Congenital microtia patients: the genetically engineered exosomes released from porous gelatin methacryloyl hydrogel for downstream small RNA profiling, functional modulation of microtia chondrocytes and tissue-engineered ear cartilage regeneration

Abstract

Background Mesenchymal stem cells (MSCs) exosomes were previously shown to be effective in articular cartilage repairing. However, whether MSCs exosomes promote mature cartilage formation of microtia chondrocytes and the underlying mechanism of action remains unknown. Additionally, some hurdles, such as the low yield and unsatisfactory therapeutic effects of natural exosomes have emerged when considering the translation of exosomes-therapeutics to clinical practices or industrial production. Herein, we investigated the roles of human adipose-derived stem cells (ADSCs) exosomes in modulating microtia chondrocytes and the underlying mechanism of action. Special attention was also paid to the mass production and functional modification of ADSCs exosomes. Results We firstly used porous gelatin methacryloyl (Porous Gelma) hydrogel with pores size of 100 to 200 μm for 3D culture of passage 2, 4 and 6 ADSCs (P2, P4 and P6 ADSCs, respectively), and obtained their corresponding exosomes (Exo 2, Exo 4 and Exo 6, respectively). In vitro results showed Exo 2 outperformed both Exo 4 and Exo 6 in enhancing cell proliferation and attenuating apoptosis. However, both Exo 4 and Exo 6 promoted chondrogenesis more than Exo 2 did. Small RNA sequencing results indicated Exo 4 was similar to Exo 6 in small RNA profiles and consistently upregulated PI3K/AKT/mTOR signaling pathway. Notably, we found hsa-miR-23a-3p was highly expressed in Exo 4 and Exo 6 compared to Exo 2, and they modulated microtia chondrocytes by transferring hsa-miR-23a-3p to suppress PTEN expression, and consequently to activate PI3K/AKT/mTOR signaling pathway. Then, we designed genetically engineered exosomes by directly transfecting agomir-23a-3p into parent P4 ADSCs and isolated hsa-miR-23a-3p-rich exosomes for optimizing favorable effects on cell viability and new cartilage formation. Subsequently, we applied the engineered exosomes to in vitro and in vivo tissue-engineered cartilage culture and consistently found that the engineered exosomes could enhance cell proliferation, attenuate apoptosis and promote cartilage regeneration. Conclusions Taken together, the porous Gelma hydrogel could be applied to exosomes mass production, and functional modification could be achieved by selecting P4 ADSCs as parent cells and genetically modifying ADSCs. Our engineered exosomes are a promising candidate for tissue-engineered ear cartilage regeneration. Graphical Abstract

Huc-MSC-derived exosomes modified with the targeting peptide of aHSCs for liver fibrosis therapy

“…Accumulating evidences has shown that exosomes are emerging as promising natural nanovectors, given their unique properties, including low immunogenicity, high biocompatibility, and low toxicity [ 32 , 33 ]. Moreover, MSC-derived exosomes are believed to be a potential therapeutic tool for liver fibrosis [ 34 ].…”

Hydrogel-exosome system in tissue engineering: A promising therapeutic strategy

“…The use of a hydrogel-exosome system to fill bone defect areas has represented a novel therapeutic approach for its adaptability. On the one hand, appropriate pretreatment of exosome donor cells, such as 3D cultivation, lentiviral engineering, overexpression or knockdown of function-related miRNAs, and hypoxic stimulation, can enhance the exosome's production, targeting capability, and functions related to bone regeneration [ [263] , [264] , [265] , [266] ]. On the other hand, various biomaterials can produce distinct chemical signals, influencing cell communication and cellular microenvironment, thereby regulating exosome secretion [ 267 ].…”

Maximizing Oxaliplatin's Impact on EGFR+ Colorectal Cancer Through Targeted Extracellular Vesicles

“…In breast cancer treatment, the targeting of EVs using anti-human CD3 and anti-human HER2 antibodies has been employed to generate SMART-Exos that dualtarget T cells to CD3 and HER2 receptors associated with breast cancer. This approach enhances the e cacy of targeting of HER2-positive breast cancer cells by T cells [8][9][10][11][12][13]. In lung cancer treatment, ligands that bind to the epidermal growth factor receptor (EGFR) can be modi ed to attach to the surface of EVs to increase their binding and uptake by EGFR-positive tumor cells.…”