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• W1996709249 abstract "The role of NK cells following solid organ transplantation remains unclear. We examined NK cells in acute allograft rejection using a high responder model (DA → Lewis) of rat orthotopic liver transplantation. Recipient‐derived NK cells infiltrated liver allografts early after transplantation. Since chemokines are important in the trafficking of cells to areas of inflammation, we determined the intragraft expression of chemokines known to attract NK cells. CCL3 was significantly increased in allografts at 6 h post‐transplant as compared to syngeneic grafts whereas CCL2 and CXCL10 were elevated in both syngeneic and allogeneic grafts. CXCL10 and CX3CL1 were significantly upregulated in allografts by day 3 post‐transplant as compared to syngeneic grafts suggesting a role for these chemokines in the recruitment of effector cells to allografts. Graft‐infiltrating NK cells were shown to be a major source of IFN‐γ, and IFN‐γ levels in the serum were markedly increased, specifically in allograft recipients, by day 3 post‐transplant. Accordingly, in the absence of NK cells the levels of IFN‐γ were significantly decreased. Furthermore, graft survival was significantly prolonged. These data suggest that IFN‐γ‐producing NK cells are an important link between the innate and adaptive immune responses early after transplantation. The role of NK cells following solid organ transplantation remains unclear. We examined NK cells in acute allograft rejection using a high responder model (DA → Lewis) of rat orthotopic liver transplantation. Recipient‐derived NK cells infiltrated liver allografts early after transplantation. Since chemokines are important in the trafficking of cells to areas of inflammation, we determined the intragraft expression of chemokines known to attract NK cells. CCL3 was significantly increased in allografts at 6 h post‐transplant as compared to syngeneic grafts whereas CCL2 and CXCL10 were elevated in both syngeneic and allogeneic grafts. CXCL10 and CX3CL1 were significantly upregulated in allografts by day 3 post‐transplant as compared to syngeneic grafts suggesting a role for these chemokines in the recruitment of effector cells to allografts. Graft‐infiltrating NK cells were shown to be a major source of IFN‐γ, and IFN‐γ levels in the serum were markedly increased, specifically in allograft recipients, by day 3 post‐transplant. Accordingly, in the absence of NK cells the levels of IFN‐γ were significantly decreased. Furthermore, graft survival was significantly prolonged. These data suggest that IFN‐γ‐producing NK cells are an important link between the innate and adaptive immune responses early after transplantation. NK cells, large granular lymphocytes with the ability to lyse selected target cells without prior sensitization, play an important role in the host immune response against neoplastic cells and virally infected cells (1Koo GC Peppard JR Mark WH Natural killer cells generated from bone marrow culture.J Immunol. 1984; 132: 2300-2304Crossref PubMed Google Scholar, 2Trinchieri G Biology of natural killer cells.Adv Immunol. 1989; 47: 187-376Crossref PubMed Scopus (2717) Google Scholar, 3Biron CA Byron KS Sullivan JL Severe herpesvirus infections in an adolescent without natural killer cells.N Engl J Med. 1989; 320: 1731-1735Crossref PubMed Scopus (1051) Google Scholar). NK cells have also been shown to inhibit the engraftment of allogeneic bone marrow (4Rolstad B Benestad HB The “natural resistance” to bone marrow allografts in normal and athymic nude rats. Rapid cytotoxic reactions both in vivo and in vitro.Eur J Immunol. 1984; 14: 793-799Crossref PubMed Scopus (23) Google Scholar, 5Murphy WJ Kumar V Bennett M Acute rejection of murine bone marrow allografts by natural killer cells and T cells. Differences in kinetics and target antigens recognized.J Exp Med. 1987; 166: 1499-1509Crossref PubMed Scopus (161) Google Scholar, 6Murphy WJ Kumar V Bennett M Rejection of bone marrow allografts by mice with severe combined immune deficiency (SCID). Evidence that natural killer cells can mediate the specificity of marrow graft rejection.J Exp Med. 1987; 165: 1212-1217Crossref PubMed Scopus (232) Google Scholar) and demonstrate strong cytotoxicity against xenogeneic targets (7Gourlay WA Chambers WH Monaco AP Maki T Importance of natural killer cells in the rejection of hamster skin xenografts.Transplantation. 