Development of novel approaches for an in vitro colon model

Abstract

Gastrointestinal conditions and diseases have a significant impact on people, and in the past couple of decades, the number of cases related to these conditions have increased steadily. Due to the heterogeneity of causes and genetic factors involved, there are no therapies for some of these ailments which, in some cases, last a lifetime. In vitro models are a valuable tool and a great starting point in the quest for better therapeutics and exploring the cellular mechanisms and physiology in a controlled and simplistic fashion. My dissertations focuses on developing novel approaches to create 3D in vitro colon crypt models.Firstly, we have developed a novel bio printing technique based on the traditional lithographic technique of screen printing to pattern layers of mammalian cells with a suitable hydrogel scaffold. Screen printing is an accessible system that can be easily incorporated into the cell culture setup in an aseptic way. This dissertations presents a detailed description of the screen printing process along with its merits and limitations. To complement the screen printing setup, we have developed four libraries of hydrogels both degradable and non-degradable to be used as bioinks in these systems. These hydrogel systems are based on poly(ethylene glycol) (PEG), poly-L-lysine (PLL), and poly-allylamine (PAA), using the vinyl sulfone- thiol click chemistry and succinimidyl amine chemistry. These hydrogels systems have shown some interesting properties like cord formation in endothelial cells, along with several in vitro applications apart from their use as bioinks in the screen printing process. These synthetic hydrogels have a huge potential to be tuned in terms of biochemical cues using various extracellular matrix proteins, growth factors, adhesive motifs, etc., and also in terms of mechanical properties and degradation rates to be tailored to suit the needs of the cells. The synthesis, characterization, properties, and applications of these hydrogel libraries are investigated in this dissertations. Finally, we developed a non-degradable PEG diacrylate based crypt shaped scaffold to be used as in vitro colon crypt model. This scaffold presents a proof of concept of a novel in vitro crypt model which can accommodate the intestinal epithelial cells (like the Caco-2 and HT29) as well as the immune cells (macrophages) acting as a closer replicate of the native tissue environment. Overall, this thesis demonstrates a novel bioprinting technique based on screen printing, libraries of synthetic hydrogels, a crypt shaped scaffold, as accessible tools to develop 3D in vitro models.