Talk:Cells - HT4
Transplantation of a temperature-sensitive, nerve growth factor-secreting, neuroblastoma cell line into adult rats with fimbria-fornix lesions rescues cholinergic septal neurons
J Neurosci Res. 1991 Feb;28(2):156-70.
Whittemore SR, Holets VR, Keane RW, Levy DJ, McKay RD. Source Miami Project, University of Miami School of Medicine, FL 33136.
The HT4 cell line was derived from infection of a mouse neuroblastoma cell line with a retrovirus that encoded the temperature-sensitive (ts) mutant of SV40 large T antigen. At nonpermissive temperature, HT4 cells differentiated with neuronal morphology, expressed neuronal antigens, synthesized nerve growth factor (NGF) mRNA, and secreted biologically active NGF in vitro. We sought to establish whether transplanted HT4 cells expressed class I major histocompatibility complex (MHC) antigens, a partial requirement for recognition by cytotoxic T lymphocytes (CTL), and thus be susceptible to xenograft rejection. Differentiated HT4 cells expressed marginally detectable levels of class I MHC antigens, but demonstrated higher levels of class I MHC expression after treatment with interferon-gamma. However, HT4 cells were resistant to direct lysis by perforin, the pore-forming protein of CTLs, and thus may have potential use in xenograft experiments. To address whether HT4 cells secrete NGF in vivo, HT4 cells were transplanted into adults rats with unilateral fimbria-fornix transections. A ts cell line derived from P4 cerebellum, BT1, that does not differentiate with neuronal phenotype or synthesize NGF in vitro, was transplanted as a control. Six weeks posttransplant. HT4 cells had integrated into host CNS without forming tumors. In BT1 transplants, the number of medial septal acetylcholinesterase (AChE)-positive cells was reduced to 26-39% of the contralateral control side, depending on the rostrocaudal level. In HT4 transplants, the number of cholinergic septal neurons was 58-78% of the contralateral side. This percentage was significantly (P less than 0.005) greater than that seen with BT1 transplants, indicating that transplanted HT4 cells secrete NGF in vivo and rescue cholinergic septal neurons following fimbria-fornix transection.
HT4 cells were derived by infecting a mouse neuroblastoma cell line with a retrovirus encoding the ts mutant of SV40 large T antigen and BTI cells were similarly derived from primary cultures of P4 rat cerebellum (D. Levy and R. McKay, unpublished results); for a detailed description of the methods used to develop these cell lines, see Frederiksen et al., 1988). All cells were grown at 33°C in Dulbecco’s modified Earl’s medium (DMEM) containing 10% FCS, 100 Uiml penicil- lin, 100 pg/ml streptomycin, and 500 pg/ml G418 in
HT4 and BTI cells were differentiated at 39°C in N5 media (Kawamoto and Barrett, 1986) containing 0.5%horse serum, the N2 serum free components (Bottenstein and Sato, 1979), and 5 ng/ml bFGF. Following 1-2 weeks under differentiating conditions, cells were immunohistochemically stained for NF, NSE, GFAP, and