Alexander Huber

The Effects of Processing Methods upon Mechanical and Biologic Properties of Porcine Dermal Extracellular Matrix Scaffolds

Biologic materials from various species and tissues are commonly used as surgical meshes or scaffolds for tissue reconstruction. Extracellular matrix (ECM) represents the secreted product of the cells comprising each tissue and organ, and therefore provides a unique biologic material for selected regenerative medicine applications. Minimal disruption of ECM ultrastructure and content during tissue processing is typically desirable. The objective of this study was to systematically evaluate effects of commonly used tissue processing steps upon porcine dermal ECM scaffold composition, mechanical properties, and cytocompatibility. Processing steps evaluated included liming and hot water sanitation, trypsin/SDS/TritonX-100 decellularization, and trypsin/TritonX-100 decellularization. Liming decreased the growth factor and glycosaminoglycan content, the mechanical strength, and the ability of the ECM to support in vitro cell growth (p ≤ 0.05 for all). Hot water sanitation treatment decreased only the growth factor content of the ECM (p ≤ 0.05). Trypsin/SDS/TritonX-100 decellularization decreased the growth factor content and the ability of the ECM to support in vitro cell growth (p ≤ 0.05 for both). Trypsin/Triton X-100 decellularization also decreased the growth factor content of the ECM but increased the ability of the ECM to support in vitro cell growth (p ≤ 0.05 for both). We conclude that processing steps evaluated in the present study affect content, mechanical strength, and/or cytocompatibility of the resultant porcine dermal ECM, and therefore care must be taken in choosing appropriate processing steps to maintain the beneficial effects of ECM in biologic scaffolds.

Mechanical properties and in vivo behavior of a biodegradable synthetic polymer microfiber - extracellular matrix hydrogel biohybrid scaffold

A biohybrid composite consisting of extracellular matrix (ECM) gel from porcine dermal tissue and biodegradable elastomeric fibers was generated and evaluated for soft tissue applications. ECM gel possesses attractive biocompatibility and bioactivity with weak mechanical properties and rapid degradation, while electrospun biodegradable poly(ester urethane)urea (PEUU) has good mechanical properties but limited cellular infiltration and tissue integration. A concurrent gel electrospray/polymer electrospinning method was employed to create ECM gel/PEUU fiber composites with attractive mechanical properties, including high flexibility and strength. Electron microscopy revealed a structure of interconnected fibrous layers embedded in ECM gel. Tensile mechanical properties could be tuned by altering the PEUU/ECM weight ratio. Scaffold tensile strengths for PEUU/ECM ratios of 67/33, 72/28 and 80/20 ranged from 80 to 187 kPa in the longitudinal axis (parallel to the collecting mandrel axis) and 41-91 kPa in the circumferential axis with 645-938% breaking strains. The 72/28 biohybrid composite and a control scaffold generated from electrospun PEUU alone were implanted into Lewis rats, replacing a full-thickness abdominal wall defect. At 4 wk, no infection or herniation was found at the implant site. Histological staining showed extensive cellular infiltration into the biohybrid scaffold with the newly developed tissue well integrated with the native periphery, while minimal cellular ingress into the electrospun PEUU scaffold was observed. Mechanical testing of explanted constructs showed evidence of substantial remodeling, with composite scaffolds adopting properties more comparable to the native abdominal wall. The described elastic biohybrid material imparts features of ECM gel bioactivity with PEUU strength and handling to provide a promising composite biomaterial for soft tissue repair and replacement.

