Langerhans cells shape postnatal oral homeostasis in a mechanical-force-dependent but microbiota and IL17-independent manner.

The postnatal interaction between microbiota and the immune system establishes lifelong homeostasis at mucosal epithelial barriers, however, the barrier-specific physiological activities that drive the equilibrium are hardly known. During weaning, the oral epithelium, which is monitored by Langerhans cells (LC), is challenged by the development of a microbial plaque and the initiation of masticatory forces capable of damaging the epithelium. Here we show that microbial colonization following birth facilitates the differentiation of oral LCs, setting the stage for the weaning period, in which adaptive immunity develops. Despite the presence of the challenging microbial plaque, LCs mainly respond to masticatory mechanical forces, inducing adaptive immunity, to maintain epithelial integrity that is also associated with naturally occurring alveolar bone loss. Mechanistically, masticatory forces induce the migration of LCs to the lymph nodes, and in return, LCs support the development of immunity to maintain epithelial integrity in a microbiota-independent manner. Unlike in adult life, this bone loss is IL-17-independent, suggesting that the establishment of oral mucosal homeostasis after birth and its maintenance in adult life involve distinct mechanisms.

Topical phage therapy in a mouse model of Cutibacterium acnes-induced acne-like lesions.

Acne vulgaris is a common neutrophil-driven inflammatory skin disorder in which Cutibacterium acnes (C. acnes) is known to play a key role. For decades, antibiotics have been widely employed to treat acne vulgaris, inevitably resulting in increased bacterial antibiotic resistance. Phage therapy is a promising strategy to combat the growing challenge of antibiotic-resistant bacteria, utilizing viruses that specifically lyse bacteria. Herein, we explore the feasibility of phage therapy against C. acnes. Eight novel phages, isolated in our laboratory, and commonly used antibiotics eradicate 100% of clinically isolated C. acnes strains. Topical phage therapy in a C. acnes-induced acne-like lesions mouse model affords significantly superior clinical and histological scores. Moreover, the decrease in inflammatory response was reflected by the reduced expression of chemokine CXCL2, neutrophil infiltration, and other inflammatory cytokines when compared with the infected-untreated group. Overall, these findings indicate the potential of phage therapy for acne vulgaris as an additional tool to conventional antibiotics.

Microbiota-dependent and -independent postnatal development of salivary immunity.

While saliva regulates the interplay between the microbiota and the oral immune system, the mechanisms establishing postnatal salivary immunity are ill-defined. Here, we show that high levels of neutrophils and neonatal Fc receptor (FcRn)-transferred maternal IgG are temporarily present in the neonatal murine salivary glands in a microbiota-independent manner. During weaning, neutrophils, FcRn, and IgG decrease in the salivary glands, while the polymeric immunoglobulin receptor (pIgR) is upregulated in a growth arrest-specific 6 (GAS6)-dependent manner independent of the microbiota. Production of salivary IgA begins following weaning and relies on CD4-help, IL-17, and the microbiota. The weaning phase is characterized by a transient accumulation of dendritic cells capable of migrating from the oral mucosa to the salivary glands upon exposure to microbial challenges and activating T cells. This study reveals the postnatal mechanisms developed in the salivary glands to induce immunity and proposes the salivary glands as an immune inductive site.

Protocol for parotidectomy and saliva analysis in mice.

The murine parotid salivary glands develop postnatally, shaping oral mucosal immunity in early and adult life. This protocol details the surgical removal of the parotid glands (parotidectomy) of mice. We also describe a protocol for saliva collection to enable manipulation and measurement of physiological and immunological salivary functions. Our saliva collection approach has been modified from published protocols to enable saliva collection from young mice, which can be challenging. For complete details on the use and execution of this protocol, please refer to Koren et al. (2020).

Shaped by the epithelium - postnatal immune mechanisms of oral homeostasis.

The first encounter of mucosal barriers with the microbiota initiates host-microbiota feedback loops instructing the tailored development of both the immune system and microbiota at each mucosal site. Once established, balanced immunological interactions enable symbiotic relationships with the microbiota in adult life. This process has been extensively investigated in the mammalian monolayer epithelium-covered intestine and lung mucosae; however, the postnatal mechanisms engaged by the oral mucosa to establish homeostasis are currently being discovered. Here, we discuss the early life dialogue between the oral mucosa and the microbiota, with particular emphasis on the central role the multilayer epithelium plays to protect the oral mucosa. These intricate and unique postnatal immunological processes shape oral homeostasis, which can potentially affect buccal and systemic health in adult life.

Maturation of the neonatal oral mucosa involves unique epithelium-microbiota interactions.

Postnatal host-microbiota interplay governs mucosal homeostasis and is considered to have life-long health consequences. The intestine monolayer epithelium is critically involved in such early-life processes; nevertheless, the role of the oral multilayer epithelium remains ill defined. We demonstrate that unlike the intestine, the neonate oral cavity is immensely colonized by the microbiota that decline to adult levels during weaning. Neutrophils are present in the oral epithelium prenatally, and exposure to the microbiota postnatally further recruits them to the preamble neonatal epithelium by γδT17 cells. These neutrophils virtually disappear during weaning as the epithelium seals. The neonate and adult epithelium display distinct turnover kinetics and transcriptomic signatures, with neonate epithelium reminiscent of the signature found in germ-free mice. Microbial reduction during weaning is mediated by the upregulation of saliva production and induction of salivary antimicrobial components by the microbiota. Collectively, unique postnatal interactions between the multilayer epithelium and microbiota shape oral homeostasis.

Niche rather than origin dysregulates mucosal Langerhans cells development in aged mice.

Unlike epidermal Langerhans cells (LCs) that originate from embryonic precursors and are self-renewed locally, mucosal LCs arise and are replaced by circulating bone marrow (BM) precursors throughout life. While the unique lifecycle of epidermal LCs is associated with an age-dependent decrease in their numbers, whether and how aging has an impact on mucosal LCs remains unclear. Focusing on gingival LCs we found that mucosal LCs are reduced with age but exhibit altered morphology with that observed in aged epidermal LCs. The reduction of gingival but not epidermal LCs in aged mice was microbiota-dependent; nevertheless, the impact of the microbiota on gingival LCs was indirect. We next compared the ability of young and aged BM precursors to differentiate to mucosal LCs. Mixed BM chimeras, as well as differentiation cultures, demonstrated that aged BM has intact if not superior capacity to differentiate into LCs than young BM. This was in line with the higher percentages of mucosal LC precursors, pre-DCs, and monocytes, detected in aged BM. These findings suggest that while aging is associated with reduced LC numbers, the niche rather than the origin controls this process in mucosal barriers.