KHYG-1

KHYG-1 Used to Study Enhanced Natural Killer Cytotoxicity

KHYG-1 is a valuable model for the study of enhanced cytotoxicity by NK cells. NK/NK T-cell lines KHYG-1, NK-92, YT, and SNT-8 were compared with a novel flow cytometric cytotoxicity assay under different culture conditions.

The cytotoxicity of KHYG-1 was compared to the other cell lines against K562, at a 3:1 and a 50:1 effector (E) to target (T) ratio, with 7 hours of co-incubation (Fig. 1). K562 cells were lysed to 46%, 0%, 1%, and 0%, by KHYG-1, NK-92, SNT-8, and YT, respectively, at a ratio of 3:1 and to 86%, 23%, 27%, and 12%, by KHYG-1, NK-92, SNT-8, and YT, at 50:1 (Fig. 1A and C). The target cells were 25% (KHYG-1), 4% (NK-92), 1% (SNT-8), and 0.5% (YT) positive for Annexin V at the 3:1 ratio and 36% (KHYG-1), 14% (NK-92), 8% (SNT-8), and 7% (YT) positive for Annexin V at 50:1 (Fig. 1B and D).

The cytotoxicity of KHYG-1 cultured under RPMI conditions was tested against other leukemia cell lines, including EM2, EM3, and HL60 in a 7-hour assay, at a 10:1 E: T ratio, with K562 as a positive control (Fig. 2). Target cells were lysed to 82%, 25%, 48%, and 31% for K562, EM2, EM3, and HL60, respectively (Fig. 2A). The frequency of Annexin-positive cells among K562, EM2, EM3, and HL60 was 43%, 20%, 26%, and 9%, respectively (Fig. 2B).

Fig. 1 KHYG-1 is a superior killer cell line against K562 under RPMI conditions. (Suck G, et al., 2005)

Fig. 2 KHYG-1 is cytotoxic against cell lines with relevance to leukemia. (Suck G, et al., 2005)

Shuterin Enhances the Cytotoxicity of the NK Leukemia Cell Line KHYG-1

NK cell therapy is an emerging tool for cancer immunotherapy. Therefore, primary NK cells should be expanded substantially, and their proliferation and cytotoxicity must be enhanced. Shuterin is a phytochemical isolated from Ficus thonningii. This study explored the possible capacity of shuterin to enhance the proliferation and activity of KHYG-1 cells (an NK leukemia cell line).

Fig. 3A shows the chemical structure of shuterin. KHYG-1 cells were treated with various doses (0, 1, 5, and 10 µM) of shuterin for 72 h and subjected to WST-8 assays (Fig. 3B). Shuterin substantially increased the proliferation of KHYG-1 cells. Shuterin markedly increased the activity of KHYG-1 cells and thus increased these cells' cytotoxicity to K562 cells. At an E: T ratio of 6:1, cytotoxicity increased from 40% to 54% within 2 h (Fig. 3C).

The secretion of IFN-γ from KHYG-1 cells was evaluated through an enzyme-linked immunosorbent assay. Shuterin markedly enhanced the secretion of IFN-γ in a dose-dependent manner (Fig. 3D). Furthermore, protein levels were analyzed by Western blotting in KHYG-1 cells treated with shuterin for 24 h and those left untreated. At a concentration of 10 μM, shuterin considerably increased granzyme A, granzyme B, FasL, and granulysin by 240%, 350%, 128%, and 187%, respectively, but not perforin (Fig. 3E, F).

Fig. 3 Shuterin enhanced the cytolytic activity of KHYG-1 cells. (Lin JT, et al., 2022)