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The transcriptomic profile of primary cells isolated from intestinal muscle layers in 2D or 3D culture conditions and with different culture media

ABSTRACT: The intestinal muscle layers execute various gut wall movements to achieve controlled propulsion and mixing of intestinal content. Engineering intestinal muscle layers with complex contractile function is critical for developing bioartificial intestinal tissue to treat patients with short bowel syndrome. A living intestinal muscle patch is created for the first time in vitro to generate three distinct motility patterns and display multiple digesta manipulations. Developing such intestinal muscle patches combines a differentiation medium and a 3D scaffold. Next-generation RNA sequencing was performed across primary cells isolated from the intestinal muscle layers that were further cultured with/without the differentiation medium or with/without the 3D scaffold to characterize the effect of the medium and the scaffold on the transcriptomic profile of the cells. We found that the successful generation of the contracting intestinal muscle patch relies on both biological factors in the medium and environmental cues from the scaffold. In cooperation, the medium and the scaffold both greatly enhance the differentiation and maturity of the enteric neurons. Separately, the medium profoundly impacts ion transport activities. The scaffold markedly reshapes cell communication and protein binding.

ORGANISM(S): Mus musculus

PROVIDER: GSE223116 | GEO | 2023/02/15

REPOSITORIES: GEO

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Project description:Background & Aims: 3D Bioprinting of an endocrine pancreas is a promising future curative treatment for selected patients with insulin secretion deficiency. In this study we present an end-to-end integrative, scalable concept extending from the molecular to the macroscopic level. Methods: A hybrid scaffold device was manufactured by 3D (bio)printing. INS-1 cells with/without endothelial cells were bioprinted in gelatin methacrylate blend hydrogel. Polycaprolactone was 3D-printed and heparin-functionalized as structural scaffold component. In vitro evaluation was performed by viability and growth assays, total mRNA sequencing, and glucose-stimulated insulin secretion. In vivo, xenotransplantation to fertilized chicken eggs was used to investigate vascularization and function, and finite element analysis modeling served to detect boundary conditions and applicability for human islets of Langerhans. Results: Insulin-secreting pseudoislets were formed and resulted in a viable and proliferative experimental model. Transcriptomics revealed upregulation of proliferative and ß-cell-specific signaling cascades, downregulation of apoptotic pathways, and overexpression of extracellular matrix proteins and VEGF induced by pseudoislet formation and 3D culture. Co-culture with human endothelial cells created a natural cellular niche resulting in enhanced insulin response after glucose stimulation. Survival and function of the pseudoislets after explantation and extensive scaffold vascularization of both the hydrogel and heparinized polycaprolactone components were demonstrated in ovo. Computer simulations of oxygen, glucose, and insulin flows were used to evaluate scaffold architectures and Langerhans islets at a future transplantation site along neurovascular structures. Conclusion: A defined end-to-end process for multidisciplinary bioconvergence research on a bioartificial endocrine pancreas was developed. A modular, patient-specific device architecture is proposed for future research studies.

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The transcriptomic profile of primary cells isolated from intestinal muscle layers in 2D or 3D culture conditions and with different culture media

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