PC12 cells grown on cellulosic filters differentiate in response to NGF and exhibit a polarity not seen when they are grown on solid substrata

Abstract

Studies were performed with cellulosic filters and standard culture plates to compare methods of cell culture and differentiation of the cell line PC12, a clone originating from a rat pheochromocytoma. PC12 cells respond to nerve growth factor (NGF) by flattening of the cell body and subsequent extension of neurite-like processes. When PC12 cells are cultured in dishes without NGF, they have a diameter of approximately 3 to 7 micron and exhibit short processes of no longer than 3 to 5 micron. If PC12 cells are grown on a cellulosic filter they have the same average soma diameter and similar short processes extending laterally, but in addition have branching processes which extend as far as 10 to 15 micron into the filter substrate. When dish-cultured and filter-cultured cells are incubated with 50 ng/ml NGF they both exhibit differentiation-specific ultrastructural changes by 3 d of treatment. In the case of dish-cultured cells, large cytoplasmic processes exhibit an increase in the number of chromaffin cell-like secretory granules by 3 d of treatment. This characteristic is also demonstrated by filter-cultured cells, but the processes containing these granules are found concentrated within the cellulosic meshwork. Thus the timing of the NGF-elicited differentiation program is similar to both filter-cultured and dish-cultured cells, but the ultrastructural consequences are different. The filter-cultured PC12 cells exhibit a polarity not demonstrated by dish-cultured cells. Growing PC12 cells on cellulosic filters is a technique useful for "anchoring" neurons without the complication of the addition of extracellular matrix components. Filter-culture may represent a more in vivo-like method for studying neuronal growth and differentiation.

Nerve growth factor-induced changes in the intracellular localization of the protein kinase C substrate B-50 in pheochromocytoma PC12 cells.

“…1 A. This morphological differentiation may be induced by the cellulose filter substrate, as recently described (Van Buskirk et al, 1988). These protrusions sometimes resemble neurites and are also strongly labeled for B-50.…”

“…PC12 cells, grown on polylysine/gelatin-coated cellulose filters (Van Buskirk et al, 1988), were fixed in 2% paraformaldehyde/0.1% acrolein in PBS for 60 min, incubated for another 60 rain in 50 mM glycine in PBS, and embedded in 20% gelatin in PBS. Cellulose filters, sandwiched between two differently colored gelatin layers, were excised from the insert and cut in small squares.…”

Fabrication of Chitosan/Polypyrrole‐coated poly(L‐lactic acid)/Polycaprolactone aligned fibre films for enhancement of neural cell compatibility and neurite growth

“…41 At the same time, CS/PPy-PLLA/PCL and PPy-PLLA/ PCL fibre films were placed in the 24-well plate and disinfected with 75% alcohol for 0.5 hour and UV lights for 12 hours. Prior to seeding, the cells were pre-cultured with NGF of 50 ng/mL for 3 days to obtain their differentiation.…”

“…Prior to seeding, the cells were pre-cultured with NGF of 50 ng/mL for 3 days to obtain their differentiation. 41 At the same time, CS/PPy-PLLA/PCL and PPy-PLLA/ PCL fibre films were placed in the 24-well plate and disinfected with 75% alcohol for 0.5 hour and UV lights for 12 hours.…”

Performance tests of a novel coaxial tube catheter in anin vitromodel of intracranial cell delivery

“…PC 12 cells were available as a cell culture suspension. These cells have the potential to differentiate into neurons in the presence of nerve growth factor [10][11][12]. The cells were harvested each week to maintain viability, using the following protocol.…”