Short Chain Fatty Acid Receptors and Blood Pressure Regulation

Abstract

The gut microbiome influences host physiology and pathophysiology through a number of pathways, one of which is microbial production of chemical metabolites which interact with host signaling pathways. Short chain fatty acids (SCFAs) are a class of gut microbial metabolites known to activate multiple signaling pathways in the host. Growing evidence indicates that the gut microbiome is linked to blood pressure, that SCFAs modulate blood pressure regulation, and that delivery of exogenous SCFAs lowers blood pressure. Given that hypertension is a key risk factor for cardiovascular disease, the examination of novel contributors to blood pressure regulation has the potential to lead to novel approaches or treatments. Thus, this review will discuss SCFAs with a focus on their host G protein-coupled receptors including G protein-coupled receptor 41 (GPR41), GPR43, and GPR109A, as well as olfactory receptor 78 (OLFR78) and OLFR558. This includes a discussion of the ligand profiles, G protein coupling, and tissue distribution of each receptor. We will also review phenotypes relevant to blood pressure regulation which have been reported to date for Gpr41, Gpr43, Gpr109a, and Olfr78 KO mice. In addition, we will consider how SCFA signaling influences physiology at baseline, and, how SCFA signaling may contribute to blood pressure regulation in settings of hypertension. In sum, this review will integrate current knowledge regarding how SCFAs and their receptors regulate blood pressure.

Introduction

Hypertension is a major risk factor for cardiovascular disease and stroke, as well as a hallmark of obesity, diabetes and metabolic syndrome^1–3. Successful hypertension prevention and treatment are key to decreasing the risk of those diseases. Based on the American Heart Association’s definition of hypertension (systolic blood pressure (SBP)⩾130 mmHg, diastolic blood pressure (DBP)⩾80 mmHg), it is estimated that 47% of US adults, approximately 121.5 million adults⁴, are hypertensive (52% for males and 43% for females^1,5). For those <65 years of age hypertension is more prevalent in males; the opposite is true for those who are ⩾65 years of age¹. Hypertension incidence also varies by race, with black men and black women both having a cumulative incidence of 76% by age 55, as compared to 54% for white men and 40% for white women⁶. Compounding these issues, 39% of US adults with hypertension are unaware that they are hypertensive¹.

Growing evidence indicates that the gut microbiota influences a wide variety of traits in the host organisms, including atherosclerosis^7,8, irritable bowel syndrome^9,10, immune disorders^11–13, and kidney disease^14–17. A primary way that microbes affect the host is via the production of metabolites that are absorbed into the host bloodstream, where they can activate signaling pathways to alter physiology. A particularly well-studied class of gut microbial metabolites are short chain fatty acids (SCFAs), which are simple straight-chain carboxylic acid molecules. In recent years, multiple groups have connected SCFAs to blood pressure regulation, and, to hypertension^18–23. Here, we will review our current understanding of SCFAs, their receptors, and their effects on physiology.

Short Chain Fatty Acids (SCFAs)

Short chain fatty acids (SCFAs) are metabolites produced by microbial fermentation of dietary carbohydrates, and are perhaps the most well-studied of the gut microbial metabolites²⁴. The term ‘SCFAs’ commonly refers to the three most abundant species of SCFAs, which are all 2-4 carbon straight-chain compounds (acetate, propionate, and butyrate) - this is how we will use the term in this review. However, SCFAs can also include shorter (formate) or longer (valerate) compounds, as well as branched compounds (isobutyrate, isovalerate, and 2-methylbutanoate).

Indigestible complex carbohydrates in the large intestine undergo fermentation by anaerobic microbes, resulting in the production of metabolites including SCFAs²⁵. The most abundant butyrate-producing bacteria in the colon belongs to the Clostridia clusters IV and XIVa, which includes species related to Firmicutes, Eubacterium and Roseburia^26–28. Acetate and propionate are primarily produced by Bacteriodetes phyla^29,30.

