GPU-accelerated OCT imaging: Real-time data processing and artifact suppression for enhanced monitoring of 3D bioprinted tissues and vascular-like networks

Optical coherence tomography(OCT)imaging technology has significant advantages in in situ and noninvasive monitoring of biological tissues.However,it still faces the following challenges:including data processing speed,image quality,and improvements in three-dimensional(3D)visualization effects.OCT technology,especially functional imaging techniques like optical coherence tomography angiography(OCTA),requires a long acquisition time and a large data size.Despite the substantial increase in the acquisition speed of swept source optical coherence tomography(SS-OCT),it still poses significant challenges for data processing.Additionally,during in situ acquisition,image artifacts resulting from interface reflections or strong reflections from biological tissues and culturing containers present obstacles to data visualization and further analysis.Firstly,a customized frequency domainfilter with anti-banding suppression parameters was designed to suppress artifact noises.Then,this study proposed a graphics processing unit(GPU)-based real-time data processing pipeline for SS-OCT,achieving a measured line-process rate of 800 kHz for 3D fast and high-quality data visualization.Furthermore,a GPU-based realtime data processing for CC-OCTA was integrated to acquire dynamic information.Moreover,a vascular-like network chip was prepared using extrusion-based 3D printing and sacrificial materials,with sacrificial material being printed at the desired vascular network locations and then removed to form the vascular-like network.OCTA imaging technology was used to monitor the progression of sacrificial material removal and vascular-like network formation.Therefore,GPU-based OCT enables real-time processing and visualization with artifact suppression,making it particularly suitable for in situ noninvasive longitudinal monitoring of 3D bioprinting tissue and vascular-like networks in microfluidic chips.

3D-bioprinted tri-layered cellulose/collagen-based drug-eluting fillers for the treatment of deep tunneling wounds

Tunneling wounds create passageways underneath the skin surface with varying sizes and shapes and can have twists and turns,making their treatment extremely difficult.Available wound care solutions only cater to superficial wounds,and untreated tunneling wounds pose major health concerns.This study aims to fulfill this challenge by fabricating tunnel wound fillers(TWFs)made of natural polymers that mimic the dermal extracellular matrix.In this study,cellulose microfibers(CMFs)derived from banana stem and fish skin-derived collagen were used to formulate bio-inks with varying CMF contents(25,50,and 75 mg).Tri-layered(CMFs,primary and secondary collagen coatings),drug-eluting(Baneocin),and cell-laden(human mesenchymal stem cells)TWFs were three-dimensional(3D)-printed and extensively characterized.CMFs showed the most suitable rheological properties for 3D printing at 50 mg concentration.The Alamar Blue data showed significantly increased cell proliferation from Day 1 to Day 7,and scratch tests used to evaluate in vitro wound healing revealed that the best coverage of the wound area was achieved using CMFs in combination with collagen and alginate.Finally,the TWF showed promising capability and tunability in terms of wound shape and size upon testing on a chicken tissue model.The results demonstrate the tremendous potential of TWFs in treating deep tunneling wounds with unique advantages,such as patient-specific customization,good wound exudate absorption capability while releasing wound healing drugs,and the inclusion of stem cells for accelerated healing and tissue regeneration.

Ca_v3.3-mediated endochondral ossification in a three-dimensional bioprinted Gel MA hydrogel

The growth plate(GP)is a crucial tissue involved in skeleton development via endochondral ossification(EO).The bone organoid is a potential research model capable of simulating the physiological function,spatial structure,and intercellular communication of native GPs.However,mimicking the EO process remains a key challenge for bone organoid research.To simulate this orderly mineralization process,we designed an in vitro sh Ca_(v)3.3 ATDC5-loaded gelatin methacryloyl(Gel MA)hydrogel model and evaluated its bioprintability for future organoid construction.In this paper,we report the first demonstration that the T-type voltage-dependent calcium channel(T-VDCC)subtype Ca_(v)3.3 is dominantly expressed in chondrocytes and is negatively correlated with the hypertrophic differentiation of chondrocytes during the EO process.Furthermore,Ca_(v)3.3 knockdown chondrocytes loaded with the Gel MA hydrogel successfully captured the EO process and provide a bioink capable of constructing layered and orderly mineralized GP organoids in the future.The results of this study could therefore provide a potential target for regulating the EO process and a novel strategy for simulating it in bone organoids.

