A potential role of TLR2 in xenograft rejection of porcine iliac endothelial cells: An in vitro study.
Animals
Biomarkers
/ metabolism
Blotting, Western
Endothelial Cells
/ immunology
Enzyme-Linked Immunosorbent Assay
Graft Rejection
/ immunology
Humans
Iliac Artery
/ immunology
In Vitro Techniques
Real-Time Polymerase Chain Reaction
Swine
Toll-Like Receptor 2
/ immunology
Transplantation, Heterologous
complement
porcine iliac endothelial cells
toll-like receptor 2
xenograft rejection
xenotransplantation
Journal
Xenotransplantation
ISSN: 1399-3089
Titre abrégé: Xenotransplantation
Pays: Denmark
ID NLM: 9438793
Informations de publication
Date de publication:
09 2019
09 2019
Historique:
received:
02
07
2018
revised:
26
03
2019
accepted:
18
04
2019
pubmed:
28
5
2019
medline:
24
9
2020
entrez:
26
5
2019
Statut:
ppublish
Résumé
Porcine vascular endothelial cells are a major participant in xenograft rejection. The Toll-like receptor 2 (TLR2) pathway plays an important role in both innate and adaptive immunity. The specific role of TLR2 in the response to a xenograft has not been reported. Whether the TLR2 pathway in pig vascular endothelial cells is involved in acute rejection needs to be investigated, and the mechanism is explored. We used a modified antibody-dependent complement-mediated cytotoxicity (ADCC) assay to conduct in vitro experiments. In porcine iliac artery endothelial cells (PIECs), siRNA was used to knock down the expression of TLR2, CXCL8, and CCL2. The effect of human serum or inactivated human serum on the expression of TLR2 was analyzed by real-time PCR and Western blotting, and transwell assays were used to assess the chemotactic efficiency of PIECs on human monocyte-macrophages (THP-1 cells) and human neutrophils. The downstream signaling pathways activated by human serum were detected by Western blotting, and the regulation of proinflammatory chemokines and cytokines by TLR2 signaling was assessed by real-time PCR and ELISA. TLR2 was significantly upregulated in PIECs after exposure to human serum, and porcine proinflammatory chemokines, CXCL8 and CCL2, were induced, at least partially, in a TLR2-dependent pattern; the upregulated chemokines participated in the chemotaxis of human neutrophils and THP-1 cells across the species barrier. (i) TLR2 is significantly upregulated in PIECs by human serum, (ii) the elevated TLR2 participates in the chemotaxis of inflammatory cells through the secretion of chemokine CCL2 and CXCL8, and (iii) blockade of TLR2 would be beneficial for xenograft survival.
Sections du résumé
BACKGROUND
Porcine vascular endothelial cells are a major participant in xenograft rejection. The Toll-like receptor 2 (TLR2) pathway plays an important role in both innate and adaptive immunity. The specific role of TLR2 in the response to a xenograft has not been reported. Whether the TLR2 pathway in pig vascular endothelial cells is involved in acute rejection needs to be investigated, and the mechanism is explored.
METHODS
We used a modified antibody-dependent complement-mediated cytotoxicity (ADCC) assay to conduct in vitro experiments. In porcine iliac artery endothelial cells (PIECs), siRNA was used to knock down the expression of TLR2, CXCL8, and CCL2. The effect of human serum or inactivated human serum on the expression of TLR2 was analyzed by real-time PCR and Western blotting, and transwell assays were used to assess the chemotactic efficiency of PIECs on human monocyte-macrophages (THP-1 cells) and human neutrophils. The downstream signaling pathways activated by human serum were detected by Western blotting, and the regulation of proinflammatory chemokines and cytokines by TLR2 signaling was assessed by real-time PCR and ELISA.
RESULTS
TLR2 was significantly upregulated in PIECs after exposure to human serum, and porcine proinflammatory chemokines, CXCL8 and CCL2, were induced, at least partially, in a TLR2-dependent pattern; the upregulated chemokines participated in the chemotaxis of human neutrophils and THP-1 cells across the species barrier.
CONCLUSIONS
(i) TLR2 is significantly upregulated in PIECs by human serum, (ii) the elevated TLR2 participates in the chemotaxis of inflammatory cells through the secretion of chemokine CCL2 and CXCL8, and (iii) blockade of TLR2 would be beneficial for xenograft survival.
