High arterial stiffness is known to be a risk factor, as well as a prognostic marker for cardiovascular disease (CVD) (1). Endothelial dysfunction plays a key role in arterial stiffness by reducing the endothelial properties of vasodilatation, accelerating the proinflammatory and prothrombotic properties (2). Endothelial dysfunction is an early event in patients with CVD and is considered as one of the several potential contributors to plaque destabilization (3).

A number of non-invasive techniques are available to assess the endothelial function and arterial stiffness of the peripheral vasculature, such as brachial artery flow-mediated vasodilatation (bFMD), pulse wave velocity (PWV) and cardio-ankle vascular index (CAVI) (4,5). These non-invasive methods were widely used clinically to predict the risk of subclinical atherosclerosis in individuals who are at a high risk of CVD (6). CAVI is an inexpensive, non-invasive, office-based method which is used to evaluate arterial stiffness in the aorta, femoral artery and tibial artery, which reflects the degree of cardiovascular disease (1,7). The advantage to CAVI is that it is not affected by blood pressure (BP) and the measurements are automatic via validated software (6,8,9).

Our group has demonstrated the cardioprotective effects of AGE by decreasing atherosclerotic plaque progression in patients with type 2 diabetes (10) and improving endothelial function (11,12). Endothelial dysfunction plays a critical role in the pathogenesis of micro- and macrovascular diseases in patients with type 2 diabetes (13). Annual screening is recommended for diabetic patients for the early detection of micro- and macrovascular complications. This study investigated the effects of AGE in individuals with type 2 diabetes on vascular elasticity and endothelial function, which was measured by CAVI, over a period of 3 months.

A total of 65 patients (38 men and 27 women; mean age 58.45±11.25 years) completed 2 study visits (baseline and 3 months), where baseline and follow-up CAVI measures were assessed (Fig. 1). Out of the 65 patients, 37 were randomized to the treatment (AGE; active) group (19 males; mean age, 59.3±10.8 years). Patients current medications, such as aspirin, hypertensive or hyperlipidemia or diabetic medications, did not change during the 3 months of the study. The baseline characteristics of the patients have been published elsewhere (14).

The primary objective of this study was reduction in CAVI over a 3-month period. As shown in Table I, CAVI was significantly reduced on the right arm by 0.64±1.09, relative to the placebo group reduction of 0.11±0.92 (P=0.04). On the left arm, CAVI was trending toward a significant reduction by 0.79±1.58, relative to 0.16±1.05 in the placebo group (P=0.07). The reduction based upon the average of both arms was 0.71±1.27 in AGE vs. 0.13± 0.94 (P=0.04). As shown Table II, the CAVI measures completed on the right and left arms for both visits were highly and significantly correlated, (Pearsons's r for visit 1 = 0.89, P<0.0001; visit 2 = 0.95, P<0.0001) indicating a high degree of consistency in the CAVI measures (Figs. 2 and 3).

To the best of our knowledge, this is the first study to demonstrate the effects of AGE on arterial stiffness by improving endothelial function, which was measured by CAVI over a period of 3 months in patients with type 2 diabetes. Diabetic patients with a high 10-year atherosclerotic cardiovascular disease (ASCVD) risk score have been noted to have high CAVI (>9) (15). CAVI is independent of blood pressure and highly reproducible, which is significantly higher in patients with coronary artery stenosis (1,6–9,16). In comparison to PWV, CAVI reflects the smooth muscle contraction rather than changes in BP, and changes over a short period of time in response to sympathetic tone and pharmacological influences (17). The current study demonstrated AGE reduced the arterial stiffness, which was demonstrated by CAVI, which inturn reduces the risk of coronary events.