3D bioprinted hyaluronic acid‑based cell‑laden scaffold for brain microenvironment simulation

Treatments for lesions in central nervous system(CNS)are always faced with challenges due to the anatomical and physiological particularity of the CNS despite the fact that several achievements have been made in early diagnosis and precision medicine to improve the survival and quality of life of patients with brain tumors in recent years.Understanding the complexity as well as role of the microenvironment of brain tumors may suggest a better revealing of the molecular mechanism of brain tumors and new therapeutic directions,which requires an accurate recapitulation of the complex microenvironment of human brain in vitro.Here,a 3D bioprinted in vitro brain matrix-mimetic microenvironment model with hyaluronic acid(HA)and normal glial cells(HEBs)is developed which simulates both mechanical and biological properties of human brain microenvironment in vivo through the investigation of the formulation of bioinks and optimization of printing process and parameters to study the effects of different concentration of gelatin(GA)within the bioink and different printing structures of the scaffold on the performance of the brain matrix-mimetic microenvironment models.The study provides experimental models for the exploration of the multiple factors in the brain microenvironment and scaffolds for GBM invasion study.

Coaxial bioprinted microfibers with mesenchymal stem cells for glioma microenvironment simulation

Due to their special anatomical and physiological features,central nervous system diseases still presented challenges,despite the fact that some advances have been made in early diagnosis and precision medicine.One of the complexities in treating tumors is the tumor microenvironment,which includes mesenchymal stem cells(MSCs)that exhibit tumor tropism and can be used for cell therapy.However,whether MSCs promote or suppress gliomas is still unclear,especially in glioma microenvironments.In this study,a coaxial microfiber was designed to mimic the tumor microenvironment and to reveal the effect of MSCs on glioma cells.The fiber shell was composed of MSCs and alginate,and the core was filled with U87 MG(glioblastoma)cells and gelatin methacrylate.This Shell-MSC/Core-U87 MG microenvironment improved the proliferation,survival,invasion,metastasis,and drug resistance of glioma cells,while simultaneously maintaining the stemness of glioma cells.In summary,coaxial extrusion bioprinted Shell-MSC/Core-U87 MG microfiber is an ideal platform for tumor and stromal cell coculture to observe tumor biological behavior in vitro.

3D-bioprinted cholangiocarcinoma-on-a-chip model for evaluating drug responses

Cholangiocarcinoma(CCA)is characterized by heterogeneous mutations and a refractory nature.Thus,the development of a model for effective drug screening is urgently needed.As the established therapeutic testing models for CCA are often ineffective,we fabricated an enabling three-dimensional(3D)-bioprinted CCA-on-a-chip model that to a good extent resembled the multicellular microenvironment and the anatomical microstructure of the hepato-vascular-biliary system to perform high-content antitumor drug screening.Specifically,cholangiocytes,hepatocytes,and vascular endotheliocytes were employed for 3D bioprinting of the models,allowing for a high degree of spatial and tube-like microstructural control.Interestingly,it was possible to observe CCA cells attached to the surfaces of the gelatin methacryloyl(GelMA)hydrogelembedded microchannels and overgrown in a thickening manner,generating bile duct stenosis,which was expected to be analogous to the in vivo configuration.Over 4000 differentially expressed genes were detected in the CCA cells in our 3D coculture model compared to the traditional two-dimensional(2D)monoculture.Further screening revealed that the CCA cells grown in the 3D traditional model were more sensitive to the antitumoral prodrug than those in the 2D monoculture due to drug biotransformation by the neighboring functional hepatocytes.This study provides proof-of-concept validation of our bioprinted CCA-on-a-chip as a promising drug screening model for CCA treatment and paves the way for potential personalized medicine strategies for CCA patients in the future.