Substances chimiques
Biomarkers
0
Toll-Like Receptor 2
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e12526Subventions
Organisme : National Key R&D Program of China
ID : 2017YFC1103704
Pays : International
Organisme : Construction of the high level Hospitals in Guangdong province:2019; Shenzhen Foundation of Science and Technology
ID : JCYJ20170306092047810
Pays : International
Organisme : Construction of the high level Hospitals in Guangdong province:2019; Shenzhen Foundation of Science and Technology
ID : JCJY20160229204849975
Pays : International
Organisme : Construction of the high level Hospitals in Guangdong province:2019; Shenzhen Foundation of Science and Technology
ID : GJHZ20170314171357556
Pays : International
Organisme : National Natural Science Foundation of China
ID : 81502410
Pays : International
Organisme : National Natural Science Foundation of Guangdong province
ID : 2016A030313028
Pays : International
Organisme : Sanming Project of Medicine in Shenzhen
ID : SZSM201412020
Pays : International
Organisme : High Level Medical Discipline Construction of Shenzhen
ID : 2016031638
Pays : International
Organisme : Shenzhen Foundation of Health and Family Planning Commission
ID : SZXJ2017021
Pays : International
Organisme : Shenzhen Foundation of Health and Family Planning Commission
ID : SZXJ2018059
Pays : International
Informations de copyright
© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Références
Satyananda V, Hara H, Ezzelarab MB, Phelps C, Ayares D, Cooper DK. New concepts of immune modulation in xenotransplantation. Transplantation. 2013;96:937-945.
Yamada K, Scalea J. Current progress in xenogeneic tolerance. Curr opin organ transplant. 2012;17:168-173.
Ebrahimi A, Rahim F. Recent immunomodulatory strategies in transplantation. Immunol Invest. 2014;43:829-837.
Yu X, Jiang Y, Lu L, et al. A crucial role of IL-17 and IFN-gamma during acute rejection of peripheral nerve xenotransplantation in mice. PLoS ONE. 2012;7:e34419.
Schneider MK, Seebach JD. Current cellular innate immune hurdles in pig-to-primate xenotransplantation. Curr opin organ transplant. 2008;13:171-177.
Cowan PJ, Roussel JC, d'Apice AJ. The vascular and coagulation issues in xenotransplantation. Curr opin organ transplant. 2009;14:161-167.
Chandra AP, Ouyang LI, Yi S, et al. Chemokine and toll-like receptor signaling in macrophage mediated islet xenograft rejection. Xenotransplantation. 2007;14:48-59.
Ezzelarab MB, Ekser B, Azimzadeh A, et al. Systemic inflammation in xenograft recipients precedes activation of coagulation. Xenotransplantation. 2015;22:32-47.
Hancock WW, Gao W, Csizmadia V, Faia KL, Shemmeri N, Luster AD. Donor-derived IP-10 initiates development of acute allograft rejection. J Exp Med. 2001;193:975-980.
Lv Q, Li C, Mo Y, He L. The role of HMGB1 in heart transplantation. Immunol Lett. 2018;194:1-3.
Okamura Y, Watari M, Jerud ES, et al. The extra domain A of fibronectin activates Toll-like receptor 4. J Bio Chem. 2001;276:10229-10233.
Kirschning CJ, Schumann RR. TLR2: cellular sensor for microbial and endogenous molecular patterns. Curr Top Microbiol Immunol. 2002;270:121-144.
Loiarro M, Ruggiero V, Sette C. Targeting TLR/IL-1R signalling in human diseases. Mediators Inflamm. 2010;2010:674363.
Takeda K, Akira S. TLR signaling pathways. Semin Immunol. 2004;16:3-9.
Lee SO, Brown RA, Kang SH, Abdel Massih RC, Razonable RR. Toll-like receptor 4 polymorphisms and the risk of gram-negative bacterial infections after liver transplantation. Transplantation. 2011;92:690-696.
Jin MS, Lee JO. Structures of TLR-ligand complexes. Curr Opin Immunol. 2008;20:414-419.
Kruger B, Krick S, Dhillon N, et al. contributes to ischemia and reperfusion injury following human kidney transplantation. Proc Natl Acad Sci USA. 2009;106:3390-3395.
Krichen H, Gorgi Y, Dhaouadi T, et al. Toll-like receptor 4 and CD14 gene polymorphisms in Tunisian kidney transplantation. Transpl Proc. 2013;45:3472-3477.
