Nicholas Bryan

Hydrogels for tissue engineering and regenerative medicine

Injectable hydrogels have become an incredibly prolific area of research in the field of tissue engineering and regenerative medicine, because of their high water content, mechanical similarity to natural tissues, and ease of surgical implantation, hydrogels are at the forefront of biomedical scaffold and drug carrier design. The aim of this review is to concisely summarise current state-of-the-art in natural and synthetic hydrogels with respect to their synthesis and fabrication, comparing and contrasting the many chemistries available for biomedical hydrogel generation using both biologic and synthetic base materials. We then discuss these hydrogels in the specific instance of several pertinent areas of TERM which have been specifically selected to demonstrate how this versatile class of materials can be modified to augment damage and disease of a seemingly limitless array of adult tissues.

Derivation and performance of an entirely autologous injectable hydrogel delivery system for cell-based therapies

A host-derived hydrogel has been designed and validated as an entirely autologous, injectable delivery system for cells with potential for cell-based therapies and tissue engineering applications. Each individual has components in their blood from which can be formed a mechanically stable hydrogel having the capacity to maintain cellular phenotype and support cellular proliferation of multiple cell types through several culture passages ex vivo. The hydrogel can be triggered to gel at the time of implantation into the patient through an injection system that facilitates a liquid injection of components of the donor plasma and cells into the site of interest. This results in stable ectopic tissue formation at the site of implantation. Our studies have demonstrated excellent integration of the neotissue with host tissues with maintenance of the phenotype of implanted cells whilst observing minimal host innate immune cell recruitment. These findings could provide the fundamental basis for new hydrogel-based biomaterial therapies, overcoming the histocompatibility factors associated with implantable biomaterials whilst providing a stable three dimensional medium for cellular growth both in vivo and ex vivo.

A scientific evidence for the efficacy of biologic implants for soft tissue reconstruction

The challenges and complications arising from abdominal surgery frequently necessitate soft tissue reconstruction or augmentation. Soft tissue repair generally has been revolutionised by the introduction of synthetic meshes, but their use is contra‐indicated in contaminated or infected fields. Biologic materials derived from devitalised allo‐ or xenogeneic tissues have been proposed as a safer alternative to synthetics and provide an extracellular scaffold necessary for the in‐growth of new blood vessels and infiltration of native stromal cells. We review the scientific evidence behind commercially available biologic prostheses in relation to the impact of tissue source, manufacturing processes and supplemental cross‐linking on in vitro and in vivo (animal model and clinical) performance.

The innate oxygen dependant immune pathway as a sensitive parameter to predict the performance of biological graft materials

Clinical performance of a biomaterial is decided early after implantation as leukocytes interrogate the graft throughout acute inflammation. High degrees of leukocyte activation lead to poor material/patient compliance, accelerated degeneration and graft rejection. A number reactive oxygen species (ROS) are released by leukocytes throughout their interaction with a material, which can be used as a sensitive measure of leukocyte activation. The aim of this study was to compare leukocyte activation by commercially available biologic surgical materials and define the extent manufacturing variables influence down-stream ROS response. Chemiluminescence assays were performed using modifications to a commercially available kit (Knight Scientific, UK). Whole blood was obtained from 4 healthy human adults at 7 day intervals for 4 weeks, combined with Adjuvant K, Pholasin (a highly sensitive ROS excitable photoprotein) and biomaterial, and incubated for 60 min with continuous chemiluminescent measurements. Leukocyte ROS inducers fMLP and PMA were added as controls. Xeno- and allogeneic dermal and small intestinal submucosal (SIS) derived biomaterials were produced commercially (Surgisis Biodesign™, Alloderm(®), Strattice(®)Firm & Pliable & Permacol™) or fabricated in house to induce variations in decellularisation and cross-linking. Statistics were performed using Waller-Duncan post hoc ranking. Materials demonstrated significant differences in leukocyte activation as a function of decellularisation reagent and tissue origin. The data demonstrated SIS was significantly more pro-inflammatory than dermis. Additionally it was deduced that SDS during decellularisation induced pro-inflammatory changes to dermal materials. Furthermore, it was possible to conclude inter-patient variation in leukocyte response. The in vitro findings were validated in vivo which confirmed the chemiluminescence observations, highlighting the potential for translation of this technique as a routine component of pre-surgical evaluation to maximise foreign body compliance.

