Chylous fluid samples were collected from pediatric patients who underwent cardiac surgery to correct structural congenital cardiac deformities (Table 1 ) with postoperative chylothorax fluid defined as: ≥80% lymphocyte count, and/or chylomicron positivity, and chyothorax fluid triglyerides (TG) > 50% serum TG levels (Columbia University IRB AAAQ6902). Patients with chromosomal anomalies were excluded.
Cells were depleted of CD133+ cells by magnetic bead isolation (Miltenyi) as described.18 (link) Nonadherent immune cells were removed after seeding and adherent CD133-negative cells were expanded and characterized by quantitative RT-PCR (qRT-PCR) and fluorescent activated cell sorting (FACS). HdLECs, isolated from neonatal dermis using CD31+ bead selection and live cell sorting for PODOPLANIN, served as normal controls.19 (link) pcLECs and HdLECs were maintained on fibronectin-coated plates in EGM-2 media (Lonza) supplemented with 18% FBS or EGM-MV2 media (Lonza), respectively.
Cells were depleted of CD133+ cells by magnetic bead isolation (Miltenyi) as described.18 (link) Nonadherent immune cells were removed after seeding and adherent CD133-negative cells were expanded and characterized by quantitative RT-PCR (qRT-PCR) and fluorescent activated cell sorting (FACS). HdLECs, isolated from neonatal dermis using CD31+ bead selection and live cell sorting for PODOPLANIN, served as normal controls.19 (link) pcLECs and HdLECs were maintained on fibronectin-coated plates in EGM-2 media (Lonza) supplemented with 18% FBS or EGM-MV2 media (Lonza), respectively.