Gruhn B, Kloppner N, Pfaffendorf-Regler N, et al. Toll-like 4 receptor variant, Asp299Gly, and reduced risk of hemorrhagic cystitis after hematopoietic stem cell transplantation. Biol Blood Marrow Transplant. 2012;18:958-963.
Li J-H, Zhao B, Zhu X-H, et al. Blockade of extracellular HMGB1 suppresses xenoreactive B cell responses and delays acute vascular xenogeneic rejection. Am J Transplant. 2015;15:2062-2074.
Ro H, Lee EW, Hong JH, et al. Roles of islet Toll-like receptors in pig to mouse islet xenotransplantation. Cell Transplant. 2013;22:1709-1722.
Li H, Wang G, Yu Y, et al. alpha-1,2-mannosidase MAN1C1 inhibits proliferation and invasion of clear cell renal cell carcinoma. J Cancer. 2018;9:4618-4626.
Bunk S, Ponnuswamy P, Trbic A, et al. IVIG induces apoptotic cell death in CD56(dim) NK cells resulting in inhibition of ADCC effector activity of human PBMC. Clin Immunol. 2018;198:62-70.
Zhou X, Zhou Y, Ding Q, et al. High level expression of B7H1 molecules by keratinocytes suppresses xeno- and allo-reactions by inducing type I regulatory T cells. Transpl Immunol. 2009;21:192-197.
Chen T, Wang J, Xue B, Kong Q, Liu Z, Yu B. Identification and characterization of a novel porcine endothelial cell-specific Tie1 promoter. Xenotransplantation. 2013;20:438-448.
Mulley WR, Li YQ, Wee JL, et al. Local expression of IDO, either alone or in combination with CD40Ig, IL10 or CTLA4Ig, inhibits indirect xenorejection responses. Xenotransplantation. 2008;15:174-183.
Chen J-C, Cai H-Y, Wang Y, et al. Up-regulation of stomatin expression by hypoxia and glucocorticoid stabilizes membrane-associated actin in alveolar epithelial cells. J Cell Mol Med. 2013;17:863-872.
Zhang P, Hou S, Chen J, et al. Smad4 deficiency in smooth muscle cells initiates the formation of aortic aneurysm. Circ Res. 2016;118:388-399.
Saggu G, Okubo K, Chen Y, et al. Cis interaction between sialylated FcgammaRIIA and the alphaI-domain of Mac-1 limits antibody-mediated neutrophil recruitment. Nat Commun. 2018;9:5058.
Bosmann M, Ward PA. Role of C3, C5 and anaphylatoxin receptors in acute lung injury and in sepsis. Adv Exp Med Biol. 2012;946:147-159.
Sultan P, Murray AG, McNiff JM, et al. Pig but not human interferon-gamma initiates human cell-mediated rejection of pig tissue in vivo. Proc Natl Acad Sci USA. 1997;94:8767-8772.
Citro A, Cantarelli E, Maffi P, et al. CXCR32/2 inhibition enhances pancreatic islet survival after transplantation. J Clin Investig. 2012;122:3647-3651.
Busch-Petersen J. Small molecule antagonists of the CXCR33 and CXCR33 chemokine receptors as therapeutic agents for the treatment of inflammatory diseases. Curr Top Med Chem. 2006;6:1345-1352.
Xiong X, Liu YU, Mei Y, et al. Novel protective role of myeloid differentiation 1 in pathological cardiac remodelling. Sci Rep. 2017;7:41857.
Baramova E, Foidart JM. Matrix metalloproteinase family. Cell Biol Int. 1995;19:239-242.
Turunen AJ, Lindgren L, Salmela KT, Kyllonen LE, Andersson S, Pesonen E. Matrix metalloproteinase-9 and graft Preservation injury in clinical renal transplantation. Transpl Proc. 2015;47:2831-2835.
Riewald M, Petrovan RJ, Donner A, Mueller BM, Ruf W. Activation of endothelial cell protease activated receptor 1 by the protein C pathway. Science. 2002;296:1880-1882.
Denis CV, Andre P, Saffaripour S, Wagner DD. Defect in regulated secretion of P-selectin affects leukocyte recruitment in von Willebrand factor-deficient mice. Proc Natl Acad Sci USA. 2001;98:4072-4077.