Effect of ionic crosslinking on morphology and thermostability of biomimetic supercritical fluids-decellularized dermal-based composite bioscaffolds for bioprinting applications.

In the present study, supercritical fluid was employed to prepare a kind of supercritical fluids-decellularized dermal-based scaffold (SFDDS) from porcine dermal tissue. Further, new composite bioscaffolds containing SFDDS were designed for bioprinting applications. Then, the effect of crosslinking functionality on microstructures and thermal properties of the composite bioscaffolds containing decellularized extracellular matrix were studied. The results of thermal stability from thermogravimetric analysis and difference thermogravimetry demonstrated the structural stability of the composite bioscaffolds. A method was designed to prepare bioinspired decellularized dermal-based composite bioscaffolds, which were further characterized by infrared spectroscopy, scanning electron microscopy, and thermogravimetry analysis.

Highly Organized Porous Gelatin-Based Scaffold by Microfluidic 3D-Foaming Technology and Dynamic Culture for Cartilage Tissue Engineering.

A gelatin-based hydrogel scaffold with highly uniform pore size and biocompatibility was fabricated for cartilage tissue engineering using microfluidic 3D-foaming technology. Mainly, bubbles with different diameters, such as 100 µm and 160 µm, were produced by introducing an optimized nitrogen gas and gelatin solution at an optimized flow rate, and N2/gelatin bubbles were formed. Furthermore, a cross-linking agent (1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide, EDC) was employed for the cross-linking reaction of the gelatin-based hydrogel scaffold with uniform bubbles, and then the interface between the close cells were broken by degassing. The pore uniformity of the gelatin-based hydrogel scaffolds was confirmed by use of a bright field microscope, conjugate focus microscope and scanning electron microscope. The in vitro degradation rate, mechanical properties, and swelling rate of gelatin-based hydrogel scaffolds with highly uniform pore size were studied. Rabbit knee cartilage was cultured, and its extracellular matrix content was analyzed. Histological analysis and immunofluorescence staining were employed to confirm the activity of the rabbit knee chondrocytes. The chondrocytes were seeded into the resulting 3D porous gelatin-based hydrogel scaffolds. The growth conditions of the chondrocyte culture on the resulting 3D porous gelatin-based hydrogel scaffolds were evaluated by MTT analysis, live/dead cell activity analysis, and extracellular matrix content analysis. Additionally, a dynamic culture of cartilage tissue was performed, and the expression of cartilage-specific proteins within the culture time was studied by immunofluorescence staining analysis. The gelatin-based hydrogel scaffold encouraged chondrocyte proliferation, promoting the expression of collagen type II, aggrecan, and sox9 while retaining the structural stability and durability of the cartilage after dynamic compression and promoting cartilage repair.

Extraction and characterization of bovine collagen Type V and its effects on cell behaviors.

Collagen Type V (Col. V) plays an essential role in cell behaviors and has attracted increasing attention in recent years. High-purity Col. V is needed for evaluating its biological properties. In this research, the enzymatic hydrolysis process was combined with ultrafiltration to purify Col. V from the bovine cornea. The purity of Col. V was determined to be above 90% by both sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and high-performance liquid chromatography methods. The effect of Col. V on cell behaviors was evaluated. The circular dichroism spectroscopy results demonstrated that the extracted Col. V exhibited a complete triple helix structure. SDS-PAGE suggested that the molecular weight of Col. V was 440 kDa. The self-assembly experiment revealed that the proportion of Col. V in the collagen mixture can affect the Col. I fiber diameter. The cell culture results implied that Col. V can inhibit fibroblasts (L929) proliferation. The L929 showed maximum mobility when the addition of Col. V was 30%. Thus, Col. V has the effect of inhibiting L929 proliferation and promoting migration. The high-purity Col. V provides useful information for further understanding its biological implications.

Design and Characterization of a Bioinspired Polyvinyl Alcohol Matrix with Structural Foam-Wall Microarchitectures for Potential Tissue Engineering Applications.

