Formation of propionate and butyrate by the human colonic microbiota.Short-chain fatty acids SCFAsprimarily acetate, propionate, and butyrate, are metabolites formed by gut microbiota from complex dietary carbohydrates. Butyrate and acetate were reported to protect against diet-induced obesity without causing hypophagia, while propionate was shown to reduce propionate in colon intake. However, the underlying mechanisms for these effects are unclear. We examined the effects of SCFA administration in mice, and propionate in colon that butyrate, propionate, and acetate all protected against diet-induced obesity and insulin resistance. Butyrate and propionate, but not acetate, induce gut hormones east german t-72 reduce food intake.
Intestinal Short Chain Fatty Acids and their Link with Diet and Human Health
Short-chain fatty acids SCFAs , primarily acetate, propionate, and butyrate, are metabolites formed by gut microbiota from complex dietary carbohydrates. Butyrate and acetate were reported to protect against diet-induced obesity without causing hypophagia, while propionate was shown to reduce food intake. However, the underlying mechanisms for these effects are unclear. We examined the effects of SCFA administration in mice, and show that butyrate, propionate, and acetate all protected against diet-induced obesity and insulin resistance.
Butyrate and propionate, but not acetate, induce gut hormones and reduce food intake. The effects of butyrate and propionate on body weight and food intake are independent of FFAR3. In addition, FFAR3 plays a minor role in butyrate stimulation of Glucagon-like peptide-1, and is not required for butyrate- and propionate-dependent induction of Glucose-dependent insulinotropic peptide.
Finally, FFAR3-deficient mice show normal body weight and glucose homeostasis. Stimulation of gut hormones and food intake inhibition by butyrate and propionate may represent a novel mechanism by which gut microbiota regulates host metabolism. These effects are largely intact in FFAR3-deficient mice, indicating additional mediators are required for these beneficial effects. November 7, ; Accepted: March 10, ; Published: This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
All authors are employees of Merck Research Laboratories, Rahway. There are no patents, products in development or marketed products to declare.
Short-chain fatty acids SCFAs are produced by microbiota in the colon and the distal small intestine from resistant starch, dietary fiber, and other low-digestible polysaccharides in a fermentation process .
Acetate, propionate, and butyrate are the predominant SCFAs in the gut lumen in humans and rodents, and are present at high mM levels . Once produced, SCFAs are readily absorbed by colonocytes. Butyrate is largely utilized by the colonic epithelium as an energy source, and propionate is primarily utilized by the liver, whereas a significant amount of acetate enters systemic circulation and reaches peripheral tissues.
In addition to acting as energy sources, SCFAs are also signaling molecules. It is well established that supplementing resistant starch and dietary fibers in diet, which raises intestinal and circulating SCFAs, confers metabolic benefits in humans. In rodent models of genetic or diet-induced obesity, supplementation of butyrate in diet  and oral administration of acetate  was shown to suppress weight gain independent of food intake suppression.
Activation of Adenosine 5'-monophosphate-activated protein kinase AMPK  ,  and increased mitochondrial function  were observed in these models, but only after chronic SCFA treatment when body weight was already significantly reduced compared to controls.
Thus, the primary mechanism underlying the resistance to obesity remains obscure. Propionate was reported to inhibit food intake in humans  , but the molecular mediators have not been identified. PYY and other peptide hormones secreted by enteroendocrine cells, such as the incretins Glucagon-like peptide 1 GLP-1 and Glucose-dependent insulinotropic polypeptide GIP , are key modulators of energy homeostasis and glucose metabolism.
In this study, we examine the effects of SCFAs on body weight, glucose metabolism, and gut hormones in wild-type and Ffar3 knockout mice. We show that butyrate and propionate suppress food intake, protect against high-fat diet-induced weight gain and glucose intolerance, and stimulate gut hormone secretion predominantly via FFAR3-independent mechanisms.
