Clay Sculpture-Inspired 3D Printed Microcage Module Using Bioadhesion Assembly for Specific-Shaped Tissue Vascularization and Regeneration.

3D bioprinting techniques have enabled the fabrication of irregular large-sized tissue engineering scaffolds. However, complicated customized designs increase the medical burden. Meanwhile, the integrated printing process hinders the cellular uniform distribution and local angiogenesis. A novel approach is introduced to the construction of sizable tissue engineering grafts by employing hydrogel 3D printing for modular bioadhesion assembly, and a poly (ethylene glycol) diacrylate (PEGDA)-gelatin-dopamine (PGD) hydrogel, photosensitive and adhesive, enabling fine microcage module fabrication via DLP 3D printing is developed. The PGD hydrogel printed micocages are flexible, allowing various shapes and cell/tissue fillings for repairing diverse irregular tissue defects. In vivo experiments demonstrate robust vascularization and superior graft survival in nude mice. This assembly strategy based on scalable 3D printed hydrogel microcage module could simplify the construction of tissue with large volume and complex components, offering promise for diverse large tissue defect repairs.

Low glutaminase and glycolysis correlate with a high transdifferentiation efficiency in mouse cortex.

Both exogenous transcriptional factors and chemical-defined medium can transdifferentiate astrocytes into functional neurons. However, the regional preference for such transdifferentiation has not been fully studied. A previously reported 5C medium was infused into the mouse cortex and striatum to determine the regional preference for transdifferentiation from astrocytes to neurons. The numbers of NeuN+ GFAP+ EdU+ cells (intermediates) and NeuN+ EdU+ cells (end products) were determined by immunofluorescence to explore the regional preference of transdifferentiation. In addition, to optimize the delivery of the transdifferentiation medium, three key growth factors, insulin, bFGF and transferrin, were loaded onto chitosan nanoparticles, mixed with gelatin methacryloyl and tested in animals with motor cortex injury. A higher transdifferentiation efficiency was identified in the mouse cortex. Differences in cellular respiration and the balance between glutaminase (Gls) and glutamine synthetase were confirmed to be key regulators. In addition, the sustained drug release system induced transdifferentiation of cortex astrocytes both in vivo and in vitro, and partially facilitated the behaviour recovery of mice with motor cortex injury. We also applied this method in pigs and obtained consistent results. In summary, low Gls and glycolysis can be used to predict high transdifferentiation efficiency, which may be useful to identify better indications for the current transdifferentiation system. In addition, the current drug delivery system has the potential to treat diseases related to cortex injuries.

Amphiphilic and fatigue-resistant organohydrogels for small-diameter vascular grafts.

Hydrogels are used in vascular tissue engineering because of their good biocompatibility. However, most natural hydrogels exhibit high swelling ratio, poor mechanical stability, and low durability, which are key limitations for wider applications. Amphiphilic and fatigue-resistant organohydrogels were fabricated here via the click chemical reaction of unsaturated functional microbial polyhydroxyalkanoates and polyethylene glycol diacrylate and a combination of two-dimensional material graphdiyne. These organohydrogels were maintained stable in body fluids over time, and their tensile moduli remained unchanged after more than 2000 cycles of cyclic stretching. The tubular scaffolds presented good biocompatibility and perfusion in vitro. After transplantation in vivo, the vascular grafts exhibited obvious cell infiltration and tissue regeneration, having a higher patency rate than the control group in 3 months. This fabrication method provides a strategy of improving and promoting the application of organohydrogels as implant materials for small-diameter vascular graft.

Graphene Oxide Functionalized Double-Layered Patch with Anti-Adhesion Ability for Abdominal Wall Defects.

BACKGROUND: Effective repair of full-thickness abdominal wall defects requires a patch with sufficient mechanical strength and anti-adhesion characteristics to avoid the formation of hernias and intra-abdominal complications such as intestinal obstruction and fistula. However, patches made from polymers or bio-derived materials may not meet these requirements and lack the bionic characteristics of the abdominal wall. MATERIALS AND METHODS: In this study, we report a consecutive electrospun method for preparing a double-layer structured nanofiber membrane (GO-PCL/CS-PCL) using polycaprolactone (PCL), graphene oxide (GO) and chitosan (CS). To expand the bio-functions (angiogenesis/reducing reactive oxygen species) of the patch (GO-PCL/NAC-CS-PCL), N-acetylcysteine (NAC) was loaded for the repair of full-thickness abdominal wall defects (2×1.5cm) in rat model. RESULTS: The double-layered patch (GO-PCL/NAC-CS-PCL) showed excellent mechanical strength and biocompatibility. After 2 months, rats treated with the patch exhibited the desired repair effect with no hernia formation, less adhesion (adhesion score: 1.50±0.50, P<0.001) and more collagen deposition (percentage of collagen deposition: 34.94%±3.31%, P<0.001). CONCLUSION: The double-layered nanomembranes presented in this study have good anti-hernia and anti-adhesion effects, as well as improve the microenvironment in vivo. It, therefore, holds good prospects for the repair of abdominal wall defects and provides a promising key as a postoperative anti-adhesion agent.

