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.

The Comparative Effects of Schwann Cells and Wharton's Jelly Mesenchymal Stem Cells on the AIM2 Inflammasome Activity in an Experimental Model of Spinal Cord Injury.

Inflammasome activation and the consequent release of pro-inflammatory cytokines play a crucial role in the development of sensory/motor deficits following spinal cord injury (SCI). Immunomodulatory activities are exhibited by Schwann cells (SCs) and Wharton's jelly mesenchymal stem cells (WJ-MSCs). In this study, we aimed to compare the effectiveness of these two cell sources in modulating the absent in melanoma 2 (AIM2) inflammasome complex in rats with SCI. The Basso, Beattie, Bresnahan (BBB) test, Nissl staining, and Luxol fast blue (LFB) staining were performed to evaluate locomotor function, neuronal survival, and myelination, respectively. Real-time polymerase chain reaction (RT-PCR), Western blotting, and enzyme-linked immunosorbent assay (ELISA) were employed to analyze the gene and protein expressions of inflammasome components, including AIM2, ASC, caspase-1, interleukin-1ß (IL-1ß), and IL-18. Both gene and protein expressions of all evaluated factors were decreased after SC or WJ-MSC treatment, with a more pronounced effect observed in the SCs group (P < 0.05). Additionally, SCs promoted neuronal survival and myelination. Moreover, the administration of 3 × 105 cells resulted in motor recovery improvement in both treatment groups (P < 0.05). Although not statistically significant, these effects were more prominent in the SC-treated animals. In conclusion, SC therapy demonstrated greater efficacy in targeting AIM2 inflammasome activation and the associated inflammatory pathway in SCI experiments compared to WJ-MSCs.

Differentiation of human endometrial stem cells encapsulated in alginate hydrogel into oocyte-like cells.

Introduction: Human endometrial mesenchymal stem cells (hEnMSCs) are a rich source of mesenchymal stem cells (MSCs) with multi-lineage differentiation potential, making them an intriguing tool in regenerative medicine, particularly for the treatment of reproductive and infertility issues. The specific process of germline cell-derived stem cell differentiation remains unknown, the aim is to study novel ways to achieve an effective differentiation method that produces adequate and functioning human gamete cells. Methods: We adjusted the optimum retinoic acid (RA) concentration for enhancement of germ cell-derived hEnSCs generation in 2D cell culture after 7 days in this study. Subsequently, we developed a suitable oocyte-like cell induction media including RA and bone morphogenetic protein 4 (BMP4), and studied their effects on oocyte-like cell differentiation in 2D and 3D cell culture media utilizing cells encapsulated in alginate hydrogel. Results: Our results from microscopy analysis, real-time PCR, and immunofluorescence tests revealed that 10 µM RA concentration was the optimal dose for inducing germ-like cells after 7 days. We examined the alginate hydrogel structural characteristics and integrity by rheology analysis and SEM microscope. We also demonstrated encapsulated cell viability and adhesion in the manufactured hydrogel. We propose that in 3D cell cultures in alginate hydrogel, an induction medium containing 10 µM RA and 50 ng/mL BMP4 can enhance hEnSC differentiation into oocyte-like cells. Conclusion: The production of oocyte-like cells using 3D alginate hydrogel may be viable in vitro approach for replacing gonad tissues and cells.

The effect of Scrophularia striata on cell attachment and biocompatibility of decellularized bovine pericardia.

Since using tissue transplantation has faced limitations all over the world, regenerative medicine has introduced decellularized tissues as natural scaffolds and researchers are trying to improve their efficiency and function. In this study, to increase cell attachment and ultimately cell proliferation on decellularized bovine pericardia, scrophularia striata extract was used. Scrophularia striata is an Iranian traditional medicinal plant. For this aim after decellularization of bovine pericardium and analysis of its morphology, it was incubated in scrophularia striata solution. Next, isolated human adipose-derived mesenchymal stem cells were cultured on the tissue. Finally, MTT assay, nitric oxide assay, and scanning electron microscopy observation were performed. MTT showed an increase in cell survival after treating the tissue with the plant extract after 48 h in a dose dependent manner significantly. The survival of cells in 0.5%, 2.5%, and 5% groups was about 5, 10 and 15 folds higher in comparison to control groups, respectively. Additionally, nitric oxide secretion in 2.5% and 5% samples was three and five folds higher than that in control group, respectively. Moreover, SEM observation indicated an impressive and dose-dependent effect of using Scrophularia striata on tissue biocompatibility. The results of this study showed that using Scrophularia striata increased cell viability and cell attachment on decellularized pericardia which could pave the way for the use of natural extracts of medicinal plants to reduce unwanted effects and make desired changes in decellularized tissues.

Tracking the Transplanted Neurosphere in Retinal Pigment Epithelium Degeneration Model.

INTRODUCTION: Retinal Pigment Epithelium (RPE) layer deterioration is a leading cause of Age-Related Macular Degeneration (AMD), i.e., the most significant reason for irreversible blindness. The present study aimed to track the Neurosphere-Derived (NS) from Bone Marrow Stromal Stem Cells (BMSCs) grafted into the sub-retinal space (destruction of the RPE layer by sodium iodate). METHODS: RPE degeneration model was performed using the injection of 5% sodium iodate performed in the retro-orbital sinus of Wistar rats. BMSCs were extracted from the examined rat femur and induced into NS, using EGF, bFGF, and B27. BrdU-NS labeled cells were transplanted into the sub-retinal space. For detecting BMSCs and NS markers, immunocytochemistry was performed. Moreover, immunohistochemical was conducted for tracking the transplanted cells in the RPE and sensory retina. RESULTS: The immunocytochemistry of BMSCs cells displayed the expression of mesenchymal stem cells markers (CD90; 99%±1), CD166 (98%±2), CD44 (99%±1). Additionally, the expression of neural lineage markers in NS, such as SOX2, OCT4, Nanog, Nestin, and Neurofilaments (68, 160, 200) revealed the differentiation from BMSCs. Tracking BrdU-NS labeled suggested these aggregations in most layers of the retina. CONCLUSION: Our study data indicated that BMSCs derived neurosphere had the potential to migrate in injured retinal and integrate into the neurosensory retina. These data can be useful in finding safe cells for replacement therapy in AMD.

Exosome loaded alginate hydrogel promotes tissue regeneration in full-thickness skin wounds: An in vivo study.

Wound healing is known as one of the most complicated biological processes for injured skin caused by surgical, trauma, burns, or diabetic diseases, which causes a nonfunctioning mass of fibrotic tissue. Recent reports have suggested that exosomes (EXOs) secreted by this type of stem cells may contribute to their paracrine effect. In this study, the EXOs were isolated from the supernatant of cultured adipose-derived stem cells (ADSCs) via ultracentrifugation and filtration. The EXO loaded in the alginate-based hydrogel was used as a bioactive scaffold to preserve the EXO in the wound site in the animal model. The physical and biochemical properties of EXO loaded Alg hydrogel were characterized and results proved that fabricated structure was biodegradable and biocompatible. This bioactive wound dressing technique has significantly improved wound closure, collagen synthesis, and vessel formation in the wound area. Results offer a new viewpoint and a cell-free therapeutic strategy, for wound healing through the application of the composite structure of EXO encapsulated in alginate hydrogel.