- Email: [email protected]
320: Donor-Specific Antibodies (DSA) Patterns after Pediatric Heart Transplantation with A + and – Crossmatch
S108
The Journal of Heart and Lung Transplantation, Vol 29, No 2S, February 2010
320 Donor-Specific Antibodies (DSA) Patterns after Pediatric Heart Transplantation with A ⴙ and – Crossmatch C. Reddy,1 E. Riddle,1 P. Checchia,1 C. Huddleston,2 S. Gandhi,2 D. Phelan,3 C. Canter.1 1Washington University School of Medicine, Saint Louis, MO; 2Washington University School of Medicine, Saint Louis, MO; 3Washington University School of Medicine, Saint Louis, MO. Purpose: Pediatric heart transplantation in the presence of DSA with a complement dependent cytotoxic crossmatch (CDCXM) can be successfully performed. The qualitative and quantitative changes in DSA after CDCXM heart transplantation are unknown. The purpose of this study was to characterize DSA behavior after ⫹CDCXM transplant and its relationship to rejection episodes. Methods and Materials: Post-transplant DSA activity in 12 ⫹CDCXM recipients from 1/07-9/09 was compared to 52 –CDCXM recipients. DSA were identified using single antigen bead Luminex assays. DSA were obtained pre-transplant, serially at the time of surveillance biopsy, and during rejection episodes within the first transplant year. Quantification of specific DSA strength was calculated by the ratio of median fluorescence intensity (MFI) to a ⫹ control value. Results: Pre-transplant DSA was present in 11/12 ⫹CDCXM patients compared to 3/50 –CDCXM patients (p⬍0.0001). Post-transplant DSA was present in 9/11 ⫹CDCXM patients and 9/32 –CDCXM (p⫽0.0035). Appearance/disappearance of individual DSA occurred in 8/11 ⫹CDCXM compared to 13/32 –CDCXM patients (p⫽ 0.088). Persistence of DSA in CDCXM ⫹ vs–was: 100 vs 54% (p⫽0.04) 1 month, 100 vs 21% (p⫽0.01) 6 months, and 100 vs 29% (p⫽0.33) 12 months after transplant (median follow-up for group⫽ 9 months). DSA ratios ranged between 0.2-2.3 pre-transplant and post-transplant without any demonstrable pattern in the rise and/or fall of ratios in association with rejection, rejection treatment, or time after transplant. Conclusions: DSA present pre-transplant is strongly associated with a ⫹CDCXM transplant. DSA persists after ⫹CDCXM transplants compared to –CDCXM transplants but may appear and disappear after transplantation in varying intensities that show no apparent correlation with rejection or time after transplant. DSA patterns after transplant would appear to be a poor marker for rejection and may represent accommodation in ⫹CDCXM transplants. 321 Il-17 Is a Mediator of Bleomycin-Induced Acute Lung Injury in Mice R.K. Braun, A.A. Martin, P. Sethupathi, M.G. Medina, C. Wigfield, R.B. Love. Loyola University Medical Center, Maywood, IL. Purpose: Murine lung T cell receptor (TCR) ␥␦ T cells are intraepithelial cells critical in regulating inflammation. We previously demonstrated in TCR ␦⫺/⫺ mice that bleomycin (blm) induced acute diffuse lung injury (ALI) is associated with delayed epithelial regeneration (1). As TCR ␥␦ T cells are a major source of IL-17 production, we tested if IL-17 knockout (ko) mice would demonstrate similar patterns of Blm-induced ALI. Methods and Materials: Wild type (WT) and IL-17 ko mice were subjected to either N/saline (control) or Blm (0.04 U) via intratracheal instillation. Bronchoalveolar (BAL) fluid was analyzed for cell count, CD4⫹, CD8⫹ and ␥␦ T cells as well as IL-17 producing T cells by flow cytometry.