As referenced above, gut microbiota produce SCFAs as a byproduct of metabolism; in fact, the concentration of SCFAs in the colonic lumen is ~100mM³¹. The ratio of acetate:propionate:butyrate in the colon is approximately 60:20:20^32,33. SCFAs transverse from the colonic lumen to the blood by monocarboxylate transporters³⁴, as well as by diffusion³⁵. SCFA levels within the circulating blood of the host are much lower than in the colonic lumen, albeit still substantial. For example, acetate, the most abundant of the SCFAs, is typically 50-500 μM in circulating plasma^32,36–38. Plasma acetate levels, as well as the ratio of acetate:propionate:butyrate, varies based on diet^25,39, but acetate is consistently the most abundant (ratios have been reported to vary from 40:26:34 to 78:17:5^39,40).

As previously stated, SCFAs are produced by the gut microbiota in quantities that significantly contribute to host circulating levels. Notably, host metabolism can also produce SCFAs, but it is unclear whether host SCFA production meaningfully contributes to host plasma levels. A study by Perry, et al found that plasma acetate, as well as plasma butyrate and propionate, are nearly undetectable in germ-free mice (that is, mice which entirely lack microbiota)³⁷. However, a study from Kimura, et al reported that plasma acetate only decreases by ~40% in germ-free animals⁴¹. Furthermore, a recent report indicated that mice treated with antibiotics had dramatically suppressed SCFAs in the colonic lumen, but no change in plasma SCFAs⁴². Thus, although there is agreement that gut microbial metabolism contributes to circulating SCFAs, the precise contribution of gut vs host metabolism remains to be fully understood.

Cell Biology of SCFA Receptors

SCFAs circulating in the blood can alter signaling pathways via several different classes of host proteins, most notably histone deacetylases (HDACs) and G protein-coupled receptors (GPCRs). HDACs remove acetyl groups from histones, increasing the affinity between histones and DNA to repress transcription⁴³. There are four classes of human HDACs based on their conserved sequences to yeast HDACs: I, II (including IIA and IIB), III, and IV⁴³ . SCFAs, primarily butyrate, are prominent inhibitors of HDAC activity^44,45. For example, in rats with cardiac hypertrophy, treatment with sodium butyrate was beneficial in reducing LV wall thickness, cardiomyocyte diameters, and hypertrophic indices⁴⁶. It was suggested these effects were mediated by decreased mRNA expression of HDAC subtypes⁴⁶. In another report, mice treated with SCFAs (acetate, butyrate, and propionate) showed reduced mRNA expression of HDAC subtypes in kidney tissue, and were protected against folic-acid nephropathy⁴⁷. Results of this study suggest SCFA-mediated HDAC inhibition contributes to the pathophysiology of acute kidney injury.

SCFAs can also act as ligands and bind to host microbial metabolite-sensing GPCRs, and this is the primary focus of this review. GPCRs are cell surface receptors containing seven transmembrane alpha helices and an extracellular amino terminus⁴⁸. Given their highly conserved sequences and structure, GPCRs are widely studied for their ability to sense a variety of stimuli⁴⁹. To date, GPCRs that have been identified to bind SCFAs include: GPR41, GPR43, GPR109A, OR51E2, and OR51E1^50–53; the expression of these GPCRs in the top 10 single cell types is shown in Figure 1.

Figure 1. Top 10 single cell types expressing (A) Gpr43, (B) Gpr41, (C) Gpr109a, (D) Or51e2 (mouse ortholog: Olfr78), and (E) Or51e1 (mouse ortholog: Olfr558).

Values are reported in normalized transcripts per million (nTPM)¹³⁴. Data available from Human Protein Atlas (v21.1.proteinatlas.org)^135–139.

GPR41 (free fatty acid receptor 3, FFAR3) and GPR43 (free fatty acid receptor 2, FFAR2) have been identified in smooth muscle cells, adipose, spleen, pancreatic tissues, and immune cells^53–59, where they play significant roles in host health and disease. The most potent agonists for GPR41 and GPR43 are acetate, propionate, and butyrate^53,60,61. Studies conflict as to whether beta-hydroxybutyrate (βHB), a ketone body, acts as an agonist or antagonist for GPR41^62–66. A 2011 study reported that βHB is an antagonist for GPR41⁶². However, a 2013 study demonstrated that βHB is a GPR41 dose-dependent agonist, and were unable to find evidence of antagonism⁶⁶. In our view, the data showing that βHB is an agonist for GPR41 is quite strong; however, it is important to note that βHB is used experimentally as both an antagonist^64,65 and an agonist⁶³, resulting in some confusion in the literature.

