Maturation of neuroblastoma cells in the presence of dimethylsulfoxide.

Abstract

Addition of dimethylsulfoxide at concentrations of 1% and 2% (vol/vol) to cells of mouse neuroblastoma clone NIE-115 in the confluent phase of growth resulted in the production of morphologicalfir differentiated cultures with extensive process formation. Cells maintained in 2% dimethylsulfoxide remained in a stable nondividing condition for periods of up to 4 weeks. A high degree of electrical excitability was found in these cells, but there was no clear correlation of this property with the level of induction of either acetylcholinesterase (acetylcholine hydrolase; EC 3.1.1.7) or tyrosine hydroxylase [L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating); EC 1.14.16.2]. In addition, intracellular levels of cyclic 3':5'-AMP were not elevated in fully morphologically and electrically differentiated cells. While cell division was markedly inhibited by 2% or higher concentrations of dimethylsulfoxide, at 1% growth continued at a somewhat slowed rate and such cultures exhibited enhanced process formation and electrical activity for a relatively short period. High concentrations (3% or 4%) of dimethylsulfoxide totally suppressed process formation and did not result in increased excitability, but cells maintained high resting potentials. The results suggest that the development of the excitable membrane in neuroblastoma cells maybe expressed independently of neurospecific enzyme induction, and does not require a sustained elevation of cyclic 3'i5'-AMP levels.Nerve cells are characterized by their unique morphological appearance, the possession of an excitable membrane, and a specialized biochemical machinery. In order to study the expression of specific neuronal properties during the maturation process it is necessary to obtain sufficient quantities of a relatively homogeneous differentiating population. Cloned cell lines isolated from the C-1300 mouse neuroblastoma may serve as a useful system for exploring certain aspects of nerve cell differentiation. The transition of a culture of neuroblastoma cells from the actively dividing state to the confluent one is characterized by the synthesis of various enzymes involved in neurotransmitter metabolism (1), an enhancement of electrical excitability (2, 22), and some degree of process formation. To achieve a further expression of these properties, cells have been treated with various agents such as aminopterin (3, 4) or dibutyryl cAMP (5-7).In the present study, we show that in the presence of dimethylsulfoxide (Me2SO) neuroblastoma cells will extend neurites and develop a highly excitable membrane. It appears that this method offers certain advantages over those most commonly used, especially in that cells appear to reach a higher level of electrical differentiation and can be maintained in this state for extended periods. In experiments where logarithmically growing cells were used, cultures were trypsinized before attaining confluency and replated in 60 mm dishes at a density of 1 X 105 cells per dish. Two days later, the medium was replaced w...

Carboxyl methylation of 21–23 kDa membrane proteins in intact neuroblastoma cells is increased with differentiation

“…The control cells continued to multiply and reached confluence by 5-6 days in culture. The DMSO-treated cells stopped multiplication and extended processes several days (4-S) after plating, in line with previous reports [11,19]. Cells were incubated at 37°C in 2.5 ml DMEM (without methionine) containing 250&i [methyl-'Hlmethionine (15 CVmmol.…”

Regulation by Neurotransmitter Receptors of Serotonergic or Catecholaminergic Neuronal Cell Differentiation

“…Me 2 SO therefore acts as a dominant epigenetic signal. Such an effect of Me 2 SO can be related to that described in the case of Friend erythroleukemia cells (24) and of NIE-115 neuroblastoma cells (25), where Me 2 SO confers a neuronal morphology. The catecholaminergic program is restricted to the induction of (i) the catecholamine metabolism, (ii) an adrenoreceptor, and (iii) NE uptake.…”

Early G1 growth arrest of hybridoma B cells by DMSO involves cyclin D2 inhibition and p21[CIP1] induction

“…The growth inhibitory properties of DMSO were usually associated with di erentiation (Kimhi et al, 1976;Kluge et al, 1976;Maeda and Sachs, 1978;Reagan et al, 1990).…”

“…Dimethylsulfoxide (DMSO) has been reported to induce both growth arrest and di erentiation of myeloid (Hiroguchi-Yamada and Yamada, 1993;Klinken et al, 1988) or neuroblastoma cell lines (Kimhi et al, 1976;Kranenburg et al, 1995) but in these cell systems it is di cult to discriminate the molecular events involved in growth arrest to those that are necessary for cellular di erentiation. We show that, in the hybridoma 7TD1 cells, like in other B cell lines such as Raji, Molt 4 or SKW6-CL4 (Sawai et al, 1990;Takase et al, 1992Takase et al, , 1994Teroaka et al, 1996) exposure to DMSO arrests the cells in G 1 phase, without inducing di erentiation or apoptosis.…”