*4.7. Cell Culture*

Adult lymphatic endothelial cells (LECs) (PromoCell) were cultured with endothelial cell growth medium (PromoCell). After starvation with serum-free medium at passage 5–6, LECs were incubated in normal medium (control group, (Na+) = 145 mmol/L) and high-sodium medium ((Na+) = 185 mmol/L) for 24 h. Urea (Sigma–Aldrich, St. Louis, MO, USA) group was used to control for potential osmolarity effects. RT-PCR was performed for *Nkcc1* mRNA quantification. Total protein was extracted for quantitation of NKCC1, SPAK, and eNOS.
