title: Indirubin, a Chinese anti-leukaemia drug, promotes neutrophilic differentiation of human myelocytic leukaemia HL-60 cells.
abstract: Indirubin, a purple vegetable dye, is a traditional Chinese medicine for myelocytic leukaemia. Indirubin inhibits cyclin-dependent protein kinases (CDKs) and is present in human urine and serum. When indirubin was present during the neutrophilic differentiation of human myelocytic leukaemia HL-60 cells, it augmented superoxide production triggered by opsonized zymosan (OZ) by the terminally differentiated HL-60 cells. It also augmented the calcium response to OZ stimulation, and HL-60 cell chemotaxis evoked by interleukin-8 (IL-8, CXCL8) and formylpeptide. In addition, indirubin induced marked IL-8 release by the cells during differentiation and the cells differentiated with indirubin had typical neutrophilic properties, deformed nuclei and granules. Use of stable cloned HL-60 cells that contained a reporter vector for monitoring the activity of the transcription factor PU.1, which acts specifically at the stage of promyelocyte differentiation into neutrophils and monocytes, revealed that indirubin has a potent promoting activity on intracellular PU.1. Indirubin enhanced the expression of typical neutrophil proteins, including granulocyte-colony stimulating factor receptor, the beta2-integrin subunit CD18, the NADPH-oxidase subunit p47phox, and the IL-8 receptor CXCR1, all are controlled by PU.1. Indirubin also inhibited CDK2-dependent phosphorylation of retinoblastoma protein during neutrophilic differentiation. These results suggest that indirubin augments the neutrophilic differentiation of human myelocytic leukaemia HL-60 cells through inhibition of CDK2 and activation of PU.1.
authors.affiliation: National Institute of Health Sciences, Tokyo, Japan. ksuzuki@nihs.go.jp
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Suzuki
authors.forename: Reiko
authors.initials: R
authors.lastname: Adachi
authors.forename: Akiko
authors.initials: A
authors.lastname: Hirayama
authors.forename: Hidemi
authors.initials: H
authors.lastname: Watanabe
authors.forename: Saki
authors.initials: S
authors.lastname: Otani
authors.forename: Yuka
authors.initials: Y
authors.lastname: Watanabe
authors.forename: Tadashi
authors.initials: T
authors.lastname: Kasahara
mesh_terms: D018821:CD18 Antigens / Q000378:metabolism; D002454:Cell Differentiation; D002633:Chemotaxis; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D018922:HL-60 Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted; D015151:Immunoblotting; D007211:Indoles / Q000494:pharmacology; D016209:Interleukin-8 / Q000378:metabolism / Q000494:pharmacology; D007951:Leukemia, Myeloid / Q000473:pathology*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects*; D010766:Phosphorylation; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000378:metabolism; D023062:Receptors, Interleukin-8A / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism; D013268:Stimulation, Chemical; D013481:Superoxides / Q000378:metabolism
chemical_list: D018821:CD18 Antigens; D007211:Indoles; D016209:Interleukin-8; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D023062:Receptors, Interleukin-8A; D016160:Retinoblastoma Protein; D013481:Superoxides; D016179:Granulocyte Colony-Stimulating Factor; D009240:N-Formylmethionine Leucyl-Phenylalanine; D018844:Cyclin-Dependent Kinases; C027185:indirubin; D004121:Dimethyl Sulfoxide