- Email: [email protected]
Value of Pretransplantation Cytokine Profiles for Predicting Acute Rejection in Renal Transplant Recipients
Value of Pretransplantation Cytokine Profiles for Predicting Acute Rejection in Renal Transplant Recipients M. Lessan-Pezeshki, A. Amirzargar, A. Fathi, M.R. Khatami, B. Einollahi, V. Pourfarziani, J. Azmandian, F. Khosravi, B. Ansaripour, and B. Nikbin ABSTRACT Episodes of acute rejection may represent an important risk factor for the development of chronic allograft nephropathy. Various studies have shown that pretransplant cytokine profiles in recipient blood are associated with transplant outcome. Serum samples were collected 24 hours before transplantation from 57 patients (38 men and 19 women of age 36 ⫾ 5 years) receiving kidneys from unrelated living donors. Additional samples were collected at 1 and 2 weeks after transplantation, as well as during every rejection episode. The immunosuppression consisted of a cyclosporine, prednisolone, and mycophenolate mofetil. Among the transplanted patients, 19 (33.3%) individuals experienced an acute rejection episode based on an increased level of serum creatinine and blood urea nitrogen during the first 14 days after transplantation. TGF-, IL-2 and IFN-␥ serum levels were determined by an ELISA method using Bindermed system kits. The mean concentration of TGF- before transplantation tended to be lower among patients with acute rejection episodes compared to those with stable graft (75,265 versus 85,394 pg/mL; P ⫽ .34) and at 1 week after transplantation (77,558 versus 84,390 pg/mL), although the differences were not significant. Among patients with rejection the mean IL-2 concentration was significantly higher before, at 1 week, and at 2 weeks after transplantation (15.0 versus 6.8 pg/mL, P ⫽ .005; 19.0 versus 4.9 pg/mL, P ⫽ .001; and 21.1 versus 4.7 pg/mL, P ⫽ .0001). The mean concentration of IFN-␥ was significantly higher pre- and at 1 and 2 weeks posttransplantation in patients with acute rejection episodes (161.1 versus 65.2, 175.6 versus 66.5 and 173.7 versus 77.1 pg/mL, all P ⬍ .001). In conclusion, evaluation of Th1 cytokines before transplantation may represent valuable predictive marker for an acute rejection episode.
C
YTOKINES PLAYa central role in directing both the magnitude and type of immune response generated against organ transplants. These substances, which are normally expressed at low levels, are rapidly upregulated at the onset of an immune response. Consequently, they are early predictors of graft dysfunction, and, in theory, might provide information about mechanisms that underlie immune attacks.1 Acute rejection episodes (ARE) are a major cause of morbidity in renal transplant recipients, and may be considered an important factor in the development of chronic allograft nephropathy.2 The T-helper (Th1) cytokines—Interleukin (IL)-2 and interferon gamma (IFN-␥)—promote the cellular immune response, specifically activating cytotoxic T lymphocytes, natural killer cells, and monocytes. These effector cells
infiltrate the graft causing cellular rejection.3 Transforming growth factor- (TGF-), a pleiotropic, multifunctional cytokine, is transported to the circulation after being produced by a wide variety of cells, including monocytes, lymphocytes, renal tubular cells, endothelial cells, and airway epithelial cells. TGF- is a fibrogenic cytokine that may be an important mediator of chronic rejection in renal transplants.4 Nicholson et al5 reported a correlation between increased TGF- mRNA and increased mRNA for collagens as well as tissue inhibitors of metalloproteinases From the Imam Khomeini Medical Center, Tehran, Iran. Address reprint requests to Mahboob Lessan-Pezeshki, Nephrology Department, Keshavarz Blvd., Tehran, Iran. E-mail: [email protected]
0041-1345/05/$–see front matter doi:10.1016/j.transproceed.2005.08.031
© 2005 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710
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Transplantation Proceedings, 37, 2982–2984 (2005)
PRETRANSPLANTATION CYTOKINE PROFILES
