Differentiation Of HL-60 Cells Research Articles

Chromatin remodeling: recruitment of histone demethylase RBP2 by Madl for transcriptional repression of a Myc target gene, telomerase reverse transcriptase

The Myc/Max/Mad network transcription factors are known to govern target gene expression through recruiting histone acetyltransferases or deacetylases. In the present study, we show that Mad1 recruits the histone demethylase RBP2 to the Myc target telomerase reverse transcriptase (hTERT) gene promoter to repress transcription. With differentiation of leukemic HL60 cells, Mad1 and RBP2 were both up-regulated, interacted, and cooccupied the hTERT promoter accompanied by histone H3-K4 demethylation. In immortalized p493-6 B cells, shutting down c-Myc led to the accumulation of Mad1 and RBP2 at hTERT promoter and diminished hTERT mRNA expression. When RBP2 was depleted, hTERT expression was significantly enhanced, coupled with dissociation of RBP2 with and increased H3-K4 methylation at the hTERT promoter in p493-6 cells. Moreover, RBP2 and Mad1 were present on the hTERT promoter in human fibroblasts having a silent hTERT gene, and RBP2 depletion resulted in gene derepression. Taken together, Mad1 recruits RBP2 to the hTERT promoter that, in turn, demethylates H3-K4, thereby contributing to a stable repression of the hTERT gene in normal or differentiated malignant cells. Our findings reveal a novel mechanism through which the Myc/Max/Mad network proteins control their target gene transcription and provide insights into mechanisms underlying telomerase silencing and activation.

Novel events in mitchondria: Increase of poly(ADP-ribosyl)ation in human promyelocytic leukemia HL-60 cell line during differentiation by nicotinamide

Poly(ADP-ribosyl)ation, synthesized mainly by poly(ADP-ribose) polymerase-1 (PARP-1) using nicotinamide adenine dinucleotide as a substrate, has been identified as a post-translational modification of nuclear proteins in many eukaryotic cells. At present, studies on poly(ADP-ribosyl)ation have focused primarily on its roles in DNA repair, replication, transcription, and cell death. It has been suggested that poly(ADP-ribosyl)ation activity occurs in organelles, including mitochondria, but its role is not fully understood. On the other hand, it has been reported that PARP-1 and poly(ADP-ribosyl)ation play an important role in the differentiation of many cell types. Recently, we have demonstrated that niacin and its related compounds induced granulocytic differentiation in human promyelocytic leukemia HL-60 cells. In this study, we investigated changes in poly(ADP-ribosyl)ation levels during granulocytic differentiation by niacin, its related compounds and well-known granulocytic inducers. Results indicated no significant changes in expression levels and cellular localization of PARP-1 protein was observed in HL-60 cells during the differentiation. However, poly(ADP-ribosyl)ation was detected in cells treated with nicotinamide (NA) compared to the controls. Intriguingly, this phenomenon was detected in the mitochondrial fractions, but not in the nucleus. In addition, nicotinic acid, which is equivalent to NA induce neither differentiation nor the change of poly(ADP-ribosyl)ation. Because NA promotes the differentiation activities in several types of cells, these results provide important clues towards elucidating the mechanisms of NA. Additionally, these findings provide new information about mitochondrial poly(ADP-ribosyl)ation.

A fluorescence-based opsonophagocytosis assay to measure the functional activity of antibody to group B Streptococcus

