## 4 Interaction Between the Gut Microbiota and Cardiovascular Diseases
### Trimethylamine N-Oxide  
TMAO is the main diet-derived metabolite, linked to an increased risk of developing atherosclerosis, thrombosis, and stroke. This compound is formed from TMA, a precursor produced in the colon by the action of bacteria from the phylum _Bacillota_, _Pseudomonad_, and _Actinomycetota_, whose species have been previously mentioned [143]. These bacteria convert dietary quaternary amines, such as L-carnitine and choline, present in foods like red meat and eggs, into TMA. Once produced, TMA is absorbed and transported via the portal vein to the liver, where it is converted into TMAO by the FMO3 enzyme. In addition, this metabolite is found in fish and seafood, so their consumption represents an exogenous and direct source of TMAO [144; 145]. Once in circulation, TMAO contributes to endothelial dysfunction, promotes foam cell formation, and enhances platelet activity, which, together, increase the risk of thrombus formation [143]. Endothelial dysfunction is an early and critical step in atherosclerosis. In this context, TMAO is known to activate the NF-kB signalling pathway, partly through a mechanism involving high-mobility group box 1 (HMGB1), a key inflammatory mediator [146; 147]. HMGB1 can be released extracellularly and interacts with the receptor for advanced glycation end-products (RAGE), initiating downstream pro-inflammatory signalling cascades. This interaction promotes the activation of NF-kB, leading to the upregulation of pro-inflammatory genes that encode adhesion molecules, chemokines, and inflammatory cytokines. Additionally, HMGB1/RAGE signalling can activate the NLRP3 inflammasome, further exacerbating the inflammatory response. Thus, TMAO-induced HMGB1 release, and the subsequent RAGE activation, constitute an important pathway, contributing to endothelial inflammation and dysfunction in atherosclerosis [146; 147; 148]. This also decreases the expression of tight junction proteins, such as occludin and zonula occludens-2 (ZO-2), as well as vascular endothelial cadherin in ECs, which increases permeability to LDL, a particle formed in the bloodstream from the metabolic conversion of VLDL secreted by the liver, as shown in Figure 4. Additionally, TMAO upregulates VCAM-1 and ICAM-1 expression, promoting monocyte adhesion [146]. Concerning foam cell formation, macrophages play a central role by accumulating lipoproteins. In this context, several studies support a crucial role of TMAO in promoting foam cell development [149; 150]. TMAO, which structurally resembles ox-LDL, promotes monocyte differentiation into macrophages and increases the expression of scavenger receptors like CD36 and SRA-1. More recently, [PERSON] and colleagues demonstrated that TMAO downregulates the expression of Nrf2 and its downstream effectors, heme oxygenase\(1\) and glutathione peroxidase \(4\), leading to oxidative stress and the reduced expression of cholesterol efflux proteins, such as ATP-binding cassette transporter A1 (ABCA1) and G1 (ABCG1), thereby promoting lipid accumulation in foam cells [151]. In addition, TMAO has been associated with an inhibition of the reverse cholesterol transport, a key protective mechanism against atherosclerosis, as depicted in Figure 4[138, 151, 152, 153]. Reverse cholesterol transport involves the efflux of excess cholesterol from peripheral cells, such as macrophages, to apolipoprotein A-I (ApoA-I), initiating the formation of nascent HDL particles via their interactions with ABCA1 and ABCG1. These HDL particles mature and deliver cholesterol to the liver through scavenger receptor class B type 1 (SR-B1), either directly or indirectly, via the cholesteryl ester transfer protein (CETP). TMAO interferes with this pathway by downregulating the expression of LXR\(\alpha\), a transcriptional regulator of ABCA1 and ABCG1, thereby impairing cholesterol efflux from macrophages and HDL biogenesis. This disruption compromises reverse cholesterol transporter efficiency and contributes to lipid accumulation and foam cell formation within atherosclerotic plaques [138, 153, 154].