Anna D. Dikina

Engineered cartilaginous tubes for tracheal tissue replacement via self-assembly and fusion of human mesenchymal stem cell constructs

There is a critical need to engineer a neotrachea because currently there are no long-term treatments for tracheal stenoses affecting large portions of the airway. In this work, a modular tracheal tissue replacement strategy was developed. High-cell density, scaffold-free human mesenchymal stem cell-derived cartilaginous rings and tubes were successfully generated through employment of custom designed culture wells and a ring-to-tube assembly system. Furthermore, incorporation of transforming growth factor-β1-delivering gelatin microspheres into the engineered tissues enhanced chondrogenesis with regard to tissue size and matrix production and distribution in the ring- and tube-shaped constructs, as well as luminal rigidity of the tubes. Importantly, all engineered tissues had similar or improved biomechanical properties compared to rat tracheas, which suggests they could be transplanted into a small animal model for airway defects. The modular, bottom up approach used to grow stem cell-based cartilaginous tubes in this report is a promising platform to engineer complex organs (e.g., trachea), with control over tissue size and geometry, and has the potential to be used to generate autologous tissue implants for human clinical applications.

Combined Administration of ASCs and BMP-12 Promotes an M2 Macrophage Phenotype and Enhances Tendon Healing

BackgroundOutcomes after intrasynovial tendon repair are highly variable. An intense inflammatory cascade followed by a delayed healing response can cause adhesion formation and repair-site failure that severely impair the function of repaired digits. No effective remedies exist to fully address these issues. Cell- and growth factor-based therapies have been shown to modulate inflammation and improve cell proliferation and matrix synthesis and therefore are promising treatment approaches for intrasynovial tendon repair.Questions/Purposes(1) Can autologous adipose-derived mesenchymal stromal cells (ASCs) and recombinant bone morphogenetic protein-12 (rBMP-12) be effectively delivered to an intrasynovial flexor tendon repair without adverse effects? (2) Do autologous ASCs modulate the inflammatory response after intrasynovial tendon injury and repair? (3) Does the combined application of autologous ASCs and rBMP-12 modulate the proliferative and remodeling responses after intrasynovial tendon injury and repair?MethodsSixteen 1- to 2-year-old female canines were used in this study. Autologous ASC sheets, with and without rBMP-12, were applied to the surface of sutured flexor tendons. Fourteen days after repair, the effects of treatment were determined using quantitative PCR (six per group) for the expression of genes related to macrophage phenotype or inflammation (IL-4, CD163, VEGF, NOS2, IL-1B, and IFNG), cell proliferation (CCND1), and tendon formation (SCX, TNMD, COL1A1 and COL3A1). Proteomics analysis (four per group) was performed to examine changes in tendon protein abundances. CD146 immunostaining and hematoxylin and eosin staining (four per group) were used to detect tendon stem or progenitor cells and to semiquantitatively evaluate cellularity at the tendon repair; analyses were done blinded to group.ResultsGross inspection and cell tracing showed that autologous ASCs and rBMP-12 were delivered to the flexor tendon repair site without the deleterious effects of adhesion and repair-site gap formation. Quantitative assessment of gene and protein expression showed effects of treatment: ASC-sheet treatment modulated the postrepair inflammatory response and facilitated healing by increasing regenerative M2 macrophages (M2 marker CD204, twofold of normal, p = 0.030), inflammatory inhibitor (prostaglandin reductase 1 [PTRG1], 1.6-fold of normal, p = 0.026), and proteins involved in tendon formation (periostin [POSTN], 1.9-fold of normal, p = 0.035). Consistently, semiquantitative and qualitative evaluations of repaired tissue showed that ASC-sheet treatment reduced mononuclear cell infiltration (12% less than nontreated tendons, p = 0.021) and introduced CD146+ stem or progenitor cells to the repair site. The combined administration of ASCs and rBMP-12 further stimulated M2 macrophages by increasing IL-4 (116-fold of normal, p = 0.002) and led to the increase of M2 effector matrix metalloproteinase-12 involved in matrix remodeling (twofold of normal, p = 0.016) and reduction of a negative regulator of angiogenesis and cell migration (StAR-related lipid transfer domain protein13 [STARD13]; 84% of normal, p = 0.000), thus facilitating the proliferative stage of tendon repair.ConclusionsASCs and BMP-12 accelerated the progression of healing in the proliferative stage of tendon repair. The effects of ASCs and BMP-12 on tendon functional recovery should be evaluated in future studies.Clinical RelevanceThe cell sheet approach is an effective, biocompatible, and surgeon-friendly approach for cell and growth factor delivery during tendon repair. Combined application of ASCs and BMP-12 may accelerate intrasynovial tendon healing while suppressing the adverse inflammatory response.

High-throughput approaches for screening and analysis of cell behaviors

The rapid development of new biomaterials and techniques to modify them challenge our capability to characterize them using conventional methods. In response, numerous high-throughput (HT) strategies are being developed to analyze biomaterials and their interactions with cells using combinatorial approaches. Moreover, these systematic analyses have the power to uncover effects of delivered soluble bioactive molecules on cell responses. In this review, we describe the recent developments in HT approaches that help identify cellular microenvironments affecting cell behaviors and highlight HT screening of biochemical libraries for gene delivery, drug discovery, and toxicological studies. We also discuss HT techniques for the analyses of cell secreted biomolecules and provide perspectives on the future utility of HT approaches in biomedical engineering.

A Modular Strategy to Engineer Complex Tissues and Organs

Abstract Currently, there are no synthetic or biologic materials suitable for long‐term treatment of large tracheal defects. A successful tracheal replacement must (1) have radial rigidity to prevent airway collapse during respiration, (2) contain an immunoprotective respiratory epithelium, and (3) integrate with the host vasculature to support epithelium viability. Herein, biopolymer microspheres are used to deliver chondrogenic growth factors to human mesenchymal stem cells (hMSCs) seeded in a custom mold that self‐assemble into cartilage rings, which can be fused into tubes. These rings and tubes can be fabricated with tunable wall thicknesses and lumen diameters with promising mechanical properties for airway collapse prevention. Epithelialized cartilage is developed by establishing a spatially defined composite tissue composed of human epithelial cells on the surface of an hMSC‐derived cartilage sheet. Prevascular rings comprised of human umbilical vein endothelial cells and hMSCs are fused with cartilage rings to form prevascular–cartilage composite tubes, which are then coated with human epithelial cells, forming a tri‐tissue construct. When prevascular– cartilage tubes are implanted subcutaneously in mice, the prevascular structures anastomose with host vasculature, demonstrated by their ability to be perfused. This microparticle–cell self‐assembly strategy is promising for engineering complex tissues such as a multi‐tissue composite trachea.