Enhanced Cell Infiltration Research Articles

3D Printing Hierarchical Porous Nanofibrous Scaffold for Bone Regeneration.

Current limitations in 3D printing pose significant challenges for the fabrication of hierarchical 3D scaffolds with nanofibrous structures that simulate the natural bone extracellular matrix (ECM) for enhanced bone regeneration. This study presents an innovative approach to 3D printing customized hierarchical porous scaffolds with nanofiber structures using biodegradable nanofibrous microspheres as the bio-ink. In vitro investigations demonstrate that the hierarchical porous architecture substantially enhances cell infiltration and proliferation rates, while the nanofiber topology provides physical cues to guide osteogenic differentiation and ECM deposition. When serving as a cell carrier, the 3D-printed nanofibrous scaffold promotes bone tissue regeneration and integration in vivo. Additionally, the facile and versatile chemical modification facilitates the precise tailoring of the scaffold's functionality. Using nanofibrous microspheres with highly biomimetic and versatile modification properties as the foundational constituent in this universal 3D printing methodology enables comprehensive manipulation of scaffolding biological properties, spanning from macroscopic external morphology to molecular-scale biochemical kinetics, thereby addressing a diverse spectrum of clinical requisites.

467 Enhancing Cell Infiltration and Controlled Growth Factor Release for a Customized 3D-Printed Bone Graft Composite

OBJECTIVES/GOALS: Annually, 1.5 million global patients receive maxillofacial reconstruction. The gold standard, involving bone particulate, lacks reproducibility. To improve this, we have developed a custom 3D-printable, porous cover-core design. This study optimizes the hydrogel core properties and growth factor (GF) release for enhanced bone regeneration. METHODS/STUDY POPULATION: Different ratios of Methacrylated Gelatin (GelMa), Methacrylated Alginate (AlgMa) and tricalcium phosphate (α²-TCP) were combined to optimize cell viability, GF sequestration and mechanical stability. Material characterization was performed using a rheometer to determine the viscoelastic properties of the blends. Release from disks loaded with FGF-containing PLGA microparticles was quantified with an ELISA kit. Furthermore, scanning electron microscopy (SEM) was conducted to quantify hydrogel porosity. In vitro studies were performed using NIH 3T3 murine fibroblasts in Corning Transwells while immunofluorescent, metabolic and osteogenic studies were performed in 96 well plates to investigate cell infiltration, cell adhesion, viability and differentiation, respectively. RESULTS/ANTICIPATED RESULTS: By adjusting the AlgGelMa ratio, we manipulated matrix properties. GelMa possesses excellent durability and cell adhesion due to intrinsic RGD-binding motifs. AlgMa enhanced swelling by 30%, growth factor sequestration by 50% in 24hrs, and matrix storage modulus without increasing the loss modulus which could cause cell migration away from the hydrogel. Varying the AlgGelMa ratio lowered pH, promoted cell infiltration, and reduced fibronectin accumulation. The addition of β-TCP is anticipated to improve cell differentiation towards an osteogenic lineage due to improved elastic modulus, calcium and phosphate ion concentration improving mineral deposition. DISCUSSION/SIGNIFICANCE: These findings suggest through the use of this composite, early cell infiltration can be increased and promoted due to FGF release, leading to increased osteointegration. Our porous cover-core design ensures efficient clot integration and early cell infiltration, enhancing osteointegration through FGF release.

Anti-Inflammatory Peptide-Conjugated Silk Fibroin/Cryogel Hybrid Dual Fiber Scaffold with Hierarchical Structure Promotes Healing of Chronic Wounds.

