Chondrogenesis of mesenchymal stromal cells on the 3D printed polycaprolactone/fibrin/decellular cartilage matrix hybrid scaffolds in the presence of piascledine.

Nowadays, cartilage tissue engineering (CTE) is considered important due to lack of repair of cartilaginous lesions and the absence of appropriate methods for treatment. In this study, polycaprolactone (PCL) scaffolds were fabricated by three-dimensional (3D) printing and were then coated with fibrin (F) and acellular solubilized extracellular matrix (ECM). After extracting adipose-derived stem cells (ADSCs), 3D-printed scaffolds were characterized and compared to hydrogel groups. After inducing the chondrogenic differentiation in the presence of Piascledine and comparing it with TGF-ß3 for 28 days, the expression of genes involved in chondrogenesis (AGG, COLII) and the expression of the hypertrophic gene (COLX) were examined by real-time PCR. The expression of proteins COLII and COLX was also determined by immunohistochemistry. Glycosaminoglycan was measured by toluidine blue staining. 3D-printed scaffolds clearly improved cell proliferation, viability, water absorption and compressive strength compared to the hydrogel groups. Moreover, the use of compounds such as ECM and Piascledine in the process of ADSCs chondrogenesis induction increased cartilage-specific markers and decreased the hypertrophic marker compared to TGF-ß3. In Piascledine groups, the expression of COLL II protein, COLL II and Aggrecan genes, and the amount of glycosaminoglycan showed a significant increase in the PCL/F/ECM compared to the PCL and PCL/F groups.

Pomegranate seed extract enhances the inhibitory effect of adipose-derived mesenchymal stem cells on breast cancer cell line in co-culture conditions.

Background and purpose: Pomegranate seed extract (PSE) possesses anticancer activities and healing effects. Adipose-derived stem cells (ADSCs) are being considered a new candidate for cancer treatment. The purpose of this study was to investigate the effect of PSE on the cell cycle and apoptosis of the MCF-7 cell line in the co-culture condition with ADSCs. Experimental approach: MCF-7 and ADSC cells (ratio 1/1) were cultured in a transwell plate with and without PSE (PSE-co-culture and co-culture groups). MCF-7 cells were cultured in monolayer without and with PSE (mono-culture and PSE-mono-culture groups). MCF-7 cell line was harvested on day 5 and cell viability, apoptotic activity, cell cycle, and gene expression were evaluated. Findings / Results: The results of the MTT assay indicated that PSE at 100 µg/mL has the highest cytotoxicity on the MCF-7 in the PSE-co-culture group. The cell cycle analysis revealed that ADSCs in combination with PSE significantly increased the population of MCF-7 cells in the G1 phase, resulting in the arrest of MCF-7 cells cycle in the G0/G1 transition. In addition, the most apoptotic MCF-7 cells (41.5%) were detected in the same group. Expression of BAX and caspase3 genes were upregulated while anti-apoptotic (BCL-2) and angiogenesis inducer (VEGF) genes were downregulated in the PSE-co-culture group compared with the other groups. Conclusion and implications: ADSCs reduced cell viability and proliferation of MCF-7 cells in co-culture conditions and adding PSE to the medium increased the apoptosis of cancer cells. This study suggests that ADSCs with PSE can suppress tumor cells.

A Review on Antibacterial Biomaterials in Biomedical Applications: From Materials Perspective to Bioinks Design.

