance tests conducted in this study, Bonferroni correction was applied to adjust the p-values, thereby controlling the Type I error rate. Additionally, when applicable, the Steel-Dwass test was utilized to address multiple comparison issues in non-parametric data. P < 0.05 (two-tailed) meant statistically significant.

Results

Comparison of vascular measurement parameters among different groups of piglets

Figures 1 and 2 show that, in the S2 and S3 groups of pigs, the neointimal thickness, neointimal area, and degree of luminal stenosis were notably inferior to those in the S1 and D0 groups (P < 0.05). However, the residual luminal area was greatly larger in the S2 and S3 groups versus the S1 and D0 groups (P < 0.05). The internal elastic lamina area, external elastic lamina area, and medial area differed slightly when the S2 group was compared with the S1 and D0 groups (P = 0.059). The neointimal thickness and neointimal area in the S2 group pigs were slightly inferior to those in the S3 group, while the residual luminal area was slightly larger, although these differences did not reach statistical significance (P = 0.082).

Fig. 1

Comparison of neointimal thickness, residual lumen area, internal elastic lamina area, and neointimal area among the groups of miniature pigs. (Data were presented as mean ± standard deviation (SD). A-D represent neointimal thickness, residual lumen area, internal elastic lamina area, and neointimal area, respectively.) Note: * indicates marked differences versus the S1 group (P < 0.05); # indicates drastic differences versus the D0 group (P < 0.05) (in Figs. 1, 2 and 4)

Fig. 2

Comparison of external elastic lamina area, medial area, and luminal stenosis among the groups of miniature pigs (A-C represent external elastic lamina area, medial area, and luminal stenosis, respectively)

Scoring of cardiovascular injury severity in each group of piglets

Figure 3 shows that the cardiovascular injury scores were 0.71 ± 0.23 for the S1 group, 0.66 ± 0.25 for the S2 group, 0.59 ± 0.18 for the S3 group, and 0.64 ± 0.15 for the D0 group. Statistical analysis revealed neglectable differences in cardiovascular injury scores among the four groups (P = 0.072).

Fig. 3

Comparison of cardiovascular injury scores among different groups of piglets

Inflammatory response score of piglets in each group

Figure 4 shows that the coronary artery wall inflammation scores were 1.18 ± 0.14 for the S1 group, 0.62 ± 0.11 for the S2 group, 0.71 ± 0.09 for the S3 group, and 1.09 ± 0.22 for the D0 group. Comparison shows that the coronary artery wall inflammation scores in the S2 and S3 groups were markedly lower than those in the S1 and D0 groups, with statistical significance (P < 0.05).

Fig. 4

Comparison of coronary artery wall inflammation scores among the groups of pigs

Scoring of endothelialization of blood vessels in each group of piglets

Figure 5 shows that the endothelialization scores of pigs in the S1 group were 2.51 ± 0.38, in the S2 group were 2.44 ± 0.26, in the S3 group were 2.39 ± 0.24, and in the D0 group were 2.60 ± 0.31. Comparison showed neglectable differences in endothelialization scores between any two groups of pigs (P = 0.085).

Fig. 5

Comparison of endothelialization scores of piglets in different groups

Blood biochemical indicators of piglets in each group before and after surgery

Figure 6 shows that the levels of ALT and AST in the liver function indicators of the four groups of pigs showed fluctuations before and after surgery, but the differences were neglectable (P > 0.05) when comparing pre-operative and post-operative levels within each group. Additionally, there were slight differences in ALT and AST levels among the four groups of pigs post-operatively (P = 0.074).

Fig. 6

Comparison of pre- and post-operative liver function indicators in different groups of miniature pigs. (A: ALT; B: AST)

Figure 7 shows that the levels of BUN and Scr in the four groups of miniature pigs showed some fluctuations before and after surgery, but the differences in comparisons before and after surgery were inconsiderable (P = 0.052). Additionally, BUN and Scr levels differed slightly among the four groups of miniature pigs after surgery (P > 0.078).

Fig. 7

Comparison of pre- and post-operative renal function indicators in different groups of miniature pigs. (A: BUN; B: SCr)

Based on Fig. 8, the levels of myocardial injury biomarkers CK and CK-MB in miniature pigs showed fluctuations before and after surgery, with no considerable differences observed in the comparisons before and after surgery (P = 0.095). Similarly, there existed neglectable differences in CK and CK-MB levels among the four groups of miniature pigs after surgery (P = 0.64).