Markers of Bleomycin-induced ALI
Total BAL cell number CD4⫹ CD8⫹ ␥␦ T cells IL-17⫹ CD4⫹ IL-17⫹ ␥␦ T cells VDL
WT control
IL-17 ko control
WT bleo
IL-17 ko bleo
p value
2.1 ⫾ 0.55 80 ⫾ 22 28 ⫾ 16 3⫾3 N.D. N.D. 0
3.0 ⫾ 0.8 44 ⫾ 38 32 ⫾ 8 5⫾4 N.D. N.D. 0
450 ⫾ 70 61,000 ⫾ 5,000 13,000 ⫾ 1,000 20,000 ⫾ 2,000 5.6 ⫾ 1.3% 76.8 ⫾ 18.9% 0.38 ⫾ 0.03
16 ⫾ 7 220 ⫾ 80 150 ⫾ 60 800 ⫾ 60 N.D. N.D. 0.07 ⫾ 0.04
0.017 0.031 0.005 0.018 — — 0.022
p value refers to treatment groups 4 & 5
Lung injury was quantified by volume density of lesion (VDL) method on histology (H&E) at day 7 (1). Results: In contrast to WT, IL-17 ko mice were protected from Blminduced ALI. Inflammation as determined by VDL and BAL cell number was significantly reduced across all T cell subpopulations and less histopathological injury was evident. Conclusions: IL-17 is a critical mediator of Blm-induced ALI demonstrated by significant decrease in inflammation. IL-17 producing T cells are a potential target in other forms of ALI. 1. Braun RK et al Inflammation ‘08 Jun;31(3):167 322 IL-17F Induces a Distinct Form of Anti-MHC Class I Induced Autoimmunity and Playing a Role in Chronic Rejection Post-Lung Transplantation H. Ilias Basha,1 V. Tiriveedhi,1 M. Takenaka,1 S. Ramachandran,1 G.A. Patterson,2 T. Mohanakumar.1,3 1Washington University School of Medicine, St. Louis, MO; 2Washington University School of Medicine, St. Louis, MO; 3Washington University School of Medicine, St. Louis, MO. Purpose: IL-17 (or IL-17A) producing Th17 cells specific to self antigens, have been implicated in the pathogenesis of de novo immunity to self antigens that leads to chronic rejection following lung transplantation (LTx). In this study, we determined the specific role for a related homodimeric Th17 cytokine, IL-17F, in the immunopathogenesis of antiMHC class I antibodies (Abs) induced immune response. Methods and Materials: Anti-MHC I Abs (anti-H2Kb) or control Abs (C1.18) were administered intrabronchially to IL-17A⫺/⫺ and Wild type C7BL/6 animals and followed for 30 days. Immunohistochemistry, T-cell stimulation studies,Chemokine and growth factor analysis by RT-PCR were performed. Results: Anti-MHC class I Ab administration in WT mice resulted in cellular infiltration characterized by CD4⫹ and CD11b⫹ cells around bronchioles and vessels, epithelial hyperplasia, fibrosis, and occlusion of distal airways by 30 days. In contrast, IL-17A⫺/⫺ mice demonstrated minimal lesions. IL-17F⫺/⫺ animals had a 30⫾14% decrease in CD4⫹ infiltrates and a 33⫾21% decrease in CD11b⫹ infiltrates around bronchioles and vessels and a 21⫹10% decrease in fibrosis. A significant decline in CD4⫹CD25⫹Foxp3⫹ T cells was also noted in anti-H2Kb treated WT (5.0% Vs 15.5%) and IL-17A⫺/⫺ (7.31% Vs 13.72%) lungs compared to control mice. Compared to WT mice, a specific increase in expression of IL-17F (9⫾3 fold), CXCL1 (4⫾1.5 fold), TRAF-6(12⫾3 fold) genes was also noted in IL-17A⫺/⫺ mice. Further, neutralization of IL-17F in IL17A⫺/⫺ mice completely abrogated the minimal lesions induced by antiMHC Class I Abs. Conclusions: Our findings suggest that IL-17F mediates a less severe yet distinct form of anti-MHC Class I induced autoimmune pathology compared to the classical IL-17A dependent Th17 pathway. Further, the combined neutralization of IL-17A and F completely abrogates the lesions in murine model of Obliterative Airway Disease and thus represents an effective therapeutic strategy for the prevention of chronic rejection following LTx. 323 Alloimmune Responses to Mismatched MHC Class I Antigens Induces Immune Responses to Self Antigens Leading to Chronic Rejection V. Tiriveedhi,1 D.H. Brand,2 D. Saini,1 S. Ramachandran,1 R. Hachem,3 E.P. Trulock,3 G.A. Patterson,4 T. Mohanakumar.1,5 1Washington University School of Medicine, St. Louis, MO; 2University of Tennessee, Memphis, TN; 3Washington University School of Medicine, St. Louis, MO; 4Washington University School of Medicine, St. Louis, MO; 5 Washington University School of Medicine, St. Louis, MO. Purpose: Chronic rejection represents the leading cause of long term graft failure following solid organ transplantation. Studies in our laboratory and others have recently identified circulating Abs to an airway epithelial self