1998; 65: 727-734Crossref PubMed Scopus (39) Google Scholar, 8Lin Y Vandeputte M Waer M Natural killer cell‐ and macrophage‐mediated rejection of concordant xenografts in the absence of T and B cell responses.J Immunol. 1997; 158: 5658-5667Crossref PubMed Google Scholar). Rejection of solid organ transplants is thought to be mediated by allospecific T lymphocytes that recognize foreign MHC molecules on donor tissue (9Krensky AM Weiss A Crabtree G Davis MM Parham P T‐lymphocyte‐antigen interactions in transplant rejection.N Engl J Med. 1990; 322: 510-517Crossref PubMed Scopus (235) Google Scholar, 10Sayegh MH Turka LA The role of T‐cell costimulatory activation pathways in transplant rejection.N Engl J Med. 1998; 338: 1813-1821Crossref PubMed Scopus (496) Google Scholar). However, we have previously demonstrated that depletion of host CD8+ T cells does not prevent rejection in fully MHC‐mismatched models of rat liver and small intestinal transplantation (11Ogura Y Martinez OM Villanueva JC et al.Apoptosis and allograft rejection in the absence of CD8+ T cells.Transplantation. 2001; 71: 1827-1834Crossref PubMed Scopus (29) Google Scholar, 12Krams SM Hayashi M Fox CK et al.CD8+ cells are not necessary for allograft rejection or the induction of apoptosis in an experimental model of small intestinal transplantation.J Immunol. 1998; 160: 3673-3680Crossref PubMed Google Scholar), indicating that in the absence of CD8+ T cells, an alternate pathway can mediate graft rejection. Recent interest has focused on the role of NK cells in allograft rejection after solid organ transplantation. NK‐cell infiltration into allografts has been described in multiple experimental transplant models (13Maier S Tertilt C Chambron N et al.Inhibition of natural killer cells results in acceptance of cardiac allografts in CD28‐/‐ mice.Nat Med. 2001; 7: 557-562Crossref PubMed Scopus (180) Google Scholar, 14Kondo T Morita K Watarai Y et al.Early increased chemokine expression and production in murine allogeneic skin grafts is mediated by natural killer cells.Transplantation. 2000; 69: 969-977Crossref PubMed Scopus (53) Google Scholar, 15Petersson E Ostraat O Ekberg H et al.Allogeneic heart transplantation activates alloreactive NK cells.Cell Immunol. 1997; 175: 25-32Crossref PubMed Scopus (39) Google Scholar, 16Fuggle SV Immunophenotypic analysis of leukocyte infiltration in the renal transplant.Immunol Lett. 1991; 29: 143-146Crossref PubMed Scopus (14) Google Scholar, 17Hsieh CL Ogura Y Obara H et al.Identification, cloning, and characterization of a novel rat natural killer receptor, RNKP30: a molecule expressed in liver allografts.Transplantation. 2004; 77: 121-128Crossref PubMed Scopus (12) Google Scholar), and alloreactive NK cells have been shown to be activated in allogeneic cardiac transplantation (15Petersson E Ostraat O Ekberg H et al.Allogeneic heart transplantation activates alloreactive NK cells.Cell Immunol. 1997; 175: 25-32Crossref PubMed Scopus (39) Google Scholar). Acute liver allograft rejection is characterized by a mixed portal tract infiltrate containing mononuclear cells. The accumulation of activated lymphocytes into the allograft is essential to the pathogenesis of tissue injury. The mechanism by which activated lymphocytes are recruited to the graft from the circulation is poorly understood but probably involves local chemotactic factors that promote the migration, positioning and retention of effector cells in the graft (18Goddard S Williams A Morland C et al.Differential expression of chemokines and chemokine receptors shapes the inflammatory response in rejecting human liver transplants.Transplantation. 