Obesity is linked to insulin resistance and metabolic syndrome and contributes to conditions such as hypertension, high serum cholesterol, low HDL cholesterol, and elevated blood sugar levels. It also independently increases the risk of cardiovascular diseases, type II diabetes, and certain cancers. Moreover, excess fat accumulation in adipose tissues prompts these tissues to release inflammatory substances, leading to a pro-inflammatory state and oxidative stress [3].

To date, no research has investigated the effects of green coffee (GC) supplementation on miRNA-133a, miRNA-155, and various inflammatory markers associated with obesity and metabolic syndrome. This study is the first to examine this potential, providing new insights into how GC supplementation might influence these biomarkers and aid in managing obesity-related health problems. To ensure that all participants receive adequate nutrition, both the placebo and green coffee groups were provided with a balanced diet to meet their needs for vitamins, minerals, and macronutrients.

In this study, patients who received 800 mg of GC daily for 6 months, alongside with a balanced diet, showed improvements in BMI, blood pressure, blood glucose levels, and HOMA-IR, and experienced a correction in dyslipidemia compared to those in the placebo group. Moreover, numerous studies in both humans and animals have highlighted the positive effects of green coffee on glucose and lipid metabolism, benefiting both healthy individuals and those with genetic metabolic disorders [39] which were in accordance with our findings. Moreover, there was heterogeneity between studies when considering the effect of GC on HOMA-IR status. However, Meng et al. [40] reported that a dose greater than 400 mg of GC significantly decreases HOMA-IR.

GC contains chlorogenic acid (CGA) as its primary phenolic compound, which is known for its antioxidant properties. CGA functions as a hypoglycemic agent through several mechanisms: (a) it enhances insulin sensitivity and action, similar to metformin; (b) it inhibits the activity of hepatic glucose-6-phosphatase; (c) it reduces intestinal glucose absorption; and (d) it stimulates glucose uptake in both insulin-sensitive and insulin-resistant adipocytes. Additionally, unlike thiazolidinediones or insulin, CGA does not lead to weight gain or other adverse side effects [41].

The impact of GC on lipid profile is primarily driven by CGA. CGA lowers total cholesterol by inhibiting lipid and cholesterol absorption in the intestines, reducing their transfer, and limiting hepatic biosynthesis [20]. Additionally, CGA enhances the expression of the PPAR-α gene and increases levels of carnitine palmitoyltransferase-1, while decreasing the expression of lipogenic factors such as sterol regulatory element-binding proteins (SREBPs). These SREBPs are crucial in regulating genes involved in glucose and lipoprotein metabolism, as well as liver inflammation [42, 43]. Clinically, synthetic PPAR-α agonists are used to treat dyslipidemia by lowering triglyceride levels and raising serum HDL-C levels, similar to the effects observed with CGA [20].

The current study found that markers of inflammation—specifically total sialic acid, homocysteine, resistin, TNF-α, hs-CRP—and the oxidative stress biomarker MDA were elevated at baseline compared to standard reference values, whereas adiponectin levels were lower. Additionally, a positive correlation was observed among these biomarkers.

Obesity is associated with a chronic, low-grade inflammatory state, characterized by elevated levels of pro-inflammatory cytokines such as TNF-α, IL-6, and CRP [44]. Macrophages` infiltration to fat tissue leads to overproduction of pro-inflammatory chemokines. This results in aggravation of localized inflammation in adipose tissue and spreading of an overall systemic inflammation that is associated with the development of obesity-related comorbidities [45].

Furthermore, obese patients show increased activity of liver c-jun N-terminal kinase, which triggers the expression of inflammatory cytokines. These cytokines then enhance the activation of transcription factors like activator protein-1, nuclear factor-κB (NF-κB), and interferon regulatory factors, leading to a reduction in insulin sensitivity [46].

In this study, administering 800 mg of GC along with a balanced diet for six months led to a decrease in total sialic acid levels compared to a placebo. This effect may be linked to improvements in lipid profile, glucose regulation, or insulin sensitivity. Additionally, the CGA component in GC helps reduce inflammation and oxidative damage, which in turn lowers the release of sialic acid.

Sialic acid (SA), also known as N-acetyl-Neuraminic acid, is released from glycoconjugates by neuraminidase and plays various physiological roles. Inflammatory and oxidative stress conditions stimulate hepatocytes to produce SA, which then acts as a signaling molecule. This signaling can trigger myocardial injury by activating the Rho/ROCK-JNK/ERK signaling pathway [45]. Increased levels of sialic acid (SA) are linked to dyslipidemia, insulin resistance, and immune responses. Additionally, desialylated LDL is more susceptible to oxidative modification and greater accumulation compared to native LDL, which plays a role in the development of atherosclerosis [46].

Hyperhomocysteinemia accelerates atherothrombosis by elevating oxidative stress and impairing vascular endothelial function. Furthermore, it has been reported that homocysteine induces insulin resistance in vitro by inhibiting insulin signaling, with this effect being mediated by oxidative stress [47]. Herein, the present study showed a significant decrease in homocysteine level in coffee green group. These results were consistent with Ochiai et al. [48], who found that ingestion of GC decreases blood homocysteine level and improve vascular endothelial function.