Traditional medical soft matrix used in a surgical treatment or in wound management was not good enough in both the structural support and interconnectivity to be applied in tissue engineering as a scaffold. Avian skeleton and feather rachises might be good reference objects to mimic in designing a scaffold material with good structural support and high interconnectivity because of its structural foam-wall microarchitectures and structural pneumaticity. In this study, a biomimetic airstream pore-foaming process was built up and the corresponding new medical soft matrix derived from polyvinyl alcohol matrix (PVAM) with air cavities inspired by avian skeleton and feather rachises was prepared. Furthermore, the resulting medical soft matrix and bovine Achilles tendon type I collagen could be employed to prepare a new collagen-containing composite matrix. Characterization, thermal stability and cell morphology of the bioinspired PVA matrix and the corresponding collagen-modified PVA composite matrix with open-cell foam-wall microarchitectures were studied for evaluation of potential tissue engineering applications. TGA, DTG, DSC, SEM and FTIR results of new bioinspired PVA matrix were employed to build up the effective system identification approach for biomimetic structure, stability, purity, and safety of target soft matrix. The bioinspired PVA matrix and the corresponding collagen-modified PVA composite matrix would be conductive to human hepatoblastoma HepG2 cell proliferation, migration, and expression which might serve as a promising liver cell culture carrier to be used in the biological artificial liver reactor.

Evaluation and Preparation of a Designed Kartogenin Drug Delivery System (DDS) of Hydrazone-Linkage-Based pH Responsive mPEG-Hz-b-PCL Nanomicelles for Treatment of Osteoarthritis.

Osteoarthritis (OA) is a chronic disease caused by the damage of articular cartilage. Kartogenin (KGN) is a well-recognized small molecule which could induce MSCs chondrogenesis and promote cartilage repair treatments. Nano-level micells could be a suitable drug carrier technology for the treatments. In this study, the acid-responsive methoxy poly(ethylene oxide)-hydrazone-poly(ε-caprolactone) copolymers, mPEG-Hz-b-PCL, were synthesized. The structure was characterized by 1H NMR. The evaluation of a designed kartogenin drug delivery system (DDS) of hydrazone-linkage-based pH responsive mPEG-Hz-b-PCL nanomicelles for treatment of osteoarthritis could be carried out.

Quantitation of Collagen Type V in Tissues by High-Performance Liquid Chromatography Coupled to Mass Spectrometry.

A method for quantifying the bovine collagen type V (Col. V) was established based on high-performance liquid chromatography coupled to mass spectrometry by the marker peptide external standard. High-purity Col. V was extracted by the acid-enzyme hydrolysis process, and the marker peptide of Col. V was identified by LCQ mass spectrometry as GPAGPMGLTGR. A broad linear range (0.01-5.00 µg/mL) with a correlation coefficient of 0.9984 was achieved, and the limit of detection and limit of quantification were found to be 3.00 × 10-3 and 6.25 × 10-3 µg/mL, respectively. The method precision was 1.49%. The recovery rate was determined as 97.1-109.6% with a relative standard deviation less than 5%. The proposed method was successfully applied for the determination of Col. V contents in the bovine heart, lung, and cornea, which were 0.72 ± 0.01%, 0.23 ± 0.01%, and 2.89 ± 0.00%, respectively. The results show that the proposed method is more suitable for measuring the content of Col. V in tissue samples compared with the enzyme-linked immunosorbent assay. The marker peptide method has high accuracy and great reproducibility, and will lay a foundation for the extraction and application of Col. V. Impact statement The accurate quantitative method for collagen type V (Col. V) is particularly important in scientific research, disease diagnosis and treatment, and industrial production. In this article, we proposed a high-performance liquid chromatography coupled to mass spectrometry method based on the external standard marker peptide to quantify bovine Col. V. This method shows a higher accuracy and recovery rate than enzyme-linked immunosorbent assay (ELISA), indicating that it is more suitable for measuring the content of Col. V in tissue samples than ELISA. The establishment of this method has laid a solid foundation for the extraction and application of Col. V.

