Journal of immunology and regenerative medicine最新文献

In islet transplantation, one of the major obstacles to optimal engraftment is the loss of islet natural vascularization and islet-specific extracellular matrix (ECM) during the islet isolation process. Thus, transplanted islets must re-establish nutritional and physical support through formation of new blood vessels and new ECM. To promote this critical process, we developed an elastin-based vasculogenic and ECM-promoting scaffold engineered for extrahepatic islet transplantation. The scaffold by design consisted of type I collagen (Coll) blended with 20wt% of elastin (E) shown to promote angiogenesis as well as de novo ECM deposition. The resulting “CollE” scaffolds had interconnected pores with a size distribution tailored to accommodate seeding of islets as well as growth of new blood vessels. In vitro, CollE scaffolds enabled prolonged culture of murine islets for up to one week while preserving their integrity, viability and function. In vivo, after only four weeks post-transplant of a marginal islet mass, CollE scaffolds demonstrated enhanced vascularization of the transplanted islets in the epididymal fat pad and promoted a prompt reversal of hyperglycemia in previously diabetic recipients. This outcome was comparable to that of kidney capsular (KC) islet transplantation, and superior to that of islets transplanted on the control collagen-only scaffolds (Coll). Crucial genes associated with angiogenesis (VEGFA, PDGFB, FGF1, and COL3A1) as well as de novo islet-specific matrix deposition (COL6A1, COL4A1, LAMA2 and FN1) were all significantly upregulated in islets on CollE scaffolds in comparison to those on Coll scaffolds. Finally, CollE scaffolds were also able to support human islet culture in vitro. In conclusion, CollE scaffolds have the potential to improve the clinical outcome of marginal islet transplantation at extrahepatic sites by promoting angiogenesis and islet-specific ECM deposition.

The host innate immune response to a surgical mesh is arguably the most important determinant of tissue remodeling and functional outcome. Macrophage phenotype and the associated secretion of pro-inflammatory or anti-inflammatory cytokines during the first 10–14 days following implantation has been strongly associated with downstream events such as chronic inflammation vs. functional tissue remodeling, respectively, and the associated clinical consequences. A persistent, pro-inflammatory (M1-like) macrophage phenotype is typically associated with fibrosis and scarring. In contrast, an early transition to a regulatory, pro-remodeling (M2-like) macrophage phenotype is predictive of organized, site-appropriate connective tissue deposition. The ratio of M2-like to M1-like macrophages in the early post-implantation period defines the microenvironmental milieu and the associated tissue response. The present study evaluated the early macrophage response to a synthetic non-resorbable (Bard^® Mesh), synthetic resorbable (TIGR^® Matrix Surgical Mesh and GORE^® BIO-A^® Tissue Reinforcement), synthetic mesh composed of the naturally occurring molecule 4-hydroxybutyrate, (Phasix™ Mesh), and a biologic surgical mesh composed of dermal extracellular matrix (Strattice™ Reconstructive Tissue Matrix); all of which are used in ventral hernia repair. The spatiotemporal distribution of pro-inflammatory (CD68⁺CD86⁺TNF-α⁺) and pro-remodeling (CD68⁺CD206⁺) macrophages, and the remodeling response in terms of vascularization, total number of infiltrating cells, presence of multinucleate giant cells (MNGC), and cell layer thickness around the implanted materials was evaluated at 3, 7, 14, 21, and 35 days post implantation. Results showed an association of the synthetic non-resorbable and resorbable meshes with a robust, pro-inflammatory response within 3 days of implantation, and an increased presence of MNGC around the mesh fibers at longer time points. Phasix™ Mesh was associated with an increased presence of M2-like macrophages immediately adjacent to the mesh fibers at earlier time points, and a favorable tissue remodeling outcome at 35 days. Results of the present study are consistent with the premise that an early shift of M1-like to M2-like macrophages is associated with favorable outcomes, including reduced fibrosis, at later time points.