In tissue engineering, three-dimensional (3D) printing is an emerging approach to producing functioning tissue constructs to repair wounds and repair or replace sick tissue/organs. It allows for precise control of materials and other components in the tissue constructs in an automated way, potentially permitting great throughput production. An ink made using one or multiple biomaterials can be 3D printed into tissue constructs by the printing process; though promising in tissue engineering, the printed constructs have also been reported to have the ability to lead to the emergence of unforeseen illnesses and failure due to biomaterial-related infections. Numerous approaches and/or strategies have been developed to combat biomaterial-related infections, and among them, natural biomaterials, surface treatment of biomaterials, and incorporating inorganic agents have been widely employed for the construct fabrication by 3D printing. Despite various attempts to synthesize and/or optimize the inks for 3D printing, the incidence of infection in the implanted tissue constructs remains one of the most significant issues. For the first time, here we present an overview of inks with antibacterial properties for 3D printing, focusing on the principles and strategies to accomplish biomaterials with anti-infective properties, and the synthesis of metallic ion-containing ink, chitosan-containing inks, and other antibacterial inks. Related discussions regarding the mechanics of biofilm formation and antibacterial performance are also presented, along with future perspectives of the importance of developing printable inks.

Electrospun Ag-decorated reduced GO-graft-chitosan composite nanofibers with visible light photocatalytic activity for antibacterial performance.

The treatment of water contaminated by bacteria is becoming a necessity. The nanomaterials possessing both intrinsic antibacterial properties and photocatalytic activity are excellent candidates for water disinfection. The powdered form of nanomaterials can be aggregated while embedding the nanomaterials into the NFs can overcome the limitation and enhance the photocatalytic activity and transition from UV-light to visiblelight. Here, graphene oxide (GO) was synthesized, grafted to chitosan, and decorated with silver nanoparticles (Ag NPs) to produce Ag-decorated reduced GO-graft-Chitosan (AGC) NPs. The blends of polyacrylonitrile (PAN) and AGC NPs were prepared in various concentrations of 0.5 wt%, 1.0 wt%, 5.0 wt%, and 10.0 wt% and used to fabricate the electrospun composite NFs. FTIR/ATR, UV-Vis, Raman, XRD, and SEM/EDAX analyses confirmed the successful preparation of the NPs and NFs. The cytotoxicity and antibacterial activity of the composite NFs were received in the order of composite NFs 10.0 wt%˃ 5.0 wt%˃ 1.0 wt%˃ 0.5 wt% in both conditions with/without light irradiation. Their cytotoxicity and antibacterial activity were more under light irradiation compared to the dark. The composite NFs (5.0 wt%) were distinguished as the optimum NFs with cell viability of 80% within 24 h and 60% within 48 h on L929 cells and inhibition zone diameter (IZD) of 12 mm for E. coli and 13 mm for S. aureus after 24 h under the light irradiation. The optimum composite NFs showed thermal stability up to 180 °C and tensile strength of 1.11 MPa with 21.71% elongation at break.

Bioprinting Via a Dual-Gel Bioink Based on Poly(Vinyl Alcohol) and Solubilized Extracellular Matrix towards Cartilage Engineering.

Various hydrogel systems have been developed as biomaterial inks for bioprinting, including natural and synthetic polymers. However, the available biomaterial inks, which allow printability, cell viability, and user-defined customization, remains limited. Incorporation of biological extracellular matrix materials into tunable synthetic polymers can merge the benefits of both systems towards versatile materials for biofabrication. The aim of this study was to develop novel, cell compatible dual-component biomaterial inks and bioinks based on poly(vinyl alcohol) (PVA) and solubilized decellularized cartilage matrix (SDCM) hydrogels that can be utilized for cartilage bioprinting. In a first approach, PVA was modified with amine groups (PVA-A), and mixed with SDCM. The printability of the PVA-A/SDCM formulations cross-linked by genipin was evaluated. On the second approach, the PVA was functionalized with cis-5-norbornene-endo-2,3-dicarboxylic anhydride (PVA-Nb) to allow an ultrafast light-curing thiol-ene cross-linking. Comprehensive experiments were conducted to evaluate the influence of the SDCM ratio in mechanical properties, water uptake, swelling, cell viability, and printability of the PVA-based formulations. The studies performed with the PVA-A/SDCM formulations cross-linked by genipin showed printability, but poor shape retention due to slow cross-linking kinetics. On the other hand, the PVA-Nb/SDCM showed good printability. The results showed that incorporation of SDCM into PVA-Nb reduces the compression modulus, enhance cell viability, and bioprintability and modulate the swelling ratio of the resulted hydrogels. Results indicated that PVA-Nb hydrogels containing SDCM could be considered as versatile bioinks for cartilage bioprinting.