2001; 72: 1957-1967Crossref PubMed Scopus (124) Google Scholar). Chemokines are low molecular weight proteins with chemotactic properties for leukocytes and are grouped into families based on their cysteine motif. They are expressed and secreted by a wide variety of cell types including lymphocytes (19Wang JD Nonomura N Takahara S et al.Lymphotactin: a key regulator of lymphocyte trafficking during acute graft rejection.Immunology. 1998; 95: 56-61Crossref PubMed Scopus (32) Google Scholar) and endothelial components of rejecting allografts (18Goddard S Williams A Morland C et al.Differential expression of chemokines and chemokine receptors shapes the inflammatory response in rejecting human liver transplants.Transplantation. 2001; 72: 1957-1967Crossref PubMed Scopus (124) Google Scholar, 20Robertson H Morley AR Talbot D Callanan K Kirby JA Renal allograft rejection: beta‐chemokine involvement in the development of tubulitis.Transplantation. 2000; 69: 684-687Crossref PubMed Scopus (65) Google Scholar, 21Adams DH Hubscher S Fear J Johnston J Shaw S Afford S Hepatic expression of macrophage inflammatory protein‐1 alpha and macrophage inflammatory protein‐1 beta after liver transplantation.Transplantation. 1996; 61: 817-825Crossref PubMed Scopus (85) Google Scholar) in response to activation (22Lukacs NW Hogaboam C Campbell E Kunkel SL Chemokines: function, regulation and alteration of inflammatory responses.Chem Immunol. 1999; 72: 102-120Crossref PubMed Scopus (83) Google Scholar). Several studies have shown that CXCL10 (IFN‐γ‐inducible protein‐10), and CX3CL1 (fractalkine), are upregulated in rejecting murine cardiac allografts (23Fairchild RL VanBuskirk AM Kondo T Wakely ME Orosz CG Expression of chemokine genes during rejection and long‐term acceptance of cardiac allografts.Transplantation. 1997; 63: 1807-1812Crossref PubMed Scopus (92) Google Scholar, 24Robinson LA Nataraj C Thomas DW et al.A role for fractalkine and its receptor (CX3CR1) in cardiac allograft rejection.J Immunol. 2000; 165: 6067-6072Crossref PubMed Scopus (155) Google Scholar). Both CXCL10 and CX3CL1 are induced by IFN‐γ (25Fraticelli P Sironi M Bianchi G et al.Fractalkine (CX3CL1) as an amplification circuit of polarized Th1 responses.J Clin Invest. 2001; 107: 1173-1181Crossref PubMed Scopus (289) Google Scholar, 26Rollins BJ Chemokines.Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The CC chemokines, CCL2 (monocyte chemoattractant protein‐1) and CCL3 (macrophage inflammatory protein‐1α), have also been detected in cardiac allografts (27Miura M Morita K Kobayashi H et al.Monokine induced by IFN‐gamma is a dominant factor directing T cells into murine cardiac allografts during acute rejection.J Immunol. 2001; 167: 3494-3504Crossref PubMed Scopus (132) Google Scholar, 28Morita K Miura M Paolone DR et al.Early chemokine cascades in murine cardiac grafts regulate T cell recruitment and progression of acute allograft rejection.J Immunol. 2001; 167: 2979-2984Crossref PubMed Scopus (121) Google Scholar). NK cells, similar to T cells, are capable of migrating in response to the chemokines CXCL10, CX3CL1, CCL2 and CCL3 (29Robertson MJ Role of chemokines in the biology of natural killer cells.J Leukoc Biol. 2002; 71: 173-183Crossref PubMed Google Scholar, 30Maghazachi AA Al‐Aoukaty A Schall TJ C‐C chemokines induce the chemotaxis of NK and IL‐2‐activated NK cells. Role for G proteins..J Immunol. 1994; 153: 4969-4977Crossref PubMed Google Scholar). Recipient‐derived NK cells can amplify the early chemokine expression in allogeneic skin grafts (14Kondo T Morita K Watarai Y et al.Early increased chemokine expression and production in murine allogeneic skin grafts is mediated by natural killer cells.Transplantation. 2000; 69: 969-977Crossref PubMed Scopus (53) Google Scholar). In addition, NK cells migrate to sites of liver infection through a CCL3‐dependent pathway and produce high levels of IFN‐γ in murine cytomegalovirus‐infected livers (31Salazar‐Mather TP Orange JS Biron CA Early murine cytomegalovirus (MCMV) infection induces liver natural killer (NK) cell inflammation and protection through macrophage inflammatory protein 1alpha (MIP‐1alpha)‐dependent pathways.J Exp Med. 1998; 187: 1-14Crossref PubMed Scopus (300) Google Scholar, 32Salazar‐Mather TP Hamilton TA Biron CA A chemokine‐to‐cytokine‐to‐chemokine cascade critical in antiviral defense.J Clin Invest. 