Poster X2 - Targeted Cellular Delivery of Nanoengineered Extracellular Vesicles for Acute Kidney Injury

Introduction: Acute kidney injury (AKI) is a critical condition characterized by a sudden decline in kidney function, often triggered by ischemia, nephrotoxins, or sepsis. This condition is associated with significant morbidity and mortality, leading to prolonged hospital stays, increased healthcare costs, and long-term complications such as chronic kidney disease and end-stage renal failure. Despite its clinical importance, current treatment options primarily focus on supportive care measures, highlighting the urgent need for effective therapeutic interventions targeting AKI's underlying pathophysiology to mitigate renal damage and promote kidney repair and regeneration. One promising approach for AKI therapy involves utilizing nanoengineered extracellular vesicles (EVs) as delivery vehicles for therapeutic agents. EVs, including exosomes and microvesicles, play crucial roles in intercellular communication and tissue homeostasis. By engineering EVs to encapsulate and deliver therapeutic cargo, we aim to enhance the targeted delivery of protective and/or regenerative factors to injured renal cells. As a therapeutic cargo, we use Sox9, which during early- AKI, has been seen to play a protective role, and during the recovery phase facilitates repair. Notably, employing cells like myeloid derived suppressor cells (MDSCs) as delivery vehicles can further enhance targeting, given their higher tropism towards inflammatory tissue.

Materials and

Methods: MDSCs were transfected using bulk electroporation with a Sham or Sox9 plasmid, then the EVs were isolated from culture media 24 h after transfection. For in vitro experiments, renal proximal tubule cells (RPTCs) were cultured until achieving around 80 - 85% confluence, then 106 EVs were added to the media and after 24 hours cells were collected and RNA isolated, qPCR was used to evaluate Sox9 expression. For in vivo experiments, a cisplatin-induced mouse model was used, where C57BL/6J male mice (8-10 weeks) received either a 25 mg/kg cisplatin or saline by intraperitoneal injection. Subsequently, 106 MemGlow 640 labeled-MDSCs were administered via tail-vein injection at 4- and 24-hours post-AKI. Brain, lungs, heart, spleen, liver and kidney, blood and urine were collected. Biodistribution in brain, lungs, spleen, heart, liver, and kidney was assessed via IVIS imaging, and kidney tissues were subjected to histological analysis, immunofluorescence staining, and qPCR to evaluate renal function and expression of AKI and inflammation markers.

Results, Conclusions, and Discussions: For in vitro studies, we have shown the ability to package SOX9 mRNA in EVs isolated from transfected MDSCs, and that the size and concentration of SOX9 EVs did not have any noticeable difference between them and those from sham transfected cells. For in vivo studies, our findings indicated a significant increase in the expression of AKI and inflammation markers after high-dose cisplatin injection, corroborated by elevation in blood urea nitrogen levels after 24 hours. Biodistribution analysis showed a significantly higher fluorescence in the kidneys with AKI and labeled MDSCs when compared to labeled MDSCs injected to non-injured mice indicating inflammation-mediated homing of MDSCs to the site of injury.

We observed preferential accumulation of labeled-MDSCs in the kidneys at 4- and 24-hours post-injury, underscoring the potential for targeted cellular delivery of engineered EVs in AKI treatment. This personalized approach holds promise for improving patient outcomes and reducing AKI-related complications, representing a significant step towards precision medicine in AKI management. Future studies will focus on exploring the therapeutic effect of Sox9 delivery to AKI kidneys.

Acknowledgements (Optional): This work was supported by the OSU Center for Medical and Engineering Innovation Interdisciplinary Pilot Grant