Self-Organized 3D Tissue Patterns

Fundamentals, Design, and Experiments

Xiaolu Zhu, Zheng Wang

• Publisher: Jenny Stanford Publishing
• Year: 2022
• ISBN:
  9781000400847
• Edition: 1
• Topic: Matemáticas
• Subtopic: Matemáticas general

Therapies for regenerating damaged tissue and organs have been attracting much attention. In order to efficiently regenerate the functions of living tissue and organs, diverse attempts have been made to utilize scaffolds to "mold" artificial tissue structures. However, the structural complexity of reconstituted tissue is limited by the mechanical precision of scaffolds, which still cause problems arising from their degradation, immunogenic reactions, and so forth. It is also being realized that ultimately the best approach might be to rely on the innate self-organizing properties of cells and the regenerative capability of the organism itself.

This book investigates the 3D-pattern formation and evolution mechanism in multipotent cells embedded in 3D semi-synthetic hydrogels and the control methodology for self-organized patterns. The authors theoretically and experimentally demonstrate several types of topological 3D-pattern formation by cells in a 3D matrix in vitro, which can be modeled and predicted by mathematical models based on the reaction-diffusion dynamics of various chemical, physical, and mechanical cues. The study, focused on the 3D pattern formation of cells, provides (i) a unique perspective for understanding the self-organized 3D tissue structures based on Turing instability, (ii) the scheme for rationally controlling the cellular self-organization via exogenous factors or tailored inner interfaces inside hydrogels, and (iii) the elaborate and sophisticated regulating method for tuning collective cellular behaviors in 3D matrices.

Chapter 1 Introduction

1.1 Research in Tissue Engineering

Tissue engineering applies principles and methods from engineering and life sciences to create artificial constructs to direct tissue regeneration or enhance tissues and organs [1, 2]. In the context of incessant development of tools and improvements, such as synthetic biology and genomic editing techniques [3], tissue engineering is playing a leading role as a multidisciplinary research branch. In order to achieve effective strategies to regenerate the functions of living tissues and organs, many cell-based tissue engineering methods have been proposed to heal or reconstitute and restore tissue functions for translational medicine. Tissue engineering emerged as a scientific field with great potentialities in the 1980s, and extensive work on it has been developed since then, which aims at regenerating skin, cartilage, bone, and many other prototypes of tissue and organ substitutes, such as nerve conduits, blood vessels, liver, and even heart. Early stage of tissue engineering was directly motivated by practical therapy demands in clinics, especially in the areas of skin replacement and cartilage repair. Skin grafts are the first engineered tissue constructs, and autologous skin grafts were the golden choice [4], yet the available donor sites of graft materials are limited. Similar to skin grafts, osteochondral transplantation techniques repair cartilage by placing the autologous donor tissue harvested from nonweight-bearing regions of the joint [5]. This technique is also limited by the rather short supply of donor tissue available. A following improved method is autologous chondrocyte implantation (ACI) that implants the autologous cells significantly expanded in vitro into the debrided cartilage defect. This method provides enough cells via in vitro expansion which alleviates the short supply of autologous donor tissue. However, sometimes there will be chondrocyte leakage without the retention by an artificial scaffold, providing an uneven chondrocyte distribution [6, 7]. Therefore, the structured scaffolds are required for better efficacy in tissue/organ regeneration.

Traditional structured scaffolds, sometimes called macroscaffolds including exogenous scaffolds and decellularized 3D matrix scaffolds, may be the most widely investigated topic. Exogenous scaffolds, such as the synthetic porous scaffolds, provide spatial three-dimensional (3D) structure for seeded cells to adhere, spread, proliferate, differentiate, meanwhile to secrete extracellular matrix (ECM), and deliver biochemical factors across a 3D space [8–10], which could mimic the physicochemical, biological, and mechanical properties of native tissues/organs [11]. After cell seeding, the geometry and stiffness of the scaffold may influence the response of cells [12–16]. To reconstitute tissue architectural features, several studies have been devoted to fabricate various scaffolds with specific structures to guide cell spreading and growing [17–19]. Nevertheless, the structural complexity of the scaffolds is limited by the manufacture precision. On the other hand, decellularized 3D matrices obtained from solid cadaveric organ can retain the intact structure of natural tissues or organs. It advances the progress in building complex organs with vascular networks. In this technique, the original cells from the solid cadaveric organ could be decellularized using biochemical technology, and the obtained acellular natural scaffold with the original organ microarchitecture is then re-cellularized. The re-cellularized organ, including vascular network [20] seeded with autologous cells, is then transplanted into the patient. The tissue engineering based on decellularized natural scaffolds with great promise, yet still encounters the lack of donor organs with specific size and available functions for the patients. Although porcine organs are probably the potential substitutes for human organs because they are similar to human organs in size and function [21], currently the problems of immunological incompatibilities still exist after the important progress that the porcine endogenous retrovirus (PERV) in pigs was inactivated by using CRISPR-Cas9 [22].

