Hermetia illucens-Derived Chitosan: A Promising Immunomodulatory Agent for Applications in Biomedical Fields

Abstract

Chitosan, renowned for its important biological properties, is a valuable pharmaceutical excipient for different therapeutic approaches. Currently, the demand for the biopolymer on the market is growing, and, for this reason, it is important to biologically characterize the biopolymer produced from an alternative source to crustaceans, specifically the bioconverter insect Hermetia illucens. In this work, insect chitosan, yielded via heterogeneous and homogeneous deacetylation from larvae, pupal exuviae, and adults, was studied as an immunomodulatory agent. The inflammatory response of immortalized human keratinocyte cells was induced by Salmonella enterica subsp. enterica serovar Typhimurium lipopolysaccharide. After that, the ability of the biopolymer to reduce the expression of the pro-inflammatory cytokines IL-6, IL-8, IL-1α, and TNF-α was tested after 6 and 24 h of treatment. Insect chitosan samples effectively downregulated cytokine expression, with improved activity obtained from heterogeneous chitosan treatments.

Figure 1

qPCR showed the expression levels of pro-inflammatory cytokines IL-1-α (a), IL-8 (b), TNF-α (c), and IL-6 (d) on HaCat cells treated with LPS and H. illucens heterogeneous chitosan samples, both bleached and unbleached. Data are mean ± SD and are expressed as a percentage of the relative mRNAs compared to unstimulated control (ctrl), arbitrarily assigned as 100%. Significant differences are indicated by *p < 0,05, **p < 0,01, ***p < 0,001. Data were analyzed with two-way ANOVA and Bonferroni post-hoc test.

Figure 2

qPCR showed the expression levels of pro-inflammatory cytokines IL-1-α (a), IL-8 (b), TNF-α (c), and IL-6 (d) on HaCat cells treated with LPS and H. illucens homogeneous chitosan samples, both bleached and unbleached. Data are mean ± SD and are expressed as a percentage of the relative mRNAs compared to unstimulated control (ctrl), arbitrarily assigned as 100%. Significant differences are indicated by *p < 0,05, **p < 0,01, ***p < 0,001. Data were analyzed with two-way ANOVA and Bonferroni post-hoc test.

Figure 3

ELISA assay showed the concentration of pro-inflammatory cytokines IL-1-α (a), IL-8 (b), TNF-α (c), and IL-6 secreted in cell supernatants of HaCat cells treated with LPS and H. illucens heterogeneous chitosan samples, both bleached and unbleached. Data are expressed as pg/mL of protein concentration ± standard deviation in each group and are representative of three different experiments. Significant differences are indicated by *p < 0,05, **p < 0,01. Data were analyzed with two-way ANOVA and Bonferroni post-hoc test.

Figure 4

ELISA assay showed the concentration of pro-inflammatory cytokines IL-1-α (a), IL-8 (b), TNF-α (c), and IL-6 secreted in cell supernatants of HaCat cells treated with LPS and H. illucens homogeneous chitosan samples, both bleached and unbleached. Data are expressed as pg/mL of protein concentration ± standard deviation in each group and are representative of three different experiments. Significant differences are indicated by *p < 0,05, **p < 0,01, ***p < 0,001. Data were analyzed with two-way ANOVA and Bonferroni post-hoc test.