@article{mora-navarro_garcia_sarker_ozpinar_enders_khan_branski_freytes_2022, title={Monitoring decellularization via absorbance spectroscopy during the derivation of extracellular matrix scaffolds}, volume={17}, ISSN={["1748-605X"]}, url={https://doi.org/10.1088/1748-605X/ac361f}, DOI={10.1088/1748-605X/ac361f}, abstractNote={Abstract
               Extracellular matrix (ECM) is a complex structure composed of bioactive molecules representative of the local tissue microenvironment. Decellularized ECM biomaterials harness these biomolecules for regenerative medicine applications. One potential therapeutic application is the use of vocal fold (VF) specific ECM to restore the VFs after injury. ECM scaffolds are derived through a process of decellularization, which aims to remove unwanted immunogenic biomolecules (e.g. DNA) while preserving the composition of the ECM. The effectiveness of the decellularization is typically assessed at the end by quantifying ECM attributes such as final dsDNA content. However, batch-to-batch variability in ECM manufacturing remains a significant challenge for the standardization, cost-effectiveness, and scale-up process. The limited number of tools available for in-process control heavily restricts the uncovering of the correlations between decellularization process parameters and ECM attributes. In this study, we developed a technique applicable to both the classical batch method and semi-continuous decellularization systems to trace the decellularization of two laryngeal tissues in real-time. We hypothesize that monitoring the bioreactor’s effluent absorbance at 260 nm as a function of time will provide a representative DNA release profile from the tissue and thus allow for process optimization. The DNA release profiles were obtained for laryngeal tissues and were successfully used to optimize the derivation of VF lamina propria-ECM (auVF-ECM) hydrogels. This hydrogel had comparable rheological properties to commonly used biomaterials to treat VF injuries. Also, the auVF-ECM hydrogel promoted the down-regulation of CCR7 by THP-1 macrophages upon lipopolysaccharide stimulation in vitro suggesting some anti-inflammatory properties. The results show that absorbance profiles are a good representation of DNA removal during the decellularization process thus providing an important tool to optimize future protocols.}, number={1}, journal={BIOMEDICAL MATERIALS}, publisher={IOP Publishing}, author={Mora-Navarro, Camilo and Garcia, Mario E. and Sarker, Prottasha and Ozpinar, Emily W. and Enders, Jeffrey R. and Khan, Saad and Branski, Ryan C. and Freytes, Donald O.}, year={2022}, month={Jan} }
 @article{mora-navarro_ozpinar_sze_martin_freytes_2021, title={Transcriptome-targeted analysis of human peripheral blood-derived macrophages when cultured on biomaterial meshes}, volume={16}, ISSN={["1748-605X"]}, url={http://dx.doi.org/10.1088/1748-605x/abdbdb}, DOI={10.1088/1748-605x/abdbdb}, abstractNote={Abstract
               Surgical meshes are commonly used to repair defects and support soft tissues. Macrophages (Mφs) are critical cells in the wound healing process and are involved in the host response upon foreign biomaterials. There are various commercially available permanent and absorbable meshes used by surgeons for surgical interventions. Polypropylene (PP) meshes represent a permanent biomaterial that can elicit both inflammatory and anti-inflammatory responses. In contrast, poly-4-hydroxybutyrate (P4HB) based meshes are absorbable and linked to positive clinical outcomes but have a poorly characterized immune response. This study evaluated the in vitro targeted transcriptomic response of human Mφs seeded for 48 h on PP and P4HB surgical meshes. The in vitro measured response from human Mφs cultured on P4HB exhibited inflammatory and anti-inflammatory gene expression profiles typically associated with wound healing, which aligns with in vivo animal studies from literature. The work herein provides in vitro evidence for the early transcriptomic targeted signature of human Mφs upon two commonly used surgical meshes. The findings suggest a transition from an inflammatory to a non-inflammatory phenotype by P4HB as well as an upregulation of genes annotated under the pathogen response pathway.}, number={2}, journal={BIOMEDICAL MATERIALS}, publisher={IOP Publishing}, author={Mora-Navarro, Camilo and Ozpinar, Emily W. and Sze, Daphne and Martin, David P. and Freytes, Donald O.}, year={2021}, month={Mar} }
