Phenolic excipients of insulin formulations induce cell death, pro-inflammatory signaling and MCP-1 release

Skin reactions at the infusion site are a common side effect of continuous subcutaneous insulin infusion therapy. We hypothesized that local skin complications are caused by components of commercial insulin formulations that contain phenol or m-cresol as excipients. The toxic potential of insulin solutions and the mechanisms leading to skin reactions were explored in cultured cells. The toxicity of insulin formulations (Apidra, Humalog, NovoRapid, Insuman), excipient-free insulin, phenol and m-cresol was investigated in L929 cells, human adipocytes and monocytic THP-1 cells. The cells were incubated with the test compounds dose- and time-dependently. Cell viability, kinase signaling pathways, monocyte activation and the release of pro-inflammatory cytokines were analyzed. Insulin formulations were cytotoxic in all cell-types and the pure excipients phenol and m-cresol were toxic to the same extent. P38 and JNK signaling pathways were activated by phenolic compounds, whereas AKT phosphorylation was attenuated. THP-1 cells incubated with sub-toxic levels of the test compounds showed increased expression of the activation markers CD54, CD11b and CD14 and secreted the chemokine MCP-1 indicating a pro-inflammatory response. Insulin solutions displayed cytotoxic and pro-inflammatory potential caused by phenol or m-cresol. We speculate that during insulin pump therapy phenol and m-cresol might induce cell death and inflammatory reactions at the infusion site in vivo. Inflammation is perpetuated by release of MCP-1 by activated monocytic cells leading to enhanced recruitment of inflammatory cells. To minimize acute skin complications caused by phenol/m-cresol accumulation, a frequent change of infusion sets and rotation of the infusion site is recommended.