Cellular Repopulation Research Articles

Myriad factors and pathways influencing tumor radiotherapy resistance

Abstract Radiotherapy is a cornerstone in the treatment of various tumors, yet radioresistance often leads to treatment failure and tumor recurrence. Several factors contribute to this resistance, including hypoxia, DNA repair mechanisms, and cancer stem cells. This review explores the diverse elements that drive tumor radiotherapy resistance. Historically, resistance has been attributed to cellular repair and tumor repopulation, but recent research has expanded this understanding. The tumor microenvironment – characterized by hypoxia, immune evasion, and stromal interactions – further complicates treatment. Additionally, molecular mechanisms such as aberrant signaling pathways, epigenetic modifications, and non-B-DNA structures play significant roles in mediating resistance. This review synthesizes current knowledge, highlighting the interplay of these factors and their clinical implications. Understanding these mechanisms is crucial for developing strategies to overcome resistance and improve therapeutic outcomes in cancer patients.

Iron Chelators Augment Large Osteochondral Allograft Osseointegration in a Preclinical Canine Model.

Osteochondral allograft transplantation (OCAT) can be a successful joint restoration treatment option for large post-traumatic articular defects but is still associated with significant revision and failure rates. Despite recent advances that have improved OCAT success, insufficient osteochondral allograft (OCA) osseointegration remains a major cause of failure. Deferoxamine (DFO) is an effective angiogenic and osteo-anabolic iron chelator that consistently promotes bone neovascularization and regeneration. This study was designed to investigate local delivery of DFO for augmenting OCA osseointegration using a preclinical canine model for OCAT in the knee and hip as commonly affected joints. On Institutional Animal Care and Use Committee (IACUC) approval, 12 purpose-bred dogs underwent OCAT of the femoral head or femoral condyles with DFO or DFO-free (controls) microspheres in recipient sites. OCA revascularization, cellular repopulation, and integration were evaluated based on functional, diagnostic imaging, microcomputed tomography, histology, and immunohistochemistry outcome measures. Local delivery of DFO into OCAT recipient sites was associated with maintained or improved joint function, superior radiographic appearance, significantly greater trabecular thickness, higher bone volume, and new bone ingrowth compared with DFO-free controls. OCA osseointegration is dependent on cellular repopulation and neovascularization, resulting in new bone ingrowth through creeping substitution, and insufficient osseointegration with resorption and subsidence of the OCA remains a major cause of failure after transplantation. The results of this study suggest that local delivery of DFO using a controlled microsphere release system may reduce resorption and improve revascularization and cellular repopulation to increase new bone ingrowth, potentially expediting OCA osseointegration after transplantation.

Cellular Enhancement of Frozen Meniscus Allograft Combining Native Meniscus and Mesenchymal Stromal Cell Injections.

This proof-of-concept study investigated an improved cell-based injection therapy combining mesenchymal stem cells (MSCs) and meniscus cells (MCs) to support superior meniscus allograft repopulation and early revival compared to injecting MSCs alone. In this controlled laboratory study, frozen meniscus allograft samples were injected vertically with a cell suspension containing different ratios of MSCs and MCs or control (lactated ringers) and cultured for 28 days. Samples were analyzed weekly for cell viability, migration, and metabolism using histological and biochemical assays. Tissue medium was analyzed for matrix metalloproteinase (MMP) expression using zymography. Cellular repopulation of frozen allografts injected with different cell suspensions was validated by immunohistochemistry. Significant higher DNA content was evidenced in grafts treated with suspensions of MCs or MC:MSC (1:4 ratio). Cell metabolic activity was significantly different between all treated groups and control group after 1 week. Allografts injected with MCs showed significantly more cell proliferation than injections with MSCs. MMP2 activity was detected in medium of all grafts cellularized with MCs with or without MSCs. Scanning electron microscopy (SEM) analysis showed resolution of the needle puncture, but not in the control group. Cell labeling of MCs upon injection of mixed MC:MSC suspensions revealed a gradual increase in the cell ratio. The findings of this study establish that injection of MCs with or without MSCs enhances the cellularity of meniscus allograft to support early graft revival and remodeling.

