1. Introduction

Cardiovascular disease (CVD), remain the number one cause of global deaths, responsible for about 17.5 million deaths worldwide annually ^[1]. As a vast multitude of cardiovascular risk factors have been identified, while some are genetic dispositions, many risk factors are modifiable and commonly arise under the current economic climate, especially from the commercialized diets and excessively sedentary lifestyles. Acquiring, maintaining, or exacerbating cardiovascular risk factors culminate in conditions of the metabolic syndrome, which include hypercholesterolemia, dyslipidemia, type-II diabetes, as well as endothelial dysfunction and cardiovascular diseases. Endothelial dysfunction and hypercholesterolemia, among other disorders, are the driving mechanisms of atherosclerosis, which is the major cause of CVDs. Indeed, clinical complications of the heart and blood vessels, such as myocardial infarction and peripheral artery disease, account for the majority of the morbidity/mortality associated with the metabolic syndrome. As the body’s lipid levels correlate with cardiovascular risk, fatty acid binding proteins are small intracellular proteins in many cell types responsible for the roles of lipid-trafficking. Many isoforms of this protein family have been identified and described for their shared mechanisms of interacting and binding with cytosolic lipids ligands for their escorts to coordinated sites of metabolism and signaling. However, the unique functions of each isoforms in specific cell types are still largely unknown. Nevertheless, the roles of certain isoforms in cell-types specialized for lipid processing, such as adipocytes, macrophages, and endothelial cells, have been implicated in the regulation of systemic homeostasis, suggesting their importance in the development of metabolic and cardiovascular disorders.

2. Fatty Acid Binding Proteins

Lipids are vital components of many biological processes and crucial in the pathogenesis of numerous common diseases, but the specific mechanisms coupling intracellular lipids to biological targets and signalling pathways are not well understood. This is particularly the case for cells burdened with high lipid storage, trafficking and signalling capacity such as adipocytes and macrophages. Here, we discuss the central role of lipid chaperones—the fatty acid-binding proteins (FABPs)—in lipid-mediated biological processes and systemic metabolic homeostasis through the regulation of diverse lipid signals, and highlight their therapeutic significance. Pharmacological agents that modify FABP function may provide tissue-specific or cell-type-specific control of lipid signalling pathways, inflammatory responses and metabolic regulation, potentially providing a new class of drugs for diseases such as obesity, diabetes and atherosclerosis.

The body’s lipids are physiologically essential. In addition to serving as effective long-term metabolic energy storages, cellular lipids can have signaling roles in many metabolic and inflammatory pathways. For instances, the eicosanoids, such as prostaglandins, are derived from fatty-acids metabolism and mediate the acute inflammatory responses ^[2]. In addition, lipid levels in adipocytes dictate their production of cytokines and adipokines, such as leptin and adiponectin, that have potent impact on inflammation and metabolism ^[3]. Pathologically, as the metabolic syndrome is closely linked to cardiovascular risk, diverse lipid signals and cells with high capacity of lipids storage, trafficking and signaling, such as adipocytes and macrophages, are crucial in the pathogenesis of CVDs. Therefore, lipids-related physiology and cardiovascular impacts crucially depend upon the specific processing and management of the bioavailability of cellular lipids. Such roles have been described for the prominently expressed fatty acid binding proteins (FABPs), lipid-chaperones that regulate many lipid-related processes. The functional aspects of the FABPs are currently being investigated to provide pharmacological or diagnostic targets for controlling the body’s lipid signaling and the associated inflammatory and metabolic mechanisms, which helps develop treatment for atherosclerosis and the metabolic syndrome.

