J Korean Med Sci.  2013 Dec;28(12):1729-1733. 10.3346/jkms.2013.28.12.1729.

Application of the Multiplex Cytokine Analysis to Monitor Xenogeneic Immune Responses to the Porcine Islet Graft in Non-Human Primate

• Kim YH
• Kim JS
• Yoon IH
• Shin JS
• Kim JM
• Kim SJ
• Park CG

• KMID: 1779412
• DOI: http://doi.org/10.3346/jkms.2013.28.12.1729

Abstract

Non-human primate studies must be conducted prior to the clinical trial of xenotransplantation. In order to develop clinically applicable immune-modulatory regimen through non-human primate studies, close monitoring of xenogeneic immune responses is required. We adopted multiplex cytokine analysis in assessment of the immune status during the course of pig-to-non-human primate islet transplantation. This study aimed to assess the feasibility of this multiplex cytokine assay in the development of immune-modulatory regimen. Using this assay, we were able to detect different cytokines with a minimal usage of blood samples, and this allowed us to detect various immunological situations in the recipients. Detection of TNF-alpha surge (347.8 pg/mL) guided us to block TNF-alpha in the early phase of transplantation. Supportive information for in vivo efficacy of cytokine neutralizing antibody could be speculated by in vitro neutralization assay (1,250 pg/mL --> 0 pg/mL). In addition, periodic monitoring of cytokines in peripheral blood allowed the detection of the infection episode prior to other routine assays. These benefits of multiplex cytokine assay may be generally applied to other pre-clinical research, which is a prerequisite for clinical trials.

Keyword

Immune Monitoring; Islet Transplantation; Translational Research; Xenotransplantation

MeSH Terms

Animals
Antibodies, Neutralizing/immunology
Blood Cell Count
Cytokines/*blood/immunology
Immunoassay/*methods
Interleukin-6/blood
Islets of Langerhans Transplantation/*immunology
Macaca mulatta
Swine
Transplantation, Heterologous
Tumor Necrosis Factor-alpha/blood/immunology
Antibodies, Neutralizing
Cytokines
Interleukin-6
Tumor Necrosis Factor-alpha

Figure

• 

  Fig. 1 Detection of TNF-α surge in an episode of acute graft rejection. A recipient monkey of porcine islets was monitored for their diabetic status with blood glucose level, porcine c-peptide and exogenously injected insulin (A) and blood cytokine levels (B).

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  Fig. 2 Ex vivo test for neutralizing efficacy of antibody drug, Humira. Multiplex cytokine assay was conducted with (left panel) or without (right panel) pre-incubation with 20 µg/mL of anti-TNF-α neutralizing antibody, Humira. Beads for each cytokines are discerned with discrete APC-fluorescence intensities. PE-intensities of beads captured with detecting antibodies are depicted.

• 

  Fig. 3 Prediction of peri-transplant immune response before the re-transplantation. (A) Cytokine levels detected in the blood of a recipient monkey before the re-transplantation of porcine islets are depicted. (B) Blood glucose levels and amounts of exogenously injected insulin are depicted.

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  Fig. 4 IL-6 surge in an episode of septic shock. Periodic monitoring of blood cytokine levels was conducted in a recipient monkey, and IL-6 levels are depicted along with the timeframes in which other signs were shown.

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