Novel anti-inflammatory liposomal formulation for the pre-ocular tear film: In vitro and ex vivo functionality studies in corneal epithelial cells

Abstract

In ocular surface inflammatory diseases, such as dry eye disease, long-term symptom relief requires targeting the inflammation itself rather than treating only the surface-associated dryness with artificial tears. Therefore, we included an anti-inflammatory agent in an unpreserved liposome-based (LP) formulation used as artificial tears. Our aim was to characterize and study its in vitro and ex vivo cell uptake and functionality. Human corneal epithelial (HCE) cells were used to study MPA-LP-induced effects after 60 min of exposure, using blank LP and non-LP MPA formulations as controls. A fluorescent labeled LP formulation was used to determine uptake by HCE cells and localization in ex vivo porcine corneas. The LP formulation complied with the required physicochemical properties and had no cytotoxicity on HCE cells after 60 min of exposure. HCE cells showed LP-associated fluorescence at 24, 48, and 72 h after 60 min of exposure, and the LP-associated fluorescence was uniformly distributed throughout the porcine corneal epithelium immediately after 5 min of exposure. MPA-LP increased protein expression and nuclear translocation of progesterone receptor in comparison with controls as determined by Western blotting and immunofluorescence. Moreover, MPA-LP significantly reduced the cell proliferation rate and IL-6 and IL-8 production 48 h after the exposure period, as determined by the alamarBlue assay and ELISA, respectively. None of these effects were evident in blank LP-exposed cells and non-LP MPA formulation reduced only IL-6 production. Our results suggest that the LP-based formulation, used to replenish the lipids of the tear film, can be loaded with anti-inflammatory agents that can be delivered into the cells and activate specific drug receptors. These agents can reduce inflammatory cytokine production and may be effective in the treatment of inflammatory processes associated with ocular surface diseases.

Appraising the Competency of Lipid Nano-Platforms as Non-invasive Paradigms for the Treatment of Ocular Inflammations: A review

“…This blank liposome was considered effective in supplement tear lipids and hence indicated as artificial tears for dry eyes. The assimilation of an antiinflammatory agent as Medroxyprogesterone acetate in the same liposomal platform was found to improve the ocular inflammatory condition by reducing cytokine production, demonstrating better results when compared to the nonliposomal drug formulation [29].…”

“…5. The cell proliferation assay was employed to determine the possible changes in proliferation rate as a tool confirming the ocular tolerability of the tested formulae [29].…”

“…The uptake of the developed nanoformulations, particularly nanoparticles, into corneal epithelial cells was studied by incubating the tested particles labeled with a fluorescent dye for a certain exposure period. After careful cell washing with phosphate buffer saline (PBS), the intensity of fluorescence is measured using fluorometric analysis or visualized by fluorescence microscopy [29].…”

Formulation Considerations for the Management of Dry Eye Disease

“…The phospholipid component of liposomes may also have a role in stabilizing the lipid layer of the tear film, as discussed above, rendering them particularly advantageous for DED therapy. Hence, liposomes have been investigated for ocular delivery of several DED drugs, such as CsA [110], medroxyprogesterone acetate [111] and sirolimus [112]. However, in practice, liposomes have poor stability on the corneal surface and provide only moderate improvement in bioavailability over traditional ophthalmic eyedrops [113].…”

Liposomes as vehicles for topical ophthalmic drug delivery and ocular surface protection

“…According to some studies SUVs present the highest permeation ratio through the corneal epithelial barrier while MLVs the lowest [14]. Besides, some studies with the lipophile fluorophore cumarin-6 have demonstrated that liposomes around 190-200 nm pass through every corneal epithelial layer and are able to reach the stroma [111]. Regarding topical ophthalmic administration, liposomes close to 200 nm are normally desired to deliver drugs to the ocular surface [112].…”