Comparison of TGF-ß3 and avocado/soybean unsaponifiable on chondrogenesis of human adipose-derived stem cells on poly (lactic-co-glycolic) acid/ hyaluronic acid hybrid scaffold.

OBJECTIVES: Avocado/soybean unsaponifible (ASU) possesses properties including chondroprotective, anticatabolic, and anabolic. The goal behind this research was to detect the effect of ASU and TGF-ß3 on the chondrogenesis of human adipose-derived stem cells (hADSCs) on poly (lactic-co-glycolic) acid (PLGA)/ hyaluronic acid (PLGA/HA) hybrid scaffold. MATERIALS AND METHODS: First hADSCs were seeded in PLGA/Hyaluronic acid scaffold and cultured in chondrogenic media. These cells were assigned into 4 groups: control, TGFß-3, ASU, and TGFß-3+ASU. The viability was assessed separately by MTT. Real-time PCR was used to quantify the expression of chondrogenic specific genes [Sox9, collagen type II (ColII), Aggrecan (AGG)] and collagen type X (ColX). Moreover, Western blotting was employed to evaluate protein expression levels of collagens type II and X. RESULTS: These findings indicated a significant increase in the proliferation and survival of hADSCs differentiated cells by ASU compared with the control group (P=0.008). Real-time PCR results revealed significant differences in the expression of AGG, SOX9, ColII, and ColX genes in the control group when compared with other groups (ASU, TGF-ß3, and TGF-ß3+ASU). ColII protein production significantly dropped in the TGF-ß3 group in comparison with the TGF-ß3+ASU group (0.000). The ColII (P=0.002) and ColX (P=0.002) protein production proved significantly higher in the TGF-ß3+ASU group compared with the ASU group. CONCLUSION: Using the synergist form TGFß-3, ASU induces chondrogenesis in hADSCs in PLGA/HA composite scaffold. This can be deduced with reduction of special markers of hyaline cartilage in comparison with ASU and decreased hypertrophic marker compared with TGF-ß3.

Evaluation Avocado Soybean Unsaponifiables Loaded in Poly (lactic-co-glycolic) Acid/Avocado Soybean Unsaponifiables-Fibrin Nanoparticles Scaffold (New Delivery System) is an Effective Factor for Tissue Engineering.

BACKGROUND: Growth factors and chemical stimulants have key role in cartilage tissue engineering, but these agents have unfavorable effects on cells. Avocado soybean unsaponifiables (ASU) has chondroprotective and anti-inflammatory effects. In this study, fibrin2nanoparticles (FNP)/ASU, as a new delivery system, with stem cells applied for cartilage tissue engineering in poly (lactic-co-glycolic) acid (PLGA) scaffold. MATERIALS AND METHODS: FNP/ASU prepared by freeze milling and freeze drying. NFP/ASU was characterized by dynamic light scattering (DLS). PLGA-NFP/ASU scaffold was fabricated and assessed by scanning electron microscope (SEM). Human adipose-derived stem cells (hADSCs) were seeded on scaffold and induced for chondrogenesis. After 14 days, cell viability and gene/protein expression evaluated. RESULTS: The results of DLS and SEM indicated that nanoparticles had high quality. The expression of type II collagen and SOX9 and aggrecan (ACAN) genes in differentiated cells in the presence of ASU was significantly increased compared with the control group (P and lt; 0.01), on the other hand, type I collagen expression was significantly decreased and western blot confirmed it. CONCLUSIONS: This study indicated FNP/ASU loaded in PLGA scaffold has excellent effect on chondrogenic differentiation of hADSCs and tissue engineering.

Herbal Remedies as Potential in Cartilage Tissue Engineering: An Overview of New Therapeutic Approaches and Strategies.