 @article{huang_ozpinar_su_tang_shen_qiao_hu_li_liang_mathews_et al._2020, title={An off-the-shelf artificial cardiac patch improves cardiac repair after myocardial infarction in rats and pigs}, volume={12}, url={http://dx.doi.org/10.1126/scitranslmed.aat9683}, DOI={10.1126/scitranslmed.aat9683}, abstractNote={Acellular patches embedded with encapsulated cell-secreted factors improve cardiac repair after acute myocardial infarction in rodents and pigs.}, number={538}, journal={Science Translational Medicine}, publisher={American Association for the Advancement of Science (AAAS)}, author={Huang, Ke and Ozpinar, Emily W. and Su, Teng and Tang, Junnan and Shen, Deliang and Qiao, Li and Hu, Shiqi and Li, Zhenhua and Liang, Hongxia and Mathews, Kyle and et al.}, year={2020}, month={Apr} }
 @article{ozpinar_frey_arthur_mora-navarro_biehl_snider_cruse_freytes_2021, title={Dermal Extracellular Matrix-Derived Hydrogels as an In Vitro Substrate to Study Mast Cell Maturation}, volume={27}, ISSN={["1937-335X"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85110277858&partnerID=MN8TOARS}, DOI={10.1089/ten.tea.2020.0142}, abstractNote={Mast cells (MCs) are pro-inflammatory tissue-resident immune cells that play a key role in inflammation. MCs circulate in peripheral blood as progenitors and undergo terminal differentiation in the tissue microenvironment where they can remain for many years. This in situ maturation results in tissue- and species-specific MC phenotypes, culminating in significant variability in response to environmental stimuli. There are many challenges associated with studying mature tissue-derived MCs, particularly in humans. In cases where cultured MCs are able to differentiate in two-dimensional in vitro cultures, there remains an inability for full maturation. Extracellular matrix (ECM) scaffolds provide for a more physiologically relevant environment for cells in vitro and have been shown to modulate the response of other immune cells such as T cells, monocytes, and macrophages. To improve current in vitro testing platforms of MCs and to assess future use of ECM scaffolds for MC regulation, we studied the in vitro response of human MCs cultured on decellularized porcine dermis hydrogels (dermis extracellular matrix hydrogel [dECM-H]). This study investigated the effect of dECM-H on cellular metabolic activity, cell viability, and receptor expression compared to collagen type I hydrogel (Collagen-H). Human MCs showed different metabolic activity when cultured in the dECM-H and also upregulated immunoglobulin E (IgE) receptors associated with MC maturation/activation compared to collagen type I. These results suggest an overall benefit in the long-term culture of human MCs in the dECM-H compared to Collagen-H providing important steps toward a model that is more representative of in vivo conditions. Mast cells (MCs) are difficult to culture in vitro as current culture conditions and substrates fail to promote similar phenotypic features observed in vivo. Extracellular matrix (ECM)-based biomaterials offer three-dimensional, tissue-specific environments that more closely resemble in vivo conditions. Our study explores the use of dermal ECM hydrogels for MC culture and shows significant upregulation of metabolic activity, cell viability, and gene expression of markers associated with MC maturation or activation compared to collagen type I-hydrogel and tissue culture plastic controls at 7 days. These results are among the first to describe MC behavior in response to ECM hydrogels.}, number={15-16}, journal={TISSUE ENGINEERING PART A}, author={Ozpinar, Emily W. and Frey, Ariana L. and Arthur, Greer K. and Mora-Navarro, Camilo and Biehl, Andreea and Snider, Douglas B. and Cruse, Glenn and Freytes, Donald O.}, year={2021}, month={Aug}, pages={1008–1022} }