Development, in vitro and ex vivo evaluation of bioadhesive colloidal systems for curcumin skin delivery aiming the antioxidant and photoprotective activities

This work aimed to develop bioadhesive colloidal systems containing curcumin (CUR) for antioxidant and photoprotective applications. Gels and emulgels were prepared and characterized as rheology, texture profile analysis, softness, bioadhesive properties using porcine skin, skin permeability and antioxidant activity. Moreover, the formulations were investigated for their photoprotective potential using UVB-irradiated L929 fibroblasts, analyzing cellular viability with or without UVB irradiation, determination of reactive oxygen species (ROS), DNA damage, and cellular migration. The presence of CUR in the systems did not influence their mechanical textural properties and the rheological behavior (p > 0.05), displaying shear thinning characteristics, hysteresis area and rheopexy. The formulations exhibited important antioxidant activity, by scavenging radicals DPPH•, ABTS•+, superoxide anion, and reducing ferric ions. Gel-containing CUR (GEL CUR) exhibited a less cytotoxic effect in L929 fibroblasts than free CUR, besides increasing cell viability in UVB exposure. The treatment with GEL CUR in UVB-irradiated cells attenuated oxidative stress, displaying the ability to scavenge ROS generation (31.3 %), decrease DNA damage (≈30 %), and promote cellular repopulation at wound-scratch assay (8 % after 48h), which was statistically superior in comparison to the control and showing relevant activity. These colloidal systems showed suitable properties for topical application, with good antioxidant activity, permeation ability and potential application for the prevention of UVB-induced skin photodamage.

Tissue formation and host remodeling of an elastomeric biodegradable scaffold in an ovine pulmonary leaflet replacement model.

In pursuit of a suitable scaffold material for cardiac valve tissue engineering applications, an acellular, electrospun, biodegradable polyester carbonate urethane urea (PECUU) scaffold was evaluated as a pulmonary valve leaflet replacement in vivo. In sheep (n = 8), a single pulmonary valve leaflet was replaced with a PECUU leaflet and followed for 1, 6, and 12 weeks. Implanted leaflet function was assessed in vivo by echocardiography. Explanted samples were studied for gross pathology, microscopic changes in the extracellular matrix, host cellular re-population, and immune responses, and for biomechanical properties. PECUU leaflets showed normal leaflet motion at implant, but decreased leaflet motion and dimensions at 6 weeks. The leaflets accumulated α-SMA and CD45 positive cells, with surfaces covered with endothelial cells (CD31+). New collagen formation occurred (Picrosirius Red). Accumulated tissue thickness correlated with the decrease in leaflet motion. The PECUU scaffolds had histologic evidence of scaffold degradation and an accumulation of pro-inflammatory/M1 and anti-inflammatory/M2 macrophages over time in vivo. The extent of inflammatory cell accumulation correlated with tissue formation and polymer degradation but was also associated with leaflet thickening and decreased leaflet motion. Future studies should explore pre-implant seeding of polymer scaffolds, more advanced polymer fabrication methods able to more closely approximate native tissue structure and function, and other techniques to control and balance the degradation of biomaterials and new tissue formation by modulation of the host immune response.

Predicting the Tumour Response to Radiation by Modelling the Five Rs of Radiotherapy Using PET Images

Despite the intensive use of radiotherapy in clinical practice, its effectiveness depends on several factors. Several studies showed that the tumour response to radiation differs from one patient to another. The non-uniform response of the tumour is mainly caused by multiple interactions between the tumour microenvironment and healthy cells. To understand these interactions, five major biologic concepts called the “5 Rs” have emerged. These concepts include reoxygenation, DNA damage repair, cell cycle redistribution, cellular radiosensitivity and cellular repopulation. In this study, we used a multi-scale model, which included the five Rs of radiotherapy, to predict the effects of radiation on tumour growth. In this model, the oxygen level was varied in both time and space. When radiotherapy was given, the sensitivity of cells depending on their location in the cell cycle was taken in account. This model also considered the repair of cells by giving a different probability of survival after radiation for tumour and normal cells. Here, we developed four fractionation protocol schemes. We used simulated and positron emission tomography (PET) imaging with the hypoxia tracer 18F-flortanidazole (18F-HX4) images as input data of our model. In addition, tumour control probability curves were simulated. The result showed the evolution of tumours and normal cells. The increase in the cell number after radiation was seen in both normal and malignant cells, which proves that repopulation was included in this model. The proposed model predicts the tumour response to radiation and forms the basis for a more patient-specific clinical tool where related biological data will be included.