The regenerative healing response of injured skeletal muscle is dependent upon an appropriately timed switch from a local type-I to a type-II immune response. Biologic scaffolds derived from extracellular matrix (ECM) have been shown to facilitate a macrophage phenotype transition that leads to downstream site-appropriate functional tissue deposition and myogenesis. However, the mechanisms by which ECM directs the switching of immune cell phenotype are only partially understood. Herein, we provide the first evidence that matrix bound nanovesicles (MBV) embedded within ECM-scaffolds are a rich and stable source of interleukin-33 (IL-33), an alarmin/cytokine with emerging reparative properties. We show that IL-33 encapsulated within MBV bypass the classical IL33/ST2 receptor signaling pathway to direct macrophage differentiation into the reparative, pro-remodeling M2 phenotype, which in turn facilitates myogenesis of skeletal muscle progenitor cells. Our results suggest the potential of IL-33⁺ MBV as a clinical therapy to augment the restorative efficacy of existing ECM-based and non-ECM based approaches.

Choosing the best ectopic in vivo model for cartilage engineering studies remains challenging and there is no clear consensus on how different models compare to one another. The use of xenogenic cells can often limit the choice to immunocompromised animals only and thus prevents the understanding of how tissue-engineered grafts perform with potential active inflammatory and immunological responses. The aim of this study was to evaluate the chondrogenic potential of a recently developed hydrogel, hyaluronan transglutaminase (HA-TG), in four mouse strains with varying immune systems: NSG, nude, NSG-SGM3 humanized and C57BL/6. The hyaluronan-based hydrogel was implanted subcutaneously for 4 weeks after an in vitro pre-culture time of 4 weeks. Scaffolds were prepared without cell seeding as the control and in combination with either human auricular chondrocytes (hAUR) or human fetal chondroprogenitor cells (hCC). We have seen that constructs were able to maintain their volumes and resisted vascularization as well as macrophage infiltration in vivo. Both hAUR and hCC maintained and produced ECM in vivo, but hAUR showed higher levels of innate collagen 2 even without mechanical stimulation. Collagen 1 and 2 deposition as well as mechanical properties of the scaffolds were comparable in all mouse strains. The C57BL/6 mouse model consistently displayed higher levels of C-reactive protein (CRP), serum amyloid A (SAA), and serum amyloid protein (SAP) in serum as a reaction to the foreign material and human cells. In addition, the number of CD68 + and CD163+ macrophages as well as CD3+ lymphocytes around the constructs in C57BL/6 mice was significantly higher than in humanized and immunocompromised mouse models. The results show that it is possible to engineer a cartilage-like graft subcutaneously not only in immunocompromised, but also in immunocompetent and humanized mouse models.

Severe musculoskeletal trauma is one of the most prevalent types of trauma in both combat-wounded and civilian patients. However, despite advances in trauma care management, mortality and complication rates remain high. Recently, systemic immune dysregulation and immunosuppression has been implicated as a main contributor to unsatisfactory success of intervention strategies and poor outcomes in a subset of trauma patients. The importance of the immune system in wound healing and tissue regeneration suggests that for optimal success of regenerative medicine strategies, this underlying systemic immune dysregulation must be addressed, highlighting a strong need for immunomodulatory approaches to augment regenerative therapies. This review addresses what is currently known about systemic immune dysregulation following severe trauma, previous attempts at immunomodulation, considerations for the development of future immunomodulatory strategies, and animal models for testing these therapeutics. The severity of immune dysregulation over time, both systemically and locally, is impacted by injury severity and patient-specific factors, such as age and co-morbidities. Varying degrees of systemic immune dysregulation may require different and personalized approaches to restore immune homeostasis. Further, the timing of treatment and the site of delivery must be considered to determine the most safe and effective intervention strategy. In order to test and develop these immunomodulatory therapeutics aimed at improving outcomes of regenerative interventions, robust pre-clinical models are needed that recapitulate systemic immune dysregulation and clinical scenarios. Overall, more fundamental research is needed to better understand the complex interactions between the musculoskeletal system and the immune system, as well as the importance of systemic and local immune function on healing. Ultimately, this information could lead to new immunomodulatory strategies to enhance regeneration for severe musculoskeletal trauma patients.