 @article{dermal extracellular matrix-derived hydrogels as an in vitro substrate to study mast cell maturation_2020, DOI={10.1089/ten.TEA.2020.0142.}, abstractNote={Mast cells (MCs) are pro-inflammatory tissue-resident immune cells that play a key role in inflammation. MCs circulate in peripheral blood as progenitors and undergo terminal differentiation in the tissue microenvironment where they can remain for many years. This  in situ  maturation results in tissue- and species-specific MC phenotypes, culminating in significant variability in response to environmental stimuli. There are many challenges associated with studying mature tissue-derived MCs, particularly in humans. In cases where cultured MCs are able to differentiate in two-dimensional  in vitro  cultures, there remains an inability for full maturation. Extracellular matrix (ECM) scaffolds provide for a more physiologically relevant environment for cells  in vitro  and have been shown to modulate the response of other immune cells such as T cells, monocytes, and macrophages. To improve current  in vitro  testing platforms of MCs and to assess future use of ECM scaffolds for MC regulation, we studied the  in vitro  response of human MCs cultured on decellularized porcine dermis hydrogels (dermis extracellular matrix hydrogel [dECM-H]). This study investigated the effect of dECM-H on cellular metabolic activity, cell viability, and receptor expression compared to collagen type I hydrogel (Collagen-H). Human MCs showed different metabolic activity when cultured in the dECM-H and also upregulated immunoglobulin E (IgE) receptors associated with MC maturation/activation compared to collagen type I. These results suggest an overall benefit in the long-term culture of human MCs in the dECM-H compared to Collagen-H providing important steps toward a model that is more representative of  in vivo  conditions. Graphical abstract [Formula: see text] Impact statement Mast cells (MCs) are difficult to culture  in vitro  as current culture conditions and substrates fail to promote similar phenotypic features observed  in vivo . Extracellular matrix (ECM)-based biomaterials offer three-dimensional, tissue-specific environments that more closely resemble  in vivo  conditions. Our study explores the use of dermal ECM hydrogels for MC culture and shows significant upregulation of metabolic activity, cell viability, and gene expression of markers associated with MC maturation or activation compared to collagen type I-hydrogel and tissue culture plastic controls at 7 days. These results are among the first to describe MC behavior in response to ECM hydrogels.}, journal={Tissue Engineering Part A}, year={2020}, month={Oct} }
 @article{badileanu_mora-navarro_martins_garcia_sze_ozpinar_gaffney_enders_branski_freytes_2020, title={Fast Automated Approach for the Derivation of Acellular Extracellular Matrix Scaffolds from Porcine Soft Tissues}, volume={6}, url={https://doi.org/10.1021/acsbiomaterials.0c00265}, DOI={10.1021/acsbiomaterials.0c00265}, abstractNote={Decellularized extracellular matrix (ECM) scaffolds derived from tissues and organs are complex biomaterials used in clinical and research applications. A number of decellularization protocols have been described for ECM biomaterials derivation, each adapted to a particular tissue and use, restricting comparisons among materials. One of the major sources of variability in ECM products comes from the tissue source and animal age. Although this variability could be minimized using established tissue sources, other sources arise from the decellularization process itself. Overall, current protocols require manual work and are poorly standardized with regard to the choice of reagents, the order by which they are added, and exposure times. The combination of these factors adds variability affecting the uniformity of the final product between batches. Furthermore, each protocol needs to be optimized for each tissue and tissue source making tissue-to-tissue comparisons difficult. Automation and standardization of ECM scaffold development constitute a significant improvement to current biomanufacturing techniques but remains poorly explored. This study aimed to develop a biofabrication method for fast and automated derivation of raw material for ECM hydrogel production while preserving ECM composition and controlling lot-to-lot variability. The main result was a closed semibatch bioreactor system with automated dosing of decellularization reagents capable of deriving ECM material from pretreated soft tissues. The ECM was further processed into hydrogels to demonstrate gelation and cytocompatibility. This work presents a versatile, scalable, and automated platform for the rapid production of ECM scaffolds.}, number={7}, journal={ACS Biomaterials Science & Engineering}, publisher={American Chemical Society (ACS)}, author={Badileanu, Andreea and Mora-Navarro, Camilo and Martins, Ana M. Gracioso and Garcia, Mario E. and Sze, Daphne and Ozpinar, Emily W. and Gaffney, Lewis and Enders, Jeffrey R. and Branski, Ryan C. and Freytes, Donald O.}, year={2020}, month={Jul}, pages={4200–4213} }
