The endothelial cells are integral to the cardiovascular system and function as gatekeepers of vascular health and homeostasis. The dysfunction of vascular endothelial cells has been recognized as a critical component in the pathophysiology of atherosclerosis [48]. Mediated by endothelial dysfunction, circulating lipoprotein particles enter the artery wall, facilitating the recruitment of monocytes/macrophages and the formation of atherogenic foam cells, ultimately triggering a series of complex pathogenic processes to promote plaque formation [48].

Triggered by various cardiovascular risk factors, endothelial cells undergo morphological and functional modifications, termed endothelial activation, manifesting increased expression of adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), E-Selectin, and chemokines/pro-inflammatory cytokines such as interleukin-8 (IL-8), IL-6, tumor necrosis factor-α (TNF-α), and IL-1β. The results of endothelial activation are increased leukocyte adhesion and infiltration into the vascular wall leading to the propagation and development of vascular inflammation [48].

Nuclear factor κB (NF-κB) is a master transcription factor responsible for inflammatory responses [49]. Peroxisome proliferator-activated receptor Gama (PPARγ) can attenuate inflammatory responses in the cardiovascular system, including endothelial cells. Growing evidence suggests that PPARγ is an upstream regulator of NF-κB in the anti-inflammatory process [50,51]. Eucalyptol is a monoterpene found naturally in many aromatic plants with anti-inflammatory effects. Pre-treatment with eucalyptol has been reported to suppress the expression of VCAM-1, E-selectin, IL-8, and IL-6 in lipopolysaccharide (LPS)-induced human umbilical vein endothelial cells (HUVECs), and this was achieved by blockade of NF-κB signaling. Importantly, by using PPARγ inhibitor or PPARγ gene silencing, LPS-induced activation of NF-κB and expression of inflammatory mediators in HUVECs could be reversed, suggesting that modulation of the PPARγ/NF-κB axis contributes to eucalyptol-mediated suppression of endothelial pro-inflammatory activation [52]. Similarly, several studies have shown that monoterpenes, including citral, citronellol, and genipin, can inhibit adhesion molecule expression in HUVECs and neutrophil/monocyte–endothelial cell adhesion by regulating the PPARγ-dependent NF-κB signaling pathway [53,54,55]. High mobility group box 1 (HMGB1) is a damage-associated molecular pattern (DAMP), secreted from endothelial cells and leukocytes that mediate inflammation, and correlates with the severity of atherosclerosis [56]. Paeoniflorin and cornuside, two natural monoterpenoids used in traditional oriental herbal medicine, were found to suppress the expression and release of HMGB1 in LPS- or lysophosphatidylcholine (LPC)-stimulated HUVECs, paralleling with reduced expression of endothelial cell-derived adhesion molecules and inflammatory factors [57,58]. The underlying mechanism may involve the induction of sirtuin 1 (SIRT1), a nicotinamide adenine dinucleotide–dependent protein deacetylase, which plays an important role in deacetylation of HMGB1 and thus inhibits HMGB1 release and subsequent NF-κB activation in HUVECs [57]. Additionally, reactive oxygen species (ROS) have been reported to induce the activation of NF-κB to promote the adhesiveness of endothelial cells [59]. Using a high glucose-induced HUVECs model, Wang et al. showed that geniposide exhibited a beneficial role in normalizing endothelial pro-inflammatory activation by inhibiting ROS overproduction, NF-κB activation, and monocyte–endothelial cell adhesion [60]. Recently, molecular docking analysis combined with in vitro cell culture-based approaches showed that amarogentin isolated from Gentianaceace plants could directly interact with AMPK to block NF-κB-mediated endothelial inflammation, indicating the pivotal role of AMPK/NF-κB in the protective effect of amarogentin on endothelial activation [26]. Notably, many monoterpenes with NF-κB inhibitory effects, such as cornuside, paeoniflorin, and geniposide, can attenuate endothelial activation through mitogen-activated protein kinase (MAPK) signaling (p38 MAPK, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK)), another major signaling pathway driving inflammatory response [57,61,62].

