Alginate hydrogel-PCL/gelatin nanofibers composite scaffold containing mesenchymal stem cells-derived exosomes sustain release for regeneration of tympanic membrane perforation.

Exosomes are among the most effective therapeutic tools for tissue engineering. This study demonstrates that a 3D composite scaffold containing exosomes can promote regeneration in rat tympanic membrane perforation (TMP). The scaffolds were characterized using scanning electron microscopy (SEM), degradation, PBS adsorption, swelling, porosity, and mechanical properties. To confirm the isolation of exosomes from human adipose-derived mesenchymal stem cells (hAMSCs), western blot, SEM, and dynamic light scattering (DLS) were performed. The Western blot test confirmed the presence of exosomal surface markers CD9, CD81, and CD63. The SEM test revealed that the isolated exosomes had a spherical shape, while the DLS test indicated an average diameter of 82.5 nm for these spherical particles. MTT assays were conducted to optimize the concentration of hAMSCs-exosomes in the hydrogel layer of the composite. Exosomes were extracted on days 3 and 7 from an alginate hydrogel containing 100 and 200 µg/mL of exosomes, with 100 µg/mL identified as the optimal value. The optimized composite scaffold demonstrated improved growth and migration of fibroblast cells. Animal studies showed complete tympanic membrane regeneration (TM) after five days. These results illustrate that a scaffold containing hAMSC-exosomes can serve as an appropriate tissue-engineered scaffold for enhancing TM regeneration.

Fabrication and optimization of multilayered composite scaffold made of sulfated alginate-based nanofiber/decellularized Wharton's jelly ECM for tympanic membrane tissue engineering.

In this study, we fabricated a novel multilayer polyvinyl alcohol (PVA)/alginate sulfate (ALG-S) nanofiber/decellularized Wharton's Jelly ECM (d-ECM) composite for tympanic membrane perforations (TMPs) tissue engineering (TE). Initially, electrospun PVA/ALG-S scaffolds with different blend ratios were fabricated. The influence of ALG-S ratio on surface morphology, mechanical, physical and biological properties of the nanofibers was studied. Secondly, 3-layer composites were developed as a combination of PVA/ALG-S nanofibers and d-ECM to take synergic advantages of electrospun mats and d-ECM. As part of the evaluation of the effects of d-ECM incorporation, the composite's mechanical properties, in vitro degradation, swelling ratio, and biological activities were assessed. The MTT assay showed that PVA/ALG-S nanofibers with 50:50 ratio provided a more desirable environment to support cell growth. A composite containing 25 mg/cm2 d-ECM was determined as the optimal composite through MTT assay, and this composite was used for animal studies inducing TMP regeneration. According to the in vivo studies, the optimal composite not only stimulated the healing of TMPs but also shortened the healing period. These results suggest that a multilayer nanofiber/hydrogel composite could be a potential platform for regenerating TMPs.

Alginate sulfate/ECM composite hydrogel containing electrospun nanofiber with encapsulated human adipose-derived stem cells for cartilage tissue engineering.

Stem cell therapy is a promising strategy for cartilage tissue engineering, and cell transplantation using polymeric scaffolds has recently gained attention. Herein, we encapsulated human adipose-derived stem cells (hASCs) within the alginate sulfate hydrogel and then added them to polycaprolactone/gelatin electrospun nanofibers and extracellular matrix (ECM) powders to mimic the cartilage structure and characteristic. The composite hydrogel scaffolds were developed to evaluate the relevant factors and conditions in mechanical properties, cell proliferation, and differentiation to enhance cartilage regeneration. For this purpose, different concentrations (1-5 % w/v) of ECM powder were initially loaded within an alginate sulfate solution to optimize the best composition for encapsulated hASCs viability. Adding 4 % w/v of ECM resulted in optimal mechanical and rheological properties and better cell viability. In the next step, electrospun nanofibrous layers were added to the alginate sulfate/ECM composite to prepare different layered hydrogel-nanofiber (2, 3, and 5-layer) structures with the ability to mimic the cartilage structure and function. The 3-layer structure was selected as the optimum layered composite scaffold, considering cell viability, mechanical properties, swelling, and biodegradation behavior; moreover, the chondrogenesis potential was assessed, and the results showed promising features for cartilage tissue engineering application.

An additive manufacturing-based 3D printed poly ɛ-caprolactone/alginate sulfate/extracellular matrix construct for nasal cartilage regeneration.

Various composite scaffolds with different fabrication techniques have been applied in cartilage tissue engineering. In this study, poly ɛ-caprolactone (PCL) was printed by fused deposition modeling method, and the prepared scaffold was filled with Alginate (Alg): Alginate-Sulfate (Alg-Sul) hydrogel to provide a better biomimetic environment and emulate the structure of glycosaminoglycans properly. Furthermore, to enhance chondrogenesis, different concentrations of decellularized extracellular matrix (dECM) were added to the hydrogel. For cellular analyses, the adipose-derived mesenchymal stem cells were seeded on the hydrogel and the results of MTT assay, live/dead staining, and SEM images revealed that the scaffold with 1% dECM had better viscosity, cell viability, and proliferation. The study was conducted on the optimized scaffold (1% dECM) to determine mechanical characteristics, chondrogenic differentiation, and results demonstrated that the scaffold showed mechanical similarity to the native nasal cartilage tissue along with possessing appropriate biochemical features, which makes this new formulation based on PCL/dECM/Alg:Alg-Sul a promising candidate for further in-vivo studies.

Can Tissue Engineering Bring Hope to the Development of Human Tympanic Membrane?

The tympanic membrane (TM), more commonly known as the eardrum, consists of a thin layer of tissue in the human ear that receives sound vibrations from outside of the body and transmits them to the auditory ossicles. The TM perforations (TMPs) are a common ontological condition, which in some cases can result in permanent hearing loss. Despite the spontaneous healing capacity of the TM to regenerate in the majority of cases of acute perforation, chronic perforations require surgical interventions. However, the disadvantages of the surgical procedure include infection, anesthetic risks, and high failure of graft patency. The tissue engineering strategy, which includes the applications of a three-dimensional (3D) scaffold, cells, and biomolecules or a combination of them for the closure of chronic perforation, has been considered as an emerging treatment. Using this approach, emerging products are currently under development to regenerate the TM structure and its properties. This research aimed to highlight the problems with the current methods of TMP treatment, and critically evaluate the tissue engineering approaches, which may overcome these drawbacks. The focus of this review is on recent literature to critically discuss the emerging advanced materials used as a 3D scaffold in the development of a TM with cellular engineering, biomolecules, cells, and the fabrications of the TM and its pathway to the clinical application. In this review, we discuss the properties of TM and the advantages and disadvantages of the current clinical products for repair and replacement of the TM. Furthermore, we provide an overview of the in vitro and preclinical studies of emerging products over the past 5 years. The results of recent preclinical studies suggest that the tissue engineering field holds significant promise.