 @article{ozpinar_frey_cruse_freytes_2021, title={Mast Cell-Biomaterial Interactions and Tissue Repair}, volume={27}, ISSN={["1937-3376"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85122126010&partnerID=MN8TOARS}, DOI={10.1089/ten.teb.2020.0275}, abstractNote={Tissue engineers often use biomaterials to provide structural support along with mechanical and chemical signals to modulate the wound healing process. Biomaterials that are implanted into the body interact with a heterogeneous and dynamic inflammatory environment that is present at the site of injury. Whether synthetically-derived, naturally-derived, or a combination of both, it is important to assess biomaterials for their ability to modulate inflammation in order to understand their potential clinical use. One important, but under-explored cell in the context of biomaterials is the mast cell (MC). MCs are granulocytic leukocytes that engage in a variety of events in both the innate and adaptive immune systems. Though highly recognized for their roles in allergic reactions, MCs play an important role in wound healing by recognizing antigens through pattern recognition receptors and the high-affinity immunoglobulin E (IgE) receptor, FcεRI, and releasing granules that affect cell recruitment, fibrosis, extracellular matrix deposition, angiogenesis, and vasculogenesis. MCs also mediate the foreign body response, contributing to the incorporation or rejection of implants. Studies of MC-biomaterial interactions can aid in the elucidation of MC roles during the host tissue response. This review is designed for those in the tissue engineering and biomaterials fields who are interested in exploring the role MCs may play in wound-biomaterial interactions and wound healing. With this review, we hope to inspire more research in the MC-biomaterial space in order to accelerate the design and construction of optimized implants.}, number={6}, journal={TISSUE ENGINEERING PART B-REVIEWS}, author={Ozpinar, Emily W. and Frey, Ariana L. and Cruse, Glenn and Freytes, Donald O.}, year={2021}, month={Dec}, pages={590–603} }
 @article{mast cell-biomaterial interactions and tissue repair_2020, DOI={ten.TEB.2020.0275}, journal={Tissue Engineering Part B: Reviews}, year={2020}, month={Nov} }
 @article{mora-navarro_badileanu_martins_ozpinar_gaffney_huntress_harrell_enders_peng_branski_et al._2020, title={Porcine Vocal Fold Lamina Propria-Derived Biomaterials Modulate TGF-β1-Mediated Fibroblast Activation in Vitro}, volume={6}, url={http://dx.doi.org/10.1021/acsbiomaterials.9b01837}, DOI={10.1021/acsbiomaterials.9b01837}, abstractNote={The vocal fold lamina propria (VFLP), one of the outermost layers of the vocal fold (VF), is composed of tissue-specific extracellular matrix (ECM) proteins and is highly susceptible to injury. Various biomaterials have been clinically tested to treat voice disorders (e.g., hydrogels, fat, and hyaluronic acid), but satisfactory recovery of the VF functionality remains elusive. Fibrosis or scar formation in the VF is a major challenge, and the development and refinement of novel therapeutics that promote the healing and normal function of the VF are needed. Injectable hydrogels derived from native tissues have been previously reported with major advantages over synthetic hydrogels, including constructive tissue remodeling and reduced scar tissue formation. This study aims to characterize the composition of a decellularized porcine VFLP-ECM scaffold and the cytocompatibility and potential antifibrotic properties of a hydrogel derived from VFLP-ECM. In addition, we isolated potential matrix-bound vesicles (MBVs) and macromolecules from the VFLP-ECM that also downregulated smooth muscle actin ACTA2 under transforming growth factor-beta 1 (TGF-β1) stimulation. The results provide evidence of the unique protein composition of the VFLP-ECM and the potential link between the components of the VFLP-ECM and the inhibition of TGF-β1 signaling observed in vitro when transformed into injectable forms.}, number={3}, journal={ACS Biomaterials Science & Engineering}, publisher={American Chemical Society (ACS)}, author={Mora-Navarro, Camilo and Badileanu, Andreea and Martins, Ana M. Gracioso and Ozpinar, Emily W. and Gaffney, Lewis and Huntress, Ian and Harrell, Erin and Enders, Jeffrey R. and Peng, Xinxia and Branski, Ryan C. and et al.}, year={2020}, month={Feb}, pages={1690–1703} }
