Transfer of IgG in the female genital tract by MHC class I-related neonatal Fc receptor (FcRn) confers protective immunity to vaginal infection Zili Lia,b, Senthilkumar Palaniyandia,b, Rongyu Zenga,b, Wenbin Tuoc, Derry C. Roopeniand, and Xiaoping Zhua,b,1 aLaboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, College Park, MD 20742; bMaryland Pathogen Research Institute, University of Maryland, College Park, MD 20742; cAnimal Parasitic Diseases Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, MD 20705; and dThe Jackson Laboratory, Bar Harbor, ME 04609 Edited by Roy Curtiss, Arizona State University, Tempe, AZ, and approved February 1, 2011 (received for review August 30, 2010) IgG is a major Ig subclass in mucosal secretions of the human and viral load. Therefore, HIV-specific IgG may be much more female genital tract, where it predominates over the IgA isotype. important in mucosal protection than previously thought. Unlike Despite the abundance of IgG, surprisingly little is known about S-IgA, the mechanism(s) by which the IgG antibody is transported where and how IgG enters the lumen of the genital tract and the across the genital epithelium and the role of IgG in genital mu- exact role local IgG plays in preventing sexually transmitted dis- cosal protection have not been investigated. Incomplete un- eases. We demonstrate here that the neonatal Fc receptor, FcRn, is derstanding of IgG transport in the genital tract and of its role in expressed in female genital tract epithelial cells of humans and mice combating genital infections has hampered the design and de- and binds IgG in a pH-dependent manner. In vitro we show that velopment of vaccines and preventative treatments for sexually FcRn mediates bidirectional IgG transport across polarized human transmitted diseases (STDs). Historically, the source of IgG in the endometrial HEC-1-A monolayers and primary human genital epi- genital tract has been attributed to simple passive paracellular thelial cells. Furthermore, endosomal acidification appears to be diffusion from the circulation or local production by epithelium- a prerequisite for FcRn-mediated IgG transcytosis; IgG transcytosis associated plasma cells (12). This view has been challenged by was demonstrated in vivo by translocation of systemically admin- increasing evidence that IgG levels in genital mucosal secretions istered IgG into the genital lumen in WT but not FcRn-KO mice. The can be affected dramatically by genital infections (6, 13–16), the MEDICAL SCIENCES biological relevance of FcRn-transported IgG was demonstrated by estrous cycle (17), and immunization (7, 8, 15, 18). Collectively, – fi passive immunization using herpes simplex virus-2 (HSV-2) speci c these observations raise the possibility of an active transport sys- fi polyclonal serum, which conferred signi cantly higher protection tem through which IgG crosses the genital epithelium. against intravaginal challenge infection by the HSV-2 186 strain in The neonatal Fc receptor for IgG (FcRn) is a heterodimer WT mice than in FcRn-KO mice. These studies demonstrate that composed of a membrane-bound 45- to 50-kDa heavy chain as- FcRn-mediated transport is a mechanism by which IgG can act lo- sociated nonconvalently with the 12-kDa β2-microglobulin (β2m) cally in the female genital tract in immune surveillance and in host (19). FcRn was identified originally in the intestine of neonatal defense against sexually transmitted diseases. rodents, but it also is expressed and is functionally active in a variety of adult tissues and cells (20–22). Furthermore, FcRn is mucosa | humoral | uterine responsible for transporting maternal IgG across the placenta and colostrum IgG across the intestine to the fetal and neonatal ost pathogens initiate infection by making contact with bloodstream, respectively (20, 21). The ability of FcRn to Mpolarized epithelial cells at mucosal surfaces. Immuno- transport IgG allows newborns to acquire humoral immunity in globulins present in mucosal secretions function as a first line of the form of maternal antibodies before their own immune system defense against mucosally transmitted pathogens (1). In the is fully matured. In addition, FcRn is capable of protecting IgG gastrointestinal and upper respiratory tracts, the major Ig species and albumin from catabolism (22, 23), and this protection con- found in the mucosal surface is secretory IgA (S-IgA), which is tributes to their being the most abundant proteins in the blood. known to result primarily from active transport of dimeric IgA by This protective function of FcRn plays an important role in the polymeric IgA receptor (pIgR) through polarized epithelium prolonging the half-life and maintaining functional levels of IgG (2). Although S-IgA generally is considered the primary com- after immunization or infection and therefore promotes long- ponent of antibody-mediated defense in the intestine, several lasting protective