Resistin is a peptide hormone secreted from human monocytes and macrophages and is involved in insulin resistance by impairing glucose tolerance. Moreover, resistin is correlated to abdominal fat depots and inflammation mediated by macrophages [49]. In this study, supplementation with GC for 6 months led to a reduction in resistin levels. These findings align with the work of Hwang et al. [50] and Huang et al. [51], who reported that CGA has anti-inflammatory effects by down-regulating iNOS, IL-1β, TNF-α, IL-6, and the chemokine CXCL1. Additionally, our results demonstrated a positive correlation between resistin and each of TNF-α, hs-CRP, total sialic acid, and homocysteine. This is consistent with the findings of Malo et al. [52], who observed that pro-inflammatory cytokines such as IL-1, IL-6, and TNF-α increase resistin expression in human peripheral blood mononuclear cells.

In the present study, hs-CRP levels were measured both at baseline and after six months of GC supplementation, compared to a placebo. CRP, an acute-phase protein, is released in response to inflammation triggered by IL-6 secretion and is considered an additional marker of metabolic syndrome, with elevated levels often seen in obesity and insulin resistance [43]. Our findings indicated that GC supplementation significantly reduced CRP levels. This reduction is likely due to CGA's ability to suppress macrophage infiltration and inhibit the production of inflammatory mediators by down-regulating NF-κB [51, 53]. Moreover, CGA diminished the expression of pro-inflammatory markers like TNF-α and IL-6, ROS and RNS in type 1  and 2 diabetes in several animal and human studies.

The present study found that supplementation with GC significantly reduced TNF-α levels compared to the placebo group. Additionally, TNF-α levels were positively correlated with other inflammatory biomarkers. These findings are consistent with the work of Tzanavari et al. [54], who reported that obesity leads to macrophage infiltration in adipose tissue and increased production of the pro-inflammatory cytokine TNF-α, which correlates with adiposity and insulin resistance. Targeting TNF-α and/or its receptors may therefore be a promising approach for treating type II diabetes and insulin resistance [51, 54]. Furthermore, CGA has been shown to exert cardioprotective effects by inhibiting the Nrf2/HO-1 and TGF-β/Smads signaling pathways [53].

GC has the ability to scavenge free radicals and enhance antioxidant capacity both in vivo and in vitro. This effect was supported by our study, which found that serum levels of MDA, a marker of lipid peroxidation, were significantly lower after six months of GC supplementation (800 mg daily) compared to the placebo. The antioxidant properties of CGA are primarily mediated through the transcription factor nuclear factor-E2-related factor 2 (Nrf2), which regulates phase II detoxifying enzymes, including superoxide dismutase, glutathione peroxidase, and glutathione reductase [51, 53, 55]. The antioxidant activity of GC plays a crucial role in preventing the oxidation of LDL-C, particularly in patients with dyslipidemia [55, 56].

In the present study, serum adiponectin level was increased by GC supplementation compared with placebo. These results were consistent with Lukitasari et al. [57], who reported that CGA can induce the transcriptional activity of PPAR-γ and consequently adiponectin production by adipose tissue. Moreover, chlorogenic acid is a potential agonist of PPAR-γ regulating glucose homeostasis and increasing insulin sensitivity of peripheral tissues thus preventing type II diabetes [58].

This study is the first to investigate the impact of green coffee bean extract supplementation on miR-133a and miR-155. Recently, miRNAs have been identified as key biological regulators with the potential to influence inflammation through various pathways [59]. Our findings revealed a significant reduction in miR-155 levels following six months of green coffee treatment compared to the placebo group. Additionally, miR-155 showed a significant correlation with fasting blood glucose (FBG), HbA1c, HOMA-IR, total sialic acid, and miR-133a. Prior to the intervention, miR-155 was up-regulated in obese patients. These results align with previous studies indicating that miR-155 contributes to adipose tissue dysfunction and insulin resistance [60, 61]. Furthermore, the study found that elevated miR-155 levels were associated with increased resistin, hs-CRP, TNF-α, and total sialic acid. These observations are consistent with earlier research showing that resistin levels are notably higher in the plasma of miR-155-/-/ApoE-/- obese mouse models compared to ApoE-/- mice, indicating a miR-155-mediated suppression of inflammation in adipose tissue [62].

Ohishi et al. [63] discovered that CGA may block CD36 via AMPK activation, resulting in decreased lipid absorption and transport. CGA also raised miR-122 levels, a liver-specific miRNA that is crucial for liver homeostasis. This indicated that CGA may suppress lipogenesis and fatty acid synthase via post-transcriptional mechanisms [64].

We also assessed miR-133a expression in obese patients with metabolic syndrome (MetS) before and after treatment with green coffee (GC). Initially, miR-133a was up-regulated in these patients, but GC supplementation for six months significantly reduced its levels. Furthermore, serum levels of miR-133a were positively correlated with fasting blood glucose (FBG) and HbA1c. These findings are consistent with previous research [63, 65, 66], which has identified circulating miR-133a as a biomarker for myocardial damage. Elevated serum levels of miR-133a have been observed in patients with severe myocardial injury and adverse cardiovascular events [67]. Additionally, Kim et al. [68] demonstrated that CGA protects against alcohol-induced liver injury in mice by modulating hepatic miRNAs that regulate mitochondrial redox systems.