| Plasmodium faiciparum malaria remains one of the most important causes of death in the tropics. Cerebral malaria (CM) is the major lethal complications of this infection. The sequestration of red blood cells infected by mature form of Plasmodium faiciparum (PRBC) in the cerebral microvasculature is considered to be the essential underlying pathological process of CM , leading to the alterations of blood flow, metabolic dysfunction and local toxicity, due to the cytoadherence of PRBC to the. endothelial cells of postcapillary venules. Understanding the molecular basis on the adhesion between cytoadherent ligands of PRBC and vascular endothelial receptors may provide insight into the pathology of severe faiciparum malaria and the possibility of new therapeutic strategies.PRBC adherence to endothelial cells is mediated by several different endothelial cell surface receptors including intercellular adhesion molecule 1(ICAM-1), CDS 6, thrombospondin(TSP), vascular cell adhesion molecule 1(VCAM-1) and endothelial leukocyte adhesion molecule l(ELAM-l). Among these adhesion molecules, ICAM-1 may play the most important role in the pathophysiology of CM, due to the up-regulated expression of ICAM-1 during the process of CM. It was shown that ICAM-1 and bound parasitized red blood cells co-localized in brain capillaries of cerebral malaria patients. Furthermore, parasites isolated from cerebral malaria patients showed the highest capacity of binding to ICAM-1. So the development of ICAM-1 analogs based on the critical contact residues for PRBCs may be engineered to bind, lyse, and kill sequestered intraerythrocytic parasites in cases of severe and complicated faiciparum malaria.Ockenhouse and others demonstrated that PRBC bound to the first amino-terminal domain of human ICAM-1, at a site distinct but partially overlapping with the recognition sites for LFA1 and human rhinoviruses. But the molecular basis on the adhesion of malaria-infected erythrocytes to ICAM-1 is still unknown. Phage display appears to represent a new approach for the identification of peptides that interfere with ICAM-1 binding to PRBC.In this study, a 12-mer random peptide library was used to screen short peptides specific to monoclonal antibody against ICAM-1 which had been identified to have the ability to block the binding of ICAM-1 to both PRBC and LFA-1. After three rounds of biopanning, the binding phages were found to be enriched by ELISA and Dot-ELISA, and the recovery rate of the third round was about one thousand times higher than that of the first one. 30 clones from the third round were randomly selected, and 26 of them were positive identified by sandwich ELISA. The insert DNA fragments of the positive clones were sequenced, and the amino acid sequences of peptides deduced from the corresponding DNA sequences of these positive clones were P1(KLYLIAEGSVAA, 18/26, 69.2%), P2(HSSHTSNTLPSV ,2/26, 7.7%), P3(HPPKLLESMQRS, 2/26, 7.7%), P4(IEVPFLKKHYTL, 1/26, 3.8%), P5(LPLSLRPNTTPF, 1/26, 3.8%) and P6(SSMTHQHARVDT, 2/26, 7.7%), respectively. Peptide KLYLIAEGSVAA was the most frequently appeared peptide, which showed 50% homogeneity to 8-17 aa of ICAM-1, and the homologous sequence K ...L....GS V might probably play significant role in the binding reaction of ICAM-1 to PRBC.Competitive ELISA, Dot-ELISA and Western blot were used to evaluate the character of phage-borne peptides binding to 15.2 antibody, resulting all these clones but clone 16 specifically bound to 15.2 antibody. 15.2 antibody recognized a 65kDa protein from phage PI and P3 which was defined as coat protein III by Western blot. The result that most of these short peptides competitively inhibited the binding of the antibody to the immobilized ICAM-1, and ICAM-1 competitively inhibit the binding of the antibody tothese peptides by competitive ELISA suggested that these peptides may be the short analogs of 1C AM-1.Bioactivity of phage-borne peptide PI and P3 binding to LFA-1 was detected by immunohistochemical staining. The results showed that PI and P3 specifically...
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