Decellularized liver-regenerative 3D printing biomaterials for cell-based liver therapies via a designed procedure combining supercritical fluids with papain-containing reagents.

BACKGROUND: The biologic scaffolds derived from decellularized tissues and organs have been successfully developed in a variety of preclinical and/or clinical studies. OBJECTIVE: The new decellularized liver-regenerative 3D printing biomaterials were designed and prepared for cell-based liver therapies. METHODS: An extraction process was employed to remove the tissue and cellular molecules from porcine liver via pretreatment of supercritical fluid of carbon dioxide (ScCO2). Varying porosities of the decellularized liver tissues were created using papain-containing reagent treatments after ScCO2. RESULTS: The resulting liver-regenerative 3D printing biomaterials of decellularized liver collagen scaffolds were characterized by Fourier transform infrared spectroscopy, thermo-gravimetric analysis, differential scanning calorimetry and scanning electron microscopy. CONCLUSIONS: The decellularized liver collagen scaffolds with good thermal stability (>150 °C) were obtained and employed as liver-regenerative 3D printing biomaterials for cell-based liver therapies.

Newly Designed Decellularized Scaffolds for Scaffold-based Gene Therapy from Elastic Cartilages via Supercritical Carbon Dioxide Fluid and Alkaline/ Protease Treatments.

BACKGROUND: Scaffold-based gene therapy provides a promising approach for tissue engineering, which was important and popular as it combined medical applications and engineering materials' knowledge. OBJECTIVE: The decellularization techniques were employed to remove the cellular components from porcine elastic cartilages, leaving a native decellularized Extracellular Matrix (dECM) composition and architecture integrity of largely insoluble collagen, elastin, and tightly bound glycosaminoglycans. For newly designed collagen scaffold samples, elastic cartilages were hydrolyzed by protease with different concentrations to gain state completely and clearly. METHODS: An extraction process of Supercritical Carbon Dioxide (ScCO2) was used to remove cellular components from porcine elastic cartilage. The dECM scaffolds with collagen must be characterized by Fourier transform infrared spectroscopy(FTIR), Thermo-Gravimetric Analysis (TGA), and Scanning Electron Microscope (SEM). RESULTS: The study provided a new treatment combined with supercritical carbon dioxide and alkaline/ protease to prepare dECM scaffolds with hole-scaffold microstructures and introduce into a potential application on osteochondral tissue engineering using scaffold-based gene therapy. The new process is simple and efficient. The pore-scaffold microstructures were observed in dECM scaffolds derived from porcine elastic cartilages. The Tdmax values of the resulting dECM scaffolds were observed at over 330oC. CONCLUSION: A series of new scaffolds were successfully obtained from porcine tissue by using ScCO2 and alkaline/enzyme treatments such as a mixing aqueous solution of NH4OH and papain. The dECM scaffolds with high thermal stability were obtained. The resulting scaffold with clean pore-scaffold microstructure could be a potential application for scaffold-based gene therapy.

Characterization of Composite Nano-Bioscaffolds Based on Collagen and Supercritical Fluids-Assisted Decellularized Fibrous Extracellular Matrix.

Nano-bioscaffolds obtained from decellularized tissues have been employed in several medical applications. Nano-bioscaffolds could provide structural support for cell attachment and a suitable environment with sufficient porosity for cell growth and proliferation. In this study, a new combined method constitutes a decellularization protocol to remove the tissue and cellular molecules from porcine dermis for preparation of nano-bioscaffolds with fibrous extracellular matrix via pre- and post-treatment of supercritical fluids. The supercritical fluids-assisted nano-bioscaffolds were characterized by peptide identification, infrared spectrum of absorption, morphology, histological observations, DNA quantification, and hemocompatibility. Further, the resulting nano-bioscaffolds could be employed to obtain new cross-linked composite nano-bioscaffold containing collagen and acellular matrix.

