Karina A. Hernandez

Tissue engineering for plastic surgeons: a primer.

A central tenet of reconstructive surgery is the principle of “replacing like with like.” However, due to limitations in the availability of autologous tissue or because of the complications that may ensue from harvesting it, autologous reconstruction may be impractical to perform or too costly in terms of patient donor-site morbidity. The field of tissue engineering has long held promise to alleviate these shortcomings. Scaffolds are the structural building blocks of tissue-engineered constructs, akin to the extracellular matrix within native tissues. Commonly used scaffolds include allogenic or xenogenic decellularized tissue, synthetic or naturally derived hydrogels, and synthetic biodegradable nonhydrogel polymeric scaffolds. Embryonic, induced pluripotent, and mesenchymal stem cells also hold immense potential for regenerative purposes. Chemical signals including growth factors and cytokines may be harnessed to augment wound healing and tissue regeneration. Tissue engineering is already clinically prevalent in the fields of breast augmentation and reconstruction, skin substitutes, wound healing, auricular reconstruction, and bone, cartilage, and nerve grafting. Future directions for tissue engineering in plastic surgery include the development of prevascularized constructs and rationally designed scaffolds, the use of stem cells to regenerate organs and tissues, and gene therapy.

Reduction of suture associated inflammation after 28 days using novel biocompatible pseudoprotein poly(ester amide) biomaterials

Sutures elicit an inflammatory response, which may impede the healing process and result in wound complications. We recently reported a novel family of biocompatible, biodegradable polymers, amino acid-based poly(ester amide)s (AA-PEA), which we have shown to significantly attenuate the foreign body inflammatory response in vitro. Two types of AA-PEA (Phe-PEA and Arg-Phe-PEA) were used to coat silk or plain-gut sutures, which were implanted in the gluteus muscle of C57BL/6 mice, while the uncoated control sutures were implanted in the contralateral side. After 3, 7, 14, and 28 days the mean area of inflammation surrounding the sutures was compared. Phe-PEA coating of silk sutures significantly decreased inflammation compared with noncoated controls (67.8 ± 17.4% after 3d [p = 0.0014], 51.6 ± 7.2% after 7d [p < 0.001], and 37.3 ± 8.3% after 28d [p = 0.0001]) when assessed via analysis of photomicrographs using digital image software. Phe-PEA coated plain-gut sutures were similarly assessed and demonstrated a significant decrease in the mean area of inflammation across all time points (54.1 ± 8.3% after 3 d, 41.4 ± 3.9% after 7 d, 71.5 ± 8.1% after 14 d, 78.4 ± 8.5%, and after 28 d [all p < 0.0001]). Arg-Phe-PEA coated silk demonstrated significantly less inflammation compared to noncoated controls (61.3 ± 9.4% after 3 d, 44.7 ± 4.7% after 7 d, 19.6 ± 8%, and 38.3 ± 6.8% after 28 d [all p < 0.0001]), as did coated plain-gut (37.4 ± 8.3% after 3 d [p = 0.0004], 55.0 ± 7.8% after 7 d [p < 0.0001], 46.0 ± 4.6% after 14 d [p < 0.0001], and 59.0 ± 7.9% after 28 d [p < 0.0001]). Both Phe-PEA and Arg-Phe-PEA coatings significantly decrease the inflammatory response to sutures in vivo for up to 28 days.

Long-Term Morphological and Microarchitectural Stability of Tissue-Engineered, Patient-Specific Auricles In Vivo.

Current techniques for autologous auricular reconstruction produce substandard ear morphologies with high levels of donor-site morbidity, whereas alloplastic implants demonstrate poor biocompatibility. Tissue engineering, in combination with noninvasive digital photogrammetry and computer-assisted design/computer-aided manufacturing technology, offers an alternative method of auricular reconstruction. Using this method, patient-specific ears composed of collagen scaffolds and auricular chondrocytes have generated auricular cartilage with great fidelity following 3 months of subcutaneous implantation, however, this short time frame may not portend long-term tissue stability. We hypothesized that constructs developed using this technique would undergo continued auricular cartilage maturation without degradation during long-term (6 month) implantation. Full-sized, juvenile human ear constructs were injection molded from high-density collagen hydrogels encapsulating juvenile bovine auricular chondrocytes and implanted subcutaneously on the backs of nude rats for 6 months. Upon explantation, constructs retained overall patient morphology and displayed no evidence of tissue necrosis. Limited contraction occurred in vivo, however, no significant change in size was observed beyond 1 month. Constructs at 6 months showed distinct auricular cartilage microstructure, featuring a self-assembled perichondrial layer, a proteoglycan-rich bulk, and rounded cellular lacunae. Verhoeffs staining also revealed a developing elastin network comparable to native tissue. Biochemical measurements for DNA, glycosaminoglycan, and hydroxyproline content and mechanical properties of aggregate modulus and hydraulic permeability showed engineered tissue to be similar to native cartilage at 6 months. Patient-specific auricular constructs demonstrated long-term stability and increased cartilage tissue development during extended implantation, and offer a potential tissue-engineered solution for the future of auricular reconstructions.

Abstract 162: in vivo microanastomosis of microvessel containing tissue-engineered constructs: the final frontier.

PurPose: Although autologous tissue transfer has been established as a reliable approach to the reconstruction of complex defects, there are associated consequences including donor site pain, functional loss, paresthesias, dysthesthia, and scarring. The ability to synthesize vascularized constructs for the management of these complex wounds would represent a quantum leap in the field of tissue engineering. In previous work we synthesized and performed an in vivo microvascular anastomosis of a collagen construct containing an unseeded internal longitudinal microchannel with inlet and outlet. Here we fabricate and microsurgically anastomose collagen constructs containing an internal endothelialized microchannel.

