Coronary artery disease (CAD) is the leading cause of morbidity, disability and death worldwide (1). Atherosclerosis as the primary cause of CAD is initiated with the damage and subsequent impairment of the functional integrity of the vascular endothelium (2). Endothelial dysfunction is a result of oxidative stress which occurs concurrently with the activation of pro-inflammatory signaling pathways and the release of cytokines and chemokines (3, 4). The modifiable risk factors for CAD include hypertension, insulin resistance, dyslipidemia, chronic inflammation, obesity and sedentary lifestyle (5, 6). The high cost and side effects of synthetic drugs have drawn attention to the use of dietary supplements and antioxidants, that might be an easy way to restore the redox state and inhibiting the oxidative stress (7, 8).

Astaxanthin (AX) is a fat-soluble carotenoid without provitamin A activity in humans (9). It belongs to xanthophylls with the chemical formula C40H52O4 (10). The natural sources of AX found in various microorganisms and marine animals including microalgae, yeast, salmon, trout, shrimp, lobster, and crayfish (11, 12). One of the most important sources of biological AX is green unicellular microalga H.pluvialis which contains more than 80% AX content in its cells (13, 14). Affirmative impacts of AX are often associated with its anti-inflammatory, antioxidative, and antiapoptotic properties (15). Some of clinical trial studies have found that AX can improve metabolic markers such as lipid profile or glycemic indices (16–18). Urakaze et al. investigated that the supplementation of 12 mg AX for 12 weeks in subjects with prediabetes lead to significant reduction in malondialdehyde-modified low-density lipoprotein, glucose levels after 120 min in oral glucose tolerance test and HbA1c (Hemoglobin A1C) compared with placebo group (19). According to an vivo study, AX consumption in ApoE⁻/⁻ mice model caused a significant decline in total cholesterol (TC) and triglycerides (TG), as well as aortic atherosclerotic plaque levels and aortic cholesterol levels (20). AX has attracted increasing interest as a multi-target pharmacological agent and it has been suggested that AX may be able to be a potential therapeutic agent against atherosclerotic cardiovascular disease (21, 22). However, according to our knowledge, no study has been conducted on the effect of AX supplementation in CAD patients. Therefore, the purpose of this study was to evaluate the effect of AX supplementation on body composition, metabolic and anthropometric parameters in patients with CAD.

In vivo studies have demonstrated that AX intake has cardiovascular protective effects, although the results of human studies regarding the beneficial effects of AX on cardiovascular risk factors are contradictory and limited. To the best of our knowledge, this study was the first randomized, double-blind, placebo controlled, clinical trial investigating the impacts of AX supplementation on metabolic parameters in CAD patients. The findings revealed that AX had no effect on the lipid profile compared to the placebo group, but it should be noted that LDL-C and TC levels decreased significantly in the AX group. However, it had no significant improvements on body composition and other metabolic and anthropometric parameters.

The results of a recent meta-analysis conducted in 2020 are in line with our findings and demonstrate that body weight and BMI changes were not statistically significant (23). Supplementation of 12-week AX (6 or 12 mg/d) in overweight middle-aged and senior subjects did not affect body weight and BMI (24). Choi et al. reported that 20 mg AX administration in overweight subjects for 12 weeks showed the same results (25). According to the food intake data, the energy intake of the patients in both groups decreased, as a result, BMI and body weight decreased in both groups. Despite of adjusting confounding factors, no substantial between-group changes were found. WC and HC are parameters that were only analyzed in this study, even though the differences between the AX-receiving group and the placebo group are not statistically significant.

Alike our results, Roustae Rad et al. investigated that lipid profile did not change significantly through consuming 10 mg AX after 12 weeks in T2DM subjects receiving metformin (26). Also, administration of 12 mg AX for 12 weeks in healthy volunteers including subjects with prediabetes exhibit the same outcomes (19). In contrast, in another clinical trial on T2DM patients revealed that 8 weeks AX administration at the dosages of 6 and 12 mg led to LDL-C reduction and only 12 mg AX cause TG and TC decrease (17). Another study on non-obese subjects with mild hyperlipidemia reported that 12 and 18 mg AX significantly diminish TG levels after 8 weeks (18). Two recent meta-analysis investigated that AX consumption was not associated with TC, LDL-C, TG reduction. However, only Xia reported an overall increase in HDL-C (23, 27). Tominaga et al. and Saito et al. showed that AX had no impact on lipid profile in healthy individuals regardless of the duration of supplementation and the AX dosage (28, 29).

It is well known that the major causative risk factors for CAD and a definite contributor to accelerated atherosclerosis is dyslipidemia (30, 31). Statins are used as the first line of treatment in both primary and secondary prevention to lower cholesterol and enhance lipid composition (32, 33). Therefore, AX might not have affected TC, TG, and LDL-C levels since the majority of the patients were taking statins and lipid profile had improved before the intervention.

