Raman Spectroscopy Based Techniques in Tissue Engineering—An Overview

Cleft lip and palate patients suffer from functional, aesthetical, and psychosocial problems due to suboptimal regeneration of skin, mucosa, and skeletal muscle after restorative cleft surgery. The field of tissue engineering and regenerative medicine (TE/RM) aims to restore the normal physiology of tissues and organs in conditions such as birth defects or after injury. A crucial factor in cell differentiation, tissue formation, and tissue function is mechanical strain. Regardless of this, mechanical cues are not yet widely used in TE/RM. The effects of mechanical stimulation on cells are not straight-forward in vitro as cellular responses may differ with cell type and loading regime, complicating the translation to a therapeutic protocol. We here give an overview of the different types of mechanical strain that act on cells and tissues and discuss the effects on muscle, and skin and mucosa. We conclude that presently, sufficient knowledge is lacking to reproducibly implement external mechanical loading in TE/RM approaches. Mechanical cues can be applied in TE/RM by fine-tuning the stiffness and architecture of the constructs to guide the differentiation of the seeded cells or the invading surrounding cells. This may already improve the treatment of orofacial clefts and other disorders affecting soft tissues.

BACKGROUND Cranioplasty is a procedure commonly performed as the next step in management for patients with decompressive craniectomy. It is commonly associated with complications that might result in implant failure and reoperation. Tissue engineering and regenerative medicine (TERM) involve the development of biological substitutes that restore tissue function and regrow and replace damaged tissues. TERM is a potential solution that might allow for improved long-term outcomes in cranioplasty. OBJECTIVE To present a case series that describes the management and treatment outcomes of using TERM in cranioplasty, using polycaprolactone-tricalcium phosphate (PCL-TCP) implants in 2 patients. METHODS In 2 young adults, large bone defects of the skull following decompressive craniectomy were reconstructed using patient-specific implants manufactured from PCL-TCP. Bone marrow aspirate was used as autologous biologic that incorporates stem cell and growth factors into the implant. RESULTS Both patients recovered without any complications following surgery. Physical examination shows favorable healing outcomes with the defect area having a hard texture upon palpation. Postoperative imaging performed at 8 and 20 months for each patient, respectively, demonstrated ossification over the defect area. CONCLUSION Satisfactory bone growth is seen with TERM cranioplasty. The PCL-TCP implant is a novel and recommendable alternative to currently available patient-specific implants used in cranioplasty. TERM cranioplasty is the potential solution to the quest of finding the ideal cranioplasty implant.

The field of tissue engineering and regenerative medicine (TERM) has exploded in the last decade. In this Year (or so) in Review, we highlight some of the high impact advances within the field over the past several years. Using the past as our guide and starting with an objective premise, we attempt so to identify recent "hot topics" and transformative publications within the field. Through this process, several key themes emerged: (1) tissue engineering: grafts and materials, (2) regenerative medicine: scaffolds and factors that control endogenous tissue formation, (3) clinical trials, and (4) novel cell sources: induced pluripotent stem cells. Within these focus areas, we summarize the highly impactful articles that emerged from our objective analysis and review additional recent publications to augment and expand upon these key themes. Finally, we discuss where the TERM field may be headed and how to monitor such a broad-based and ever-expanding community.

Tissue engineering and regenerative medicine (TERM) as an emerging field is a multidisciplinary science and combines basic sciences such as biomaterials science, biology, genetics and medical sciences to achieve functional TERM-based products to regenerate or replace damaged or diseased tissues or organs. Probiotics are useful microorganisms which have multiple effective functions on human health. They have some immunomodulatory and biocompatibility effects and improve wound healing. In this article, we describe the latest findings on probiotics and their pro-healing properties on various body systems that are useable in regenerative medicine. Therefore, this review presents a new perspective on the therapeutic potential of probiotics for TERM.

