Microfluidic Synthesis of miR-200c-3p Lipid Nanoparticles: Targeting ZEB2 to Alleviate Chondrocyte Damage in Osteoarthritis

Dong Zheng,1,* Tong Chen,2,* Kaiyuan Yang,1 Guangrong Yin,1 Yang Chen,3 Jianchao Gui,4 Chao Xu,1 Songwei Lv5 1Department of Orthopedics, The Affiliated Changzhou No.2 People’s Hospital with Nanjing Medical University, The Third Affiliated Hospital of Nanjing Medical University, Changzhou, People’s Republic of China; 2Department of Orthopedics, The First Affiliated Hospital of Nanjing Medical University, Nanjing, People’s Republic of China; 3Changzhou Productivity Development Center, Changzhou, People’s Republic of China; 4Department of Sports Medicine and Joint Surgery, Nanjing First Hospital, Nanjing Medical University, Nanjing, People’s Republic of China; 5School of Pharmacy, Changzhou University, Changzhou, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chao Xu; Songwei Lv, Email [email protected]; [email protected]: Osteoarthritis (OA) is a degenerative joint disease characterized by articular cartilage degeneration. Chondrocyte inflammation, apoptosis, and extracellular matrix degradation accelerated OA progression. MicroRNA (miRNA) has the potential to be a therapeutic method for osteoarthritis. However, it is difficult to penetrate the cell to exercise its biological function, and its extracellular effect is unclear.Methods: lipo-AgPEI-miR-200c-3p was created by combining miR-200c-3p with silver nitrate polyvinylimine nanoparticles on a microfluidic device. The drug release curve, stability, temperature sensitivity, cytotoxicity, and the impact of lipo-AgPEI-miR-200c-3p on the expression of proteins linked to matrix disintegration, apoptosis, and inflammatory factors were all detected.Results: Results showed that the particle size of Lipo-AgPEI-miR-200c-3p was about 130 nm, the Zeta potential was lowered to 1.08± 0.12 mV. Lipo-AgPEI-miR-200c-3p could increase cell viability, prevent cell apoptosis, and decrease the expression levels of TNF-α, IL-6, IL-1β, and MCP-1 in ADTC5 cells following LPS stimulation. MMP3, MMP13, and ADAMTS-4 expression was downregulated whereas collagen II expression was upregulated. The ZEB2 expression was greatly elevated following LPS stimulation and dramatically decreased following transfection of miR-200c-3p. Collagen II expression rose following transfection of si-ZEB2, whereas the expression levels of inflammatory factors, apoptosis-related proteins, MMP3, MMP13, and ADAMTS-4 decreased. The dual luciferase experiment demonstrated that ZEB2 was the target gene of miR-200c-3p.Conclusion: The synergistic effect of AgPEI and miR-200c-3p can inhibit the inflammatory response, apoptosis, and matrix degradation of chondrocytes. Lipo-AgPEI-miR-200c-3p can also improve transfection efficiency and obtain good physicochemical properties of drugs. miR-200c-3p may be crucial in the development of OA and can influence the target gene ZEB2, control the inflammatory response, apoptosis, and chondrocyte matrix breakdown.Keywords: microfluidic synthesis, miR-200c-3p, lipid nanoparticles, osteoarthritis, ZEB2