Neuronal traits of clonal cell lines derived by fusion of dorsal root ganglia neurons with neuroblastoma cells.

Abstract

In an attempt to immortalize the gene products of single neurons, somatic cell hybrids were produced by fusion of embryonic rat dorsal root ganglion (DRG) neurons with mouse neuroblastoma cells. Embryonic day 13 rat DRGs were fused with mouse neuroblastoma cells deficient in hypoxanthine phosphoribosyltransferase (HPRT; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8). The hybrid cells were selected in medium with 100 IM hypoxanthine/1 !LM aminopterin/12 ,LM thymidine to eliminate the neuroblastoma cells and with cis-hydroxyproline to retard fibroblast growth. Of the 17 lines derived, 4 manifested neuronal properties and were cloned. These lines retain both rat and mouse chromosomes and synthesize characteristic rat and mouse isoenzymes. Neuronal gangliosides, action potentials, and extensive neurite-like processes are exhibited by these hybrid cells, properties characteristic of DRG neurons but not of the neuroblastoma parent. Each line manifests a unique combination of action-potential properties and cell-surface markers, suggesting the selective expression of subsets of DRG neuronal genes.All of these neuronal properties are expressed constitutively, without the need for chemical induction or mitotic inhibition, and stably, without diminution after at least 5 months in culture. These lines may prove useful in the identification and isolation of gene products that characterize individual or small subsets of DRG neurons.

Detailed Characterization of Neuroprotection by a Rescue Factor Humanin against Various Alzheimer's Disease-Relevant Insults

“…F11 cells, the hybrid cells of a rat embryonic day 13 (E13) primary cultured neuron and a mouse neuroblastoma, were grown in Ham's F-12 (Life Technologies, Gaithersburg, MD) plus 18% fetal bovine serum (FBS) (Hyclone, L ogan, UT) and antibiotics, as described previously (Platika et al, 1985;Yamatsuji et al, 1996;Huang et al, 2000;Niikura et al, 2001). F11 cells (7 ϫ 10 4 cells per well in a six-well plate cultured in Ham's F-12 plus 18% FBS for 12-16 hr) were transfected with plasmids encoding FAD genes by lipofection [1 g of FAD cDNA, 2 l of LipofectAM I N E, and 4 l of PLUS reagent (Life Technologies)] in the absence of serum for 3 hr and were incubated with Ham's F-12 plus 18% FBS for 2 hr.…”

“…We next examined the effect of HN on F11 cell death by other APP mutants: A617G-APP and L648P-APP. F11 cells are neuronal hybrid cells established by fusing rat primary cultured E13 neurons with mouse NTG18 neuroblastoma cells and carry typical characteristics of primary cultured neurons, including generation of action potentials (Platika et al, 1985). As shown in Figure 3A, sHN dose-dependently suppressed death of F11 cells induced by A617G-APP and L648P-APP, and Ն10 M sHN completely inhibited F11 cell death by both APP mutants (IC 50 values were 100 nM to 1 M against both mutants, which were comparable with the IC 50 of the sHN effects against V642I-APP and NL-APP).…”

Nonspecific association of 2',3'-cyclic nucleotide 3'-phosphodiesterase with the rat forebrain postsynaptic density fraction

“…HiB5 cells can differentiate into neuronal cells with neurite-like processes in vitro in serum-free defined N2 medium containing PDGF at non-permissive temperature of 39 o C (Cho et al, 2001; Sung et al, 2001). F11 is a hybrid cell line between neuroblastoma N18TG2 and dorsal root ganglionic neurons (Platika et al, 1985). F11 cells differentiate into neurons in the presence of cAMP or prostaglandins and express neuronal markers including receptor for bradykinin, prostaglandin, opioids and N-and L-type voltage-dependent Ca 2+ channels (Francel et al, 1987;Boland and Dingledine 1990;Fan et al, 1992;Cruciani et al, 1993), suggesting that F11 cells are similar to dorsal root ganglionic neurons.…”

“…Thus, CNP expression confirmed in neuronal cell lines. Two cell lines were chosen for this purpose; HiB5, a hippocampal stem cell line which has been established from the rat E16 hippocampus (Renfranz et al, 1991) and F11, a hybrid cell line between neuroblastoma N18TG2 and dorsal root ganglionic neurons (Platika et al, 1985). Differentiation of HiB5 and F11 stem cells were induced by platelet-derived growth factor (PDGF) and cAMP, respectively (Cho et al, 2001;Sung et al, 2001), and their homogenates were subjected to immunoblot analysis with polyclonal or monoclonal α-CNP antibody.…”

Human Neuroma Contains Increased Levels of Semaphorin 3A, Which Surrounds Nerve Fibers and Reduces Neurite ExtensionIn Vitro

“…F11 cells are derived from a fusion of mouse embryonic neuroblastoma cells and rat dorsal root ganglion neurons (Platika et al, 1985). Neurite outgrowth in these cells can be induced by withdrawal of serum and the addition of forskolin to the culture medium.…”