Adam B. Nover

Silk microfiber-reinforced silk hydrogel composites for functional cartilage tissue repair

Cartilage tissue lacks an intrinsic capacity for self-regeneration due to slow matrix turnover, a limited supply of mature chondrocytes and insufficient vasculature. Although cartilage tissue engineering has achieved some success using agarose as a scaffolding material, major challenges of agarose-based cartilage repair, including non-degradability, poor tissue-scaffold integration and limited processing capability, have prompted the search for an alternative biomaterial. In this study, silk fiber-hydrogel composites (SF-silk hydrogels) made from silk microfibers and silk hydrogels were investigated for their potential use as a support material for engineered cartilage. We demonstrated the use of 100% silk-based fiber-hydrogel composite scaffolds for the development of cartilage constructs with properties comparable to those made with agarose. Cartilage constructs with an equilibrium modulus in the native tissue range were fabricated by mimicking the collagen fiber and proteoglycan composite architecture of native cartilage using biocompatible, biodegradable silk fibroin from Bombyx mori. Excellent chondrocyte response was observed on SF-silk hydrogels, and fiber reinforcement resulted in the development of more mechanically robust constructs after 42 days in culture compared to silk hydrogels alone. Thus, we demonstrate the versatility of silk fibroin as a composite scaffolding material for use in cartilage tissue repair to create functional cartilage constructs that overcome the limitations of agarose biomaterials, and provide a much-needed alternative to the agarose standard.

Modern breast cancer detection: a technological review

Breast cancer is a serious threat worldwide and is the number two killer of women in the United States. The key to successful management is screening and early detection. What follows is a description of the state of the art in screening and detection for breast cancer as well as a discussion of new and emerging technologies. This paper aims to serve as a starting point for those who are not acquainted with this growing field.

Microbubbles as Biocompatible Porogens for Hydrogel Scaffolds

In this study, we explored the application of lipid-shelled, gas-filled microbubbles as a method for creating on-demand microporous hydrogels for cartilage tissue engineering. The technique allowed for homogenous distribution of cells and micropores within the scaffold, increasing the absorption coefficient of large solutes (70kDa dextran) over controls in a concentration-dependent manner. The stability of the gas phase of the microbubbles depended on several factors, including the initial size distribution of the microbubble suspension, as well as the temperature and pressure during culture. Application of pressure cycles provided controlled release of the gas phase to generate fluid-filled micropores with remnant lipid. The resulting microporous agarose scaffolds were biocompatible, leading to a twofold increase in engineered cartilage properties (E(Y)=492±42kPa for the bubble group vs. 249±49kPa for the bubble-free control group) over a 42-day culture period. Our results suggest that microbubbles offer a simple and robust method of modulating mass transfer in cell-seeded hydrogels through mild pressurization, and the methodology may be expanded in the future to include focused ultrasound for improved spatio-temporal control.

Porous titanium bases for osteochondral tissue engineering.

UNLABELLED Tissue engineering of osteochondral grafts may offer a cell-based alternative to native allografts, which are in short supply. Previous studies promote the fabrication of grafts consisting of a viable cell-seeded hydrogel integrated atop a porous, bone-like metal. Advantages of the manufacturing process have led to the evaluation of porous titanium as the bone-like base material. Here, porous titanium was shown to support the growth of cartilage to produce native levels of Youngs modulus, using a clinically relevant cell source. Mechanical and biochemical properties were similar or higher for the osteochondral constructs compared to chondral-only controls. Further investigation into the mechanical influence of the base on the composite material suggests that underlying pores may decrease interstitial fluid pressurization and applied strains, which may be overcome by alterations to the base structure. Future studies aim to optimize titanium-based tissue engineered osteochondral constructs to best match the structural architecture and strength of native grafts. STATEMENT OF SIGNIFICANCE The studies described in this manuscript follow up on previous studies from our lab pertaining to the fabrication of osteochondral grafts that consist of a bone-like porous metal and a chondrocyte-seeded hydrogel. Here, tissue engineered osteochondral grafts were cultured to native stiffness using adult chondrocytes, a clinically relevant cell source, and a porous titanium base, a material currently used in clinical implants. This porous titanium is manufactured via selective laser melting, offering the advantages of precise control over shape, pore size, and orientation. Additionally, this manuscript describes the mechanical influence of the porous base, which may have applicability to porous bases derived from other materials.

A puzzle assembly strategy for fabrication of large engineered cartilage tissue constructs

Engineering of large articular cartilage tissue constructs remains a challenge as tissue growth is limited by nutrient diffusion. Here, a novel strategy is investigated, generating large constructs through the assembly of individually cultured, interlocking, smaller puzzle-shaped subunits. These constructs can be engineered consistently with more desirable mechanical and biochemical properties than larger constructs (~4-fold greater Young׳s modulus). A failure testing technique was developed to evaluate the physiologic functionality of constructs, which were cultured as individual subunits for 28 days, then assembled and cultured for an additional 21-35 days. Assembled puzzle constructs withstood large deformations (40-50% compressive strain) prior to failure. Their ability to withstand physiologic loads may be enhanced by increases in subunit strength and assembled culture time. A nude mouse model was utilized to show biocompatibility and fusion of assembled puzzle pieces in vivo. Overall, the technique offers a novel, effective approach to scaling up engineered tissues and may be combined with other techniques and/or applied to the engineering of other tissues. Future studies will aim to optimize this system in an effort to engineer and integrate robust subunits to fill large defects.