Polypropylene Surgical Mesh Coated with Extracellular Matrix Mitigates the Host Foreign Body Response

Surgical mesh devices composed of synthetic materials are commonly used for ventral hernia repair. These materials provide robust mechanical strength and are quickly incorporated into host tissue; factors that contribute to reduced hernia recurrence rates. However, such mesh devices cause a foreign body response with the associated complications of fibrosis and patient discomfort. In contrast, surgical mesh devices composed of naturally occurring extracellular matrix (ECM) are associated with constructive tissue remodeling, but lack the mechanical strength of synthetic materials. A method for applying a porcine dermal ECM hydrogel coating to a polypropylene mesh is described herein with the associated effects upon the host tissue response and biaxial mechanical behavior. Uncoated and ECM coated heavy-weight BARD™ Mesh were compared to the light-weight ULTRAPRO™ and BARD™ Soft Mesh devices in a rat partial thickness abdominal defect overlay model. The ECM coated mesh attenuated the pro-inflammatory response compared to all other devices, with a reduced cell accumulation and fewer foreign body giant cells. The ECM coating degraded by 35 days, and was replaced with loose connective tissue compared to the dense collagenous tissue associated with the uncoated polypropylene mesh device. Biaxial mechanical characterization showed that all of the mesh devices were of similar isotropic stiffness. Upon explanation, the light-weight mesh devices were more compliant than the coated or uncoated heavy-weight devices. This study shows that an ECM coating alters the default host response to a polypropylene mesh, but not the mechanical properties in an acute in vivo abdominal repair model.

Abdominal wall reconstruction by a regionally distinct biocomposite of extracellular matrix digest and a biodegradable elastomer

Current extracellular matrix (ECM) derived scaffolds offer promising regenerative responses in many settings, however in some applications there may be a desire for more robust and long lasting mechanical properties. A biohybrid composite material that offers both strength and bioactivity for optimal healing towards native tissue behavior may offer a solution to this problem. A regionally distinct biocomposite scaffold composed of a biodegradable elastomer (poly(ester urethane)urea) and porcine dermal ECM gel was generated to meet this need by a concurrent polymer electrospinning/ECM gel electrospraying technique where the electrosprayed component was varied temporally during the processing. A sandwich structure was achieved with polymer fiber rich upper and lower layers for structural support and an ECM‐rich inner layer to encourage cell ingrowth. Increasing the upper and lower layer fiber content predictably increased tensile strength. In a rat full thickness abdominal wall defect model, the sandwich scaffold design maintained its thickness whereas control biohybrid scaffolds lacking the upper and lower fiber‐rich regions failed at 8 weeks. Sandwich scaffold implants also showed higher collagen content 4 and 8 weeks after implantation, exhibited an increased M2 macrophage phenotype response at later times and developed biaxial mechanical properties better approximating native tissue. By employing a processing approach that creates a sheet‐form scaffold with regionally distinct zones, it was possible to improve biological outcomes in body wall repair and provide the means for further tuning scaffold mechanical parameters when targeting other applications. Copyright

Phenotypic changes in cultured smooth muscle cells: limitation or opportunity for tissue engineering of hollow organs?

Smooth muscle cells (SMCs) are typically used as a cell source for the reconstruction of hollow organs by conventional tissue engineering techniques. However, the necessity for and advantage of the use of tissue‐specific SMCs are unknown. The present study investigated the phenotypic changes that occur following isolation and in vitro expansion of rat SMC populations isolated from three different tissues: the aorta, oesophagus and urinary bladder. rSMCs were isolated by enzymatic dispersion and expanded by conventional cell culture techniques, yielding microscopically homogeneous populations. SMC phenotypes were monitored according to their expression of marker proteins during the first two passages. Two of the three SMC populations (rSMC‐a and rSMC‐e) showed a marked change in their marker protein profiles during the first two passages, which resulted in a homogeneous phenotype that was neither fully contractile nor fully synthetic. SMCs from the urinary bladder did not show such a shift. Differences between the three rSMC populations were observed with regard to proliferative activity and gene expression patterns, suggesting the retention of some tissue‐specific cell characteristics. In summary, phenotypic changes in SMCs occur as a result of conventional cell isolation and expansion techniques, thus questioning the necessity for a tissue‐specific cell source for regenerative medicine applications. Copyright