Previously reported ligand profiles and EC₅₀’s for GPR41 and GPR43 vary by both assay and species tested, but, acetate, propionate and butyrate are detected in the micromolar range^67,68 GPR41’s reported EC₅₀ for acetate, propionate, and butyrate are: 393-1072μM, 6-127μM, and 33-158μM, respectively. Reported EC₅₀’s for GPR43 for acetate, propionate, and butyrate are: 35-431μM, 14-290μM, and 28-371μM, respectively^67,68. In addition, these receptors do have additional (weaker) ligands^67,68. As stated above, circulating levels of acetate range from 50-500 μM in circulating plasma^32,36–38, and thus it is likely that physiological changes in acetate may alter GPR41 and/or GPR43 signaling. SCFAs which bind to these two receptors activate cell signaling through interactions with G proteins coupled to the receptor.

Additional reports find that GPR109A (alternatively referred to as HCAR2 or NIACR1) is also a metabolite-sensing receptor, albeit not as extensively studied as the previously mentioned SCFA receptors^59,69–71. GPR109A recognizes both niacin and butyrate, but binds butyrate with relatively low affinity⁶⁹. Indeed, it has been demonstrated in GTPyS assays that niacin activates GPR109A with an EC₅₀ ~ 250nM⁷⁰, although concentrations of butyrate needed to activate the receptor are within the millimolar range, at ~1 mmol/L⁵⁹. Similar to GPR41 and GPR43, GPR109A signals by activating the G_i/o protein, thus inhibiting adenylyl cyclase^53,59,69. Studies have demonstrated that GPR109A is primarily expressed in immune cells⁵⁰ and the intestinal lumen⁵², and expression of this receptor in the presence of its ligands plays a role in regulating host immune function and metabolism.

Olfactory receptors (OR) constitute the largest group of GPCRs, and recent findings demonstrate that two receptors within this family, OLFR78 and OLFR558, respond to SCFAs^50,72,73. Of note, a new unified nomenclature has been proposed for olfactory receptor naming across species, and has been adapted for some but not all species⁷⁴. In this article we will use the widely used root ‘Olfr’ nomenclature for murine receptors. Although olfactory receptors comprise the largest gene family in the genome⁷⁵, only three olfactory receptors have 1:1 orthology among placental mammals⁷⁶. OLFR78 (human ortholog: OR51E2) and OLFR558 (human ortholog: OR51E1) are two of these well-conserved receptors. OLFR78 is activated by acetate with an EC₅₀ of 2.01mM - 2.35mM, and propionate with an EC₅₀ of 0.63-0.92mM^50,73,77,78. OLFR558 is activated by butyrate with an EC₅₀ of ~0.2 mM (when co-transfected with the G_olf protein)^73,79.

Importantly, the ligand profile of OLFR78 is very similar to its human ortholog OR51E2⁸⁰, and the ligand profile of OFLR558 is very similar to its human ortholog OR51E1^73,79, with both OLFR558 and OR51E1 having additional (structurally related) ligands. Some groups^81,82 have reported that beta-ionone (β-ionone) and/or androstenone derivatives also act as ligands for OR51E2. There are also reports that lactate is a partial agonist for OLFR78^77,83,84; however, lactate does not activate OR51E2^83,85. Both OLFR78 and OLFR558 activation increases cAMP production, and in olfactory tissues, this happens via coupling to the olfactory G protein (G_olf)⁸⁶. In vitro, OLFR78 (and other ORs) likely can also couple to G_s; for both G_s and G_olf, OR activation leads to increases in cAMP. Importantly, it was recently demonstrated that OLFR78 (and OR51E2) heterodimerize with GPR132, and that the heteromer receptor is sensitive to lactate⁸⁵. The idea that some olfactory receptors dimerize to other GPCRs to influence cell signaling pathways is a noteworthy development in OR research.

