{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,28]],"date-time":"2026-04-28T20:43:30Z","timestamp":1777409010356,"version":"3.51.4"},"reference-count":26,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2026,4,8]],"date-time":"2026-04-08T00:00:00Z","timestamp":1775606400000},"content-version":"vor","delay-in-days":7,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,4,1]],"date-time":"2026-04-01T00:00:00Z","timestamp":1775001600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["AI175479"],"award-info":[{"award-number":["AI175479"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["DK065988"],"award-info":[{"award-number":["DK065988"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100031234","name":"Michigan Medicine, University of Michigan","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100031234","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000897","name":"Cystic Fibrosis Foundation","doi-asserted-by":"publisher","award":["ESTHER24R0"],"award-info":[{"award-number":["ESTHER24R0"]}],"id":[{"id":"10.13039\/100000897","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Current Protocols"],"published-print":{"date-parts":[[2026,4]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Robust inflammatory responses to viral infection are mediated by immune cell populations, including monocytes and dendritic cells. However, severe acute respiratory syndrome coronavirus 2 (SARS\u2010CoV\u20102) does not replicate efficiently in these cell types and instead preferentially infects epithelial cell subsets in the airway. Because of this, stimulation of inflammatory cytokine responses from immune cell populations during SARS\u2010CoV\u20102 infection depends not only on interactions with viral particles but also interactions with infected epithelial cells. In this article, we describe two co\u2010culture systems to study inflammatory cytokine responses generated by epithelial\u2010immune interaction during SARS\u2010CoV\u20102 infection in vitro. Basic Protocol 1 describes how to set up a partially primary co\u2010culture system consisting of SARS\u2010CoV\u20102\u2010infected Vero\u2010E6 cells and primary human peripheral blood mononuclear cells (PBMCs) to observe release of the inflammasome\u2010regulated cytokine interleukin\u20101\u03b2 (IL\u20101\u03b2). Basic Protocol 2 details a primary, human co\u2010culture system that consists of SARS\u2010CoV\u20102\u2010infected primary human airway epithelia (HAE) grown at an air\u2013liquid interface (ALI) and primary human PBMCs, and how to observe IL\u20101\u03b2 and IL\u20106 crosstalk between these cell populations during infection. In these Basic Protocols, we include a description of the use of inhibitors in these systems to perturb cytokine responses. We also provide Support Protocols for the culture of HAE and Vero\u2010E6 and for the isolation, storage, and preparation of PBMCs prior to use in these systems. \u00a9 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC.<\/jats:p>\n                  <jats:p>\n                    <jats:bold>Basic Protocol 1<\/jats:bold>\n                    : SARS\u2010CoV\u20102 infection in a Vero\u2010E6+PBMC co\u2010culture system\n                  <\/jats:p>\n                  <jats:p>\n                    <jats:bold>Support Protocol 1<\/jats:bold>\n                    : Vero\u2010E6 culture and passaging\n                  <\/jats:p>\n                  <jats:p>\n                    <jats:bold>Support Protocol 2<\/jats:bold>\n                    : Isolation and cryopreservation of PBMCs for use in co\u2010culture\n                  <\/jats:p>\n                  <jats:p>\n                    <jats:bold>Basic Protocol 2<\/jats:bold>\n                    : SARS\u2010CoV\u20102 infection in a primary HAE+PBMC co\u2010culture system\n                  <\/jats:p>\n                  <jats:p>\n                    <jats:bold>Support Protocol 3<\/jats:bold>\n                    : Establishment and maturation of HAE grown at an ALI\n                  <\/jats:p>","DOI":"10.1002\/cpz1.70365","type":"journal-article","created":{"date-parts":[[2026,4,8]],"date-time":"2026-04-08T14:15:08Z","timestamp":1775657708000},"source":"Crossref","is-referenced-by-count":0,"title":["Co\u2010Culture Systems to Study Epithelial\u2010Immune Interactions During SARS\u2010CoV\u20102 