1. Nanoparticle-Based Gene Delivery Systems for Anti-Inflammatory Therapy Michelle Gee, YSP Student, Wellesley High School Yasmeen Elaywan, YSP Student, Boston Latin School Dr. Huyen Tran, Postdoctoral Research Associate, Northeastern University Prof. Mansoor Amiji, Department of Pharmaceutical Sciences, Northeastern University

2. Inflammatory Diseases Cancer Pulmonary disease Rheumatoid arthritis Type II diabetes Fatty liver disease Inflammatory bowel disease Atherosclerosis Alzheimer’s Parkinson’s Chronic Inflammation

3. Macrophages in Inflammation Macrophage: phagocytic immune cell Can be stimulated to inflammatory M1 or anti-inflammatory M2 In inflammatory diseases macrophages are prominently M1 Stimulate macrophages to M2 as possible treatment M1 M2 Bosurgi et al., Front Immunol. 2011; 2: 62.

4. Why miRNA-223 and Nanoparticles? miRNA-223 Inhibits expression of pro-inflammatory cytokines Verify that miR-223 converts macrophages to anti- inflammatory M2 Nanoparticles Protect miRNA and allow it to enter cell Targeted delivery- HAPEI Cy5-labeled miRNA -ve charge HAPEI-miRNA JJJJJJ

5. Methods I. Nanoparticle Preparation and Characterization II. Peritoneal Macrophage Isolation & Culture III. Peritoneal Macrophage Transfection with miRNA-223 Nanoparticles IV. Polarization StudyIV. Anti-Inflammatory Effect Study

6. Nanoparticle Characterization 235 d nm Z-Average: 200.6 d nm 25 bp Ladder miRNA HAPEI/miRNA HAPEI/miRNA/PAA A B C

7. Uptake Study in Peritoneal Macrophages DAPI-nuclei Cy5-HAPEI Merged Control 1 h 3 h 5 h 10 um

8. miRNA-223 Levels in Peritoneal Macrophages Blank-HAPEI Untreated HAPEI/miRNA-223 Lipofectamine/miRNA-223 Taqman Assay

9. Polarization Study iNOS (M1 inflammatory marker) Arg (M2 anti-inflammatory marker) M2M1 Untreated Macrophage LPS IFN- γ HAPEI/miRNA Nanoparticle qPCR

10. Inflammatory Cytokine Expression Levels in Peritoneal Macrophages IL-1 β Expression Levels IL-6 Expression Levels TNF- α Expression Levels LPS (inflammatory) M2 Untreated Macrophage HAPEI/miRNA Nanoparticle qPCR M2

11. Conclusions and Further Research Conclusions Nanoparticles prepared and entered cells miRNA-223 treated cells expressed lower levels of inflammatory cytokines and polarized from M1 to M2 Promising treatment for inflammatory diseases Future Research In vivo studies Disease markers

12. Acknowledgements Amiji Lab Professor Mansoor Amiji Dr. Huyen Tran Dr. George Matthaiolampakis Swathi Krishnan Dandan Ling Amit Mirchandani Yue “Jerry” Zhang Northeastern University Young Scholars Program Claire Duggan Maureen Cabrera Madeline Leger Supported by: