Environmental Enteric Dysfunction is Associated with Carnitine Deficiency and Altered Fatty Acid Oxidation

Richard D Semba, Indi Trehan, Ximin Li, Ruin Moaddel, M Isabel Ordiz, Kenneth M Maleta, Klaus Kraemer, Michelle Shardell, Luigi Ferrucci, Mark Manary, Richard D Semba, Indi Trehan, Ximin Li, Ruin Moaddel, M Isabel Ordiz, Kenneth M Maleta, Klaus Kraemer, Michelle Shardell, Luigi Ferrucci, Mark Manary

Abstract

Background: Environmental enteric dysfunction (EED), a condition characterized by small intestine inflammation and abnormal gut permeability, is widespread in children in developing countries and a major cause of growth failure. The pathophysiology of EED remains poorly understood.

Methods: We measured serum metabolites using liquid chromatography-tandem mass spectrometry in 400 children, aged 12-59months, from rural Malawi. Gut permeability was assessed by the dual-sugar absorption test.

Findings: 80.7% of children had EED. Of 677 serum metabolites measured, 21 were negatively associated and 56 were positively associated with gut permeability, using a false discovery rate approach (q<0.05, p<0.0095). Increased gut permeability was associated with elevated acylcarnitines, deoxycarnitine, fatty acid β-oxidation intermediates, fatty acid ω-oxidation products, odd-chain fatty acids, trimethylamine-N-oxide, cystathionine, and homocitrulline, and with lower citrulline, ornithine, polyphenol metabolites, hippurate, tryptophan, and indolelactate.

Interpretation: EED is a syndrome characterized by secondary carnitine deficiency, abnormal fatty acid oxidation, alterations in polyphenol and amino acid metabolites, and metabolic dysregulation of sulfur amino acids, tryptophan, and the urea cycle. Future studies are needed to corroborate the presence of secondary carnitine deficiency among children with EED and to understand how these metabolic derangements may negatively affect the growth and development of young children.

Keywords: Acylcarnitines; Carnitine; Environmental enteric dysfunction; Fatty acid oxidation; Hippurate; Polyphenols; Tryptophan; Urea cycle.

Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Figures

Fig. 1
Fig. 1
The carnitine shuttle. Carnitine enters the cell through active transport by the high affinity carnitine transporter, organic cation transporter novel 2 (OCTN2). Long-chain fatty acid-CoA in the cytosol exchanges CoA for carnitine by the action of carnitine palmitoyltransferase I (CPT1) in the outer mitochondrial membrane. Acylcarnitine moves into the mitochondrial matrix by facilitated diffusion through a transporter, carnitine-acylcarnitine translocase (CACT), on the inner mitochondrial membrane, in exchange for carnitine. In the mitochondrial matrix, the acyl group is transferred to mitochondrial coenzyme A by carnitine palmitoyltransferase II (CPT2). Carnitine is then free to cycle back to the cytosol through the transporter. Carnitine acyltransferase (CAT) removes CoA from acetyl-CoA that is formed from β-oxidation to form acetylcarnitine. Acetylcarnitine can exit the mitochondria via CACT and enter the blood via OCTN2.
Fig. 2
Fig. 2
Volcano plot showing relationship of partial Spearman correlations between gut permeability, as measured by the L:M ratio, and serum metabolites, adjusted for age, sex, and village. Horizontal line indicates significance at p-value of 0.0096, which corresponds to a q-value

Fig. 3

Serum metabolite profile in secondary…

Fig. 3

Serum metabolite profile in secondary carnitine deficiency. With low carnitine and a defective…

Fig. 3
Serum metabolite profile in secondary carnitine deficiency. With low carnitine and a defective carnitine shuttle, β-oxidation is blocked and intermediates of blocked β-oxidation accumulate. To compensate for blocked β-oxidation, ω-oxidation, which is usually a minor fatty acid oxidation pathway in endoplasmic reticulum, is upregulated with accumulation of ω-oxidation products.

