Mucus production, host-microbiome interactions, hormone sensitivity, and innate immune responses modeled in human cervix chips

Abstract

Modulation of mucus production by the human ecto- and endo-cervical epithelium by steroid hormones and associated interactions with commensal microbiome play a central role in the physiology and pathophysiology of the female reproductive tract. However, most of our knowledge about these interactions is based on results from animal studies or in vitro models that fail to faithfully mimic the mucosal environment of the human cervix. Here we describe microfluidic organ-on-a-chip (Organ Chip) models of the human cervical mucosa that recreate the cervical epithelial-stromal interface with a functional epithelial barrier and produce abundant mucus that has compositional, biophysical, and hormone-responsive properties similar to the living cervix. Use of continuous fluid flow promoted ecto-cervical differentiation, whereas use of periodic flow including periods of stasis stimulated endo-cervical specialization. Similar results with minor differences were obtained using epithelial cells isolated from three donors each from a different ethnic background (African American, Hispanic, and Caucasian). When the endo-Cervix Chips were co-cultured with living Lactobacillus crispatus and Gardnerella vaginalis bacterial communities to respectively mimic the effects of human host interactions with optimal (healthy) or non-optimal (dysbiotic) microbiome, significant differences in tissue innate immune responses, barrier function, cell viability, and mucus composition were detected reminiscent of those observed in vivo. Thus, human Cervix Chips provide a physiologically relevant experimental in vitro model to study cervical mucus physiology and its role in human host-microbiome interactions as well as a potential preclinical testbed for development of therapeutic interventions to enhance women's health.

Modulation of dysbiotic vaginal complications by cervical mucus revealed in linked human vagina and cervix chips

“…We recently described a human Vagina Chip lined by primary human vaginal epithelium interfaced across an extracellular matrix (ECM)-coated porous membrane with underlying stromal fibroblasts cells that enables analysis of human host-microbiome interactions in the vaginal microenvironment, 9 as well as a human Cervix Chip containing a primary cervical epithelium interfaced with stromal cervical fibroblasts that produces cervical mucus with physical and chemical properties similar to those observed in vivo . 10 Here, we collected mucus-containing effluents from the outflow of the epithelial channel of the Cervix Chip (’cervical chip mucus’ containing 4.01 ± 3.04 mg/ml of mucus glycoproteins) for 7 days and then perfused it through the epithelial channel of a Vagina Chip to simulate the natural flow of mucus in the reproductive tract in vivo ( Fig. 1A ).…”

“…The first is a human vagina-on-a-chip (Vagina Chip) that is lined by primary, hormone-sensitive, vaginal epithelium interfaced with underlying stromal fibroblasts that we have shown recapitulates the pathophysiology of a dysbiotic vaginal epithelium when co-cultured with a G. vaginalis containing microbiome and that enables analysis of human host-microbiome interactions in vitro. 9 The second is a human Cervix Chip lined by primary cervical epithelium interfaced with cervical fibroblasts 10 that produces abundant cervical mucus with compositional, biophysical, and hormone-responsive properties similar to those observed in vivo . To model and study the effect of cervical mucus on vaginal responses in vitro, we co-cultured dysbiotic microbiome in the human Vagina Chip in the presence or absence of mucus-containing effluents that were transferred from the epithelial channel of the human Cervix Chips.…”

Advanced Technologies for Studying Microbiome–Female Reproductive Tract Interactions: Organoids, Organoids-on-a-Chip, and Beyond

“…98 One notable advancement was made by Izadifar et al, who developed a two-channel cervical chip lined with primary human cervical ECs. 18 This innovation, along with the human vaginal chip, has provided insights into mucus physiology and pathophysiology. 89 A simplified alternative, the mucus chip, has also emerged, offering insights into mucus penetration for drug delivery.…”

“…The ideal system ought to be able to model the complex interplay between mucus, mucins, and host-microbe interactions within the FRT, as this is essential for reproductive health and immunity. 18 However, it will depend on the researchers' goals which model will suit best. The review provides an overview of the available models and discusses what features can be integrated to create more representative in vitro models.…”

Breaking the clean room barrier: exploring low-cost alternatives for microfluidic devices

“…Labs-on-a-chip (LOCs), also known as microfluidic devices, have revolutionized biomedical and chemical analysis by providing efficient, portable, and highly miniaturized solutions ( Najjar et al, 2022 ; Izadifar et al, 2023 ; Zhao et al, 2023 ). The microfluidic design of these devices enables precise control of fluid flows and processes, leading to improved accuracy and repeatability of results ( Karthik et al, 2022 ; Verma & Pandya, 2022 ).…”