Constructive Remodeling Research Articles

Matrix Metalloproteinase-Responsive Controlled Release of Self-Assembly Nanoparticles Accelerates Heart Valve Regeneration In Situ by Orchestrating Immunomodulation.

In situ tissue engineering heart valves (TEHVs) are the most promising way to overcome the defects of existing valve prostheses. Despite their promising prospects, the clinical translation of TEHVs remains a formidable challenge, mainly due to unpredictable host interactions post-implantation. An immunomodulatory idea based on hydrogel encapsulation of nanoparticle-coated heart valve scaffolds is introduced. Specifically, galactose-modified human serum albumin nanoparticles (miR-93@HSA NPs) to deliver microRNA-93 mimics are utilized, which target macrophages and induce their differentiation into the anti-inflammatory M2 subtype, fostering a conducive immune microenvironment. Matrix metalloproteinase (MMP)-responsive hydrogel is used to encapsulate the nanoparticles, enabling targeted and sustained release. Results show that the miR-93@HSA NPs exhibit excellent ability to induce macrophage polarization toward the M2 phenotype. A decellularized valve modified with hydrogel reveals MMP-response release of the miR-93@HSA NPs. In vitro, the immunomodulatory heart valve possesses good endocytocompatibility and effectively reprograms macrophages when cocultured with HUVECs or RAW264.7 macrophages. In vivo, this valve scaffold promises to mitigate early inflammatory damage and provide a pro-endothelialization niche for scaffolds' constructive remodeling. With the use of cell coculture systems and transcriptome sequencing, the mechanism of immune-modulating scaffold accelerating endothelialization is being elucidated. The immunomodulatory heart valve scaffold holds promising potential for clinical translation.

Evaluation of Bi-layer Silk Fibroin Grafts for Inlay Vaginoplasty in a Rat Model.

Autologous tissues derived from bowel, buccal mucosa and skin are primarily used to repair or replace diseased vaginal segments as well as create neovaginas for male-to-female transgenders. These grafts are often limited by scarce tissue supply, donor site morbidity and post-operative complications. Bi-layer silk fibroin (BLSF) biomaterials represent potential alternatives for vaginoplasty given their structural strength and elasticity, low immunogenicity, and processing flexibility. The goals of the current study were to assess the potential of acellular BLSF scaffolds for vaginal tissue regeneration in respect to conventional small intestinal submucosal (SIS) matrices in a rat model of vaginoplasty. Inlay vaginoplasty was performed with BLSF and SIS scaffolds (N = 21 per graft) in adult female rats for up to 2months of implantation. Nonsurgical controls (N = 4) were investigated in parallel. Outcome analyses included histologic, immunohistochemical and histomorphometric evaluations of wound healing patterns; µ-computed tomography (CT) of vaginal continuity; and breeding assessments. Animals in both scaffold cohorts exhibited 100% survival rates with no severe post-operative complications. At 2months post-op, µ-CT analysis revealed normal vaginal anatomy and continuity in both graft groups similar to controls. In parallel, BLSF and SIS grafts also induced comparable constructive remodeling patterns and were histologically equivalent in their ability to support formation of vascularized vaginal neotissues with native tissue architecture, however with significantly less smooth muscle content. Vaginal tissues reconstructed with both implants were capable of supporting copulation, pregnancy and similar amounts of live births. BLSF biomaterials represent potential "off-the-shelf" candidates for vaginoplasty.

Efficacy of the Sub-Urethral Transobturator KIM System® for Female Urinary Incontinence: Long Term Results.