Chronic wounds resulting from diabetes, pressure, radiation therapy, and other factors continue to pose significant challenges in wound healing. To address this, this study introduces a novel hybrid fibroin fibrous scaffold (FFS) comprising randomly arranged fibroin fibers and vertically aligned cryogel fibers (CFs). The fibroin scaffold is efficiently degummed at room temperature and simultaneously formed a porous structure. The aligned CFs are produced via directional freeze-drying, achieved by controlling solution concentration and freezing polymerization temperature. The incorporation of aligned CFs into the expanded fibroin fiber scaffold leads to enhanced cell infiltration both in vitro and in vivo, further elevating the hybrid scaffold's tissue compatibility. The anti-inflammatory peptide 1 (AP-1) is also conjugated to the hybrid fibrous scaffold, effectively transforming the inflammatory status of chronic wounds from pro-inflammatory to pro-reparative. Consequently, the FFS-AP1+CF group demonstrates superior granulation tissue formation, angiogenesis, collagen deposition, and re-epithelialization during the proliferative phase compared to the commercial product PELNAC. Moreover, the FFS-AP1+CF group displays epidermis thickness, number of regenerated hair follicles, and collagen density closer to normal skin tissue. These findings highlight the potential of random fibroin fibers/aligned CFs hybrid fibrous scaffold as a promising approach for skin tissue filling and tissue regeneration.

Injectable Microporous Annealed Crescent-Shaped (MAC) Particle Hydrogel Scaffold for Enhanced Cell Infiltration.

Hydrogels are widely used for tissue engineering applications to support cellular growth, yet the tightly woven structure often restricts cell infiltration and expansion. Consequently, granular hydrogels with microporous architectures have emerged as a new class of biomaterial. Particularly, the development of microporous annealed particle (MAP) hydrogel scaffolds has shown improved stability and integration with host tissue. However, the predominant use of spherically shaped particles limits scaffold porosity, potentially limiting the level of cell infiltration. Here, a novel microporous annealed crescent-shaped particle (MAC) scaffold that is predicted to have improved porosity and pore interconnectivity in silico is presented. With microfluidic fabrication, tunable cavity sizes that optimize interstitial void space features are achieved. In vitro, cells incorporated into MAC scaffolds form extensive 3D multicellular networks. In vivo, the injectable MAC scaffold significantly enhances cell infiltration compared to spherical MAP scaffolds, resulting in increased numbers of myofibroblasts and leukocytes present within the gel without relying on external biomolecular chemoattractants. The results shed light on the critical role of particle shape in cell recruitment, laying the foundation for MAC scaffolds as a next-generation granular hydrogel for diverse tissue engineering applications.

Biostimulatory Micro-Fragmented Nanofiber-Hydrogel Composite Improves Mesenchymal Stem Cell Delivery and Soft Tissue Remodeling.

Functional microgels are preferred stem cell carriers due to the ease of delivery through minimally invasive injection and seamless integration with the surrounding host tissue. A biostimulatory nanofiber-hydrogel composite (NHC) has been previously developed through covalently crosslinking a hyaluronic acid hydrogel network with surface-functionalized poly (ε-caprolactone) nanofiber fragments. The NHC mimics the microarchitecture of native soft tissue matrix, showing enhanced cell infiltration, immunomodulation, and proangiogenic properties. Here, injectability of the pre-formed NHC is improved by mechanical fragmentation, making it into micro-fragmented NHC (mfNHC) in a granular gel form as a stem cell carrier to deliver mesenchymal stem cells (MSCs) for soft tissue remodeling. The mfNHC shows a similar storage modulus but a significantly reduced injection force, as compared with the corresponding bulk NHC. When injected subcutaneously in a rat model, mfNHC-MSC constructs initiate an elevated level of host macrophage infiltration, more pro-regenerative polarization, and subsequently, improved angiogenesis and adipogenesis response when compared to mfNHC alone. A similar trend of host cell infiltration and pro-angiogenic response is detected in a swine model with a larger volume injection. These results suggest a strong potential for use of the mfNHC as an injectable carrier for cell delivery and soft tissue remodeling.