SCFAs and Blood Pressure Regulation

Numerous studies have found an association between gut microbes and hypertension. For example, there are shifts in the gut microbiota of hypertensive subjects (‘dysbiosis’) in mice, in rats, and in humans^5,87–89. In addition, transplanting microbes from a hypertensive subject into germ-free mice (which do not have any native microbes) can transfer the hypertensive phenotype⁹⁰. Intriguingly, another study has shown that a high salt diet (a known risk factor for hypertension) depletes a specific strain of gut microbes, and depleting this strain can prevent salt-sensitive hypertension⁸⁷. SCFA transporters also link SCFAs to blood pressure regulation. Yang, et al. found that decreased expression of colonic Slc5a8 (which can transport butyrate) is associated with more butyrate in the cecal content but less butyrate in the circulation in the spontaneously hypertensive rat (SHR)⁹¹.

Changes in SCFAs have also been reported in Dahl rats. One study reported that dietary salt is associated with increased fecal levels of acetate, propionate, and isobutyrate (but not butyrate) in Dahl rats⁹². Another study reported that dietary salt did not alter plasma acetate, valerate or butyrate⁹³; however, it is not surprising that fecal levels may change without altering plasma levels. On the other hand, it was reported that fecal transplants can alter plasma SCFAs in Dahl salt-sensitive rats⁹⁴. Together, it seems that the connections between gut microbes, SCFAs, and blood pressure are not always clear. In fact, several groups have attempted to measure blood pressure in animals without any gut microbes (germ-free models), but results have varied with some reporting decreased blood pressure⁹⁵, a trend toward increased blood pressure⁹⁶, or no change in blood pressure⁹⁷. Although some of these differences may be due to the difficulties of working with germ-free models without compromising their germ-free status (to date, telemetry blood pressure measurements have not been performed in germ-free animals), it is also possible that differing findings may be due to the fact that germ-free mice are ‘missing’ not just one signaling pathway, but a multitude of pathways (due to the absence of all gut microbial metabolites), and likewise, that altering fecal or plasma levels of a SCFA can affect multiple receptors. Thus, in the following sections, we will focus on specific effects which can be assigned to specific SCFAs and/or specific GPCRs, in the hopes of beginning to disentangle these physiologically important yet interconnected and complex signaling pathways.

SCFAs and vasodilation

A key regulator of blood pressure is vasoconstriction and vasodilation of blood vessels⁵. In 1928, acetate was first reported as a vasodilator which drops BP due to vasodilation⁹⁸. Acetate was used as a pH buffer in dialysis solutions until the 1980s, when it was recognized that acetate was contributing to hypotension in patients via a vasodilatory effect^99–102. Numerous studies have now documented dose-dependent vasodilation of acetate, propionate, and butyrate which promote hypotension^{20,22,23,80,103}. This effect is markedly attenuated after denudation of the epithelium¹⁰³. Butyrate is also reported to dilate phenylephrine pre-constricted rat mesenteric and gracilis muscle arteries ex vivo⁶⁴.

As a result of these vasodilatory actions, acetate, propionate, and butyrate have blood pressure lowering effects. When delivered acutely (for example, via i.v. or i.p.), SCFAs cause acute hypotension which develops in seconds and recovers over minutes^80,104. Chronic intake of acetate¹⁰⁵, butyrate^106,107, or propionate¹⁹ also lowers blood pressure. Consistent with these findings, SCFAs lower BP even without prebiotic sources in the diet¹⁰⁸. Recent evidence indicates that Dendrobium officinale ultrafine powder (DOFP) alters intestinal flora, increases fecal and serum SCFAs, and enhances vascular endothelial vasodilation function in metabolic hypertension¹⁰⁹. This effect was blocked by the eNOS inhibitor L-NAME¹⁰⁹. In humans, it was reported that higher levels of plasma but not fecal butyrate was positively associated with ambulatory arterial stiffness index (AASI)¹¹⁰. Although no association between bacterial α and β diversity and AASI was found, two main bacteria taxa were associated with human AASI: Lactobacillus spp. and Clostridium app¹¹⁰. However, the role of these two bacteria taxa in human arterial stiffness needs to be determined. Of note, there are no published studies showing that SCFAs can lower blood pressure in humans as an intervention (however, there is a promising preprint in this area¹¹¹).