Tissue Engineering and Regenerative Medicine Strategies in Meniscus Lesions

The persistent societal challenge of organ donor shortage, driven by factors such as population growth, increased lifespan, and higher disease prevalence, leaves thousands awaiting life-saving transplants. Tissue Engineering and Regenerative Medicine (TERM) emerge as promising solutions, with 3D biofabrication technologies capable of creating functional tissues and preclinical testing models. These technologies, spanning soft tissue structures to musculoskeletal replacements, aim to address the critical shortage of organ donors. Melt electrowriting (MEW), enable precise recreation of native tissue equivalents due to its high resolution and fine control over architectures. MEW, and other biofabrication technologies automate the generation of biologically functional products with structural organization using living cells, bioactive molecules, and biomaterials. Tailoring tissue architecture, porosity, and structural organization is crucial for directing cell differentiation toward specific target tissues. While offering fine control over cell alignment and differentiation, ensuring the durability of engineered constructs is vital. Hydrogels, commonly used in TERM, provide a suitable environment for cell growth but often lack mechanical strength. Techniques such as MEW reinforce hydrogels, addressing this challenge. Vascularization within thick tissue structures remains a hurdle, necessitating strategies like modifying scaffold porosity and incorporating growth factors. To incorporate biochemical cues, atmospheric-pressure plasma jet (APPJ) functionalization was used to covalently immobilize transforming growth factor beta-1 (TGFβ1), improving hydrophilicity, and chondrogenic differentiation. Results indicated enhanced compressive modulus and glycosaminoglycan production compared to conventional TGFβ1 supply in-medium. Structural cues were then explored to guide cell behavior using MEW. Advances in 3D fiber scaffold manufacturing were discussed for their potential in repairing cardiac tissues. Methacrylated hydroxyl-functionalized PCL was investigated to covalently bond MEW microfibers to methacrylated silk fibroin hydrogels, resulting in more durable constructs with improved mechanical strength for articular cartilage tissue engineering. The introduction of z-porosity into MEW box structure scaffolds using isomalt as a sacrificial material demonstrated minimal impact on structural integrity, offering a promising method for creating z-directional pores crucial for cellular interconnectivity and vascularization. MEW and extrusion-based bioprinting were converged to create complex tissue patterns with native-inspired pre-vascular pathways. Optimal bioinks were identified for myocardial cellular arrangement and vascular cell connectivity. Preservation methods for tissue-engineered constructs were compared, highlighting hypothermic preservation as a promising approach for short-term preservation and transportation. Building upon myocardial tissue engineering approaches explored in this thesis, steps were taken to generate a full-thickness transmural tissue engineered construct. Co-culturing cardiomyocytes with fibroblasts and optimizing culture substrates improved alignment and functionality, showcasing the potential for sophisticated in vitro cardiac models with native-like architecture. In conclusion, this thesis explored MEW as a versatile technology, combining it with other fabrication strategies to achieve biochemical- and/or structural-guided tissue regeneration. By manipulating design and technologies, functional tissues for a broad range of target applications were generated, including contractile, muscular, myocardial tissue, and fibrous, dense articular cartilage tissue.

Advancements, Strategies, and Challenges in Organoid-Based Drug Evaluation for Tissue Engineering and Regenerative Medicine

The increasing demand for organ replacements in a growing world with an aging population as well as the loss of tissues and organs due to congenital defects, trauma and diseases has resulted in rapidly evolving new approaches for tissue engineering and regenerative medicine (TERM). The extracellular matrix (ECM) is a crucial component in tissues and organs that surrounds and acts as a physical environment for cells. Thus, ECM has become a model guide for the design and fabrication of scaffolds and biomaterials in TERM. However, the fabrication of a tissue/organ replacement or its regeneration is a very complex process and often requires the combination of several strategies such as the development of scaffolds with multiple functionalities and the simultaneous delivery of growth factors, biochemical signals, cells, genes, immunomodulatory agents, and external stimuli. Although the development of multifunctional scaffolds and biomaterials is one of the most studied approaches for TERM, all these strategies can be combined among them to develop novel synergistic approaches for tissue regeneration. In this review we discuss recent advances in which multifunctional scaffolds alone or combined with other strategies have been employed for TERM purposes.

Injectable soft-tissue augmentation by tissue engineering and regenerative medicine with human mesenchymal stromal cells, platelet-rich plasma and hyaluronic acid scaffolds