We also show that FFAR3 is not required for normal body weight and glucose homeostasis. As expected, mice on control HFD gained weight steadily over time. At the end of four weeks, propionate-fed mice showed reduced fasting glycemia, and both butyrate- and propionate-fed mice showed significantly improved oral glucose tolerance, while the acetate-fed group did not Figure 1B. These data are consistent with improvements in insulin sensitivity secondary to body weight reduction.
In a separate cohort, food intake and locomotor activity were measured for nine days after mice were switched to control or SCFA-supplemented HFD. Locomotor activity was not altered by butyrate or acetate, and tended to be increased by propionate feeding Figure 2C. An eight-day dose titration study showed that the minimum efficacious dose for suppression of weight gain is 2. Acetate did not lead to a significant inhibition of weight gain during the first week Figure 1A and was not included in the dose titration study.
Collectively, these data indicate that butyrate and propionate inhibit weight gain partially via suppressing food intake, while the inhibition of weight gain by acetate is independent of changes in food intake and locomotor activity, suggesting increased metabolic rate or reduced absorptive efficiency.
A Body weight was measured weekly, and four-week cumulative weight gain is expressed as a percentage of initial body weight. B Oral glucose tolerance test was performed in overnight fasted mice four weeks after diet switch. Blood glucose levels and total glucose area-under-the-curve AUC are shown.
C, D Plasma levels of insulin and leptin were determined in overnight fasted mice four weeks after diet switch. Daily food intake, cumulative food intake, and cumulative locomotor activity are shown.
Eight-day cumulative body weight change and food intake are shown. We selected the dose in the acute studies to match the amount of SCFAs consumed in a typical meal in the dietary supplementation experiment.
Thus acetate, propionate, and butyrate supplemented at 3. Plasma levels of the incretins GLP-1 and GIP were reported to peak at approximately ten minutes after an oral glucose challenge  and at 0. We therefore examined gut hormones at ten minutes and one hour after oral SCFA challenge.
Oral administration of sodium butyrate in mice significantly increased plasma levels of GLP-1 and GIP ten minutes after dosing Figure 3A-C , and levels of both hormones normalized to baseline by sixty minutes post-dosing data not shown.
PYY also showed a moderate increase ten minutes after oral butyrate administration Figure 3D. An SCFA admixture mimicking the endogenous proportions present in colon acetate mpk, propionate 80mpk, and butyrate 60mpk also elicited a modest increase in GIP.
Since butyrate and propionate, the SCFAs that preferentially activate FFAR3, are more effective than acetate in suppressing weight gain and stimulating gut hormones, we examined the contribution of FFAR3 to these effects. Ffar3 knockout mice showed no significant difference in body weight compared to wild-type littermates on standard chow diet and after one week of HFD feeding Figure 4A. The mice were then switched to HFD supplemented with butyrate and propionate for eight days.
Both butyrate and propionate inhibited weight gain and food intake in Ffar3 knockouts to the same extent as in wild-type mice Figure 4B-D. A Body weight of three-month-old Ffar3 knockouts and wild-type littermates on standard chow diet and one week after switching to HFD. Cumulative body weight change and daily food intake are shown. This expression pattern is consistent with a potential role in peptide secretion from L, K, and other enteroendocrine cells.
GIP levels in Ffar3 knockouts trended lower than wildtype controls under basal and butyrate-stimulated conditions, but the degree of stimulation by butyrate was similar between genotypes, and that by propionate showed a slight increase in knockouts Figure 5D, E. Stimulation of PYY and insulin by butyrate was blunted in this cohort, likely due to exposure to HFD for four weeks, and showed no significant difference between genotypes Figure 5F, G.
Interestingly, butyrate significantly reduced plasma ghrelin levels in Ffar3 knockouts, while this effect did not reach statistical significance in control mice Figure 5H. Data are normalized against Rplp0 mRNA.
These data suggest that FFAR3 is dispensable for normal energy homeostasis and glucose metabolism. B, C Plasma leptin levels were determined in overnight fasted Ffar3 knockouts and wild-type littermates maintained on standard chow diet or HFD.
D Blood glucose was measured in ad libitum fed mice maintained on HFD three hours after the start of the light phase.