An in vitro assay designed to measure the functional activity of vaccine-induced antibody is a necessary component of any vaccine development program. Because traditional efficacy studies of vaccines to prevent neonatal diseases caused by group B Streptococcus (GBS) are unlikely given the effectiveness of current antibiotics and screen-based surveillance practices, the ability to efficiently and effectively measure functional antibody responses may be of particular importance. GBS, like other encapsulated bacterial pathogens, are susceptible to opsonization by specific antibody and complement and subsequent killing by the host’s effector cells. The in vitro opsonophagocytosis and killing assay (OPA) mimics this in vivo defense strategy and has been used for decades to measure the functionality of natural and/or vaccine-induced GBS-specific antibody. Here we describe a fluorescence-based OPA (flOPA) that measures the ability of specific antibody to opsonize fixed, fluorescently labeled GBS or antigen-coated fluorescent microspheres for uptake by differentiated HL-60 cells in the presence of complement. Compared to the classical OPA, the flOPA is standardized with respect to effector cells, complement, and antigenic targets. The GBS flOPA is also less time-intensive and has the potential to measure antibody to multiple antigens simultaneously. Quantitative functional antibody determinations using the flOPA may serve as a surrogate measure of GBS vaccine effectiveness in lieu of traditional phase 3 efficacy trials.

ICAT as a potential enhancer of monocytic differentiation: implications from the comparative proteome analysis of the HL60 cell line stimulated by all‐trans retinoic acid and NSC67657

A novel sterol mesylate compound (NSC67657) was recently identified and reported by National Cancer Institute that could efficiently induce the differentiation of HL60 cells into monocytes in vitro and in vivo. The expression of many proteins would have been changed during the differentiation process, and some proteins may have played key roles in the differentiation of HL60 cell line induced by this drug. Therefore, we treated HL60 cells with NSC67657 and all-trans retinoic acid (ATRA) to identify the differentially expressed proteins and determine their functions in cellular differentiation. Of the 45 differentially expressed protein spots investigated, 24 were either elevated or decreased in both the monocytic and granulocytic differentiating HL60 cells, 8 showed significant changes only when induced by NSC67657, and 13 showed significant changes only when induced by ATRA. After verification by RT-PCR, Western blotting, and immunocytochemistry, only the protein ICAT was found to be elevated by NSC67657 treatment alone. Although the over-expression of ICAT is not sufficient to induce the differentiation of HL60 cells into monocytes, it did increase the proportion of CD14+ cells in cells pretreated with NSC67657. Successful application of multiple techniques including two-dimensional gel electrophoresis, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, Western blotting, and eukaryotic electroporation revealed that proteomic and molecular biological analyses provide valuable tools in drug development research.

Monocytic differentiation of leukemic HL‐60 cells induced by co‐treatment with TNF‐α and MK886 requires activation of pro‐apoptotic machinery

The block of hematopoietic differentiation program in acute myeloid leukemia cells can be overcome by differentiating agent like retinoic acid, but it has several side effects. A study of other differentiation signaling pathways is therefore useful to predict potential targets of anti-leukemic therapy. We demonstrated previously that the co-treatment of HL-60 cells with Tumor necrosis factor-alpha (TNF-alpha) (1 ng/mL) and inhibitor of 5-lipoxygenase MK886 (5 microm) potentiated both monocytic differentiation and apoptosis. In this study, we detected enhanced activation of three main types of mitogen-activated protein kinases (MAPKs) (p38, c-Jun amino-terminal kinase [JNK], extracellular signal-regulated kinase [ERK]), so we assessed their role in differentiation using appropriate pharmacologic inhibitors. The inhibition of pro-apoptotic MAPKs (p38 and JNK) suppressed the effect of MK886 + TNF-alpha co-treatment. On the other hand, down-regulation of pro-survival ERK pathway led to increased differentiation. Those effects were accompanied by increased activation of caspases in cells treated by MK886 + TNF-alpha. Pan-caspase inhibitor ZVAD-fmk significantly decreased both number of apoptotic and differentiated cells. The same effect was observed after inhibition of caspase 9, but not caspase 3 and 8. To conclude, we evidenced that the activation of apoptotic processes and pathways supporting apoptosis (p38 and JNK MAPKs) is required for the monocytic differentiation of HL-60 cells.

