Volume 78, Issue 2, Pages (July 2010)

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Volume 78, Issue 2, Pages 152-159 (July 2010) Recombinant human C1-inhibitor prevents acute antibody-mediated rejection in alloimmunized baboons  Xavier Tillou, Nicolas Poirier, Stéphanie Le Bas-Bernardet, Jeremy Hervouet, David Minault, Karine Renaudin, Fabio Vistoli, Georges Karam, Mohamed Daha, Jean Paul Soulillou, Gilles Blancho  Kidney International  Volume 78, Issue 2, Pages 152-159 (July 2010) DOI: 10.1038/ki.2010.75 Copyright © 2010 International Society of Nephrology Terms and Conditions

Figure 1 Specificity and cytotoxicity of the induced alloantibody (alloAb). (a) Kinetics of anti-donor immunoglobulin G (IgG) development after primary immunization analyzed by fluorescence-activated cell sorting (FACS) each week. (b) Appearance of specific IgG, measured by FACS, against donor but not third-party or autologous cells in immunized animals at day 28 after primary immunization in comparison with day 0. (c) Complement-mediated cytotoxicity of immunized recipient serum against donor (n=6) but not third-party cells (n=3) at the plateau of immunization. (d) Depletion of anti-donor class I Abs in immunized recipient sera by donor platelet absorption abolished cytotoxicity, proving the induced alloAb to be specific for donor major histocompatability complex class I (n=6). (e) Anti-donor alloAb cytotoxicity is inhibited by increased recombinant human C1-inhibitor (rhC1INH) concentration (n=3). Kidney International 2010 78, 152-159DOI: (10.1038/ki.2010.75) Copyright © 2010 International Society of Nephrology Terms and Conditions

Figure 2 Clinical and biological effects of recombinant human C1-inhibitor (rhC1INH) administration in presensitized transplanted recipients. (a) Renal allograft survival in nonimmunized baboons (n=3), immunized baboons without therapy (n=4), in those treated with 100U/kg of rhC1INH (n=2), or those with 200U/kg of rhC1INH (n=4). *P=0.0134 versus the nonimmunized group. **P=0.0058 versus the nontreated immunized group. (b) Daily diuresis of immunized baboons without therapy (n=4), treated with 100U/kg of rhC1INH (n=2) or 200U/kg of rhC1INH (n=4). (c) Daily creatinine levels and (d) blood urea nitrogen measurement after transplantation in immunized recipients without therapy (n=4), in those treated with 100U/kg of rhC1INH (n=2) or in those treated with 200U/kg of rhC1INH (n=4). Kidney International 2010 78, 152-159DOI: (10.1038/ki.2010.75) Copyright © 2010 International Society of Nephrology Terms and Conditions

Figure 3 Analysis of kidney allograft histology. Histology (hematoxylin and eosin staining) and immunostaining of infiltrating cells (CD3 and CD20) and complement deposition (C5b-9, C4d, and C3c) in kidney biopsies or grafts explanted from untreated nonimmunized recipients, untreated immunized recipients, and recombinant human C1-inhibitor (rhC1INH) (200U/kg)-treated immunized recipients at early phase (days 2–3) or later point (1 week). Kidney International 2010 78, 152-159DOI: (10.1038/ki.2010.75) Copyright © 2010 International Society of Nephrology Terms and Conditions

Figure 4 Complement monitoring. (a) C1 inhibitor levels in immunized transplant recipients treated with recombinant human C1-inhibitor (rhC1INH) at 100U/kg (n=2; dashed line) or 200U/kg (n=2; solid line). (b) Total immunoglobulin G, (c) C3, and (d) C4 plasma level measurement after transplantation in immunized recipients without therapy (n=4; dotted line) or in those treated with rhC1INH (n=4; solid line). (e) The 50% complement hemolysis (CH50) analyzed by enzyme-linked immunosorbent assay, (f) Functional CH50 kinetics analyzed during the first few hours after transplantation by sheep erythrocyte assay on two immunized recipients treated with 200U/kg of rhC1INH, (g) the 50% alternate pathway (AP50), (h) the 50% mannose-binding lectin pathway (MBL 50) and (i) MBL plasma level measurement after transplantation in immunized recipients without therapy (n=4; dotted line) or in those treated with 100U/kg of rhC1INH (n=2; solid line). Kidney International 2010 78, 152-159DOI: (10.1038/ki.2010.75) Copyright © 2010 International Society of Nephrology Terms and Conditions