Viral IL-10 gene therapy inhibits TNF-α and IL-1β, not IL-6, in the newborn endotoxemic mouse,☆☆

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Abstract

Introduction: Modulation of the inflammatory cascade within the liver of critically ill infants may improve the chance of survival. Using gene therapy, the authors hypothesized that augmented local production of the counter-regulatory cytokine viral interleukin-10 (IL-10) in vivo will modulate the critical cytokines in the inflammatory response. The purpose of the present study was to determine whether replication-defective adenovirus-mediated viral IL-10 (vIL-10) gene transfer and expression within the liver can achieve this goal in newborn mice. Materials and Methods: Four-week-old Balbc mice were administered (intraperitoneally) 1 × 109 plaque-forming units (pfu) per milliliter of an adenovirus vector (E1ab-deleted) that encodes the sv40 promoter and the BCRF1 cDNA, or of control vector dl434 that expresses no foreign gene. Forty-eight hours later the mice were challenged with 50 μg/kg of lipopolysaccharide; (LPS) they were killed 1, 2, 6, or 24 hours later (six at each time point). Southern blot analysis was performed on genomic DNA isolated from the liver, lung, and kidney to assess gene transfer of BCRF1. Homogenized liver protein was analyzed for tumor necrosis factor-α (TNF-α), IL-1β, IL-6, and recombinant vIL-10. Results: Southern blot analysis confirmed successful gene transfer to the liver but not to the lung, kidney, or dl434-transduced liver in mice that received adenovectors. Viral IL-10 levels within the liver ranged from 14 to 18 ng/mL. In controls, INF-α production was elevated at early time points, to 18,000 pg/mL, but decreased rapidly by 24 hours after LPS challenge. The TNF-α levels of animals treated with Ad5svBCRF1 were significantly lower than those of controls throughout the course of study (P < .0001). After the LPS challenge, hepatic IL-1β decreased, from a maximum of 800 pg/mL (2 hours) to 411 pg/mL (24 hours). Inhibition of IL-1β by vIL-10 occurred at 1 hour (P < .016) and 2 hours (P < .001) only. Hepatic production of IL-6 after LPS challenge ranged from 7 to 8,000 pg/mL in all groups and was not altered by vIL-10 gene therapy. Conclusion: In vivo administration of adenovectors encoding BCRF1 to newborn mice results in efficient hepatic transduction and expression of recombinant vIL-10. The Kupffer cell response to LPS is suppressed with respect to TNF-α and IL-1β, but not IL-16. In vivo modulation of hepatic cytokine responses is achievable using gene products that mimic cellular cytokines. This is an effective model for the selective evaluation of therapeutic gene products for gene therapy of sepsis.

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Presented in part at the 26th Annual Meeting of the American Pediatric Surgical Association, Boca Raton, Florida, May 1995.

☆☆

Supported by the Ethicon-Society of University Surgeons Research Award (K.E.D.) and National Institutes of Health Grant No. DK43111 (A.S.).

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