Time-course analysis of C3a and C5a quantifies the coupling between the upper and terminal Complement pathways in vitro

Abstract

An in vitro zymosan-activation of the Complement system, through the lectin and alternative pathways, was performed in pooled human serum over a 24h time-course. Activation was quantitatively monitored by measuring the concentration of the upper Complement pathway fragment, C3a and the terminal pathway fragment, C5a. Upper Complement showed a maximum activation of 39% and the time-to-maximum activation reduced 8-fold, as a highly non-linear function of the zymosan dose. The C3a:C5a molar ratio rose to a maximum of 1100:1, before terminal pathway activation was initiated; indicating a flux threshold. This threshold appears to be exceeded once more than 31% of C3 molecules are activated. Above this threshold, significant activation of terminal pathway was observed; reducing the molar ratio to 17:1. The C5a/C3a molar ratio was used to determine the terminal pathway activation relative to total Complement activation and ranged from 0.1-0.8%. This depicts upper Complement activation to be 49-fold larger than terminal activation, a figure consistent with the observed density of the membrane attack complex in the membrane of cells. Our results thus indicate that the relative activity of opsonisation is ~50-fold greater than membrane attack complex formation, in vitro, in the pooled serum phenotype. The results suggest a potential clinical application, where an in vitro analysis of a patient on admission, or prior to a surgical procedure, would indicate their upper Complement activation capacity, with activation of C3 measured thereafter, or post-operatively. A patient with an exhausted upper Complement capacity may be vulnerable to infections and complications, such as sepsis.

Reduced order modeling and analysis of the human complement system

“…The complement model described the initiation of the alternative and lectin pathways, 244 and the downstream integration of the lectin and classical pathways ( Fig. 1 Morad et al [34] and is given in Table T2. In this study, we assumed zymosan A 281 differentially activated the lectin pathway, consistent with the study of Morad et al [34] 282 and several previous studies [35][36][37].…”

“…1 Morad et al [34] and is given in Table T2. In this study, we assumed zymosan A 281 differentially activated the lectin pathway, consistent with the study of Morad et al [34] 282 and several previous studies [35][36][37]. However, the role of zymosan may be more 283 complex, as it may activate all three complement pathways under certain conditions.…”

“…3). The data used for model validation was taken 307 from the study of Morad et al [34] and is given in Table T2. Six validation cases were 308 considered, three for C3a and C5a each, respectively.…”

Improvement in the blood compatibility of polyvinylidene fluoride membranes via in situ cross‐linking polymerization

“…Whether or not complement activation occurs after blood contact with a biomaterial also reflects whether the material has good blood compatibility. After complement activation, anaphylatoxins such as C3a and C5a are produced, and thus, the concentration of these anaphylatoxins can be detected in plasma after contact with a material, reflecting the degree of complement activation …”

“…After complement activation, anaphylatoxins such as C3a and C5a are produced, and thus, the concentration of these anaphylatoxins can be detected in plasma after contact with a material, reflecting the degree of complement activation. 49,50 The results of contact activation, including complement activation, are shown in Figure 5. As shown in Figure 5A, PF4 concentrations were no larger than the value for noncontacted plasma for all modified membranes, indicating that platelet activation did not occur after contact of the modified membranes with blood.…”