Endoplasmic reticulum (ER) stress, autophagy, and inflammation are tightly integrated biological processes in atherosclerosis’s pathogenesis [79]. ER homeostasis is crucial in determining cell survival or death based on the cellular stress factors present. When stimulated by pathological insults, intracellular unfolded protein responses can be induced to protect ER homeostasis. However, excessive and prolonged stimuli disturbing ER homeostasis can cause persistent ER stress responses, triggering inflammatory cascades and cell death events [79]. Thus, ER stress is acknowledged as a danger signal for inflammation. Paeoniflorin extracted from the traditional Chinese herb Paeonia lactiflora was reported to be able to modulate ER stress during endothelial pro-inflammatory activation [65]. Pre-treatment with paeoniflorin significantly reduced the expression of ER stress markers glucose regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), and spliced X-box binding protein-1 (XBP-1), as well as improved ultrastructural abnormalities of the ER. These events ultimately contributed to the attenuation of NF-κB-dependent production of inflammatory mediators in LPS-induced HUVECs [65]. Similarly, catalpol, another natural monoterpene, could also alleviate endothelial inflammation, at least in part, by inhibiting ER stress [66]. Autophagy is an evolutionarily conserved lysosomal catabolic process engaged in degrading dysfunctional or surplus protein aggregates and organelles to maintain cellular homeostasis. Nevertheless, aberrant autophagy causes detrimental effects on cellular homeostasis and thus leads to the induction of inflammation and facilitates the pathophysiology of atherosclerosis [79]. In addition to regulating ER stress, paeoniflorin was found to promote autophagy and attenuate endothelial pro-inflammatory activation through a SIRT1-dependent mechanism [64].

Oxidative stress refers to an imbalance favoring the production of ROS over intrinsic antioxidant mechanisms, which leads to extensive cellular and molecular damage. Endothelial cells are susceptible to risk factors of oxidative stress, such as oxidized lipids, homocysteine, angiotensin II, hyperglycemia, and inflammatory mediators. These risk factors result in the overproduction of ROS followed by endothelial cell damage, dysfunction, pro-inflammatory activation, and apoptosis, eventually promoting the development of atherosclerosis [80]. Accordingly, targeting endothelial oxidative stress by attenuating ROS overproduction and/or improving intracellular antioxidant activity would be beneficial for protecting against endothelial oxidative injury and atherosclerosis.

Studies have demonstrated that many monoterpenes can alleviate endothelial oxidative stress by either inhibiting pro-oxidant enzymes or by enhancing antioxidant enzymes to maintain cellular redox balance. For example, geraniol, paeoniflorin and harpagoside could suppress endothelial ROS production through the downregulation of NADPH oxidases (NOXs) and cyclooxygenase (COX), which are the predominant sources of ROS in the vasculature [81,82,83]. Other monoterpenes, such as thymoquinone, perillaldehyde, citronellal, geniposide, and monotropein, mitigate endothelial oxidative stress by strengthening the activities of antioxidant enzymes, including superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) [67,77,84,85,86].