Human hemokinin-1 and human hemokinin-1(4–11), mammalian tachykinin peptides, suppress proliferation and induce differentiation in HL-60 cells

Human hemokinin-1 (h HK-1) and its truncated form h HK-1(4–11) are mammalian tachykinin peptides encoded by the TAC4 gene identified in human, and the biological functions of these peptides have not been well investigated. The tachykinins have shown immuno-regulatory activities in humans. In the present study, we investigated the effects of h HK-1 and h HK-1(4–11) on the proliferation and differentiation of a human promyelocyte leukemia cell line, HL-60. It is noteworthy that h HK-1 (1–300 μM) displayed inhibitory effects on the proliferation of HL-60 cells in a dose- and time-dependent manner. The effect of suppressing proliferation induced by these peptides was accompanied by an accumulation of cell cycle in the S phase. Moreover, this peptide induced differentiation of HL-60 cells by significantly increasing the NBT-reduction activity. The effects induced by h HK-1(4–11) on HL-60 cells were similar to that of h HK-1, indicating that it is the active fragment of h HK-1. However these effects induced by h HK-1 or h HK-1(4–11) were not antagonized by the NK 1 receptor antagonist SR140333 or the NK 2 receptor antagonist SR48968. All the results indicated that h HK-1 and h HK-1(4–11) were able to significantly inhibit proliferation and induce differentiation and S phase arrest of a human promyelocyte leukemia cell line HL-60, which may not be mediated through the activation of classical tachykinin NK 1 receptors and tachykinin NK 2 receptors. Our observations also implied that h HK-1 and h HK-1(4–11) could act as immunomodulatory factors in cancer chemotherapy.

5-Lipoxygenase inhibitors potentiate 1α,25-dihydroxyvitamin D3-induced monocytic differentiation by activating p38 MAPK pathway

The treatment of human promyelocytic leukemia cell lines HL-60, and to some extent NB-4, with 1alpha,25-dihydroxyvitamin D(3) (VD3) induces differentiation toward the monocytic/macrophage lineage, demonstrated by the increased expression of CD11b and CD14, and the production of opsonized zymosan particles (OZP)-stimulated reactive oxygen species (ROS). Moreover, in more sensitive HL-60 cells, increased expression of 5-lipoxygenase (5-LPO), Mcl-1, IkappaB, and c-Jun, accompanied by the activation of p38 MAPK, was detected. These VD3 effects on HL-60 cell differentiation were significantly potentiated by 5-LPO inhibitors MK-886 and AA-861 and were inverted by SB202190 (SB), a p38 MAPK inhibitor. The inhibition of differentiation by SB was demonstrated by a reduction of CD14 expression and by a decrease in OZP-activated ROS production. These results indicated that p38 MAPK pathway is involved in 5-LPO inhibitors-dependent potentiation of VD3-induced monocytic differentiation.

Flotillins Interact with PSGL-1 in Neutrophils and, upon Stimulation, Rapidly Organize into Membrane Domains Subsequently Accumulating in the Uropod

BackgroundNeutrophils polarize and migrate in response to chemokines. Different types of membrane microdomains (rafts) have been postulated to be present in rear and front of polarized leukocytes and disruption of rafts by cholesterol sequestration prevents leukocyte polarization. Reggie/flotillin-1 and -2 are two highly homologous proteins that are ubiquitously enriched in detergent resistant membranes and are thought to shape membrane microdomains by forming homo- and hetero-oligomers. It was the goal of this study to investigate dynamic membrane microdomain reorganization during neutrophil activation.Methodology/Principal FindingsWe show now, using immunofluorescence staining and co-immunoprecipitation, that endogenous flotillin-1 and -2 colocalize and associate in resting spherical and polarized primary neutrophils. Flotillins redistribute very early after chemoattractant stimulation, and form distinct caps in more than 90% of the neutrophils. At later time points flotillins accumulate in the uropod of polarized cells. Chemotactic peptide-induced redistribution and capping of flotillins requires integrity and dynamics of the actin cytoskeleton, but does not involve Rho-kinase dependent signaling related to formation of the uropod. Both flotillin isoforms are involved in the formation of this membrane domain, as uropod location of exogenously expressed flotillins is dramatically enhanced by co-overexpression of tagged flotillin-1 and -2 in differentiated HL-60 cells as compared to cells expressing only one tagged isoform.Flotillin-1 and -2 associate with P-selectin glycoprotein ligand 1 (PSGL-1) in resting and in stimulated neutrophils as shown by colocalization and co-immunoprecipitation. Neutrophils isolated from PSGL-1-deficient mice exhibit flotillin caps to the same extent as cells isolated from wild type animals, implying that PSGL-1 is not required for the formation of the flotillin caps. Finally we show that stimulus-dependent redistribution of other uropod-located proteins, CD43 and ezrin/radixin/moesin, occurs much slower than that of flotillins and PSGL-1.Conclusions/SignificanceThese results suggest that flotillin-rich actin-dependent membrane microdomains are importantly involved in neutrophil uropod formation and/or stabilization and organize uropod localization of PSGL-1.

