Weak response of porcine C5a receptor towards human C5a in miniature pig endothelial cells and PMNs
| Autor: | Bumrae Cho, Hee Jung Kang, Curie Ahn, Yoe Sik Bae, Min Soo Kim, Hyori Kim, Kye Sook Yi, Junho Chung, Sukmook Lee, Insu Ha, Jae Yong Kim, Jaeseok Yang, Yoon Ho Kang, Seung Yong Hwang, Ki Tae Bang |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
Neutrophils
Swine Molecular Sequence Data Transplantation Heterologous Immunology Complement C5a chemical and pharmacologic phenomena Biology C5a receptor Cell Line Flow cytometry medicine Animals Humans Secretion Amino Acid Sequence Cytoskeleton Cell adhesion Receptor Anaphylatoxin C5a Aorta Transplantation medicine.diagnostic_test Reverse Transcriptase Polymerase Chain Reaction Cell adhesion molecule hemic and immune systems Chemotaxis Molecular biology Recombinant Proteins Gene Expression Regulation Swine Miniature Endothelium Vascular Intracellular |
| Zdroj: | Xenotransplantation. 14:563-571 |
| ISSN: | 1399-3089 0908-665X |
| DOI: | 10.1111/j.1399-3089.2007.00421.x |
| Popis: | Background: The anaphylatoxin C5a is a potent inflamma- tory molecule generated during complement activation. Although some reports have implicated C5a in xenograft rejection, to date, the molec- ular compatibility between human C5a and porcine C5a receptor (C5aR) has been little studied. To examine the need for pC5aR-deficient pig in xenotransplantaion, we aimed to look at the degree of direct interaction between human C5a (recipient side) and porcine endothelial cells (PECs) and porcine polymorphonuclear neutrophils (PMNs) (donor side). Methods: Following the treatment of human C5a to isolated porcine PMNs, transmigration of PMNs was measured by Transwell system and superoxide generation by cytochrome c reduction assay. Next, the effects of human C5a on several intracellular signaling pathways were further checked; actin cytoskeletal change was observed under a confocal microscope after staining with Alexa Fluor-546-phalloidin, intracellular calcium mobilization was measured by spectrofluorophotometer. The degree of direct effect of human C5a on porcine PMNs was compared with that in human PMNs. Finally, microarray was performed to monitor the effect of human C5a on gene expression of PEC and the expression of several candidate proteins was checked by flow cytometry. Results: We found that human C5a was able to induce chemotaxis, superoxide generation, actin cytoskeletal change, and intracellular cal- cium mobilization in porcine PMNs. However, higher concentration of human C5a was required to stimulate porcine PMNs in comparison with activating human PMNs. The amino acid sequences of porcine C5aR with those of human C5aR showed a sequence homology of only 67%. To elucidate the effect of human C5a to PECs, microarray analysis following the treatment of PECs with human C5a was performed. These data showed that human C5a did not significantly affect gene tran- scription patterns in PECs. Additionally, treatment of PECs with human C5a also did not induce protein expression of several cell adhesion molecules, including vascular cell adhesion molecule-1, intercellular adhesion molecule-1, P-selectin, and E-selectin, or secretion of inter- leukin-8 from PECs. Conclusions: These results suggest that human C5a may play a minor role on PEC activation possibly due to molecular incompatibility across the species barrier. |
| Databáze: | OpenAIRE |
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