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(TIMP1 and TIMP2).5 The authors suggested that TGF- may be a profibrotic cytokine in human renal allografts. Cohen and Nast 6 found that, unlike mRNAs for other inflammatory cytokines, levels of TGF mRNA strongly correlate with interstitial fibrosis, a hallmark of chronic rejection. The aim of this study was to predict acute AREs in renal transplant recipients by measurement of cytokine profiles before transplantation. PATIENTS AND METHODS The subjects were 57 patients (38 men and 19 women, mean age 36 ⫾ 5 years) who underwent living-unrelated kidney transplantation between September 2002 and August 2003. In each case, a serum sample (5 mL of blood without anticoagulants) was collected 24 hours before as well as 1 and 2 weeks after transplantation. All patients were placed on cyclosporine (5 mg/kg initially followed by maintenance doses of 2 to 2.5 mg/kg; target cyclosporine levels of 50 to 150 ng/mL); prednisolone (120 mg/d initially, tapering to 10 mg/d during 6 months after transplantation), and mycophenolate mofetil (1000 mg twice daily). All patients had a negative lymphocytotoxic crossmatch before transplantation. For this study, an ARE was defined as an elevated serum creatinine and elevated blood urea nitrogen. Nineteen (33.3%) of the 57 patients experienced an ARE in the first 14 days after transplantation, whereas the grafts in the other 38 patients (66.7%) remained stable during this period. Serum levels of IL-2, IFN-␥, and TGF- were determined at each of the three time points using an ELISA method (Bender Medsystem Kits, Germany). We compared the findings in the ARE(⫹) group versus ARE(⫺) group. Statistical analysis was performed using SPSS 11.5. P values ⬍.05 were considered significant.
RESULTS
The cytokine findings for the ARE(⫹) and ARE(⫺) groups are summarized in Table 1 Patients who developed ARE in the first 2 weeks (n ⫽ 19) showed significantly higher serum IL-2 levels than the ARE(⫺) group (n ⫽ 38) at 1 day before transplantation (15.0 versus 6.8 pg/mL, respectively; P ⫽ .005) as well as at 1 week after transplantation (19.0 versus 4.9 pg/mL, respectively; P ⫽ .001) and 2 weeks after transplantation (21.1 versus 4.7 pg/mL, respectively; P ⫽ .0001). The ARE(⫹) group also showed significantly higher serum IFN-␥ levels at all three time points; before transplantation, 161.1 versus 65.2 pg/mL, respectively, P ⫽ .001; 1 week, 175.6 versus 66.5 pg/mL, respectively, P ⫽ .001; 2 weeks, 173.7 versus 77.1 pg/mL, respectively, P ⫽ .001). There were trends toward higher mean TGF- concentra-
tion in the ARE(⫹) group before transplantation (85394 versus 75265 pg/mL, respectively; P ⫽ .34) and at 1 week after transplantation (84,390 versus 77,558 pg/mL, respectively; P ⫽ .6); however, the differences were not significant. DISCUSSION
In organ transplantation, donor–recipient histoincompatibility induces an inflammatory immune response that often results in an ARE. Renal tissue expression of TGF- has long been associated with renal damage. Immunohistochemical studies have shown that in chronic allograft nephropathy renal tubular cells produce this factor in large amounts. In one study, investigators found that sustained TGF- expression in rodent renal tissue contributes to the development of glomerulosclerosis and tubulointerstitial fibrisis.7 Sadeghi et al8 observed that increased IFN-␥ prior to transplantation is indicative of an early ARE after transplantation. Rostaing et al9 reported that the ARE group showed a significantly greater frequency of IL-2/ IFN-␥ double-positive cells prior to transplantation. Cho et al10 recorded levels of IL-2, IL-2–receptor, and tumor necrosis factor alpha (TNF-␣) that were approximately 150% to 200% above pretransplant levels at 1 month after transplantation. In this investigation, we compared cytokine levels in two groups of patients; those who experienced an ARE during the first 2 weeks after transplantation (n ⫽ 19), and those who had stable grafts with no evidence of renal dysfunction during this period (n ⫽ 38). In all 57 cases, donor selection was based on blood group matching and negative white blood cell crossmatch results between recipients and donors. Histocompatibility matching between donor and recipient was not considered. We evaluated IL-2, IFN-␥ and TGF- levels before and after renal transplantation to assess whether elevation of these cytokines can signal ARE facilitating an early diagnosis of allograft rejection. The data indicate that a Th1 cytokine pattern namely, significantly elevated IL-2 and IFN-␥ levels, at 1 day prior to renal transplantation was predictive of an early ARE. On the other hand, the mean concentration of TGF- before transplantation was slightly lower among patients with an acute rejection episode than those with stable grafts (85,394 versus 75,265 pg/mL, P ⫽ .34) or at 1 week after transplantation (84,390 versus 77,558 pg/mL); however, the differences was not significant. At 2 weeks posttransplantation, the ARE(⫹) group showed slightly higher levels of TGF- than the ARE(⫺) group.