Fig. 8

Comparison of pre- and post-operative myocardial injury biomarkers in various groups of miniature pigs. (A: CK; B: CK-MB)

Discussion

Currently, the treatment of coronary artery stenosis requires making rational choices based on different disease conditions. Mild stenosis can be managed with medication, primarily aimed at reducing myocardial oxygen consumption and controlling lipid levels to improve symptoms. When coronary artery stenosis exceeds 70%, coronary artery intervention can be considered, involving balloon angioplasty or the placement of drug-eluting stents to control the disease [20, 21]. Ideally, drug-eluting stents should exhibit excellent biocompatibility in their application as carriers, ensuring minimal local inflammatory reactions or mild responses, and avoiding complications such as thrombus formation and stent thrombosis. Therefore, selecting high-quality drug-eluting stents is crucial for effective treatment [22]. The study randomly divided 20 miniature pigs into four groups: S1 (low-dose rapamycin-eluting stent), S2 (medium-dose rapamycin-eluting stent), S3 (high-dose rapamycin-eluting stent), and D0 (bare metal stent), with 5 pigs per group. The direct effects of different drug-eluting stents were analyzed post-implantation using intravascular ultrasound. From vascular measurement parameters, it was observed that in S2 and S3 groups, the neointimal thickness, residual lumen area, neointimal area, and degree of luminal narrowing were drastically inferior to those in S1 and D0 groups (P < 0.05). However, the area surrounded by internal elastic lamina, area surrounded by external elastic lamina, and medial area differed inconsiderably between S1 and D0 groups (P > 0.05). Similar to the findings of Lee et al. (2009) [23], our results suggested that relative to low-dose rapamycin, medium and high doses of rapamycin-eluting stents effectively suppress neointimal hyperplasia in miniature pigs, demonstrating significant preventive effects in coronary artery stenosis. The poorer performance of the low-dose rapamycin-eluting stent may be attributed to its lower drug concentration, which may not reach the biological threshold required for efficacy [24]. Additionally, it was observed that the neointimal thickness and area in the S2 group were slightly lower than those in the S3 group, while the residual lumen area was slightly larger in the S2 group compared to the S3 group, although these differences were not statistically significant. The reason for this may be related to the mechanism of action of higher concentrations of rapamycin in local tissues. Higher concentrations of rapamycin may exert a stronger antiproliferative effect, thereby inhibiting neointimal hyperplasia. However, this may also induce local inflammatory responses or other side effects, which could have influenced the neointimal thickness and area to some extent. Furthermore, higher rapamycin concentrations may reduce neointimal formation by inhibiting smooth muscle cell migration and proliferation, while potentially leading to a relative increase in lumen size. Although these differences were not statistically significant, they suggest the possibility of an optimal dose range for rapamycin concentration in achieving therapeutic effects. Further studies will help validate the optimal concentration for the prevention and treatment of coronary artery stenosis.

Liu et al. (2022) [25] evaluated the effects of Biomagicrapamycin eluting bioresorbable stents on a porcine coronary artery model and found that these stents exhibited similar efficacy to Firebird2 rapamycin-eluting cobalt-chromium alloy stents in coronary arteries with mild to moderate inflammation. In this study, it was observed that the coronary artery wall inflammation scores in the S2 and S3 groups of miniature pigs were markedly lower than those in the S1 and D0 groups (P < 0.05). These findings are consistent with the aforementioned study, suggesting that medium and high doses of rapamycin-eluting stents effectively inhibit local vascular inflammation in miniature pigs with coronary artery stenosis, demonstrating anti-inflammatory effects. Furthermore, this study found that pairwise comparisons of cardiovascular injury scores (P = 0.072) and endothelialization scores (P = 0.085) among the four groups of miniature pigs did not show statistical significance (P > 0.05). This finding is consistent with the results of Diaz-Rodriguez et al. (2021) [26], who investigated the use of cobalt-chromium discs and stents coated with CD31 mimetic peptides in coronary artery implantation in pigs. Their study suggested that medium and high doses of rapamycin-eluting stents promote vascular homeostasis and arterial wall healing in pigs with coronary artery stenosis, preventing stent narrowing and thrombosis formation, thus potentially enhancing the biocompatibility of metal stents. Comparison of blood biochemical indicators before and after surgery in the various groups of miniature pigs revealed neglectable differences (P > 0.05) in liver function indicators (ALT, AST), kidney function indicators (BUN, SCr), and myocardial injury markers (CK, CK-MB). This further confirms the safety of rapamycin-eluting stents in the treatment of coronary artery stenosis, as they do not impair liver and kidney function or myocardial cells in the model.

Conclusion

This study randomized 20 miniature pigs into four groups: S1 (low-dose rapamycin-eluting stents), S2 (medium-dose rapamycin-eluting stents), S3 (high-dose rapamycin-eluting stents), and D0 (bare-metal stents), with 5 pigs per group. The effects of different drug coatings on stents were analyzed using intravascular ultrasound imaging after implantation. The results indicated that relative to low-dose rapamycin, medium and high-dose rapamycin-eluting stents effectively inhibit neointimal hyperplasia and local vascular inflammatory reactions in miniature pigs, significantly preventing coronary artery stenosis. Importantly, these stents do not induce damage to liver function, kidney function, or myocardial cells, demonstrating high safety. However, this study did not establish clear criteria for rapamycin dosage, and differences in the therapeutic effects of medium and high doses of rapamycin were observed. Future research will further refine drug concentrations and explore optimal dosing strategies for rapamycin. In summary, this study provides a theoretical basis for the prevention and treatment of coronary artery stenosis.