Characteristics and Preparation of Designed Alginate-Based Composite Scaffold Membranes with Decellularized Fibrous Micro-Scaffold Structures from Porcine Skin.

Alginate-based composite scaffold membranes with various ratios of decellularized extracellular matrices could be designed and obtained from porcine skin tissue by using supercritical carbon dioxide fluid technology. Retention of decellularized extracellular matrix (dECM) and scaffold-structure integrity was observed. This work provides a simple and time-saving process for the preparation of biomedical alginate-based composite scaffold membranes with fibrous dECM micro-scaffolds, which were further characterized by Fourier transform infrared spectroscopy (FTIR), thermo-gravimetric analysis (TGA), and scanning electron microscope (SEM). The introduction of fibrous dECM micro-scaffolds enhanced the thermal stability and provided expected effects on the biological properties of the designed composite scaffold membranes in regenerative applications.

Biological evaluations of decellularized extracellular matrix collagen microparticles prepared based on plant enzymes and aqueous two-phase method.

For patients with extensive full-thickness burns who do not have sufficient autologous split-thickness skin for skin grafts, the application of biological skin substitutes may be considered. The aim of this study was to find an optimal new type method for the production of a biovital skin substitute based on acellular dermal matrix (ADM) and preclinical evaluations. In this work, 25 methods of ADM production were assessed. The proposed methods are based on the use of the following enzymes: papain, Carica papaya lipase (CPL), and purification using a polymer/salt aqueous two-phase system. The obtained ADM samples were characterized via scanning electron microscopy (SEM), porosity measurement and water vapor transmission test. Results showed that the collagen bundles of ADM microparticles were intact and orderly. Through differential scanning calorimetry (DSC), thermo gravimetric analysis (TGA) and biocompatibility tests, the results indicated that the proportion of papain and CPL was the same and 5 h processing time are the optimum conditions for ADM preparation and the material showed good biocompatibility. Our results suggested that the potential of developing this kind of decellularization process to manufacture ADM scaffolds for clinical application.

Microstructure and Thermal Property of Designed Alginate-Based Polymeric Composite Foam Materials Containing Biomimetic Decellularized Elastic Cartilage Microscaffolds.

This study presents a designed alginate-based polymeric composite foam material containing decellularized elastic cartilage microscaffolds from porcine elastic cartilage by using supercritical fluid and papain treatment for medical scaffold biomaterials. The microstructure and thermal property of the designed alginate-based polymeric composite foam materials with various controlled ratios of alginate molecules and decellularized elastic cartilage microscaffolds were studied and characterized by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and differential thermal gravimetric analysis (TGA/DTG). The microstructure and thermal property of the composite foam materials were affected by the introduction of decellularized elastic cartilage microscaffolds. The designed alginate-based polymeric composite foam materials containing decellularized elastic cartilage microscaffolds were ionically cross-linked with calcium ions by soaking the polymeric composite foam materials in a solution of calcium chloride. Additional calcium ions further improved the microstructure and thermal stability of the resulting ionic cross-linked alginate-based polymeric composite foam materials. Furthermore, the effect of crosslinking functionality on microstructures and thermal properties of the resulting polymeric composite foam materials were studied to build up useful information for 3D substrates for cultivating and growing cartilage cells and/or cartilage tissue engineering.

Anti-inflammation and anti-apoptosis effects of pearl extract gel on UVB irradiation HaCaT cells.