Repopulation of decellularized organ scaffolds with human pluripotent stem cell-derived pancreatic progenitor cells

Diabetes is an emerging global epidemic that affects more that 285 million people worldwide. Engineering of endocrine pancreas tissue holds great promise for the future of diabetes therapy. Here we demonstrate the feasibility of re-engineering decellularized organ scaffolds using regenerative cell source. We differentiated human pluripotent stem cells (hPSC) toward pancreatic progenitor (PP) lineage and repopulated decellularized organ scaffolds with these hPSC-PP cells. We observed that hPSCs cultured and differentiated as aggregates are more suitable for organ repopulation than isolated single cell suspension. However, recellularization with hPSC-PP aggregates require a more extensive vascular support, which was found to be superior in decellularized liver over the decellularized pancreas scaffolds. Upon continued culture for nine days with chemical induction in the bioreactor, the seeded hPSC-PP aggregates demonstrated extensive and uniform cellular repopulation and viability throughout the thickness of the liver scaffolds. Furthermore, the decellularized liver scaffolds was supportive of the endocrine cell fate of the engrafted cells. Our novel strategy to engineer endocrine pancreas construct is expected to find potential applications in preclinical testing, drug discovery and diabetes therapy.

Elastin Barrier Membranes for Guided Tissue Regeneration Technologies

This article discusses the prospects for the use of new elastin barrier membranes manufactured using adapted technologies for the selective isolation of the elastin component from the extracellular xenogenic matrix of the pericardium ligamentous apparatus: (1) by high-temperature extraction under pressure; (2) cyanogen bromide method. A commercial material, Geistlich Bio–Gide® membrane (BG), was used as a control comparison group. It is shown that the materials of group (1) have a high degree of biocompatibility, exceeding the indicators of the control group BG. Based on the results of an study in a model of subcutaneous heterotopic implantation in rats, it was shown that elastin BM has a chemoattractant effect on the mesenchymal recipient cells and, unlike the control, is able to integrate to a high degree into the surrounding recipient tissues. At the same time, the materials of group (1) had a pronounced proangiogenic effect. Thus, it has been shown that elastin BM groups (1) have a medium-term barrier function and are able to induce full-fledged cellular repopulation and local neoangiogenesis, which can be useful in clinical practice, primarily in GTR technologies (with gingival flap augmentation) or when used together with other BM as an angiogenesis inducer to ensure formation of the vascular bed in GBR technologies of bone tissue.

Abstract 10957: Six Month Patency of Bioengineered Human Acellular Vessels in a Primate Model of Coronary Artery Bypass Grafting