Background/Objectives: Female stress urinary incontinence (SUI) surgical treatment has changed since its beginning. Selecting materials that promote constructive tissue remodelling helps to maintain continence after surgery and minimizes complications. To analyze the long-term urinary continence results in women who underwent SUI surgical correction using the transobturator mid-urethral sling KIM system® (Knotless Incontinence Mesh) and analyze the complications associated with this SUI treatment. Materials and Methods: Prospective study of the first 1000 patients undergoing SUI surgery with the Contasure KIM® sling between April 2007 and December 2018. Results and complications were recorded. Group A represented 94.2% of the sample and were the continent patients after surgery (GA = 942), while Group B accounted for incontinent patients after surgery (5.8%) (GB = 58). Results: The mean age at operation was 60 years. Eutocic deliveries (p = 0.0022), high blood pressure (p = 0.0190), anxiety (p = 0.0084), hemorrhoidectomies (p = 0.0016) and hysterectomies (p = 0.0002) were higher in GB. No differences between groups were found regarding body mass index (GA 26.50; GB 26.93) (p = 0.220649), food or drug allergies (p = 0.0.6547), dystocia (p = 0.2365), diabetes mellitus (p = 0.1715), pelvic surgery (p = 0.8842), other surgery (p = 0.8801) or concomitant treatments that would have an impact on bladder function. Correction of SUI was achieved in 94.2% of cases. Continence persisted over time in 97.98% of continent patients at 4-year follow-up and 90.90% of patients at last follow-up. Mesh caused erosion of the urethra in 0.3% of patients and extruded to the vagina in 0.42%. De novo urinary urgency (2.8%), significant chronic pain (3.6%) and urinary tract infections (3.0%) after surgery were lower than complications reported in reviewed publications. Pain was treated with medication, and all patients were pain-free at the one-year follow-up visit. Conclusions: The mid-urethral transobturator sling KIM system® has good initial and long-term results in patients with stress urinary incontinence, with a low recurrence rate and minimal complications.

Contribution of Tregs to the promotion of constructive remodeling after decellularized extracellular matrix material implantation

Host remodeling of decellularized extracellular matrix (dECM) material through the appropriate involvement of immune cells is essential for achieving functional organ/tissue regeneration. As many studies have focused on the role of macrophages, only few have evaluated the role of regulatory T cells (Tregs) in dECM remodeling. In this study, we used a mouse model of traumatic muscle injury to determine the role of Tregs in the constructive remodeling of vascular-derived dECM. According to the results, a certain number of Tregs could be recruited after dECM implantation. Notably, using anti-CD25 to reduce the number of Tregs recruited by the dECM was significantly detrimental to material remodeling based on a significant reduction in the number of M2 macrophages. In addition, collagen and elastic fibers, which maintain the integrity and mechanical properties of the material, rapidly degraded during the early stages of implantation. In contrast, the use of CD28-SA antibodies to increase the number of Tregs recruited by dECM promoted constructive remodeling, resulting in a decreased inflammatory response at the material edge, thinning of the surrounding fibrous connective tissue, uniform infiltration of host cells, and significantly improved tissue remodeling scores. The number of M2 macrophages increased whereas that of M1 macrophages decreased. Moreover, Treg-conditioned medium further enhanced material-induced M2 macrophage polarization in vitro. Overall, Treg is an important cell type that influences constructive remodeling of the dECM. Such findings contribute to the design of next-generation biomaterials to optimize the remodeling and regeneration of dECM materials.

Biodegradable Multi-layered Silk Fibroin-PCL Stent for the Management of Cervical Atresia: In Vitro Cytocompatibility and Extracellular Matrix Remodeling In Vivo.

Cervical atresia is a rare congenital Müllerian duct anomaly that manifests as the absence or deformed nonfunctional presence of the cervix. Herein, a multi-layered biodegradable stent is fabricated using a homogeneous blend of silk fibroin with polycaprolactone using hexafluoroisopropanol as a common solution. Briefly, a concentric cylinder of 3D honeycomb layer is sandwiched within electrospun sheets for fixing at the cervico-uterine junction to pave the way of cervical reconstruction. An average length of 40 mm with 3 mm diameter is fabricated for the hybrid stent design. SEM evidences an evenly distributed pore architecture of the electrospun layer, and mechanical characterization of stent reveals a tensile strength of 1.7 ± 0.2 MPa, with a Young's modulus of 5.9 ± 0.1 MPa. Physico-chemical characterization confirms the presence of silk fibroin and poly caprolactone within the engineered stent. Following 14 days of pepsin enzymatic degradation, 18% degradation and a contact angle measurement of 97° are observed. In vitro cytocompatibility studies are performed using site-specific primary human cervical squamous, columnar epithelial cells, and human endometrial stromal cells. The study demonstrates non-cytotoxic cells' viability (no significant toxicity), improved cell anchoring, adherence among the stent layers, and proliferation in the 3D microenvironment. Furthermore, in vivo subcutaneous studies in the rodent model indicate that the implanted stent undergoes constructive remodeling, neo-tissue creation, neo-vasculature formation, and re-epithelialization while maintaining patency for 2 months.