2000; 105: 985-993Crossref PubMed Scopus (210) Google Scholar). In the current study, we examine the role of NK cells in acute allograft rejection using a high responder model of rat orthotopic liver transplantation (OLTx). We demonstrate that increased expression of the chemokines CCL2, CCL3 and CXCL10 are detected in liver grafts as early as 6 h post‐transplant. This is accompanied by the accumulation of recipient‐derived NK cells. Importantly, our data also indicate that graft‐infiltrating NK cells are a major source of IFN‐γ and that liver allograft survival is prolonged in the absence of NK cells. These results suggest that recipient‐derived NK cells recruited to the allograft early after transplantation produce IFN‐γ and facilitate the ensuing adaptive immune response that culminates in graft rejection. Inbred male Dark Agouti (DA) rats (RT1a) and Lewis rats (RT1l), weighing 220–239 g, were purchased from Harlan (Indianapolis, IN). All animals were housed in accordance with institutional animal care and had access to water and standard laboratory chow ad libitum. Lewis rats were grafted with Lewis livers in the syngeneic group or with DA livers in the allogeneic group. Lewis recipients reject DA livers with a median survival time of 10 days (range: 9–12 days) (33Egawa H Martinez OM Quinn MB et al.Acute liver allograft rejection in the rat. An analysis of the immune response.Transplantation. 1995; 59: 97-102Crossref PubMed Scopus (29) Google Scholar). Donor and recipient surgeries were carried out under anesthesia with isoflurane (Abbott Laboratories, North Chicago, IL). OLTx was performed with a modification of Kamada and Calne's technique (34Kamada N Calne RY Orthotopic liver transplantation in the rat. Technique using cuff for portal vein anastomosis and biliary drainage.Transplantation. 1979; 28: 47-50Crossref PubMed Scopus (688) Google Scholar) without hepatic artery reconstruction. The liver was perfused with 15 mL of lactated Ringer's solution at 4°C through the catheter placed in the abdominal aorta, and the excised graft was stored in lactated Ringer's solution at 4°C. Cold ischemic time was ≤90 min. Upon completion of the recipient's hepatectomy, the graft was transplanted orthotopically. The anhepatic phase was ≤16 min. No immunosuppression was given to the recipient rats in this study. In some experiments, NK cells were depleted in vivo by one intraperitoneal injection (50 μL) of rabbit anti‐asialo‐GM1 antibodies (AGM1) (Wako Chemicals, Richmond, VA) the day before transplantation. This protocol completely depletes NK cells for 7 days after transplantation. The recipient rats were sacrificed at 6, 12 and 24 h and days 2, 3, 5 and 7 post‐transplant. Liver tissue was snap frozen for mRNA analysis. When indicated, a portion of the liver graft was utilized for the isolation of liver infiltrating mononuclear cells (LIMC). LIMC were prepared as previously described (11Ogura Y Martinez OM Villanueva JC et al.Apoptosis and allograft rejection in the absence of CD8+ T cells.Transplantation. 2001; 71: 1827-1834Crossref PubMed Scopus (29) Google Scholar). After systemic heparinization followed by exsanguination, the graft was perfused in situ via the portal vein with 15 mL of Ca2+‐free PBS to remove residual blood from the liver graft. The graft was subsequently removed and perfused through the portal vein with HBSS (GIBCO BRL, Grand Island, NY) containing 0.5 mg/mL collagenase (Sigma, St. Louis, MO), 0.02 mg/mL DNase I (Sigma) and 10% fetal calf serum (FCS). The liver tissue was cut into small pieces, resuspended in HBSS/collagenase/DNase I solution, and completely digested on a tilting device in a 37°C incubator for 30 min. The digested liver‐cell suspension was filtered through a nylon mesh (100 μm pore size) to remove debris, and LIMC were isolated by centrifugation over Ficoll (Ficoll‐Paque Plus, Amersham Pharmacia Biotech, Piscataway, NJ). Isolated LIMC were washed and counted. Cells were incubated with specific mAbs; fluorescein isothiocyanate (FITC)‐anti‐αβTCR (R7.3, Serotec Inc, Raleigh NC), ‐anti‐RT1Aa,b (C3, BD PharMingen, San Diego, CA) and PE‐anti‐NKR‐P1 (10/78, Serotec) for 30 min on ice and then washed twice in fluorescence‐activated cell sorting (FACS) buffer (PBS, 1% FCS, 0.1% sodium azide). To distinguish between cells of donor or recipient origin, FITC‐anti‐RT1Aa,b mAb was used to detect MHC class I expressed on donor cells (RT1a) but not on recipient cells (RT1l). Flow cytometric analysis was performed on a FACScan flow cytometer using CellQuest software (Becton Dickinson, San Jose, CA). The corresponding isotype matched antibodies (Dako, Carpinteria, CA) were used for negative controls and to set appropriate quadrants. Total RNA was isolated from liver grafts using TRIzol (GIBCO BRL) (35Hayashi M Martinez OM Krams SM Burns W Esquivel CO Characterization of allograft rejection in an experimental model of small intestinal transplantation.J Gastrointest Surg. 1998; 2: 325-332Crossref PubMed Scopus (22) Google Scholar). RNA integrity was confirmed by detection of the 28S and 18S RNA bands following electrophoresis in a 1.0% agarose gel. The concentration of RNA was measured by spectrophotometry (Beckman DU640B, Beckman Instruments, Inc., Fullerton, CA). RPA experiments (RiboQuant Multi‐Probe RNase Protection Assay System, BD PharMingen) were performed according to the manufacturer's recommendations. Probes containing rat CCL2, CCL3, CXCL10, CX3CL1, rGAPDH and r/mL32 (Torrey Pines Biolabs, La Jolla, CA) were used to detect chemokines. Anti‐sense riboprobes were prepared by in vitro transcription with either T7 (CCL2, CXCL10, CX3CL1 and r/mL32) (BD PharMingen) or SP6 (CCL3 and rGAPDH) (Invitrogen, Carlsbad, CA) RNA polymerase with the incorporation of [α32P]UTP (NEN Life Science Products, Boston, MA) at 37°C according to the manufacturer's directions (BD PharMingen) using the PharMingen RiboQuant in vitro transcription kit. Total RNA (10.0 μg) was hybridized with a [α32P]UTP‐labeled RNA probe (0.5−1.0 × 106 cpm each probe/sample) followed by RNase digestion. Protected bands were resolved on a 5.0% nondenaturing polyacrylamide gel and exposed to X‐ray film. The undigested probe set was run also as a marker for each experiment. The intensity of the protected band was determined by densitometry (Image Analyzer IS 2000, Alpha Innotech, San Leandro, CA), and each value was normalized against its corresponding GAPDH band intensity. The levels of IFN‐γ in rat serum were measured by ELISA using commercially available antibodies (BioSource International, Camarillo, CA). All serum samples were analyzed in triplicate. This assay was determined to have a sensitivity of 10 pg/mL using recombinant rat IFN‐γ as a standard (BioSource Int.). LIMC were cultured at a density of 106 cells/mL in RPMI 1640 medium supplemented with 10% FCS, 50 μM 2‐mercaptoethanol, 10 mM Hepes, 100 U/mL penicillin, 100 μg/mL streptomycin (GIBCO BRL) and 1000 U/mL human recombinant IL‐2 (Biological Resources Branch, National Cancer Institute (NCI)) for 24 h at 37°C. Brefeldin A (10 μg/mL; Sigma) was added during the last 6 h to inhibit cytokine secretion. Cells were washed, and surface staining using FITC‐anti‐αβTCR or ‐anti‐NKR‐P1 (10/78, Serotec) mAb was performed in FACS buffer plus brefeldin A for 30 min. Cells were then washed, fixed for 15 min using 250 μL of Cytofix (BD PharMingen), permeabilized for 15 min in permeabilization buffer (FACS buffer/0.5% saponin) and then incubated with PE‐anti‐rat IFN‐γ mAb (DB‐1, BD PharMingen) or PE‐mouse IgG1 (Dako) as an isotype control for 30 min at 4°C. After one wash with permeabilization buffer, cells were washed in FACS buffer without saponin. Stained cells were acquired by FACS can immediately and analyzed with CellQuest software. Data are expressed as mean values ± SD. Statistical analyses were performed by Student's t‐test or Welch's t‐test, where appropriate. In previous studies, we determined that CD8+ T cell depletion does not prolong the survival of liver allografts in a high‐responder OLTx (11Ogura Y Martinez OM Villanueva JC et al.Apoptosis and allograft rejection in the absence of CD8+ T cells.Transplantation. 