Introduction: Limitations of coronary artery bypass grafting (CABG) include the availability, quality, and long-term patency of autologous saphenous vein grafts. An “off-the-shelf” human acellular vessel (HAV) bioengineered from banked human vascular cells, decellularized before implantation, and capable of host cellular regeneration may address these issues. Therefore, we assessed early patency and tolerability of a small diameter HAV as a CABG conduit in a baboon surgical model. Methods: Adult male, non-immunosuppressed baboons (n = 3; 30.5 - 37.0 kg) underwent CABG via sternotomy with cardiopulmonary bypass. The right coronary artery (RCA) was ligated and bypassed with a 3.5 mm inner diameter HAV. Intraoperative blood flow was determined by transit-time flow measurement (TTFM). HAV patency was assessed by computed tomography angiography (CTA) at 1, 3 and 6 months, and confirmed by left heart catheterization at the 6 month study endpoint. Explanted tissues were processed for histological analysis of host cellular repopulation and HAV remodeling. Results: Three animals were successfully bypassed with a 3.5 mm HAV as aorta-RCA CABG conduits with resulting TTFM blood flow rates of 22 ± 7.7 mL/min. CTA at 1, 3 and 6 months demonstrated patency of the HAV with no significant stenosis or dilatation. There were no HAV related adverse events for the duration of the study. Histological staining of 6 month explanted tissues showed recellularization and remodeling of the HAV with a microcapillary-containing neo-adventitial layer, alpha smooth muscle actin expressing cells within the HAV wall, and endothelialization of the HAV lumen. Conclusion: A small diameter, readily available HAV was successfully implanted in a primate model of CABG and remained patent up to 6 months with no evidence of dilatation. In addition, the HAVs were thoroughly recellularized and remodeled by host cells. These bioengineered vessels may prove to be durable long-term conduit options in CABG.

Paper 18: Effect of Bone Marrow Aspirate Concentrate on Osteochondral Allograft Transplantation Incorporation: A Prospective, Randomized, Single Blind Investigation

Objectives:Osteochondral allograft transplantation for cartilage defects of the knee has demonstrated excellent long-term clinical outcomes and survival, which largely depends on successful graft osseointegration. Biologics have been suggested as a viable adjunct to enhance successful healing in several musculoskeletal applications. In the context of OCA, early results have suggested that BMAC may improve cellular repopulation and viability within the osseus portion of an implanted graft. However, few clinical studies to date have investigated the impact of BMAC on patient outcomes following OCA. The purpose of this study was to investigate the effect of bone marrow aspirate concentrate (BMAC) on osseointegration and patient-reported outcome metrics (PROMs) after osteochondral allograft transplantation in a prospective, randomized controlled single-blinded trial.Methods:Patients undergoing osteochondral allograft transplantation of the knee were consented and enrolled. Prior to surgery, patients were randomized into either the BMAC or sham incision groups. In the BMAC group, the osteochondral allograft plug was soaked in BMAC for a minimum of 2 minutes prior to implantation. All patients underwent postoperative computed tomography (CT) scanning at 6 months postoperatively and completed PROMs preoperatively, 6 months, and 1 year postoperatively. Two board-eligible orthopaedic surgeons blinded to treatment allocation independently assessed and graded each CT according to the ACTOCA system proposed by Gelber et al.Results:Thirty-six patients enrolled between April 2018 to December 2020 (17 female, 19 male) were included for analysis. There were no significant differences between the BMAC and non-BMAC groups in graft signal density (Grader 1: p=0.283, Grader 2: p=0.467), osseous integration (both graders: p=0.489), surface percentage with discernible cleft (Grader 1: 0.287, Grader 2: 0.469), or intra-articular fragments (Grader 1: p=0.617, Grader 2: p=0.810) (Table 1). A significantly greater number of patients receiving BMAC demonstrating cystic changes <3 mm (Grader 1: p=0.015, Grader 2: p=0.05) (Figure 1). At 1 year, BMAC patients reported significantly better WOMAC Pain (87.82±14.26 vs 75.80±15.56, p=0.043) and trended towards improved PROMIS Pain (54.14±8.31 vs 61.79±5.24, p=0.09).Conclusions:Patients receiving BMAC soaked OCA grafts demonstrated no difference from controls with respect to graft signal intensity, osseous integration, intra-articular fragments, or discernible graft-host clefts at 6-months postoperatively. BMAC patients had a significantly greater occurrence of small (<3 mm) cystic changes. At 1 year, BMAC patients reported significantly less pain than controls on WOMAC Pain, with similar trends on PROMIS Pain Interference.Table 1.Postoperative Imaging Characteristics (ACTOCA) by Grader.Figure 1.Distribution of cystic changes between BMAC and control groups.