Adenosine-treated bioprinted muscle constructs prolong cell survival and improve tissue formation

It is crucial to maintaining the viability of biofabricated human-sized tissues to ensure their successful survival and function after transplantation.Adenosine is a purine nucleoside that has the function to suppress cellular metabolism and has been previously proposed as a method to prolong cell viability under hypoxia.In this study,we optimized the dose concentration of adenosine for incorporation into bioprinted constructs to preserve long-term cell viability in vitro.Our results showed that muscle cells(C2C12)containing 6,7,or 8 mM adenosine maintained high cell viability for 20 days under hypoxic conditions(0.1%O2),whereas cells without adenosine treatment showed 100%cell death after 11 days.After 20 days under hypoxic conditions,muscle cells treated with adenosine proliferated and differentiated when transferred to normoxic conditions.From these adenosine concentrations,6 mM was picked as the optimized adenosine concentration for further investigations due to its most effective results on improving cell viability.The bioprinted muscle constructs containing adenosine(6 mM)maintained high cell viability for 11 days under hypoxic conditions,while the control constructs without adenosine had no live cells.For in vivo validation,the bioprinted constructs with adenosine implanted under the dorsal subcutaneous space in mice,showed the enhanced formation of muscle tissue with minimal central necrosis and apoptosis,when compared to the constructs without adenosine.These positive in vitro and in vivo results demonstrate that the use of adenosine is an effective approach to preventing the challenge of hypoxia-induced necrosis in bioprinted tissues for clinical translation.

Bioprinted mesenchymal stem cell microfiber-derived extracellular vesicles alleviate unilateral renal ischemia-reperfusion injury and fibrosis by inhibiting tubular epithelial cells ferroptosis

Renal unilateral ischemia-reperfusion injury(UIRI)constitutes a significant global health challenge,with poor recovery leading to chronic kidney disease and subsequent renal fibrosis.Extracellular vesicles(EVs)present substantial potential benefits for renal diseases.However,the limited yield and efficacy of EVs produced through traditional methodologies(2D-EVs)severely restrict their widespread application.Moreover,the efficient and effective strategies for using EVs in UIRI treatment and their mechanisms remain largely unexplored.In this study,we propose an innovative approach by integrating bioprinted mesenchymal stem cell microfiber extracellular vesicles production technology(3D-EVs)with a tail vein injection method,introducing a novel treatment strategy for UIRI.Our comparison of the biological functions of 2D-EVs and 3D-EVs,both in vitro and in vivo,reveals that 3D-EVs significantly outperform 2D-EVs.Specifically,in vitro,3D-EVs demonstrate a superior capacity to enhance the proliferation and migration of NRK-52E cells and mitigate hypoxia/reoxygenation(H/R)-induced injuries by reducing epithelial-mesenchymal transformation,extracellular matrix deposition,and ferroptosis.In vivo,3D-EVs exhibit enhanced therapeutic effects,as evidenced by improved renal function and decreased collagen deposition in UIRI mouse kidneys.We further elucidate the mechanism by which 3D-EVs derived from KLF15 ameliorate UIRI-induced tubular epithelial cells(TECs)ferroptosis through the modulation of SLC7A11 and GPX4 expression.Our findings suggest that bioprinted mesenchymal stem cells microfiberderived EVs significantly ameliorate renal UIRI,opening new avenues for effective and efficient EV-based therapies in UIRI treatment.

Multi-modal imaging for dynamic visualization of osteogenesis and implant degradation in 3D bioprinted scaffolds

In situ monitoring of bone regeneration enables timely diagnosis and intervention by acquiring vital biological parameters.However,an existing gap exists in the availability of effective methodologies for continuous and dynamic monitoring of the bone tissue regeneration process,encompassing the concurrent visualization of bone formation and implant degradation.Here,we present an integrated scaffold designed to facilitate real-time monitoring of both bone formation and implant degradation during the repair of bone defects.Laponite(Lap),CyP-loaded mesoporous silica(CyP@MSNs)and ultrasmall superparamagnetic iron oxide nanoparticles(USPIO@SiO2)were incorporated into a bioink containing bone marrow mesenchymal stem cells(BMSCs)to fabricate functional scaffolds denoted as C@M/GLU using 3D bioprinting technology.In both in vivo and in vitro experiments,the composite scaffold has demonstrated a significant enhancement of bone regeneration through the controlled release of silicon(Si)and magnesium(Mg)ions.Employing near-infrared fluorescence(NIR-FL)imaging,the composite scaffold facilitates the monitoring of alkaline phosphate(ALP)expression,providing an accurate reflection of the scaffold’s initial osteogenic activity.Meanwhile,the degradation of scaffolds was monitored by tracking the changes in the magnetic resonance(MR)signals at various time points.These findings indicate that the designed scaffold holds potential as an in situ bone implant for combined visualization of osteogenesis and implant degradation throughout the bone repair process.