Caused by acute radiation skin reaction and injury, receiving radiotherapy treatment process is often performed side-effects on cancer patients. The clinical manifestations of skin irritation, itching, peeling, pigmentation, ulcer bleeding and other symptoms, in addition to causing patient discomfort and affecting quality of life, may increase the risk of local or systemic infection, and lead to interruption of radiation therapy. At present, for acute radiation dermatitis, there is no uniform treatment, and the various methods are evaluated variously. In this study, the authors focus on broken pearls using room temperature super extraction system, the water extraction process of wet-grinding method, nano-scale pearl, along with a large number of high purity natural amino acid extracts in the water. The room-temperature super-extraction system (RTSES) can be extracted from a relatively high-volume of pearl extract. We use pearl extract as the main component of experimental material, and the blending of pearl extract and poly (γ-glutamic acid) is used to form biodegradable composite hydrogels. This study aims to evaluate the use of RTSES to extract the major active components of pearl and enhance their anti-inflammation and anti-apoptosis effects. The possible effect of pearl extract on inducing apoptosis in human keratinocyte cells (HaCaT) under the exposure of low dose UVB has been investigated. Various concentrations of pearl extracts have been used to study the effect of low dosage UVB on HaCaT cells. The results show that pearl extract has no toxic effect on HaCaT cells. Combining the pearl extract and poly (γ-glutamic acid) hydrogels with UVB irradiation would decrease the inflammation and apoptosis of HaCaT cells. The commercial pearl extract has the potential to inhibit radiation dermatitis occurring within keratinocyte cells.

Optically-guided scalpel with light-scattering module for carpal tunnel surgical procedure via minimally invasive surgery.

A novel technique and product for carpal tunnel microscopic surgical procedures using a modified medical device containing a head as a surgical scalpel under light guidance were designed and studied. The novel design of the medical device was developed and applied in a new carpal tunnel microscopic surgical procedure in place of the traditional carpal tunnel surgical procedure. Biological and clinical evaluations of carpal tunnel surgical procedure using the designed optically guided medical device were studied. For commercialized reasons, some guidance was considered, such as ISO 10993-1:2009(E), for the biological evaluation of the device. Furthermore, a clinical evaluation was carried out. The designed optically guided medical device could provide a powerful medical device for carpal tunnel syndrome and related applications.

New designed nerve conduits with a porous ionic cross-linked alginate/chitisan structure for nerve regeneration.

A new fabrication process for designing nerve conduits with a porous ionic cross-linked alginate/chitosan composite for nervous regeneration could be prepared. New designed nerve conduits with a porous ionic cross-linked alginate/chitosan composite were developed for nervous regeneration. Nerve conduits (NCs) represent a promising alternative to conventional treatments for peripheral nerve repair. NCs composed of various polysaccharides such as sodium alginate were designed and prepared by lyophilization as potential matrices for tissue engineering. The use of a porous ionic cross-linked alginate/chitosan composite could provide penetration channels that would lead to the products' increasing penetration rate properties. Furthermore, the use of a porous ionic cross-linked alginate/chitosan composite also has a highly cross-linked structure, which would give the products relatively good mechanical properties. Furthermore, the drug could be incorporated into nerve conduits as a new drug-carrying system for nerve regeneration because of its porous and cross-linked structures.

Fermented broth in tyrosinase- and melanogenesis inhibition.

Fermented broth has a long history of applications in the food, pharmaceutical and cosmetic industries. Recently, the use of fermented broth in skin care products is in ascendance. This review investigates the efficacy of fermented broth in inhibiting tyrosinase and melanogenesis. Possible active ingredients and hypopigmentation mechanisms of fermented broth are discussed, and potential applications of fermented broth in the cosmetic industry are also addressed.

Carbodimide cross-linked and biodegradation-controllable small intestinal submucosa sheets.