Conformational changes of exogenous deposited fibronectin on decellularized aortic conduits after implantation

Fibronectin is an abundant and ubiquitous extracellular matrix component that regulates a wide range of cell functions by directly interacting with cell surface integrin receptors. In a prior study, we observed that fibronectin can accelerate the autologous cellular repopulation of implanted decellularized grafts, enhancing their adhesion capacity and biocompatibility, through fibronectin fibrillogenesis. The supportive effect of deposited fibronectin was shown to be dissimilar in different regions of implanted decellularized grafts. This led us to question whether there is any difference in conformation of fibronectin deposited on the luminal and adventitial surfaces of decellularized aortic conduits after implantation. In this study, we doubly labelled fibronectin with a fluorescent resonance energy transfer (FRET) pair and use this to analyse differences in conformational changes of adsorbed fibronectin at different sites of the decellularized aortic conduits.

Impact of freeze-thaw cytoablation on aqueous outflow patterns in ex vivo anterior chamber perfusion cultures and whole eyes

Background: Porcine eyes have been widely used as ex vivo models in glaucoma research, as they share similar features with human eyes. Freeze-thawing is a non-invasive technique that has been used to obliterate living cells in anterior segment ex vivo cultures, to prepare them for further research such as cellular repopulation. This technique has previously been shown to reduce the intraocular pressure (IOP) in porcine eyes. The aim of this study was to investigate whether freeze-thaw cytoablation causes corresponding canalogram outflow changes in perfused anterior segment cultures (AFT) and whole porcine eyes (WFT). We hypothesized that the known IOP drop in AFT after trabecular meshwork ablation by freeze-thaw would be accompanied by a similarly large change in the distal outflow pattern. Methods: Two-dye (fluorescein and Texas red) reperfusion canalograms were used to compare the outflow time before and after two -80°C cycles of freeze-thaw. We assigned 28 freshly enucleated porcine eyes to four groups: perfused anterior segment dye controls (ACO, n = 6), perfused whole eye dye controls (WCO, n = 6), freeze-thaw treated anterior segment cultures (AFT, n = 10), and freeze-thaw treated whole eyes (WFT, n = 6). Results: In control groups ACO and WCO, the two different dyes had similar filling times. In AFT, the outflow pattern and filling times were unchanged. In WFT, the temporal superior quadrant filled more slowly (p = 0.042) while all others remained unchanged. The qualitative appearance of distal outflow spaces was altered only in some eyes. Conclusions: Freeze-thaw cytoablation caused neither loss nor leakage of distal outflow structures. Surprisingly, the loss of an intact trabecular meshwork over the entire circumference did not result in a general acceleration of quadrant outflow times. The results validate freeze-thawing as a method to generate an extracellular matrix without major structural changes.

Inflammatory and regenerative processes in bioresorbable synthetic pulmonary valves up to two years in sheep–Spatiotemporal insights augmented by Raman microspectroscopy

In situ heart valve tissue engineering is an emerging approach in which resorbable, off-the-shelf available scaffolds are used to induce endogenous heart valve restoration. Such scaffolds are designed to recruit endogenous cells in vivo, which subsequently resorb polymer and produce and remodel new valvular tissue in situ. Recently, preclinical studies using electrospun supramolecular elastomeric valvular grafts have shown that this approach enables in situ regeneration of pulmonary valves with long-term functionality in vivo. However, the evolution and mechanisms of inflammation, polymer absorption and tissue regeneration are largely unknown, and adverse valve remodeling and intra- and inter-valvular variability have been reported. Therefore, the goal of the present study was to gain a mechanistic understanding of the in vivo regenerative processes by combining routine histology and immunohistochemistry, using a comprehensive sheep-specific antibody panel, with Raman microspectroscopy for the spatiotemporal analysis of in situ tissue-engineered pulmonary valves with follow-up to 24 months from a previous preclinical study in sheep. The analyses revealed a strong spatial heterogeneity in the influx of inflammatory cells, graft resorption, and foreign body giant cells. Collagen maturation occurred predominantly between 6 and 12 months after implantation, which was accompanied by a progressive switch to a more quiescent phenotype of infiltrating cells with properties of valvular interstitial cells. Variability among specimens in the extent of tissue remodeling was observed for follow-up times after 6 months. Taken together, these findings advance the understanding of key events and mechanisms in material-driven in situ heart valve tissue engineering. Statement of significanceThis study describes for the first time the long-term in vivo inflammatory and regenerative processes that underly in situ heart valve tissue engineering using resorbable synthetic scaffolds. Using a unique combinatorial analysis of immunohistochemistry and Raman microspectroscopy, important spatiotemporal variability in graft resorption and tissue formation was pinpointed in in situ tissue-engineered heart valves, with a follow-up time of up to 24 months in sheep. This variability was correlated to heterogenous regional cellular repopulation, most likely instigated by region-specific differences in surrounding tissue and hemodynamics. The findings of this research contribute to the mechanistic understanding of in situ tissue engineering using resorbable synthetics, which is necessary to enable rational design of improved grafts, and ensure safe and robust clinical translation.