It is estimated that by 2023, approximately 20% of the population of Western Europe and North America will suffer from a degenerative joint disease commonly known as osteoarthritis (OA). During the development of OA, pro-inflammatory cytokines are one of the major causes that drive the production of inflammatory mediators and thus of matrix-degrading enzymes. OA is a challenging disease for doctors due to the limitation of the joint cartilage's capacity to repair itself. Though new treatment approaches, in particular with mesenchymal stem cells (MSCs) that integrate the tissue engineering (TE) of cartilage tissue, are promising, they are not only expensive but more often do not lead to the regeneration of joint cartilage. Therefore, there is an increasing need for novel, safe, and more effective alternatives to promote cartilage joint regeneration and TE. Indeed, naturally occurring phytochemical compounds (herbal remedies) have a great anti-inflammatory, anti-oxidant, and anabolic potential, and they have received much attention for the development of new therapeutic strategies for the treatment of inflammatory diseases, including the prevention of age-related OA and cartilage TE. This paper summarizes recent research on herbal remedies and their chondroinductive and chondroprotective effects on cartilage and progenitor cells, and it also emphasizes the possibilities that exist in this research area, especially with regard to the nutritional support of cartilage regeneration and TE, which may not benefit from non-steroidal anti-inflammatory drugs (NSAIDs).

The Effects of Fibrin-icariin Nanoparticle Loaded in Poly (lactic-co-glycolic) Acid Scaffold as a Localized Delivery System on Chondrogenesis of Human Adipose-derived Stem Cells.

BACKGROUND: Nowadays, cartilage tissue engineering is the best candidate for regeneration of cartilage defects. This study evaluates the effect of fibrin/icariin (ICA) nanoparticles (F/I NPs) on chondrogenesis of stem cells. MATERIALS AND METHODS: F/I NPs were characterized by Dynamic Light Scattering DLS. Poly (lactic-co-glycolic) acid (PLGA)-F/I NP scaffold was fabricated and assessed by scanning electron microscope. Human adipose-derived stem cells (hADSCs) were seeded on scaffold and induced for chondrogenesis. After 14 days, cell viability and gene expression were analyzed by the 3-(4, 5- dimethylthiazol-2yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. MTT assay and real-time polymerase chain reaction (RT-PCR). RESULTS: The size and surface charge of F/I NP were about 28-30 nm and - 17, respectively. The average of pore size of PLGA and PLGA-fibrin/ICA was 230 and 340 µm, respectively. Cell viability of differentiated cells in P/F group was higher than others significantly (P ≤ 0.05). Furthermore, quantitative RT-PCR analysis demonstrated that ICA upregulated cartilaginous-specific gene expression. Furthermore, the results of the expression of type I collagen revealed that ICA downregulated this gene significantly (P < 0.01). CONCLUSIONS: The results indicated that F/I NP could be a potential factor for chondrogenesis of stem cells and downregulation of fibrocartilage marker.

Fabrication of chitosan/agarose scaffolds containing extracellular matrix for tissue engineering applications.

One of the most effective approaches for treatment of cartilage involves the use of porous three-dimensional scaffolds, which are useful for improving not only cellular adhesion but also mechanical properties of the treated tissues. In this study, we manufactured a composite scaffold with optimum properties to imitate nasal cartilage attributes. Cartilage extracellular matrix (ECM) was used in order to improve the cellular properties of the scaffolds; while, chitosan and agarose were main materials that are used to boost the mechanical and rheological properties of the scaffolds. Furthermore, we explored the effect of the various weight ratios of chitosan, agarose, and ECM on the mechanical and biomedical properties of the composite scaffolds using the Taguchi method. The resulting composites display a range of advantages, including good mechanical strength, porous morphology, partial crystallinity, high swelling ratio, controlled biodegradability rate, and rheological characteristics. Additionally, we performed the cytotoxicity tests to confirm the improvement of the structure and better cell attachments on the scaffolds. Our findings illustrate that the presence of the ECM in chitosan/agarose structure improves the biomedical characteristics of the final scaffold. In addition, we were able to control the mechanical properties and microstructure of the scaffolds by optimizing the polymers' concentration and their resulting interactions. These results present a novel scaffold with simultaneously enhanced mechanical and cellular attributes comparing to the scaffolds without ECM for nasal cartilage tissue engineering applications.