2001; 71: 1827-1834Crossref PubMed Scopus (29) Google Scholar). We did, however, note a marked infiltration of NKR‐P1+ NK cells in these liver allografts. To expand upon this observation, NK‐cell infiltration was examined in a model where fully allogeneic donor DA (RT1a) livers were transplanted into Lewis (RT1l) recipients, and LIMC were isolated according to our previously published procedures (11Ogura Y Martinez OM Villanueva JC et al.Apoptosis and allograft rejection in the absence of CD8+ T cells.Transplantation. 2001; 71: 1827-1834Crossref PubMed Scopus (29) Google Scholar). LIMC were labeled with mAbs against NKR‐P1, αβTCR and RT1Aa,b for analysis by two‐color flow cytometry. Twenty‐four hours after transplantation, 18.6 ± 3.0% (n = 3) of the LIMC isolated from allografts were αβTCR+ T cells, while 56.4 ± 6.2% (n = 3) were αβTCR− NKR‐P1+ NK cells. A representative experiment is shown in Figure 1A. To determine if the NK cells were of donor or recipient origin, LIMC were labeled with FITC‐anti‐RT1Aa,b mAb specific for donor MHC class I (RT1a) in combination with PE‐anti‐NKR‐P1 mAb. At 24 h post‐transplant, 22.9 ± 6.1% (n = 3) of the LIMC were NK cells of recipient origin (RT1Aa,b,−). A representative experiment is shown in Figure 1B. We included only the NKR‐P1high‐positive sub‐sets to eliminate NKT cell and monocyte populations, since hepatic NKT cells and activated monocytes exist mainly in NKR‐P1dim‐positive subsets in the rat (36Steiniger B Stehling O Scriba A Grau V Monocytes in the rat: phenotype and function during acute allograft rejection.Immunol Rev. 2001; 184: 38-44Crossref PubMed Scopus (35) Google Scholar, 37Matsuura A Kinebuchi M Chen HZ et al.NKT cells in the rat: organ‐specific distribution of NK T cells expressing distinct V alpha 14 chains.J Immunol. 2000; 164: 3140-3148Crossref PubMed Scopus (42) Google Scholar). As previously discussed, αβTCR− NKR‐P1+ NK cells are a major component of the infiltrating cells in liver allografts. To evaluate the kinetics of NK‐cell infiltration after transplantation, LIMC isolated from allografts were analyzed for the proportion of NK cells at eight time points post‐transplant. The proportion of NK in LIMC cells peaked at 12–24 h post‐transplant (56.4 ± 6.2%; n = 3) and decreased to 35.6 ± 3.0% (n = 3) by day 2 as the proportion of T cells increased in the allograft. NK cells of recipient origin (NKR‐P1+ RT1Aa,b−), infiltrated the grafts very early post‐transplant constituting over half of the NK cells at 6 h post‐transplant (Figure 2A). The proportion of recipient‐derived NK cells in allografts peaked at 12 h post‐transplant, decreased by day 3, then increased again at day 5 and comprised 29.5 ± 7.0% of the total number of infiltrating cells at day 7 post‐transplant (Figure 2B). Conversely, in syngeneic liver grafts the overall proportion of NK cells and T cells were 32.2 ± 2.1% and 31.2 ± 4.0%, respectively, prior to transplant. NK cells peaked at 12 h post‐transplant (NK cells, 49.7 ± 3.3% and T cells 19.9 ± 5.4%) and then decreased to pre‐transplant levels by day 3 post‐transplant (NK cells, 30.6 ± 3.3% and T cells 32.4 ± 3.4%) and remained constant (NK cells, 34.0 ± 4.3% and T cells 30.4 ± 4.4%). In syngeneic grafts, it is not possible to separately quantitate the numbers of host and infiltrating NK and T cells thus these numbers reflect the total cell pool. These data suggest that there is a bimodal infiltration of NK cells into liver allografts, early as a result of nonspecific surgical stress, including ischemia/reperfusion injury and later at the time that effector cells are infiltrating the allograft and mediating rejection. Because NK cells were recruited to liver allografts rapidly following transplantation and because chemokines are important in the trafficking of lymphoid cells to areas of inflammation, we determined the