Infection"],"prefix":"10.1002","volume":"6","author":[{"given":"Scott H.","family":"Randell","sequence":"first","affiliation":[{"name":"Marsico Lung Institute University of North Carolina at Chapel Hill, Chapel Hill  North Carolina"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9552-5325","authenticated-orcid":false,"given":"Katherine C.","family":"Barnett","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology University of Michigan Medical School  Ann Arbor Michigan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2026,4,8]]},"reference":[{"key":"e_1_2_11_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.chom.2022.12.005"},{"key":"e_1_2_11_3_1","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jiaf030"},{"key":"e_1_2_11_4_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41587\u2010020\u20100602\u20104"},{"key":"e_1_2_11_5_1","doi-asserted-by":"publisher","DOI":"10.1007\/978\u20101\u201062703\u2010125\u20107_8"},{"key":"e_1_2_11_6_1","doi-asserted-by":"publisher","DOI":"10.1002\/cpz1.453"},{"key":"e_1_2_11_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2020.02.052"},{"key":"e_1_2_11_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2020.05.042"},{"key":"e_1_2_11_9_1","doi-asserted-by":"publisher","DOI":"10.1152\/ajplung.00377.2003"},{"key":"e_1_2_11_10_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41586\u2010020\u20102180\u20105"},{"key":"e_1_2_11_11_1","doi-asserted-by":"crossref","unstructured":"Lan K. Verma S. C. Murakami M. Bajaj B. &Robertson E. S.(2007).Isolation of human peripheral blood mononuclear cells (PBMCs). Current Protocol Microbiology APPENDIX 4 APPENDIX 4C.https:\/\/doi.org\/10.1002\/9780471729259.mca04cs6","DOI":"10.1002\/9780471729259.mca04cs6"},{"key":"e_1_2_11_12_1","doi-asserted-by":"publisher","DOI":"10.1002\/cptx.31"},{"key":"e_1_2_11_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.addr.2023.114866"},{"key":"e_1_2_11_14_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.autrev.2020.102537"},{"key":"e_1_2_11_15_1","doi-asserted-by":"publisher","DOI":"10.1002\/cpmc.105"},{"key":"e_1_2_11_16_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.abm8108"},{"key":"e_1_2_11_17_1","doi-asserted-by":"publisher","DOI":"10.1111\/eci.13429"},{"key":"e_1_2_11_18_1","doi-asserted-by":"publisher","DOI":"10.2174\/1566524021666210408103921"},{"key":"e_1_2_11_19_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467\u2010022\u201030893\u20105"},{"key":"e_1_2_11_20_1","doi-asserted-by":"publisher","DOI":"10.1007\/978\u20101\u201061779\u2010120\u20108_18"},{"key":"e_1_2_11_21_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.abb7314"},{"key":"e_1_2_11_22_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41572\u2010025\u201000662\u2010x"},{"key":"e_1_2_11_23_1","doi-asserted-by":"publisher","DOI":"10.3201\/eid2705.210023"},{"key":"e_1_2_11_24_1","doi-asserted-by":"publisher","DOI":"10.1124\/jpet.106.111344"},{"key":"e_1_2_11_25_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.sbi.2023.102619"},{"key":"e_1_2_11_26_1","doi-asserted-by":"publisher","DOI":"10.1164\/rccm.202001\u20100179LE"},{"key":"e_1_2_11_27_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41586\u2010020\u20102012\u20107"}],"container-title":["Current Protocols"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/currentprotocols.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/cpz1.70365","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/currentprotocols.onlinelibrary.wiley.com\/doi\/full-xml\/10.1002\/cpz1.70365","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/currentprotocols.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/cpz1.70365","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,28]],"date-time":"2026-04-28T10:41:57Z","timestamp":1777372917000},"score":1,"resource":{"primary":{"URL":"https:\/\/currentprotocols.onlinelibrary.wiley.com\/doi\/10.1002\/cpz1.70365"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4]]},"references-count":26,"journal-issue":{"issue":"4","published-print":{"date-parts":[[2026,4]]}},"alternative-id":["10.1002\/cpz1.70365"],"URL":"https:\/\/doi.org\/10.1002\/cpz1.70365","archive":["Portico"],"relation":{},"ISSN":["2691-1299","2691-1299"],"issn-type":[{"value":"2691-1299","type":"print"},{"value":"2691-1299","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4]]},"article-number":"e70365"}}