The FABPs are small (12–15 kDa) cytosolic proteins abundantly expressed in tissues with active lipid metabolism, such as the heart and liver, or cell types specialized for lipid storage, trafficking and signaling, such as adipocytes and macrophages ^[4]. They are a multigene family, well-conserved, and known to be central in a variety of metabolic and cardiovascular disorders, including obesity, diabetes and atherosclerosis ^[5]. Structurally, all members of the FABP family share a β-barrel signature that consists of a water-filled cavity and a site that binds specific lipid-ligand unique for each member ^[6]. Nine members of the FABP family have been identified (FABP 1–9) with 20–70% sequence homology among members and each named according to the most abundantly expressing tissue in addition to their designed number. For instance, FABP3 is also heart-type FABP which is most abundant in cardiomyocytes; FABP4 and 5 are adipocyte- and epidermal-type and expressed most prominently in the respective tissues ^[7]. Despite the unique tissue-expression pattern of each member, in general, tissues with active lipid metabolism tend to express FABPs and, often, more than one isoform. For instance, the small intestine where active absorption of diet lipid take place express prominently FABP2, but also FABP1 (liver FABP) and FABP6 (ileal FABP) ^[8].

Functionally, the FABPs are known to reversibly interact with hydrophobic ligands with various affinities and mediate their escorts to coordinated sites of lipid metabolisms or signaling, typically serving as intracellular lipid chaperones. Reports up-to-date have documented some of the targeted sites to be lipid droplets for storage, plasma membrane in lipid import and export, mitochondria for lipid-metabolism, as well as the endoplasmic reticulum for phospholipid biosynthesis, specific enzymes for the production of lipid-derived signaling molecules, and the nucleus where their physical interaction with the peroxisome proliferator-activated receptors have been reported ^[9]. For instance, FABP1 was shown to regulate PPARα in mammalian renal COS-7 cells ^[10]. However, the promoter of the FABP1 gene itself contains a peroxisome-proliferator response element and, accordingly, FABP1 transcript level was shown to be regulated by intracellular fatty acids, dicarboxylic acids and retinoic acid ^[11]. The degree of expression may reflect the lipid-metabolizing capacity of a given tissue or cell type and can be modulated upon changes in the bioavailability of lipids, as in lipid exposure or usage processes (^[12]). While there is a strong regulatory connection between FABP expression and lipid related signaling, the exact function of different FABPs member remain poorly understood as their general mechanism is associated with a vast scope of complex lipid-related regulatory pathways. For example, FABP2s are abundantly expressed in the small intestine and known to bind absorbed saturated long-chain fatty acids with a high affinity. The lipid-intracellular trafficking of FABP2 is thought to be within the lipid-uptake, lipid-sensor, and lipoprotein synthesis pathways. However, complete ablation for FABP2 in mice did not compromise fat absorption but resulted in larger livers and higher triglyceride levels in males and the opposite in females ^[13]. In another example, studies have shown epidermal FABP5 influencing cell-survival pathways through PPAR-δ ^[14] As the exact functions of each unique FABP members are still under investigation, large resources and attention in metabolic and cardiovascular research are being directed at three members among others: FABP3, 4, and 5.

2.1. FABP3

FABP3 (heart-FABP) is expressed most abundantly in myocardiocytes and skeletal muscle. As lipid chaperone, myocardial FABP3 is essential for the metabolic homeostasis of cardiac function. Physiologically, 70% of the energy in the heart are derived from the oxidation of fatty acids within the mitochondria and peroxisomes, which require lipid-trafficking mechanisms ^[15]. Increased exposures to fatty-acids were shown to upregulate H-FABP in myocytes ^[16]. FABP3 is also influenced by the metabolic essential PPAR-α agonists ^[17]. Diabetic patients, within whom fatty acids become the primary source of energy, also exhibited upregulated cardiac FABP3 ^[18]. In addition, FABP3-deficient mice showed elevated plasma free fatty acids and were compromised for cardiac fatty acid uptake, resulting in reduced exercise tolerance and a switch toward rapid glucose usage in the heart that leads to cardiac hypertrophy ^[19]. Meanwhile, similarly to other FABP members, FABP3 was also found in a multitude of other tissues to a lesser extent, brain, testis, kidneys, adrenal glands, and others ^[6]. For this notion, the unique function of FABP3 remains complex and unclear. For instance, while the FABP3 in skeletal muscles mediate the uptake and escorting of fatty acids to the mitochondrial oxidation system as similar as cardiac FABP3, increased apoptosis and exacerbated cardiac dysfunction has been associated with FABP3 overexpression in myocardiocytes ^[20].