Polypropylene‐containing synthetic mesh devices in soft tissue repair: A meta‐analysis

The acute and chronic host tissue response to synthetic and biologic mesh devices for abdominal hernia repair is thought to ultimately determine clinical outcomes such as adhesion formation, device shrinkage, cellular response, and neotissue formation. A meta-analysis of 38 publications was performed to assess these outcomes in six different treatment groups depending on mesh composition: polypropylene (PP), PP in combination with nonabsorbable polymers, PP in combination with absorbable polymers, non-PP polymers, non-PP in combination with absorbable polymers, and natural materials. Despite showing the least device shrinkage, meshes made entirely from PP generally showed the most adverse host tissue response. PP devices with an absorbable component elicited a more beneficial host response with respect to connective tissue adhesion and tissue inflammation than devices made from PP alone. These devices also provided a high level of mechanical stability resulting in a reduced level of adhesion formation and device shrinkage postapplication. However, the compositional heterogeneity within certain groups, that is, devices of non-PP polymers, non-PP in combination with absorbable polymers, and natural materials, did not allow for a more detailed evaluation or the identification of a single composition with superior host tissue response characteristics.

Histopathologic host response to polypropylene-based surgical mesh materials in a rat abdominal wall defect model.

Composite polypropylene-based surgical mesh materials including various synthetic polymers and naturally occurring biomaterials have been developed to ameliorate device-associated inflammatory response and associated reduced compliance of pure polypropylene meshes. This study evaluated the histomorphologic response of three composite polypropylene-based surgical meshes, Revive™, a polycarbonate polyurethane reinforced monofilamentous polypropylene scaffold, Assure™, a polycarbonate polyurethane reinforced monofilamentous polypropylene scaffold with a resorbable anti-adhesion layer of lactide caprolactone copolymer, and Proceed™, a polypropylene mesh modified with oxidized cellulose, in a soft tissue repair model in the rat. The host inflammatory response and neotissue formation were evaluated by semiquantitative histologic scoring including the amount of cellular infiltration, angiogenesis, presence of multinucleate giant cells, fibrous connective tissue formation, and host neo-extracellular matrix deposition for up to 26 weeks. All three composite surgical mesh materials showed good integration with host tissue as indicated by rapid cellular infiltration, abundant neo-vascularization, minimal shrinkage, and the lack of visible mesh degradation. The devices elicited a similar inflammatory response and the presence of a mild foreign body response in spite of the different composition and morphology of these composite mesh materials.

Biological Scaffolds for Regenerative Medicine

This chapter evaluates biological scaffold materials in comparison to conventional synthetic scaffold materials, with a focus on intact acellular extracellular matrix (ECM) scaffold materials. Collagens, glycosaminoglycans, chitosans, and other components of the extracellular matrix are used as implantable scaffold materials. Collagen is the most common and abundant naturally occurring scaffold material that can be extracted from various tissues such as tendons, ligaments, and other connective tissues, solubilized, and reconstituted into fibers of various geometries that could, in turn, be transformed into a variety of shapes and sizes to mimic body structures such as heart valves, blood vessels, and skin. Allogeneic and xenogeneic collagen is generally recognized as “self” tissue when used as a biological scaffold material regardless of its species of origin, and it is subjected to the fundamental biological processes of degradation and integration into adjacent host tissues when left in its native ultrastructure. Intact ECM could be isolated from a large variety of different tissues, including heart valves, blood vessels, skin, nerves, skeletal muscle, tendons, ligaments, small intestine, urinary bladder, and liver. These biological scaffolds could be harvested from several different species including tissues from human, porcine, bovine, and equine or from cells grown in vitro . ECM degradation leads to an initial decrease in overall strength during the early phase of in vivo remodeling, followed by an increase in strength due to the deposition of site-specific ECM and the formation of functional site-appropriate neotissue by infiltrating cells in response to their experienced mechanical stresses.