Fig. 4

Metabolism of sulfur amino acids.…

Fig. 4

Metabolism of sulfur amino acids. Methionine is metabolized through transmethylation produces homocysteine. Homocysteine…

Fig. 4
Metabolism of sulfur amino acids. Methionine is metabolized through transmethylation produces homocysteine. Homocysteine is remethylated through 5-methyl tetrahydrofolate (THF) to produce methionine. During inflammation, homocysteine undergoes transsulfuration to produce cystathione and eventually taurine. Children with increased gut permeability had elevated serum cystathione and taurine, suggesting an increase in the transsulfuration pathway.

Fig. 5

Abnormalities in serum metabolites known…

Fig. 5

Abnormalities in serum metabolites known to be associated with abnormal gut microbiome. Metabolites…

Fig. 5
Abnormalities in serum metabolites known to be associated with abnormal gut microbiome. Metabolites known to be related to dietary polyphenols and an abnormal gut microbiome were decreased in children with increased gut permeability. Metabolites known to be associated with amino acid fermentation by an abnormal gut microbiome were increased in children with increased gut permeability.
Fig. 3
Fig. 3
Serum metabolite profile in secondary carnitine deficiency. With low carnitine and a defective carnitine shuttle, β-oxidation is blocked and intermediates of blocked β-oxidation accumulate. To compensate for blocked β-oxidation, ω-oxidation, which is usually a minor fatty acid oxidation pathway in endoplasmic reticulum, is upregulated with accumulation of ω-oxidation products.
Fig. 4
Fig. 4
Metabolism of sulfur amino acids. Methionine is metabolized through transmethylation produces homocysteine. Homocysteine is remethylated through 5-methyl tetrahydrofolate (THF) to produce methionine. During inflammation, homocysteine undergoes transsulfuration to produce cystathione and eventually taurine. Children with increased gut permeability had elevated serum cystathione and taurine, suggesting an increase in the transsulfuration pathway.
Fig. 5
Fig. 5
Abnormalities in serum metabolites known to be associated with abnormal gut microbiome. Metabolites known to be related to dietary polyphenols and an abnormal gut microbiome were decreased in children with increased gut permeability. Metabolites known to be associated with amino acid fermentation by an abnormal gut microbiome were increased in children with increased gut permeability.