Despite the functional complexity, FABP3 is currently utilized as a clinical biomarker for cardiac injury and heart failure, particularly in the diagnosis of myocardial infarction (MI). MI is characterized by the death of heart tissues from injuries commonly due to atherosclerotic-mediated cardiac ischemia. The current diagnostic approaches for MI include assessing initial chest pain, characteristic electrocardiography, and the detection of biomarkers for myocardial injury ^[21]. During a heart injury, myocardiocytes suffer a breakdown of cellular and subcellular components, which is followed by the releases of these biomolecules from the injured cells into the circulatory system. Of clinical significance is the leakage of the cytosolic myocardial proteins; their releases serve as a pathological biomarker that can be detected and measured in blood to enable early and efficient clinical assessment of cardiac injury. The more effective biomarkers are more cardiac specific and more abundantly expressed in the myocardiocytes. Clinical guidelines for MI dictates that detecting/excluding MI within the first 6 h of chest pain would bring about the most effective clinical responses ^[22]. Currently, the only gold-standard biomarker for MI is the cardiac troponin, particularly the cardiac-specific subunits I and T, which can be detected 2–4 h after the onset of chest pain ^[23]. FABP3 has been proposed as an effective biomarker of myocardial injury. Under normal conditions, the cytosolic to plasma presence ratio of myocardial FABP3 is significantly high, with negligible plasma concentration of FABP3 ^[24]. Within 30 min of chest pain, blood FABP3 begins to rise and peak in a few hours before returning to baseline via renal clearance in about 24 h. This early release of FABP3 from injured myocardium has been observed in both animal models ^[25] and MI patients ^[26]. Recently our research group has demonstrated that patients with PAD have elevated plasma levels of FABP3. Our data demonstrated that the circulating levels of FABP3 increase as the severity of PAD worsens. However, extensive research is still required to demonstrated utilizing FABP3 as a biomarker for PAD ^[27].

2.2. FABP4 and 5

FABP4 (adipose-FABP) have been described for their roles in the development of the metabolic syndrome through their mechanisms in adipocytes as well as macrophages. Both the differentiation of functional adipocytes ^[28] and macrophages ^[29] involved the regulation of FABP4, and cellular signals regulating FABP4 in adipocytes and macrophages include fatty acids, agonists of PPAR-γ, insulin, lipopolysaccharide, and oxLDL ^[30]. Reduced lipolysis efficiency was observed in adipocyte-specific-FABP4-deficient mice ^[31], and these mice were also found with ameliorated insulin resistance during diet-induced obesity (^[32],^[33]). Moreover, apolipoprotein-E-deficient mice with FABP4 deficiency were found to be protected against atherosclerosis with or without induction by high-cholesterol Western diets ^[34], but how FABP4 deficiency can alter insulin resistance and lipid metabolism remain to be revealed. FABP4 was also shown to be released from adipocytes into blood. While their biological roles in blood remains unknown, serum FABP4 has been suggested as a potential biomarker for metabolic syndrome and CVDs ^[35]. In macrophages, FABP4 was found to modulate inflammation and cholesterol concentration. Administration of the cholesterol-lowering atorvastatin was found to suppress FABP4 expression in macrophages in vitro ^[36]. In addition, enhanced cholesterol efflux was observed in macrophages with elevated PPAR-γ activity from FABP4-deficient mice from a study that demonstrated macrophage’s FABP4 playing a role in foam-cell formation through regulating the PPAR-γ–liver X receptor-α (LXR-α)–ATP-binding cassette A1 (ABCA1) pathway. The same study also found suppressed production of cytokines and pro-inflammatory enzymes, such as TNF-α and COX2, respectively in macrophages of mice deficient for FABP4 ^[37].

