A Reg Family Protein Is Overexpressed in Islets From a Patient With New-Onset Type 1 Diabetes and Acts as T-Cell Autoantigen in NOD Mice

  1. Werner Gurr1,
  2. Reza Yavari1,
  3. Li Wen1,
  4. Margaret Shaw2,
  5. Conchi Mora3,
  6. Laurence Christa4 and
  7. Robert S. Sherwin1
  1. 1Department of Internal Medicine, Yale University, New Haven, Connecticut
  2. 2Center for Veterinary Sciences, University of Cambridge, Cambridge, U.K.
  3. 3Laboratory for Research in Diabetes, University of Barcelona, Barcelona, Spain
  4. 4Molecular Virology and Liver Carcinogenesis, Hospital Necker for Sick Children, Paris, France

    Abstract

    Genes overexpressed in pancreatic islets of patients with new-onset type 1 diabetes are potential candidates for novel disease-related autoantigens. RT-PCR-based subtractive hybridization was used on islets from a patient who died at the onset of type 1 diabetes, and it identified a type 1 diabetes-related cDNA encoding hepatocarcinoma-intestine-pancreas/pancreatic-associated protein (HIP/PAP). This protein belongs to the family of Reg proteins implicated in islet regeneration; its gene contains a putative interleukin-6 (IL-6) response element. Islets from healthy cadaveric human donors released HIP/PAP protein into the culture medium, and this release was enhanced by the addition of IL-6. The expression pattern of mouse homologues of HIP/PAP was determined in pancreata of prediabetic and diabetic NOD mice. Both groups showed positive immunostaining for HIP/PAP in islets and ductal epithelium. To test whether HIP/PAP is a target of islet-directed autoimmunity, we measured splenic T-cell responses against HIP/PAP in NOD mice. Spontaneous proliferation was detected after 4 weeks. Lymphocytes from islet infiltrates and pancreatic lymph nodes from 7- to 10-week-old NOD mice were used to establish an HIP/PAP-specific I-Ag7-restricted T-cell line, termed WY1, that also responded to mouse islets. WY1 cells homed to islets of NOD-SCID mice and adoptively transferred disease when coinjected with purified CD8+ cells from diabetic NOD mice. Our conclusion was that differential cloning of Reg from islets of a type 1 diabetic patient and the response of Reg to the cytokine IL-6 suggests that HIP/PAP becomes overexpressed in human diabetic islets because of the local inflammatory response. HIP/PAP acts as a T-cell autoantigen in NOD mice. Therefore, autoimmunity to HIP/PAP might create a vicious cycle, accelerating the immune process leading to diabetes.

    Footnotes

    • Address correspondence and reprint requests to Robert S. Sherwin, Department of Internal Medicine, Yale University, New Haven, CT 06520-8020. E-mail: robert.sherwin{at}yale.edu.

      Received for publication 14 March 2001 and accepted in revised form 25 October 2001.

      AEC, 3-amino-9-ethylcarbazol; Bruff’s/FCS, Bruff’s medium supplemented with 3% FCS; CRD, carbohydrate recognition domain; FACS, fluorescence-activated cell sorter; FITC, fluorescein isothiocyanate; HIP-CRD, CRD of HIP/PAP; HIP/PAP, hepatocarcinoma-intestine-pancreas/pancreatic-associated protein; HRP, horseradish peroxidase; IL-6, interleukin-6; PBS-T, PBS with 0.05% Tween 20; PE, phycoerythrin; SSC, sodium chloride-sodium citrate.

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