Ocular and systemic bio-distribution of rhodamine-conjugatedliposomes loaded with VIP injected into the vitreous of Lewis rats

[摘要] Purpose: Local delivery of therapeutic molecules encapsulated withinliposomes is a promising method to treat ocular inflammation. Thepurpose of the present study was to define the biodistribution ofrhodamine-conjugated liposomes loaded with vasoactive intestinal peptide(VIP), an immunosuppressive neuropeptide, following their intravitreal(IVT) injection in normal rats.Methods: Healthy seven- to eight-week-old Lewis male rats wereinjected into the vitreous with empty rhodamine-conjugated liposomes(Rh-Lip) or with VIP-loaded Rh-Lip (VIP-Rh-Lip; 50 mM of lipids with anencapsulation efficiency of 3.0±0.4 mmol VIP/mol lipids).Twenty-four h after IVT injection, the eyes, the cervical, mesenteric,and inguinal lymph nodes (LN), and spleen were collected. The phenotypeand distribution of cells internalizing Rh-Lip and VIP-Rh-Lip werestudied. Determination of VIP expression in ocular tissues and lymphoidorgans and interactions with T cells in cervical LN was performed onwhole mounted tissues and frozen tissue sections by immunofluorescenceand confocal microscopy.Results: In the eye, 24 h following IVT injection, fluorescentliposomes (Rh-Lip and VIP-Rh-Lip) were detected mainly in the posteriorsegment of the eye (vitreous, inner layer of the retina) and to a lesserextent at the level of the iris root and ciliary body. Liposomes wereinternalized by activated retinal Müller glial cells, ocular tissueresident macrophages, and rare infiltrating activated macrophages. Inaddition, fluorescent liposomes were found in the episclera andconjunctiva where free VIP expression was also detected. In lymphoidorgans, Rh-Lip and VIP-Rh-Lip were distributed almost exclusively in thecervical lymph nodes (LN) with only a few Rh-Lip-positive cells detectedin the spleen and mesenteric LN and none in the inguinal LN. In thecervical LN, Rh-Lip were internalized by resident ED3-positivemacrophages adjacent to CD4 and CD8-positive T lymphocytes. Some ofthese T lymphocytes in close contact with macrophages containingVIP-Rh-Lip expressed VIP.Conclusions: Liposomes are specifically internalized by retinalMüller glial cells and resident macrophages in the eye. A limitedpassage of fluorescent liposomes from the vitreous to the spleen via theconventional outflow pathway and the venous circulation was detected.The majority of fluorescent liposomes deposited in the conjunctivafollowing IVT injection reached the subcapsular sinus of the cervical LNvia conjuntival lymphatics. In the cervical LN, Rh-Lip were internalizedby resident subcapsular sinus macrophages adjacent to T lymphocytes.Detection of VIP in both macrophages and T cells in cervical LN suggeststhat IVT injection of VIP-Rh-Lip may increase ocular immune privilege bymodulating the loco-regional immune environment. In conclusion, ourobservations suggest that IVT injection of VIP-loaded liposomes is apromising therapeutic strategy to dampen ocular inflammation bymodulating macrophage and T cell activation mainly in the loco-regionalimmune system.

[学科分类] 生物化学/生物物理