Jialiu Zeng1 Kevin Smith1 Orian Shirihai2 Mark Grinstaff1

1, Boston University, Boston, Massachusetts, United States
2, University of California, Los Angeles, Los Angeles, California, United States

We have developed a novel polymeric pH-activable, acidifying nanoparticle (acNP) that restores the pH of compromised lysosomes to rescue autophagic flux and cellular function in pancreatic beta cells (INS1 cells) under lipotoxicity. In beta cells, chronic exposure to high levels of fatty acids (lipotoxicity) leads to an inhibition of autophagic flux and subsequent cellular dysfunction, which has been recently associated with impaired lysosomal acidification and elevated lysosomal pH. Therefore, restoration of lysosomal pH is essential in alleviating the block in autophagy and promote proper cellular quality control and function. In this study, we designed an acidic nanoparticle (acNP) that contains caged acid which can be released upon moderate pH changes (pH 6.0) to enable controlled acidification of the impaired lysosomes under lipotoxicity. Rhodamine labelled acNPs demonstrate dose dependent uptake into lysosomes of INS1 cells. pH-activation of acNPs in dysfunctional lysosomes at pH 6.0 environment demonstrate acidification of lysosomes and restored lysosomal pH with minimal cytotoxicity. acNPs also increased lysosomal cathepsin enzyme activity, and decreased both autophagic proteins LC3II and p62 levels, indicating a rescue of lysosomal function and autophagic flux due to restoration of lysosomal acidity. Additionally, acNPs restored glucose-stimulated insulin secretion that is reduced in INS1 cells and mouse islets under lipotoxicity. These results indicate that acidifying lysosomes with acNPs improved lysosomal function and autophagic flux in INS1 cells under lipotoxicity, and are of therapeutic interest for pathologies associated with lysosomal acidity impairment such as type II diabetes and non-alcoholic fatty liver disease (NAFLD).