Topical cathelicidin (LL‐37) an innate immune peptide induces acute olfactory epithelium inflammation in a mouse model

Abstract

Background Cathelicidin (LL-37) is an endogenous innate immune peptide that is elevated in patients with chronic rhinosinusitis (CRS). The role of LL-37 in olfactory epithelium (OE) inflammation remains unknown. We hypothesized that 1) LL-37 topically delivered would elicit profound OE inflammation, and 2) LL-37 induced inflammation is associated with increased infiltration of neutrophils and mast cells. Methods To test our hypothesis we challenged C57BL/6 mice intranasally with increasing concentrations of LL-37. At 24 hours tissues were examined histologically and scored for inflammatory cell infiltrate, edema, and secretory hyperplasia. In separate experiments, fluorescently conjugated LL-37 was instilled and tissues were examined at 0.5 and 24 hours. To test our last hypothesis, we performed tissue myeloperoxidase (MPO) assays for neutrophil activity and immunohistochemistry for tryptase to determine the mean number of mast cells per mm2. Results LL-37 caused increased inflammatory cell infiltrate, edema, and secretory cell hyperplasia of the sinonasal mucosa with higher LL-37 concentrations yielding significantly more inflammatory changes (p < 0.01). Fluorescent LL-37 demonstrated global sinonasal epithelial binding and tissue distribution. Further, higher concentrations of LL-37 led to significantly greater MPO levels with dose-dependent increases in mast cell infiltration (p < 0.01). Conclusions LL-37 has dramatic inflammatory effects in the OE mucosa that is dose-dependent. The observed inflammatory changes in the olfactory mucosa were associated with the infiltration of both neutrophils and mast cells. Our biologic model represents a new model to further investigate the role of LL-37 in OE inflammation.

Prevention of sinonasal inflammation by a synthetic glycosaminoglycan

“…Mice were maintained supine during and following inoculations to facilitate retention in the nasal cavity. [32,38] The animals were observed for 24 h for pain or discomfort and subsequently sacrificed 24 h post-administration for tissue processing and fluorescence imaging using a Nikon A1R confocal microscope.…”

“…GM-0111 or HA was then instilled at a dose of 800 μg (10 mg/mL, 40 μL per nare over 40 min, 10 μL for each 10 min interval), immediately followed by a single dose of 320 μM LL-37 (40 μL per nare over 40 min, 10 μL for each 10 min interval, 115 μg total dose). [32] The animals were subsequently sacrificed 24 h post-instillation of LL-37, and sinonasal tissues were processed for histological, immunohistochemical (IHC), and biochemical analyses.…”

“…Briefly, heads were bisected in the sagittal plane at the nasal septum, and sinonasal tissues were processed for histological or biochemical as previously reported. [32]…”

“…RS is an inflammatory respiratory disease that affects up to 16% of the United States population and is in dire need of new effective therapies. [23–31] We recently reported that intranasal administration of the inflammatory modulating peptide LL-37 causes profound sinonasal mucosal inflammation in the mouse with similar clinical signs to human RS,[32] as LL-37 causes cell death in the airways [33] and is elevated in sinus epithelial cells [34] and the nasal mucosa cultures of patients with RS. [35,36] Employing this model and based on prior evidence, we hypothesized that GM-0111 would (1) effectively coat and penetrate the sinuses through topical, intranasal delivery and (2) prevent sinonasal inflammation by protecting the sinonasal epithelium against inflammatory cell infiltration and cell death.…”

“…[35,36] Employing this model and based on prior evidence, we hypothesized that GM-0111 would (1) effectively coat and penetrate the sinuses through topical, intranasal delivery and (2) prevent sinonasal inflammation by protecting the sinonasal epithelium against inflammatory cell infiltration and cell death. [21,32]. …”