Poster #004, Children's Hospital Los Angeles
The Cellular Uptake of Microplastics within Proximal Tubular Kidney Cells colocalize in the endosomes and lysosomes
Mentors: Mohammadreza Kasravi, MD; PI: Stefano Da Sacco, PhD
Microplastics are in the water people drink to the food people eat and the clothes that people wear everyday. They are a growing concern because it is almost impossible for them to fully degrade due to their synthetic polymer chains. Instead, they only break down into smaller and smaller particles dispersing and contaminating every environment. When these microplastics are ingested or inhaled, they accumulate in organs, including the kidneys. Studies have previously reported that the absorption of microplastics in the kidneys is linked to oxidative stress, cell apoptosis, and inflammation. While these effects have been tested, there is not yet a complete understanding of their cellular entry. To figure out how they are uptaken, proximal tubular cells (PTC) were treated with microplastics. It can be hypothesized that endocytosis is causing the cellular uptake, with the particles ultimately ending up in the endosomes and lysosomes. PTC were given red fluorescent polystyrene microplastics (1 µm) and left to absorb at various time points of 1, 3, 6, and 24 hours. PTC were then fixed and stained with primary antibodies (EEA1, LAMP1, Rab7, Rab11) and secondary antibodies (555 anti rabbit and 448 anti mouse). EEA1 is to mark early endosomes, LAMP1 to mark lysosomes, Rab 7 to track late endosomes, and Rab11 to track the excretion of the microplastics. Confocal fluorescence microscopy was used to image the antibodies and analyze whether the microplastics were being absorbed by the endosomes and lysosomes. Data will be quantified with ImageJ by using Manders coefficient to measure the colocalization of the microplastics’ red fluorescent markers with the green fluorescent endosomes and lysosomes. Results are expected to show that the microplastics will colocalize the most at the 24 hour mark compared to time less than that because the microplastics will need time to be engulfed by the PTC.