In vitro DIFFERENTIATION OF A MOUSE NEUROBLASTOMA

Abstract

Abstract. M\Iouse tumor C1300 has been established in tissue culture. The cells have a round cell morphology in both the subcutaneous tumor and in suspension culture. However, when given a surface on which to attach, they send out processes up to 3 mm in length and assume the morphology of mature neurons. The attached cells are stained by the Bodian silver procedure for neurons, whereas the cells grown in suspension are not. Electron microscopy reveals that the attached cells contain neurofilaments, neurotubules, and densecore vesicles indicative of nerve fibers. Both free-floating and attached cells have tyrosine hydroxylase activity characteristic of sympathetic nervous tissue. Apparently cell attachment can induce morphological differentiation from an anaplastic round cell to a cell which has many properties of a mature neuron.The analysis of differentiated function is hampered by the fact that the end cell usually does not divide. In the case of extremely heterogeneous cell populations such as those found in the endocrine, nervous, immune, and hepatic tissues, further advances require a way of establishing clones of end cells which express their differentiated phenotype in tissue culture.1' 2 Such cells are most easily derived from neoplasms and have been used successfully in the study of endocrine,3 hepatic,4 and endoreticularl cell function. This communication extends these studies to the nervous system by describing a cloned, tissue-culture adapted, mouse cell line which can be induced to differentiate to a cell which has some properties of a mature neuron.Materials and Methods.-Tumor lines: Mouse tumor C1300 was obtained from Jackson Laboratories. This cell line has been previously described as a round cell tumor, possibly a neuroblastoma.5' 6 The spontaneous neoplasm originated in the body cavity in mouse strain A/J in 1940 and has been maintained by subcutaneous transfer in A/J mice. The solid tumor was adapted to tissue culture conditions by dispersing the cells in modified Eagle's medium' containing 20% fetal calf serum. The cell cultures were maintained at 370C in an 85% air, 15% CO2 incubator. When grown in plastic Petri dishes, the established cell line had a doubling time of 17 hr. Cells were cloned twice by spreading dilute cell suspensions on solid agar (0.5% agar, 4.5 mgm/ml trypticase soy broth in Eagle's modified medium plus 20% fetal calf serum) and picking visible colonies with a platinum loop after a 2-week incubations Clones had doubling times comparable to the initial explant, and were homogeneous with respect to cell morphology, staining characteristics, and karyotype.9Staining and electron microscopy: For light microscopy, cells were grown in Petri dishes with glass cover slips. After 2 to 4 weeks, the cover slips were washed several times in Eagle's medium and the attached cells stained according to the Giemsa,10 and Bodian10 11 procedures. Cells growing in suspension were centrifuged, washed twice in modified Eagle's medium, spread on a slide, and stained as described above.

Herpes simplex virus type 1 latency-associated transcript inhibits apoptosis and promotes neurite sprouting in neuroblastoma cells following serum starvation by maintaining protein kinase B (AKT) levels

“…Removal of serum from certain neuroblastoma cells induces a low percentage of these cells to sprout neurites and develop a differentiated morphology (Schubert et al, 1969;Seeds et al, 1970). Certain growth factors also stimulate neurite sprouting (Evangelopoulos et al, 2005;Lee et al, 2006).…”

The uptake of alpha-foetoprotein by C-1300 mouse neuroblastoma cells

“…Proliferating cell lines of neuroblastoma express several neuronal characteristics such as process formation (Schubert et al, 1969), neurotransmitter synthesis (Biedler et al, 1978;Pons et al, 1982), high acetylcholinesterase and electrical activities. They lack, however, the ability to synapse between themselves (Zagon et al, 1978) and have a less complex ganglioside pattern than is found for neurons (Stoolmiller, 1973).…”

A sequence of changes in cytoskeletal components during neuroblastoma differentiation

“…Despite some abnormalities, due to their neoplastic character, they have proved very useful in the study of formation of neurites, the appearance of enzymes that synthesize neurotransmitters and the acquisition of membrane excitability [15][16][17]. However, if studies on brain maturation have shown large modulations in the various cytoskeletal components, comparable studies 'concerning the neuroblastoma are generally limited to the isotubulins and report moderate changes in their microheterogeneity concomitant with cell differentiation.…”