The liver harbors a rich, diverse spectrum of innate and adaptive immune cells. In homeostasis, these immune cells perform host defense against gut-derived pathogens and mediate tolerance to self-antigens. Following tissue injury there are complex interactions within the immune cell compartment which regulate liver regeneration and repair. Partial hepatectomy (PHx) and acetaminophen induced liver injury (AILI) are clinically relevant models of liver injury, which are commonly used to study liver regeneration. Here we discuss how the innate and adaptive immune systems influence liver regeneration and repair following acute hepatic injury.

Extracellular matrix (ECM) hydrogels represent a growing subset of injectable biomaterials for tissue engineering and regenerative medicine applications. Whilst two-dimensional scaffolds composed of ECM have been in routine clinical use for decades, the clinical translation of ECM hydrogels has been limited to date. Evaluation of the stability and function of ECM hydrogels after sterilization and storage is necessary to advance therapeutic use of these materials. In the present study, a new form of ECM material, specifically, ‘lyophilized digest’ was introduced to mitigate potential storage issues and investigate the effects of sterilization. Both ECM powders and lyophilized digests were subjected to sterilization by gamma irradiation, electron beam irradiation, ethylene oxide and supercritical carbon dioxide prior to hydrogel formation. The efficacy and effect of sterilization upon mechanical properties, macrophage response and stem cell chemotaxis was determined. Independent of the form of ECM exposed to sterilization, irradiation at 30 kGy altered ECM properties and inhibited subsequent gelation. Interestingly, the form of the ECM exposed to sterilization had a dramatic impact upon hydrogel rheological properties. Hydrogels formed from sterilized powders had reduced mechanical properties whereas hydrogels formed from sterilized lyophilized digest had mechanical properties equivalent to the non-sterilized control. Sterilization did not affect the bioactivity of ECM hydrogels as measured by stem cell chemotaxis and macrophage response. Together these data provide practical insight required for the translation of ECM based hydrogels.

Fibrosis is a process in which an accumulation of extracellular matrix (ECM) leads to an impaired function of the affected organ. Pulmonary fibrosis is the end-stage of several lung diseases, characterized by scarring of the lungs. Although macrophages are known to be important players in ECM homeostasis, their ability to respond to fibrosis-related morphological and mechanical changes of the ECM is relatively unexplored. In this study we aimed to elucidate the effect of ECM stiffness and morphology on macrophage polarization, by using a collagen type I-based in vitro system. Collagen morphology, but not stiffness, affected the relative expression of CD206 (the mannose receptor) and Ym1 (a murine marker of pro-healing M2 macrophages). Higher expression of Ym1 was found when macrophages were cultured on fibrous collagen. Globular collagen led to higher expression of CD206, a marker known to be upregulated on alveolar macrophages in idiopathic pulmonary fibrosis. Moreover, macrophages exhibited distinct differences in shape with actin-rich protrusions on fibrous collagen and more filopodia on globular collagen. In addition to these cytoskeletal changes, transmigration was higher when macrophages were cultured on fibrous collagen. Together these findings indicate that macrophages are sensitive to collagen morphology, responding with subtle changes in marker expression, shape and behavior rather than a complete polarization switch. This study emphasizes the complex interaction between macrophages and their surroundings, and the need for further exploration of both mechanical and morphological aspects.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a severe fibrotic component that compromises treatment, alters the immune cell profile and contributes to patient mortality. It has been shown that early on in this process, dynamic changes in tissue biomechanics play an integral role in supporting pancreatic cancer development and progression. Despite the acknowledgement of its importance, a granular view of how stromal composition changes during the course of PDAC progression remains largely unknown. To mimic the quasi-mesenchymal phenotype and pronounced desmoplastic response observed clinically, we utilized a genetically engineered mouse model of PDAC that is driven by a Kras^G12D mutation and loss of Tgfbr2 expression. Application of compartment resolved proteomics revealed that PDAC progression in this KTC model is associated with dynamic stromal alterations that are indicative of a wound healing program. We identified an early provisional matricellular fibrosis that was accompanied by markers of macrophage activation and infiltration, consistent with the inflammatory phase of wound healing. At 20 weeks a proliferative phenotype was observed with increased fibroblast markers, further collagen deposition and loss of basement membrane and native cell markers.