 @article{effects of polarized macrophages on the in vitro gene expression after co-culture of human pluripotent stem cell-derived cardiomyocytes_2019, url={http://dx.doi.org/10.1016/j.regen.2019.100018}, DOI={10.1016/j.regen.2019.100018}, abstractNote={A promising approach to rescue cardiac function after a myocardial infarction (MI) is to apply an engineered heart tissue (EHT) onto the infarcted area. After the onset of MI, a dynamic inflammatory environment develops comprising of the temporal recruitment of macrophages (Mϕs), and their interactions with the cells of the damaged myocardium. There is limited knowledge about the interactions between this inflammatory environment and the cells that could potentially be used to create an EHT, such as pluripotent stem cell derived-cardiomyocytes. In the present study, a cell-based system was used to study the in vitro interactions between lipopolysaccharide (LPS) and interferon-gamma (IFNγ)-activated Mϕs, and interleukin 4 (IL4) and interleukin 13 (IL13)-activated Mϕs and human embryonic stem cell-derived cardiomyocytes (hESC-CMs). Using a co-culture system, gene expression profiles of key markers of both the Mϕs and the hESC-CMs were obtained, as well as the protein secretion. Additionally, the effects of Mϕ polarizing cytokines on hESC-CMs with or without the presence of Mϕs were studied. Mϕs co-cultured with hESC-CMs showed no significant changes in their gene expression profile after two days in culture. hESC-CMs, however, were noted to have an overall decrease in expression of cardiac-related genes upon exposure to both Mϕ subtypes in co-culture. Gene expression of Bone morphogenetic protein-2 (BMP2), Bone morphogenetic protein-4 (BMP4) and GATA-binding protein-4 (GATA4) were also affected by Mϕ exposure and by inflammatory signals such as LPS and IFNγ. This study represents an important step towards the design of advanced in vitro testing platforms to further study the effect of Mϕs and inflammatory signals on EHTs in vitro.}, journal={Journal of Immunology and Regenerative Medicine}, year={2019}, month={Jun} }
 @article{liang_huang_su_li_hu_dinh_wrona_shao_qiao_vandergriff_et al._2018, title={Mesenchymal Stem Cell/Red Blood Cell-Inspired Nanoparticle Therapy in Mice with Carbon Tetrachloride-Induced Acute Liver Failure}, volume={12}, ISSN={1936-0851 1936-086X}, url={http://dx.doi.org/10.1021/ACSNANO.8B00553}, DOI={10.1021/acsnano.8b00553}, abstractNote={Acute liver failure is a critical condition characterized by global hepatocyte death and often time needs a liver transplantation. Such treatment is largely limited by donor organ shortage. Stem cell therapy offers a promising option to patients with acute liver failure. Yet, therapeutic efficacy and feasibility are hindered by delivery route and storage instability of live cell products. We fabricated a nanoparticle that carries the beneficial regenerative factors from mesenchymal stem cells and further coated it with the membranes of red blood cells to increase blood stability. Unlike uncoated nanoparticles, these particles promote liver cell proliferation in vitro and have lower internalization by macrophage cells. After intravenous delivery, these artificial stem cell analogs are able to remain in the liver and mitigate carbon tetrachloride-induced liver failure in a mouse model, as gauged by histology and liver function test. Our technology provides an innovative and off-the-shelf strategy to treat liver failure.}, number={7}, journal={ACS Nano}, publisher={American Chemical Society (ACS)}, author={Liang, Hongxia and Huang, Ke and Su, Teng and Li, Zhenhua and Hu, Shiqi and Dinh, Phuong-Uyen and Wrona, Emily A. and Shao, Chen and Qiao, Li and Vandergriff, Adam C. and et al.}, year={2018}, month={Jun}, pages={6536–6544} }
 @article{gaffney_warren_wrona_fisher_freytes_2017, title={Macrophages' role in tissue disease and regeneration}, volume={62}, journal={Macrophages: origin, functions and biointervention}, author={Gaffney, L. and Warren, P. and Wrona, E. A. and Fisher, M. B. and Freytes, D. O.}, year={2017}, pages={245–271} }