expression of chemokines in liver grafts. Total RNA was isolated from both syngeneic and allogeneic grafts at 0, 6, 12 and 24 h and days 3 and 7 post‐transplant and analyzed by RPA (Figure 3). The chemokines CCL2 and CXCL10 were elevated as early as 6 h post‐transplant in both syngeneic and allogeneic grafts, suggesting that these chemokines were increased in liver tissue due to nonspecific surgical stress and ischemia/reperfusion injury. Expression of CCL3 was detected by 6 h post‐transplant in both syngeneic and allogeneic grafts (Figure 3A) but was significantly increased (p = 0.015) in the allografts as compared to the syngeneic grafts (Figure 3B, bottom left panel). CXCL10 and CX3CL1 were significantly upregulated (p = 0.014 and 0.013, respectively) in the allografts at day 3 post‐transplant as compared to the syngeneic grafts (Figure 3A, B, top right panel and bottom right panel), suggesting a role for these chemokines in promoting the recruitment of effector cells to allogeneic tissue. To define the role of NK cells after liver transplantation we treated a group of rat liver allograft recipients (n = 7) with a single dose (50 μL) AGM1 or control rabbit serum (500 μg), on the day prior to transplantation. Using this protocol NK cells are quickly (by 24 h) depleted from the periphery, and begin to reappear in the blood 7 days later (Figure 4B). Depletion of NK cells was confirmed by flow cytometry in all graft recipients treated with AGM1. We determined, by flow cytometry, that AGM1 does bind to rat NK cells, NKT cells and the majority of T cells, however, NK cells are AGM1bright as compared to T cells (data not shown). Interestingly, AGM1 treatment depletes virtually all of the NK cells and NKT cells, yet spares the majority of T cells (Figure 4A, B). Allograft recipients depleted of NK cells had significantly prolonged survival (p = 0.018) as compared to control allograft recipients (n = 7) (Figure 4). These data suggest that NK cells have a role in the early events post‐transplantation that contribute to liver allograft rejection. We and others have demonstrated that there is increased expression of inflammatory cytokines during rejection of liver allografts (33Egawa H Martinez OM Quinn MB et al.Acute liver allograft rejection in the rat. An analysis of the immune response.Transplantation. 1995; 59: 97-102Crossref PubMed Scopus (29) Google Scholar, 38Hoffmann MW Wonigeit K Steinhoff G Herzbeck H Flad HD Pichlmayr R Production of cytokines (TNF‐alpha, IL‐1‐beta) and endothelial cell activation in human liver allograft rejection.Transplantation. 1993; 55: 329-335Crossref PubMed Scopus (106) Google Scholar, 39Shirwan H Cosenza CA Wang HK Wu GD Makowka L Cramer DV Prevention of orthotopic liver allograft rejection in rats with a short‐term brequinar sodium therapy. Analysis of intragraft cytokine gene expression.Transplantation. 1994; 57: 1072-1080Crossref PubMed Scopus (18) Google Scholar). To specifically analyze the IFN‐γ levels post‐transplantation, serum was obtained from recipients of liver grafts (n = 3–5) at 0, 6, 12 and 24 hours and 2, 3, 5 and 7 days after transplant. IFN‐γ was generally below the level of detection in recipients of syngeneic grafts at all time points (Figure 5A). In contrast, the levels of IFN‐γ increased by day 1 post‐transplantation in recipients of allogeneic liver grafts and peaked at 3 days post‐transplantation. To assess the role of NK cells in the production of IFN‐γ we depleted NK cells, in vivo, by injecting AGM1 into a group (n = 3) of allograft recipients. The levels of IFN‐γ in the serum was significantly reduced (p = 0.001) by >70%, 3 days after transplantation (the peak day for IFN‐γ production), in NK‐cell‐depleted graft recipients as compared to recipients of liver allografts treated with control rabbit serum (Figure 5B). To further analyze the cellular production of IFN‐γ in the allografts, LIMC were isolated from allografts, post‐transplant and IFN‐γ production was assessed in permeabilized cells and analyzed by flow cytometry. In a representative experiment, 2 day post‐transplant, 