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INNOVATIVE APPROACHES FOR 3D CELL CULTURES IN BIOTECHNOLOGY

Overview

Studying a biological phenomenon requires selecting an appropriate experimental model, and in vitro cell culture systems have proven to be highly useful over the years for various types of experiments. In recent years, also with the goal of reducing animal use in research, several 3D culture systems have been developed that increasingly replicate the physiology of living organisms.

Aims and Content

Learning Outcomes

The aim of this course is to offer an overview of the main aspects of three-dimensional models of cell cultures, from spheroids to organoids and 3D-printing.

Aims and Learning Outcomes

Active participation in the course and practical activities will enable the student to:

• Describe the dynamic reciprocity between cells and the extracellular matrix in determining cell fate, and compare 2D and 3D cultures in terms of environmental signaling.
• Distinguish between spheroids, organoids, hydrogels, and rigid scaffolds, explaining their advantages, limitations, and main applications in biological and biotechnological research.
• Set up three-dimensional cell cultures under static conditions in the laboratory using hydrogels, scaffolds, and systems for spheroid formation.
• Describe the principles of dynamic cultures and illustrate the functioning of bioreactors as tools to simulate more complex physiological microenvironments.
• Observe and understand the operation of a 3D bioprinter for biological applications, and take part in a demonstration of acellular printing.
• Understand the potential of microfluidic devices and organ-on-chip systems, even if not directly used, as advanced tools for physiological modeling.
• Critically analyze current scientific literature on 3D cell culture systems.

Prerequisites

Students will be required to have a basic understanding of chemistry, biochemistry, cell biology, and 2D cell cultures.

Teaching Methods

The course includes 24 hours of lectures and 16 hours of practical laboratory activities. The theoretical lessons aim to provide the conceptual foundations of three-dimensional cell culture models and the related technologies. The practical sessions allow students to apply the acquired knowledge by setting up 3D cell cultures on various supports (hydrogels, scaffolds, spheroids), performing microscopic observations, and attending demonstrations involving 3D printing tools and bioreactors.

Syllabus/Content

Lectures:

• The "dynamic reciprocity" between cells and the extracellular matrix in determining cell fate
• Modulation of microenvironmental signals (cell adhesion, mechanical forces, soluble factors): comparison between 2D and 3D cultures
• 2D vs. 3D cultures: main differences, advantages, and limitations
• Definition of spheroids and organoids: characteristics, applications, and challenges
• Use of organoids to study innervation and vascularization processes
• Biomaterials for 3D cultures: physicochemical and structural properties of hydrogels and rigid scaffolds
• 3D printing technologies applied to the generation of tissue-specific models
• Bioreactors for dynamic cultures: operating principles and experimental advantages
• Overview of microfluidic devices and organ-on-chip systems
• Simulated microgravity models (Random Positioning Machine, Rotating Wall Vessel): principles and applications in ground-based biological research
• Guided discussion and analysis of scientific articles on 3D models

Laboratory Activities:

• Microscopic observation of rigid scaffolds used in static, dynamic, and perfusion 3D cultures
• Preparation of cell cultures in hydrogels and on rigid scaffolds, including staining and cell identification
• Formation of spheroids using the hanging drop technique
• Introduction to the 3D printer: components and operation (laminar flow, UV light, compressed air, touch screen)
• Practical demonstration of 3D printing: preparation of tools, setting of printing parameters, model printing, and ink crosslinking using calcium chloride
• Observation of bioreactors used for 3D cultures in the laboratory and explanation of the research projects in which they are applied

Recommended Reading/Bibliography

Exam preparation is based primarily on selected scientific articles and materials provided by the instructor. All reference materials, including lecture slides and updated scientific literature, will be made available on the AulaWeb platform at the beginning of the course.

Teachers and Exam Board

• SARA TAVELLA
• CHIARA GENTILI

Exam Description

The exam is written and covers the entire program addressed during the course, including both lectures and laboratory activities. The test consists of 30 multiple-choice questions designed to assess students' understanding and ability to apply the concepts covered in the different parts of the course.

Assessment Methods

Assessment will be based on the accuracy of the answers provided in the multiple-choice final exam. The test is designed to evaluate the student's actual understanding of the course content and their ability to connect concepts, apply acquired knowledge, and use appropriate scientific terminology. Critical understanding of three-dimensional models and the ability to place them within the relevant application contexts will also be assessed.