Table 1. Comparison of the Groups’ Serum Cytokine Levels Before Transplantation and at 1 and 2 Weeks After Transplantation IFN-␥ (pg/mL)
ARE(⫹) (n ⫽ 19) ARE(⫺) (n ⫽ 38) P
TGF- (pg/mL)
IL-2 (pg/mL)
C0
C1
C2
C0
C1
C2
C0
C1
C2
161.1 65.2 .001
175.6 66.5 .001
173.7 77.1 .001
15.0 6.8 .005
19.0 4.9 .001
21.1 4.7 .001
75,265 85,394 .3
77,558 84,390 .6
91,581 67,920 .08
Abbreviations: C0, 1 day before transplantation; C1, 1 week posttransplantation; C2, 2 weeks posttransplantation; ARE, acute rejection episode; IFN-␥, interferon gamma; IL, interleukin; TGF-, transforming growth factor-beta.
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We suggest that this observation may have been due to the effects of cyclosporine or to immune reactivity. In conclusion, our data support the findings of other investigators that Th1 cytokines are valuable predictors of an early ARE after renal transplantation. REFERENCES 1. Dallman MJ: Cytokines and transplantation: Th1/Th2 regulation of the immune response to solid organ transplantation in the adult. Curr Opin Immunol 7:632, 1995 2. Tenjani A, Sullivan EK: The impact of acute rejection on chronic rejection: a report of the north American pediatric renal transplant cooperative study. Pediatric Transplant 4:107, 2000 3. Delios MM, Josien R, Manghetti M: Predominant Th1 cell infiltration in acute rejection episodes of human kidney grafts. Kidney Int 51:1876, 1997 4. Terrell TG, Working PK, Chow CP: Int Rev Exp Pathol 34:43, 1993
LESSAN-PEZESHKI, AMIRZARGAR, FATHI ET AL 5. Nicholson ML, Bicknell GR, Williams ST, et al: Is TGF- a profibrotic cytokine in human renal transplant. Transplant Proc 30:592, 1998 6. Cohen AH, Nast CC: TGF in renal allograft rejection. Miner Electrlyte Metab 24:197, 1998 7. Ymamoto T, Noble NA, Miller DE: Sustained expression of TGF-Beta underlies development of progressive kidney fibrosis. Kidney Int 45:916, 1994 8. Sadeghi M, Daniel V, Weimer R, et al: Pre-transplant Th1 and post-transplant Th2 cytokine patterns are associated with early acute rejection in renal transplant recipients. Clin Transplant 17:151, 2003 9. Rostaing L, Puyoo O, Tkaczuk J, et al: Differences in type 1 and type 2 intra cytoplasmic cytokines, detected by flowcytometry, according to immunosupressin in stable renal allograft. Clin Transplant 13:400, 1999 10. Cho WH, Kim HT, Sohn CY, et al: Significance of IL-2, IL-2R, IL-6 and TNF-␣ as a diagnostic test of acute rejection after renal transplantation. Transplant Proc 30:2967, 1998
C
YTOKINES PLAYa central role in directing both the magnitude and type of immune response generated against organ transplants. These substances, which are normally expressed at low levels, are rapidly upregulated at the onset of an immune response. Consequently, they are early predictors of graft dysfunction, and, in theory, might provide information about mechanisms that underlie immune attacks.1 Acute rejection episodes (ARE) are a major cause of morbidity in renal transplant recipients, and may be considered an important factor in the development of chronic allograft nephropathy.2 The T-helper (Th1) cytokines—Interleukin (IL)-2 and interferon gamma (IFN-␥)—promote the cellular immune response, specifically activating cytotoxic T lymphocytes, natural killer cells, and monocytes. These effector cells
infiltrate the graft causing cellular rejection.3 Transforming growth factor- (TGF-), a pleiotropic, multifunctional cytokine, is transported to the circulation after being produced by a wide variety of cells, including monocytes, lymphocytes, renal tubular cells, endothelial cells, and airway epithelial cells. TGF- is a fibrogenic cytokine that may be an important mediator of chronic rejection in renal transplants.4 Nicholson et al5 reported a correlation between increased TGF- mRNA and increased mRNA for collagens as well as tissue inhibitors of metalloproteinases From the Imam Khomeini Medical Center, Tehran, Iran. Address reprint requests to Mahboob Lessan-Pezeshki, Nephrology Department, Keshavarz Blvd., Tehran, Iran. E-mail: [email protected]