Simple application of adipose-derived stem cell-derived extracellular vesicles coating enhances cytocompatibility and osteoinductivity of titanium implant.

Surface modification using bioactive molecules is frequently performed to improve the biological properties of medical metal biomaterial titanium (Ti) implants. Developmental evidence suggests that mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) served as potent bioactive component. As a subset of MSC-EV, adipose-derived stem cell-derived extracellular vesicles (ADSC-EVs) could be obtained from abundant adipose tissue. Meanwhile, it possesses multiple regenerative properties and might be used to endow biological activities to medical Ti implant. Here, we present a simple ADSC-EV coating strategy based on physisorption of fibronectin. This ADSC-EV functionalized Ti implants (EV-Ti) revealed enhanced osteoblast compatibility and osteoinductive activity. Cell spreading area of EV-Ti group was 1.62- and 1.48-fold larger than that of Ti group after 6 and 12 h of cell seeding, respectively. Moreover, EV-Ti promoted alkaline phosphatase, collagen 1 and osteocalcin gene expression in osteoblast by 1.51-, 1.68- and 1.82-fold compared with pristine Ti, respectively. Thus, the MSC-EVs modification method reported here provide a clinically translatable strategy to promote the bioactivity of Ti implants.

Dietary Intake of N-3 and N-6 Polyunsaturated Fatty Acids and Risk of Cancer: Meta-Analysis of Data from 32 Studies.

BACKGROUND: Large epidemiological studies have yielded conflicting results regarding the relationship between polyunsaturated fatty acids (PUFAs) and cancers. Here, we performed a meta-analysis to examine the link between dietary intake of n-3 and n-6 PUFAs and cancer risk. MATERIALS AND METHODS: We performed a search on PubMed, EMBASE, and the Cochrane Library. Studies that reported adjusted relative risk (RR) estimates with 95% confidence intervals (CI) for the associations of interest were included. RESULTS: Thirty-two studies involving 1,445,732 participants were included. Colorectal, breast and prostate cancer had been analyzed in our study. Specifically, for colorectal cancer, total n-3 PUFAs, marine n-3 PUFAs, α-linolenic acids (ALA) and n-6 PUFAs were not associated with the risk of it (RR 1.04, 95%CI 0.85-1.28; RR 0.99, 95%CI 0.89-1.09; RR 1.05, 95%CI 0.93-1.19; RR 1.02, 95%CI 0.94-1.11, respectively). For breast cancer, only marine n-3 PUFAs, but not total n-3 PUFAs, ALA, and n-6 PUFAs, was associated with a lower risk of it (RR 0.70, 95%CI 0.55-0.91). For prostate cancer, ALA and n-6 PUFAs also have no association with the risk of it. CONCLUSIONS: Most subtypes of PUFAs are probably not related to cancers. However, additional high-quality trials are warranted to corroborate the findings of this meta-analysis.

Multi-Layered Polyamide/Collagen Scaffolds with Topical Sustained Release of N-Acetylcysteine for Promoting Wound Healing.

BACKGROUND: Impaired wound healing might be associated with many issues, especially overactive of reactive oxygen species (ROS), deficiency of blood vessels and immature of epidermis. N-acetylcysteine (NAC), as an antioxidant, could solve these problems by inhibiting overreactive of ROS, promoting revascularization and accelerating re-epithelialization. How to deliver NAC in situ with a controllable releasing speed still remain a challenge. MATERIALS AND METHODS: In this study, we combined collagen (Col) with N-acetylcysteine to perform the characteristics of sustained release and chemically crosslinked Col/NAC composite with polyamide (PA) nanofibers to enhance the mechanical property of collagen and fabricated this multi-layered scaffold (PA-Col/NAC scaffold). The physical properties of the scaffolds such as surface characteristics, water absorption and tensile modulus were tested. Meanwhile, the ability to promote wound healing in vitro and in vivo were investigated. RESULTS: These scaffolds were porous and performed great water absorption. The PA-Col/NAC scaffold could sustainably release NAC for at least 14 days. After cell implantation, PA-Col/NAC scaffold showed better cell proliferation and cell migration than the other groups. In vivo, PA-Col/NAC scaffolds could promote wound healing best among all the groups. CONCLUSION: The multi-layered scaffolds could obviously accelerate the process of wound healing and exert better and prolonged effects.