3-D bioprinted human-derived skin organoids accelerate full-thickness skin defects repair

The healing of large skin defects remains a significant challenge in clinical settings.The lack of epidermal sources,such as autologous skin grafting,limits full-thickness skin defect repair and leads to excessive scar formation.Skin organoids have the potential to generate a complete skin layer,supporting in-situ skin regeneration in the defect area.In this study,skin organoid spheres,created with human keratinocytes,fibroblasts,and endothelial cells,showed a specific structure with a stromal core surrounded by surface keratinocytes.We selected an appropriate bioink and innovatively combined an extrusion-based bioprinting technique with dual-photo source cross-linking technology to ensure the overall mechanical properties of the 3D bioprinted skin organoid.Moreover,the 3D bioprinted skin organoid was customized to match the size and shape of the wound site,facilitating convenient implantation.When applied to full-thickness skin defects in immunodeficient mice,the 3D bioprinted human-derived skin organoid significantly accelerated wound healing through in-situ regeneration,epithelialization,vascularization,and inhibition of excessive inflammation.The combination of skin organoid and 3D bioprinting technology can overcome the limitations of current skin substitutes,offering a novel treatment strategy to address large-area skin defects.

3D bioprinted breast cancer model reveals stroma-mediated modulation of extracellular matrix and radiosensitivity

Deciphering breast cancer treatment resistance remains hindered by the lack of models that can successfully capture the four-dimensional dynamics of the tumor microenvironment.Here,we show that microextrusion bioprinting can reproducibly generate distinct cancer and stromal compartments integrating cells relevant to human pathology.Our findings unveil the functional maturation of this millimeter-sized model,showcasing the development of a hypoxic cancer core and an increased surface proliferation.Maturation was also driven by the presence of cancer-associated fibroblasts(CAF)that induced elevated microvascular-like structures complexity.Such modulation was concomitant to extracellular matrix remodeling,with high levels of collagen and matricellular proteins deposition by CAF,simultaneously increasing tumor stiffness and recapitulating breast cancer fibrotic development.Importantly,our bioprinted model faithfully reproduced response to treatment,further modulated by CAF.Notably,CAF played a protective role for cancer cells against radiotherapy,facilitating increased paracrine communications.This model holds promise as a platform to decipher interactions within the microenvironment and evaluate stroma-targeted drugs in a context relevant to human pathology.

Redirecting differentiation of mammary progenitor cells by 3D bioprinted sweat gland microenvironment

Background:Mammary progenitor cells(MPCs)maintain their reproductive potency through life,and their specific microenvironments exert a deterministic control over these cells.MPCs provides one kind of ideal tools for studying engineered microenvironmental influence because of its accessibility and continually undergoes postnatal developmental changes.The aim of our study is to explore the critical role of the engineered sweat gland(SG)microenvironment in reprogramming MPCs into functional SG cells.Methods:We have utilized a three-dimensional(3D)SG microenvironment composed of gelatin-alginate hydrogels and components from mouse SG extracellular matrix(SG-ECM)proteins to reroute the differentiation of MPCs to study the functions of this microenvironment.MPCs were encapsulated into the artificial SG microenvironment and were printed into a 3D cell-laden construct.The expression of specific markers at the protein and gene levels was detected after cultured 14 days.Results:Compared with the control group,immunofluorescence and gene expression assay demonstrated that MPCs encapsulated in the bioprinted 3D-SG microenvironment could significantly express the functional marker of mouse SG,sodium/potassium channel protein ATP1a1,and tend to express the specific marker of luminal epithelial cells,keratin-8.When the Shh pathway is inhibited,the expression of SG-associated proteins in MPCs under the same induction environment is significantly reduced.Conclusions:Our evidence proved the ability of differentiated mouse MPCs to regenerate SG cells by engineered SG microenvironment in vitro and Shh pathway was found to be correlated with the changes in the differentiation.These results provide insights into regeneration of damaged SG by MPCs and the role of the engineered microenvironment in reprogramming cell fate.