 @inbook{gaffney_warren_wrona_fisher_freytes_2017, title={Macrophages’ role in tissue disease and regeneration}, volume={62}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85019009634&partnerID=MN8TOARS}, DOI={10.1007/978-3-319-54090-0_10}, abstractNote={Inflammation is an essential component of the normal mammalian host tissue response and plays an important role during cardiovascular and musculoskeletal diseases. Given the important role of inflammation on the host tissue response after injury, understanding this process represents essential aspects of biomedical research, tissue engineering, and regenerative medicine. Macrophages are central players during the inflammatory response with an extensive role during wound healing. These cells exhibit a spectrum of activation states that span from pro-inflammatory to pro-healing phenotypes. The phenotype of the macrophages can have profound influences on the progression of disease or injury. As such, understanding and subsequent modulation of macrophage phenotype represents an exciting target area for regenerative medicine therapies. In this chapter, we describe the role of macrophages in specific cases of injury and disease. After myocardial infarction, a biphasic response of pro- and anti-inflammatory macrophages are involved in the remodeling process. In volumetric muscle loss, there is an intricate communication between inflammatory cells and progenitor cells affecting repair processes. Osteoarthritis is characterized by increased levels of pro-inflammatory macrophages over an extended period of time with significant impact on the progression of the disease. By harnessing the complex role of macrophages, enhanced therapeutic treatments can be developed that enhance the normal healing response as well as help the survival of therapeutic cells delivered to the site of injury.}, booktitle={Results and Problems in Cell Differentiation}, author={Gaffney, L. and Warren, P. and Wrona, E.A. and Fisher, M.B. and Freytes, D.O.}, year={2017}, pages={245–271} }
 @misc{gaffney_wrona_freytes_2018, title={Potential Synergistic Effects of Stem Cells and Extracellular Matrix Scaffolds}, volume={4}, ISSN={["2373-9878"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85045213284&partnerID=MN8TOARS}, DOI={10.1021/acsbiomaterials.7b00083}, abstractNote={In recent years, extracellular matrix (ECM)-derived biomaterials have been used as scaffolds to help regenerate diseased or damaged tissues. These biomaterials are prepared by decellularization of a tissue of interest by chemical or physical removal of the cellular components. The goal of the decellularization process is to remove cells without disturbing tissue-specific composition, growth factor content, and, in some cases, the mechanical properties. As decellularization can be achieved without significantly affecting the native architecture of the tissue or organ of interest, it provides a scaffold material with native-like composition and structure. ECM scaffolds promote constructive remodeling through several mechanisms that include chemotactic properties, growth factor release, and modulation of the immune response. Constructive remodeling by ECM scaffolds relies, in part, on the recruitment of neighboring or circulating cells to the wound site. However, this is a relatively lengthy process, and the...}, number={4}, journal={ACS BIOMATERIALS SCIENCE & ENGINEERING}, author={Gaffney, Lewis and Wrona, Emily A. and Freytes, Donald O.}, year={2018}, month={Apr}, pages={1208–1222} }
 @misc{wrona_peng_amin_branski_freytes_2016, title={Extracellular Matrix for Vocal Fold Lamina Propria Replacement: A Review}, volume={22}, ISSN={["1937-3376"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85020490144&partnerID=MN8TOARS}, DOI={10.1089/ten.teb.2016.0015}, abstractNote={The vocal folds (VFs) are exposed to a number of injurious stimuli that frequently lead to aberrant structural alterations and altered biomechanical properties that clinically manifest as voice disorders. Therapies to restore both structure and function of this delicate tissue are ideal. However, such methods have not been adequately developed. Our group and others hypothesize that tissue engineering and regenerative medicine approaches, previously described for other tissue systems, hold significant promise for the VFs. In this review, we explore the concept of tissue engineering as it relates to the VFs, as well as recent studies employing both naturally and synthetically derived biomaterials, including those from laryngeal and nonlaryngeal sources, in combination with stem cells for a tissue-engineered approach to VF repair.}, number={6}, journal={TISSUE ENGINEERING PART B-REVIEWS}, author={Wrona, Emily A. and Peng, Robert and Amin, Milan R. and Branski, Ryan C. and Freytes, Donald O.}, year={2016}, month={Dec}, pages={421–429} }