Arterial stiffness increases with age through the loss of elastin and collagen fibers, which results in increased blood pressure (18). Kobayashi et al (19) demonstrated the significant association between endothelial dysfunction and increased arterial stiffness. Tomiyama et al (20) demonstrated the significant association of bFMD endothelial measurements with arterial stiffness assessed by PWV. Previous studies have indicated that treatments targeting to reduce arterial stiffness and wave reflections can reduce the risk of CVD along with a reduction in BP (21). Larijani et al (11) demonstrated the beneficial effects of AGE and CoQ10 on vascular elasticity and endothelial function in firefighters (mean reduction of PWV, 1.21; 95% CI, −2.1 to −0.32; P=0.005). Ried and Fakler (22) demonstrated the potential effect of garlic in lowering BP through a meta-analysis, including 20 clinical trials with hypertensive individuals (8–9 mmHg of SBP and 6–7 mmHg of DBP; P<0.0001). Furthermore, Breithaupt-Grögler et al (23) demonstrated the protective effects of garlic extract on arterial stiffness assessed by PWV (active vs. placebo; 8.3±1.46 vs. 9.8±2.45 m/sec; P<0.0001) and pressure-standardized elastic vascular resistance (EVR) (active vs. placebo; 0.63±0.21 vs. 0.9±0.44 m2 · sec-2 · mm Hg-1; P<0.0001) compared to the placebo. The results of the current study are consistent with those of previous studies and the mean of both right and left CAVI was significantly improved in AGE group relative to the placebo (0.71+1.27 vs. 0.13+0.94; P=0.04).

Oxidative stress and systemic inflammation play a key role in endothelial dysfunction, which impairs the pathways leading to the production of endothelial-derived relaxing factors, such as nitric oxide (NO), prostacyclin, tissue plasminogen activator and vasoconstrictors (eg, leukotrienes and endothelin-1) (24). Increased oxidative stress accelerates the production of reactive oxygen species (ROS), leading to the inactivation of two anti-atherosclerotic enzymes, such as endothelial nitric oxide synthase (eNOS) and prostacyclin synthases (25,26). Endogenous NO is a potent vasodilator and is produced by two pathways. The first one is by the oxidation of L-arginine in the vascular endothelium by eNOS and the other is by reducing the dietary nitrate (NO3^-) to nitrite (NO2^-) to NO (27,28). The uncoupling of NOS is a mechanism which plays a critical role in endothelial dysfunction, resulting in the generation of high levels of superoxide (O2^-), leading to the formation of potent oxidant peroxynitrite (ONOO^-), which is highly toxic, damaging biomolecules, including proteins, lipids and DNA (22,29). Previous studies have used bFMD as an index of endothelial NO regulation of vascular tone, and targeting ROS with vitamin C and dietary nitrate supplement improved vascular function in a number of conditions known to be associated with excess oxidative stress (e.g., type II diabetes, hypertension and CAD) (30–32).

AGE contains water soluble S-allyl cysteine (SAC) and S-allymercaptocysteine (SAMC), which have potent antioxidant properties to protect the vascular endothelium from oxidative stress (11) and has also been reported to have a cholesterol-lowering effect (33). The thiol components (γ-glutamylcysteine) of garlic have the ability to reduce blood pressure by modulating NO, H2S and endothelial synthesis as previously described (21,34–36). Furthermore, AGE has been shown to exert an anti-inflammatory effect by decreasing the expression of CD36 on foam cells and oxidized LDL uptake by macrophages (37). Taken together, the antioxidant and anti-inflammatory properties of garlic increase the enzymatic activity of endothelial cells, such as eNOS, catalase, glutathione peroxidase and superoxide reductase to maintain vascular hemostasis, which may be useful for the prevention of CVD (38).

The current study has several limitations. First, the sample size was relatively small and follow-up was relatively short-term to demonstrate the effects of AGE on BP. Second, patients were under different therapies for hyperlipidemia, hypertension and type 2 diabetes mellitus at different doses. Due to our small sample size, a separate analysis by different background medications was not performed.

In conclusion, this study indicates that at the end of 3 months, the change in CAVI was significantly greater in the AGE group than in the placebo group. Further studies however, are required to evaluate whether AGE has the ability to improve arterial stiffness and endothelial function and thereby decrease adverse cardiovascular events.