Microstructure and Thermal Property of Designed Alginate-Based Polymeric Composite Foam Materials Containing Biomimetic Decellularized Elastic Cartilage Microscaffolds

This study presents a designed alginate-based polymeric composite foam material containing decellularized elastic cartilage microscaffolds from porcine elastic cartilage by using supercritical fluid and papain treatment for medical scaffold biomaterials. The microstructure and thermal property of the designed alginate-based polymeric composite foam materials with various controlled ratios of alginate molecules and decellularized elastic cartilage microscaffolds were studied and characterized by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and differential thermal gravimetric analysis (TGA/DTG). The microstructure and thermal property of the composite foam materials were affected by the introduction of decellularized elastic cartilage microscaffolds. The designed alginate-based polymeric composite foam materials containing decellularized elastic cartilage microscaffolds were ionically cross-linked with calcium ions by soaking the polymeric composite foam materials in a solution of calcium chloride. Additional calcium ions further improved the microstructure and thermal stability of the resulting ionic cross-linked alginate-based polymeric composite foam materials. Furthermore, the effect of crosslinking functionality on microstructures and thermal properties of the resulting polymeric composite foam materials were studied to build up useful information for 3D substrates for cultivating and growing cartilage cells and/or cartilage tissue engineering.

Three-Dimensional-Printed External Scaffolds Mitigate Loss of Volume and Topography in Engineered Elastic Cartilage Constructs

Objective: A major obstacle in the clinical translation of engineered auricular scaffolds is the significant contraction and loss of topography that occur during maturation of the soft collagen-chondrocyte matrix into elastic cartilage. We hypothesized that 3-dimensional-printed, biocompatible scaffolds would “protect” maturing hydrogel constructs from contraction and loss of topography. Design: External disc-shaped and “ridged” scaffolds were designed and 3D-printed using polylactic acid (PLA). Acellular type I collagen constructs were cultured in vitro for up to 3 months. Collagen constructs seeded with bovine auricular chondrocytes (BAuCs) were prepared in 3 groups and implanted subcutaneously in vivo for 3 months: preformed discs with (“Scaffolded/S”) or without (“Naked/N”) an external scaffold and discs that were formed within an external scaffold via injection molding (“Injection Molded/SInj”). Results: The presence of an external scaffold or use of injection molding methodology did not affect the acellular construct volume or base area loss. In vivo, the presence of an external scaffold significantly improved preservation of volume and base area at 3 months compared to the naked group ( P < 0.05). Construct contraction was mitigated even further in the injection molded group, and topography of the ridged constructs was maintained with greater fidelity ( P < 0.05). Histology verified the development of mature auricular cartilage in the constructs within external scaffolds after 3 months. Conclusion: Custom-designed, 3D-printed, biocompatible external scaffolds significantly mitigate BAuC-seeded construct contraction and maintain complex topography. Further refinement and scaling of this approach in conjunction with construct fabrication utilizing injection molding may aid in the development of full-scale auricular scaffolds.

Ear wound healing in MRL/MpJ mice is associated with gut microbiome composition and is transferable to non-healer mice via microbiome transplantation

Objective Adult elastic cartilage has limited repair capacity. MRL/MpJ (MRL) mice, by contrast, are capable of spontaneously healing ear punctures. This study was undertaken to characterize microbiome differences between healer and non-healer mice and to evaluate whether this healing phenotype can be transferred via gut microbiome transplantation. Methods We orally transplanted C57BL/6J (B6) mice with MRL/MpJ cecal contents at weaning and as adults (n = 57) and measured ear hole closure 4 weeks after a 2.0mm punch and compared to vehicle-transplanted MRL and B6 (n = 25) and B6-transplanted MRL (n = 20) mice. Sex effects, timing of transplant relative to earpunch, and transgenerational heritability were evaluated. In a subset (n = 58), cecal microbiomes were profiled by 16S sequencing and compared to ear hole closure. Microbial metagenomes were imputed using PICRUSt. Results Transplantation of B6 mice with MRL microbiota, either in weanlings or adults, improved ear hole closure. B6-vehicle mice healed ear hole punches poorly (0.25±0.03mm, mm ear hole healing 4 weeks after a 2mm ear hole punch [2.0mm—final ear hole size], mean±SEM), whereas MRL-vehicle mice healed well (1.4±0.1mm). MRL-transplanted B6 mice healed roughly three times as well as B6-vehicle mice, and half as well as MRL-vehicle mice (0.74±0.05mm, P = 6.9E-10 vs. B6-vehicle, P = 5.2E-12 vs. MRL-vehicle). Transplantation of MRL mice with B6 cecal material did not reduce MRL healing (B6-transplanted MRL 1.3±0.1 vs. MRL-vehicle 1.4±0.1, p = 0.36). Transplantation prior to ear punch was associated with the greatest ear hole closure. Offspring of transplanted mice healed significantly better than non-transplanted control mice (offspring:0.63±0.03mm, mean±SEM vs. B6-vehicle control:0.25±0.03mm, n = 39 offspring, P = 4.6E-11). Several microbiome clades were correlated with healing, including Firmicutes (R = 0.84, P = 8.0E-7), Lactobacillales (R = 0.65, P = 1.1E-3), and Verrucomicrobia (R = -0.80, P = 9.2E-6). Females of all groups tended to heal better than males (B6-vehicle P = 0.059, MRL-transplanted B6 P = 0.096, offspring of MRL-transplanted B6 P = 0.0038, B6-transplanted MRL P = 1.6E-6, MRL-vehicle P = 0.0031). Many clades characteristic of female mouse cecal microbiota vs. males were the same as clades characteristic of MRL and MRL-transplanted B6 mice vs. B6 controls, including including increases in Clostridia and reductions in Verrucomicrobia in female mice. Conclusion In this study, we found an association between the microbiome and tissue regeneration in MRL mice and demonstrate that this trait can be transferred to non-healer mice via microbiome transplantation. We identified several microbiome clades associated with healing.

