Recently we showed that donor-specific tolerance to MHC-matched islet allografts in diabetic NOD mice could be induced by simultaneous islet and bone marrow transplantation. Mononuclear cell infiltration surrounding the islets was also found in tolerated grafts. In this study, we compared gene expression in the tolerated and rejected islet grafts by using Affymetrix Murine U74A oligonucleotide arrays. To confirm the results of microarray analysis, we performed real-time PCR and RNase protection assay on selected genes. Of over 12,000 genes studied, 57 genes were expressed at consistently higher levels in tolerated islet grafts, and 524 genes in rejected islet grafts. Genes from a variety of functional clusters were found to be different between rejected and tolerated grafts. In the rejected islet grafts, a number of T-cell surface markers and of cytotoxicity-related genes were highly expressed. Also in the rejected grafts, a number of cytokines and chemokines and their receptors were highly expressed. The differential expression of selected genes found by microarray analysis was also confirmed by real-time PCR and RNase protection assay. Our results indicated that gene microarray analysis can help us to detect gene expression differences representative of the biologic mechanisms of tolerance and rejection.
AbdiR.; SmithR. N.; MakhloufL.; NajafianN.; LusterA. D.; AuchinclossH.Jr.; SayeghM. H.The role of CC chemokine receptor 5 (CCR5) in islet allograft rejection.Diabetes51: 2489–2495; 2002.
2.
AkalinE.; HendrixR. C.; PolavarapuR. G.; PearsonT. C.; NeylanJ. F.; LarsenC. P.; LakkisF. G.Gene expression analysis in human renal allograft biopsy samples using high-density oligoarray technology.Transplantation72: 948–953; 2001.
3.
ArmstrongH. E.; BoltonE. M.; McMillanI.; SpencerS. C.; BradleyJ. A.Prolonged survival of actively enhanced rat renal allografts despite accelerated cellular infiltration and rapid induction of both class I and class II MHC antigens.J. Exp. Med.165: 891–907; 1987.
4.
BrownP. O.; BotsteinD.Exploring the new world of the genome with DNA microarrays.Nat. Genet.21: 33–37; 1999.
ColvinB. L.; ThomsonA. W.Chemokines, their receptors, and transplant outcome.Transplantation74: 149–155; 2002.
7.
DallmanM. J.; WoodK. J.; MorrisP. J.Specific cytotoxic T cells are found in the nonrejected kidneys of blood-transfused rats.J. Exp. Med.165: 566–571; 1987.
8.
DiamondA. S.; GillR. G.An essential contribution by IFN-gamma to CD8+ T cell-mediated rejection of pancreatic islet allografts.J. Immunol.165: 247–255; 2000.
GaoW.; FaiaK. L.; CsizmadiaV.; SmileyS. T.; SolerD.; KingJ. A.; DanoffT. M.; HancockW. W.Beneficial effects of targeting CCR5 in allograft recipients.Transplantation72: 1199–1205; 2001.
11.
GrandalianoG.; GesualdoL.; RanieriE.; MonnoR.; StalloneG.; SchenaF. P.Monocyte chemotactic peptide-1 expression and monocyte infiltration in acute renal transplant rejection.Transplantation63: 414–420; 1997.
12.
HanD.; XuX.; PastoriR. L.; RicordiC.; KenyonN. S.Elevation of cytotoxic lymphocyte gene expression is predictive of islet allograft rejection in nonhuman primates.Diabetes51: 562–566; 2002.
13.
HancockW. W.; LuB.; GaoW.; CsizmadiaV.; FaiaK.; KingJ. A.; SmileyS. T.; LingM.; GerardN. P.; GerardC.Requirement of the chemokine receptor CXCR3 for acute allograft rejection.J. Exp. Med.192: 1515–1520; 2000.
14.
HaoL. M.; WangY.; GillR. G.; La RosaF. G.; TalmageD. W.; LaffertyK. J.Role of lymphokine in islet allograft rejection.Transplantation49: 609–614; 1990.
15.
HerbelinA.; GombertJ. M.; LepaultF.; BachJ. F.; ChatenoudL.Mature mainstream TCR alpha beta+CD4+ thymocytes expressing L-selectin mediate “active tolerance” in the nonobese diabetic mouse.J. Immunol.161: 2620–2628; 1998.
16.
HutchinsonI. V.The role of transforming growth factor-beta in transplant rejection.Transplant. Proc.31: 9S–13S; 1999.
17.
KobayashiH.; NovickA. C.; TomaH.; FairchildR. L.Chronic antagonism of Mig inhibits cellular infiltration and promotes survival of class II MHC disparate skin allografts.Transplantation74: 387–395; 2002.
18.
