Vascularized bone tissue engineering: approaches for potential improvement.

Significant advances have been made in bone tissue engineering (TE) in the past decade. However, classical bone TE strategies have been hampered mainly due to the lack of vascularization within the engineered bone constructs, resulting in poor implant survival and integration. In an effort toward clinical success of engineered constructs, new TE concepts have arisen to develop bone substitutes that potentially mimic native bone tissue structure and function. Large tissue replacements have failed in the past due to the slow penetration of the host vasculature, leading to necrosis at the central region of the engineered tissues. For this reason, multiple microscale strategies have been developed to induce and incorporate vascular networks within engineered bone constructs before implantation in order to achieve successful integration with the host tissue. Previous attempts to engineer vascularized bone tissue only focused on the effect of a single component among the three main components of TE (scaffold, cells, or signaling cues) and have only achieved limited success. However, with efforts to improve the engineered bone tissue substitutes, bone TE approaches have become more complex by combining multiple strategies simultaneously. The driving force behind combining various TE strategies is to produce bone replacements that more closely recapitulate human physiology. Here, we review and discuss the limitations of current bone TE approaches and possible strategies to improve vascularization in bone tissue substitutes.

Progenitor cells: role and usage in bone tissue engineering approaches for spinal fusion.

Advancement of in vitro osteogenesis, or the production of bone, is a complex process that has significant clinical implications. Surgical intervention of several spinal disorders entails decompression of the spinal cord and nerves which can lead to subsequent biomechanical instability of the spine. Spinal arthrodesis (fusion) is often required to correct this instability and necessary to eliminate the resulting pathological motion of vertebral segments. Therefore, the achievement of proper spinal fusion, is a critical determinant of treatment efficacy. This chapter focuses on the molecular and cellular components that are involved in bone growth and healing. Mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) are the precursor cells essential for the formation of the five different types of bone cells: osteoprogenitor cells, osteoblasts, osteoclasts, osteocytes and lining cells. Similarly, endothelial progenitor cells (EPCs) differentiate into endothelial cells, which are essential in angiogenesis and neovascularization. MSCs tri-lineage potential (osteogenic, chondrogenic and adipogenic lineages) have made them the focus of most experimental approaches. Here, we describe their individual roles, as well as pose novel concepts on how their collective role may be the optimal strategy to improve upon in vitro osteogenesis and whether this could also be translated to improved bone formation in vivo. Further, we discuss the various molecular markers that are available for cell identification and the tissue engineering strategies that could replicate the osteoinductive, osteoconductive and osteoproductive milieuthat is available in autograft. Finally, we present a broad primer on the possible integration of cellular, molecular and tissue engineering strategies to improve osteogenesis and the future trends that may bring the promise seen in the laboratory to fruition in preclinical animal models.

Engineering articular cartilage with spatially-varying matrix composition and mechanical properties from a single stem cell population using a multi-layered hydrogel.

Despite significant advances in stem cell differentiation and tissue engineering, directing progenitor cells into three-dimensionally (3D) organized, native-like complex structures with spatially-varying mechanical properties and extra-cellular matrix (ECM) composition has not yet been achieved. The key innovations needed to achieve this would involve methods for directing a single stem cell population into multiple, spatially distinct phenotypes or lineages within a 3D scaffold structure. We have previously shown that specific combinations of natural and synthetic biomaterials can direct marrow-derived stem cells (MSC) into varying phenotypes of chondrocytes that resemble cells from the superficial, transitional, and deep zones of articular cartilage. In this current study, we demonstrate that layer-by-layer organization of these specific biomaterial compositions creates 3D niches that allow a single MSC population to differentiate into zone-specific chondrocytes and organize into a complex tissue structure. Our results indicate that a three-layer polyethylene glycol (PEG)-based hydrogel with chondroitin sulfate (CS) and matrix metalloproteinase-sensitive peptides (MMP-pep) incorporated into the top layer (superficial zone, PEG:CS:MMP-pep), CS incorporated into the middle layer (transitional zone, PEG:CS) and hyaluronic acid incorporated in the bottom layer (deep zone, PEG:HA), creates native-like articular cartilage with spatially-varying mechanical and biochemical properties. Specifically, collagen II levels decreased gradually from the superficial to the deep zone, while collagen X and proteoglycan levels increased, leading to an increasing gradient of compressive modulus from the superficial to the deep zone. We conclude that spatially-varying biomaterial compositions within single 3D scaffolds can stimulate efficient regeneration of multi-layered complex tissues from a single stem cell population.

Unique biomaterial compositions direct bone marrow stem cells into specific chondrocytic phenotypes corresponding to the various zones of articular cartilage.

Numerous studies have reported generation of cartilage-like tissue from chondrocytes and stem cells, using pellet cultures, bioreactors and various biomaterials, especially hydrogels. However, one of the primary unsolved challenges in the field has been the inability to produce tissue that mimics the highly organized zonal architecture of articular cartilage; specifically its spatially varying mechanical properties and extra-cellular matrix (ECM) composition. Here we show that different combinations of synthetic and natural biopolymers create unique niches that can "direct" a single marrow stem cell (MSC) population to differentiate into the superficial, transitional, or deep zones of articular cartilage. Specifically, incorporating chondroitin sulfate (CS) and matrix metalloproteinase-sensitive peptides (MMP-pep) into PEG hydrogels (PEG:CS:MMP-pep) induced high levels of collagen II and low levels of proteoglycan expression resulting in a low compressive modulus, similar to the superficial zone. PEG:CS hydrogels produced intermediate-levels of both collagen II and proteoglycans, like the transitional zone, while PEG:hyaluronic acid (HA) hydrogels induced high proteoglycan and low collagen II levels leading to high compressive modulus, similar to the deep zone. Additionally, the compressive moduli of these zone-specific matrices following cartilage generation showed similar trend as the corresponding zones of articular cartilage, with PEG:CS:MMP-pep having the lowest compressive modulus, followed by PEG:CS while PEG:HA had the highest modulus. These results underscore the potential for composite scaffold structures incorporating these biomaterial compositions such that a single stem-progenitor cell population can give rise to zonally-organized, functional articular cartilage-like tissue.