Poster N19 - Dose-Dependency of Mesenchymal Stem Cell Extracellular Matrix Nanoparticles with Antimicrobial Effects

Introduction: Acute Respiratory Distress Syndrome (ARDS) is a prevalent pulmonary disorder with high morbidity rates in critically conditioned patients [1]. The pathogenesis of ARDS is largely due to lung infections which damage alveoli as fluid leaks into air sacs. These infections are the highest risk factors for ARDS patients, with sepsis being the most common cause [2], and the inflammation associated with injured lungs often leads to worsened outcomes in critically ill or infected patients. Current pharmacological treatments for ARDS also lack effectiveness [3] and without a cure, these symptoms can only be mitigated with mechanical ventilation that also disrupts lung tissue. This ventilation further increases morbidity as ARDS patients develop ventilator-associated pneumonia (VAP); drug-resistant pathogens such as Escherichia coli (E.coli) and Pseudomonas aeruginosa (P.aeruginosa) often cause these bacterial infections [4]. The mesenchymal stem cell extracellular matrix (MSC ECM) contains proteins such as collagen, glycosaminoglycans, and growth factors that aid wound healing without triggering foreign inflammatory responses. The MSC secretome also secretes peptides and cytokines that suppress fibrosis and inhibit bacterial growth [5]. Our solution will harness these antimicrobial and pro-regenerative properties by electrospraying the MSC ECM into nanoparticles that have the proper zeta potential and size to reach the distal regions of the lung. We hypothesize that increased dosage of MSC ECM nanoparticles will further enhance the pro-regenerative and antimicrobial properties of this treatment to reach targeted distal regions of the lung while encouraging wound healing at the sites of injury.

Materials and

Methods: Decellularized MSC ECM was obtained using 0.1% Sodium Deoxycholate and DNAse I on confluent MSC cultures, then lyophilized into an ECM powder. The powder was digested in a glacial acetic acid solution and then electrosprayed into nanoparticles. In previous work [6], antibacterial effects were evaluated against E.coli, P. aeruginosa, and Staphylococcus aureus (Staph.aureus) after 24 hours of incubation with MSC ECM nanoparticles at concentrations of 0.5 ng/ml and 4 ng/ml. Investigating the same strains and now evaluating the dose dependency, we increased concentrations to 0.2 ug/ml and an additional double dose was administered at the third hour of the inhibition assay.

Results, Conclusions, and Discussions: MSC ECM nanoparticles demonstrated both pro-regenerative and antimicrobial effects. The average zeta potential was -11.16 (± 0.49) and the average size was 273.4 nm (± 20.9), allowing the nanoparticles to surpass mucus via repulsion and reach the distal lungs [7]. Decellularization retained ECM proteins and reduced DNA in the ECM by 93.6% [6]. This reduction in DNA is vital to suppress immune responses that could disrupt the delivery and effectiveness of foreign MSC ECM nanoparticles. The MSC ECM nanoparticles were deemed not cytotoxic and had proliferative effects from the MTT results. Preliminary results have suggested that MSC ECM nanoparticles inhibit the growth of P. aeruginosa (Figure 1).
The initial data suggests the MSC ECM nanoparticles are sufficient in stimulating cellular growth and inhibiting bacterial growth. Future work will be completed to analyze the pro-regenerative effects of MSC ECM nanoparticles at increased concentrations to determine the significance of dose dependency with current methods. Additional studies could be conducted to administer these nanoparticles into in vivo lung infection with aerosol delivery to regenerate injured tissue while preventing bacterial infection.

Acknowledgements (Optional): [1] Diamond M, Peniston HL, Sanghavi DK, et al. Acute Respiratory Distress Syndrome. [Updated 2024 Jan 31]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan-. Available from: https://www.ncbi.nlm.nih.gov/books/NBK436002/
[2] Xu H, Sheng S, Luo W, Xu X, Zhang Z. Acute respiratory distress syndrome heterogeneity and the septic ARDS subgroup. Front Immunol. 2023 Nov 14;14:1277161. doi: 10.3389/fimmu.2023.1277161. PMID: 38035100; PMCID: PMC10682474.
[3] Ragel EJ, Harris LK, Campbell RA. Acute respiratory distress syndrome: potential of therapeutic interventions effective in treating progression from COVID-19 to treat progression from other illnesses-a systematic review. BMJ Open Respir Res. 2023 Sep;10(1):e001525. doi: 10.1136/bmjresp-2022-001525. PMID: 37657844; PMCID: PMC10476125.
[4] Moretti M, Van Laethem J, Minini A, Pierard D, Malbrain MLNG. Ventilator-associated bacterial pneumonia in coronavirus 2019 disease, a retrospective monocentric cohort study. J Infect Chemother. 2021 Jun;27(6):826-833. doi: 10.1016/j.jiac.2021.01.011. Epub 2021 Jan 23. PMID: 33583739; PMCID: PMC7826005.
[5] Wang F, Li Y, Wang B, Li J, Peng Z. The safety and efficacy of mesenchymal stromal cells in ARDS: a meta-analysis of randomized controlled trials. Crit Care. 2023 Jan 20;27(1):31. doi: 10.1186/s13054-022-04287-4. PMID: 36670442; PMCID: PMC9857915.
[6] Wandling EN, Rhoads K, Ohman DE, Heise RL. Electrosprayed Mesenchymal Stromal Cell Extracellular Matrix Nanoparticles Accelerate Cellular Wound Healing and Reduce Gram-Negative Bacterial Growth. Pharmaceutics. 2023 Apr 19;15(4):1277. doi: 10.3390/pharmaceutics15041277. PMID: 37111762; PMCID: PMC10142868.
[7] Link PA, Ritchie AM, Cotman GM, Valentine MS, Dereski BS, Heise RL. Electrosprayed extracellular matrix nanoparticles induce a pro-regenerative cell response. J Tissue Eng Regen Med. 2018 Dec;12(12):2331-2336. doi: 10.1002/term.2768. Epub 2018 Nov 20. PMID: 30367566.