{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,16]],"date-time":"2026-10-16T02:02:29Z","timestamp":1792116149359,"version":"4.4.0"},"reference-count":70,"publisher":"National Academy of Sciences","issue":"21","license":[{"start":{"date-parts":[[2024,5,15]],"date-time":"2024-05-15T00:00:00Z","timestamp":1715731200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100018818","name":"National Research, Development and Innovation Office","doi-asserted-by":"publisher","award":["K143766"],"award-info":[{"award-number":["K143766"]}],"id":[{"id":"10.13039\/501100018818","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003825","name":"Magyar Tudom\u00e1nyos Akad\u00e9mia","doi-asserted-by":"publisher","award":["NAP2022-I-6\/2022"],"award-info":[{"award-number":["NAP2022-I-6\/2022"]}],"id":[{"id":"10.13039\/501100003825","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100018818","name":"National Research, Development and Innovation Office","doi-asserted-by":"publisher","award":["PD138930"],"award-info":[{"award-number":["PD138930"]}],"id":[{"id":"10.13039\/501100018818","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100020916","name":"E\u00f6tv\u00f6s Lor\u00e1nd Research Network","doi-asserted-by":"publisher","award":["SA-111\/2021"],"award-info":[{"award-number":["SA-111\/2021"]}],"id":[{"id":"10.13039\/100020916","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100018818","name":"National Research, Development and Innovation Office","doi-asserted-by":"publisher","award":["FK143233"],"award-info":[{"award-number":["FK143233"]}],"id":[{"id":"10.13039\/501100018818","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001602","name":"Science Foundation Ireland","doi-asserted-by":"publisher","award":["Eye-D-21\/SPP\/3732"],"award-info":[{"award-number":["Eye-D-21\/SPP\/3732"]}],"id":[{"id":"10.13039\/501100001602","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001602","name":"Science Foundation Ireland","doi-asserted-by":"publisher","award":["16\/RC\/3948"],"award-info":[{"award-number":["16\/RC\/3948"]}],"id":[{"id":"10.13039\/501100001602","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000781","name":"EC | European Research Council","doi-asserted-by":"publisher","award":["864522"],"award-info":[{"award-number":["864522"]}],"id":[{"id":"10.13039\/501100000781","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.pnas.org"],"crossmark-restriction":true},"short-container-title":["Proc. Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[2024,5,21]]},"abstract":"<jats:p>Blood\u2013brain barrier (BBB) models derived from human stem cells are powerful tools to improve our understanding of cerebrovascular diseases and to facilitate drug development for the human brain. Yet providing stem cell\u2013derived endothelial cells with the right signaling cues to acquire BBB characteristics while also retaining their vascular identity remains challenging. Here, we show that the simultaneous activation of cyclic AMP and Wnt\/\u03b2-catenin signaling and inhibition of the TGF-\u03b2 pathway in endothelial cells robustly induce BBB properties in vitro. To target this interaction, we present a small-molecule cocktail named cARLA, which synergistically enhances barrier tightness in a range of BBB models across species. Mechanistically, we reveal that the three pathways converge on Wnt\/\u03b2-catenin signaling to mediate the effect of cARLA via the tight junction protein claudin-5. We demonstrate that cARLA shifts the gene expressional profile of human stem cell\u2013derived endothelial cells toward the in vivo brain endothelial signature, with a higher glycocalyx density and efflux pump activity, lower rates of endocytosis, and a characteristic endothelial response to proinflammatory cytokines. Finally, we illustrate how cARLA can improve the predictive value of human BBB models regarding the brain penetration of drugs and targeted nanoparticles. Due to its synergistic effect, high reproducibility, and ease of use, cARLA has the potential to advance drug development for the human brain by improving BBB models across laboratories.<\/jats:p>","DOI":"10.1073\/pnas.2316006121","type":"journal-article","created":{"date-parts":[[2024,5,15]],"date-time":"2024-05-15T12:47:42Z","timestamp":1715777262000},"update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":36,"title":["Synergistic induction of blood\u2013brain barrier properties"],"prefix":"10.1073","volume":"121","author":[{"given":"Gerg\u0151","family":"Porkol\u00e1b","sequence":"first","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"},{"name":"Doctoral School of Biology, University of Szeged, Szeged H-6720, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4847-9618","authenticated-orcid":false,"given":"M\u00e1ria","family":"M\u00e9sz\u00e1ros","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6210-1689","authenticated-orcid":false,"given":"Anik\u00f3","family":"Szecsk\u00f3","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"},{"name":"Doctoral School of Biology, University of Szeged, Szeged H-6720, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4694-3891","authenticated-orcid":false,"given":"Judit