A Bioengineered Organized Collagen-Based Microfibrous Implant Designed for Tendon Regeneration

Category:Basic Sciences/Biologics; SportsIntroduction/Purpose:Tendon healing is a slow and complicated process that results in inferior structural and functional properties compared to healthy tissue. It may be possible to improve outcomes of tendon healing with enhancement of biological repair through the development of tissue engineered medical products (TEMPs). Although many tendons heal with satisfactory outcomes, others do not, leading to pain and additional cost. Current augments for repair lack either native tissue structure or composition, limiting their potential effectiveness. For example, in tendon healing, restoring native organized and dense collagen composition is a primary goal; however, most TEMPs, such as lyophilized collagen or electrospun pure collagen fibers, are mechanically weak and unstable. For optimizing stability and biocompatibility, collagen can be combined with a high-performance co-biopolymer, such as poly(D,L-lactide) (PDLLA).Methods:The purpose of this study was to demonstrate the efficacy of a novel biointegrative construct, composed of type I collagen and PDLLA for Achilles tendon repair. PDLLA is biocompatible, has shown to support cell growth, and degrades to CO2 and H2O in vivo. The electrospun copolymer constructs were post-processed with low-temperature annealing to enhance their porosity and mechanical stability. The constructs were first tested for their ability to absorb blood and platelet-rich-plasma (PRP). Additionally, the constructs were tested in vivo by implantation in a rabbit Achilles tendon injury model by creating a 7 mm midsubstance Achilles defect. For each tendon defect, a copolymer construct was circumferentially wrapped around the defect and sutured into place. Finally, human cadaver testing was performed to assess implant handling and fixation characteristics.Results:Collagen-PDLLA constructs absorbed more than ten times their weight in blood and PRP. Histologically, rabbit tendons showed rapid implant cellularization by 2-8 weeks, along with de novo collagen deposition. By 16 weeks, dense collagenous connective tissue was seen integrating at the tendon-implant interface and throughout the implant. Movin-Bonar scores for the collagen-PDLLA-implanted and sham-operated tendons were identical from 16-72 weeks post-operatively, indicating a return to normal tendon morphology. Overall, surgical application of the collagen-PDLLA constructs demonstrated dense collagenous fibrous connective tissue ingrowth into and around the implant. Moreover, cadaver surgical implantation of the implant in a minimally invasive Achilles tendon repair and arthroscopic rotator cuff repair, allowed easy graft orientation and delivery as well as excellent handleability and suture fixation to the native tendon.Conclusion:Electrospun biointegrative implants were tissue-engineered to enhance cell infiltration for promoting tissue integration and functional remodeling. When implanted in the rabbit Achilles tendon, the implants demonstrated new in situ tissue generation and remodeled into dense, regularly oriented connective tissue at the tendon-implant interface. Collagenous ingrowth may be crucial in tendon protection and significantly progresses towards improving clinical outcomes following tendon injury.Moreover, the implants demonstrated additional clinical utility by absorbing greater than ten times their weight in biologic fluids which demonstrates the potential to combine this novel biointegrative implant with additional fluid based biologic therapies.

Engineering the Composition of Microfibers to Enhance the Remodeling of a Cell-Free Vascular Graft.

The remodeling of vascular grafts is critical for blood vessel regeneration. However, most scaffold materials have limited cell infiltration. In this study, we designed and fabricated a scaffold that incorporates a fast-degrading polymer polydioxanone (PDO) into the microfibrous structure by means of electrospinning technology. Blending PDO with base polymer decreases the density of electrospun microfibers yet did not compromise the mechanical and structural properties of the scaffold, and effectively enhanced cell infiltration. We then used this technique to fabricate a tubular scaffold with heparin conjugated to the surface to suppress thrombosis, and the construct was implanted into the carotid artery as a vascular graft in animal studies. This graft significantly promoted cell infiltration, and the biochemical cues such as immobilized stromal cell-derived factor-1α further enhanced cell recruitment and the long-term patency of the grafts. This work provides an approach to optimize the microfeatures of vascular grafts, and will have broad applications in scaffold design and fabrication for regenerative engineering.

Adapting the Pore Size of Individual, 3D-Printed CPC Scaffolds in Maxillofacial Surgery.

