Ibrahim Fatih Cengiz

Preparation and characterization of collagen/PLA, chitosan/PLA, and collagen/chitosan/PLA hybrid scaffolds for cartilage tissue engineering

In this study, three-dimensional (3D) porous scaffolds were developed for the repair of articular cartilage defects. Novel collagen/polylactide (PLA), chitosan/PLA, and collagen/chitosan/PLA hybrid scaffolds were fabricated by combining freeze-dried natural components and synthetic PLA mesh, where the 3D PLA mesh gives mechanical strength, and the natural polymers, collagen and/or chitosan, mimic the natural cartilage tissue environment of chondrocytes. In total, eight scaffold types were studied: four hybrid structures containing collagen and/or chitosan with PLA, and four parallel plain scaffolds with only collagen and/or chitosan. The potential of these types of scaffolds for cartilage tissue engineering applications were determined by the analysis of the microstructure, water uptake, mechanical strength, and the viability and attachment of adult bovine chondrocytes to the scaffolds. The manufacturing method used was found to be applicable for the manufacturing of hybrid scaffolds with highly porous 3D structures. All the hybrid scaffolds showed a highly porous structure with open pores throughout the scaffold. Collagen was found to bind water inside the structure in all collagen-containing scaffolds better than the chitosan-containing scaffolds, and the plain collagen scaffolds had the highest water absorption. The stiffness of the scaffold was improved by the hybrid structure compared to plain scaffolds. The cell viability and attachment was good in all scaffolds, however, the collagen hybrid scaffolds showed the best penetration of cells into the scaffold. Our results show that from the studied scaffolds the collagen/PLA hybrids are the most promising scaffolds from this group for cartilage tissue engineering.

Quantitative assessment of the regenerative and mineralogenic performances of the zebrafish caudal fin

The ability of zebrafish to fully regenerate its caudal fin has been explored to better understand the mechanisms underlying de novo bone formation and to develop screening methods towards the discovery of compounds with therapeutic potential. Quantifying caudal fin regeneration largely depends on successfully measuring new tissue formation through methods that require optimization and standardization. Here, we present an improved methodology to characterize and analyse overall caudal fin and bone regeneration in adult zebrafish. First, regenerated and mineralized areas are evaluated through broad, rapid and specific chronological and morphometric analysis in alizarin red stained fins. Then, following a more refined strategy, the intensity of the staining within a 2D longitudinal plane is determined through pixel intensity analysis, as an indicator of density or thickness/volume. The applicability of this methodology on live specimens, to reduce animal experimentation and provide a tool for in vivo tracking of the regenerative process, was successfully demonstrated. Finally, the methodology was validated on retinoic acid- and warfarin-treated specimens, and further confirmed by micro-computed tomography. Because it is easily implementable, accurate and does not require sophisticated equipment, the present methodology will certainly provide valuable technical standardization for research in tissue engineering, regenerative medicine and skeletal biology.

Segmental and regional quantification of 3D cellular density of human meniscus from osteoarthritic knee

The knee menisci have important roles in the knee joint. Complete healing of the meniscus remains a challenge in the clinics. Cellularity is one of the most important biological parameters that must be taken into account in regenerative strategies. However, knowledge on the 3D cellularity of the human meniscus is lacking in the literature. The aim of this study was to quantify the 3D cellular density of human meniscus from the osteoarthritic knee in a segmental and regional manner with respect to laterality. Human lateral menisci were histologically processed and stained with Giemsa for histomorphometric analysis. The cells were counted in an in‐depth fashion. 3D cellular density in the vascular region (27 199 cells/mm3) was significantly higher than in the avascular region (12 820 cells/mm3). The cells were observed to possess two distinct morphologies, roundish or flattened. The 3D density of cells with fibrochondrocyte morphology (14 705 cells/mm3) was significantly greater than the 3D density of the cells with fibroblast‐like cell morphology (5539 cells/mm3). The best‐fit equation for prediction of the 3D density of cells with fibrochondrocyte morphology was found to be:

Tissue Engineering and Regenerative Medicine Strategies for the Treatment of Osteochondral Lesions

There is an unmet clinical need for repairing osteochondral (OC) lesions. Tissue engineering and regenerative medicine (TERM) strategies advance with the possibility of regenerating different tissues by means of using cells, scaffolds and growth factors, alone or together. The use of bioreactor systems for developing mature tissues in vitro is also appealing. This book chapter aims to overview the main aspects related to structure and functions of articular cartilage, subchondral bone, and bone. The components of the tissue engineering and the most relevant reports on their use for treating OC lesions are concisely covered. Several treatment strategies are available; however, the gold standard does not exist. The biofunctional knowledge of these tissues has been uncovered by the development of advanced characterization techniques including medical imaging allowing visualization from sub-cellular to macro level. These techniques have been helping scientists not only to understand how these tissues function but also to develop multiscale TERM strategies. Thus, this hot topic is also briefly discussed.

