Volume 114, Issue 5, Pages 947-955 (May 1998) Rotavirus infection of cultured intestinal epithelial cells induces secretion of CXC and CC chemokines  Antonella Casola*, Mary K. Estes‡, Sue E. Crawford‡, Pearay L. Ogra*, Peter B. Ernst*, Roberto P. Garofalo*, Sheila E. Crowe§  Gastroenterology  Volume 114, Issue 5, Pages 947-955 (May 1998) DOI: 10.1016/S0016-5085(98)70314-2 Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 1 Immunofluorescence staining of rotavirus-infected HT29 cells. Cells were infected with rotavirus at MOI of 1 for 24 hours and stained with a mouse antiserum raised against SA11 strain as the primary antibody and a fluorescein isothiocyanate–conjugated goat anti-mouse F(ab')2 antibody as the secondary antibody. (A) Mock-infected control cells. (B) Rotavirus-infected cells. Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 1 Immunofluorescence staining of rotavirus-infected HT29 cells. Cells were infected with rotavirus at MOI of 1 for 24 hours and stained with a mouse antiserum raised against SA11 strain as the primary antibody and a fluorescein isothiocyanate–conjugated goat anti-mouse F(ab')2 antibody as the secondary antibody. (A) Mock-infected control cells. (B) Rotavirus-infected cells. Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 2 Comparison of chemokine release by intestinal epithelial cells after cytokine stimulation and rotavirus infection. (A) Caco-2 and (B) HT29 cells were infected with rotavirus (MOI of 1) or treated with TNF-α (20 ng/mL, ▩) or IL-1β (10 ng/mL, ▨ ). 2, Control; ■, SA11. Cell supernatants were harvested 24 hours later, and chemokine levels were assayed by ELISA. Values are expressed as fold induction compared with basal levels of mock-infected control cells or cells treated with vehicle only (n = 3). Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 3 Dose-dependent release of chemokines after rotavirus infection. HT29 cells were infected with rotavirus at different MOI, ranging from 0.1 to 10. Cell supernatants were harvested at 24 hours after infection, and chemokine levels were assayed by ELISA. Values are expressed as fold induction compared with basal levels of mock-infected control cells (n = 3). Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 4 Kinetics of chemokine secretion after rotavirus infection. Confluent monolayers of HT29 cells were infected with rotavirus at a MOI of 1. At the indicated times, cell supernatants were harvested and assayed for (A) IL-8, (B) GRO-α, and (C) RANTES by ELISA. 2, Mock-infected control cells; ■, rotavirus-infected cells. Results are expressed as means ± SD (n = 3–5). *P < 0.05, **P < 0.01 compared with mock infected. Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 5 Kinetics of change in chemokine mRNA abundance in response to rotavirus infection by reverse-transcription PCR. Total RNA was extracted from HT29 cells infected with rotavirus (MOI of 1) for the indicated times. β-Actin is included as an internal control. Lane 1, molecular weight markers; lane 2, positive control (cDNA); lane 3, mock-infected HT29 cells; lane 4, 6-hour infection; lane 5, 12-hour infection; lane 6, 24-hour infection; lane 7, 48-hour infection. Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions

Fig. 6 Effect of viral inactivation and VLPs on IL-8 production. HT29 cells were infected with live rotavirus (SA11) or psoralen/UV-treated virus (Psor./UV), both at an MOI of 1, or treated with VLPs. Cell supernatants were harvested 24 hours after infection, and IL-8 secretion was assayed by ELISA. Results are expressed as means ± SD (n = 3). *P < 0.01 compared with mock-infected control cells. Gastroenterology 1998 114, 947-955DOI: (10.1016/S0016-5085(98)70314-2) Copyright © 1998 American Gastroenterological Association Terms and Conditions