Carnosic acid and promotion of monocytic differentiation of HL60-G cells initiated by other agents.
BACKGROUND
Carnosic acid is a plant-derived polyphenol food preservative with chemoprotective effects against carcinogens when tested in animals. Recently, we showed that carnosic acid potentiates the effects of 1alpha,25-dihydroxyvitamin D3 (1alpha,25[OH]2D3) and of all-trans-retinoic acid (ATRA) on differentiation of human leukemia cells. We now examine the mechanisms associated with carnosic acid-induced enhancement of cell differentiation (in subline HL60-G) initiated by 1alpha,25(OH)2D3, ATRA, or 12-O-tetradecanoylphorbol-13-acetate (TPA).


METHODS
We evaluated monocytic differentiation markers (CD11b, CD14, and monocytic serine esterase), cell cycle parameters, and cell proliferation rates after treatment of cells with different agents with or without carnosic acid. We also assessed the abundance of the vitamin D receptor (VDR), retinoid X receptor (RXR)-alpha, retinoic acid receptor (RAR)-alpha, and cell cycle-associated proteins by immunoblot analysis (p27, early growth response gene [EGR]-1, and p35Nck5a), the expression of corresponding genes by reverse transcription-polymerase chain reaction (RT-PCR), and the activity of VDR by electrophoretic mobility shift analysis. The two-sided nonparametric Kruskal-Wallis one-way analysis-of-variance test with Dunn's adjustment was used for statistical analyses.


RESULTS
Monocytic differentiation induced by low (1 nM) concentrations of 1alpha,25(OH)2D3, ATRA, or TPA was enhanced by carnosic acid (10 microM), as shown by the increased expression of monocytic serine esterase (P<.001, P<.001, and P =.043, respectively) and of CD11b (P =.008, P =.046, and P =.041, respectively). Increased expression of CD14 was seen only for 1alpha,25(OH)2D3 and ATRA (P =.009 and P =.048, respectively) and also for several cell cycle-associated proteins. Carnosic acid in combination with 1alpha,25(OH)2D3 and ATRA resulted in decreased cell proliferation and blocked the cell cycle transition from G1 to S phase (P<.05). Carnosic acid alone increased the expression of VDR and RXR-alpha, but the expression was greatly enhanced in the presence of 1alpha,25(OH)2D3 and ATRA. In combination with TPA, carnosic acid potentiated the expression of VDR and RAR-alpha.


CONCLUSIONS
Carnosic acid enhances a program of gene expression consistent with 1alpha,25(OH)2D3-, ATRA-, or TPA-induced monocytic differentiation of HL60-G cells.
Danilenko
M
M
Department of Pathology and Laboratory Medicine, UMD-New Jersey Medical School, Newark 07103, USA.
Wang
X
X
Studzinski
G P
GP
D045784:Abietanes; D000704:Analysis of Variance; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology*; D015153:Blotting, Western; D002117:Calcitriol / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000032:analysis; D004224:Diterpenes / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006098:Granulocytes / Q000187:drug effects*; D018922:HL-60 Cells; D006801:Humans; D009000:Monocytes / Q000187:drug effects*; D010936:Plant Extracts / Q000494:pharmacology*; D018167:Receptors, Calcitriol / Q000032:analysis; D018168:Receptors, Retinoic Acid / Q000008:administration & dosage / Q000032:analysis; D047488:Retinoid X Receptors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000032:analysis; D014212:Tretinoin / Q000494:pharmacology
D045784:Abietanes; D000970:Antineoplastic Agents; D000975:Antioxidants; D004268:DNA-Binding Proteins; D004224:Diterpenes; D010936:Plant Extracts; D018167:Receptors, Calcitriol; D018168:Receptors, Retinoic Acid; D047488:Retinoid X Receptors; D014157:Transcription Factors; D014212:Tretinoin; D002117:Calcitriol; C018381:salvin