The in vivo evaluation of tissue‐based biomaterials in a rat full‐thickness abdominal wall defect model

Hernias are defects in which an anatomical fascia is breached resulting in ectopic positioning of an organ into an orifice which routinely does not contain it. Intervention often involves repositioning translocated organs and repair of damaged fascia using exogenous grafts. Despite hernia prevalence, repairs can still fail due to postoperative complications, such as chronic pain and decreased mobility. This study compared repair capacities and characterized the foreign body response elicited by a number of hernia repair grafts to deduce their bulk inflammatory properties while also concluding the point in their fabrication when these are inferred. Materials derived from human dermis (Alloderm(®) ), porcine dermis (Permacol™, patch A, patch D and Strattice(®) ), porcine small-intestinal submucosa (Surgisis™) and a synthetic (multifilament Surgipro™) were implanted into a rat full-thickness abdominal wall excision model, incubated for up to 2 years and characterized histopathologically. Surgisis™ resorbed the fastest of the materials tested (1-3 months) resulting in a mechanically stable parietal peritoneum. Decellularization using sodium dodecyl sulfate (patch A) stimulated a large early inflammatory response which ultimately may have contributed to increased resorption of porcine dermal matrix however the remaining materials typically persisted throughout the 2-year incubation. Cross-linking porcine dermis using 1,6-hexamethylene disocyanate (vs. an identical noncross-linked counterpart) showed no difference in cell recruitment or material integration over 2 years. Typically Strattice(®) and Alloderm(®) recruited larger early populations of cells than Permacol™; however, over extended periods of time in vivo this response normalized.

Elucidating the contribution of the elemental composition of fetal calf serum to antigenic expression of primary human umbilical-vein endothelial cells in vitro

One of the major obstacles to obtaining human cells of a defined and reproducible standard suitable for use as medical therapies is the necessity for FCS (fetal calf serum) media augmentation in routine cell culture applications. FCS has become the supplement of choice for cell culture research, as it contains an array of proteins, growth factors and essential ions necessary for cellular viability and proliferation in vitro. It is, however, a potential route for the introduction of zoonotic pathogens and makes defining the cell culture milieu impossible in terms of reproducibility, as the precise composition of each batch of serum not only changes but is in fact extremely variable. The present study determined the magnitude of donor variations in terms of elemental composition of FCS and the effect these variations had on the expression of a group of proteins associated with the antigenicity of primary human umbilical-vein endothelial cells, using a combination of ICPMS (inductively coupled plasma MS) and flow cytometry. Statistically significant differences were demonstrated for a set of trace elements in FCS, with correlations made to variations in antigenic expression during culture. The findings question in detail the suitability of FCS for the in vitro supplementation of cultures of primary human cells due to the lack of reproducibility and modulations in protein expression when cultured in conjunction with sera from xenogeneic donors.

In vitro activation of human leukocytes in response to contact with synthetic hernia meshes.

OBJECTIVES Evaluation of an in vitro chemiluminescent screen to predict leukocyte ROS in response to surgical materials. DESIGN AND METHODS 6 surgical meshes; manufacture and knitting variations of polypropylene (PP), polyester terephtalate (PET) and polyglycolic acid (PGA) trialled healthy human blood (n=5). Materials and blood were incubated with pholasin. Pholasin emits photons in the presence of reactive oxygen species; secreted by activated leukocytes. RESULTS Multifilament-PGA mesh stimulated the greatest ROS response from blood derived human leukocytes. Multifilament-PET light weight and multifilament-PP meshes stimulated similar levels of ROS production which were greater than monofilament-PP light, monofilament-PP and monofilament-PET light meshes. Data demonstrated statistical variations in trans-donor response to the materials. CONCLUSIONS An in vitro chemiluminescent assay can be used to assess leukocyte respiratory burst response to biomaterials. PGA mesh elicited the greatest ROS response. PP and PET monofilament meshes induce less ROS than multifilament equivalents. In vitro results correlate with previously published clinical responses to these materials.