The epidermal-type FABP5 is most prominently expressed in the skin cells, but it is also expressed in multiple other tissues, including adipocytes and macrophages, as well as tongue, brain, kidney, liver, lung, and testis ^[30]. In adipocytes, FABP5 expression is significantly minimal compared to FABP4 ^[38], but loss of FABP4 induce the upregulation of FABP5 that, in fact, masked the phenotypic effects of FABP4-deficiency ^[32]. Unsurprisingly, FABP4 compensatory upregulation is not observed in adipocytes of FABP5 deficient mice due to the already higher presence of FABP4 in adipocytes ^[5]. In macrophages, the expression ratio of FABP4 and 5 is about identical, but no compensatory FABP5 expression is observed in FABP4-deficient mice ^[39]. Due to this wide and complicate pattern of tissue expression and regulation, the unique function of FABP5 remains unclear. Nevertheless, several in vivo phenotypes regarding FABP5 expression are relevance to the metabolic syndrome. Transgenic mice overexpressing adipose-specific FABP5 exhibited enhanced lipolysis ^[40] and reduced insulin sensitivity ^[5]. On the other hand, increased insulin sensitivity was observed in adipocytes from FABP5 deficient mice ^[5]. FABP5 deficient mice also appeared healthy, without any changes in the normal epidermal fatty-acid composition ^[41].

As macrophages accumulation in adipose tissues characterize the enhanced inflammatory response and risks for insulin resistance and CVDs in obesity ^[42], the roles of FABP4 and 5 in both adipocytes and macrophages contribute to the inflammatory and metabolic aspects of the metabolic syndrome and atherosclerosis ^[4]. In general, adipocytes and macrophages in mice deficient for these FABPs were more insulin sensitive. Obese mice deficient for both FABP4 and 5 exhibited reduced tissue fatty-acid composition and did not develop insulin resistance ^[43]. Even when the ApoE^-/- model was integrated, these mice showed less atherosclerosis development and increased survival compared to wild-type and either the individual FABP-deficient counterparts ^[44]. The mice deficient for FABP4 and/or FABP5 also exhibited increased fatty acids levels in plasma ^[45], suggesting that the intracellular bioavailability of lipids, rather than the total body’s amount, is more relevant to the development of metabolic and cardiovascular disorders. Recently, we were able to demonstrate that FABP4 levels were elevated in diabetic patients with PAD ^[46]. The elevation in FABP4 was independent of confounding factors such as age, sex, or prior history of CAD. Therefore, our work raises the possibility of utilizing FABP4 as a biomarker for diagnosing PAD in diabetic patients.

2.3. FABP in Endothelium

Although glucose is the primary source of energy in endothelium ^[47], ECs have been demonstrated for extensive lipid-processing mechanisms, including lipid uptakes and transport, FA metabolisms pathways for the synthesis of deoxyribonucleotide triphosphate fueling proliferation and lipid-derived arachidonic acid metabolites. Moreover, cholesterols metabolism was also shown in ECs for their expression of the Niemann–Pick disease type C (NPC) 1 and NPC2 proteins, which mediate cholesterol uptake and trafficking ^[48]. The activities of endothelial mammalian target of rapamycin (mTOR), which is central player of a signaling network regulating cell growth and proliferation, is dependent on intracellular cholesterol trafficking. Pharmacological blockade of cholesterol trafficking by itraconazole or by silencing NPC 1 and 2 led to inhibition of mTOR activity in ECs ^[48]. These observations suggest ECs to be active sources of lipid signaling and metabolism. However, few studies have been able to demonstrate the expression and activities of FABP in the endothelium. Nevertheless, two FABP members have been identified in ECs. The expression of FABP3 and FABP5 have been described in the microvascular of cardiac tissues and skeletal muscles. In addition, FABP4 and FABP5 expressions were found in the microvasculars of other organs with active fatty acids metabolism, including the liver and adipose tissues ^[49][50]. Moreover, Masouyé et al. presented that ECs are capable of fluctuating their expression of FABPs depending on environmental factors, such as between tissue and culture conditions. This study detected FABP5 by immunohistochemistry in cultured human umbilical vein ECs (HUVECs), but not in the endothelium of the umbilical vein from which the HUVECs were isolated ^[51].