Optimization of Vasculogenesis within Naturally-Derived, Biodegradable Hybrid Hydrogel Scaffolds

IntroductIon: Cellular ingrowth and neovascularization of acellular tissue-regeneration scaffolds represent the ratelimiting steps of permanent integration.1 In previous work, we have demonstrated that naturally-derived, biodegradable hybrid hydrogel scaffolds fabricated from a 10:1 w/w combination of alginate and type I collagen allow for maximal human umbilical vein endothelial cell (HUVEC) adherence and invasion in an in vitro wound healing model. We next sought to stimulate endothelial tubule formation within these scaffolds.

A Novel Three-Dimensional Platform to Investigate Neoangiogenesis, Transendothelial Migration, and Metastasis of MDAMB-231 Breast Cancer Cells.

Background: A crucial step in the progression of cancer involves the transendothelial migration of tumor cells into the bloodstream and invasion at distant sites. Most in vitro models of malignant cell behavior do not account for the presence of and interaction with vascular cells. Three-dimensional platforms to further explore the factors responsible for metastatic cellular behavior are under intensive investigation. Methods: Hydrogels with encapsulated MDAMB-231 breast cancer cells were fabricated with a central microchannel. The microchannel was lined with a co-culture of human umbilical vein endothelial cells and human aortic smooth muscle cells. For comparison, co-culture–seeded microchannels without breast cancer cells (MDAMB-negative) were fabricated. Results: After 7 and 14 days, the endoluminal lining of encapsulated MDAMB-231 co-culture–seeded microchannels demonstrated aberrant endothelial cell and smooth muscle cell organization and breast cancer cell transendothelial migration. MDAMB-231 cells performed matrix remodeling, forming tumor aggregates within the bulk, migrating preferentially toward the hydrogel “neovessel.” In contrast, MDAMB-negative constructs demonstrated maintenance of an intact endoluminal lining composed of endothelial cells and smooth muscle cells that organized into discrete layers. Furthermore, the thicknesses of the endoluminal lining of MDAMB-negative constructs were significantly greater than encapsulated MDAMB-231 co-culture–seeded constructs after 7 and 14 days (p = 0.012 and p < 0.001, respectively). Conclusion: The authors have created a powerful tool that may have tremendous impact on furthering our understanding of cancer recurrence and metastasis, shedding light on these poorly understood phenomena.

Abstract P29: Optimization of Functional, Perfusable Vascular Networks within Tissue Engineered Hydrogels

eleCtroPhysiology: SSEP negative peak amplitudes were significantly decreased (p<.05) at POD 14, 28, 37, and 79 when compared to baseline with significant recovery at POD 79. SSEP stimulation thresholds were significantly increased (p<.05) at POD 37 and 79 with significant recovery occurring at POD 79. Abductor pollicis brevis thresholds obtained from Tc-MEP stimulation were significantly increased at POD 37 and 79. CNAPs were first measured at POD 42 and confirmed with stimulation-response curve. Nerve conduction velocity was 40% baseline at POD 90 with significantly increased CNAP stimulation thresholds (p<.05).

Abstract 50: fabrication of tissue-engineered human constructs for patient specific auricles.

PurPose: The reconstruction of pediatric microtia using autologous donor cartilage is limited by significant obligatory donor site, pain and scarring as well as frequent suboptimal aesthetic outcome. Tissue-engineering allows for the creation of anatomically correct auricular constructs and the minimization or even elimination of the previously mentioned complications. In previous work, we fabricated patient specific, high fidelity tissue-engineered frameworks composed of type I collagen and bovine auricular chondrocytes that not only maintained shape and size over 12 weeks, but also exhibited proteoglycan and elastin deposition, with mechanical properties indistinguishable from native auricular cartilage. As a bridge to clinical translation, we have now synthesized human auricular chondrocyte (HAC) constructs in order to determine the optimal chrondrocyte passage and seeding density for the fabrication of patient specific tissue-engineered auricles.

Abstract 153: A Novel 3D Platform to Investigate Neoangiogenesis, Transendothelial Migration and Metastasis of Breast Cancer Cells.

PurPose: Breast cancer remains the most common cancer afflicting women and is the second leading cause of death from cancer. A crucial step in the progression of this disease is the transendothelial migration of tumor cells into the blood stream or lymphatic system. The factors guiding this process remain poorly understood. The development an in vitro biomimetic platform to further investigate these factors is under intensive investigation. In previous work we synthesized a tissue-engineered scaffold containing an endothelialized internal loop microchannel for microsurgical anastomosis and in vivo perfusion utilizing a sacrificial microfiber technique. Here we design a novel 3D platform to investigate tumor cell behavior in the presence of vascular cells in order to better understand the cell-cell and cell-matrix interactions that drive neoangiogenesis, invasion, metastasis and ultimately tumor progression.

Abstract 197: NOVEL BIODEGRADABLE POLY ESTER-AMIDE (PEA) POLYMER COATING SIGNIFICANTLY REDUCES SUTURE ASSOCIATED INFLAMMATORY RESPONSE

Introduction: Despite their ubiquitous use, surgical sutures are foreign bodies which induce a local immune reaction within the adjacent tissues. This in ammatory reaction predisposes the patient to infection and leads to complications including the development of suture granulomas, tunneling, spitting and abscess formation. We introduce a novel biocompatible and biodegradable (poly (ester-amide) 8-Phe-4 polymer (PEA)) material with anti-in ammatory properties that can be used to coat sutures. We hypothesized that two widely used sutures, plain-gut and silk, would elicit a reduced in ammatory response after being coated with PEA when compared to non-coated controls.