{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T02:17:53Z","timestamp":1790821073200,"version":"4.1.0"},"reference-count":44,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2021,9,16]],"date-time":"2021-09-16T00:00:00Z","timestamp":1631750400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000928","name":"Welch Foundation","doi-asserted-by":"publisher","award":["BI-0028"],"award-info":[{"award-number":["BI-0028"]}],"id":[{"id":"10.13039\/100000928","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Nutrients"],"abstract":"<jats:p>\u03b2-Hydroxy-\u03b2-methylbutyrate (HMB), a leucine metabolite, is used as a nutritional ingredient to improve skeletal muscle health. Preclinical studies indicate that this supplement also elicits significant benefits in the brain; it promotes neurite outgrowth and prevents age-related reductions in neuronal dendrites and cognitive performance. As orally administered HMB elicits these effects in the brain, we infer that HMB crosses the blood\u2013brain barrier (BBB). However, there have been no reports detailing the transport mechanism for HMB in BBB. Here we show that HMB is taken up in the human BBB endothelial cell line hCMEC\/D3 via H+-coupled monocarboxylate transporters that also transport lactate and \u03b2-hydroxybutyrate. MCT1 (monocarboxylate transporter 1) and MCT4 (monocarboxylate transporter 4) belonging to the solute carrier gene family SLC16 (solute carrier, gene family 16) are involved, but additional transporters also contribute to the process. HMB uptake in BBB endothelial cells results in intracellular acidification, demonstrating cotransport with H+. Since HMB is known to activate mTOR with potential to elicit transcriptomic changes, we examined the influence of HMB on the expression of selective transporters. We found no change in MCT1 and MCT4 expression. Interestingly, the expression of LAT1 (system L amino acid transporter 1), a high-affinity transporter for branched-chain amino acids relevant to neurological disorders such as autism, is induced. This effect is dependent on mTOR (mechanistic target of rapamycine) activation by HMB with no involvement of histone deacetylases. These studies show that HMB in systemic circulation can cross the BBB via carrier-mediated processes, and that it also has a positive influence on the expression of LAT1, an important amino acid transporter in the BBB.<\/jats:p>","DOI":"10.3390\/nu13093220","type":"journal-article","created":{"date-parts":[[2021,9,18]],"date-time":"2021-09-18T00:13:47Z","timestamp":1631924027000},"page":"3220","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":16,"title":["A Proton-Coupled Transport System for \u03b2-Hydroxy-\u03b2-Methylbutyrate (HMB) in Blood\u2013Brain Barrier Endothelial Cell Line hCMEC\/D3"],"prefix":"10.3390","volume":"13","author":[{"given":"Kei","family":"Higuchi","sequence":"first","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9230-0144","authenticated-orcid":false,"given":"Sathish","family":"Sivaprakasam","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Souad R.","family":"Sennoune","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiro","family":"Ogura","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yangzom D.","family":"Bhutia","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2127-3190","authenticated-orcid":false,"given":"Ricardo","family":"Rueda","sequence":"additional","affiliation":[{"name":"Abbott Nutrition, Research & Development, 18004 Granada, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5491-7266","authenticated-orcid":false,"given":"Suzette L.","family":"Pereira","sequence":"additional","affiliation":[{"name":"Abbott Nutrition, Research & Development, Columbus, OH 43219, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vadivel","family":"Ganapathy","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, 3601 4th Street, Lubbock, TX 79430, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2021,9,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"765","DOI":"10.1042\/BCJ20200686","article-title":"Whole-body metabolic fate of branched-chain amino acids","volume":"478","author":"Blair","year":"2021","journal-title":"Biochem. J."