However, the TC and LDL-C levels were significantly decreased only in the intra-group analysis of AX group. The capacity of AX to upregulate SREBP-2 and therefore boost the gene expression of LDLR and HMGR may have contributed to the ability of AX to reduce cholesterol levels (34). SIRT1 involve in regulating the homeostasis of the body’s total cholesterol as a positive regulator of the liver X receptor (LXR) (35). It has been demonstrated that SIRT1 also regulates PPARα expression, which helps to control lipid metabolism (36, 37).

The current study also declared that AX intake did not significantly improve serum levels of insulin, FBS, and HOMA-IR index. A recent meta-analysis revealed that FBS did not decrease after AX supplementation compare with placebo group. According to yang et al. study, the plasma levels of glucose in apoE knockout (apoE⁻/⁻) mice, a mouse model for atherosclerosis, which were fed a high-fat/high-cholesterol diet supplemented with AX-rich Hematococcus pluvialis extract for 4 weeks did not improved significantly compared with control group (34). Alike our result, Saito and Tominaga et al. demonstrated that 12 mg, 6 and 12 mg AX did not affect FBS levels in healthy volunteers, respectively (28, 29). Yoshida et al. also reported the similar result which FBS did not change significantly in subjects with mild hyperlipidemia (18). However, the blood glucose levels have dropped significantly or even slightly in trials on diabetic subjects (16, 17).

SIRT1 play an important role in controlling cellular physiological processes (38). It has been identified as a novel homoeostasis regulator in the human cardiovascular system through protecting against aging and endothelial inflammation, inducing resistance against oxidative stress and hypertrophic, inhibiting apoptosis of cardiomyocytes, and modulating cardiac energy metabolism (39, 40). Furthermore, transcription factor SIRT1 plays an important role in energy and lipid metabolism, inflammation and insulin sensitivity (41). Nishida et al. reported that AX treatment induced upregulation of the gene expression of the mitochondrial SRIT1 and mitochondrial biogenesis in mice (42). In our investigation no significant change in serum level of SIRT1 following AX supplementation were observed even after adjusting for confounders. Non-significant alterations in lipid profile and glycemic indices may be associated with negligible changes in SIRT1. The impact of AX on SIRT1 levels has not been studied in any RCT. Furthermore, drawing a definite conclusion on the effects of AX on SIRT1 is not feasible due to the insufficient number of studies in this field.

TNF-α is a pro-inflammatory cytokine found in atherosclerotic lesions and can have a direct effect on vascular endothelial cells and induce the expression of adhesive molecules in leukocytes and other inflammatory cells (43). In our study the changes of TNF-α levels were not significant and in line with our study. Park et al. demonstrated that AX administration at two doses of 2 and 8 mg for 8 weeks did not affect TNF-α in healthy women (44). Although, 6 and 12 mg AX consumption for 8 weeks in diabetic patients could alleviate diabetes-associated systemic inflammatory stress by reducing TNF-α circulation (17). Considering that more than 80% of the participants in our study had normal range of TNF-α at the baseline, a higher dose should probably be used to have a greater impact on TNF-α factor. Alternatively, the result of non-significant change in TNF-α levels in our study is in line with the hypothesis of the recent meta-analysis about the effect of carotenoids supplementation on inflammation, which suggests that higher baseline levels of pro-inflammatory cytokines indicate a potential for a greater response to carotenoid supplementation such as AX (45).

SIRT1 directly can deacetylate the p65 subunit of the NF-κB complex to suppress NF-κB activation. Furthermore, SIRT1 promotes oxidative energy generation by activating of AMP-activated protein kinase (AMPK), Peroxisome proliferator-activated receptor α (PPARα) and Peroxisome proliferator-activated receptor gamma coactivator 1-alpha. These factors concurrently inhibit NF-κB signaling and led to decline inflammation (46). If this pathway is blocked, TNF-α production can also decrease.

The small sample size may be the reason for the lack of noticeable changes in the parameters of present study. The follow-up duration in our study was short and extending the duration of the intervention might show potential benefits. Moreover, we were not able to measure the concentration of AX in the plasma in order to evaluate its bioavailability. Future trials may be able to better understand the impacts of AX by assessing certain genes involved in the regulation of metabolic variables.

The trial was completely conducted using a double-blind, random allocation approach. Patients who had recovered from COVID-19 even before the intervention were excluded from the study due to prevent disturbances in the measured factors, especially the inflammatory index.

The data indicate that the levels of LDL-C and TC have decreased after using AX supplements for 8 weeks, while other metabolic variables have not significantly changed. However, more clinical trials with various AX supplementation doses and durations are required to obtain a definitive conclusion.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The studies involving human participants were reviewed and approved by Tabriz University of medical science. The patients/participants provided their written informed consent to participate in this study.

MH, MA, and MC designed the study. MH, MC, and BS coordinated the collection of the samples and data collection from study participants. MH carried out the statistical analysis and wrote the first draft of the article. MH, MC, MA, BS, and SK participated in revising it critically for important intellectual content. All authors contributed to the article and approved the submitted version.

Tabriz University of Medical Sciences supported this study (grant number 65868).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.