The Effect of Titanium Surface Topography on Adherent Macrophage Integrin and Cytokine Expression

Immunomodulatory biomaterials have the potential to stimulate an immune response able to promote constructive and functional tissue remodeling, as opposed to persistent inflammation and scar tissue formation. This study examined the effects of titanium surface modification on integrin expression and concurrent cytokine secretion by adherent macrophages in vitro in an attempt to delineate the molecular events involved in biomaterial-mediated immunomodulation. Non-polarised (M0) and inflammatory polarised (M1) macrophages were cultured on a relatively smooth (machined) titanium surface and two proprietary modified rough titanium surfaces (blasted and fluoride-modified) for 24 h. The physiochemical characteristics of the titanium surfaces were assessed by microscopy and profilometry, while macrophage integrin expression and cytokine secretion were determined using PCR and ELISA, respectively. After 24 h adhesion onto titanium, integrin α1 expression was downregulated in both M0 and M1 cells on all titanium surfaces. Expression of integrins α2, αM, β1 and β2 increased in M0 cells cultured on the machined surface only, whereas in M1 cells, expression of integrins α2, αM and β1 all increased with culture on both the machined and rough titanium surfaces. These results correlated with a cytokine secretory response whereby levels of IL-1β, IL-31 and TNF-α increased significantly in M1 cells cultured on the titanium surfaces. These results show that adherent inflammatory macrophages interact with titanium in a surface-dependent manner such that increased levels of inflammatory cytokines IL-1β, TNF-α and IL-31 secreted by M1 cells were associated with higher expression of integrins α2, αM and β1.

Harnessing the synergy of perfusable muscle flap matrix and adipose-derived stem cells for prevascularization and macrophage polarization to reconstruct volumetric muscle loss.

Muscle flaps must have a strong vascular network to support a large tissue volume and ensure successful engraftment. We developed porcine stomach musculofascial flap matrix (PDSF) comprising extracellular matrix (ECM) and intact vasculature. PDSF had a dominant vascular pedicle, microcirculatory vessels, a nerve network, well-retained 3-dimensional (3D) nanofibrous ECM structures, and no allo- or xenoantigenicity. In-depth proteomic analysis demonstrated that PDSF was composed of core matrisome proteins (e.g., collagens, glycoproteins, proteoglycans, and ECM regulators) that, as shown by Gene Ontology term enrichment analysis, are functionally related to musculofascial biological processes. Moreover, PDSF-human adipose-derived stem cell (hASC) synergy not only induced monocytes towards IL-10-producing M2 macrophage polarization through the enhancement of hASCs' paracrine effect but also promoted the proliferation and interconnection of both human skeletal muscle myoblasts (HSMMs) and human umbilical vein endothelial cells (HUVECs) in static triculture conditions. Furthermore, PDSF was successfully prevascularized through a dynamic perfusion coculture of hASCs and HUVECs, which integrated with PDSF and induced the maturation of vascular networks in vitro. In a xenotransplantation model, PDSF demonstrated myoconductive and immunomodulatory properties associated with the predominance of M2 macrophages and regulatory T cells. In a volumetric muscle loss (VML) model, prevascularized PDSF augmented neovascularization and constructive remodeling, which was characterized by the predominant infiltration of M2 macrophages and significant musculofascial tissue formation. These results indicate that hASCs' integration with PDSF enhances the cells' dual function in immunomodulation and angiogenesis. Owing in part to this PDSF-hASC synergy, our platform shows promise for vascularized muscle flap engineering for VML reconstruction.

Biocompatibility and biodistribution of matrix-bound nanovesicles in vitro and in vivo.