Extracellular Hsp72, an endogenous DAMP, is released by virally infected airway epithelial cells and activates neutrophils via Toll-like receptor (TLR)-4

BackgroundNeutrophils play an important role in the pathophysiology of RSV, though RSV does not appear to directly activate neutrophils in the lower airways. Therefore locally produced cytokines or other molecules released by virally-infected airway epithelial cells are likely responsible for recruiting and activating neutrophils. Heat shock proteins (HSPs) are generally regarded as intracellular proteins acting as molecular chaperones; however, HSP72 can also be released from cells, and the implications of this release are not fully understood.MethodsHuman bronchial epithelial cells (16HBE14o-) were infected with RSV and Hsp72 levels were measured by Western blot and ELISA. Tracheal aspirates were obtained from critically ill children infected with RSV and analyzed for Hsp72 levels by ELISA. Primary human neutrophils and differentiated HL-60 cells were cultured with Hsp72 and supernatants analyzed for cytokine production. In some cases, cells were pretreated with polymyxin B prior to treatment with Hsp72. IκBα was assessed by Western blot and EMSA's were performed to determine NF-κB activation. HL-60 cells were pretreated with neutralizing antibody against TLR4 prior to Hsp72 treatment. Neutrophils were harvested from the bone marrow of wild type or TLR4-deficient mice prior to treatment with Hsp72.ResultsInfection of 16HBE14o- with RSV showed an induction of intracellular Hsp72 levels as well as extracellular release of Hsp72. Primary human neutrophils from normal donors and differentiated HL-60 cells treated with increasing concentrations of Hsp72 resulted in increased cytokine (IL-8 and TNFα) production. This effect was independent of the low levels of endotoxin in the Hsp72 preparation. Hsp72 mediated cytokine production via activation of NF-κB translocation and DNA binding. Using bone marrow-derived neutrophils from wild type and TLR4-mutant mice, we showed that Hsp72 directly activates neutrophil-derived cytokine production via the activation of TLR4.ConclusionCollectively these data suggest that extracellular Hsp72 is released from virally infected airway epithelial cells resulting in the recruitment and activation of neutrophils.

Retinoic acid inhibits sodium butyrate-induced monocytic differentiation of HL60 cells while synergistically inducing granulocytoid differentiation

The human myeloid leukemia cell line HL60 is an in vitro model to study myeloid differentiation. HL60 cells differentiate along different cell type lineages in response to a variety of compounds. The direction of differentiation is usually inducer-specific. However, the response of HL60 cells to sodium n-butyrate (NaB) is pleiotropic. NaB induces HL60 along the monocytic, neutrophilic, eosinophilic, and basophilic pathways. In this study we saw that physiologic concentrations of all-trans-retinoic acid (RA) switched the direction of NaB-induced differentiation from monocytic to granulocytic. We showed previously (Breitman & He, Cancer Res 1990: 50: 6268-6273) that combinations of RA and NaB synergistically induce HL60 to cells that reduce nitroblue tetrazolium. The present study shows that this synergy was even greater if the parameter measured was mature granulocytes. Our results raise the possibility that the endogenous RA in the serum used to grow cells in culture may affect the direction of differentiation of HL60 cells induced by NaB. Furthermore, our results may provide additional rationale for the use of combinations of RA and NaB in the treatment of some leukemias.

