Abstract
Indole, which is produced by the intestinal microbiota from l-tryptophan, is recovered at millimolar concentrations in the human feces. Indoxyl sulfate (IS), the main indole co-metabolite, can be synthesized by the host tissues. Although indole has been shown to restore intestinal barrier function in experimental colitis, little is known on the effects of indole and IS on colonic epithelial cell metabolism and physiology. In this study, we compared the effects of indole and IS on the human colonic epithelial HT-29 Glc−/+ and Caco-2 cell lines, exposed to these compounds for 1–48 h. Indole, but not IS, was cytotoxic at 5 mM, altering markedly colonocyte proliferation. Both molecules, used up to 2.5 mM, induced a transient oxidative stress in colonocytes, that was detected after 1 h, but not after 48 h exposure. This was associated with the induction after 24 h of the expression of glutathione reductase, heme oxygenase, and cytochrome P450 (CYP)1B1. Indole and IS used at 2.5 mM impaired colonocyte respiration by diminishing mitochondrial oxygen consumption and maximal respiratory capacity. Indole, but not IS, displayed a slight genotoxic effect on colonocytes. Indole, but not IS, increased transepithelial resistance in colonocyte monolayers. Indole and IS used at 2.5 mM, induced a secretion of the pro-inflammatory interleukin-8 after 3 h incubation, and an increase of tumor necrosis factor-α secretion after 48 h. Although our results suggest beneficial effect of indole on epithelial integrity, overall they indicate that indole and IS share adverse effects on colonocyte respiration and production of reactive oxygen species, in association with the induction of enzymes of the antioxidant defense system. This latter process can be viewed as an adaptive response toward oxidative stress. Both compounds increased the production of inflammatory cytokines from colonocytes. However, only indole, but not IS, affected DNA integrity in colonocytes. Since colonocytes little convert indole to IS, the deleterious effects of indole on colonocytes appear to be unrelated to its conversion to IS.
Similar content being viewed by others
References
Adesso S, Ruocco M, Rapa SF, Dal Piaz F, Di Iorio BR, Popolo A, Autore G, Nishijima F, Pinto A, Marzocco S (2019) Effect of indoxyl sulfate on the repair and intactness of intestinal epithelial cells: role of reactive oxygen species’ release. Int J Mol Sci 20(9):2280
Agus A, Planchais J, Sokol H (2018) Gut microbiota regulation of tryptophan metabolism in health and disease. Cell Host Microbe 23(6):716–724
Andriamihaja M, Chaumontet C, Tome D, Blachier F (2009) Butyrate metabolism in human colon carcinoma cells: Implications concerning its growth-inhibitory effect. J Cell Physiol 218:58–65
Andriamihaja M, Lan A, Beaumont M, Audebert M, Wong X, Yamada K, Yin Y, Tomé D, Carrasco-Pozo C, Gotteland M, Kong X, Blachier F (2015) The deleterious metabolic and genotoxic effects of the bacterial metabolite p-cresol on colonic epithelial cells. Free Radic Biol Med 85:219–227
Armand L, Andriamihaja M, Gellenoncourt S, Bitane V, Lan A, Blachier F (2019) In vitro impact of amino acid-derived bacterial metabolites on colonocyte mitochondrial activity, oxidative stress response and DNA integrity. Biochimica Et Biophysica Acta (BBA) - General Subjects 1863(8):1292–1301
Banoglu E, Jha GG, King RS (2001) Hepatic microsomal metabolism of indole to indoxyl, a precursor of indoxyl sulfate. Eur J Drug Metab Pharmacokinet 26(4):235–240
Bansal T, Alaniz RC, Wood TK, Jayaraman A (2010) The bacterial signal indole increases epithelial-cell tight-junction resistance and attenuates indicators of inflammation. Proc Natl Acad Sci USA 107(1):228–233
Barzilai A, Yamamoto K-I (2004) DNA damage responses to oxidative stress. DNA Repair 3(8):1109–1115
Beaumont M, Andriamihaja M, Lan A, Khodorova N, Audebert M, Blouin J-M, Grauso M, Lancha L, Benetti P-H, Benamouzig R, Tomé D, Bouillaud F, Davila A-M, Blachier F (2016) Detrimental effects for colonocytes of an increased exposure to luminal hydrogen sulfide: the adaptive response. Free Radical Biol Med 93:155–164
Bellezza I, Giambanco I, Minelli A, Donato R (2018) Nrf2-Keap1 signaling in oxidative and reductive stress. Biochimica Et Biophysica Acta. Mol Cell Res 1865(5):721–733
Blachier F, Mariotti F, Huneau JF, Tomé D (2007) Effects of amino acid-derived luminal metabolites on the colonic epithelium and physiopathological consequences. Amino Acids 33(4):547–562