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Nie, C., He, T., Zhang, W., Zhang, G., and Ma, X. (2018). Branched chain amino acids: Beyond nutrition metabolism. Int. J. Mol. Sci., 19.","DOI":"10.3390\/ijms19040954"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"101198","DOI":"10.1016\/j.arr.2020.101198","article-title":"Branched chain amino acids, aging and age-related health","volume":"64","author":"Cogger","year":"2020","journal-title":"Ageing Res. Rev."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1186\/1743-7075-8-11","article-title":"\u03b2-Hydroxy-\u03b2-methylbutyrate (HM\u03b2) supplementation stimulates skeletal muscle hypertrophy in rats via the mTOR pathway","volume":"8","author":"Pimentel","year":"2011","journal-title":"Nutr. Metab. (Lond.)"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"368","DOI":"10.1016\/j.nutres.2014.02.003","article-title":"\u03b2-Hydroxy-\u03b2-methylbutyrate facilitates PI3K\/Akt-dependent mammalian target of rapamycin and FoxO1\/3a phosphorylations and alleviates tumor necrosis factor \u03b1\/interferon \u03b3-induced MuRF-1 expression in C2C12 cells","volume":"34","author":"Kimura","year":"2014","journal-title":"Nutr. Res."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"211","DOI":"10.2478\/hukin-2019-0070","article-title":"Mechanism of action and the effect of \u03b2-hydroxy-\u03b2-methylbutyrate (HMB) supplementation on different types of physical performance\u2014A systematic review","volume":"68","author":"Kaczka","year":"2019","journal-title":"J. Hum. Kinet."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"10019","DOI":"10.1096\/fj.201900665RR","article-title":"Gut microbiota mediates the protective effects of dietary \u03b2-Hydroxy-\u03b2-methylbutyrate (HMB) against obesity induced by high-fat diets","volume":"33","author":"Duan","year":"2019","journal-title":"FASEB J."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"11560","DOI":"10.1158\/0008-5472.CAN-06-1950","article-title":"SLC5A8 triggers tumor cell apoptosis through pyruvate-dependent inhibition of histone deacetylases","volume":"66","author":"Thangaraju","year":"2006","journal-title":"Cancer Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"27601","DOI":"10.1074\/jbc.M110.102947","article-title":"Blockade of dendritic cell development by bacterial fermentation products butyrate and propionate through a transporter (Slc5a8)-dependent inhibition of histone deacetylases","volume":"285","author":"Singh","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"869","DOI":"10.1016\/j.coph.2013.08.006","article-title":"Transporters and receptors for short-chain fatty acids as the molecular link between colonic bacteria and the host","volume":"13","author":"Ganapathy","year":"2013","journal-title":"Curr. Opin. Pharmacol."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Sivaprakasam, S., Bhutia, Y.D., Ramachandran, S., and Ganapathy, V. (2017). Cell-surface and nuclear receptors in the colon as targets for bacterial metabolites and its relevance to colon health. Nutrients, 9.","DOI":"10.3390\/nu9080856"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Saito, R., Vilchez, J.D., Giron, M.D., Cabrera, E., Campos, N., Manzano, M., Rueda, R., and Lopez-Pedrosa, J.M. (2015). \u03b2-Hydroxy-\u03b2-methylbutyrate (HMB) promotes neurite outgrowth in Neuro2a cells. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0135614"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1016\/j.neurobiolaging.2016.01.004","article-title":"\u03b2-Hydroxy-\u03b2-methylbutyrate ameliorates aging effects in the dendritic tree of pyramidal neurons in the medial prefrontal cortex of both male and female rats","volume":"40","author":"Kougias","year":"2016","journal-title":"Neurobiol. Aging"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"93","DOI":"10.1016\/j.physbeh.2016.12.025","article-title":"\u03b2-Hydroxy-\u03b2-methylbutyrate (HMB) ameliorates age-related deficits in water maze performance, especially in male rats","volume":"170","author":"Kougias","year":"2017","journal-title":"Physiol. Behav."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"84","DOI":"10.1007\/s11095-019-2626-3","article-title":"Transport mechanisms for the nutritional supplement \u03b2-hydroxy-\u03b2-methylbutyrate (HMB) in mammalian cells","volume":"36","author":"Ogura","year":"2019","journal-title":"Pharm. Res."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"466","DOI":"10.1124\/pr.119.018762","article-title":"Monocarboxylate transporters (SLC16): Function, regulation, and role in health and disease","volume":"72","author":"Felmlee","year":"2020","journal-title":"Pharmacol. Rev."