0041-1345/05/$–see front matter doi:10.1016/j.transproceed.2005.08.031
© 2005 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710
2982
Transplantation Proceedings, 37, 2982–2984 (2005)
PRETRANSPLANTATION CYTOKINE PROFILES
2983
(TIMP1 and TIMP2).5 The authors suggested that TGF- may be a profibrotic cytokine in human renal allografts. Cohen and Nast 6 found that, unlike mRNAs for other inflammatory cytokines, levels of TGF mRNA strongly correlate with interstitial fibrosis, a hallmark of chronic rejection. The aim of this study was to predict acute AREs in renal transplant recipients by measurement of cytokine profiles before transplantation. PATIENTS AND METHODS The subjects were 57 patients (38 men and 19 women, mean age 36 ⫾ 5 years) who underwent living-unrelated kidney transplantation between September 2002 and August 2003. In each case, a serum sample (5 mL of blood without anticoagulants) was collected 24 hours before as well as 1 and 2 weeks after transplantation. All patients were placed on cyclosporine (5 mg/kg initially followed by maintenance doses of 2 to 2.5 mg/kg; target cyclosporine levels of 50 to 150 ng/mL); prednisolone (120 mg/d initially, tapering to 10 mg/d during 6 months after transplantation), and mycophenolate mofetil (1000 mg twice daily). All patients had a negative lymphocytotoxic crossmatch before transplantation. For this study, an ARE was defined as an elevated serum creatinine and elevated blood urea nitrogen. Nineteen (33.3%) of the 57 patients experienced an ARE in the first 14 days after transplantation, whereas the grafts in the other 38 patients (66.7%) remained stable during this period. Serum levels of IL-2, IFN-␥, and TGF- were determined at each of the three time points using an ELISA method (Bender Medsystem Kits, Germany). We compared the findings in the ARE(⫹) group versus ARE(⫺) group. Statistical analysis was performed using SPSS 11.5. P values ⬍.05 were considered significant.
RESULTS
The cytokine findings for the ARE(⫹) and ARE(⫺) groups are summarized in Table 1 Patients who developed ARE in the first 2 weeks (n ⫽ 19) showed significantly higher serum IL-2 levels than the ARE(⫺) group (n ⫽ 38) at 1 day before transplantation (15.0 versus 6.8 pg/mL, respectively; P ⫽ .005) as well as at 1 week after transplantation (19.0 versus 4.9 pg/mL, respectively; P ⫽ .001) and 2 weeks after transplantation (21.1 versus 4.7 pg/mL, respectively; P ⫽ .0001). The ARE(⫹) group also showed significantly higher serum IFN-␥ levels at all three time points; before transplantation, 161.1 versus 65.2 pg/mL, respectively, P ⫽ .001; 1 week, 175.6 versus 66.5 pg/mL, respectively, P ⫽ .001; 2 weeks, 173.7 versus 77.1 pg/mL, respectively, P ⫽ .001). There were trends toward higher mean TGF- concentra-
tion in the ARE(⫹) group before transplantation (85394 versus 75265 pg/mL, respectively; P ⫽ .34) and at 1 week after transplantation (84,390 versus 77,558 pg/mL, respectively; P ⫽ .6); however, the differences were not significant. DISCUSSION
In organ transplantation, donor–recipient histoincompatibility induces an inflammatory immune response that often results in an ARE. Renal tissue expression of TGF- has long been associated with renal damage. Immunohistochemical studies have shown that in chronic allograft nephropathy renal tubular cells produce this factor in large amounts. In one study, investigators found that sustained TGF- expression in rodent renal tissue contributes to the development of glomerulosclerosis and tubulointerstitial fibrisis.7 Sadeghi et al8 observed that increased IFN-␥ prior to transplantation is indicative of an early ARE after transplantation. Rostaing et al9 reported that the ARE group showed a significantly greater frequency of IL-2/ IFN-␥ double-positive cells prior to transplantation. Cho et al10 recorded