The small intestinal submucosa (SIS) is an acellular collagen-based matrix, primarily composed of fibrillar collagens (types I, II, and V). They enhance healing due to a minimal immune response. A good degradation rate is the degradation of materials equal to the rate of remodeling in the host. The SIS should apply a good degradation rate and cytocompatibility. In this study, a series of SIS with different degradation rates is obtained by treatment with Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Morphology, composition, degradable ratio, mechanical properties and cytocompatibility of the SIS are evaluated. We determined a 20 µm thickness and 60 µm pore size of the native SIS. The degradable ratio of the native SIS was approximately 90% in the presence of 0.25 mg/ml collagenase for 24 hours. The storage modulus of the native SIS was 388 MPa. The degradable ratio of the SIS decreased to 6% and the storage modulus increased to 777 MPa after being treated with 30 mM EDC for 24 hours. In cytocompatibility assay, cell numbers on the native SIS were similar as on the treated SIS due to the non-toxicity of the EDC treatment process. This SIS exhibited collagenase resistance, stronger mechanical strength and good cytocompatibility after the EDC treatment concluded. The cross-linked SIS could be utilized as a potential cell carrier for tissue engineering application.

Designed drug-release systems having various breathable polyurethane film-backed hydrocolloid acrylated adhesive layers for moisture healing.

A series of designed drug-release systems were prepared and established for clear moisture healing. These systems were designed to have an interpenetrating polymer network (IPN) structure, which contained a breathable polyurethane film, hydrocolloidlayer, and polyacrylate adhesive layer. Breathable polyurethane film (2000 g/m(2)/24 hr) with high moisture permeability was employed as a base for new drug-release systems or wound dressings. All drug-release systems having a polyurethane film-backed hydrocolloid acrylated adhesive layer showed an increase of water uptakes with increasing time. After 114 hours, high water uptakes of drug-release systems with 20% hydrocolloid components were observed in the values of 160, 1100, and 1870% for different additional hydrocolloid components of carboxymethylcellulose, sodium alginate, and carbomer U10, respectively. New drug-release systems of polyurethane film-backed hydrocolloid/adhesive layers could be designed and established for wound care managements.

Designed hydrocolloid interpenetrating polymeric networks for clinical applications of novel drug-carrying matrix systems using Tris (6-isocyanatohexyl) isocyanurate and hydroxypropylmethylcellulose.

Hydroxypropyl methylcellulose (HPMC) was employed in this study to design controllable drug release systems because of its non-toxic nature, swelling properties. New interpenetrating polymer networks (IPN) of HPMC / tri-isocyanate crosslinked polyurethane (TCPU) could be prepared on the surfaces of IPN materials. To design "Novel Drug-carrying Matrix Systems", incorporation of novel structure is important to the possible formation of drug-carrying spaces within the material, which was achieved by using Tris (6-isocyanatohexyl) isocyanurate with three soft hexyl arms in this study. A series of novel drug-carrying matrix systems prepared by crosslinking reaction could be candidates for an excellent and smart potential material. When the polymeric networks were established on the surfaces of resulting materials, the developed hydrophilic interpenetrating polymeric structures of HPMC/ polyurethane could provide good wettability to the wound dressings, particularly for moisture healing application. The materials containing HPMC/polyurethane networks using 1% cross-linking agent showed a water uptake value of 5.1% after one hour, which has great potential for use as wound dressings for moisture healing. Furthermore, a new drug delivery system of hydrophilic IPN was successfully designed and established.

Rapid and sensitive controlled release monitoring method of biomedical combined products with IDM for pain management and cancer treatment.

New designed combined products and its determination for pain management and cancer treatment were studied. A rapid and sensitive stability indicating HPLC method had been developed and validated for the determination of Indomethacin(IDM) in a transdermal patch. This analytical method was successfully applied to the determination of Indomethacin in a transdermal patch and can be used for routine quality control analysis. Chromatographic separation was achieved isocratically on an Inertsil® C8-3 column utilizing a mobile phase of acetonitrile / 0.01 M monobasic sodium phosphate and 0.01 M dibasic sodium phosphate buffer (pH 3) (65:35, v/v) at the flow rate of 1 mL/min with UV detection at the wavelength of 210 nm. The system suitability was performed, and the result showed that Indomethacin(IDM) and its impurity were separated. The calibration curve of Indomethacin(IDM) was linear in the range of 0.1~15 ppm (r = 0.9989, n = 3).