Deamino-hydroxy-phoslactomycin B, a biosynthetic precursor of phoslactomycin, induces myeloid differentiation in HL-60 cells

During the screening for novel differentiation inducers, we found that a culture broth of Streptomyces sp. HK-803 induced myeloid differentiation of HL-60 cells. The active substance was identified as deamino-hydroxy-phoslactomycin B (HPLM) by mass spectrometry, and synthesized HPLM also induced the differentiation of HL-60 cells. HPLM showed greater inhibition of protein phosphatase 2A (PP2A) activity than phoslactomycin B (PLMB); however, PLMB and okadaic acid did not induce differentiation. Moreover, treatment with ATRA and 1α, 25(OH) 2D 3 induced retinoic acid receptor-β and 1α, 25(OH) 2 D 3 24-hydroxylase, respectively, whereas HPLM did not, suggesting that HPLM is a novel differentiation inducer.

Gi-Dependent Cell Signaling Responses of the Human P2Y14Receptor in Model Cell Systems

Eight G protein-coupled receptors comprise the P2Y receptor family of cell signaling proteins. The goal of the current study was to define native cell signaling pathways regulated by the uridine nucleotide sugar-activated P2Y(14) receptor (P2Y(14)-R). The P2Y(14)-R was stably expressed in human embryonic kidney (HEK) 293 and C6 rat glioma cells by retroviral infection. Nucleotide sugar-dependent P2Y(14)-R activation was examined by measuring inhibition of forskolin-stimulated cAMP accumulation. The effect of P2Y(14)-R activation on mitogen-activated protein kinase signaling also was studied in P2Y(14)-HEK293 cells and in differentiated HL-60 human myeloid leukemia cells. UDP-Glc, UDP-galactose, UDP-glucuronic acid, and UDP-N-acetylglucosamine promoted inhibition of forskolin-stimulated cAMP accumulation in P2Y(14)-HEK293 and P2Y(14)-C6 cells, and this signaling effect was abolished by pretreatment of cells with pertussis toxin. Inhibition of cAMP formation by nucleotide sugars also was observed in direct assays of adenylyl cyclase activity in membranes prepared from P2Y(14)-C6 cells. UDP-Glc promoted concentration-dependent and pertussis toxin-sensitive extracellular signal-regulated kinase (ERK) 1/2 phosphorylation in P2Y(14)-HEK293 cells. P2Y(14)-R mRNA was not observed in wild-type HL-60 cells but was readily detected in dimethyl sulfoxide-differentiated cells. Consistent with this observation, no effect of UDP-Glc was observed in wild-type HL-60 cells, but UDP-Glc-promoted pertussis toxin-sensitive activation of ERK1/2 occurred after differentiation. These results illustrate that the human P2Y(14)-R signals through G(i) to inhibit adenylyl cyclase, and P2Y(14)-R activation also leads to ERK1/2 activation. This work also identifies two stable P2Y(14)-R-expressing cell lines and differentiated HL-60 cells as model systems for the study of P2Y(14)-R-dependent signal transduction.

2-(1-Hydroxethyl)-4,8-dihydrobenzo[1,2-b:5,4-b′]dithiophene-4,8-dione (BTP-11) enhances the ATRA-induced differentiation in human leukemia HL-60 cells

2-(1-Hydroxethyl)-4,8-dihydrobenzo[1,2-b:5,4-b′]dithiophene-4,8-dione (BTP-11) is a potent enhancer for all-trans retinoic acid (ATRA)-induced differentiation in HL-60 cells. Combination of BTP-11 and ATRA cut down the concentration of ATRA significantly, and that BTP-11 promoted the progression of ATRA-induced into the terminal granulocytic differentiation. Further, Western blot analysis revealed that combination of BTP-11 and ATRA decreased cyclin D/CDK4 and increased C/EBPɛ protein expression to arrest the cells into G0/G1 phase leading to granulocytic maturation. These results confirmed that BTP-11 is a potent enhancer for ATRA-induced differentiation of HL-60 cells, and the great developmental potential of BTP-11 will be expected.