A 3D-bioprinted scaffold with doxycycline-controlled BMP2-expressing cells for inducing bone regeneration and inhibiting bacterial infection

Large bone defects face a high risk of pathogen exposure due to open wounds,which leads to high infection rates and delayed bone union.To promote successful repair of infectious bone defects,fabrication of a scaffold with dual functions of osteo-induction and bacterial inhibition is required.This study describes creation of an engineered progenitor cell line(C3H10T1/2)capable of doxycycline(DOX)-mediated release of bone morphogenetic protein-2(BMP2).Three-dimensional bioprinting technology enabled creation of scaffolds,comprising polycaprolactone/mesoporous bioactive glass/DOX and bioink,containing these engineered cells.In vivo and in vitro experiments confirmed that the scaffold could actively secrete BMP2 to significantly promote osteoblast differentiation and induce ectopic bone formation.Additionally,the scaffold exhibited broad-spectrum antibacterial capacity,thereby ensuring the survival of embedded engineered cells when facing high risk of infection.These findings demonstrated the efficacy of this bioprinted scaffold to release BMP2 in a controlled manner and prevent the occurrence of infection;thus,showing its potential for repairing infectious bone defects.

Bioprinted constructs that simulate nerve-bone crosstalk to improve microenvironment for bone repair

Crosstalk between nerves and bone is essential for bone repair,for which Schwann cells(SCs)are crucial in the regulation of the microenvironment.Considering that exosomes are critical paracrine mediators for intercellular communication that exert important effects in tissue repair,the aim of this study is to confirm the function and molecular mechanisms of Schwann cell-derived exosomes(SC-exos)on bone regeneration and to propose engineered constructs that simulate SC-mediated nerve-bone crosstalk.SCs promoted the proliferation and differentiation of bone marrow mesenchymal stem cells(BMSCs)through exosomes.Subsequent molecular mechanism studies demonstrated that SC-exos promoted BMSC osteogenesis by regulating the TGF-βsignaling pathway via let-7c-5p.Interestingly,SC-exos promoted the migration and tube formation performance of endothelial progenitor cells.Furthermore,the SC-exos@G/S constructs were developed by bioprinting technology that simulated SC-mediated nerve-bone crosstalk and improved the bone regeneration microenvironment by releasing SC-exos,exerting the regulatory effect of SCs in the microenvironment to promote innervation,vascularization,and osteogenesis and thus effectively improving bone repair in a cranial defect model.This study demonstrates the important role and underlying mechanism of SCs in regulating bone regeneration through SC-exos and provides a new engineered strategy for bone repair.

3D-bioprinted microenvironments for sweat gland regeneration

The development of 3D bioprinting in recent years has provided new insights into the creation of in vitro microenvironments for promoting stem cell-based regeneration.Sweat glands(SGs)are mainly responsible for thermoregulation and are a highly differentiated organ with limited regenerative ability.Recent studies have focused on stem cell-based therapies as strategies for repairing SGs after deep dermal injury.In this review,we highlight the recent trend in 3D bioprinted native-like microenvironments and emphasize recent advances in functional SG regeneration using this technology.Furthermore,we discuss five possible regulatory mechanisms in terms of biochemical factors and structural and mechanical cues from 3D bioprinted microenvironments,as well as the most promising regulation from neighbor cells and the vascular microenvironment.