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"337","DOI":"10.1016\/j.mam.2012.05.003","article-title":"The SLC16 gene family\u2014Structure, role and regulation in health and disease","volume":"34","author":"Halestrap","year":"2013","journal-title":"Mol. Aspects Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"333","DOI":"10.1111\/j.1471-4159.2011.07208.x","article-title":"Quantitative targeted absolute proteomics of human blood\u2013brain transporters and receptors","volume":"117","author":"Uchida","year":"2011","journal-title":"J. Neurochem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/s00429-013-0672-x","article-title":"Monocarboxylate transporters in temporal lobe epilepsy: Roles of lactate and ketogenic diet","volume":"220","author":"Lauritzen","year":"2015","journal-title":"Brain Struct. Funct."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2481","DOI":"10.1016\/j.bbamcr.2016.03.013","article-title":"Monocarboxylate transporters in the brain and in cancer","volume":"1863","author":"Sboarina","year":"2016","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"H11478","DOI":"10.1152\/ajpheart.00166.2006","article-title":"Vacuolar type H+-ATPases at the plasma membrane regulate pH and cell migration in microvascular endothelial cells","volume":"291","author":"Rojas","year":"2006","journal-title":"Am. J. Physiol. Heart Circ. Physiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1152\/physrev.00025.2013","article-title":"The organic anion transporter (OAT) family: A systems biology perspective","volume":"95","author":"Nigam","year":"2015","journal-title":"Physiol. Rev."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"391","DOI":"10.1038\/sj.bjp.0704875","article-title":"Transport and uptake of nateglinide in Caco-2 cells and its inhibitory effect on human monocarboxylate transporter MCT1","volume":"137","author":"Okamura","year":"2002","journal-title":"Br. J. Pharmacol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1113\/jphysiol.2002.026500","article-title":"Structure, function and immunolocalization of a proton-coupled amino acid transporter (hPAT1) in the human intestinal cell line Caco-2","volume":"546","author":"Chen","year":"2003","journal-title":"J. Physiol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1802","DOI":"10.1111\/j.1476-5381.2011.01438.x","article-title":"The SLC36 family of proton-coupled amino acid transporters and their potential role in drug transport","volume":"164","author":"Thwaites","year":"2011","journal-title":"Br. J. Pharmacol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1758","DOI":"10.1096\/fj.03-1387fje","article-title":"A novel bifunctionality: PAT1 and PAT2 mediate electrogenic proton\/amino acid and electroneutral proton\/fatty acid symport","volume":"18","author":"Foltz","year":"2004","journal-title":"FASEB J."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"908","DOI":"10.1158\/0008-5472.CAN-13-2034","article-title":"Blocking lactate export by inhibiting the Myc target MCT1 disables glycolysis and glutathione synthesis","volume":"74","author":"Doherty","year":"2014","journal-title":"Cancer Res."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"427","DOI":"10.1016\/j.bbrc.2017.10.025","article-title":"Identification of a selective inhibitor of human monocarboxylate transporter 4","volume":"495","author":"Futagi","year":"2018","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1006\/bbrc.1997.6588","article-title":"Identification of a unique monocarboxylate transporter (MCT3) in retinal pigment epithelium","volume":"234","author":"Yoon","year":"1997","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"282","DOI":"10.1101\/gad.180968.111","article-title":"A monocarboxylate transporter required for hepatocyte secretion of ketone bodies during fasting","volume":"26","author":"Hugo","year":"2012","journal-title":"Genes Dev."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1236","DOI":"10.1101\/gad.327056.119","article-title":"MYC promotes tryptophan uptake and metabolism by the kynurenine pathway in colon cancer","volume":"33","author":"Venkateswaran","year":"2019","journal-title":"Genes Dev."