levels of IL-2, IL-2–receptor, and tumor necrosis factor alpha (TNF-␣) that were approximately 150% to 200% above pretransplant levels at 1 month after transplantation. In this investigation, we compared cytokine levels in two groups of patients; those who experienced an ARE during the first 2 weeks after transplantation (n ⫽ 19), and those who had stable grafts with no evidence of renal dysfunction during this period (n ⫽ 38). In all 57 cases, donor selection was based on blood group matching and negative white blood cell crossmatch results between recipients and donors. Histocompatibility matching between donor and recipient was not considered. We evaluated IL-2, IFN-␥ and TGF- levels before and after renal transplantation to assess whether elevation of these cytokines can signal ARE facilitating an early diagnosis of allograft rejection. The data indicate that a Th1 cytokine pattern namely, significantly elevated IL-2 and IFN-␥ levels, at 1 day prior to renal transplantation was predictive of an early ARE. On the other hand, the mean concentration of TGF- before transplantation was slightly lower among patients with an acute rejection episode than those with stable grafts (85,394 versus 75,265 pg/mL, P ⫽ .34) or at 1 week after transplantation (84,390 versus 77,558 pg/mL); however, the differences was not significant. At 2 weeks posttransplantation, the ARE(⫹) group showed slightly higher levels of TGF- than the ARE(⫺) group.
Table 1. Comparison of the Groups’ Serum Cytokine Levels Before Transplantation and at 1 and 2 Weeks After Transplantation IFN-␥ (pg/mL)
ARE(⫹) (n ⫽ 19) ARE(⫺) (n ⫽ 38) P
TGF- (pg/mL)
IL-2 (pg/mL)
C0
C1
C2
C0
C1
C2
C0
C1
C2
161.1 65.2 .001
175.6 66.5 .001
173.7 77.1 .001
15.0 6.8 .005
19.0 4.9 .001
21.1 4.7 .001
75,265 85,394 .3
77,558 84,390 .6
91,581 67,920 .08
Abbreviations: C0, 1 day before transplantation; C1, 1 week posttransplantation; C2, 2 weeks posttransplantation; ARE, acute rejection episode; IFN-␥, interferon gamma; IL, interleukin; TGF-, transforming growth factor-beta.
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We suggest that this observation may have been due to the effects of cyclosporine or to immune reactivity. In conclusion, our data support the findings of other investigators that Th1 cytokines are valuable predictors of an early ARE after renal transplantation. REFERENCES 1. Dallman MJ: Cytokines and transplantation: Th1/Th2 regulation of the immune response to solid organ transplantation in the adult. Curr Opin Immunol 7:632, 1995 2. Tenjani A, Sullivan EK: The impact of acute rejection on chronic rejection: a report of the north American pediatric renal transplant cooperative study. Pediatric Transplant 4:107, 2000 3. Delios MM, Josien R, Manghetti M: Predominant Th1 cell infiltration in acute rejection episodes of human kidney grafts. Kidney Int 51:1876, 1997 4. Terrell TG, Working PK, Chow CP: Int Rev Exp Pathol 34:43, 1993
LESSAN-PEZESHKI, AMIRZARGAR, FATHI ET AL 5. Nicholson ML, Bicknell GR, Williams ST, et al: Is TGF- a profibrotic cytokine in human renal transplant. Transplant Proc 30:592, 1998 6. Cohen AH, Nast CC: TGF in renal allograft rejection. Miner Electrlyte Metab 24:197, 1998 7. Ymamoto T, Noble NA, Miller DE: Sustained expression of TGF-Beta underlies development of progressive kidney fibrosis. Kidney Int 45:916, 1994 8. Sadeghi M, Daniel V, Weimer R, et al: Pre-transplant Th1 and post-transplant Th2 cytokine patterns are associated with early acute rejection in renal transplant recipients. Clin Transplant 17:151, 2003 9. Rostaing L, Puyoo O, Tkaczuk J, et al: Differences in type 1 and type 2 intra cytoplasmic cytokines, detected by flowcytometry, according to immunosupressin in stable renal allograft. Clin Transplant 13:400, 1999 10. Cho WH, Kim HT, Sohn CY, et al: Significance of IL-2, IL-2R, IL-6 and TNF-␣ as a diagnostic test of acute rejection after renal transplantation. Transplant Proc 30:2967, 1998