Blachier F, Beaumont M, Andriamihaja M, Davila AM, Lan A, Grauso M, Armand L, Benamouzig R, Tomé D (2017) Changes in the luminal environment of the colonic epithelial cells and physiopathological consequences. Am J Pathol 187(3):476–486
Brial F, Le Lay A, Dumas M-E, Gauguier D (2018) Implication of gut microbiota metabolites in cardiovascular and metabolic diseases. Cell Mol Life Sci 75(21):3977–3990
Bulus H, Oguztuzun S, Güler Simsek G, Kilic M, Ada AO, Göl S, Kocdogan AK, Kaygın P, Bozer B, Iscan M (2019) Expression of CYP and GST in human normal and colon tumor tissues. Biotech Histochem 94(1):1–9
Chappell CL, Darkoh C, Shimmin L, Farhana N, Kim D-K, Okhuysen PC, Hixson J (2016) Fecal indole as a biomarker of susceptibility to cryptosporidium infection. Infect Immun 84(8):2299–2306
Cheng T-H, Ma M-C, Liao M-T, Zheng C-M, Lu K-C, Liao C-H, Hou Y-C, Liu W-C, Lu C-L (2020) Indoxyl sulfate, a tubular toxin, contributes to the development of chronic kidney disease. Toxins 12(11):684
Darkoh C, Chappell C, Gonzales C, Okhuysen P (2015) A rapid and specific method for the detection of indole in complex biological samples. Appl Environ Microbiol 81:8093–8097
Gabor F, Stangl M, Wirth M (1998) Lectin-mediated bioadhesion: Binding characteristics of plant lectins on the enterocyte-like cell lines Caco-2, HT-29 and HCT-8. J Control Release 55(2–3):131–142
Gillam EM, Notley LM, Cai H, De Voss JJ, Guengerich FP (2000) Oxidation of indole by cytochrome P450 enzymes. Biochemistry 39(45):13817–13824
Grauso M, Lan A, Andriamihaja M, Bouillaud F, Blachier F (2019) Hyperosmolar environment and intestinal epithelial cells: impact on mitochondrial oxygen consumption, proliferation, and barrier function in vitro. Sci Rep 9(1):11360
Gryp T, De Paepe K, Vanholder R, Kerckhof FM, Van Biesen W, Van de Wiele T, Verbeke F, Speeckaert M, Joossens M, Couttenye MM, Vaneechoutte M, Glorieux G (2020) Gut microbiota generation of protein-bound uremic toxins and related metabolites is not altered at different stages of chronic kidney disease. Kidney Int 97(6):1230–1242
Hayes JD, Dinkova-Kostova AT (2014) The Nrf2 regulatory network provides an interface between redox and intermediary metabolism. Trends Biochem Sci 39(4):199–218
Hobby GP, Karaduta O, Dusio GF, Singh M, Zybailov BL, Arthur JM (2019) Chronic kidney disease and the gut microbiome. Am J Physiol Renal Physiol 316(6):F1211–F1217
Jamka M, Kokot M, Kaczmarek N, Bermagambetova S, Nowak JK, Walkowiak J (2021) The effect of sodium butyrate enemas compared with placebo on disease activity, endoscopic scores, and histological and inflammatory parameters in inflammatory bowel diseases: a systematic review of randomised controlled trials. Complement Med Res 28(4):344–356
Jastroch M, Divakaruni AS, Mookerjee S, Treberg JR, Brand MD (2010) Mitochondrial proton and electron leaks. Essays Biochem 47:53–67
Ji Y, Yin W, Liang Y, Sun L, Yin Y, Zhang W (2020) Anti-inflammatory and anti-oxidative activity of indole-3-acetic acid involves induction of HO-1 and neutralization of free radicals in RAW264.7 cells. Int J Mol Sci 21(5):1579
King LJ, Parke DV, Williams RT (1966) The metabolism of [2-14C] indole in the rat. Biochem J 98(1):266–277
Lano G, Burtey S, Sallée M (2020) Indoxyl sulfate, a uremic endotheliotoxin. Toxins 12(4):229
Lee J-H, Lee J (2010) Indole as an intercellular signal in microbial communities. FEMS Microbiol Rev 34(4):426–444
Leong SC, Sirich TL (2016) Indoxyl sulfate-review of toxicity and therapeutic strategies. Toxins (basel) 8(12):358
Leschelle X, Delpal S, Goubern M, Blottière HM, Blachier F (2000) Butyrate metabolism upstream and downstream acetyl-CoA synthesis and growth control of human colon carcinoma cells: butyrate metabolism and colon cell growth. Eur J Biochem 267(21):6435–6442
Liu X, Blouin JM, Santacruz A, Lan A, Andriamihaja M, Wilkanowicz S, Benetti PH, Tomé D, Sanz Y, Blachier F, Davila AM (2014) High-protein diet modifies colonic microbiota and luminal environment but not colonocyte metabolism in the rat model: the increased luminal bulk connection. Am J Physiol Gastrointest Liver Physiol 307(4):G459–G470
Louis P, Hold GL, Flint HJ (2014) The gut microbiota, bacterial metabolites and colorectal cancer. Nat Rev Microbiol 12(10):661–672
McNabney SM, Henagan TM (2017) Short chain fatty acids in the colon and peripheral tissues: a focus on butyrate, colon cancer, obesity and insulin resistance. Nutrients 9(12):1348
