Yonago Acta medica 1992;35:221–230

A Hypoxanthine Phosphoribosyl Transferable Deficient Mutant from a Cell Line Derived from a Mouse Model with Niemann-Pick Disease Type C

Shinjiro Akaboshi, Jun Tohyama, Shigeki Miyawaki*, Kousaku Oho and Kenzo Takeshita

Division of Child Neurology, Institute of Neurological Sciences, Faculty of Medicine, Tottori University, Yonago 683 and *Research Laboratories, Nippon Shinyaku Co., Kyoto 607, Japan

An SPM-3T3 cell line, which is derived from a sphingomyelinosis (Niemann-Pick) mouse fetus, shows the characteristic abnormalities observed in cultured fibroblasts from patients with Niemann-Pick disease type C. Because hypoxanthine phosphoribosyl transferable (HPRT) deficient sublines from this cell line should be very useful in genetically analyzing diseases that appear in mice and humans, we attempted to isolate HPRT deficient SPM-3T3 cells. However, our attempt to isolate 6-thioguanine (6TG) (10 µg/mL) resistant cells directly from mutagenized SPM-3T3 cells was unsuccessful. As a result, we later applied a two-step selection process. First, we attempted to select a low concentration (1 µg/mL) of 6TG for a longer period, and second, we attempted a selection by administering a high concentration (10 µg/mL) of 6TG for 10 days. From among the 16 × 10⁷ mutagenized cells, 16 resistant clones were selected. One clone, which showed rapid growth similar to that exhibited by parental cells, was chosen ad the standard. This clone (SPM-3T3-6TG^r) exhibited a severe deficiency of HPRT activity. The cell line should be useful in genetically analyzing conditions that show a defect in the intracellular transport of exogenously derived cholesterol.

Key words: Niemann-Pick disease; hypoxanthine phosphoribosyl transferase; 6-thioguanine; cholesterol

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