Two recent studies reported that that acute delivery of SCFAs not only decreases mean arterial pressure (MAP), but simultaneously suppresses heart rate (HR)^64,104, and these studies suggest that SCFAs lowers HR via modulating sympathetic tone¹⁰⁴. Acetate is also reported to affect cardiac function¹⁰⁴. Propionate is shown to induce cardiac protection by significantly attenuating cardiac hypertrophy, fibrosis, vascular dysfunction, and hypertension in angiotensin II infused hypertension mice and apolipoprotein E KO mice models¹⁹. Butyrate is shown to lower BP and decrease HR after administration into the colon in rat. However, the hypotensive effect was dramatically decreased by subphrenic vagotomy and pretreatment with βHB (used as an antagonist of Gpr41/43 in this study, but also reported as an agonist by other groups – see “Cell Biology of SCFA Receptors” section above)⁶⁴. Finally, SCFAs are reported to have protective effects on endothelial dysfunction induced by angiotensin II in rat aortic endothelial cells and rat aortas¹¹².

Effects of SCFAs on blood pressure regulation via GPCRs

In this section we will review current literature^113–117 regarding the roles of specific SCFA GPCRs in blood pressure regulation (Figure 2).

Figure 2. Ligands and cardiovascular phenotypes of SCFAs GPCR receptors, as well as sites of expression which have been suggested to contribute to these phenotypes.

This figure summarizes current knowledge regarding the ligands and sites of expression for GPR41, GPR43, GPR109A, OLFR78, and OLFR558. In addition, cardiovascular phenotypes in KO mice are outlined. Gpr41 KO mice exhibit increased SBP¹⁰³, with higher pulse pressure, end-diastolic pressure (DP), tau, and perivascular fibrosis¹⁰⁸. Gpr43 KO mice have an increased heart/body weight, pulse pressure, and perivascular fibrosis¹⁰⁸. Gpr109a KO mice are reported to have a higher pulse pressure, end-DP, as well as perivascular fibrosis¹⁰⁸. Olfr78 KO mice have a lower plasma renin activity^39,49. Images in this figure are from Servier Medical Art (smart.servier.com).

GPR41 is expressed in the vascular endothelium¹¹⁸, and, in the autonomic and sensory ganglia in mouse and human^62,119 indicating that it is well-positioned to modulate vascular tone in response to SCFAs. To date, two studies have examined blood pressure in Gpr41 whole-animal KO mice, both with similar findings: Natarajan et al¹⁰³ and Kaye et al¹⁰⁸. Using blood pressure telemetry, Natarajan et al reported that whole-animal Gpr41 KO have increased systolic pressure and increased pulse pressure, as well as an increased heart weight/body weight ratio¹⁰³. Although Kaye, et al did not publish systolic pressure data, they did report increased pulse pressure as measured by cardiac catherisation¹⁰⁸; given that increased pulse pressure is indicative of isolated systolic hypertension, it seems likely that systolic pressure was also elevated in these animals. Unlike the Natarajan study, Kaye, et al did not observe an increase in the heart weight/body weight ratio. However, Natarajan et al specify that heart weight/body weight was increased in 6-month old KOs¹⁰³ (Kaye, et al do not report the age of mice that were examined). If Kaye, et al examined younger mice this could explain the lack of significance in this measurement (and it does appear that Kaye, et al see a trend toward an increase)¹⁰⁸. Finally, Kaye, et al reported that Gpr41 KO have an increase in end-diastolic pressure, Tau, and perivascular fibrosis; these parameters were not examined in the Natarajan, et al study.