In addition to structured scaffolds and decellularized matrices, a spectrum of approaches based on the self-organization process of cells have been studied and developed over the past three decades for tissue engineering and regenerative medicine. Central to this translational endeavor is taking advantage of cells’ natural capacity to synthesize the ECM, self-assembly into tissue architectures and concurrently respond to signals [23–26]. Self-organization based engineering approaches relieve the limitations that the scaffoldbased tissue engineering methods usually have probably included the non-synchronization of the scaffold degradation to neotissue (soft-tissue) formation and the concern of the immunogenicity due to scaffold creation, seeding, or degradation [27, 28]. Also, selforganization paradigm for fabricating tissues is usually realized by using the synthetic hydrogels as ECM. The hydrogels are easier to be produced relative to the decellularized 3D matrices that require donors. To get the controllable self-organization of cells, specifically synthesized ECM materials, external forces, specially tailored microfabrication [29], and the mathematical modeling for molecular and cellular activities [30], programmable control of endogenous gene networks [31] have been utilized as effective approaches.

The objective of this chapter is to describe advances in tissue and organ morphogenesis using diverse technologies from different disciplines. Traditional tissue grafting and cell implantation for skin or cartilage at early development stage of tissue engineering is described first. Then the commonly used structured scaffolds with gross morphology or structure features of native tissues are reviewed. The synthetic scaffolds and decellularized 3D matrices are the two main categories of the structured scaffolds here, and their features, developments, and limitations are, respectively, discussed. Subsequently, the massive efforts pursuing the controllable selforganizing scheme for tissue and organ regeneration are reviewed. The advantages and performances of the guidance by computational approaches for tissue/organ morphogenesis driven by selforganization scheme are also discussed in the following chapters. Rationally regulating self-organization exhibits high potential for complex tissue and organoid regeneration, which is an important step toward clinical translation.

1.2 Traditional Tissue Grafting and Typical Cell Implantation for Skin or Cartilage

Tissue engineering that combines knowledge from molecular biology, materials science, biomechanics, and medicine, intends to produce tissue constructs to repair or replace native tissues compromised by trauma, pathology, or age [32]. The brief history and progress of tissue engineering can be found in Berthiaume et al. [33]. Early studies were motivated by practical therapy demands, especially in the areas of skin replacement and cartilage repair, which are the most representative tissue engineering treatments. Skin grafts are the first engineered tissue constructs, and the early-stage skin grafts [27, 34–36] were mainly used for wounds having a diameter larger than 1 cm or that extend deep into the dermis, thereby requiring special treatment for closure. Autologous skin grafts remain the best choice [4], but the available donor sites are limited. Alternatively, the early tissue-engineered skin constructs were also produced by in vitro coculturing the keratinocytes isolated from patients [34–36] with a feeder layer of mouse mesenchymal cells. The keratinocytes grew ex vivo and expanded the coverage area a thousand-fold in several weeks. This achievement led to the first cell-based tissueengineered product called Epicel, for treating patients suffering from catastrophic burn injuries. But this product does not have a dermis and has a thickness filled with a few cells, which limited its common usage. To make a skin-equivalent tissue with full thickness, researchers developed a composite skin product named Apligraf that reconstituted both dermis and epidermis [27]. Since then, other analogous products incorporating bovine type I collagen sponge as substrates have also been developed. However, because living cells are used, immunological rejection may occur. Some alternative types of skin grafts were developed, which contained no extra living cells before they were placed at the wound surface. Their main mission was to guide and stimulate the body’s repair and regenerative processes. They used a porous acellular matrix located at the wound surface, thus allowing the migration and revascularization of host cells such as endothelial cells, neural cells, and fibroblasts into it [37].

Similar to skin grafts, the development of cartilage grafts is also driven by clinical demands. The demand for engineered and regenerative cartilages has been growing because of the increasing prevalence of degenerative joint diseases (e.g., osteoarthritis). Additionally, the spontaneous repair for the injured articular cartilage is still limited even for young and healthy individuals [38]. Articular cartilage is a thick, avascular, aneural tissue that located at the ends of long bones [39, 40]. Since the articular cartilage naturally possesses no vasculature, it is hard to regrow and get the neotissue (softtissue) regenerated. Artificial cartilage repairing approaches have been investigated for over 30 years, and important successes have been achieved in clinics. The transplant of autologous donor tissue harvested from a non-weight-bearing region to the damaged site can help to repair cartilages. However, similar to the case with autologous skin grafts, this approach is subject to the availability of cells and donor sites. At early stage, microfracture [41] was introduced in the late 1980s and early 1990s to treat the cartilage defects. This technique can enhances migration of mesenchymal stem cells (MSCs) from bone marrow to the site of a cartilage defect by penetrating the subchondral bone [42]. However, microfracture often leads to fibrocartilage formation and treatment failure was expected in a term more than 5 years after surgery [43, 44]. A following improved method is ACI. In this method, a chondrocyte population, collected from the patient, is expanded in vitro, yielding ~12–48 million cells; then the chondrocyte population is implanted into the debrided cartilage defect. This technique showed prolonged improvement compared to autologous osteochondral transplantation in excess of years postoperatively [45, ...

Table of contents

• Cover Page
• Half Title Page
• Title Page
• Copyright Page
• Table of Contents
• Preface
• 1. Introduction
• 2. Fundamentals of Three-Dimensional Cell Culture in Hydrogels
• 3. Three-Dimensional Patterns of Tissues Emerging in Hydrogels
• 4. Constructing 3D Tissue Structures via Cellular Self-Assembly at Patterned Interfaces inside Hydrogel
• 5. Modeling Cellular Self-Assembly at Patterned Interfaces inside Hydrogel via Turing’s Reaction-Diffusion Frame
• 6. Tuning Cellular Behaviors during Self-Organization of Cells in Hydrogel by Changing Inner Nano-Structure of Hydrogel
• Index