Advanced regenerative strategies for human knee meniscus

The meniscus tissue has important roles in the function and biomechanics of the knee. Despite the great advances in the treatment of meniscus lesions, the clinical need is still not fulfilled. To overcome the challenges of regeneration, tissue engineering-based strategies have been attempted with limited success. The process of meniscus tissue regeneration is very complex and has many parameters that are evident only to a certain degree. Today, the regenerative strategies have been advancing beyond the traditional tissue engineering concept by growing the utilization of the expertises of complementary areas that include, but not limited to, bioreactor engineering, bioprinting coupled to reverse engineering, biology, nanotechnology and gene therapy approaches. Herein, the recent reported advanced strategies involving bioreactors, self-assembling process, and somatic gene therapy for meniscus regeneration are overviewed.

Meniscal Repair: Indications, Techniques, and Outcome

The current trend when dealing with meniscus tears is “preservation whenever possible.” This way, meniscal repair techniques have significantly developed in recent years. There have been advances in both surgical techniques (e.g., techniques for root tear repair) and related devices. However, anatomical knowledge is crucial, and learning curve time for any technique should be considered. Classification of tears, preoperative planning, and surgical training (including cadaver courses) are extremely important.

Treatments of Meniscus Lesions of the Knee: Current Concepts and Future Perspectives

The present preference in the clinical management of meniscus lesions is to preserve it by repairing whenever possible or substituting the tissue. Still, meniscectomy continues to be one of the most frequent orthopedic procedures regardless of the fact that it may lead to a series of early degenerative events in the knee. Surgical and technological advances enabled to extend the indications for meniscus repair. The outcome of meniscus repair is influenced by several factors. Classification of meniscus lesions remains a challenge while there have been some attempts in building consensus around it. Substitution of meniscus tissue has been performed to avoid or minimize the possible degenerative effects occurring in the absence of meniscus. Meniscus allograft transplantation has demonstrated its use as a replacement strategy of large lesions. In partial lesions, the use of acellular scaffolds has provided an improved clinical outcome when the insertional horns and the peripheral rim are preserved. However, the current scaffolds have shown some limitations, and the neotissue is different from the native meniscus. Tissue engineers thus envision going beyond the partial meniscus regeneration. Nowadays, it is aimed to develop a new generation of meniscal implants for total meniscus regeneration, which not only meet the biomechanical requirements but also the biological requirements both in the short- and long-term. Moreover, these might be patient/injury-specific regarding the size and shape as well as being cultivated with autologous cells and biologically enhanced. Herein, the clinical management of meniscus lesions and advanced tissue engineering strategies are reviewed.Lay SummaryMeniscus injuries are the most frequent injuries in the knee. Given the increased understanding of the consequences of meniscectomy, which is still one of the most frequent orthopedic procedures, the clinical management of meniscus changed towards favoring repair or substitution. The future of meniscus substitution and regeneration is strongly supported by the clinical need. This study reviews the current concepts and provides future perspectives on the clinical management of meniscus lesions, and tissue engineering and regenerative medicine strategies to update and guide researchers and surgeons.

Histology-Ultrastructure-Biology

The menisci are nonuniform and heterogeneous structures with segmental variations according to its biology and function. It is a fibrocartilaginous tissue that plays a central role on the joint kinematics. However, they are also the most frequently injured structures of the human knee. In the advent of new technologies including tissue engineering and regenerative medicine, it is critical to deeper understand its ultrastructure and physiological function aiming to find ways for not only to enhance the repair and/or replacement approached but also to develop novel regenerative strategies. Herein, the presented work aims to describe the most relevant anatomic and biologic features of the meniscus.

Basics of the Meniscus

The meniscus is a fibro-cartilaginous tissue located between the femoral condyles and the tibial plateau in the knee. The presence of the meniscus tissue is vital for the proper function of the knee. Meniscal injuries are very frequent cases in the orthopaedics, and they have limited self-healing capacity. The importance of the basic science of meniscus has been acknowledged for the meniscus development of regenerative strategies, and the knowledge is increasing over time. Herein, the biology, anatomy, and biochemistry of meniscus tissue are overviewed.

Building the Basis for Patient-Specific Meniscal Scaffolds

The current strategies for the total or partial meniscus replacement with allograft transplantation or scaffold implantation need to be improved to overcome the limitations in the clinics. In addition to the required biological and biomechanical performance of the implants, the size and the shape of the implant are critical for the success of the treatment. The commercial implants are re-sized by cutting at the time of surgery according to the patient’s need; however, not completely in a 3D manner. The meniscal implants should advance beyond the traditional biomaterial concept by being patient-specific not only in terms of size and shape but regarding the cells and biologics derived from the patient. Herein, we overview the recent reports related to manufacturing of patient-specific meniscal implants.