Impact of freeze-thaw cytoablation on aqueous outflow patterns in ex vivo anterior chamber perfusion cultures and whole eyes

Background: Porcine eyes have been widely used as ex vivo models in glaucoma research, as they share similar features with human eyes. Freeze-thawing is a non-invasive technique that has been used to obliterate living cells in anterior segment ex vivo cultures, to prepare them for further research such as cellular repopulation. This technique has previously been shown to reduce the intraocular pressure (IOP) in porcine eyes. The aim of this study was to investigate whether freeze-thaw cytoablation causes corresponding canalogram outflow changes in perfused anterior segment cultures (AFT) and whole porcine eyes (WFT). We hypothesized that the known IOP drop in AFT after trabecular meshwork ablation by freeze-thaw would be accompanied by a similarly large change in the distal outflow pattern. Methods: Two-dye (fluorescein and Texas red) reperfusion canalograms were used to compare the outflow time before and after two -80°C cycles of freeze-thaw. We assigned 28 freshly enucleated porcine eyes to four groups: perfused anterior segment dye controls (ACO, n = 6), perfused whole eye dye controls (WCO, n = 6), freeze-thaw treated anterior segment cultures (AFT, n = 10), and freeze-thaw treated whole eyes (WFT, n = 6). Results: In control groups ACO and WCO, the two different dyes had similar filling times. In AFT, the outflow pattern and filling times were unchanged. In WFT, the temporal superior quadrant filled more slowly (p = 0.042) while all others remained unchanged. The qualitative appearance of distal outflow spaces was altered only in some eyes. Conclusions: Freeze-thaw cytoablation caused neither loss nor leakage of distal outflow structures. Surprisingly, the loss of an intact trabecular meshwork over the entire circumference did not result in a general acceleration of quadrant outflow times. The results validate freeze-thawing as a method to generate an extracellular matrix without major structural changes.

In vivo Analysis of the Resistance of the Meshes to Escherichia coli Infection.

Background: The mesh infection is mostly related to the gram-negative bacteria, such as Escherichia coli (E. coli) for emergency surgery of incarcerated hernia. However, few study investigated the effects of E. coli concentration, mesh materials and antibiotic prophylaxis on mesh infection after hernioplasty. The aim of this study was to evaluate the bacterial resistance to E. coli for three different materials of mesh, and to measure the minimum E. coli concentration for mesh infection with and without antibiotic prophylaxis in a rat model.Methods: Three types of mesh (polytetrafluoroethylene, polypropylene, and biologic meshes) were used in the repair of an acute ventral hernia rat model in the setting of different concentrations of E. coli loads and antibiotics. At the 8th day after surgery, mesh samples were sent for microbiologic and histologic analyses.Results: The positive rates of bacterial culture increased with E. coli concentration. The biologic mesh showed better bacterial resistance compared to polytetrafluoroethylene mesh and polypropylene mesh when the concentration of E. coli ranges from 106 CFU/ml to 108 CFU/ml (P = 0.002 and P = 0.029, respectively). Prophylactical ceftriaxone treatment could not decrease the colonization rate of E. coli at 106 CFU/ml or 108 CFU/ml in each group (P > 0.05). The scores of neovascularization in polypropylene mesh and biologic mesh were similar, which was higher than that of polytetrafluoroethylene mesh (P < 0.05). Compared with other meshes, biologic mesh showed better tolerance to 106 CFU/ml E. coli with respect to inflammation, depth of inflammation, neovascularization, cellular repopulation and foreign body giant cells.Conclusion: The biologic mesh had better E. coli resistance compared to polytetrafluoroethylene mesh and polypropylene mesh when the E. coli concentration is higher than 106 CFU/ml in rats. Antibiotic prophylaxis was useful when the contamination was not particularly severe.