Matrix-bound nanovesicles (MBV) are a distinct subtype of extracellular vesicles that are firmly embedded within biomaterials composed of extracellular matrix (ECM). MBV both store and transport a diverse, tissue specific portfolio of signaling molecules including proteins, miRNAs, and bioactive lipids. MBV function as a key mediator in ECM-mediated control of the local tissue microenvironment. One of the most important mechanisms by which MBV in ECM bioscaffolds support constructive tissue remodeling following injury is immunomodulation and, specifically, the promotion of an anti-inflammatory, pro-remodeling immune cell activation state. Recent in vivo studies have shown that isolated MBV have therapeutic efficacy in rodent models of both retinal damage and rheumatoid arthritis through the targeted immunomodulation of pro-inflammatory macrophages towards an anti-inflammatory activation state. While these results show the therapeutic potential of MBV administered independent of the rest of the ECM, the in vitro and in vivo safety and biodistribution profile of MBV remain uncharacterized. The purpose of the present study was to thoroughly characterize the pre-clinical safety profile of MBV through a combination of in vitro cytotoxicity and MBV uptake studies and in vivo toxicity, immunotoxicity, and imaging studies. The results showed that MBV isolated from porcine urinary bladder are well-tolerated and are not cytotoxic in cell culture, are non-toxic to the whole organism, and are not immunosuppressive compared to the potent immunosuppressive drug cyclophosphamide. Furthermore, this safety profile was sustained across a wide range of MBV doses. STATEMENT OF SIGNIFICANCE: Matrix-bound nanovesicles (MBV) are a distinct subtype of bioactive extracellular vesicles that are embedded within biomaterials composed of extracellular matrix (ECM). Recent studies have shown therapeutic efficacy of MBV in models of both retinal damage and rheumatoid arthritis through the targeted immunomodulation of pro-inflammatory macrophages towards an anti-inflammatory activation state. While these results show the therapeutic potential of MBV, the in vitro and in vivo biocompatibility and biodistribution profile of MBV remain uncharacterized. The results of the present study showed that MBV are a well-tolerated ECM-derived therapy that are not cytotoxic in cell culture, are non-toxic to the whole organism, and are not immunosuppressive. Collectively, these data highlight the translational feasibility of MBV therapeutics across a wide variety of clinical applications.

Silk fibroin bioscaffold from Bombyx mori and Antheraea assamensis elicits a distinct host response and macrophage activation paradigm in vivo and in vitro

Biomaterials composed of silk fibroin from both mulberry and non-mulberry silkworm varieties have been investigated for their utility in tissue engineering and drug delivery, but these studies have largely excluded any evaluation of host immune response. The present study compares the macrophage activation response towards mulberry (Bombyx mori, BM) and non-mulberry (Antheraea assamensis, AA) silk types, individually and as a blend (BA) in a partial thickness rat abdominal wall defect model and in vitro primary murine bone marrow-derived macrophage (BMDM) assay. Biologic materials composed of liver extracellular matrix (LECM) and small intestinal submucosa (SIS) ECM that are recognized for constructive tissue remodeling, and polypropylene mesh that is associated with pro-inflammatory macrophage phenotype activation are used as controls in the animal model. The AA silk graft shows a host response similar to SIS with few foreign body multinucleate giant cells, vascularization, high CD206 expression, and high M2-like: M1-like macrophage phenotype ratio. Exposure to AA silk degradation products in vitro induces a higher arginase: iNOS ratio in both naive BMDM and pro-inflammatory activated BMDM; and higher Fizz1: iNOS ratio in pro-inflammatory activated BMDM. These data suggest that the AA silk supports a pro-remodeling macrophage response with potential therapeutic applications.

Titanium Implant Surface Effects on Adherent Macrophage Phenotype: A Systematic Review.

Immunomodulatory biomaterials have the potential to stimulate an immune response able to promote constructive and functional tissue remodeling responses as opposed to persistent inflammation and scar tissue formation. As such, the controlled activation of macrophages and modulation of their phenotype through implant surface modification has emerged as a key therapeutic strategy. Methods: Online databases were searched for in vitro studies between January 1991 and June 2020 which examined the effect of titanium implant surface topography on the adherent macrophage phenotype at either the gene or protein level. Results: Thirty-nine studies were subsequently included for review. Although there was significant heterogeneity between studies, treatment of titanium surfaces increased the surface roughness or hydrophilicity, and hence increased macrophage attachment but decreased cell spreading. Physical coating of the titanium surface also tended to promote the formation of cell clusters. Titanium and titanium-zirconium alloy with a micro- or nano-scale rough topography combined with a hydrophilic surface chemistry were the most effective surfaces for inducing an anti-inflammatory phenotype in adherent macrophages, as indicated by significant changes in cytokine gene expression and or cytokine secretion profiles. Conclusions: The published data support the hypothesis that incorporation of specific topographical and physiochemical surface modifications to titanium can modulate the phenotypic response of adherent macrophages.

