# Roles of D-Amino Acids on the Bioactivity of Host Defense Peptides
## 5 Mechanisms of Bioactivities
### Immunological Effects of d-AA-Containing HDPs
#### 5.5.1 Immune Modulation  
HDPs have been demonstrated to be modulators of a variety of immune responses, such as inflammation and recruitment of dendritic cells, and they are notably able to affect the adaptive immune system by acting as peptide-based vaccine. Human defensins have been shown to stimulate the migration of dendritic cells to papilloma-associated pre-neoplastic epithelium [87] and could elicit an inflammatory leukocyte response, stimulate interferon \(\gamma\) production and significantly enhance the antibody response to tumor antigens [88]. Intratumoral injection with a lactoferrin-derived HDP contributed significantly to the infiltration of inflammatory cells into tumor xenografts and subsequently resulted in complete tumor ablation. Mice cured of the initial tumor graft exhibited strong resistance to subsequent tumor re-implantation attempts by the same tumor cells but not by others. Moreover, the resistance was transferable via spleen cell transplantation thereby demonstrating the potential of HDPs as vaccines [86].  
Although few studies have been carried out, d-AA-containing HDPs, both as all-d enantiomer and as diastereomers, have also been shown to be capable of modulating immune responses. [PERSON] et al. [47] demonstrated the antiinflammatory effect of an all-d protaetiamycine analogue. The d-enantiomer possessed potent selective bacterial lytic activity and could inhibit nitrogen oxide production (NO) and lipopolysaccharide-stimulated macrophage-mediated inflammation and cytokine production. Interestingly, although the direct antimicrobial activity was nearly the same for the d- and l-enantiomers, the antiinflammatory activity of the d-enantiomer was considerably higher than that of the l-enantiomer (Figure S4). Fluorescence analysis revealed that when compared to the l-enantiomer, the d-enantiomer had stronger binding to LPS, a finding that correlates well with the d-enantiomer antiinflammatory activity.  
A similar antiinflammatory effect has been observed by [PERSON] et al. [52] when testing a series of diastereomer AMPs. The diastereomers effectively inhibited LPS activation of macrophages, NO production and inducible NO synthase (iNOS) mRNA expression while at the same time maintaining selective bacterial lysis activity. Increases in the hydrophobicity and \(\alpha\)-helicity correlated positively with the antiinflammatory activity but not with the bacterial lytic activity. This observation hints at the challenges facing the design of multi-functional HDPs because previous studies had established that an increase in hydrophobicity and helicity was detrimental to the selective membrane disruption of HDPs [49; 53]. An interesting side note from the study of [PERSON] et al. [52] is that when testing the resistance of diastereomers against proteolytic enzymes, the authors found that a minimum d-AA content of 33% was required to provide the diastereomers with complete immunity against enzyme digestion.  
As of now, there seems to be no report of immune cell migration activation or vaccine-like properties of d-AA-containing HDPs in animals, neither as an enantiomer nor as a diastereomer. [PERSON] et al. [89] tested a number of temperin-related peptides and found that the tested peptides were unable to induce the chemotactic migration of human neutrophils and monocytes when the peptide sequence was reversed or when d-AAs were incorporated into the peptides to generate the all-d enantiomers. This finding indicates receptor-dependent activation of this type of immune response that is chirally dependent.