Purpose: Topical application of eye drops for dry eye disease (DED) suffers from low bioavailability due to rapid drug elimination. Using lysozyme as a prototype, we determined that solid silica nanoparticles (SiNPs) are promising drug carriers that may facilitate sustained drug release. Current drug formulations are also ineffective or carry concerning side effects. Lacritin, a human tear glycoprotein, is downregulated in DED patients. As a mitogen that promotes basal tearing, it is a promising biologic. N-94 is a peptide fragment of lacritin that can be conjugated to SiNPs at physiological pH. While lacritin reduces human corneal epithelial cell (HCEC) death induced by pro-inflammatory cytokines, validation was required with N-94. This study assessed whether SiNPs can facilitate a gradual release of N-94 and whether released N-94 is cytoprotective against pro-inflammatory cytokines implicated in DED progression. 

Study Design: N-94 release kinetics from SiNPs was studied in triplicate. To verify biocompatibility and anti-inflammatory properties in vitro, the cellular metabolic activity and lactate dehydrogenase (LDH) release of primary HCEC cultures were assessed in response to independent treatment of N-94-SiNPs or co-treatment of released N-94 with pro-inflammatory cytokine insult. Three biological and technical replicates were performed. The data were analyzed via two-way ANOVAs and post-hoc tests.

Methods: Solid SiNPs were synthesized following the Stöber process. Fluorescently labelled N-94 was conjugated to SiNPs and its release over time was quantified via fluorescence spectrophotometry. MTT assays measured cellular metabolic activity and LDH assays measured cell death. HCECs were treated with 1–1000 nM of N-94-SiNPs for 16 h and 20 h to determine biocompatibility. We optimized an insult combination of IFNγ and TNFα that decreased cellular metabolic activity to 65–75% of that of the untreated control. Insulted cells were then co-treated with 1–1000 nM of released N-94 for 16 h and 20 h.

Results: SiNPs mediated the sustained release of N-94. N-94-SiNPs were generally non-toxic on HCECs and biocompatible in vitro. Certain doses of N-94 released from SiNPs were cytoprotective on HCECs by reducing the cytotoxic effects of pro-inflammatory cytokines. 

Conclusions: This study suggests that SiNPs can deliver N-94 gradually to HCECs and that the released N-94 has anti-inflammatory properties, reducing the harmful effects of the cytokines that mediate DED. Further development of this system may involve embedding the N-94-SiNPs into hydrogel contact lenses as secondary drug carriers. The bactericidal abilities of N-94 can reduce the risk of contact lens-related infections and the contacts would assist with sustained drug delivery.

Lacritin peptide (N-94)-conjugated solid silica nanoparticles as a novel drug delivery system for dry eye disease

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