GPR43 is highly expressed in immune cells¹²⁰, and has also been reported in blood vessels⁵⁰. Gpr43 KO mice have increased heart to body weight and higher perivascular fibrosis, but without a difference in BP as measured via cardiac catheterisation¹⁰⁸. Nakai, et al. provided the first human evidence that Gpr43 is differentially modulated in essential hypertension: they found that hypertensive subjects had lower levels of Gpr43 expression in immune cells¹²¹. It was also reported that Gpr41 and Gpr43 expression in circulating immune cells is negatively associated with human arterial stiffness; presumably, decreased Gpr41 and Gpr43 decreases the response to BP-lowering SCFAs such as butyrate¹¹⁰. These data suggest that GPR43 signaling deficiency helps to drive the pro-inflammatory phenotype in hypertensive subjects¹²¹. Intriguingly, Ang et al found that GPR41 and GPR43 can heterodimerize in monocytes and macrophages, and that the heterodimer exhibited increased Ca²⁺ signaling and β-arrestin-2 recruitment. The enhanced signaling was attenuated by antagonizing GPR43, inhibiting G_αq inhibition (YM254890), or inhibiting G_αi¹²². The possibility of GPR41-GPR43 heterodimers adds an additional level of complexity to studies of GPR41 and GPR43.

GPR109A is widely expressed in white and brown adipose tissue, keratinocytes, various immune cells, and likely in microglia¹²³. Regarding blood pressure regulation, GPR109A in the rostral ventrolateral medulla (RVLM) plays a role in central blood pressure control, where activation by its ligand nicotinic acid (NA) leads to Ca²⁺-dependent L-glutamate release, subsequently increasing neuronal oxidative stress and sympathetic activity¹²³.

Gpr109a KO mice were reported to have elevated end-diastolic pressure and pulse pressure, as well as perivascular fibrosis. However, Gpr109a KO mice demonstrated no differences in BP compared to WT mice at baseline (measured via cardiac catheterisation)¹⁰⁸. Double KO of Gpr43/Gpr109a have significantly larger heart to body weight ratios, and increased end-diastolic pressure and pulse pressure¹⁰⁸, but without changes in BP at baseline (as measured by tail-cuff)¹⁰⁸. Gpr109a is also expressed in the retinal epithelium where it plays a role in the inflammatory pathway of diabetic retinopathy¹²⁴. Gpr109a expression is upregulated in diabetic retinopathy, suggesting that increased Gpr109a expression (and elevation of its ligand βHB) may help fight inflammation¹²⁴. Notably, Gpr109a has relatively high expression in immune cells (as does Gpr43), and SCFA GPCR expression in this cell type may also be quite important in blood pressure regulation. For example, immune cell infiltration in renal tissue¹²⁵ and perivascular adipose tissue¹²⁶ is required for the progression of hypertension. Immune cells are an important component of blood pressure regulation^125,126, where they ultimately alter blood pressure by altering parameters such as peripheral vascular resistance, cardiac output, or renal sodium absorption¹²⁶. An important effect of immune cell infiltration in the vasculature is the vascular remodeling, including changes in lumen diameter and in media-to-lumen ratio¹²⁷. Tissue-specific knockout models will be required to definitely determine whether the origin of a change in vascular function is due to expression of a SCFA GPCR in an immune cell, or, in vascular cells.

OLFR78 is expressed in vascular smooth muscle cells in a variety of organs; intriguingly, it is only found in a subset of the smooth muscle cells of each vessel⁵⁰. OLFR78 also localizes to renal afferent arterioles, where it has been shown to impact renin release⁵⁰. Specifically, it was reported that isolated renal glomeruli with attached juxtaglomerular apparati (JGA) from Olfr78 WT mice release renin in response to propionate, but this response is absent in JGA/glomeruli from Olfr78 whole-body KO mice⁵⁰. Consistent with this, it was shown that Olfr78 KO mice have lower plasma renin activity^36,50. Recently, it was reported that both Olfr78 whole-animal KO mice and Olfr78^fl/fl Renin-Cre KO mice have decreased renin protein levels associated with glomeruli³⁶. Despite these changes in renin, however, Olfr78 KO mice are normotensive when blood pressure is measured via telemetry³⁶. As the most closely related olfactory receptor of Olfr78, Olfr558 is widely expressed in many tissues including kidney and heart^73,128. Although a role for OLFR558 in blood pressure regulation has not been reported, Olfr558 expression is found in vascular smooth muscle cells in kidney, heart¹²⁸, and in the retina¹²⁹, suggesting that Olfr558 may play a role in blood pressure regulation.

