Skip to main content

Characterization of Flucloxacillin-specific CD8+ T-cells in a mouse model

Drug-induced liver injury (DILI) involving the adaptive immune system is a serious complication in both patient treatment and drug development. Flucloxacillin, a -lactam antibiotic effective against -lactamase-producing bacteria, is a common cause of immunological DILI through cholestasis. We have recently isolated CD8+ flucloxacillin-specific T-cells from patients with DILI, which indicates that T-cells participate in the pathogenesis of the disease. There are currently no animal models of DILI where the adaptive immune system has shown to damage liver, and therefore, it is difficult to explore the mechanistic basis of the tissue injury. Thus, the aim of this study was to use C57/Bl6 CD4 deficient mice with a mutation in the gene encoding for MHC class II molecules to explore the immunogenicity of flucloxacillin and whether flucloxacillin-responsive CD8+ T-cells damage hepatocytes. In initial experiments sensitization was achieved through epicutaneous application (1g/ml; in 70% DMSO; 50L). Proliferation, IFN- and granzyme-B secretion from draining lymph node cells was measured ex vivo following treatment with flucloxacillin. Flucloxacillin-specific CD8+ T-cell mediated killing of hepatocytes was measured using the ApoTox-Glo kit (Promega, UK). In separate experiments, flucloxacillin-specific T-cells were forced to migrate to the mesenteric lymph nodes using retinoic acid, prior to administration of oral flucloxacillin for 10 days, followed by analysis of plasma biomarkers of liver injury. CD8+ T-cells from draining lymph nodes of flucloxacillin-treated mice proliferated in a concentration-dependent manner following ex vivo secondary stimulation. The proliferative response was associated with IFN- and granzyme-B secretion. In contrast, proliferative responses and cytokine secretion was not detected with cells from vehicle control mice. Flucloxacillin-specific hepatocyte toxicity and apoptosis was observed when CD8+ T-cells were cultured with dendritic cells and flucloxacillin for 24h, washed and transferred to the hepatocyte cultures. Flucloxacillin sensitization followed by 10 day oral exposure resulted in marked gall bladder swelling and mild elevations in ALT. Other plasma biomarkers of liver damage were negative. In conclusion, these data show successful sensitization of mice against flucloxacillin. Thus, the C57/Bl6 CD4 deficient mouse represents a promising model to study the role of the adaptive immune system in DILI.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Dean Naisbitt.

Rights and permissions

Open Access  This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made.

The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.

To view a copy of this licence, visit https://creativecommons.org/licenses/by/4.0/.

The Creative Commons Public Domain Dedication waiver (https://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Nattrass, R., Faulkner, L., Jenkins, R. et al. Characterization of Flucloxacillin-specific CD8+ T-cells in a mouse model. Clin Transl Allergy 4, P41 (2014). https://doi.org/10.1186/2045-7022-4-S3-P41

Download citation

Keywords

  • Retinoic Acid
  • Cholestasis
  • Lactam
  • Adaptive Immune System
  • Drain Lymph Node