Mimoun S, Andriamihaja M, Chaumontet C, Atanasiu C, Benamouzig R, Blouin JM, Tomé D, Bouillaud F, Blachier F (2012) Detoxification of H(2)S by differentiated colonic epithelial cells: Implication of the sulfide oxidizing unit and of the cell respiratory capacity. Antioxid Redox Signal 17(1):1–10
Parada Venegas D, De la Fuente MK, Landskron G, González MJ, Quera R, Dijkstra G, Harmsen HJM, Faber KN, Hermoso MA (2019) Short chain fatty acids (SCFAs)-mediated gut epithelial and immune regulation and its relevance for inflammatory bowel diseases. Front Immunol 10:277
Poesen R, Mutsaers HAM, Windey K, van den Broek PH, Verweij V, Augustijns P, Kuypers D, Jansen J, Evenepoel P, Verbeke K, Meijers B, Masereeuw R (2015) The influence of dietary protein intake on mammalian tryptophan and phenolic metabolites. PLoS ONE 10(10):e0140820
Portune KJ, Beaumont M, Davila A-M, Tomé D, Blachier F, Sanz Y (2016) Gut microbiota role in dietary protein metabolism and health-related outcomes: the two sides of the coin. Trends Food Sci Technol 57:213–232
Reuter S, Gupta SC, Chaturvedi MM, Aggarwal BB (2010) Oxidative stress, inflammation, and cancer: How are they linked? Free Radical Biol Med 49(11):1603–1616
Rosenberg DW, Mankowski DC (1994) Induction of cyp2e-1 protein in mouse colon. Carcinogenesis 15:73–78
Schroeder JC, DiNatale BC, Murray IA, Flaveny CA, Liu Q, Laurenzana EM, Lin JM, Strom SC, Omiecinski CJ, Amin S, Perdew GH (2010) The uremic toxin 3-indoxyl sulfate is a potent endogenous agonist for the human aryl hydrocarbon receptor. Biochemistry 49(2):393–400
Shimada Y, Kinoshita M, Harada K, Mizutani M, Masahata K, Kayama H, Takeda K (2013) Commensal bacteria-dependent indole production enhances epithelial barrier function in the colon. PLoS ONE 8(11):e80604
Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175(1):184–191
Singh R, Chandrashekharappa S, Bodduluri SR, Baby BV, Hegde B, Kotla NG, Hiwale AA, Saiyed T, Patel P, Vijay-Kumar M, Langille MGI, Douglas GM, Cheng X, Rouchka EC, Waigel SJ, Dryden GW, Alatassi H, Zhang H-G, Haribabu B et al (2019) Enhancement of the gut barrier integrity by a microbial metabolite through the Nrf2 pathway. Nat Commun 10(1):89
Tebay LE, Robertson H, Durant ST, Vitale SR, Penning TM, Dinkova-Kostova AT, Hayes JD (2015) Mechanisms of activation of the transcription factor Nrf2 by redox stressors, nutrient cues, and energy status and the pathways through which it attenuates degenerative disease. Free Radical Biol Med 88(0 0):108–146
Vanholder R, Baurmeister U, Brunet P, Cohen G, Glorieux G, Jankowski J, European Uremic Toxin Work Group (2008) A bench to bedside view of uremic toxins. J Am Soc Nephrol JASN 19(5):863–870
Vanholder R, Schepers E, Pletinck A, Nagler EV, Glorieux G (2014) The uremic toxicity of indoxyl sulfate and p-cresyl sulfate: a systematic review. J Am Soc Nephrol 25(9):1897–1907
Wardyn JD, Ponsford AH, Sanderson CM (2015) Dissecting molecular cross-talk between Nrf2 and NF-κB response pathways. Biochem Soc Trans 43(4):621–626
Whitfield-Cargile CM, Cohen ND, Chapkin RS, Weeks BR, Davidson LA, Goldsby JS, Hunt CL, Steinmeyer SH, Menon R, Suchodolski JS, Jayaraman A, Alaniz RC (2016) The microbiota-derived metabolite indole decreases mucosal inflammation and injury in a murine model of NSAID enteropathy. Gut Microbes 7(3):246–261
Zapletal O, Tylichová Z, Neča J, Kohoutek J, Machala M, Milcová A, Pokorná M, Topinka J, Moyer MP, Hofmanová J, Kozubík A, Vondráček J (2017) Butyrate alters expression of cytochrome P450 1A1 and metabolism of benzo[a]pyrene via its histone deacetylase activity in colon epithelial cell models. Arch Toxicol 91(5):2135–2150
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
None of the authors declare conflict of interest.
Research involving human participants and/or animals
Not applicable.
Informed consent
Not applicable.
Additional information
Handling editor: G. Wu.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Armand, L., Fofana, M., Couturier-Becavin, K. et al. Dual effects of the tryptophan-derived bacterial metabolite indole on colonic epithelial cell metabolism and physiology: comparison with its co-metabolite indoxyl sulfate. Amino Acids 54, 1371–1382 (2022). https://doi.org/10.1007/s00726-021-03122-4
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00726-021-03122-4