Browsing by Author "Santoro, Marco"

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    3D tissue-engineered model of Ewing's sarcoma
    (Elsevier, 2014) Lamhamedi-Cherradi, Salah-Eddine; Santoro, Marco; Ramammoorthy, Vandhana; Menegaz, Brian A.; Bartholomeusz, Geoffrey; Iles, Lakesla R.; Amin, Hesham M.; Livingston, J. Andrew; Mikos, Antonios G.; Ludwig, Joseph A.; Bioengineering
    Despite longstanding reliance upon monolayer culture for studying cancer cells, and numerous advantages from both a practical and experimental standpoint, a growing body of evidence suggests that more complex three-dimensional (3D) models are necessary to properly mimic many of the critical hallmarks associated with the oncogenesis, maintenance and spread of Ewing's sarcoma (ES), the second most common pediatric bone tumor. And as clinicians increasingly turn to biologically-targeted therapies that exert their effects not only on the tumor cells themselves, but also on the surrounding extracellular matrix, it is especially important that preclinical models evolve in parallel to reliably measure antineoplastic effects and possible mechanisms of de novo and acquired drug resistance. Herein, we highlight a number of innovative methods used to fabricate biomimetic ES tumors, encompassing both the surrounding cellular milieu and the extracellular matrix (ECM), and suggest potential applications to advance our understanding of ES biology, preclinical drug testing, and personalized medicine.
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    Development of a High-Throughput 3D Tumor Model for Bone Sarcomas
    (2016-10-21) Santoro, Marco; Mikos, Antonios G; Ludwig, Joseph A
    Preclinical drug testing commonly relies on the use of two-dimensional (2D) cultures, which allow for rapid drug screening in vitro. However, 2D cultures are unable to capture the complexity of the native three-dimensional (3D) tumor microenvironment, resulting in a lack of correspondence between preclinical data and clinical trial outcomes. The establishment of high-throughput 3D models able to describe distinctive aspects of the tumor niche would advance our understanding of tumor biology and would allow for drug testing in a physiologically relevant setup. Along this rationale, this thesis focuses on the development of a high-throughput 3D tumor model of bone sarcoma based on tissue-engineered polymeric scaffolds in combination with a flow perfusion bioreactor. First, we investigated the effects of flow perfusion on a 3D culture of Ewing sarcoma (ES) cells. We found that increasing levels of flow-derived shear stress promoted the secretion of insulin-like growth factor-1 (IGF-1) which, in turn, resulted in shear stress-dependent cell sensitivity to the IGF-1 receptor (IGF-1R) blockade, a central player in ES progression. We then leveraged these findings to culture ES cells on 3D-printed scaffolds under flow perfusion conditions. By designing 3D scaffolds with a defined porosity gradient, ES cells were exposed to a shear stress gradient that resulted in a gradient in cell response. In this way we sought to model variable levels of shear stress present within ES tumors due to intratumoral heterogeneity. In the final part of this thesis we investigated how the simultaneous presence of mesenchymal stem cells (MSCs) and flow perfusion affected drug sensitivity and phenotype of ES cells. We showed that the presence of MSCs within the coculture induces a progressive inhibition of cell growth and resistance to the IGF-1R blockade, highlighting the role of mechanically-sensitive mesodermal stroma on ES drug resistance. Overall, in this thesis we present a tissue-engineered tumor model that reliably mimics key features of the bone microenvironment, specifically the effects of biomechanical stimulation and of tumor-stroma interactions. The model hereby developed is amenable to further mechanistic studies on tumor biology and allows for a more accurate high-throughput screening of novel drug candidates.
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    Direct and indirect co-culture of chondrocytes and mesenchymal stem cells for the generation of polymer/extracellular matrix hybrid constructs
    (Elsevier, 2014) Levorson, Erica J.; Santoro, Marco; Kasper, F. Kurtis; Mikos, Antonios G.; Bioengineering; Chemical and Biomolecular Engineering
    In this work, the influence of direct cell–cell contact in co-cultures of mesenchymal stem cells (MSCs) and chondrocytes for the improved deposition of cartilage-like extracellular matrix (ECM) within nonwoven fibrous poly(∊-caprolactone) (PCL) scaffolds was examined. To this end, chondrocytes and MSCs were either co-cultured in direct contact by mixing on a single PCL scaffold or produced via indirect co-culture, whereby the two cell types were seeded on separate scaffolds which were then cultured together in the same system either statically or under media perfusion in a bioreactor. In static cultures, the chondrocyte scaffold of an indirectly co-cultured group generated significantly greater amounts of glycosaminoglycan and collagen than the direct co-culture group initially seeded with the same number of chondrocytes. Furthermore, improved ECM production was linked to greater cellular proliferation and distribution throughout the scaffold in static culture. In perfusion cultures, flow had a significant effect on the proliferation of the chondrocytes. The ECM contents within the chondrocyte-containing scaffolds of the indirect co-culture groups either approximated or surpassed the amounts generated within the direct co-culture group. Additionally, within bioreactor culture there were indications that chondrocytes had an influence on the chondrogenesis of MSCs as evidenced by increases in cartilaginous ECM synthetic capacity. This work demonstrates that it is possible to generate PCL/ECM hybrid scaffolds for cartilage regeneration by utilizing the factors secreted by two different cell types, chondrocytes and MSCs, even in the absence of juxtacrine signaling.
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    Gelatin carriers for drug and cell delivery in tissue engineering
    (Elsevier, 2014) Santoro, Marco; Tatara, Alexander M.; Mikos, Antonios G.; Bioengineering; Chemical and Biomolecular Engineering
    The ability of gelatin to form complexes with different drugs has been investigated for controlled release applications. Gelatin parameters, such as crosslinking density and isoelectric point, have been tuned in order to optimize gelatin degradation and drug delivery kinetics. In recent years, focus has shifted away from the use of gelatin in isolation toward the modification of gelatin with functional groups and the fabrication of material composites with embedded gelatin carriers. In this review, we highlight some of the latest work being performed in these areas and comment on trends in the field. Specifically, we discuss gelatin modifications for immune system evasion, drug stabilization, and targeted delivery, as well as gelatin composite systems based on ceramics, naturally-occurring polymers, and synthetic polymers.