Retinoic acid induces nuclear accumulation of Raf1 during differentiation of HL-60 cells

All trans-retinoic acid (RA) is a standard therapeutic agent used in differentiation induction therapy treatment of acute promyelocytic leukemia (APL). RA and its metabolites use a diverse set of signal transduction pathways during the differentiation program. In addition to the direct transcriptional targets of the nuclear RAR and RXR receptors, signals derived from membrane receptors and the Raf–MEK–ERK pathway are required. Raf1 phosphorylation and the prolonged activation of Raf1 persisting during the entire differentiation process are required for RA-dependent differentiation of HL-60 cells. Here we identify a nuclear redistribution of Raf1 during the RA-induced differentiation of HL-60 cells. In addition, the nuclear accumulation of Raf1 correlates with an increase in Raf1 phosphorylated at serine 621. The serine 621 phosphorylated Raf1 is predominantly localized in the nucleus. The RA-dependent nuclear accumulation of Raf1 suggests a novel nuclear role for Raf1 during the differentiation process.

Nargenicin enhances 1,25-dihydroxyvitamin D 3- and all- trans retinoic acid-induced leukemia cell differentiation via PKCβI/MAPK pathways

A major goal in the treatment of acute myeloid leukemia (AML) is to achieve terminal differentiation and prevent drug resistance and side effects. Combined treatment with low doses of ATRA or 1,25-(OH) 2D 3 that do not induce toxicity with another drug is one useful strategy for the treatment of AML. Actinomycetes are the well known sources of antibiotics and bioactive molecules. Previously, we isolated nargenicin from the culture broth of an actinomycete isolate, Nocardia sp. CS682. In this study, we evaluated the effects of nargenicin on cellular differentiation in a human myeloid leukemia HL-60 cell system. Nargenicin inhibited cell proliferation and induced HL-60 cell differentiation when administered in combination with 1,25-(OH) 2D 3 or ATRA. In addition, western blot analyses and kinase inhibitor studies demonstrated that nargenicin primarily enhanced leukemia cell differentiation via PKCβ1/MAPK pathways. The results of this study indicate that nargenicin has the ability to induce differentiation and suggest that it may be useful for the treatment of neoplastic diseases.

Expression and subcellular localization of syntaxin 11 in human neutrophils

Syntaxin 11 mutations lead to familial hemophagocytic lymphohistiocytosis (FHL), characterized by uncontrolled hyperinflammation. This study examines the expression and subcellular localization of syntaxin 11 in human neutrophils as major inflammatory cells. The materials included human peripheral blood neutrophils, HL-60 cells. The methods used were RT-PCR, Western blot, immunocytochemistry, subcellular fractionation, HL-60 cell differentiation. We have found that human peripheral blood neutrophils express syntaxin 11 mRNA and protein. Syntaxin 11 was upregulated during neutrophil differentiation of HL-60 cells. Syntaxin 11, identified as a membrane-bound protein, was broadly located in the plasma membrane and granules, with a predominant location in azurophilic granules of resting human neutrophils. A secondary location of syntaxin 11 was in specific and tertiary granules, which resulted in translocation to the plasma membrane on cell activation conditions that promoted the release of these organelles. These data indicate that human neutrophils express syntaxin 11 and call attention to the possible involvement of neutrophils in familial hemophagocytic lymphohistiocytosis pathology.

