Alginate microsphere compositions dictate different mechanisms of complement activation with consequences for cytokine release and leukocyte activation

Abstract

The inflammatory potential of 12 types of alginate-based microspheres was assessed in a human whole blood model. The inflammatory potential could be categorized from low to high based on the four main alginate microsphere types; alginate microbeads, liquefied core poly-L-ornithine (PLO)-containing microcapsules, liquefied core poly-L-lysine (PLL)-containing microcapsules, and solid core PLLcontaining microcapsules. No complement or inflammatory cytokine activation was detected for the Ca/Ba alginate microbeads. Liquefied core PLO-and PLL-containing microcapsules induced significant fluid phase complement activation (TCC), but with low complement surface deposition (anti-C3c), and a low proinflammatory cytokine secretion, with exception of an elevated MCP-1(CCL2) secretion. The solid core PLL-containing microcapsules generated lower TCC but a marked complement surface deposition and significant induction of the proinflammatory cytokines interleukin (IL-1)β, TNF, IL-6, the chemokines IL-8 (CXCL8), and MIP-1α (CCL3) and MCP-1(CCL2).Inhibition with compstatin (C3 inhibitor) completely abolished complement surface deposition, leukocyte adhesion and the proinflammatory cytokines. The C5 inhibitions partly lead to a reduction of the proinflammatory cytokines. The leukocyte adhesion was abolished by inhibitory antibodies against CD18 and partly reduced by CD11b, but not by CD11c. Anti-CD18 significantly reduced the (IL-1)β, TNF, IL-6 and MIP-1α and anti-CD11b significantly reduced the IL-6 and VEGF secretion. MCP-1 was strongly activated by anti-CD18 and anti-CD11b. In conclusion the initial proinflammatory cytokine responses are driven by the microspheres potential to trigger complement C3 (C3b/iC3b) deposition, leukocyte activation and binding through complement receptor CR3 (CD11b/CD18).MCP-1 is one exception dependent on the fluid phase complement activation mediated through CR3.

Alginate microbeads are coagulation compatible, while alginate microcapsules activate coagulation secondary to complement or directly through FXII

“…We further demonstrated that the inflammatory cytokine responses were linked at the complement activation, and particularly the complement on the microcapsules surface [4, 5]. These patterns of activation have corresponded well with the in vivo findings in various mice models investigating Ca/Ba Beads and poly-L-lysine microcapsules as reviewed in [2].…”

“…The residual coagulation activation by inhibition at the C5 level by eculizumab could thus reflect the contribution from C3a, and further points to C5a as the most potent TF inductor. Since also the formation of C3b/iC3b, cell-adhesion and cytokines is prevented by compstatin [4], we cannot exclude also their involvement. The involvement of inflammatory cytokines on monocyte TF expression would however most likely be secondary to the effect of the rapidly produced complement activation products [3].…”

“…We have previously shown that the inflammatory cytokine release is dependent on the complement deposition of C3b/iC3b to the biomaterial surface, which promotes subsequent cell-adhesion involving complement receptor 3 [4]. This response can be prevented by complement inhibition at the level of complement protein C3 [5].…”

Iron oxide nanoparticles induce cytokine secretion in a complement-dependent manner in a human whole blood model

“…The C3 inhibitor compstatin analog Cp40 completely inhibited the cytokine response, whereas C5 inhibitor eculizumab only partly reduced the production of the inflammatory mediators, which is in agreement with previous findings. 26,27 While compstatin analog Cp40 inhibits both the formation of C3a and C5a, eculizumab only inhibited the formation of C5a, thus our data could be explained by a dual involvement of C3a and C5a for optimal cytokine response. In a study by Orning et al, 27 the differences between compstatin and eculizumab were also shown to be connected to the C3b/iC3b formation with subsequent cell-adhesion (by complement receptor CR3).…”

“…26,27 While compstatin analog Cp40 inhibits both the formation of C3a and C5a, eculizumab only inhibited the formation of C5a, thus our data could be explained by a dual involvement of C3a and C5a for optimal cytokine response. In a study by Orning et al, 27 the differences between compstatin and eculizumab were also shown to be connected to the C3b/iC3b formation with subsequent cell-adhesion (by complement receptor CR3). Although this study investigated non-phagocytable microcapsules and the effects of cell adhesion, it also illustrates the possibility for the involvement of other complement activating products such as C3b/iC3b.…”