Transcriptomic Regulation of Macrophages by Matrix-Bound Nanovesicle-Associated Interleukin-33.

The innate immune response, particularly the phenotype of responding macrophages, has significant clinical implications in the remodeling outcome following implantation of biomaterials and engineered tissues. In general, facilitation of an anti-inflammatory (M2-like) phenotype is associated with tissue repair and favorable outcomes, whereas pro-inflammatory (M1-like) activation can contribute to chronic inflammation and a classic foreign body response. Biologic scaffolds composed of extracellular matrix (ECM) and, more recently, matrix-bound nanovesicles (MBV) embedded within the ECM are known to direct macrophages toward an anti-inflammatory phenotype and stimulate a constructive remodeling outcome. The mechanisms of MBV-mediated macrophage activation are not fully understood, but interleukin-33 (IL-33) within the MBV appears critical for M2-like activation. Previous work has shown that IL-33 is encapsulated within the lumen of MBV and stimulates phenotypical changes in macrophages independent of its canonical surface receptor stimulation-2 (ST2). In the present study, we used next-generation RNA sequencing to determine the gene signature of macrophages following exposure to MBV with and without intraluminal IL-33. MBV-associated IL-33 instructed an anti-inflammatory phenotype in both wild-type and st2-/- macrophages by upregulating M2-like and downregulating M1-like genes. The repertoire of genes regulated by ST2-independent IL-33 signaling were broadly related to the inflammatory response and crosstalk between cells of both the innate and adaptive immune systems. These results signify the importance of the MBV intraluminal protein IL-33 in stimulating a pro-remodeling M2-like phenotype in macrophages and provides guidance for the designing of next-generation biomaterials and tissue engineering strategies. Impact statement The phenotype of responding macrophages is predictive of the downstream remodeling response to an implanted biomaterial. The clinical impact of macrophage phenotype has motivated studies to investigate the factors that regulate macrophage activation. Matrix-bound nanovesicles (MBV) embedded within the extracellular matrix direct macrophages toward an anti-inflammatory (M2)-like phenotype that is indicative of a favorable remodeling response. Although the mechanisms of MBV-mediated macrophage activation are not fully understood, the intraluminal protein interleukin-33 (IL-33) is clearly a contributing signaling molecule. The present study identifies those genes regulated by MBV-associated IL-33 that promote a pro-remodeling M2-like macrophage activation state and can guide future therapies in regenerative medicine.

The molecular mechanism of constructive remodeling of a mechanically-loaded polymer

Large or repeated mechanical loads usually degrade polymers by accelerating fragmentation of their backbones but rarely, they can cause new backbone bonds to form. When these new bonds form faster than the original bonds break, mechanical degradation may be arrested or reversed in real time. Exploiting such constructive remodeling has proven challenging because we lack an understanding of the competition between bond-forming and bond-breaking reactions in mechanically-stressed polymers. Here we report the molecular mechanism and analysis of constructive remodeling driven by the macroradical products of mechanochemical fragmentation of a hydrocarbon backbone. By studying the changing compositions of a random copolymer of styrene and butadiene sheared at 10 °C in the presence of different additives we developed an approach to characterizing this growth/fracture competition, which is generalizable to other underlying chemistries. Our results demonstrate that constructive remodeling is achievable under practically relevant conditions, requires neither complex chemistries, elaborate macromolecular architectures or free monomers, and is amenable to detailed mechanistic interrogation and simulation. These findings constitute a quantitative framework for systematic studies of polymers capable of autonomously counteracting mechanical degradation at the molecular level.