A biocompatible vascularized graphene oxide (GO)-collagen chamber with osteoinductive and anti-fibrosis effects promotes bone regeneration in vivo.

The survival of transplanted cells and tissues in bone regeneration requires a microenvironment with a vibrant vascular network. A tissue engineering chamber can provide this in vivo. However, the commonly used silicone chamber is biologically inert and can cause rejection reactions and fibrous capsule. Studies have revealed that collagen is highly biocompatible and graphene oxide (GO) could regulate osteogenic activity in vivo. Besides, GO can be cross-linked with natural biodegradable polymers to construct scaffolds. Methods: A vascularized GO-collagen chamber model was built by placing vessels traversing through the embedded tissue-engineered grafts (osteogenic-induced bone mesenchymal stem cells -gelatin) in the rat groin area. Osteogenic activity and inflammatory reactions were assessed using different methods including micro-CT scanning, Alizarin red staining, and immunohistochemical staining. Results: After one month, in vivo results showed that bone mineralization and inflammatory responses were significantly pronounced in the silicone model or no chamber (control) groups. Vascular perfusion analysis confirmed that the GO-collagen chamber improved the angiogenic processes. Cells labeled with EdU revealed that the GO-collagen chamber promoted the survival and osteogenic differentiation of bone mesenchymal stem cells. Conclusion: Overall, the novel biocompatible GO-collagen chamber exhibited osteoinductive and anti-fibrosis effects which improved bone regeneration in vivo. It can, therefore, be applied to other fields of regenerative medicine.

Efficacy of Acellular Dermal Matrix in Capsular Contracture of Implant-Based Breast Reconstruction: A Single-Arm Meta-analysis.

BACKGROUND: A large number of clinical studies have suggested that acellular dermal matrix (ADM) can decrease the incidence of capsular contracture in implant-based breast reconstruction. Yet, there is currently no high-level epidemiological evidence to prove this. The goal of this meta-analysis was to clarify the efficacy of ADM in capsular contracture, and provide a reference value for plastic surgeons. METHODS: We systematically performed a search on PubMed, EMBASE, and the Cochrane Library to identify eligible studies from inception up to October 1, 2019. A random-effects model was used to obtain a pooled incidence rate. We conducted subgroup analysis according to geographic region, type of ADM, body mass index (BMI), duration of follow-up, and proportion of participants who have received radiotherapy. RESULTS: A total of 18 studies involving 2941 cases were included. Overall, the pooled incidence rate of capsular contracture was 2.4% (95% CI 1.2-3.9%). The results from subgroup analyses indicated an even lower incidence in North America (1.6%, 95% CI 0.5-3.3%) and in human-derived ADM (HADM) (1.2%, 95% CI 0.2-3.0%). In addition, the results showed that the patients with BMI < 24, or who have received radiotherapy, were more prone to capsular contracture. CONCLUSION: The application of ADM can effectively reduce the incidence of capsular contracture in implant-based breast reconstruction. And we infer that it might also apply to breast augmentation. However, additional high-quality trials are warranted to corroborate the findings of this meta-analysis. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

Tuning the mechanics of 3D-printed scaffolds by crystal lattice-like structural design for breast tissue engineering.

Breast tissue engineering is a promising alternative to standard treatments for breast defects. Although there is a consensus that the mechanical property of the scaffold should best match the reconstructed tissue, the simulation of the soft and elastic tactility of native breast tissues using conventional materials and architecture design requires further study. Previous research has shown that the crystal microstructure-like design can drastically alter the mechanical properties of the constructed scaffolds. In this study, we designed and additive manufactured four kinds of breast scaffolds using polyurethane and termed their architectures as N5S4, N9S8, N7S6 and N4S6. The basic unit cell of each scaffold was similar to a lattice structure from the isometric crystal system. The scaffolds possessed identical porosity but different mechanical properties in which the compressive modulus of the softest scaffolds (N5S4) were similar to that of native breast tissue. When applied in the construction of tissue-engineered breast combining with delayed fat injection technique in nude rat models, the soft scaffolds(N5S4) performed better compared to its stiff counterpart (N4S6), as higher adipose survival, vascularization and milder fibrosis could be observed in N5S4 scaffolds . Lastly, using finite element analysis, we further investigated the influence of the unit cell architectures on the mechanical properties of the scaffolds and simulated the deformation as well as stress distribution patterns of the implanted scaffolds in detail. Thus, a crystal lattice-like architecture design was introduced to tune the mechanical properties of the scaffolds and match the requirements for tissue engineering applications.