References

    1. Alp H., Orbak Z., Akçay F. Plasma and urine carnitine levels and carnitine supplementation in children with malnutrition. J. Trop. Pediatr. 1999;45:294–296.
    1. Bickler S.W., Ring J., De Maio A. Sulfur amino acid metabolism limits the growth of children living in environments of poor sanitation. Med. Hypotheses. 2011;77:380–382.
    1. Breiman L. Random forests. Mach. Learn. 2001;45:5–32.
    1. Campbell D.I., Murch S.H., Elia M. Chronic T cell-mediated enteropathy in rural west African children: relationship with nutritional status and small bowel function. Pediatr. Res. 2003;54:306–311.
    1. Carroll J.E., Brooke M.H., Shumate J.B. Carnitine intake and excretion in neuromuscular diseases. Am. J. Clin. Nutr. 1981;34:2693–2698.
    1. Chickos J.S., Way B.A., Wilson J. Analysis of 3-hydroxydodecanedioic acid for studies of fatty acid metabolic disorders: preparation of stable isotope standards. J. Clin. Lab. Anal. 2002;16:115–120.
    1. Crane R.J., Jones K.D., Berkley J.A. Environmental enteric dysfunction: an overview. Food Nutr. Bull. 2015;36(1 Suppl):S76–S87.
    1. Crill C.M., Helms R.A. The use of carnitine in pediatric nutrition. Nutr. Clin. Pract. 2007;22:204–213.
    1. da Rosa M.S., Seminotti B., Ribeiro C.A. 3-Hydroxy-3-methylglutaric and 3-methylglutaric acids impair redox status and energy production and transfer in rat heart: relevance for the pathophysiology of cardiac dysfunction in 3-hydroxy-3-methylglutaryl-coenzyme A lyase deficiency. Free Radic. Res. 2016;50:997–1010.
    1. DeHaven C., Evans A., Dai H. Organization of GC/MS and LC/MS metabolomics data into chemical libraries. J. Cheminform. 2010;2:9.
    1. Demarquoy J., Georges B., Rigault C. Radioisotopic determination of l-carnitine content in foods commonly eaten in Western countries. Food Chem. 2004;2004(86):137–142.
    1. Denno D.M., VanBuskirk K., Nelson Z.C. Use of the lactulose to mannitol ratio to evaluate childhood environmental enteric dysfunction: a systematic review. Clin. Infect. Dis. 2014;59(Suppl. 4):S213–S219.
    1. Dorland L., Ketting D., Bruinvis L. Medium- and long-chain 3-hydroxymonocarboxylic acids: analysis by gas chromatography combined with mass spectrometry. Biomed. Chromatogr. 1991;5:161–164.
    1. Dror D.K., Allen L.H. The importance of milk and other animal-source foods for children in low-income countries. Food Nutr. Bull. 2011;32:227–243.
    1. Evans A.M., Bridgewater B.R., Liu Q. High resolution mass spectrometry improves data quantity and quality as compared to unit mass resolution mass spectrometry in high-throughput profiling metabolomics. Metabolomics. 2014;4:132.
    1. Evans A.M., DeHaven C.D., Barrett T. Integrated, nontargeted ultrahigh performance liquid chromatography/electrospray ionization tandem mass spectrometry platform for the identification and relative quantification of the small-molecule complement of biological systems. Anal. Chem. 2009;81:6656–6667.
    1. Friedman J., Hastie T., Tibshirani R. Regularization paths for generalized linear models via coordinate descent. J. Stat. Softw. 2010;33:1–22.
    1. Friedman J.H. Multivariate adaptive regression splines (with discussion) Ann. Stat. 1991;19:1–141.
    1. Friedman J.H. Greedy function approximation: a gradient boosting machine. Ann. Stat. 2001;29:1189–1232.
    1. Giesbertz P., Ecker J., Haag A. An LC-MS/MS method to quantify acylcarnitine species including isomeric and odd-numbered forms in plasma and tissues. J. Lipid Res. 2015;56:2029–2039.
    1. Guerrant R.L., Leite A.M., Pinkerton R. Biomarkers of environmental enteropathy, inflammation, stunting, and impaired growth in children in northeast Brazil. PLoS One. 2016;2016(11)
    1. Jenkins B., West J.A., Koulman A. A review of odd-chain fatty acid metabolism and the role of pentadecanoic acid (c15:0) and heptadecanoic acid (c17:0) in health and disease. Molecules. 2015;20:2425–2444.
    1. Kelly P., Besa E., Zyambo K. Endomicroscopic and transcriptomic analysis of impaired barrier function and malabsorption in environmental enteropathy. PLoS Negl. Trop. Dis. 2016;10