 @article{pallotta_sun_wrona_freytes_2017, title={BMP protein-mediated crosstalk between inflammatory cells and human pluripotent stem cell-derived cardiomyocytes}, volume={11}, ISSN={1932-6254}, url={http://dx.doi.org/10.1002/TERM.2045}, DOI={10.1002/term.2045}, abstractNote={Following cardiac injury, the ischaemic heart tissue is characterized by the invasion of pro‐inflammatory (M1) and pro‐healing (M2) macrophages. Any engineered cardiac tissue will inevitably interact with the inflammatory environment found at the site of myocardial infarction at the time of implantation. However, the interactions between the inflammatory and the cardiac repair cells remain poorly understood. Here we recapitulated in vitro some of the important cellular events found at the site of myocardial injury, such as macrophage recruitment and their effect on cardiac differentiation and maturation, by taking into account the involvement of paracrine‐mediated signalling. By using a 3D inverted invasion assay, we found that cardiomyocyte (CM) conditioned medium can trigger the recruitment of pro‐inflammatory (M1) macrophages, through a mechanism that involves, in part, CM‐derived BMP4. Pro‐inflammatory (M1) macrophages were also found to affect CM proliferation and differentiation potential, in part due to BMP molecules secreted by macrophages. These effects involved the activation of the canonical outside‐in signalling pathways, such as SMAD1,5,8, which are known to be activated during myocardial injury in vivo. In the present study we propose a new role for CM‐ and macrophage‐derived BMP proteins during the recruitment of macrophage subtypes and the maturation of repair cells, representing an important step towards creating a functional cardiac patch with superior therapeutic properties. Copyright © 2015 John Wiley & Sons, Ltd.}, number={5}, journal={Journal of Tissue Engineering and Regenerative Medicine}, publisher={Wiley}, author={Pallotta, Isabella and Sun, Bruce and Wrona, Emily A. and Freytes, Donald O.}, year={2017}, month={May}, pages={1466–1478} }
 @article{wrona_peng_born_amin_branski_freytes_2015, title={Derivation and characterization of porcine vocal fold extracellular matrix scaffold}, volume={126}, ISSN={0023-852X}, url={http://dx.doi.org/10.1002/LARY.25640}, DOI={10.1002/lary.25640}, abstractNote={Objectives/HypothesisTo optimize decellularization of porcine vocal folds (VF) and quantify human bone marrow‐derived mesenchymal stem cell (BM‐MSC) interactions with this matrix to provide a foundation for regenerative approaches to VF repair.Study Design and MethodsVocal folds were dissected from porcine larynges and three decellularization protocols were compared, each consisting of washes and mechanical agitations with different combinations of reagents. DNA content was analyzed via Quant‐iT Picogreen assay and hematoxylin and eosin staining. Bone marrow‐derived MSCs were then seeded onto the decellularized VF matrices. Morphology, metabolic activity, DNA content, and gene expression were assessed using LIVE/DEAD Cell Viability, alamarBlue Cell Viability Assay, Quant‐iT Picogreen assay, and quantitative polymerase chain reaction, respectively.ResultsThe most successful decellularization protocol removed 95% DNA content within 1 day, compared to several days required for previously described protocols. Histology confirmed the retention of extracellular matrix (ECM) and its components, including glycosaminoglycans, collagen, and fibrin, while void of nuclear/cellular content. Decellularized scaffolds were then seeded with BM‐MSCs. Similar DNA quantities were observed after 24 hours of seeding within the VF‐ECM scaffold when compared to cells on tissue culture plastic (TCP). LIVE/DEAD staining of the seeded VF‐ECM confirmed excellent cell viability, and the metabolic activity of BM‐MSCs increased significantly on VF‐ECM compared to TCP. Endoglin gene expression decreased, suggestive of differentiation.ConclusionPorcine VFs can be efficiently decellularized within 5 hours using a combination of sodium deoxycholate and peracetic acid. Decellularized VF‐ECM supported attachment and growth of human BM‐MSCs, with evidence of differentiation.Level of EvidenceN/A Laryngoscope, 126:928–935, 2016}, number={4}, journal={The Laryngoscope}, publisher={Wiley}, author={Wrona, Emily A. and Peng, Robert and Born, Hayley and Amin, Milan R. and Branski, Ryan C. and Freytes, Donald O.}, year={2015}, month={Sep}, pages={928–935} }
