Kamehameha Schools Kapalama Honolulu, Hawaii, United States
Background: In the Native Hawaiian and Pacific Islander populations, kidney disease has an incidence rate of 921 per million population each year (Xiang et al., 2020). Part of this is due to the fact that 80-90% of the Pacific Islander population is dependent on imported foods, containing high amounts of ultra-processed foods (Francis, 2019). Ultra-processed foods often contain high levels of salt (NaCl) (Woods, 2019), which, at healthy levels in the kidney, supports blood filtration, reduces protein excretion, and protects glomerular pressure. However, at high levels, it directly disrupts kidney function by reducing the glomerular filtration rate and the ability to regulate nutrients in the body (Koo et al., 2018). For this study, human embryonic kidney cells (HEK-293) were chosen due to their simplicity and relevance. The cells are harvested from a terminated female fetus of unknown parentage (Synthego). In this study, HEK-293 viability, proliferation, migration, and morphology were observed when treated with three levels of NaCl.
Objective: • Evaluate the effects of varying concentrations of NaCl on HEK-293 cell viability, proliferation, migration, and morphology. Hypothesis: • When HEK-293 cells are exposed to higher concentrations of NaCl than what is already present from the FBS, they will experience cellular stress, resulting in a decrease in cell proliferation, viability, and migration as well as changes in the morphology
Design/Methods: Cell Culture: • HEK-293: DMEM + 10% FBS + 1% pen/strep + 1.6 ml amphotericin B cell death. Control and treatments: • C1: DMEM only & C2: DMEM + Solvent • Three treatment concentrations (2, 4, 8 mM) Viability/Proliferation Assay: • Seed cells in a well plate (Control & Experimental n=3) • Cells were treated with the three treatment concentrations 24 hours after being seeded • 48 hours after treatment, the cells were counted using the automated cell counter Morphology (via photos): • Images were taken at hour t=24 and t=48 Migration Assay: • Seeded cells and scratched 24 hours after (Control & Experimental n=3) • Allowed cells to adhere and then treated the wells • Imaging was conducted on t=0, t=24, t=48, & t=72
Results: Experimental Cell Morphology: • As NaCl increased, the structure of the cells changed from triangular to circular. • Suggests that higher levels of NaCl in the environment affect the cell membrane and allow for apoptosis to occur Experimental Proliferation Assay: • Decline in cell proliferation as NaCl treatments increased • The lowest concentration of NaCl was shown not to inhibit cell proliferation as significantly as higher concentrations. • 4 mM and 8 mM were shown to significantly decrease cell proliferation. Experimental Viability Assay: • As concentrations of NaCl increased, the percentage of living cells decreased, and the percentage of dead cells increased. • 4 mM and 8 mM significantly decreased the number of living cells. • 8 mM was shown to have the greatest effect on the percentage of living cells and dead cells. • 4 mM and 8 mM were shown to significantly decrease cell proliferation. Experimental Cell Migration: • Control cells were able to undergo the proper migration (gap size decreased (t=72) compared to t=0). • Cells with 2 mM were able to migrate slowly, and there was a slight decrease in the gap. • 4 mM and 8 mM showed cells were not able to migrate as well compared to control and 2 mM. • The gap for 4 mM and 8 mM stayed consistent throughout the assay.
Conclusion(s): • In the presence of high levels of NaCl, cell proliferation and viability can decrease. • Cell membrane structure can change significantly and possibly affect the structures of the cell organelles • Cell migration is inhibited in the presence of high concentrations of NaCl • The hypothesis was supported that at high levels of NaCl, there is a decrease in cell proliferation, viability, and migration as well as changes in the morphology.