6 - Toxicology of nanoformulations and materials in tissue engineering

The existing 3D printing methods exhibit certain fabrication-dependent limitations for printing curved constructs that are relevant for many tissues. Four-dimensional (4D) printing is an emerging technology that is expected to revolutionize the field of tissue engineering and regenerative medicine (TERM). 4D printing is based on 3D printing, featuring the introduction of time as the fourth dimension, in which there is a transition from a 3D printed scaffold to a new, distinct, and stable state, upon the application of one or more stimuli. Here, we present an overview of the current developments of the 4D printing technology for TERM, with a focus on approaches to achieve temporal changes of the shape of the printed constructs that would enable biofabrication of highly complex structures. To this aim, the printing methods, types of stimuli, shape-shifting mechanisms, and cell-incorporation strategies are critically reviewed. Furthermore, the challenges of this very recent biofabrication technology as well as the future research directions are discussed. Our findings show that the most common printing methods so far are stereolithography (SLA) and extrusion bioprinting, followed by fused deposition modelling, while the shape-shifting mechanisms used for TERM applications are shape-memory and differential swelling for 4D printing and 4D bioprinting, respectively. For shape-memory mechanism, there is a high prevalence of synthetic materials, such as polylactic acid (PLA), poly(glycerol dodecanoate) acrylate (PGDA), or polyurethanes. On the other hand, different acrylate combinations of alginate, hyaluronan, or gelatin have been used for differential swelling-based 4D transformations. TERM applications include bone, vascular, and cardiac tissues as the main target of the 4D (bio)printing technology. The field has great potential for further development by considering the combination of multiple stimuli, the use of a wider range of 4D techniques, and the implementation of computational-assisted strategies.

Tissue engineering and regenerative medicine (TERM) is an innovative area of research and development that includes biomaterials, cells, scaffolds, scaffolding structures, and even bioactive substances to restore, repair or regenerate damaged tissues along with organs. The scope of synthetic skin expansion has now progressed to engineering kidneys, hearts, and lungs. Use of stem cell therapies or extracellular matrices for organ healing and regeneration are other; notable shifts toward tissue engineering paradigm are towards healing adn regeneration rather than the development of reconstructive surgery. While research has shown promise, there are still concerns regarding its clinical use citing complications such as immune system rejection of newly implanted tissues, ethical issues and the economical creation of engineered tissues. The efforts being made in Asia demonstrate emerging potential in TERM. Enhanced collaboration across disciplines is essential to pave new ways towards realizing current restrictions and bold influencing ideas alongside breakthroughs.

Mastication is important processes for better quality of life. After teeth extraction or trauma, tumor resection, skeletal malformations, infection and periodontal disease, alveolar bone resorption is occured and bone graft is often needed. A present gold standard of the bone graft is autologous bone graft, but it has drawbacks such as heavy burden on the patient. Tissue engineering and regenerative medicine (TERM) could be used to reduce morbidity and have been lately attracted considerable attention to restore tissue and function lost or damaged. In addition, the leading part for clinical application is somatic stem cells. Especially mesenchymal stem cells (MSCs) are easily isolated and extracted from various tissues including bone marrow, adipose, muscle, dermis, neural tissue, deciduous teeth, and dental pulp. It has the ability of self-renewal, trans-differentiation potential, plasticity, and immunosuppressive effects. It was also applied in oral and maxillofacial surgery fields. We developed the study of bone tissue engineering, which was named as “injectable tissue engineered bone (TEB)”, with MSCs as the stem cells and platelet rich plasma (PRP) as scaffolds and growth factors, and clinical trials were conducted with it. The results were good and reliable. In this time, it would introduce about application in bone regeneration for dental implants using tissue engineering technology. In the future, regenerative medicine with stem cells would be applied for not only dental field but also general disease as a promising treatment.

Kefiran is an exopolysaccharide produced by the microflora of kefir grains used to produce the fermented milk beverage kefir. The health-promoting and physicochemical properties of kefiran led to its exploration for a range of applications, mainly in the food industry and biomedical fields. Aiming to explore its potential for tissue engineering and regenerative medicine (TERM) applications, the kefiran biopolymer obtained through three different extraction methodologies was fully characterized and compared. High-quality kefiran polysaccharides were recovered with suitable yield through different extraction protocols. The methods consisted of heating the kefir grains prior to recovering kefiran by centrifugation and differed mainly in the precipitation steps included before lyophilization. Then, kefiran scaffolds were successfully produced from each extract by cryogelation and freeze-drying. In all extracts, it was possible to identify the molecular structure of the kefiran polysaccharide through 1H-NMR and FTIR spectra. The kefiran from extraction 1 showed the highest molecular weight (~3000 kDa) and the best rheological properties, showing a pseudoplastic behavior; its scaffold presented the highest value of porosity (93.2% ± 2), and wall thickness (85.8 µm ± 16.3). All extracts showed thermal stability, good injectability and desirable viscoelastic properties; the developed scaffolds demonstrated mechanical stability, elastic behavior, and pore size comprised between 98–94 µm. Additionally, all kefiran products proved to be non-cytotoxic over L929 cells. The interesting structural, physicochemical, and biological properties showed by the kefiran extracts and cryogels revealed their biomedical potential and suitability for TERM applications.

The role of polysaccharide-based biodegradable soft polymers in the healthcare sector