A 3D-Bioprinted dual growth factor-releasing intervertebral disc scaffold induces nucleus pulposus and annulus fibrosus reconstruction

Regeneration of Intervertebral disc(IVD)is a scientific challenge because of the complex structure and composition of tissue,as well as the difficulty in achieving bionic function.Here,an anatomically correct IVD scaffold composed of biomaterials,cells,and growth factors were fabricated via three-dimensional(3D)bioprinting technology.Connective tissue growth factor(CTGF)and transforming growth factor-β3(TGF-β3)were loaded onto polydopamine nanoparticles,which were mixed with bone marrow mesenchymal stem cells(BMSCs)for regenerating and simulating the structure and function of the nucleus pulposus and annular fibrosus.In vitro experiments confirmed that CTGF and TGF-β3 could be released from the IVD scaffold in a spatially controlled manner,and induced the corresponding BMSCs to differentiate into nucleus pulposus like cells and annulus fibrosus like cells.Next,the fabricated IVD scaffold was implanted into the dorsum subcutaneous of nude mice.The reconstructed IVD exhibited a zone-specific matrix that displayed the corresponding histological and immunological phenotypes:primarily type II collagen and glycosaminoglycan in the core zone,and type I collagen in the surrounding zone.The testing results demonstrated that it exhibited good biomechanical function of the reconstructed IVD.The results presented herein reveal the clinical application potential of the dual growth factors-releasing IVD scaffold fabricated via 3D bioprinting.However,the evaluation in large mammal animal models needs to be further studied.

Encapsulated three-dimensional bioprinted structure seeded with urothelial cells:a new construction technique for tissue-engineered urinary tract patch

Background:Traditional tissue engineering methods to fabricate urinary tract patch have some drawbacks such as compromised cell viability and uneven cell distribution within scaffold.In this study,we combined three-dimensional(3D)bioprinting and tissue engineering method to form a tissue-engineered urinary tract patch,which could be employed for the application on Beagles urinary tract defect mode to verify its effectiveness on urinary tract reconstruction.Methods:Human adipose-derived stem cells(hADSCs)were dropped into smooth muscle differentiation medium to generate induced microtissues(ID-MTs),flow cytometry was utilized to detect the positive percentage for CD44,CD105,CD45,and CD34 of hADSCs.Expression of vascular endothelial growth factor A(VEGFA)and tumor necrosis factor-stimulated gene-6(TSG-6)in hADSCs and MTs were identified by Western blotting.Then the ID-MTs were employed for 3D bioprinting.The bioprinted structure was encapsulated by transplantation into the subcutaneous tissue of nude mice for 1 week.After retrieval of the encapsulated structure,hematoxylin and eosin and Masson’s trichrome staining were performed to demonstrate the morphology and reveal collagen and smooth muscle fibers,integral optical density(IOD)and area of interest were calculated for further semiquantitative analysis.Immunofluorescent double staining of CD31 andα-smooth muscle actin(α-SMA)were used to reveal vascularization of the encapsulated structure.Immunohistochemistry was performed to evaluate the expression of interleukin-2(IL-2),α-SMA,and smoothelin of the MTs in the implanted structure.Afterward,the encapsulated structure was seeded with human urothelial cells.Immunofluorescent staining of cytokeratins AE1/AE3 was applied to inspect the morphology of seeded encapsulated structure.Results:The semi-quantitative assay showed that the relative protein expression of VEGFA was 0.355±0.038 in the hADSCs vs.0.649±0.150 in the MTs(t=3.291,P=0.030),while TSG-6 expression was 0.492±0.092 in the hADSCs vs.1.256±0.401 in the MTs(t=3.216,P=0.032).The semi-quantitative analysis showed that the mean IOD of IL-2 in the MT group was 7.67±1.26,while 12.6±4.79 in the hADSCs group,but semi-quantitative analysis showed that there was no statistical significance in the difference between the two groups(t=1.724,P=0.16).The semi-quantitative analysis showed that IOD was 71.7±14.2 in non-induced MTs(NI-MTs)vs.35.7±11.4 in ID-MTs for collagen fibers(t=3.428,P=0.027)and 12.8±1.9 in NI-MTs vs.30.6±8.9 in ID-MTs for smooth muscle fibers(t=3.369,P=0.028);furthermore,the mean IOD was 0.0613±0.0172 in ID-MTs vs.0.0017±0.0009 in NI-MTs forα-SMA(t=5.994,P=0.027),while 0.0355±0.0128 in ID-MTs vs.0.0035±0.0022 in NI-MTs for smoothelin(t=4.268,P=0.013),which indicate that 3D bioprinted structure containing ID-MTs could mimic the smooth muscle layer of native urinary tract.After encapsulation of the urinary tract patch for additional cell adhesion,urothelial cells were seeded onto the encapsulated structures,and a monolayer urothelial cell was observed.Conclusion:Through 3D bioprinting and tissue engineering methods,we provided a promising way to fabricate tissue-engineered urinary tract patch for further investigation.