    1. Kendler B.S. Carnitine: an overview of its role in preventive medicine. Prev. Med. 1986;15:373–390.
    1. Khan L., Bamji M.S. Tissue carnitine deficiency due to dietary lysine deficiency: triglyceride accumulation and concomitant impairment in fatty acid oxidation. J. Nutr. 1979;109:24–31.
    1. Koeth R.A., Wang Z., Levison B.S. Intestinal microbiota metabolism of l-carnitine, a nutrient in red meat, promotes atherosclerosis. Nat. Med. 2013;19:576–585.
    1. Kosek M.N., Mduma E., Kosek P.S. Plasma tryptophan and the kynurenine-tryptophan ratio are associated with the acquisition of statural growth deficits and oral vaccine underperformance in populations with environmental enteropathy. Am.J.Trop. Med. Hyg. 2016;95:928–937.
    1. Krebs N.F., Miller L.V., Hambidge K.M. Zinc deficiency in infants and children: a review of its complex and synergistic interactions. Paediatr. Int. Child Health. 2014;34:279–288.
    1. Lees H.J., Swann J.R., Wilson I.D. Hippurate: the natural history of a mammalian-microbial cometabolite. J. Proteome Res. 2013;12:1527–1546.
    1. Lerner A., Gruener N., Iancu T.C. Serum carnitine concentrations in coeliac disease. Gut. 1993;34:933–935.
    1. Lord R.S., Bralley J.A., editors. Laboratory Evaluations for Integrative and Functional Medicine. Metametrix Institute; Duluth, Georgia: 2008.
    1. Mamedov I., Zolkina I., Nikolaeva E. Carnitine insufficiency in children with inborn errors of metabolism: prevalence and treatment efficacy. J. Pediatr. Endocrinol. Metab. 2015;28:1299–1304.
    1. Martin F.P., Ezri J., Cominetti O. Urinary metabolic phenotyping reveals differences in the metabolic status of healthy and inflammatory bowel disease (IBD) children in relation to growth and disease activity. Int. J. Mol. Sci. 2016;17(8)
    1. Minkler P.E., Stoll M.S., Ingalls S.T. Validated method for the quantification of free and total carnitine, butyrobetaine, and acylcarnitines in biological samples. Anal. Chem. 2015;87:8994–9001.
    1. Miura Y. The biological significance of ω-oxidation of fatty acids. Proc. Jpn. Acad. Ser. B Phys. Biol. Sci. 2013;89:370–382.
    1. Naseri M., Mottaghi Moghadam Shahri H., Horri M. Absolute and relative carnitine deficiency in patients on hemodialysis and peritoneal dialysis. Iran. J. Kidney Dis. 2016;10:36–43.
    1. Nuss E.T., Tanumihardjo S.A. Quality protein maize for Africa: closing the protein inadequacy gap in vulnerable populations. Adv. Nutr. 2011;2:217–224.
    1. Olson A.L., Nelson S.E., Rebouche C.J. Low carnitine intake and altered lipid metabolism in infants. Am. J. Clin. Nutr. 1989;49:624–628.
    1. Onkenhout W., Venizelos V., van der Poel P.F. Identification and quantification of intermediates of unsaturated fatty acid metabolism in plasma of patients with fatty acid oxidation disorders. Clin. Chem. 1995;41:1467–1474.
    1. Ordiz M.I., Stephenson K., Agapova S. Environmental enteric dysfunction and the fecal microbiota in Malawian children. Am.J.Trop. Med. Hyg. 2016 (Epub ahead of print)
    1. Pimpão R.C., Ventura M.R., Ferreira R.B. Phenolic sulfates as new and highly abundant metabolites in human plasma after ingestion of a mixed berry fruit purée. Br. J. Nutr. 2015;113:454–463.
    1. Poesen R., Claes K., Evenepoel P. Microbiota-derived phenylacetylglutamine associates with overall mortality and cardiovascular disease in patients with CKD. J. Am. Soc. Nephrol. 2016;27:3479–3487.
    1. Puisac B., Arnedo M., Casale C.H. Differential HMG-CoA lyase expression in human tissues provides clues about 3-hydroxy-3-methylglutaric aciduria. J. Inherit. Metab. Dis. 2010;33:405–410.
    1. Rachakonda V., Gabbert C., Raina A. Serum metabolomic profiling in acute alcoholic hepatitis identifies multiple dysregulated pathways. PLoS One. 2014;9
    1. Reuter S.E., Evans A.M. Carnitine and acylcarnitines: pharmacokinetic, pharmacological and clinical aspects. Clin. Pharmacokinet. 2012;51:553–572.