The Journal of Heart and Lung Transplantation, Vol 29, No 2S, February 2010
320 Donor-Specific Antibodies (DSA) Patterns after Pediatric Heart Transplantation with A ⴙ and – Crossmatch C. Reddy,1 E. Riddle,1 P. Checchia,1 C. Huddleston,2 S. Gandhi,2 D. Phelan,3 C. Canter.1 1Washington University School of Medicine, Saint Louis, MO; 2Washington University School of Medicine, Saint Louis, MO; 3Washington University School of Medicine, Saint Louis, MO. Purpose: Pediatric heart transplantation in the presence of DSA with a complement dependent cytotoxic crossmatch (CDCXM) can be successfully performed. The qualitative and quantitative changes in DSA after CDCXM heart transplantation are unknown. The purpose of this study was to characterize DSA behavior after ⫹CDCXM transplant and its relationship to rejection episodes. Methods and Materials: Post-transplant DSA activity in 12 ⫹CDCXM recipients from 1/07-9/09 was compared to 52 –CDCXM recipients. DSA were identified using single antigen bead Luminex assays. DSA were obtained pre-transplant, serially at the time of surveillance biopsy, and during rejection episodes within the first transplant year. Quantification of specific DSA strength was calculated by the ratio of median fluorescence intensity (MFI) to a ⫹ control value. Results: Pre-transplant DSA was present in 11/12 ⫹CDCXM patients compared to 3/50 –CDCXM patients (p⬍0.0001). Post-transplant DSA was present in 9/11 ⫹CDCXM patients and 9/32 –CDCXM (p⫽0.0035). Appearance/disappearance of individual DSA occurred in 8/11 ⫹CDCXM compared to 13/32 –CDCXM patients (p⫽ 0.088). Persistence of DSA in CDCXM ⫹ vs–was: 100 vs 54% (p⫽0.04) 1 month, 100 vs 21% (p⫽0.01) 6 months, and 100 vs 29% (p⫽0.33) 12 months after transplant (median follow-up for group⫽ 9 months). DSA ratios ranged between 0.2-2.3 pre-transplant and post-transplant without any demonstrable pattern in the rise and/or fall of ratios in association with rejection, rejection treatment, or time after transplant. Conclusions: DSA present pre-transplant is strongly associated with a ⫹CDCXM transplant. DSA persists after ⫹CDCXM transplants compared to –CDCXM transplants but may appear and disappear after transplantation in varying intensities that show no apparent correlation with rejection or time after transplant. DSA patterns after transplant would appear to be a poor marker for rejection and may represent accommodation in ⫹CDCXM transplants. 321 Il-17 Is a Mediator of Bleomycin-Induced Acute Lung Injury in Mice R.K. Braun, A.A. Martin, P. Sethupathi, M.G. Medina, C. Wigfield, R.B. Love. Loyola University Medical Center, Maywood, IL. Purpose: Murine lung T cell receptor (TCR) ␥␦ T cells are intraepithelial cells critical in regulating inflammation. We previously demonstrated in TCR ␦⫺/⫺ mice that bleomycin (blm) induced acute diffuse lung injury (ALI) is associated with delayed epithelial regeneration (1). As TCR ␥␦ T cells are a major source of IL-17 production, we tested if IL-17 knockout (ko) mice would demonstrate similar patterns of Blm-induced ALI. Methods and Materials: Wild type (WT) and IL-17 ko mice were subjected to either N/saline (control) or Blm (0.04 U) via intratracheal instillation. Bronchoalveolar (BAL) fluid was analyzed for cell count, CD4⫹, CD8⫹ and ␥␦ T cells as well as IL-17 producing T cells by flow cytometry.
