Date Published: September 20, 2017
Publisher: Springer International Publishing
Author(s): Matheus O. Costa, Roman Nosach, John C.S. Harding.
Most interactions between pathogenic microorganisms and their target host occur on mucosal surfaces of internal organs such as the intestine. In vitro organ culture (IVOC) provides an unique tool for studying host-pathogen interactions in a controlled environment. However, this technique requires a complex laboratory setup and specialized apparatus. In addition, issues arise when anaerobic pathogens are exposed to the hyperoxic environment required for intestinal culture. The objective of this study was to develop an accessible 3D–printed device that would allow manipulation of the gas mixture used to supply the tissue culture media separately from the gas mixture exposed to the mucosal side of explants.
Porcine colon explants from 2 pigs were prepared (n = 20) and cultured for 0h, 8h, 18h and 24h using the device. After the culture period, explants were fixed in formalin and H&E stained sections were evaluated for histological defects of the mucosa. At 8h, 66% of samples displayed no histological abnormalities, whereas samples collected at 18h and 24h displayed progressively increasing rates of superficial epithelial erosion and epithelial metaplasia.
The 3D–design reported here allows investigators to setup intestinal culture explants while manipulating the gas media explants are exposed to, to support tissue viability for a minimal of 8h. The amount of media necessary and tissue contamination are potential issues associated with this apparatus.
The online version of this article (10.1186/s41205-017-0018-z) contains supplementary material, which is available to authorized users.
The intestinal mucosa is an active host-microorganism interaction interface, and when innate and humoral immune defenses mechanisms fail, infection and disease result. Understanding virulence mechanisms of pathogenic microorganisms, and their consequences, provides an alternative target for disease prevention and mitigation . In vitro models provide advantages for studying virulence mechanisms, given the controlled environment and ease of sample collection. In vitro organ culture (IVOC) involves excision of a small volume of tissue from a healthy donor, and its subsequent culture in appropriate media in the laboratory for a number of days. IVOCs preserve important tissue characteristics, such as different cell types and tissue architecture, while providing a controlled and accessible environment to study host-pathogen interactions [2, 3]. To prevent acidification of the culture media and maintain tissue viability, intestinal IVOCs require a hyperoxic gas environment [4–6]. Sealed chambers are usually placed within incubators for culture, an expensive setup that requires extensive bench space and is potentially an explosion hazard due to the high oxygen content [6, 7]. In addition, this setup becomes a challenge when studying host interactions with anaerobic pathogens, due to the hyperoxic environment. Here, we describe and evaluate a 3D–printed device to allow the use of different gas mixtures in the culture media, compared to the mucosal aspect of the tissue.
This study describes a low-cost, 3D–printed device used for intestinal explant culture without the need of a hyperoxic chamber or incubator. 3D–printing required approximately 2 h including manual removal of leftover filament and printing debris. No leakage was observed after 24 h of continuous liquid media circulation through the circuit, and the media was maintained constantly at 37 °C.
The design described here consistently supported porcine colon explants for up to 8 h with minimal disruption of tissue viability. It also provides an alternative culture method that does not require the use of a hyperoxic chamber, and is particularly suited for situations where a gas environment other than atmospheric is needed on the mucosal aspect of the explant.