    1. Roberts L.D., Boström P., O'Sullivan J.F. β-Aminoisobutyric acid induces browning of white fat and hepatic β-oxidation and is inversely correlated with cardiometabolic risk factors. Cell Metab. 2014;19:96–108.
    1. Russell W.R., Duncan S.H., Scobbie L. Major phenylpropanoid-derived metabolites in the human gut can arise from microbial fermentation of protein. Mol. Nutr. Food Res. 2013;57:523–535.
    1. Seakins J.W. The determination of urinary phenylacetylglutamine as phenylacetic acid. Studies on its origin in normal subjects and children with cystic fibrosis. Clin. Chim. Acta. 1971;35:121–131.
    1. Semba R.D., Shardell M., Sakr Ashour F.A. Child stunting is associated with low circulating essential amino acids. EBioMedicine. 2016;6:246–252.
    1. Semba R.D., Shardell M., Trehan I. Metabolic alterations in children with environmental enteric dysfunction. Sci. Rep. 2016;6:28009.
    1. Semba R.D., Zhang P., Gonzalez-Freire M. The association of serum choline with linear growth failure in young children from rural Malawi. Am. J. Clin. Nutr. 2016;104:191–197.
    1. Sgambat K., Moudgil A. Carnitine deficiency in children receiving continuous renal replacement therapy. Hemodial. Int. 2016;20:63–67.
    1. Shekhawat P.S., Sonne S., Carter A.L. Enzymes involved in l-carnitine biosynthesis are expressed by small intestinal enterocytes in mice: implications for gut health. J. Crohns Colitis. 2013;7:e197–e205.
    1. Shekhawat P.S., Srinivas S.R., Matern D. Spontaneous development of intestinal and colonic atrophy and inflammation in the carnitine-deficient jvs (OCTN2(−/−)) mice. Mol. Genet. Metab. 2007;92:315–324.
    1. Sigalet D.L., Martin G.R., Poole A. Differential sugar absorption as a marker for adaptation in short bowel syndrome. J. Pediatr. Surg. 2000;35:661–664.
    1. Sonne S., Shekhawat P.S., Matern D. Carnitine deficiency in OCTN2 −/− newborn mice leads to a severe gut and immune phenotype with widespread atrophy, apoptosis and a pro-inflammatory response. PLoS One. 2012;7
    1. Stanley C.A. Carnitine deficiency disorders in children. Ann. N. Y. Acad. Sci. 2004;1033:42–51.
    1. Storey J.D. A direct approach to false discovery rates. J. R. Stat. Soc. Ser. B. 2002;64:479–498.
    1. Storey J.D., Taylor J.E., Siegmund D. Strong control, conservative point estimation, and simultaneous conservative consistency of false discovery rates: a unified approach. J. R. Stat. Soc. Ser. B. 2004;66:187–205.
    1. Tanner L.M., Näntö-Salonen K., Rashed M.S. Carnitine deficiency and l-carnitine supplementation in lysinuric protein intolerance. Metabolism. 2008;57:549–554.
    1. Tanphaichitr V., Broquist H.P. Lysine deficiency in the rat: concomitant impairment in carnitine biosynthesis. J. Nutr. 1973;103:80–87.
    1. Tein I. Disorders of fatty acid oxidation. Handb. Clin. Neurol. 2013;113:1675–1688.
    1. Trehan I., Kelly P., Shaikh N. New insights into environmental enteric dysfunction. Arch. Dis. Child. 2016;101:741–744.
    1. van der Laan M.J., Polley E.C., Hubbard A.E. Super learner. Stat. Appl. Genet. Mol. Biol. 2007;6 (Article 25)
    1. Watanabe K., Petri W.A., Jr. Environmental enteropathy: elusive but significant subclinical abnormalities in developing countries. EBioMedicine. 2016;10:25–32.
    1. Weisz A.J., Manary M.J., Stephenson K. Abnormal gut integrity is associated with reduced linear growth in rural Malawian children. J. Pediatr. Gastroenterol. Nutr. 2012;55:747–750.
    1. Wierzbicki A.S., Mayne P.D., Lloyd M.D. Metabolism of phytanic acid and 3-methyl-adipic acid excretion in patients with adult Refsum disease. J. Lipid Res. 2003;44:1481–1488.
    1. Winter S.C., Szabo-Aczel S., Curry C.J. Plasma carnitine deficiency. Clinical observations in 51 pediatric patients. Am. J. Dis. Child. 1987;141:660–665.
    1. Yu J., Ordiz M.I., Stauber J. Environmental enteric dysfunction includes a broad spectrum of inflammatory responses and epithelial repair processes. Cell. Mol. Gastroenterol. Hepatol. 2015;2:158–174.e1.

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