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Singh, N., and Ecker, G.F. (2018). Insights into the structure, function, and ligand discovery of the large neutral amino acid transporter 1, LAT1. Int. J. Mol. Sci., 19.","DOI":"10.3390\/ijms19051278"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1481","DOI":"10.1016\/j.cell.2016.11.013","article-title":"Impaired amino acid transport at the blood brain barrier is a cause of autism spectrum disorder","volume":"167","author":"Tarlungeanu","year":"2016","journal-title":"Cell"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"e1036","DOI":"10.1002\/mgg3.1036","article-title":"Abnormalities in the genes that encode large amino acid transporters increase the risk of autism spectrum disorder","volume":"8","author":"Cascio","year":"2020","journal-title":"Mol. Genet. Genomic Med."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1002\/cphy.c170014","article-title":"Short-chain fatty acid transporters: Role in colonic homeostasis","volume":"8","author":"Sivaprakasam","year":"2017","journal-title":"Compr. Physiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1872","DOI":"10.1096\/fj.04-3458fje","article-title":"Blood\u2013brain barrier-specific properties of a human adult brain endothelial cell line","volume":"19","author":"Weksler","year":"2005","journal-title":"FASEB J."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"862","DOI":"10.1177\/0271678X16630991","article-title":"In vitro models of the blood\u2013brain barrier: An overview of commonly used brain endothelial cell culture models and guidelines for their use","volume":"36","author":"Helms","year":"2016","journal-title":"J. Cereb. Blood Flow Metab."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1021\/mp3004308","article-title":"Quantitative targeted absolute proteomic analysis of transporters, receptors and junction proteins for validation of human cerebral microvascular endothelial cell line hCMEC\/D3 as a human blood\u2013brain barrier model","volume":"10","author":"Ohtsuki","year":"2013","journal-title":"Mol. Pharm."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/j.1471-4159.2005.03168.x","article-title":"Monocarboxylate transporters in the central nervous system: Distribution, regulation and function","volume":"94","author":"Pierre","year":"2005","journal-title":"J. Neurochem."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"176","DOI":"10.1038\/jcbfm.2014.206","article-title":"Lactate transport and signaling in the brain: Potential therapeutic targets and roles in body-brain interaction","volume":"35","author":"Bergersen","year":"2015","journal-title":"J. Cereb. Blood Flow Metab."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"20135","DOI":"10.1074\/jbc.RA119.009093","article-title":"Monocarboxylate transporter 4 (MCT4) is a high affinity transporter capable of exporting lactate in high-lactate microenvironments","volume":"294","author":"Sandoval","year":"2019","journal-title":"J. Biol. Chem."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"423","DOI":"10.1016\/j.neuroscience.2008.06.015","article-title":"Subcellular localization of transporters along the rat blood\u2013brain barrier and blood-cerebral-spinal fluid barrier by in vivo biotinylation","volume":"155","author":"Roberts","year":"2008","journal-title":"Neuroscience"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"14706","DOI":"10.1038\/ncomms14706","article-title":"Adipocytes promote malignant growth of breast tumours with monocarboxylate transporter 2 expression via \u03b2-hydroxybutyrate","volume":"8","author":"Huang","year":"2017","journal-title":"Nat. Commun."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"C984","DOI":"10.1152\/ajpcell.00146.2011","article-title":"The short-chain fatty acid butyrate is a substrate of breast cancer resistance protein","volume":"301","author":"Goncalves","year":"2011","journal-title":"Am. J. Physiol. Cell Physiol."}],"container-title":["Nutrients"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6643\/13\/9\/3220\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:00:41Z","timestamp":1760166041000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6643\/13\/9\/3220"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,9,16]]},"references-count":44,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2021,9]]}},"alternative-id":["nu13093220"],"URL":"https:\/\/doi.org\/10.3390\/nu13093220","relation":{},"ISSN":["2072-6643"],"issn-type":[{"value":"2072-6643","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,9,16]]}}}