P.","family":"Vigh","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"},{"name":"Doctoral School of Biology, University of Szeged, Szeged H-6720, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fruzsina R.","family":"Walter","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3024-1254","authenticated-orcid":false,"given":"Ricardo","family":"Figueiredo","sequence":"additional","affiliation":[{"name":"GenXPro GmbH, Frankfurt am Main 60438, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ildik\u00f3","family":"K\u00e1lomista","sequence":"additional","affiliation":[{"name":"In Vitro Metabolism Laboratory, Gedeon Richter, Budapest H-1103, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2435-3444","authenticated-orcid":false,"given":"Zs\u00f3fia","family":"Hoyk","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gaszton","family":"Vizsnyiczai","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-4258-8636","authenticated-orcid":false,"given":"Ilona","family":"Gr\u00f3f","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8379-404X","authenticated-orcid":false,"given":"Jeng-Shiung","family":"Jan","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering, National Cheng Kung University, Tainan 70101, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0481-5026","authenticated-orcid":false,"given":"Fabien","family":"Gosselet","sequence":"additional","affiliation":[{"name":"Laboratoire de la Barri\u00e9re H\u00e9mato-Enc\u00e9phalique, Universit\u00e9 d\u2019Artois, Lens 62307, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2949-3510","authenticated-orcid":false,"given":"Melinda K.","family":"Pirity","sequence":"additional","affiliation":[{"name":"Institute of Genetics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Monika","family":"Vastag","sequence":"additional","affiliation":[{"name":"In Vitro Metabolism Laboratory, Gedeon Richter, Budapest H-1103, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Natalie","family":"Hudson","sequence":"additional","affiliation":[{"name":"Smurfit Institute of Genetics, Trinity College Dublin, Dublin D02 VF25, Ireland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthew","family":"Campbell","sequence":"additional","affiliation":[{"name":"Smurfit Institute of Genetics, Trinity College Dublin, Dublin D02 VF25, Ireland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8864-1184","authenticated-orcid":false,"given":"Szilvia","family":"Veszelka","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6084-6524","authenticated-orcid":false,"given":"M\u00e1ria A.","family":"Deli","sequence":"additional","affiliation":[{"name":"Institute of Biophysics, Biological Research Centre, Hungarian Research Network, Szeged H-6726, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"341","published-online":{"date-parts":[[2024,5,15]]},"reference":[{"key":"e_1_3_4_1_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2008.01.003"},{"key":"e_1_3_4_2_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.nbd.2009.07.030"},{"key":"e_1_3_4_3_2","doi-asserted-by":"publisher","DOI":"10.1038\/jcbfm.2012.126"},{"key":"e_1_3_4_4_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41593-019-0497-x"},{"key":"e_1_3_4_5_2","doi-asserted-by":"publisher","DOI":"10.1152\/physrev.00050.2017"},{"key":"e_1_3_4_6_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41576-022-00466-9"},{"key":"e_1_3_4_7_2","doi-asserted-by":"publisher","DOI":"10.1177\/0271678X16630991"},{"key":"e_1_3_4_8_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1471-4159.2011.07208.x"},{"key":"e_1_3_4_9_2","doi-asserted-by":"publisher","DOI":"10.1021\/mp200129p"},{"key":"e_1_3_4_10_2","doi-asserted-by":"publisher","DOI":"10.1021\/acs.molpharmaceut.0c00178"},{"key":"e_1_3_4_11_2","doi-asserted-by":"publisher","DOI":"10.1038\/nbt.2247"},{"key":"e_1_3_4_12_2","doi-asserted-by":"publisher","DOI":"10.1038\/srep04160"},{"key":"e_1_3_4_13_2","doi-asserted-by":"publisher","DOI":"10.1111\/jnc.13923"},{"key":"e_1_3_4_14_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.stemcr.2017.02.021"},{"key":"e_1_3_4_15_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.stemcr.2019.08.002"},{"key":"e_1_3_4_16_2","doi-asserted-by":"publisher","DOI":"10.1126\/sciadv.aau7375"},{"key":"e_1_3_4_17_2","doi-asserted-by":"publisher","DOI":"10.1126\/sciadv.1701679"},{"key":"e_1_3_4_18_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.stem.2019.05.011"},{"key":"e_1_3_4_19_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-019-10588-0"},{"key":"e_1_3_4_20_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.biomaterials.2018.10.023"},{"key":"e_1_3_4_21_2","doi-asserted-by":"publisher","DOI":"10.1096\/fj.202001507RR"},{"key":"e_1_3_4_22_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0099733"},{"key":"e_1_3_4_23_2","doi-asserted-by":"publisher","DOI":"10.1177\/0271678X18820584"},{"key":"e_1_3_4_24_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-020-00198-0"},{"key":"e_1_3_4_25_2","doi-asserted-by":"publisher","DOI":"10.3390\/pharmaceutics13060892"},{"key":"e_1_3_4_26_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