Manipulation Immune System via Immortal Bone Marrow Stem Cells
Circa 2008 face_with_colon_three
Adult bone marrow contains a small pool of hematopoietic (HSC) stem cells that can maintain themselves and all other blood lines throughout mammalian lifetimes (1,2). The ability to maintain HSC in tissue cultures would enable the introduction of gain and loss of function mutations in hematopoietic systems. Our failure to expand HSC has hindered the use of this method. Even in a cytokine-optimal environment, BM suspension cultured HSCs are rapidly lost despite the rapid growth of progenitors, more differentiated cells, and even despite vigorous proliferation for at least 3 weeks (3,4). The phenomenon of stem cells exhaustion, or senescence, may be the limit that makes it impossible to extend HSC culture beyond a certain period.
Iscove and Nawa (9), through serial transplantation studies that carefully assessed the input and output content of HSC for each transfer generation, showed that mouse HSC expand in vivo up to at least 8000-fold. Recent improvements in in vitro techniques and the use of new growth factors have allowed mouse HSC to expand up to 30 times ex vivo. Since it is unclear to what extent the external conditions of culture can be improved alternative, but not mutually-exclusive efforts have been made to change HSC’s intrinsic properties. Humphries and Savageau, along with their colleagues, conducted seminal experiments that showed ectopic expression in BM cells of the transcription factor HOXB4 supported HSC survival and growth in vivo as well as in vitro. They found that by closely monitoring the HSC levels in their cultures of HOXB4 transduced BM cell cultures, HSCs could grow up to 41 times in 2-week liquid cultures. HOXB4 is a member of the large family of transcription factors called HOX. They are important for basic developmental processes and also play a role in maintaining different stem cell compartments.
We have built on Humphries, Savageau, and their colleagues’ findings to establish long-term murine BM culture of HOXB4 transduced cells and monitor their stem cell content in order to determine how genetically modified HSCs and their multipotent primate progenitors can be expanded for experimental purposes. We also established BM cells transduced by constructs encoding Nucleoporin 98 NUP-HOXB4 fusion proteins (NUP cell) for comparison and to follow Humphries et. al. (14) showed that the ectopic expressions of similar fusions induced in vivo a more robust expansion of HSC.
Source:
https://academic.oup.com/intimm/article/20/9/1211/667462