immunity. The interaction of FcRn and IgG is lines of evidence indicate that IgG also is present in mucosal markedly pH dependent, with binding of IgG at acidic (≤6.5) and secretions and support a role for IgG in host mucosal defense. release of IgG at neutral (7.0–7.5) conditions (23). As such, FcRn is First, IgG is present in secretions of the human oral mucosa, likely to function primarily within the acidified endosomes where small and large intestines, and lungs (3, 4). Second, the numbers IgG binding likely occurs after fluid phase uptake. of local IgG-secreting cells in mucosal tissues and the levels of Although FcRn plays critical roles in the acquisition of ma- fi antigen-speci c IgG in mucosal secretions are dramatically in- ternal humoral immunity in early life and in the regulation of – creased after both mucosal and systemic immunization (5 7). IgG catabolism in adults, its function in IgG-mediated immunity Third, the isotype pattern and concentrations of IgG in mucosal in the female genital tract remains to be established. Because secretions are clearly distinct from those of serum, suggesting an active and selective representation of IgG in the mucosal site (8). IgG is the predominant Ig subclass in the human female genital Author contributions: Z.L., W.T., and X.Z. designed research; Z.L., S.P., R.Z., and X.Z. tract, where its concentration exceeds that of IgA (9). IgG also has performed research; D.C.R. contributed new reagents/analytic tools; Z.L., W.T., D.C.R., been detected in uterine-cervical fluids (10) and vaginal washes and X.Z. analyzed data; and Z.L., W.T., D.C.R., and X.Z. wrote the paper. (5, 11). The importance of IgG in genital infections has been The authors declare no conflict of interest. exemplified by the predominance of human immunodeficiency This article is a PNAS Direct Submission. virus (HIV-1)–specific IgG responses at the mucosal surfaces of 1To whom correspondence should be addressed. E-mail: [email protected]. – HIV-1 infected women (6, 11). In these studies, there was an This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. inverse correlation between the amount of mucosal IgG present 1073/pnas.1012861108/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1012861108 PNAS Early Edition | 1of6 Downloaded by guest on September 27, 2021 IgG is the major Ig isotype in the genital tract, we hypothesized epithelial cell line HeLa was used as a negative control (25). The that FcRn in polarized genital epithelial cells functions as an 45-kDa band detected in all four genital epithelial cell lines intracellular trafficking receptor and mediates transcytosis of (Fig. 1B) was consistent with the known molecular weight of the IgG across genital tract epithelium, leading to enriched IgG in hFcRn heavy chain and comigrated with FcRn heavy chain ex- genital mucosal secretions. Our findings show that FcRn in the pressed by Caco-2 cells (Fig. 1B, lane 6). female genital epithelial of both humans and mice is capable The expression of FcRn in human uterine and vaginal tissues of transporting IgG both apically and basolaterally. Moreover, was confirmed further by immune staining using anti-hFcRn FcRn-transcytosed viral-specific IgG conferred protection against antibodies (25). FcRn was localized to both uterine and vaginal mucosal challenge with the model pathogen HSV-2. These results tissues, with the most intense staining in the epithelium (Fig. reveal an unexpected and important role for FcRn acting at the 1C). Immune staining in the stroma was weak in the uterus and female genital epithelium in immune surveillance and host de- almost undetectable in the vagina. Many scattered cells in the fense against STDs. uterine lamina propria also showed weak cytoplasmic staining (Fig. 1C). No staining was seen using an irrelevant rabbit IgG Results (Fig. 1C), illustrating the specificity of the anti-hFcRn anti- Analysis of FcRn Expression in Human Female Genital Epithelial Cells. bodies. Together these results show that human vaginal, ecto- To determine if FcRn is expressed in female genital epithelial cervical, endocervical, and endometrial epithelia express the 45- cells, RT-PCR analysis was performed on total RNA obtained kDa heavy chain of FcRn. The presence of abundant FcRn from four human female genital epithelial cell lines (End1, mRNA and protein in human female genital epithelial cells and endocervical; Ect1, exocervical; HEC-1-A, uterine; and VK2, its observed cellular distribution are consistent with data from vaginal) and two control human cell lines (the intestinal epi- enterocytes of human small intestine (20). thelial cell line HT29 and macrophage-like cell line THP-1). Human FcRn cDNA was amplified as a single 296-bp band in all Receptor-Mediated Transcytosis of IgG in HEC-1-A Cells. We selected cell lines examined, and the sequences of the PCR products were the HEC-1-A cell to study IgG transcytosis because it expresses confirmed by DNA sequencing (Fig.