Interleukin 2 (IL- 2) is a T cell growth factor. In the present study, human IL- 2 cDNA was cloned from total RNA of activated tonsillar mononuclear cells (TMNC) following reverse transcription and PCR amplification u...Interleukin 2 (IL- 2) is a T cell growth factor. In the present study, human IL- 2 cDNA was cloned from total RNA of activated tonsillar mononuclear cells (TMNC) following reverse transcription and PCR amplification using a pair of synthesized primers. DNA sequence analysis demonstrated that the IL- 2 cDNA cloned from the Chinese tonsil donor is identical with the data reported so far, reflecting the high structural conservation of this gene. The human IL- 2 cDNA was inserted into XM6 to construct a recombinant retroviral expression vector XM6-IL2, having human IL-2 cDNA driven by the 5' LTR of MMLV. This vector was successfully passaged through ψ 2 and PA317 cells to yield high producer lines of ecotropic and amphotroplc infectious viruses. The murine myeloma cell line SP2/0 after being infected by retrovirus released from high titer PA317 constitutlvely secreted IL- 2 activity into the culture medium when assayed for T cell proliferative capacity. Activated human T cells are also exposed to the infectious retrovirus XM6- IL2. In sharp contrast to the parallel controls, the infected T cells with or without the furtheraddition of 500 units/ ml of exogenous IL- 2 proliferated and formed colonies of significant size under the selection pressure of G418. However, their growth in vitro could only be maintained for about 3 weeks. These facts demonstrated that gene transfer of human IL- 2 cDNA viaretrovirus is far from being sufficient to maintain the relatively long-termgrowth and clonal expansion of human T cell subpopulations in vitro.展开更多
文摘Interleukin 2 (IL- 2) is a T cell growth factor. In the present study, human IL- 2 cDNA was cloned from total RNA of activated tonsillar mononuclear cells (TMNC) following reverse transcription and PCR amplification using a pair of synthesized primers. DNA sequence analysis demonstrated that the IL- 2 cDNA cloned from the Chinese tonsil donor is identical with the data reported so far, reflecting the high structural conservation of this gene. The human IL- 2 cDNA was inserted into XM6 to construct a recombinant retroviral expression vector XM6-IL2, having human IL-2 cDNA driven by the 5' LTR of MMLV. This vector was successfully passaged through ψ 2 and PA317 cells to yield high producer lines of ecotropic and amphotroplc infectious viruses. The murine myeloma cell line SP2/0 after being infected by retrovirus released from high titer PA317 constitutlvely secreted IL- 2 activity into the culture medium when assayed for T cell proliferative capacity. Activated human T cells are also exposed to the infectious retrovirus XM6- IL2. In sharp contrast to the parallel controls, the infected T cells with or without the furtheraddition of 500 units/ ml of exogenous IL- 2 proliferated and formed colonies of significant size under the selection pressure of G418. However, their growth in vitro could only be maintained for about 3 weeks. These facts demonstrated that gene transfer of human IL- 2 cDNA viaretrovirus is far from being sufficient to maintain the relatively long-termgrowth and clonal expansion of human T cell subpopulations in vitro.