Utilizing 3D bioprinted platelet-rich fibrin-based materials to promote the regeneration of oral soft tissue

Oral soft tissue defects remain difficult to treat owing to the limited efficacy of available treatment materials.Although the injectable platelet-rich fibrin(i-PRF)is a safe,autologous source of high levels of growth factors that is often employed to promote the regeneration of oral soft tissue,its effectiveness is restrained by difficulties in intraoperative shaping together with the burst-like release of growth factors.We herein sought to develop a bioactive bioink composed of i-PRF,alginate and gelatin capable of promoting the regeneration of the oral soft tissue.This bioink was successfully applied in 3D bioprinting and exhibited its ability to be shaped to individual patient needs.Importantly,we were also able to significantly prolong the duration of multiple growth factors release as compared to that observed for i-PRF.The growth factor bioavailability was further confirmed by the enhanced proliferation and viability of printed gingival fibroblasts.When deployed in vivo in nude mice,this bioink was further confirmed to be biocompatible and to drive enhanced angiogenic activity.Together,these data thus confirmthe successful production of an i-PRF-containing bioink,which is suitable for the individualized promotion of the regeneration of oral soft tissue.

3D Bioprinted Skin Substitutes for Accelerated Wound Healing and Reduced Scar

The shortage of skin for grafting continues to be a major problem in the treatment of serious skin injuries.3D bioprinting provides a new way to solve this problem.However,current 3D printed skin is less effective in treatment of large wounds because of severe shrinkage and scarring.In this study,bionically designed bilayer skin was fabricated using an extrusion-based bioprinter and a gelatin/sodium alginate/gelatin methacrylate hydrogel with excellent physical and biological properties.Full-thickness skin wounds were created in the back of nude mice and treated with bioprinted skin or hydrogel.Bioprinted skin accelerated wound healing,reduced wound contraction and scarring,and facilitated wound skin epithelialization compared with the bioprinted hydrogel or untreated wound.The skin from the wound was collected 28 days after grafting for histology and immunofluorescence analysis.The thickness of the dermis and epidermis of the bioprinted skin was similar to that of nude mice.Microvascular formation in the dermis and dense keratinocytes in the epidermis of the bioprinted skin were observed.This study provides a potential treatment strategy for reducing skin contraction and scar in large skin wounds.

Three-dimensional-engineered bioprinted in vitro human neural stem cell self-assembling culture model constructs of Alzheimer’s disease

The pathogenic cascade of Alzheimer’s disease(AD)characterized by amyloid-β protein accumulation is still poorly understood,partially owing to the limitations of relevant models without in vivo neural tissue microenvironment to recapitulate cell-cell interactions.To better mimic neural tissue microenvironment,three-dimensional(3D)core-shell AD model constructs containing human neural progenitor cells(NSCs)with 2% matrigel as core bioink and 2% alginate as shell bioink have been bioprinted by a co-axial bioprinter,with a suitable shell thickness for nutrient exchange and barrier-free cell interaction cores.These constructs exhibit cell self-clustering and-assembling properties and engineered reproducibility with long-term cell viability and self-renewal,and a higher differentiation level compared to 2D and 3D MIX models.The different effects of 3D bioprinted,2D,and MIX microenvironments on the growth of NSCs are mainly related to biosynthesis of amino acids and glyoxylate and dicarboxylate metabolism on day 2 and ribosome,biosynthesis of amino acids and proteasome on day 14.Particularly,the model constructs demonstrated Aβ aggregation and higher expression of Aβ and tau isoform genes compared to 2D and MIX controls.AD model constructs will provide a promising strategy to facilitate the development of a 3D in vitro AD model for neurodegeneration research.