KogaS.; AuerbachM. B.; EngemanT. M.; NovickA. C.; TomaH.; FairchildR. L.T cell infiltration into class II MHC-disparate allografts and acute rejection is dependent on the IFN-gamma-induced chemokine Mig.J. Immunol.163: 4878–4885; 1999.
19.
LepaultF.; GagneraultM. C.Characterization of peripheral regulatory CD4+ T cells that prevent diabetes onset in nonobese diabetic mice.J. Immunol.164: 240–247; 2000.
20.
McHughR. S.; WhittersM. J.; PiccirilloC. A.; YoungD. A.; ShevachE. M.; CollinsM.; ByrneM. C.CD4(+)CD25(+) immunoregulatory T cells: Gene expression analysis reveals a functional role for the glucocorticoid-induced TNF receptor.Immunity16: 311–323; 2002.
21.
O'ConnellP. J.; Pacheco-SilvaA.; NickersonP. W.; MuggiaR. A.; BastosM.; KelleyV. R.; StromT. B.Unmodified pancreatic islet allograft rejection results in the preferential expression of certain T cell activation transcripts.J. Immunol.150: 1093–1104; 1993.
22.
OzasaT.; NewtonM. R.; DallmanM. J.; ShimizuS.; GrayD. W.; MorrisP. J.Cytokine gene expression in pancreatic islet grafts in the rat.Transplantation64: 1152–1159; 1997.
23.
PattisonJ.; NelsonP. J.; HuieP.; vonL. I.; FarshidG.; SibleyR. K.; KrenskyA. M.RANTES chemokine expression in cell-mediated transplant rejection of the kidney.Lancet343: 209–21l; 1994.
24.
PaulukatJ.; BosmannM.; NoldM.; GarkischS.; KampferH.; FrankS.; RaedleJ.; ZeuzemS.; PfeilschifterJ.; MuhlH.Expression and release of IL-18 binding protein in response to IFN-gamma.J. Immunol.167: 7038–7043; 2001.
25.
SaiuraA.; MatakiC.; MurakamiT.; UmetaniM.; WadaY.; KohroT.; AburataniH.; HariharaY.; HamakuboT.; YamaguchiT.; HasegawaG.; NaitoM.; MakuuchiM.; KodamaT.A comparison of gene expression in murine cardiac allografts and isografts by means DNA microarray analysis.Transplantation72: 320–329; 2001.
26.
SaiuraA.; KohroT.; YamamotoT.; IzumiA.; WadaY.; AburataniH.; SugawaraY.; HamakuboT.; TaniguchiT.; NaitoM.; KodamaT.; MakuuchiM.Detection of an up-regulation of a group of chemokine genes in murine cardiac allograft in the absence of interferongamma by means of DNA microarray.Transplantation73: 1480–1486; 2002.
27.
SaiuraA.; SugawaraY.; HariharaY.; SataM.; HamakuboT.; KodamaT.; MakuuchiM.Gene expression profile during acute rejection in rat-to-mouse concordant cardiac xenograft by means of DNA microarray.Transpl. Int.15: 535–540; 2002.
28.
SarwalM.; ChangS.; BarryC.; ChenX.; AlizadehA.; SalvatierraO.; BrownP.Genomic analysis of renal allograft dysfunction using cDNA microarrays.Transplant. Proc.33: 297–298; 2001.
29.
ShapiroA. M.; LakeyJ. R.; RyanE. A.; KorbuttG. S.; TothE.; WarnockG. L.; KnetemanN. M.; RajotteR. V.Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen.N. Engl. J. Med.343: 230–238; 2000.
30.
StegallM.; ParkW.; KimD.; KremersW.Gene expression during acute allograft rejection: Novel statistical analysis of microarray data.Am. J. Transplant.2: 913–925; 2002.
31.
StrehlauJ.; PavlakisM.; LipmanM.; ShapiroM.; VasconcellosL.; HarmonW.; StromT. B.Quantitative detection of immune activation transcripts as a diagnostic tool in kidney transplantation.Proc. Natl. Acad. Sci. USA94: 695–700; 1997.
WuT.; Levay-YoungB.; HeussN.; SozenH.; KirchhofN.; SutherlandD. E.; HeringB.; GuoZ.Inducing tolerance to MHC-matched allogeneic islet grafts in diabetic NOD mice by simultaneous islet and bone marrow transplantation under nonirradiative and nonmyeloablative conditioning therapy.Transplantation74: 22–27; 2002.
34.
XuG. P.; SharmaV. K.; LiB.; BologaR.; LiY.; MouradianJ.; WangJ.; SerurD.; RaoV.; StenzelK. H.Intragraft expression of IL-10 messenger RNA: A novel correlate of renal allograft rejection.Kidney Int.48: 1504–1507; 1995.