 @article{spiller_wrona_romero-torres_pallotta_graney_witherel_panicker_feldman_urbanska_santambrogio_et al._2016, title={Differential gene expression in human, murine, and cell line-derived macrophages upon polarization}, volume={347}, ISSN={0014-4827}, url={http://dx.doi.org/10.1016/J.YEXCR.2015.10.017}, DOI={10.1016/j.yexcr.2015.10.017}, abstractNote={The mechanisms by which macrophages control the inflammatory response, wound healing, biomaterial-interactions, and tissue regeneration appear to be related to their activation/differentiation states. Studies of macrophage behavior in vitro can be useful for elucidating their mechanisms of action, but it is not clear to what extent the source of macrophages affects their apparent behavior, potentially affecting interpretation of results. Although comparative studies of macrophage behavior with respect to cell source have been conducted, there has been no direct comparison of the three most commonly used cell sources: murine bone marrow, human monocytes from peripheral blood (PB), and the human leukemic monocytic cell line THP-1, across multiple macrophage phenotypes. In this study, we used multivariate discriminant analysis to compare the in vitro expression of genes commonly chosen to assess macrophage phenotype across all three sources of macrophages, as well as those derived from induced pluripotent stem cells (iPSCs), that were polarized towards four distinct phenotypes using the same differentiation protocols: M(LPS,IFN) (aka M1), M(IL4,IL13) (aka M2a), M(IL10) (aka M2c), and M(-) (aka M0) used as control. Several differences in gene expression trends were found among the sources of macrophages, especially between murine bone marrow-derived and human blood-derived M(LPS,IFN) and M(IL4,IL13) macrophages with respect to commonly used phenotype markers like CCR7 and genes associated with angiogenesis and tissue regeneration like FGF2 and MMP9. We found that the genes with the most similar patterns of expression among all sources were CXCL-10 and CXCL-11 for M(LPS,IFN) and CCL17 and CCL22 for M(IL4,IL13). Human PB-derived macrophages and human iPSC-derived macrophages showed similar gene expression patterns among the groups and genes studied here, suggesting that iPSC-derived monocytes have the potential to be used as a reliable cell source of human macrophages for in vitro studies. These findings could help select appropriate markers when testing macrophage behavior in vitro and highlight those markers that may confuse interpretation of results from experiments employing macrophages from different sources.}, number={1}, journal={Experimental Cell Research}, publisher={Elsevier BV}, author={Spiller, Kara L. and Wrona, Emily A. and Romero-Torres, Saly and Pallotta, Isabella and Graney, Pamela L. and Witherel, Claire E. and Panicker, Leelamma M. and Feldman, Ricardo A. and Urbanska, Aleksandra M. and Santambrogio, Laura and et al.}, year={2016}, month={Sep}, pages={1–13} }
 @inbook{pallotta_wrona_sun_freytes_2015, title={Role of mesenchymal stem cells, macrophages, and biomaterials during myocardial repair}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84955381540&partnerID=MN8TOARS}, DOI={10.1007/978-3-319-18045-8_1}, booktitle={Biomaterials in Regenerative Medicine and the Immune System}, author={Pallotta, I. and Wrona, E.A. and Sun, B. and Freytes, D.O.}, year={2015}, pages={1–15} }
 @book{natural cardiac extracellular matrix hydrogels for cultivation of human stem cell-derived cardiomyocytes_2014, volume={1181}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84925883447&partnerID=MN8TOARS}, DOI={10.1007/978-1-4939-1047-2_7}, abstractNote={Biomaterial scaffolds made of natural and synthetic materials are designed to serve as a structural and informational template for cell attachment and tissue formation. The use of native extracellular matrix (ECM) is of special interest for the culture of cardiac stem and progenitor cells due to the presence of intrinsic regulatory factors regulating cardiac function. We describe here how to obtain native ECM hydrogels from porcine hearts for the culture of human embryonic, induced pluripotent, and somatic stem cells for cardiac tissue engineering and regenerative medicine applications.}, journal={Methods in Molecular Biology}, year={2014}, pages={69–81} }