Protein-Coated Nanoparticles Are Internalized by the Epithelial Cells of the Female Reproductive Tract and Induce Systemic and Mucosal Immune Responses

Abstract

The female reproductive tract (FRT) includes the oviducts (fallopian tubes), uterus, cervix and vagina. A layer of columnar epithelium separates the endocervix and uterus from the outside environment, while the vagina is lined with stratified squamous epithelium. The mucosa of the FRT is exposed to antigens originating from microflora, and occasionally from infectious microorganisms. Whether epithelial cells (ECs) of the FRT take up (sample) the lumen antigens is not known. To address this question, we examined the uptake of 20–40 nm nanoparticles (NPs) applied vaginally to mice which were not treated with hormones, epithelial disruptors, or adjuvants. We found that 20 and 40 nm NPs are quickly internalized by ECs of the upper FRT and within one hour could be observed in the lymphatic ducts that drain the FRT, as well as in the ileac lymph nodes (ILNs) and the mesenteric lymph nodes (MLNs). Chicken ovalbumin (Ova) conjugated to 20 nm NPs (NP-Ova) when administered vaginally reaches the internal milieu in an immunologically relevant form; thus vaginal immunization of mice with NP-Ova induces systemic IgG to Ova antigen. Most importantly, vaginal immunization primes the intestinal mucosa for secretion of sIgA. Sub-cutaneous (s.c) boosting immunization with Ova in complete Freund's adjuvant (CFA) further elevates the systemic (IgG1 and IgG2c) as well as mucosal (IgG1 and sIgA) antibody titers. These findings suggest that the modes of antigen uptake at mucosal surfaces and pathways of antigen transport are more complex than previously appreciated.

Chitosan enhances nanoparticle delivery from the reproductive tract to target draining lymphoid organs

“…In a separate study by Howe et al, PS NPs (≤40 nm) were imaged in the ILLN by microscopy after vaginal dosing but the robustness of the delivery was not quantified or discussed. 4 Here, we quantify for the first time that only a low frequency of about 1 in 4 nodes (25%) accumulates 20 nm- or 200 nm-sized NP that are dosed vaginally. In this case, we also observed only limited NP-cell interactions in the vaginal tissue and the dLNs, which may support evidence of NP transport by the paracellular route.…”

“…However, traversing this multilayered squamous epithelium to reach the iliac (ILLNs) and inguinal lymph nodes (IGLNs) that drain the lower FRT can be a formidable obstacle for large macromolecules and particulate antigens. 1, 2 While ultrasmall nanoparticles (NPs) (≤40 nm) may appear in the dLNs following intravaginal (IVG) administration, 3, 4 the robustness of NP delivery to the draining lymph nodes (dLNs) from the vaginal routes with respect to reproducibility and frequency of positive LNs in animal treatment groups has not been reported. As such, questions still remain about mechanisms of NP delivery across the vaginal epithelium to the dLNs.…”

Immunogenicity of antigen‐conjugated biodegradable polydiacetylene liposomes administered mucosally

“…NPs larger than 200 nm have been mostly used for antigen and drug delivery because they can carry a large amount of cargo. In regard to antigen delivery, studies showing superior immunogenicity of both large and small NPs have been published, however, due to the varying properties of NPs used in these studies (such as NP chemistry, charge, antigen encapsulation, NP rigidity, and so forth), it is difficult to draw a definitive conclusion that NP size alone determines their immunogenicity. Moreover, NPs have been used in various prime‐boost immunization schemes, and are administered with (or without) adjuvants, thus additionally affecting their immunogenicity and making comparisons between studies even more challenging.…”

“…It is known, however, that NPs are internalized more efficiently by cultured cells compared to microparticles . NPs < 50 nm penetrate mucus covering and are internalized at mucosal surfaces more efficiently than larger NPs . Additionally, dendritic cells (DC) internalize NP‐associated antigens more efficiently than soluble antigens .…”

“…Ova‐conjugated NPs (∼40 nm) administered intradermally induce superior cellular and humoral immune responses compared to larger particles . Using Ova‐conjugated polystyrene NPs (NP‐Ova), it was shown that s.c. NP‐Ova administration without adjuvants induces high serum IgG1 antibody titers, while mucosal NP‐Ova administration induces mixed serum IgG1/IgG2c titers and primes the intestinal mucosa for secretion of IgA . Most importantly, mucosal priming followed by s.c. boosting induced superior systemic and mucosal memory compared to p.o.…”