CARD29: An Acellular Biologic Extracellular Matrix (ECM) Envelope for Cardiovascular Implantable Electronic Devices: Preclinical Evaluation

Background: Cardiovascular implantable electronic device (CIED) implant sites usually heal with hypovascular fibrotic capsule formation which can lead to complications including device movement, infection risk and reoperative challenges. An ECM CIED envelope at implant may promote stabilization, constructive tissue remodeling, reduced fibrosis and increased vascularity. Methods: Ten rabbits received single-chamber pacemakers (PMs) (St. Jude Medical) in ECM envelopes (CanGaroo® Envelope, Aziyo Biologics, Silver Spring, MD) and 10 received identical PMs without envelopes. One PM+ECM animal was evaluated at week 2, and remaining animals were evaluated at either week 13 or 26. Implant site tissues were analyzed by gross observation and histopathology. Results: PM+ECM animals had a 5-fold reduction in PM subcutaneous movement (migration or device flipping) compared to the PM only group. Histology showed progressive ECM envelope resorption by 13 and 26 weeks, with increased vascularization over time compared to PM only group at 13 and 26 weeks (Fig 1). PM+ECM animals had a lower average capsule thickness compared to PM only group - mean of 349 ± 165µm (mean ± SD) versus 417 ± 152µm at 13 weeks and a mean of 427 ± 141µm versus 439 ± 91µm at 26 weeks. Neovascularization score (blinded pathologist, 0-4 point scale) demonstrated a significantly higher PM+ECM group average score vs. PM only (2.1 vs 1.7, p<.05, Student’s t-test). Conclusion: Porcine small intestinal submucosa ECM envelopes promoted PM implant stabilization with implant site healing remarkable for reduced capsule thickness and increased vascularity, compared to PMs without envelopes in a preclinical model. ECM envelopes may provide meaningful clinical benefits for CIED implant pockets and warrant further investigation. Figure 1: Representative gross images of the subcutaneous pocket. 1A) Images demonstrating remodeling of the ECM envelope over time at 2, 13, and 26 weeks with increasing levels of vascularization. 1B) Comparison images from PMs without envelope implant at 13 and 26 weeks show lower vascularization levels and a flipped device at 26 weeks.

Late-stage rescue of visually guided behavior in the context of a significantly remodeled retinitis pigmentosa mouse model

Patients with progressive neurodegenerative disorder retinitis pigmentosa (RP) are diagnosed in the midst of ongoing retinal degeneration and remodeling. Here, we used a Pde6b-deficient RP gene therapy mouse model to test whether treatment at late disease stages can halt photoreceptor degeneration and degradative remodeling, while sustaining constructive remodeling and restoring function. We demonstrated that when fewer than 13% of rods remain, our genetic rescue halts photoreceptor degeneration, electroretinography (ERG) functional decline and inner retinal remodeling. In addition, in a water maze test, the performance of mice treated at 16 weeks of age or earlier was indistinguishable from wild type. In contrast, no efficacy was apparent in mice treated at 24 weeks of age, suggesting the photoreceptors had reached a point of no return. Further, remodeling in the retinal pigment epithelium (RPE) and retinal vasculature was not halted at 16 or 24 weeks of age, although there appeared to be some slowing of blood vessel degradation. These data suggest a novel working model in which restoration of clinically significant visual function requires only modest threshold numbers of resilient photoreceptors, halting of destructive remodeling and sustained constructive remodeling. These novel findings define the potential and limitations of RP treatment and suggest possible nonphotoreceptor targets for gene therapy optimization.

Repopulation of decellularised porcine pulmonary valves in the right ventricular outflow tract of sheep: Role of macrophages