E Oral glucose tolerance test after overnight fasting and F intraperitoneal insulin tolerance test after five-hour daytime fasting in Ffar3 knockouts and wild-type littermates maintained on HFD. The integral role of gut microbiota in the physiological regulation of host energy metabolism has attracted considerable attention. A number of studies have shown that obesity and metabolic disorders are associated with profound changes in gut microbiota .
However, mechanistic insights are lacking, and whether microbiota dysfunction plays a causal role in the pathogenesis of metabolic diseases is unclear.
In particular, how microbiota-derived metabolites, such as SCFAs, interact with host nutrient sensing pathways to modulate energy metabolism is poorly understood. Although SCFAs have been postulated to regulate gut hormone secretion, in vivo evidence was scant, and the downstream signaling pathway was not characterized.
In this study, we systematically examined the effects of each major SCFA naturally present in the colon—butyrate, propionate, and acetate—on energy metabolism and gut hormones. We found that all three SCFAs protected against diet-induced obesity, with butyrate and propionate being more effective than acetate. Butyrate and propionate regulate body weight at least partially by inhibiting food intake, consistent with their stimulatory effects on anorexigenic gut hormones. In contrast, acetate inhibited weight gain independent of food intake suppression and had no acute effect on gut hormones.
Our finding on the hypophagic effect of butyrate differs from a previous report, which concluded butyrate supplementation led to hyperphagia . As Gao et al. However, the current data cannot rule out potential impact on food intake due to altered palatability by butyrate and propionate supplementation.
Although mice fed SCFA-supplemented diets did not display overt signs of malaise, additional studies will be required to formally address this possibility. In addition to effects on food intake, changes in energy expenditure likely also contribute to body weight regulation by SCFAs. Butyrate-treated mice showed an increased capacity for cold-induced adaptive thermogenesis .
Systemic administration of propionate acutely increases heart rate . And acetate-treated rats were reported to have increased oxygen consumption .
Future studies will be needed to determine the contributions of these mechanisms to energy homeostasis in the chronic setting. SCFA levels in the gut lumen were reported to reach high mM levels in the colon of human and pigs, but are much lower in jejunum and ileum . In the current study, the doses of SCFAs chosen were in the high mM range, likely achieving supraphysiological levels in vivo, especially in the proximal intestine.
Therefore, the effects on GLP-1 and GIP may reflect direct stimulation of enteroendocrine cells in the proximal small intestine, where both L cells and K cells can be found. On the other hand, L cells that express PYY are only present in the colon and distal small intestine, suggesting an indirect mode of action of butyrate on this gut hormone. Butyrate- and propionate-dependent inhibition of food intake and weight gain was intact in Ffar3 knockout mice, implicating other endogenous mediators in these effects.
It's worth noting that Ffar3 knockouts on control HFD showed modest hyperphagia but had normal body weight and adiposity. This is consistent with a previous report showing accelerated intestinal transit and increased fecal energy excretion in an independent Ffar3 knockout line  , suggesting FFAR3 is required for normal gut motility and nutrient absorption.
The mechanisms responsible for increasing food intake and body weight normalization in the absence of FFAR3 are unknown.
SCFAs were shown to regulate leptin secretion from the adipose tissue. However, the role of FFAR3 in mediating these effects remains controversial  , . We showed that Ffar3 knockouts maintain normal plasma leptin levels, suggesting that leptin has no major role in normalizing energy homeostasis in Ffar3 knockouts.
Ffar3 knockout mice were recently reported to have reduced resting heart rate and sympathetic activity . However, this effect is expected to reduce energy expenditure and cannot explain the reduced feed efficiency of Ffar3 knockouts.
Despite an intact anorectic response to butyrate, Ffar3 knockouts showed an attenuation of butyrate-stimulated GLP-1 release. Fasting GLP-1 levels were normal in Ffar3 knockouts. Conversely, Ffar3 knockouts showed largely normal GIP stimulation by butyrate and propionate and an increased sensitivity to acute ghrelin suppression by butyrate.