Three dimensional (3D) printing allows additive manufacturing of patient specific scaffolds with varying pore size and geometry. Such porous scaffolds, made of 3D-printable bone-like calcium phosphate cement (CPC), are suitable for bone augmentation due to their benefit for osteogenesis. Their pores allow blood-, bone- and stem cells to migrate, colonize and finally integrate into the adjacent tissue. Furthermore, the pore size affects the scaffold’s stability. Since scaffolds in maxillofacial surgery have to withstand high forces within the jaw, adequate mechanical properties are of high clinical importance. Although many studies have investigated CPC for bone augmentation, the ideal porosity for specific indications has not been defined yet. We investigated 3D printed CPC cubes with increasing pore sizes and different printing orientations regarding cell migration and mechanical properties in comparison to commercially available bone substitutes. Furthermore, by investigating clinical cases, the scaffolds’ designs were adapted to resemble the in vivo conditions as accurately as possible. Our findings suggest that the pore size of CPC scaffolds for bone augmentation in maxillofacial surgery necessarily needs to be adapted to the surgical site. Scaffolds for sites that are not exposed to high forces, such as the sinus floor, should be printed with a pore size of 750 µm to benefit from enhanced cell infiltration. In contrast, for areas exposed to high pressures, such as the lateral mandible, scaffolds should be manufactured with a pore size of 490 µm to guarantee adequate cell migration and in order to withstand the high forces during the chewing process.

Biomanufacturing organized collagen-based microfibers as a Tissue ENgineered Device (TEND) for tendon regeneration

Approximately 800, 000 surgical repairs are performed annually in the U.S. for debilitating injuries to ligaments and tendons of the foot, ankle, knee, wrist, elbow and shoulder, presenting a significant healthcare burden. To overcome current treatment shortcomings and advance the treatment of tendon and ligament injuries, we have developed a novel electrospun Tissue ENgineered Device (TEND), comprised of type I collagen and poly(D,L-lactide) (PDLLA) solubilized in a benign solvent, dimethyl sulfoxide (DMSO). TEND fiber alignment, diameter and porosity were engineered to enhance cell infiltration leading to promote tissue integration and functional remodeling while providing biomechanical stability. TEND rapidly adsorbs blood and platelet-rich-plasma (PRP), and gradually releases growth factors over two weeks. TEND further supported cellular alignment and upregulation of tenogenic genes from clinically relevant human stem cells within three days of culture. TEND implanted in a rabbit Achilles tendon injury model showed new in situ tissue generation, maturation, and remodeling of dense, regularly oriented connective tissue in vivo. In all, TEND’s organized microfibers, biological fluid and cell compatibility, strength and biocompatiblility make significant progress towards clinically translating electrospun collagen-based medical devices for improving the clinical outcomes of tendon injuries.

Encapsulation of Calcium Phosphates on Electrospun Nanofibers for Tissue Engineering Applications

In the field of tissue engineering, electrospinning is a versatile technique that provides nanofibers with structure similar to that of the extracellular matrix owing to their flexible functionalization. Considerable developments in electrospinning have been made to produce engineered electrospun nanofibers for different biomedical applications. Various biopolymers possess good biocompatibility and biodegradability and are nontoxic in nature. Modification of these biopolymers can enhance or elicit certain properties. One technique of modification is the incorporation of certain inorganic ions or components that can enhance its specific functional characteristics such as mineralization, osseointegration, and bioactivity. Incidentally, calcium phosphate (CaP) materials have proven to be suitable and versatile for biopolymer incorporation and exploration because of their inherent bioactivity and being key mineral constituents of bone and teeth. The addition of CaP materials to polymers enhances cell infiltration, differentiation, and biomineralization. We aim to provide a broad overview of CaP material (particularly hydroxyapatite (HA))-incorporated electrospun nanocomposite fibers and their possible applications in tissue engineering. Some key polymer/HA composites were discussed in detail, and a brief discussion on other polymer/HA composites was also provided. Finally, we discussed the future perspectives of this interesting and emerging composite material fabricated via electrospinning.

