Mechanisms of inhibiting human leukemia cell lines by serum of rats treated with compound banmao capsule

Abstract

Compound banmao capsule (CBC) is a traditional Chinese medicinal formula composed of extracts from 11 organisms. The present study investigated the mechanism of CBC on the biological behavior of human leukemia cell lines using seropharmacological methods. CBC-containing rat serum was prepared by intragastrical administration of CBC to rats. The proliferation of human leukemia HL60 and K562 cell lines was assayed by measuring cell viability with the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium method, while cell cycle distribution and the rate of apoptosis were evaluated with flow cytometry. The mRNA expression of vascular endothelial growth factor A (VEGF-A) and chemotactic and inflammatory genes in human leukemia cell lines was examined using reverse transcription quantitative-polymerase chain reaction methods. It was revealed that the proliferation of K562 and HL60 cells was significantly inhibited by the CBC-containing rat serum at 72 h. The CBC-containing serum also promoted the apoptosis of K562 and HL60 cell lines. The CBC-containing serum altered the cell cycle progression of K562 and HL60, increasing the proportion of the cells in G1 phase and decreasing the proportion of the cells in S phase. Attenuated expression of VEGF-A and a decreasing trend in the expression of chemotactic and inflammatory genes were identified following treatment with CBC-containing serum in HL60 and K562 cells. In conclusion, CBC-containing serum exerted an inhibitory effect on the growth of K562 and HL60 cells by decreasing cellular proliferation, promoting apoptosis and cell cycle arrest, and decreasing the expression of VEGF-A, and chemotactic and inflammatory genes.

Keywords: HL60; K562; chemotactic and inflammatory genes; seropharmacology; vascular endothelial growth factor A.

Figure 1.

Effects of CBC-containing rat serum on the viability of human leukemia cell lines (A) K562 and (B) HL60. ***P<0.01 vs. control serum-treated cells. CBC, compound banmao capsule. Error bars represent the standard error of absorbance values of 6 wells.

Figure 2.

Effects of CBC-containing rat serum on the apoptosis of human leukemia cell lines HL60 and K562. HL60 cells treated with (A) FBS (no treatment), (B) control rat serum or (C) CBC-containing rat serum; K562 cells treated with (D) FBS, (E) control rat serum or (F) CBC-containing rat serum. CBC, compound banmao capsule; FBS, fetal calf serum; PI, propidium iodide; FITC, fluorescein isothiocyanate.

Figure 3.

Effects of CBC-containing rat serum on the cell cycle of human leukemia cell lines HL60 and K562. Subsequent to treatment with CBC-containing or vehicle control rat serum, the cells were processed with cell cycle analysis by flow cytometry. K562 cells treated with (A) control rat serum or (B) CBC-containing rat serum; (C) histogram of the differences in cell cycle distribution between (A) and (B). HL60 cells treated with (D) control rat serum or (E) CBC-containing rat serum; (F) histogram of the differences in cell cycle distribution between (D) and (E). CBC, compound banmao capsule; ctrl, control.

Figure 4.

Effects of CBC-containing rat serum on the mRNA expression of VEGF-A in human leukemia cell lines. The differences in VEGF-A mRNA expression between cells treated with control rat serum or CBC-containing rat serum were assessed in (A) K562 cells and (B) HL60 cells. ***P<0.01 compared with the control. Error bars represent the standard error of triplicate wells. CBC, compound banmao capsule; VEGF-A, vascular endothelial growth factor A.

Figure 5.

Effects of CBC-containing rat serum on the expression of chemotactic and inflammatory genes in human leukemia cell lines. The fold changes of mRNA (CBC serum/control) in (A) HL60 cells and (B) K562 cells subsequent to CBC-containing serum treatment. CBC, compound banmao capsule; CCR, C-C motif chemokine receptor; CCL, C-C motif chemokine ligand; DDK, Dickkopf; HMGB, high mobility group box; CXCL, C-X-C motif chemokine ligand; DDX, DEAD box helicase; CCND, cyclin D1; IL, interleukin.