Human meniscus allograft augmentation by allogeneic mesenchymal stromal/stem cell injections.

Meniscus allograft transplantations (MATs) represent established surgical procedures with proven outcomes. Yet, storage as frozen specimens and limited cellular repopulation may impair graft viability. This proof‐of‐concept study tests the feasibility of injecting allogeneic mesenchymal stromal/stem cells (MSCs) in meniscus allograft tissue. We investigated the injectable cell quantity, survival rate, migration, and proliferation ability of MSCs up to 28 days of incubation. In this controlled laboratory study, seven fresh‐frozen human allografts were injected with human allogeneic MSCs. Cells were labeled and histological characteristics were microscopically imaged up to 28 days. Mock‐injected menisci were included as negative controls in each experiment. Toluidine blue staining demonstrated that a 100‐µl volume can be injected while retracting and rotating the inserted needle. Immediately after injection, labeled MSCs were distributed throughout the injection channel and eventually migrated into the surrounding tissues. Histological assessment revealed that MSCs cluster in disc‐like shapes, parallel to the intrinsic lamination of the meniscus and around the vascular network. Quantification showed that more than 60% of cells were present in horizontally injected grafts and more than 30% were observed in vertically injected samples. On Day 14, cells adopted a spindle‐shaped morphology and exhibited proliferative and migratory behaviors. On Day 28, live/dead ratio assessment revealed an approximately 80% cell survival. The study demonstrated the feasibility of injecting doses of MSCs (>0.1 million) in meniscus allograft tissue with active cell proliferation, migration, and robust cell survival.

Optically activated and interrogated plasmonic hydrogels for applications in wound healing.

We disclose the use of hybrid materials featuring Au/Ag core/shell nanorods in porous chitosan/polyvinyl alcohol scaffolds for applications in tissue engineering and wound healing. The combination of Au and Ag in a single construct provides synergistic opportunities for optical activation of functions as near infrared laser tissue bonding, and remote interrogation to return parameters of prognostic relevance in wound healing monitoring. In particular, the bimetallic component ensures optical tunability, enhanced shelf life and photothermal stability, serves as a reservoir of germicidal silver cations, and changes in near-infrared and visible color according to the environmental level of oxidative stress. At the same time, the polymeric blend is ideal to bind connective tissue upon photothermal activation, and to support fabrication processes that provide high porosity, such as electrospinning, thus putting all the premises for cellular repopulation and antimicrobial protection.

Flow with variable pulse frequencies accelerates vascular recellularization and remodeling of a human bioscaffold.

Despite significant advances in vascular tissue engineering, the ideal graft has not yet been developed and autologous vessels remain the gold standard substitutes for small diameter bypass procedures. Here, we explore the use of a flow field with variable pulse frequencies over the regeneration of an ex vivo-derived human scaffold as vascular graft. Briefly, human umbilical veins were decellularized and used as scaffold for cellular repopulation with human smooth muscle cells (SMC) and endothelial cells (EC). Over graft development, the variable flow, which mimics the real-time cardiac output of an individual performing daily activities (e.g., resting vs. exercising), was implemented and compared to the commonly used constant pulse frequency. Results show marked differences on SMC and EC function, with changes at the molecular level reflecting on tissue scales. First, variable frequencies significantly increased SMC proliferation rate and glycosaminoglycan production. These results can be tied with the SMC gene expression that indicates a synthetic phenotype, with a significant downregulation of myosin heavy chain. Additionally and quite remarkably, the variable flow frequencies motivated the re-endothelialization of the grafts, with a quiescent-like structure observed after 10 days of conditioning, contrasting with the low surface coverage and unaligned EC observed under constant frequency (CF). Besides, the overall biomechanics of the generated grafts (conditioned with both pulsed and CFs) evidence a significant remodeling after 55 days of culture, depicted by high burst pressure and Young's modulus. These last results demonstrate the positive recellularization and remodeling of a human-derived scaffold toward an arterial vessel.