Biofabrication of poly(l-lactide-co-ε-caprolactone)/silk fibroin scaffold for the application as superb anti-calcification tissue engineered prosthetic valve

In this study, electrospun scaffolds were fabricated by blending poly(l-lactide-co-ε-caprolactone) (PLCL) and silk fibroin (SF) with different ratios, and further the feasibility of electrospun PLCL/SF scaffolds were evaluated for application of tissue engineered heart valve (TEHV). Scanning electron microscopy (SEM) results showed that the surface of PLCL/SF electrospun scaffolds was smooth and uniform while the mechanical properties were appropriate as valve prosthesis. In vitro cytocompatibility evaluation results demonstrated that all of the PLCL/SF electrospun scaffolds were cytocompatible and valvular interstitial cells (VICs) cultured on PLCL/SF scaffolds of 80/20 & 70/30 ratios exhibited the best cytocompatibility. The in vitro osteogenic differentiation of VICs including alkaline phosphatase (ALP) activity and quantitative polymerase chain reaction (qPCR) assays indicated that PLCL/SF scaffolds of 80/20 & 90/10 ratios behaved better anti-calcification ability. In the in vivo calcification evaluation model of rat subdermal implantation, PLCL/SF scaffolds of 80/20 & 90/10 ratios presented better anti-calcification ability, which was consistent with the in vitro results. Moreover, PLCL/SF scaffolds of 80/20 & 70/30 ratios showed significantly enhanced cell infiltration and M2 macrophage with higher CD206+/CD68+ ratio. Collectively, our data demonstrated that electrospun scaffolds with the PLCL/SF ratio of 80/20 hold great potential as TEHV materials.

Engineering vascularized dermal grafts by integrating a biomimetic scaffold and Wharton's jelly MSC-derived endothelial cells.

Tissue engineering aims to generate functional tissue constructs with the necessary scaffold properties for cell colonization and the establishment of a vascular network. However, treatment of tissue defects using synthetic scaffolds remains a challenge mainly due to insufficient and slow vascularization. Our previous study developed a macroporous silk fibroin scaffold with a nanofibrous microstructure, and demonstrated that the nanofibrous structure can promote the viability of endothelial cells (ECs) and guide cell migration. Further studies are needed to clarify the effect of scaffold microstructures on cell-mediated vascularization. Here, we investigated the efficacy of EC-seeded nanofibrous scaffolds in improving vascularization in vivo. ECs derived from induced human Wharton's Jelly mesenchymal stem cells served as a potential source for cell transplantation. The cell-seeded scaffolds were implanted into dermal defects of SD rats, demonstrating that the multiscale hierarchical design significantly improved the capacity of transplanted cells to promote and accelerate neovascularization and dermal reconstruction via enhancing cell infiltration, collagen deposition and growth factor expression. Our findings provide new insight into the development of degradable macroporous composite materials with 3D microstructures as tissue engineering scaffolds with enhanced vascularization functions, and also provide new treatment options for cell transplantation.

Multichannel Bioactive Silk Nanofiber Conduits Direct and Enhance Axonal Regeneration after Spinal Cord Injury.

After a spinal cord injury, axonal regeneration over long distances is challenging due to the lack of physical guidance cues and bioactive signals. In this study, a multichannel bioactive silk fibroin nanofiber conduit was fabricated to improve spinal cord injury repair by enhancing axonal regeneration. The conduit was composed of longitudinally oriented silk fibroin nanofibers and then functionalized with laminin. In vitro, the bioactive conduits could promote neuron-like development and directional neurite extension of PC12 cells by providing a bioactive stimulus and physical guidance. In a spinal cord injury model in Sprague-Dawley rats, the biofunctionalized conduits displayed superior integration with the host tissue due to enhanced cell infiltration and tissue ingrowth. The glial scar was significantly reduced, allowing axonal ingrowth along with the channel direction. Compared to a single-channel conduit, the multichannel conduit improved spinal cord regeneration by boosting tissue ingrowth and axonal regeneration, indicating that the conduit architectures play critical roles in spinal cord regeneration. These silk fibroin conduits, along with the multichannel architecture, nanoscale cues, and the ability to bind bioactive compounds, represent promising candidates for spinal cord regeneration.

Mimicking the Organic and Inorganic Composition of Anabolic Bone Enhances Human Mesenchymal Stem Cell Osteoinduction and Scaffold Mechanical Properties.