Long-term storage and preservation of tissue engineered articular cartilage

With limited availability of osteochondral allografts, tissue engineered cartilage grafts may provide an alternative treatment for large cartilage defects. An effective storage protocol will be critical for translating this technology to clinical use. The purpose of this study was to evaluate the efficacy of the Missouri Osteochondral Allograft Preservation System (MOPS) for room temperature storage of mature tissue engineered grafts, focusing on tissue property maintenance during the current allograft storage window (28 days). Additional research compares MOPS to continued culture, investigates temperature influence, and examines longer‐term storage. Articular cartilage constructs were cultured to maturity using adult canine chondrocytes, then preserved with MOPS at room temperature, in refrigeration, or kept in culture for an additional 56 days. MOPS storage maintained desired chondrocyte viability for 28 days of room temperature storage, retaining 75% of the maturity point Youngs modulus without significant decline in biochemical content. Properties dropped past this time point. Refrigeration maintained properties similar to room temperature at 28 days, but proved better at 56 days. For engineered grafts, MOPS maintained the majority of tissue properties for the 28‐day window without clearly extending that period as it had for native grafts. These results are the first evaluating engineered cartilage storage.

High intensity focused ultrasound as a tool for tissue engineering: Application to cartilage

This article promotes the use of High Intensity Focused Ultrasound (HIFU) as a tool for affecting the local properties of tissue engineered constructs in vitro. HIFU is a low cost, non-invasive technique used for eliciting focal thermal elevations at variable depths within tissues. HIFU can be used to denature proteins within constructs, leading to decreased permeability and potentially increased local stiffness. Adverse cell viability effects remain restricted to the affected area. The methods described in this article are explored through the scope of articular cartilage tissue engineering and the fabrication of osteochondral constructs, but may be applied to the engineering of a variety of different tissues.

Characterization of Depth-Dependent Mechanical Properties in Bio-Titanium Hybrid Osteochondral Tissue Engineered Constructs

With cartilage autografts and allografts in short supply, tissue engineered osteochondral (OC) grafts offer an alternative [1]. These constructs are comprised of a chondrocyte-seeded hydrogel region and a porous, bone-like base. Our laboratory has shown growth of more robust osteochondral constructs on clinically-relevant metal substrates (eg. tantalum) as opposed to devitalized bone, and these constructs have been evaluated in vivo [1,2]. Due to the presence of the base, it is expected that transport of nutrients and chemical factors in OC constructs will differ from transport in chondral-only constructs (Fig. 1, bottom-left). Depth-dependent mechanical properties of chondral-only constructs have been measured, yielding a “U-shaped” strain profile, in which the construct is stiffest on the edges and softest in the center. However, depth-dependent properties have not been measured in tissue engineered OC grafts [3].Copyright

Chondroitinase-ABC Digestion and Dynamic Loading Increase Tension-Compression Nonlinearity in Tissue-Engineered Cartilage

Native articular cartilage exhibits tension-compression nonlinearity (TCN), where the compressive modulus is lower than its relatively high tensile modulus [1–2]. TCN produces in restricted lateral expansion of the tissue upon axial compression. We previously demnostrated that osmotic swelling can be used to measure the TCN of engineered cartilage by placing the tissue in an initial state of tensile strain. Incremental application of compression can be used to study the tissue’s mechanical properties as it transitions from tension to compression [3]. Although engineered cartilage is able to achieve the Young’s modulus (EY) and glycosaminoglycan (GAG) content of native tissue, the collagen content and dynamic modulus (G*) consistently underperform the native tissue. Removing GAG with chondroitinase ABC (cABC) has been shown to significantly decrease the tissue properties immediately after digestion but the properties rebound, with improved collagen content and G* compared to undigested controls [4]. Furthermore, we have previously shown that cABC digestion significantly increases TCN in engineered cartilage [3]. Dynamic loading (DL) has been shown to significantly increase the mechanical properties without significantly altering biochemical composition of engineered cartilage, however the mechanism through which DL modulates the mechanical strength of engineered cartilage may be due in part to improved extracellular matrix (ECM) organization [5]. We therefore hypothesize that cABC digestion and DL will improve the tensile properties of engineered cartilage.Copyright

Prolonged Treatment of Ultra-Low Dose Chondroitinase ABC Improves Matrix Production in Engineered Cartilage

Articular cartilage is the load bearing soft tissue of diarthrodial joints, and mechanical loading maintains the integrity of the tissue. The predominant extracellular matrix constituents, proteoglycans and collagen, allow cartilage to support the high compressive and tensile loads experienced in diurnal loading. Our laboratory has been successful in cultivating engineered cartilage constructs with a compressive equilibrium modulus and glycosaminglycan (GAG) content near native values [1, 2]. Many approaches to cultivating engineered cartilage have been limited by low collagen production in vitro, an impediment for attaining native functional load-bearing properties [3].© 2013 ASME