Silica and Nano-materials Induce Red Blood Cell and Liposome Membrane Disruption That is Regulated By Cholesterol Content

Abstract

<p>Inhalation of respirable particles such as silica, has been documented to induce chronic lung diseases such as silicosis, which has been affecting humans since antiquity. Other, smaller materials such as engineered nano-materials (ENM) may trigger lung disease similar to silicosis, as seen in murine models. Within innate immune cells, such as alveolar macrophages, lysosomal membrane permeability (LMP) is proposed as a key and rate-limiting step in silica and ENM-induced inflammation. LMP activates the NLRP3 inflammasome and leads to the maturation and secretion of the pro-inflammatory cytokines IL-1ß and IL-18. In order to better study the membranolytic effects of particles, simpler, non-phagocytic cells like red blood cells (RBC) have been used. In this work, human RBC were treated with crystalline silica (CS), ZnO (ENM) or TiO<sub>2</sub> (ENM), and changes to RBC membrane order were analyzed by fluorescence lifetime imaging microscopy of the membrane incorporated probe, Di-4-ANEPPDHQ. RBC treated with methyl-ß-cyclodextrin to extract out cholesterol had a lower fluorescence lifetime of Di-4-ANEPPDHQ, which indicates a decrease in membrane order. Treatment with CS and TiO<sub>2</sub> resulted in a significant increase in fluorescence lifetime, indicating that these particles cause localized increases to lipid order. On the other hand, ZnO caused a decrease to the  lipid order with a lower fluorescence lifetime. These same three particles were used to treat liposomes composed of either phosphatidylcholine or phosphatidylserine lipids. All three materials induced an increase in lipid order measured by the time-resolved anisotropy of Di-4-ANEPPDHQ. CS and TiO<sub>2</sub> changed the order of PC liposomes, while ZnO treatment changed the lipid order of PS liposomes. Addition of cholesterol to these liposomes reduced the effects caused by TiO<sub>2</sub> and ZnO treatments. Taken together these results demonstrate a particle-membrane interaction that can disrupt lipid order and that this disruption can be reduced by the presence of cholesterol in the membrane. Selective modulation of lysosomal cholesterol content may be a potential therapeutic intervention for particle-induced inflammatory disease.  </p><p> </p><div> <div> <div> <p>The research within was funded by the National Institutes of Health grants R01ES023209, 1F32ES027324, P20GM103546, and P30GM103338. It was also funded by the M J Murdock Charitable Trust and the National Science Foundation grant CHE-1531520.</p> </div> </div> </div>