Copper/Tin Nanocomposites-Loaded Exosomes Induce Apoptosis and Cell Cycle Arrest at G0/G1 Phase in Skin Cancer Cell Line.

This study aims to explore the efficacy of Copper/Tin (CuS/SnS) nanocomposites loaded into exosomes against skin cancer A431 cell line. CuS/SnS nanocomposites (S1, S2, S3) were synthesized and characterized, then loaded into exosomes (Exo) (S1-Exo, S2-Exo and S3-Exo) and characterized. After that, the loaded samples were investigated in vitro against A431 using cytotoxicity, apoptosis, and cell cycle assays. CuS/SnS nanocomposites were indexed to hexagonal CuS structure and orthorhombic α-SnS phase and showed nano-rode shape. The exosomes loaded with nanocomposites were regular and rounded within the size of 120 nm, with no signs of broken exosomes or leakage of their contents. The cytotoxicity assay indicated the enhanced cytotoxic of S1-Exo versus the free nano-form S1 on A431. Interestingly, S1-Exo recorded 1.109 times more than DOX in its anti-skin cancer capacity. Moreover, S1-Exo recorded 40.2% for early apoptosis and 22.1% for late apoptosis. Furthermore, it displayed impact in arresting the cancer cell cycle at G0/G1 phase and reducing G2/M phase. Noteworthy, loaded nanocomposites were safe against normal HSF skin cells. In conclusion, the loaded CuS/SnS nanocomposites into the exosomes could be of great potential as anti-skin cancer candidates through induction of apoptosis and promotion of the cell cycle arrest at G0/G1 phase.

Stem cells derived exosomes as biological nano carriers for VCR sulfate for treating breast cancer stem cells.

Due to vincristine sulfate's (VCR sulfate) toxicity and non-specific targeting, which might adversely damage healthy cells, its clinical application is restricted. In this study, we loaded VCR sulfate on exosomes generated from mesenchymal stem cells (MSCs) to enhance its targeted distribution. Exosomes are able to deliver molecules to specific cells and tissues and have therapeutic potential. In this study, we isolated exosomes from MSCs, and using probe-sonication approach loaded them with VCR sulfate. Using SRB assay, the cytotoxicity of VCR sulfate-Exo was assessed in T47D breast cancer cells, and the results were contrasted with those of free VCR sulfate. Then We labeled markers (CD44+/CD24-) in the cell line to assess the targeting effectiveness of VCR sulfate-Exo using flow cytometry. Our results showed that the cytotoxicity of VCR sulfate-Exo was nearly the same as that of VCR sulfate. Flow cytometry analysis revealed that VRC sulfate-Exo was more effectively targeted to MSCs than free VCR sulfate. Our study shows that loading VCR sulfate to MSCs-derived exosomes can improve their targeted delivery and lessen their side effects. Additional research is required to determine VCR sulfate-Exo's in vivo effectiveness and safety and improve the loading and delivery strategies.

Comparison between stem cell therapy and stem cell derived exosomes on induced multiple sclerosis in dogs.

BACKGROUND: Multiple sclerosis (MS) is a chronic condition that primarily manifests as demyelination of neuronal axons in the central nervous system, due to the loss or attack of oligodendroglia cells that form myelin. Stem cell therapy has shown promising results for the treatment of MS due to its capability to halt the immune attack, stop apoptosis and axonal degeneration, and differentiate into oligodendrocytes. Stem cell-derived Exosomes (Exosomes) have shown great capabilities for neuronal diseases as they have growth factors, complex sets of miRNA, enzymes, proteins, major peptides, lipids, and macromolecules with anti-inflammatory, angiogenesis, and neurogenesis activities. METHODS: This study aimed to compare the healing properties of stem cells, against Exosomes for the treatment of an experimentally induced MS dog model. Dog models of MS received either a single treatment of stem cells or a single treatment of Exosomes intrathecally and the treatment process was evaluated clinically, radiologically, histopathologically, and electron microscopy and cerebrospinal fluid analysis. RESULTS: showed marked amelioration of the clinical signs in both treated groups compared to the control one, magnetic resonance scans showed the resolution of the hyperintense lesions at the end of the study period, the histopathology and electron microscopy showed marked healing properties and remyelination in treated groups with superiority of the stem cells compared to Exosomes. CONCLUSIONS: Although stem cell results were superior to Exosomes therapy; Exosomes have proven to be effective and safe important actors in myelin regeneration, and their use in diseases like MS helps to stimulate remyelination.

Effect of combined intrathecal/intravenous injection of bone marrow derived stromal cells in platelet-rich plasma on spinal cord injury in companion animals.

Background: Companion animals are prone to spinal cord injuries commonly associated with severe locomotor and sensory complications, which can escalate to a state of irreversible paralysis. Stem cell therapies propose a hope for treating spinal cord injuries via differentiation into neurons and associated glial cells, halting the immune attacks, inhibiting apoptosis and necrosis, and secretion of neurotrophic factors that stimulate the regeneration process. Aim: The study aims to evaluate the use of autologous bone marrow derived stromal cells in platelet-rich plasma carrier for selected clinical cases having chronic spinal cord injuries in dogs and cats via a one-time combined intrathecal/intravenous injection. Methods: Cells were injected in five dogs and three cats suffering from disc protrusion leading to spinal cord injury and in thosewho did not respond to conventional treatment during a clinical trial. Results: Results indicated that the transplanted cells led to the restoration of the weight bearing locomotor function and spinal reflexes in a period less than 90 days with physical rehabilitation. The treatment showed minor changes in the magnetic resonance images of extruded discs. Conclusion: This study concluded that the combined intrathecal/intravenous injection of bone marrow stromal cells is a safe and promising procedure for treating chronic spinal cord injuries in companion animals.