The CCL2/CCR2 axis is critical to recruiting macrophages into acellular nerve allograft bridging a nerve gap to promote angiogenesis and regeneration

Acellular nerve allografts (ANAs) are increasingly used to repair nerve gaps following injuries. However, these nerve scaffolds have yet to surpass the regenerative capabilities of cellular nerve autografts; improved understanding of their regenerative mechanisms could improve design. Due to their acellular nature, both angiogenesis and diverse cell recruitment is necessary to repopulate these scaffolds to promote functional regeneration. We determined the contribution of angiogenesis to initial cellular repopulation of ANAs used to repair nerve gaps, as well as the signaling that drives a significant portion of this angiogenesis. Wild-type (WT) mice with nerve gaps repaired using ANAs that were treated with an inhibitor of VEGF receptor signaling severely impaired angiogenesis within ANAs, as well as hampered cell repopulation and axon extension into ANAs. Similarly, systemic depletion of hematogenous-derived macrophages, but not neutrophils, in these mice models severely impeded angiogenesis and subsequent nerve regeneration across ANAs suggesting hematogenous-derived macrophages were major contributors to angiogenesis within ANAs. This finding was reinforced using CCR2 knockout (KO) models. As macrophages represented the majority of CCR2 expressing cells, a CCR2 deficiency impaired angiogenesis and subsequent nerve regeneration across ANAs. Furthermore, an essential role for CCL2 during nerve regeneration across ANAs was identified, as nerves repaired using ANAs had reduced angiogenesis and subsequent nerve regeneration in CCL2 KO vs WT mice. Our data demonstrate the CCL2/CCR2 axis is important for macrophage recruitment, which promotes angiogenesis, cell repopulation, and subsequent nerve regeneration and recovery across ANAs used to repair nerve gaps.

Povidone-iodine Does Not Affect Acellular Dermal Matrix Integration in Patients Undergoing 2-staged, Prepectoral, Breast Reconstructive Surgery

Background:Povidone-iodine has been traditionally used as an antimicrobial agent to irrigate the breast pocket and rinse the prosthesis before placement in the pocket. Use of povidone-iodine with breast implants was banned from 2000 to 2017. During this period, acellular dermal matrix (ADM) was introduced to breast surgery. In nonclinical studies, povidone-iodine can impair collagen synthesis and kill fibroblasts. Cellular repopulation was critical for ADM integration. Whether povidone-iodine impacts ADM integration was unknown.Methods:Patients who underwent immediate, prepectoral, 2-staged breast reconstruction were included in this retrospective study. Study population was divided into povidone-iodine–treated patients and triple-antibiotic–treated patients. The breast pockets were rinsed with the antimicrobial agent, and the prostheses and ADMs were presoaked in the agent perioperatively. At implant exchange, the extent of ADM integration was clinically assessed. ADM integration was defined as >25% of matrix vascularization. ADM integration and postoperative complications were compared between the groups.Results:A total of 111 patients (257 reconstructions) were included—58 patients (111 reconstructions) were exposed to povidone-iodine and 53 patients (97 reconstructions) to triple-antibiotic solution. ADM integration was noted in 97% of breasts in each group. Integrated matrices appeared healthy, had no signs of foreign body reaction, and demonstrated punctate bleeding. Complications did not differ between the groups, including the rate of infections, seroma, and expander loss.Conclusion:Irrigation of the breast pocket and presoaking of the prosthesis and ADM with povidone-iodine appear to have no adverse consequences on clinical outcomes and did not impede matrix integration.