18% of NK cells and 9% of T cells within the allograft produced IFN‐γ (Figure 6, top right and lower right panels, respectively), while 2% of NK cells and 0.4% of T cells in the nontransplanted Lewis rat liver were IFN‐γ positive (data not shown). Furthermore, the proportion of IFN‐γ+ NKR‐P1+ cells was markedly greater (8.3%, middle top panel) than that of IFN‐γ+αβTCR+ cells (3.4%, middle bottom panel) among the total LIMC. Similar results were observed at 12 h post‐transplant (absolute number = 0.6 million IFN‐γ+ NK cells/g liver and 0.24 million IFN‐γ+ T cells/g liver), and 3 days post‐transplant (0.55 million IFN‐γ+ NK cells/g liver and 0.2 million IFN‐γ+ T/g liver) as were observed on day 2 post‐transplant (0.6 million IFN‐γ+ NK cells/g and 0.38 million IFN‐γ+ T cells/g). Specifically, there are a greater number of NK cells expressing IFN‐γ than T cells. In contrast, on day 7 post‐transplant there are more T cells than NK cells producing IFN‐γ (0.4 million IFN‐γ+ NK cells/g and 2.45 million IFN‐γ+ T cells/g). Donor NK cells are the source of 20–25% of the NK cell derived IFN‐γ on day 2 post‐transplant (data not shown). These data further indicate that NK cells are a major source of IFN‐γ in liver allografts early after transplantation but not at the time of rejection. We demonstrate that recipient‐derived NK cells are recru" @default.
• W1996709249 created "2016-06-24" @default.
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• W1996709249 date "2005-09-01" @default.
• W1996709249 modified "2023-09-30" @default.
• W1996709249 title "IFN‐γ, Produced by NK Cells that Infiltrate Liver Allografts Early After Transplantation, Links the Innate and Adaptive Immune Responses" @default.
• W1996709249 cites W1536891815 @default.
• W1996709249 cites W1556867369 @default.
• W1996709249 cites W1570007714 @default.
• W1996709249 cites W1593015928 @default.
• W1996709249 cites W1593435314 @default.
• W1996709249 cites W1604338287 @default.
• W1996709249 cites W1660408415 @default.
• W1996709249 cites W1969095061 @default.
• W1996709249 cites W1983082877 @default.
• W1996709249 cites W1985009368 @default.
• W1996709249 cites W1994121614 @default.
• W1996709249 cites W2002276647 @default.
• W1996709249 cites W2008221423 @default.
• W1996709249 cites W2014794643 @default.
• W1996709249 cites W2020421485 @default.
• W1996709249 cites W2020968325 @default.
• W1996709249 cites W2021377837 @default.
• W1996709249 cites W2025123869 @default.
• W1996709249 cites W2044996210 @default.
• W1996709249 cites W2053217797 @default.
• W1996709249 cites W2056883845 @default.
• W1996709249 cites W2059647731 @default.
• W1996709249 cites W2066086221 @default.
• W1996709249 cites W2066425602 @default.
• W1996709249 cites W2067137484 @default.
• W1996709249 cites W2069669865 @default.
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• W1996709249 cites W2075542703 @default.
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• W1996709249 cites W2077705267 @default.
• W1996709249 cites W2082535269 @default.
• W1996709249 cites W2092919093 @default.
• W1996709249 cites W2104976130 @default.
• W1996709249 cites W2116493322 @default.
• W1996709249 cites W2118637098 @default.
• W1996709249 cites W2120446140 @default.
• W1996709249 cites W2121266155 @default.
• W1996709249 cites W2127908593 @default.
• W1996709249 cites W2128409610 @default.
• W1996709249 cites W2131282129 @default.
• W1996709249 cites W2132507383 @default.
• W1996709249 cites W2143480395 @default.
• W1996709249 cites W2144789797 @default.
• W1996709249 cites W2160886090 @default.
• W1996709249 cites W2164359892 @default.
• W1996709249 cites W2165234706 @default.
• W1996709249 cites W2336153715 @default.
• W1996709249 cites W2406749751 @default.
• W1996709249 cites W2430442937 @default.
• W1996709249 cites W287378556 @default.
• W1996709249 cites W4211264883 @default.
• W1996709249 cites W4244132088 @default.
• W1996709249 cites W4290250305 @default.
• W1996709249 cites W4295201567 @default.
• W1996709249 cites W4313342469 @default.
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