N-acetyl cysteine-loaded graphene oxide-collagen hybrid membrane for scarless wound healing.

Wound dressings composed of natural polymers, such as type I collagen, possess good biocompatibility, water holding capacity, air permeability, and degradability, and can be used in wound repair. However, due to the persistent oxidative stress in the wound area, the migration and proliferation of fibroblasts might be suppressed, leading to poor healing. Thus, collagen-containing scaffolds are not suitable for accelerated wound healing. Antioxidant N-acetyl cysteine (NAC) is known to reduce the reactive oxygen species (ROS) and has been widely used in the clinic. Theoretically, the carboxyl group of NAC allows loading of graphene oxide (GO) for sustained release and may also enhance the mechanical properties of the collagen scaffold, making it a better wound-dressing material. Herein, we demonstrated an innovative approach for a potential skin-regenerating hybrid membrane using GO incorporated with collagen I and NAC (N-Col-GO) capable of continuously releasing antioxidant NAC. Methods: The mechanical stability, water holding capacity, and biocompatibility of the N-Col-GO hybrid membrane were measured in vitro. A 20 mm rat full-skin defect model was created to evaluate the repair efficiency of the N-Col-GO hybrid membrane. The vascularization and scar-related genes in the wound area were also examined. Results: Compared to the Col only scaffold, N-Col-GO hybrid membrane exhibited a better mechanical property, stronger water retention capacity, and slower NAC release ability, which likely promote fibroblast migration and proliferation. Treatment with the N-Col-GO hybrid membrane in the rat wound model showed complete healing 14 days after application which was 22% faster than the control group. HE and Masson staining confirmed faster collagen deposition and better epithelization, while CD31 staining revealed a noticeable increase of vascularization. Furthermore, Rt-PCR demonstrated decreased mRNA expression of profibrotic and overexpression of anti-fibrotic factors indicative of the anti-scar effect. Conclusion: These findings suggest that N-Col-GO drug release hybrid membrane serves as a better platform for scarless skin regeneration.

Biomimetic open porous structured core-shell microtissue with enhanced mechanical properties for bottom-up bone tissue engineering.

Background: Microtissues constructed with hydrogels promote cell expansion and specific differentiation by mimicking the microarchitecture of native tissues. However, the suboptimal mechanical property and osteogenic activity of microtissues fabricated by natural polymers need further improvement for bone reconstruction application. Core-shell designed structures are composed of an inner core part and an outer part shell, combining the characteristics of different materials, which improve the mechanical property of microtissues. Methods: A micro-stencil array chip was used to fabricate an open porous core-shell micro-scaffold consisting of gelatin as shell and demineralized bone matrix particles modified with bone morphogenetic protein-2 (BMP-2) as core. Single gelatin micro-scaffold was fabricated as a control. Rat bone marrow mesenchymal stem cells (BMSCs) were seeded on the micro-scaffolds, after which they were dynamic cultured and osteo-induced in mini-capsule bioreactors to fabricate microtissues. The physical characteristics, biocompatibility, osteo-inducing and controlled release ability of the core-shell microtissue were evaluated in vitro respectively. Then microtissues were tested in vivo via ectopic implantation and orthotopic bone implantation in rat model. Results: The Young's modulus of core-shell micro-scaffold was nearly triple that of gelatin micro-scaffold, which means the core-shell micro-scaffolds have better mechanical property. BMSCs rapidly proliferated and retained the highest viability on core-shell microtissues. The improved osteogenic potential of core-shell microtissues was evidenced by the increased calcification based on von kossa staining and osteo-relative gene expression. At 3months after transplantation, core-shell microtissue group formed the highest number of mineralized tissues in rat ectopic subcutaneous model, and displayed the largest amount of new bony tissue deposition in rat orthotopic cranial defect. Conclusion: The novel core-shell microtissue construction strategy developed may become a promising cell delivery platform for bone regeneration.

