Purification and Characterization of a T Lymphocyte-Derived Differentiation-Inducing Factor for Human Promyelocytic Cell Line (HL-60) and Its Relationship to Lymphotoxin

• Published in: MICROBIOLOGY and IMMUNOLOGY Vol. 33; no. 6; pp. 489 - 501
• Main Authors: Shimizu, Yukitoshi, Nakamura, Takehiko, Niki, Tamotsu, Hemmi, Hiromichi, Sugamura, Kazuo
• Format: Journal Article
• Language: English
• Published: Tokyo Blackwell Publishing Ltd 1989; Center For Academic Publications Japan; Center for Academic Publications Japan
• Subjects: Animals; Biological and medical sciences; Cell Differentiation; Cell Line, Transformed; Chromatography, Ion Exchange; Cricetinae; Cytotoxicity Tests, Immunologic; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Human T-lymphotropic virus; Humans; Immunobiology; Isoelectric Focusing; Lymphoid cells: ontogeny, maturation, markers, receptors, circulation and recirculation; Lymphokines - isolation & purification; Lymphokines - metabolism; Lymphotoxin-alpha - metabolism; Neutralization Tests; T-Lymphocytes - metabolism; Virus; Characterization; Promyelocytic leukemia; Lymphotoxin; Cell line; Purification; T-Lymphocyte; Retroviridae; Cell transformation; HTLV-I virus; Myeloid cell; Colony stimulating factor
• ISSN: 0385-5600; 1348-0421
• DOI: 10.1111/j.1348-0421.1989.tb01998.x

The human T cell leukemia virus type I (HTLV-I)-transformed T lymphocyte cell line MT-2 constitutively produces differentiation-inducing factor (DIF) for the human promyelocytic leukemia cell line HL-60. Purification and characterization of DIF derived from MT-2 were performed here to identify T cell-derived DIF. DIF was purified from conditioned medium of the MT-2 cell culture with serum-free medium by utilizing the sequential chromatographies of anion-exchange (mono-Q) column, gel filtration (superose-12) column, and hydrophobic (phenyl 5PW) column and finally the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). In the SDS-PAGE, one major band with a molecular weight of 56, 000 and one minor band with 15, 000 were stained with Coomassie Brilliant Blue and both showed DIF activity after extraction from gels. DIF had an isoelectric point of 5.8. In all purification steps, DIF activity for HL-60 and cytotoxic activity to the sublines of mouse L-929 were not able to be separated. Further, DIF was neutralized by antibody against lymphotoxin (LT) but not by antibody against tumor necrosis factor. These results indicate that the major DIF activity derived from MT-2 is LT.