Human β ngf

Cells were seeded according to calculated doubling times to control for initial increases in proliferation. WT SH-SY5Y (30,000 cells/well) or PAG1^TM- SH-SY5Y (9000 cells/well) cells were seeded in 6-well plates containing RPMI 1640 + 10% FBS. Cells were grown for 2–4 d until fully adhered. Spent media were aspirated and replaced with differentiation media composed of RPMI 1640 + 2% FBS, 10 µM 9-cis-RA (Sigma-Aldrich) and/or 5 nM human β-NGF (Peprotech). Cells were grown in differentiation media for 8 d; treatment media were replaced every other day. On day 8, neurite tracings, length quantifications, and cell body measurements were performed using ImageJ (Rueden et al., 2017 (link)). Only neurites longer than one cell body length were quantified to eliminate lamellipodia/filopodia. Representative images were taken on day 8 using a Zeiss Invertoskop light microscope with an Amscope MD500 camera.

Poly(lactic-co-glycolic acid) (PLGA, LA/GA = 50:50, M_w = 100 kDa) and PDLLA (M_w = 100 kDa)
were supplied by Lakeshore Biomaterials. The human β-NGF with
the ELISA kit was supplied by Peprotech Inc., and human GDNF with
the ELISA kit was supplied by R&D Systems, Inc. The culture medium
and other reagents were purchased from Invitrogen, Inc. All chemicals
were used as purchased.