Markers of Bleomycin-induced ALI
Total BAL cell number CD4⫹ CD8⫹ ␥␦ T cells IL-17⫹ CD4⫹ IL-17⫹ ␥␦ T cells VDL
WT control
IL-17 ko control
WT bleo
IL-17 ko bleo
p value
2.1 ⫾ 0.55 80 ⫾ 22 28 ⫾ 16 3⫾3 N.D. N.D. 0
3.0 ⫾ 0.8 44 ⫾ 38 32 ⫾ 8 5⫾4 N.D. N.D. 0
450 ⫾ 70 61,000 ⫾ 5,000 13,000 ⫾ 1,000 20,000 ⫾ 2,000 5.6 ⫾ 1.3% 76.8 ⫾ 18.9% 0.38 ⫾ 0.03
16 ⫾ 7 220 ⫾ 80 150 ⫾ 60 800 ⫾ 60 N.D. N.D. 0.07 ⫾ 0.04
0.017 0.031 0.005 0.018 — — 0.022
p value refers to treatment groups 4 & 5
Lung injury was quantified by volume density of lesion (VDL) method on histology (H&E) at day 7 (1). Results: In contrast to WT, IL-17 ko mice were protected from Blminduced ALI. Inflammation as determined by VDL and BAL cell number was significantly reduced across all T cell subpopulations and less histopathological injury was evident. Conclusions: IL-17 is a critical mediator of Blm-induced ALI demonstrated by significant decrease in inflammation. IL-17 producing T cells are a potential target in other forms of ALI. 1. Braun RK et al Inflammation ‘08 Jun;31(3):167 322 IL-17F Induces a Distinct Form of Anti-MHC Class I Induced Autoimmunity and Playing a Role in Chronic Rejection Post-Lung Transplantation H. Ilias Basha,1 V. Tiriveedhi,1 M. Takenaka,1 S. Ramachandran,1 G.A. Patterson,2 T. Mohanakumar.1,3 1Washington University School of Medicine, St. Louis, MO; 2Washington University School of Medicine, St. Louis, MO; 3Washington University School of Medicine, St. Louis, MO. Purpose: IL-17 (or IL-17A) producing Th17 cells specific to self antigens, have been implicated in the pathogenesis of de novo immunity to self antigens that leads to chronic rejection following lung transplantation (LTx). In this study, we determined the specific role for a related homodimeric Th17 cytokine, IL-17F, in the immunopathogenesis of antiMHC class I antibodies (Abs) induced immune response. Methods and Materials: Anti-MHC I Abs (anti-H2Kb) or control Abs (C1.18) were administered intrabronchially to IL-17A⫺/⫺ and Wild type C7BL/6 animals and followed for 30 days. Immunohistochemistry, T-cell stimulation studies,Chemokine and growth factor analysis by RT-PCR were performed. Results: Anti-MHC class I Ab administration in WT mice resulted in cellular infiltration characterized by CD4⫹ and CD11b⫹ cells around bronchioles and vessels, epithelial hyperplasia, fibrosis, and occlusion of distal airways by 30 days. In contrast, IL-17A⫺/⫺ mice demonstrated minimal lesions. IL-17F⫺/⫺ animals had a 30⫾14% decrease in CD4⫹ infiltrates and a 33⫾21% decrease in CD11b⫹ infiltrates around bronchioles and vessels and a 21⫹10% decrease in fibrosis. A significant decline in CD4⫹CD25⫹Foxp3⫹ T cells was also noted in anti-H2Kb treated WT (5.0% Vs 15.5%) and IL-17A⫺/⫺ (7.31% Vs 13.72%) lungs compared to control mice. Compared to WT mice, a specific increase in expression of IL-17F (9⫾3 fold), CXCL1 (4⫾1.5 fold), TRAF-6(12⫾3 fold) genes was also noted in IL-17A⫺/⫺ mice. Further, neutralization of IL-17F in IL17A⫺/⫺ mice completely abrogated the minimal lesions induced by antiMHC Class I Abs. Conclusions: Our findings suggest that IL-17F mediates a less severe yet distinct form of anti-MHC Class I induced autoimmune pathology compared to the classical IL-17A dependent Th17 pathway. Further, the combined neutralization of IL-17A and F completely abrogates the lesions in murine model of Obliterative Airway Disease and thus represents an effective therapeutic strategy for the prevention of chronic rejection following LTx. 323 Alloimmune Responses to Mismatched MHC Class I Antigens Induces Immune Responses to Self Antigens Leading to Chronic Rejection V. Tiriveedhi,1 D.H. Brand,2 D. Saini,1 S. Ramachandran,1 R. Hachem,3 E.P. Trulock,3 G.A. Patterson,4 T. Mohanakumar.1,5 1Washington University School of Medicine, St. Louis, MO; 2University of Tennessee, Memphis, TN; 3Washington University School of Medicine, St. Louis, MO; 4Washington University School of Medicine, St. Louis, MO; 5 Washington University School of Medicine, St. Louis, MO. Purpose: Chronic rejection represents the leading cause of long term graft failure following solid organ transplantation. Studies in our laboratory and others have recently identified circulating Abs to an airway epithelial self