Engineered bone graft designs have been largely inspired by adult bone despite functionally significant differences from the composition of anabolic bone in both the mineralized and non-mineralized fractions. Specifically, anabolic bone contains hydroxyapatite with ionic substitutions that facilitate bone turnover and relatively rare collagens type VI and XII that are important for normal bone development. In this work, human mesenchymal stem cells (hMSCs) were cultured in lyophilized collagen type I scaffolds mineralized with hydroxyapatite containing Mg2+ substitutions, then induced to deposit an extracellular matrix (ECM) containing collagens VI and XII by exposure to GW9662, a PPARγ inhibitor. Delivery of GW9662 was accomplished through either Supplemented Media or via composite microspheres embedded in the scaffolds for localized delivery. Furthermore, hMSCs and scaffolds were cultured in both static and perfuse conditions to investigate the interaction between GW9662 treatment and perfusion and their effects on ECM deposition trends. Perfusion culture enhanced cell infiltration into the scaffold, deposition of collagen VI and XII, as well as osteogenic differentiation, as determined by gene expression of osteopontin, BMP2, and ALP. Furthermore, scaffold mineral density and compressive modulus were increased in response to both GW9662 treatment and perfusion after 3 weeks of culture. Local delivery of GW9662 with drug-eluting microspheres had comparable effects to systemic delivery in the perfusate. Together, these results demonstrate a strategy to create a scaffold mimicking both organic and inorganic characteristics of anabolic bone and its potential as a bone graft.

Evaluation of scaffold microstructure and comparison of cell seeding methods using micro-computed tomography-based tools.

Micro-computed tomography (micro-CT) provides a means to analyse and model three-dimensional (3D) tissue engineering scaffolds. This study proposes a set of micro-CT-based tools firstly for evaluating the microstructure of scaffolds and secondly for comparing different cell seeding methods. The pore size, porosity and pore interconnectivity of supercritical CO2 processed poly(l-lactide-co-ɛ-caprolactone) (PLCL) and PLCL/β-tricalcium phosphate scaffolds were analysed using computational micro-CT models. The models were supplemented with an experimental method, where iron-labelled microspheres were seeded into the scaffolds and micro-CT imaged to assess their infiltration into the scaffolds. After examining the scaffold architecture, human adipose-derived stem cells (hASCs) were seeded into the scaffolds using five different cell seeding methods. Cell viability, number and 3D distribution were evaluated. The distribution of the cells was analysed using micro-CT by labelling the hASCs with ultrasmall paramagnetic iron oxide nanoparticles. Among the tested seeding methods, a forced fluid flow-based technique resulted in an enhanced cell infiltration throughout the scaffolds compared with static seeding. The current study provides an excellent set of tools for the development of scaffolds and for the design of 3D cell culture experiments.

A Thixotropic, Cell-Infiltrative Nanocellulose Hydrogel That Promotes in Vivo Tissue Remodeling.

Injectable gels have been used in minimally invasive surgery for tissue regeneration and treatment of inflammatory diseases. However, polymeric hydrogels often fail in cell infiltration, because of the presence of dense, cross-linked molecular networks and a lack of bioactivity, which causes delayed tissue remodeling. Here, we report a thixotropic, cell-infiltrative hydrogel of biofunctionalized nanocellulose that topologically enhances cell infiltration and biochemically upregulates cellular activity for the promotion of tissue remodeling. Biodegradable, sulfonated nanocellulose forms a nanofibrous hydrogel, mimicking cellular microenvironments through cross-linking between nanocellulose and gelatin. Resulting nanocellulose hydrogels showed thixotropy, allowing for single syringe injection. Nanofiber-based hydrogels possess high molecular permeability, which is due to nanoporous structures. Sulfonate groups on nanocellulose increase protein adsorption and induce cellular extension in vitro. Highly sulfonated nanocellulose hydrogels enhanced cell infiltration and vascularization upon implantation into rats. Macrophage polarization to M2 was observed in nanocellulose hydrogels, which may be involved in tissue remodeling. Injectable, biofunctionalized nanocellulose gels have enormous potential as artificial biomatrices to heal inflammatory diseases through manipulation of the immune system and promotion of tissue remodeling.