We can also gain insight into how SCFA GPCRs function by considering how SCFA GPCR expression is altered by various stimuli. For example, a study by Nakai, et al assayed the expression of GPR41, GPR43 and GPR109A in immune cells of humans with and without hypertension using qPCR¹³⁰. While they did not observe changes in GPR41 or GPR109A expression, they found a decrease in GPR43 expression in hypertensive subjects. GPR43 expression was strongly negatively correlated with blood pressure, and this association was significant even after adjusting for age, body mass index, and sex. Interestingly, this same study found that hypertensive subjects had increased plasma acetate and butyrate, which was counter to what the authors had expected. Based on their own previous work^105,108, they presumed that higher levels of acetate and butyrate would lower blood pressure, not raise it. Intriguingly, they propose that increased levels of SCFAs may be ineffective in these patients due to the concomitant downregulation of GPR43. A separate study by Weber, et al¹³¹ examined the renal expression of Gpr41, Gpr43 and Olfr78 in a mouse model of hypertension (the Angiotensin II-infusion model). They found a decrease in Gpr41 and Gpr43 expression but an increase in Olfr78 expression by qPCR. They also used commercially available antibodies to examine protein level changes; although to our knowledge these antibodies have not been validated, they saw similar changes by western blot. Finally, a study by Brunskill¹³², et al used microarray to identify “genes that confer the identity of the renin cell” and found not only that Olfr558 is enriched in renin cells, but that the expression of Olfr558 is upregulated in mice treated with captopril (an angiotensin converting enzyme inhibitor).

Future Directions and Challenges

SCFA signaling is complex, and this complexity presents a clear challenge. First, there are multiple SCFAs; in fact, in addition to the three major SCFAs—acetate, propionate, and butyrate - SCFAs can induce shorter or longer, as well as branched compounds. Second, these SCFAs activate multiple receptors including but not limited to GPR41, GPR43, GPR109A, OLFR78, and OLFR558, some of which are found in multiple tissues, and some of which can heterodimerize. Each SCFA can activate multiple receptors, and each receptor can be activated by one or more than one SCFA. Moreover, multiple bacteria can produce each SCFA. Apparently, this pathway is more of a ‘web’ than a linear pathway. Thus, understanding the role of each individual member of this web (each SCFA and each host protein) is a major challenge.

To meet this challenge, we suggest that future studies which reduce aspects of this complexity will provide valuable clues: for example, tissue-specific KOs of a SCFA receptor, and/or, dosing animals with one SCFA at a time to understand the role of each SCFA. KO studies for SCFA receptors should also consider whether there may be compensatory changes in the remaining receptors which could influence interpretation of the results, and SCFA levels should be routinely measured for studies examining SCFA GPCRs to rule out (or rule in?) changes in SCFAs themselves. We must then use this information to inform and understand our interpretation of data from the more complex, intact system.

Looking ahead, this area holds great promise given that SCFA levels can be altered via dietary interventions. Thus, a nutritional therapeutic strategy – for example, the use of prebiotics and/or probiotics^4,133 - could be developed to prevent or attenuate hypertension. In future studies, it will be important to better understand the functional roles of these pathways in both normotension and hypertension, and to further explore how we may be able to strategically target SCFA signaling pathways to promote health.

Conclusion

Both SCFAs and SCFA GPCRs are known to influence blood pressure regulation, but the involvement of multiple SCFAs and multiple SCFA receptors makes this area of research complex. However, studies using both acute and chronic SCFA treatment as well as studies in KO animals have begun to unravel this complexity.