The pro-angiogenic role of hypoxia inducible factor stabilizer FG-4592 and its application in an in vivo tissue engineering chamber model.

Tissue engineering is a promising technology used as an alternative to organ/tissue transplantation which is often limited by donor shortage. The construction of large-sized engineered tissue requires a fast and sufficient vascularization process. Previous studies have shown that hypoxia-inducible factor (HIF) -1α may promote the vascularization process implying that stabilized HIF-1α can be applied in the engineering of large-sized tissue. However, the toxicity and off-target effect of previously reported HIF-1α stabilizers limit their clinical application. FG-4592, a small molecule specific HIF stabilizer, was previously investigated as an anti-anemia drug in a phase-III clinical trial. Here we found that FG-4592 promoted tube formation in an in vitro model of angiogenesis by stabilizing HIF-1α and activating vascular endothelial growth factor (VEGF). When FG-4592 immobilized fibrin gel scaffold was implanted into a subcutaneous tissue engineering chamber, the vascularization process was significantly enhanced through the similar mechanisms which was verified in vitro. We conclude that FG-4592 may serve as a pro-angiogenic molecule for the construction of large-sized engineered tissue where intensive angiogenesis is required.

Sustained release of N-acetylcysteine by sandwich structured polycaprolactone/collagen scaffolds for wound healing.

PCL (poly-caprolactone) nanofibers have good biocompatibility and high porosity, which are usually utilized for application in wound dressings. However, wound healing could be hindered by the overproduction of reactive oxygen species (ROS) and different factors. Pure nanofibers cannot satisfy these requirements of wound healing. N-acetylcysteine (NAC), as an antioxidant, meets the requirements for wound healing by resisting the overproduction of ROS and by promoting angiogenesis and maturation of the epidermis. In this study, we prepared a sandwich structured PCL-Col/NAC scaffold using the molding method, which consisted of PCL nanofibers at the core and NAC-loaded collagen on both sides. The hydroscopicity and tensile modulus of PCL-Col/NAC scaffolds showed best performance of these properties among groups. Meanwhile, the drug release profiles of PCL-Col/NAC scaffolds were investigated using the HPLC method and the results suggested a sustained drug release of NAC for PCL-Col/NAC scaffolds. In addition, PCL-Col/NAC scaffolds presented better properties than the control groups in cell migration and proliferation. The in vivo wound healing therapy effect was studied using an oval (2 × 1 cm) full-thickness skin defect wound model for SD rats. After 21 days, gross view and histological analysis showed a favorable beneficial therapeutic effect as well as better epidermal maturation compared with the control groups. CD31 immunohistology results revealed relatively more new vessels in the PCL-Col/NAC group than the control groups. This study developed novel PCL-Col/NAC scaffolds with an excellent hydroscopicity, tensile modulus and the ability to promote epidermal maturation and angiogenesis, demonstrating its promising potential in wound healing treatment. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2019.

An in vivo comparative study of the gelatin microtissue-based bottom-up strategy and top-down strategy in bone tissue engineering application.

Tissue-engineered bone grafts (TEBGs) represent a promising treatment for bone defects. Nevertheless, drawbacks of the current construction strategy (top-down [TD] strategy) such as limited transmission of nutrients and nonuniform distribution of seeded cells, result in an unsatisfied therapeutic effect on large segmental bone defects. Theoretically, tissue-engineered microtissue (TEMT)-based bottom-up (BU) strategy is effective in preserving seed cells and vascularization, thus being regarded as a better alternative for TEBGs. Yet, there are few studies focusing on the comparison of the in vivo performance of TEBGs fabricated by TD or BU strategy. Here, we developed an ectopic bone formation rat model to compare the performance of these two construction strategies in vivo. TEBGs made from gelatin TEMT (BU strategy) and bulk tissue (BT; TD strategy) were seeded with equal number of rat bone marrow-derived mesenchymal stem cells and fabricated in 5 mm polydimethylsiloxane chambers. The grafts were implanted into subcutaneous pockets in the same rat. Four weeks after implantation, microcomputed tomography and hematoxylin and eosin staining results demonstrated that more bony tissue was formed in the microtissue (MT) group than in the BT group. CD31 staining further confirmed that there were more blood vessels in the MT group, indicating that the BU strategy was superior in inducing angiogenesis. This comparative study provides evidence that the BU construction strategy is more effective for in vivo application and bone defect treatment by bone tissue engineering. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 678-688, 2019.