The primary objective was to evaluate performance of low concentration SDS decellularised porcine pulmonary roots in the right ventricular outflow tract of juvenile sheep. Secondary objectives were to explore the cellular population of the roots over time. Animals were monitored by echocardiography and roots explanted at 1, 3, 6 (n = 4) and 12 months (n = 8) for gross analysis. Explanted roots were subject to histological, immunohistochemical and quantitative calcium analysis (n = 4 at 1, 3 and 12 months) and determination of material properties (n = 4; 12 months). Cryopreserved ovine pulmonary root allografts (n = 4) implanted for 12 months, and non-implanted cellular ovine roots were analysed for comparative purposes. Decellularised porcine pulmonary roots functioned well and were in very good condition with soft, thin and pliable leaflets. Morphometric analysis showed cellular population by 1 month. However, by 12 months the total number of cells was less than 50% of the total cells in non-implanted native ovine roots. Repopulation of the decellularised porcine tissues with stromal (α-SMA+; vimentin+) and progenitor cells (CD34+; CD271+) appeared to be orchestrated by macrophages (MAC 387+/ CD163low and CD163+/MAC 387−). The calcium content of the decellularised porcine pulmonary root tissues increased over the 12-month period but remained low (except suture points) at 401 ppm (wet weight) or below. The material properties of the decellularised porcine pulmonary root wall were unchanged compared to pre-implantation. There were some changes in the leaflets but importantly, the porcine tissues did not become stiffer. The decellularised porcine pulmonary roots showed good functional performance in vivo and were repopulated with ovine cells of the appropriate phenotype in a process orchestrated by M2 macrophages, highlighting the importance of these cells in the constructive tissue remodelling of cardiac root tissues.

Integration and functional performance of a decellularised porcine superflexor tendon graft in an ovine model of anterior cruciate ligament reconstruction

The objective was to evaluate the performance of decellularised porcine superflexor tendon (pSFT) as an anterior cruciate ligament (ACL) reconstruction device. The ACL of adult sheep was reconstructed with decellularised pSFT or ovine allograft SFT and animals sacrificed at 4, 12 and 26 weeks (n = 4 per group) for biological evaluation and 26 weeks (n = 6) for biomechanical evaluation of the grafts. Both grafts showed good in vivo performance with no major differences at macroscopic evaluation post euthanasia. Histopathology revealed an inflammatory reaction to both grafts at 4 weeks, which reduced by 26 weeks. There was advanced cellular ingrowth from 12 weeks, ligamentisation of intra-articular grafts, ossification and formation of Sharpey's fibers at the graft/bone junctions. Immunohistochemistry showed that at 4 and 12 weeks, the host response was dominated by CD163+ M2 macrophages and a cell infiltrate comprising α-SMA + myofibroblasts, CD34+ and CD271+ progenitor cells. At 26 weeks the biomechanical properties of decellularised pSFT and oSFT grafts were comparable, with all grafts failing in the intra-articular region. This study provides new insight into constructive remodelling of tendons used for ACL replacement and evidence of integration and functional performance of a decellularised xenogeneic tendon with potential as an alternative for ACL reconstruction.

Functionalized Silk Vascular Grafts with Decellularized Human Wharton's Jelly Improves Remodeling via Immunomodulation in Rabbit Jugular Vein.

Cell-free polymeric tissue-engineered vascular grafts (TEVGs) have shown great promise towards clinical translation; however, their limited bioactivity and remodeling ability challenge this cause. Here, a novel cell-free bioresorbable small diameter silk TEVG system functionalized with decellularized human Wharton's jelly (dWJ) matrix is developed and successfully implanted as interposition grafts into rabbit jugular vein. Implanted TEVGs remain patent for two months and integrate with host tissue, demonstrating neo-tissue formation and constructive remodeling. Mechanistic analysis reveals that dWJ matrix is a reservoir of various immunomodulatory cytokines (Interleukin-8, 6, 10, 4 and tumor necrosis factor alpha (TNF-α)), which aids in upregulating M2 macrophage-associated genes facilitating pro-remodeling behavior. Besides, dWJ treatment to human endothelial cells upregulates the expression of functional genes (cluster of differentiation 31 (CD31), endothelial nitric oxide synthase (eNOS), and vascular endothelial (VE)-cadherin), enables faster cell migration, and elevates nitric oxide (NO) production leading to the in situ development of endothelium. The dWJ functionalized silk TEVGs support increased host cell recruitment than control, including macrophages and vascular cells. It endows superior graft remodeling in terms of a dense medial layer comprising smooth muscle cells and elevates the production of extracellular matrix proteins (collagen and elastin). Altogether, these findings suggest that dWJ functionalization imitates the usefulness of cell seeding and enables graft remodeling.

Tissue response, macrophage phenotype, and intrinsic calcification induced by cardiovascular biomaterials: Can clinical regenerative potential be predicted in a rat subcutaneous implant model?