Conductive hydrogel based on chitosan-aniline pentamer/gelatin/agarose significantly promoted motor neuron-like cells differentiation of human olfactory ecto-mesenchymal stem cells.

Developing a simple produces for efficient derivation of motor neurons (MNs) is essential for neural tissue engineering studies. Stem cells with high capacity for neural differentiation and scaffolds with the potential to promote motor neurons differentiation are promising candidates for neural tissue engineering. Recently, human olfactory ecto-mesenchymal stem cells (OE-MSCs), which are isolated easily from the olfactory mucosa, are considered a new hope for neuronal replacement due to their neural crest origin. Herein, we synthesized conducting hydrogels using different concentration of chitosan-g-aniline pentamer, gelatin, and agarose. The chemical structures, swelling and deswelling ratio, ionic conductivity and thermal properties of the hydrogel were characterized. Scaffolds with 10% chitosan-g-aniline pentamer/gelatin (S10) were chosen for further investigation and the potential of OE-MSCs as a new source for programming to motor neuron-like cells investigated on tissue culture plate (TCP) and conductive hydrogels. Cell differentiation was evaluated at the level of mRNA and protein synthesis and indicated that conductive hydrogels significantly increased the markers related to motor neurons including Hb-9, Islet-1 and ChAT compared to TCP. Taken together, the results suggest that OE-MSCs would be successfully differentiated into motor neuron-like cells on conductive hydrogels and would have a promising potential for treating motor neuron-related diseases.

Hybrid and Composite Scaffolds Based on Extracellular Matrices for Cartilage Tissue Engineering.

IMPACT STATEMENT: Scaffolds fabricated from extracellular matrix (ECM) derivatives are composed of conducive structures for cell attachment, proliferation, and differentiation, but generally do not have proper mechanical properties and load-bearing capacity. In contrast, scaffolds based on synthetic biomaterials demonstrate appropriate mechanical strength, but the absence of desirable biological properties is one of their main disadvantages. To integrate mechanical strength and biological cues, these ECM derivatives can be conjugated with synthetic biomaterials. Hence, hybrid scaffolds comprising both advantages of synthetic polymers and ECM derivatives can be considered a robust vehicle for tissue engineering applications.

Plastination of decalcified bone by a new resin technique.

BACKGROUND: The scope of this study was to preserve whole detailed structure of dissected and decalcified bones, taken from used cadavers, by a new plastination technique. MATERIALS AND METHODS: Specimens we used in this study were sheep femurs and human bones including pelvis, femur, tibia, and fibula. Bones, at first, fixed with 5% formalin and were decalcified with 5% nitric acid, and then were fixed again and washed under the tap water. The resulted flexible bones were dehydrated in -25°C acetone and degreased them in +25°C acetone. Then, the experimental and control specimen were placed in the vacuum chamber for forced impregnation with our new flexible unsaturated polyester resin (UP89 method) and silicon resin (S10 method), respectively. Finally, the strength and flexibility of plastinated decalcified specimens were investigated by tensometer, and the weight diversity was measured by digital balance. RESULTS: Plastinated bones prepared by this technique were found to be dried, non-fragile, durable, odorless, non-greasy, and demonstrating all detailed structures of the bones. Tensile and weight tests results indicated that plastinated decalcified femurs have owned higher flexibility and strength but lesser weight than plastinated undecalcified femurs. The characteristics of both experimental and control groups of plastinated decalcified specimens were found to have no significant difference. CONCLUSIONS: Our synthesized resin found to be much more economical than conventional plastination method. In more details, properties of these new products were the same as, S10 method, from points of strength, flexibility and weight, but, since the money cost for producing them was about one fifth that of S10 method.