-020-00222-3"},{"key":"e_1_3_4_27_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2016950118"},{"key":"e_1_3_4_28_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature09522"},{"key":"e_1_3_4_29_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature09513"},{"key":"e_1_3_4_30_2","doi-asserted-by":"publisher","DOI":"10.1038\/nn.4288"},{"key":"e_1_3_4_31_2","doi-asserted-by":"publisher","DOI":"10.1038\/325253a0"},{"key":"e_1_3_4_32_2","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.23-27-09254.2003"},{"key":"e_1_3_4_33_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrn1824"},{"key":"e_1_3_4_34_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1164594"},{"key":"e_1_3_4_35_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0805165106"},{"key":"e_1_3_4_36_2","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.06489"},{"key":"e_1_3_4_37_2","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.200806024"},{"key":"e_1_3_4_38_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1017192108"},{"key":"e_1_3_4_39_2","doi-asserted-by":"publisher","DOI":"10.1172\/JCI76431"},{"key":"e_1_3_4_40_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0070233"},{"key":"e_1_3_4_41_2","doi-asserted-by":"publisher","DOI":"10.1186\/2045-8118-10-16"},{"key":"e_1_3_4_42_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-019-56075-w"},{"key":"e_1_3_4_43_2","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.70992"},{"key":"e_1_3_4_44_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1911532117"},{"key":"e_1_3_4_45_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-020-00197-1"},{"key":"e_1_3_4_46_2","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.115.6.1725"},{"key":"e_1_3_4_47_2","doi-asserted-by":"publisher","DOI":"10.1242\/jcs.107.5.1347"},{"key":"e_1_3_4_48_2","doi-asserted-by":"publisher","DOI":"10.1113\/expphysiol.1995.sp003877"},{"key":"e_1_3_4_49_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature25739"},{"key":"e_1_3_4_50_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.pneurobio.2020.101937"},{"key":"e_1_3_4_51_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuint.2008.12.002"},{"key":"e_1_3_4_52_2","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.36187"},{"key":"e_1_3_4_53_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-021-04369-3"},{"key":"e_1_3_4_54_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-04521-7"},{"key":"e_1_3_4_55_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-020-69096-7"},{"key":"e_1_3_4_56_2","doi-asserted-by":"publisher","DOI":"10.1093\/neuonc\/noab022"},{"key":"e_1_3_4_57_2","doi-asserted-by":"publisher","DOI":"10.1021\/jm901036q"},{"key":"e_1_3_4_58_2","doi-asserted-by":"publisher","DOI":"10.1208\/s12248-021-00609-6"},{"key":"e_1_3_4_59_2","doi-asserted-by":"publisher","DOI":"10.3109\/1061186X.2014.888070"},{"key":"e_1_3_4_60_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.ddtec.2016.09.003"},{"key":"e_1_3_4_61_2","doi-asserted-by":"publisher","DOI":"10.1021\/acsami.1c08454"},{"key":"e_1_3_4_62_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-020-00191-7"},{"key":"e_1_3_4_63_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-021-00291-y"},{"key":"e_1_3_4_64_2","doi-asserted-by":"publisher","DOI":"10.3389\/fphys.2021.642812"},{"key":"e_1_3_4_65_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-020-60688-x"},{"key":"e_1_3_4_66_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-019-0123-z"},{"key":"e_1_3_4_67_2","doi-asserted-by":"publisher","DOI":"10.1177\/0271678X21992638"},{"key":"e_1_3_4_68_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12987-022-00377-1"},{"key":"e_1_3_4_69_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.biomaterials.2022.121983"},{"key":"e_1_3_4_70_2","unstructured":"G. Porkol\u00e1b Data from \u201cSynergistic induction of blood-brain barrier properties.\u201d Gene Expression Omnibus (GEO). https:\/\/www.ncbi.nlm.nih.gov\/geo\/query\/acc.cgi?acc=GSE224846. Deposited 8 February 2023."}],"container-title":["Proceedings of the National Academy of Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pnas.org\/doi\/pdf\/10.1073\/pnas.2316006121","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,5,15]],"date-time":"2024-05-15T12:48:24Z","timestamp":1715777304000},"score":1,"resource":{"primary":{"URL":"https:\/\/pnas.org\/doi\/10.1073\/pnas.2316006121"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,5,15]]},"references-count":70,"journal-issue":{"issue":"21","published-print":{"date-parts":[[2024,5,21]]}},"alternative-id":["10.1073\/pnas.2316006121"],"URL":"https:\/\/doi.org\/10.1073\/pnas.2316006121","relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/2023.02.09.527899","asserted-by":"object"}]},"ISSN":["0027-8424","1091-6490"],"issn-type":[{"value":"0027-8424","type":"print"},{"value":"1091-6490","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,5,15]]},"assertion":[{"value":"2023-09-14","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-04-05","order":1,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2024-05-15","order":2,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e2316006121"}}