The host immune response to an implanted biomaterial, particularly the phenotype of infiltrating macrophages, is a key determinant of biocompatibility and downstream remodeling outcome. The present study used a subcutaneous rat model to compare the tissue response, including macrophage phenotype, remodeling potential, and calcification propensity of a biologic scaffold composed of glutaraldehyde-fixed bovine pericardium (GF-BP), the standard of care for heart valve replacement, with those of an electrospun polycarbonate-based supramolecular polymer scaffold (ePC-UPy), urinary bladder extracellular matrix (UBM-ECM), and a polypropylene mesh (PP). The ePC-UPy and UBM-ECM materials induced infiltration of mononuclear cells throughout the thickness of the scaffold within 2 days and neovascularization at 14 days. GF-BP and PP elicited a balance of pro-inflammatory (M1-like) and anti-inflammatory (M2-like) macrophages, while UBM-ECM and ePC-UPy supported a dominant M2-like macrophage phenotype at all timepoints. Relative to GF-BP, ePC-UPy was markedly less susceptible to calcification for the 180 day duration of the study. UBM-ECM induced an archetypical constructive remodeling response dominated by M2-like macrophages and the PP caused a typical foreign body reaction dominated by M1-like macrophages. The results of this study highlight the divergent macrophage and host remodeling response to biomaterials with distinct physical and chemical properties and suggest that the rat subcutaneous implantation model can be used to predict in vivo biocompatibility and regenerative potential for clinical application of cardiovascular biomaterials.

Tissue‐Engineered Vascular Grafts: Emerging Trends and Technologies

AbstractVascular tissue engineering has made prodigious progress in recent years by converging multidisciplinary approaches. Latest technological advancements foster the development of next‐generation tissue‐engineered vascular grafts (TEVGs) for treating various vasculopathies. While traditional therapeutic methods rely on bypassing the severely damaged vessels with synthetic counterparts with no growth potential, contemporary perspectives focus on biodegradable conduits bestowing an inherent remodeling capability. This review highlights emerging innovative trends and technologies adopted to pragmatically fulfill current scientific needs while improving overall TEVG performance in pre‐clinical and clinical settings. A comprehensive overview of various milestones achieved in the past few decades is first summarized, followed by an appraisal of the significant hurdles for clinical translation. The latest techniques to rationally address critical challenges, viz., intimal hyperplasia, thrombosis, constructive graft remodeling, and adequate neo‐tissue formation are discussed. Finally, an update on ongoing clinical trials is provided and future perspectives required to persuade TEVGs to become a clinical reality are delineated.

Mimicking Physiologically Relevant Hepatocyte Zonation Using Immunomodulatory Silk Liver Extracellular Matrix Scaffolds toward a Bioartificial Liver Platform.

Mimicking nativelike metabolic zonation is indispensable to develop an efficient bioartificial liver model, as it facilitates physiological cues, hepatocyte polarity, and phenotypic functions. The present study shows the first evidence of hepatocyte metabolic heterogeneity in an in vitro liver model encompassing liver extracellular matrix (ECM)-functionalized silk scaffolds (LECM-SF) by altering ECM proportion. Upon static culture, individual LECM-SF scaffold supports differential synthetic and metabolic functions of cultured primary neonatal rat hepatocytes (PNRHs), owing to discrete biophysical attributes. A single in vitro liver system comprising PNRHs seeded LECM-SF scaffolds assisting periportal to pericentral gradient functions is stacked and matured in a perfusion bioreactor to simulate oxygen gradient. The scaffold with high ECM supports periportal-specific albumin synthesis, urea secretion, and bile duct formation, albeit scaffold with low ECM supports pericentral-specific cytochrome P450 activity. Extensive physicochemical characterizations confirmed the stability and interconnected porous network of scaffolds, signifying cellular infiltration and bidirectional nutrient diffusion. Furthermore, scaffolds demonstrate minimal thrombogenicity, reduced foreign-body response, and enhanced pro-remodeling macrophage activation, supporting constructive tissue remodeling. The developed liver model with zone-specific functions would be a promising avenue in bioartificial liver and drug screening.