Nasal Septum Deviation as the Consequence of BMP-Controlled Changes to Cartilage Properties

The nasal septum cartilage is a specialized hyaline cartilage important for normal midfacial growth. Abnormal midfacial growth is associated with midfacial hypoplasia and nasal septum deviation (NSD). However, the underlying genetics and associated functional consequences of these two anomalies are poorly understood. We have previously shown that loss of Bone Morphogenetic Protein 7 (BMP7) from neural crest (BMP7ncko) leads to midfacial hypoplasia and subsequent septum deviation. In this study we elucidate the cellular and molecular abnormalities underlying NSD using comparative gene expression, quantitative proteomics, and immunofluorescence analysis. We show that reduced cartilage growth and septum deviation are associated with acquisition of elastic cartilage markers and share similarities with osteoarthritis (OA) of the knee. The genetic reduction of BMP2 in BMP7ncko mice was sufficient to rescue NSD and suppress elastic cartilage markers. To our knowledge this investigation provides the first genetic example of an in vivo cartilage fate switch showing that this is controlled by the relative balance of BMP2 and BMP7. Cellular and molecular changes similar between NSD and knee OA suggest a related etiology underlying these cartilage abnormalities.

Bioprinted Cartilage

This chapter chronicles the difficulties in bioprinting any of the three types of cartilage, with emphasis on the articular cartilage, which is so often damaged in our knees and hips. The chapter sets out the disparity between the apparent simplicity of cartilage (no blood vessels, no nerves, and composed of a single, sparse cell type) and the complexity of its zonal architecture (disparate cell structure and orientation in different regions of the same cartilage tissue). The chapter presents examples of the bioprinting of all three cartilage types: articular cartilage (belonging to the hyaline family of cartilage), elastic cartilage (such as is found in our ears), and fibrocartilage (such as the meniscus in our knees). The chapter discusses the promise of decellularized extracellular matrix (which is extracellular matrix—the cells’ immediate environment—with the cells removed) as a bioink.

Метод количественного описания фиксированных гистологических образцов с помощью цифровой голографической микроскопии

A novel method for the quantitative characterization of fixed histological samples based on the statistical analysis of their phase images obtained using digital holographic microscopy is developed and presented. The proposed approach allows for fully automated processing of reconstructed phase images and obtaining quantitative data of morphological and optical characteristics of histological tissues structures. The method was validated on three histological samples of different types of tissues: ciliated columnar epithelium, elastic cartilage, and liver.

Development of a Method for Scaffold-Free Elastic Cartilage Creation

Microtia is a congenital aplasia of the auricular cartilage. Conventionally, autologous costal cartilage grafts are collected and shaped for transplantation. However, in this method, excessive invasion occurs due to limitations in the costal cartilage collection. Due to deformation over time after transplantation of the shaped graft, problems with long-term morphological maintenance exist. Additionally, the lack of elasticity with costal cartilage grafts is worth mentioning, as costal cartilage is a type of hyaline cartilage. Medical plastic materials have been transplanted as alternatives to costal cartilage, but transplant rejection and deformation over time are inevitable. It is imperative to create tissues for transplantation using cells of biological origin. Hence, cartilage tissues were developed using a biodegradable scaffold material. However, such materials suffer from transplant rejection and biodegradation, causing the transplanted cartilage tissue to deform due to a lack of elasticity. To address this problem, we established a method for creating elastic cartilage tissue for transplantation with autologous cells without using scaffold materials. Chondrocyte progenitor cells were collected from perichondrial tissue of the ear cartilage. By using a multilayer culture and a three-dimensional rotating suspension culture vessel system, we succeeded in creating scaffold-free elastic cartilage from cartilage progenitor cells.