Porcine dermis implants in soft-tissue reconstruction: current status

Soft-tissue reconstruction for a variety of surgical conditions, such as abdominal wall hernia or pelvic organ prolapse, remains a challenge. There are numerous meshes available that may be simply categorized as either synthetic or biologic. Within biologic meshes, porcine dermal meshes have come to dominate the market. This review examines the current evidence for their use and the limitations of knowledge. Although there is increasing evidence to support their safety, long-term follow-up studies that support their efficacy are lacking. Numerous clinical trials that remain ongoing may help elucidate their precise role in soft-tissue reconstruction.

Characterisation and comparison of the host response of 6 tissue-based surgical implants in a subcutaneous in vivo rat model.

Background Hernia repair often involves fascial augmentation using biologic prostheses. Small processing changes during preparation modulate host tissue response, which influence material efficacy and longevity. In this pilot study, a rat model was used to determine the specific influence of tissue origin, decellularisation treatment and 1,6-hexamethylene diisocyanate (HMDI) cross-linking. Methods Materials (1 cm2) were implanted subcutaneously into 6-week-old Wistar rats (4 materials per animal, n=6/material per time point) for 2, 5, 7, 14 and 28 days. Histologic processing was carried out after resin infiltration, observing classical histopathology and pathologic indexing. Materials comprised 6 tissue-based grafts covering both experimental and commercial porcine decellularised dermal and small intestinal submucosal materials. Results Subcutaneous delivery of biologics demonstrated material-specific inflammatory/host responses. Controlled variations of the Permacol™ manufacturing process showed sodium dodecyl sulfate (SDS) was the most proinflammatory decellularisation reagent, and HMDI cross-linking had no effect on host response. All materials remained recoverable after 28 days, although Surgisis™ had partially resorbed. Conclusion Differences in host responses exist between biologic implants for hernia repair in this rat model. It is postulated that these modifications are induced during processing and may have an effect on the clinical outcome of hernia repair.

A review of biocompatibility in hernia repair; considerations in vitro and in vivo for selecting the most appropriate repair material

PurposeRepair of hernia typically makes use of a prosthetic material; synthetic or biologic in nature. Any material which enters the body is subject to interrogation by the inflammation and immune system in addition to numerous other cell families, the outcome of which ultimately determines the success of the repair. In this review, we discuss the fundamental biology which occurs in situ when a biomaterial associates with a tissue, compare and contrast the techniques available to predict this in vitro, and review how features of hernia repair materials specifically may manipulate tissue interrogation and integration. Finally, we conclude our article by examining how biocompatibility impacts surgical practise and how a better understanding of the manner by which materials and tissues interact could benefit hernia repair.Materials and methodsA review of the literature was conducted using appropriate scientific search engines in addition to inclusion of findings from the groups’ primary research.ResultsUsing pre-clinical assays to anticipate the biocompatibility of a medical device is critical; however, to maximise the scientific power of in vitro findings, we must carefully consider the in vivo niche of the cells with which we are working. Excessive in vitro culture or contact to non-self materials can add compounding complexity to studies involving leucocytes for instance; therefore, we must ensure careful and stringent assay design when developing techniques for assaying pre-clinical biocompatibility. Furthermore, many of the features associated with hernia repair material design specifically, included to enhance their mechanical or biodegradation characteristics, are inadvertently instructive to cells, and therefore, throughout the prototype stages of a materials development, regular biocompatibility assessment must be performed.ConclusionThe biocompatibility of a material is rate limiting in its ability to function as a medical device. The future of hernia repair materials will rely on close cohesion between the surgical and scientific communities to ensure the most robust biocompatibility assessment techniques, and models are utilised to predict the efficacy of a given material in a particular surgical application.