The nuclear factor E2-related factor 2 (Nrf2) is a central transcription factor responsible for intracellular redox homeostasis [80]. Under basal conditions, Nrf2 binds to Kelch ECH associating protein 1 (Keap1) in the cytoplasm, promoting Nrf2 ubiquitination and degradation. Upon oxidative stress stimulation, Nrf2 dissociates from Keap1 and translocates into the nucleus, thereby inducing the expressions of phase II detoxifying enzymes and antioxidant enzymes for cellular defense [80]. A recent study demonstrated that paeoniflorin, a traditional Chinese herbal substance belonging to monoterpenoid, suppressed mitochondrial ROS production and restored mitochondrial functional damage in tert-butyl hydroperoxide (TBHP)-induced HUVECs [90]. Furthermore, a mechanistic study indicated that paeoniflorin directly interacted with cytoplasm Nrf2, resulting in the nuclear translocation of Nrf2 and activation of Nrf2-mediated antioxidant signaling, thus relieving TBHP-induced endothelial oxidative stress [90]. Another traditional Chinese herbal component with atheroprotective effects, geniposide, was found to attenuate H₂O₂-induced endothelial oxidative stress by activating the Nrf2 antioxidant pathway. Different from the paeoniflorin mentioned above, geniposide stimulated Nrf2 nuclear translocation by modulating AMPK/mechanistic target of rapamycin (mTOR) signaling [68]. Similarly, monoterpenes, such as eucalyptol, geraniol, and catalpol, have also been shown to have anti-oxidative stress effects via Nrf2 activation [42,87,92]. miR-21 is an endogenous miRNA implicated in various pathophysiological processes, including cardiovascular disorders [99]. Accumulating evidence suggests that the miR-21/phosphatase and tensin homolog (PTEN) pathway regulates cell survival and death in the cardiovascular system, including endothelial cells, which makes miR-21/PTEN a therapeutic target for cardiovascular disease [100,101]. Zhou et al. reported that pre-treatment with geniposide decreased ox-LDL-induced oxidative stress in HUVECs by reducing NOX2 expression and by upregulating antioxidant enzyme activities [67]. Using gain- and loss-of-function approaches, the investigators further revealed that geniposide could modulate the miR-21/PTEN pathway to restore the balance between intracellular oxidant and antioxidant states, thus preventing ox-LDL-induced endothelial oxidative injury [67]. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a well-known receptor for ox-LDL that plays a vital role in atherosclerosis. Homocysteine, a risk factor of atherosclerosis, has been reported to induce the expression of LOX-1 in endothelial cells and to promote ROS generation and oxidative injury [102,103]. Picroside II, a monoterpenoid isolated from traditional Chinese medicine Picrorhiza scrophulariiflora, was reported to counteract homocysteine-induced LOX-1 expression in a SIRT1-dependent manner and thus ameliorated HUVEC oxidative stress [76]. In addition, catalpol and monotropein, two naturally occurring monoterpenoids, were found to attenuate endothelial oxidative stress, at least in part, by inhibiting NF-κB activation, suggesting inflammation as a target of these compounds to regulate oxidative stress [66,77].

Endothelium-derived NO plays essential roles in normal endothelial functions. NO is a multifunctional signaling molecule controlling cardiovascular homeostasis, including regulating vasomotor tone, modulation of platelet activation and leukocyte adhesion, and manipulating local cell growth [104]. All established risk factors for atherosclerosis, such as hyperlipidemia, diabetes mellitus, hypertension, and smoking, are found to be associated with diminished NO production [105]. Emerging evidence has also indicated that NO bioavailability dysfunction is implicated in the initiation and development of atherosclerosis [105]. Therefore, improvement of endothelial NO production provides a potential strategy for preventing and managing atherosclerosis.

Abundant evidence has demonstrated the implication of modulating the NO pathway in the effect of natural monoterpenes on endothelial dysfunction and atherosclerosis. NO is synthesized by nitric oxide synthase (NOS) using L-arginine as the substrate [104]. NOS includes different isoforms that play contrasting roles in atherosclerosis, with endothelial NOS (eNOS) being atheroprotective and inducible NOS (iNOS) being pro-atherogenic [105]. Thus, modulating NO by adjusting the eNOS/iNOS ratio is crucial for protecting against endothelial dysfunction and atherosclerosis. Catalpol, a monoterpenoid extracted from the root of the traditional Chinese herb Rehmanniae radix, could maintain the balance of endothelial NO by inhibiting the NF-κB/iNOS pathway and by activating the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt)/eNOS pathway. As a result, catalpol improved cell viability, enhanced endothelial integrity, and reduced inflammatory response in advanced glycation end-product (AGE)-treated endothelial cells [94]. Furthermore, two other monoterpenoids, paeoniflorin and eucalyptol, can upregulate SIRT1 expression to regulate the eNOS/iNOS ratio and to protect against endothelial dysfunction [88,93]. Notably, the activation of eNOS by these monoterpenoids depends on either eNOS Ser1177 phosphorylation [94] or upregulation of eNOS expression [88,93]. Oxidative stress plays a crucial role in determining NO bioavailability [104]. During endothelial dysfunction, oxidative stress mediates ONOO⁻ formation and oxidizes eNOS cofactor tetrahydrobiopterin (BH4), causing BH4 deficiency and eNOS uncoupling. Ultimately, it leads to increased ROS generation and reduced NO bioavailability [104]. Perillaldehyde, a major component in essential oil isolated from Perilla frutescens that has been used in traditional Chinese medicine, was reported to prevent endothelial dysfunction and to attenuate the growth of atherosclerosis [85]. Mechanically, perillaldehyde alleviated ROS-mediated oxidative stress and rescued BH4 deficiency, thereby promoting eNOS recoupling and improving endothelial dysfunction [85]. Other monoterpenoids, such as citronellal and aucubin, similarly induced eNOS recoupling and ameliorated vascular endothelial injury through an anti-oxidative mechanism [91,98].