In vivo engineered extracellular matrix scaffolds with instructive niches for oriented tissue regeneration

Implanted scaffolds with inductive niches can facilitate the recruitment and differentiation of host cells, thereby enhancing endogenous tissue regeneration. Extracellular matrix (ECM) scaffolds derived from cultured cells or natural tissues exhibit superior biocompatibility and trigger favourable immune responses. However, the lack of hierarchical porous structure fails to provide cells with guidance cues for directional migration and spatial organization, and consequently limit the morpho-functional integration for oriented tissues. Here, we engineer ECM scaffolds with parallel microchannels (ECM-C) by subcutaneous implantation of sacrificial templates, followed by template removal and decellularization. The advantages of such ECM-C scaffolds are evidenced by close regulation of in vitro cell activities, and enhanced cell infiltration and vascularization upon in vivo implantation. We demonstrate the versatility and flexibility of these scaffolds by regenerating vascularized and innervated neo-muscle, vascularized neo-nerve and pulsatile neo-artery with functional integration. This strategy has potential to yield inducible biomaterials with applications across tissue engineering and regenerative medicine.

Dual Porosity Protein-based Scaffolds with Enhanced Cell Infiltration and Proliferation

3D dual porosity protein-based scaffolds have been developed using the combination of foaming and freeze-drying. The suggested approach leads to the production of large, highly porous scaffolds with negligible shrinkage and deformation compared to the conventional freeze-drying method. Scanning electron microscopy, standard histological processing and mercury intrusion porosimetry confirmed the formation of a dual network in the form of big primary pores (243 ± 14 µm) embracing smaller secondary pores (42 ± 3 µm) opened onto their surface, resembling a vascular network. High interconnectivity of the pores, confirmed by micro-CT, is shown to improve diffusion kinetics and support a relatively uniform distribution of isolated human dental pulp stem cells within the scaffold compared to conventional scaffolds. Dual network scaffolds indicate more than three times as high cell proliferation capability as conventional scaffolds in 14 days.

Low expression of microRNA-21 inhibits trophoblast cell infiltration through targeting PTEN.

We investigate whether microRNA-21 could increase the infiltration ability of trophoblast cells via regulating PTEN expression, thus participating in the occurrence and development of preeclampsia. MicroRNA-21 expression in the placenta tissues of preeclampsia women and normal pregnant women was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The effects of microRNA-21 on cell proliferation and infiltration were examined by cell counting kit-8 (CCK-8) and transwell assay, respectively. The dual-luciferase reporter gene assay was used to determine the binding relationship between microRNA-21 and PTEN. Western blot was performed to detect PTEN and microRNA-21 in trophoblasts. QRT-PCR results showed that the microRNA-21 expression was significantly lower in the placenta of the preeclampsia women than those of normal pregnant women. Overexpression of microRNA-21 in HTR-8/SVneo cells had no effect on cell proliferation, but enhanced cell infiltration ability. Inhibition of microRNA-21 in trophoblasts showed the opposite effects. The results of luciferase activity assay and Western blot showed that microRNA-21 could target PTEN and downregulate its expression. Overexpression of PTEN in HTR-8/SVneo cells partially reversed the enhanced invasive ability induced by microRNA-21 overexpression. Low expression of microRNA-21 attenuated cell infiltration of trophoblasts via direct regulating PTEN expression.

A facile method for fabricating nano/microfibrous three‐dimensional scaffold with hierarchically porous to enhance cell infiltration

ABSTRACTThe integration of electrospinning and traditional needle punching technologies were demonstrated for fabricating a new three‐dimensional (3D) scaffold to enhance the cellular infiltration. This 3D scaffold combined the advantages of electrospun polycaprolactone (PCL) nanofibrous mats and carded chitosan microfibrous webs to construct hierarchically porous, which have three regions: the large pores (about 500 μm) on the nanofibrous mats generated by needle punching, could induce the massive cells infiltration into the inner scaffold; the small pores (30–200 μm) of the microfibrous webs allowed space for continuous cellular infiltration; and the inherent smaller pores of the nanofibrous mats (1–17 μm) which play a key role in providing a platforms for cell adhesion and proliferation. In addition, the 3D scaffold exhibited appropriate mechanical properties and significantly high initial cell attachment, proliferation, and infiltration, suggesting that its great potential in tissue engineering. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019, 136, 47046