Proficiency of Carboxymethylcellulose as a Cryoprotectant. Clinical and Histological Evaluation of Cryopreserved Heterogenous Mesenchymal Stem Cell-Exosomal Hydrogel on Critical Size Skin Wounds in Dogs.

Background: Fresh stem cell exosomes are usually obtained and reused in the same individual. It cannot be kept viable for a long period of time regardless of the lengthy preparation time. Freezing is typically used to preserve the viability of perishable materials and increase their lifetime. Regrettably, normal freezing of biomaterials leads to cell damage. Therefore, a cryoprotectant can save the cells from the conventional cryodamage. Sodium carboxymethylcellulose (NA-CMC) is a powdery substance that is used to manufacture bio-safe hydrofilm gels because of its high viscosity, cytocompatibility, and nonallergenic nature. Materials and Methods: Sterile CMC hydrogel was prepared, part of which was loaded with exosomal solution derived from MSCs. The gel was kept at -20°C for preservation. Two bilateral full-thickness circular skin wounds of 2-cm diameter were created on the back of experimental dogs. The wounds were at least 2.5 cm apart. Treatment started 24 hours after wound creation. Group I received CMC gel solely, whereas group II received frozen CMC exosomal gel. The gel was applied 4 times, a single application per day with 1- day interval. Results: Clinically, the frozen exosomal gel significantly promoted wound healing with no scaring. Histologically, enhanced dermal fibroblasts and organized collagen deposition were seen in the treated group. Conclusion: CMC proved to be an efficient cryoprotectant and a suitable vehicle for exosomes. Deep freezing was proven to conserve the viability, extended the preservation, and facilitated the usage of exosomal gel. This technique of preserved cell-free therapy is inexpensive, time-saving, and proficient and seems suitable for treating cutaneous wounds.

Preservation techniques of stem cells extracellular vesicles: a gate for manufacturing of clinical grade therapeutic extracellular vesicles and long-term clinical trials.

Extracellular vesicles (EVs) are nanosized vesicles released by different cells and have been separated from most of the body fluids. These vesicles play a central role in cell-to-cell communications as carry a distinct cargo including proteins, RNA species, DNAs, and lipids that are meant to be shipped and exchanged between cells at both systemic and paracrine levels. They serve in regulating normal physiological processes. EVs released from stem cells exert similar therapeutic effect to their originating cells. Clinical application of EVs requires the preparation of sufficient and viable active therapeutic EVs as well as implementing suitable methods for long-term preservation to expedite both their clinical and commercial uses. Cryopreservation is the most common method used to preserve decomposable biomaterials. However, cryopreservation causes cryoinjury to cells which therefore necessitate the use of cryoprotectants. Two types of cryoprotectants exist: penetrating and non-penetrating. In freeze drying, the watery content is sublimed from the product after it is frozen. This drying process is pertinent to thermo-liable substances and those unstable in aqueous solutions for prolonged storage periods. In spray drying technique, the solution containing EVs is firstly atomized, then droplets are rapidly converted into a dry powder using heated gas. Even with the exposure to high temperatures of the drying gas, spray drying is considered suitable for heat-sensitive materials. EVs are considered a promising cell-free therapy, but the lack of proper preservation limits its benefits. Preservation of EVs will initiate a vast amount of clinical trials on different species and different clinical problems.

Evaluation of treatment of experimentally induced canine model of multiple sclerosis using laser activated non-expanded adipose derived stem cells.

Multiple sclerosis (MS) is a progressive demyelinating disease of the central nervous system that destroys oligodendrocytes. This work aims to evaluate the treatment of experimentally induced MS in dogs using laser activated non-expanded adipose derived stem cells. The results showed amelioration of the clinical signs over time confirmed by the resolution of the previous lesions on MRI. Positive migration of the injected cells to the site of lesion, increased remyelination detected by Myelin Basic Proteins, positive differentiation into Olig2 positive oligodendrocytes, prevented the glial scar formation and restored axonal architecture. The study concluded that treatment using laser activated stem cells holds a promising therapeutic option for treatment of MS in a canine model.

Histological Evaluation of Experimentally Induced Critical Size Defect Skin Wounds Using Exosomal Solution of Mesenchymal Stem Cells Derived Microvesicles.

BACKGROUND AND OBJECTIVES: The present study investigated whether MSCs derived microvesicles (MVs) or (Exosomes) can exert therapeutic effects on an experimental model of cutaneous injury and explored the underlying involving mechanisms. METHODS AND RESULTS: Three bilateral full thickness circular wounds were created on the back of two groups of dogs using 2-cm dermal punch. The wounds were at least 2.5 cm apart. Saline was subcutaneously injected in 4 places around each wound area in group-I (control), whereas an equal volume of exosomal solution of MSCs derived MVs was similarly injected in group-II. The findings demonstrated that MSCs derived MVs had significantly promoted cutaneous wound healing, collagen synthesis, and vascularization at wound sites. The application of the exosomal solution had not only promoted the generation of newly formed vessels, but also have accelerated their development and maturation leading to a faster healing process. CONCLUSIONS: MSC-Exosomes appeared to be a superior candidate for treating cutaneous wounds than their originator cells, and may represent a promising opportunity to develop a novel cell-free therapy approach that might overcome the obstacles and risks associated with the use of native or engineered stem cells transplantation therapy.