Apoptosis is a well-known programmed cell death pathway involved in various physiological and pathological processes. Two molecular pathways are acknowledged to regulate apoptosis: mitochondrial-dependent intrinsic pathway and death receptor-mediated extrinsic pathway. The mitochondrial-dependent apoptosis pathway is induced by cellular damage or stress, which upregulates B cell leukemia/lymphoma 2 (BCL2)-associated agonist of cell death (Bad) and facilitates the insertion of BCL2-associated X protein (Bax) into the mitochondrial outer membrane. This further causes cytochrome c to be released into the cytoplasm, followed by interaction between cytochrome c and apoptotic peptidase activating factor 1 (Apaf-1) to induce apoptosis through caspase activation [106]. The extrinsic apoptotic pathway is initiated by binding extracellular death ligands (such as TNF-α) to their respective cell-surface death receptors. The activation of the death receptors results in the formation of the death-inducing signaling complex mediated by an adaptor protein, thereby triggering caspase activation and the apoptosis process [107]. While apoptosis serves as a fundamental mechanism for physiological homeostasis, uncontrolled apoptosis may lead to cellular dysfunction and death. It has been observed that increased endothelial apoptosis is closely associated with atherosclerosis, and pro-atherosclerotic factors such as ox-LDL, oxidative stress, and low shear stress have been shown to induce vascular endothelial cell apoptosis [106,108]. Therefore, inhibition of endothelial apoptosis may represent a promising strategy to cope with atherosclerosis.

Accumulating evidence demonstrates the reduction of endothelial apoptosis as one of the mechanisms by which natural monoterpenes protect against endothelial injury and atherosclerosis. The anti-apoptotic effect of monoterpenes involves the modulation of intrinsic and/or extrinsic apoptotic pathways in endothelial cells. For instance, the beneficial impact of geniposide against atherosclerosis and endothelial dysfunction was related to the attenuation of endothelial apoptosis due to upregulation of anti-apoptotic protein Bcl-2, downregulation of pro-apoptotic protein Bax and caspase-3, and maintaining mitochondrial membrane potential [67,68]. Similarly, many other monoterpenes have also been shown to ameliorate endothelial cell apoptosis by modulating the Bcl-2/Bax ratio, caspase-9, caspase-3, the release of cytochrome c, and mitochondrial function [64,66,77,78,83,90,97,109]. All this evidence suggests a mitochondrial-dependent intrinsic pathway as a target in the effect of these monoterpenes. Furthermore, the extrinsic apoptotic pathway is likely implicated in the anti-endothelial apoptotic effect of monoterpenes. Monotropein, an active monoterpenoid isolated from the roots of Morinda officinalis, was reported to suppress the phosphorylation of NF-κB and activating protein-1 (AP-1), which further inhibited the expression of pro-inflammatory cytokine TNF-α and reduced cell apoptosis in H₂O₂-induced HUVECs. These studies indicated that the protective effect of monotropein on endothelial cells might involve the regulation of the TNF-mediated mitochondrial-independent apoptotic pathway [77]. Similarly, picroside II, the main active constituent of Picrorhiza scrophulariiflora belonging to monoterpenoid, was found to decrease the caspase-3 activity and the cleaved caspase-3 protein level to inhibit apoptosis in homocysteine-treated HUVECs, which might also be related to the attenuation of TNF-α production [76]. In addition, another study showed that harpagoside, a monoterpenoid extracted from the traditional Chinese herb Scrophulariae Radix, prevented angiotensin II (Ang II)-induced endothelial apoptosis via inactivation of caspase-8/caspase-9/caspase-3, suggesting that harpagoside could exert an anti-apoptosis effect by targeting both intrinsic and extrinsic apoptotic pathways [83].