MicroRNAs181 regulate the expression of p27Kip1 in human myeloid leukemia cells induced to differentiate by 1,25-dihydroxyvitamin D3

Human myeloid leukemia cells exposed to 1,25-dihydroxyvitamin D3 (1,25D), a major cancer chemopreventive agent, acquire features of normal monocytes and arrest in the G1 phase of the cell cycle, due to the upregulation of p27Kip1 and p21Cip1, but the mechanism is not clear. Here evidence is provided that an exposure of HL60 and U937 cells to low (1-10 nM) concentrations of 1,25D decreases the expression of miR181a and miR181b in a concentration and time-dependent manner. Since the predicted miR181 targets include the 3’-UTR of p27Kip1, we expressed pre-miR181a in these cells, and found that the elevation of cellular miR181a levels abrogates the 1,25D-induced increase in p27Kip1 at both mRNA and protein levels. In contrast, transfection of pre-miR181a resulted in a slight elevation of p21Cip1 expression. Importantly, transfection of pre-miR181a blunted the effect of 1,25D on the expression of monocytic differentiation markers, and reduced the G1 block in 1,25D-treated cells, while transfection of anti-miR181a increased 1,25D-induced differentiation. Together, these data show that miR181a participates in 1,25D-induced differentiation of HL60 and U937 cells, and suggest that a high constitutive expression of members of miR181 family may contribute to the malignant phenotype in the myeloid lineage.

Gene expression analysis of human promyelocytic leukemia HL-60 cell differentiation and cytotoxicity induced by natural and synthetic retinoids

Aims This study analyzed gene expression profiles of human promyelocytic leukemia HL-60 cells treated with natural and synthetic retinoids ( ATRA, RII and R9158), in an attempt to investigate the structure–function relationship of the retinoids in inducing cell differentiation and cytotoxicity. Main methods Flow cytometry was used to determine cell cycle changes in HL-60 cells following treatment (1.0 μM) with natural and synthetic retinoids ( ATRA, RII and R9158), and cDNA microarrays were used to monitor the gene expression profiles of HL-60 cells treated with the various retinoids. Key findings Consistent with retinoid-induced cell differentiation, treatment with these three retinoids correlated with an increase in the percentage of cells arrested in the G1/G0 phase of the cell cycle. Microarray analysis showed upregulation of known differentiation genes, adhesion molecules, and the oxidase activation pathway following retinoid treatment. Differential expression of several genes was observed in HL-60 cells treated with the three retinoids. For example, tissue remodeling protein genes, ubiquitin genes, and signal transduction genes were highly expressed in ATRA- and R9158-treated HL-60 cells, but remained unchanged in HL-60 cells treated with RII. Significance The above findings suggest that the differentiation of HL-60 cells induced by the three retinoids occurs through similar pathways, and that there exists a structure–function relationship regarding retinoids and the induction of cell differentiation and cytotoxicity.

Antitumor and Immunomodulating Effects of a Fragment of HL-60 Cell Differentiation Factor in Mice with Lewis Pulmonary Carcinoma

A six-member HLDF6 peptide, fragment of HL-60 cell differentiation factor, exhibited antimetastatic and immunomodulating effects.

Fibronectin promotes the phorbol 12-myristate 13-acetate-induced macrophage differentiation in myeloid leukemia cells

The extracellular matrix plays a critical role in macrophage maturation. In this study, the HL60 cell line was used as a model of leukemic myeloid cell differentiation. We assessed the ability of HL60 cells cultured on fibronectin substratum prior to phorbol 12-myristate 13-acetate (PMA) induction to differentiate into terminally differentiated macrophages. Beside their distinctive macrophage morphology, they expressed antigen receptors CD14, TLR2, TLR4 and CD68, and displayed enhanced phagocytic activity and production of reactive oxygen species. Expression of CD13, CD33, CD15 and alpha-naphthyl-acetate esterase was also maintained, however, differentiated HL60 cells were HLA-DR and CD1a negative. Here, we describe the enhanced capacity of fibronectin-adherent HL60 cells to differentiate into macrophages in response to PMA.