Ascorbic acid inhibits TPA-induced HL‑60 cell differentiation by decreasing cellular H2O2 and ERK phosphorylation

  • Authors:
    • Giou‑Teng Yiang
    • Jen‑Ni Chen
    • Tsai‑Kun Wu
    • Hsueh‑Fang Wang
    • Yu‑Ting Hung
    • Wei‑Jung Chang
    • Chinshuh Chen
    • Chyou‑Wei Wei
    • Yung‑Luen Yu
  • View Affiliations

  • Published online on: July 20, 2015     https://doi.org/10.3892/mmr.2015.4091
  • Pages: 5501-5507
Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )


Abstract

Retinoic acid (RA), vitamin D and 12-O‑tetradecanoyl phorbol-13-acetate (TPA) can induce HL-60 cells to differentiate into granulocytes, monocytes and macrophages, respectively. Similar to RA and vitamin D, ascorbic acid also belongs to the vitamin family. High‑dose ascorbic acid (>100 µM) induces HL‑60 cell apoptosis and induces a small fraction of HL‑60 cells to express the granulocyte marker, CD66b. In addition, ascorbic acid exerts an anti‑oxidative stress function. Oxidative stress is required for HL‑60 cell differentiation following treatment with TPA, however, the effect of ascorbic acid on HL‑60 cell differentiation in combination with TPA treatment remains to be fully elucidated. The aim of the present study was to investigate the cellular effects of ascorbic acid treatment on TPA-differentiated HL-60 cells. TPA-differentiated HL-60 cells were used for this investigation, this study and the levels of cellular hydrogen peroxide (H2O2), caspase activity and ERK phosphorylation were determined following combined treatment with TPA and ascorbic acid. The results demonstrated that low‑dose ascorbic acid (5 µM) reduced the cellular levels of H2O2 and inhibited the differentiation of HL‑60 cells into macrophages following treatment with TPA. In addition, the results of the present study further demonstrated that low‑dose ascorbic acid inactivates the ERK phosphorylation pathway, which inhibited HL‑60 cell differentiation following treatment with TPA.

Introduction

HL-60 cells, belonging to human leukemia cells, have been widely used for differentiation investigations. Previous studies have revealed that HL-60 cells can be induced to differentiate into granulocytes, monocytes and macrophages by treating the cells with various agents, including retinoic acid (RA), dimethyl sulphoxide (DMSO), vitamin D and 12-O-tetradecanoyl phorbol-13-acetate (TPA) (1,2). Several previous studies have indicated that the mitogen-activated protein kinase (MAPK) signaling pathways, c-Jun N-terminal kinase (JNK), p38 and extracellular signal-regulated kinase (ERK), are important for HL-60 cell differentiation (37). RA and DMSO can induce HL-60 cells to differentiate into granulocytes via the ERK phosphorylation signaling pathway (810), vitamin D can induce HL-60 cells to differentiate into monocytes via the EKR, JNK and p38 phosphorylation signaling pathways (4,11,12) and TPA can induce HL-60 cells to differentiate into macrophages via the ERK phosphorylation signaling pathway (13,14). In addition, previous studies have demonstrated that ERK5 is associated with vitamin D-differentiated HL-60 cells, while ERK/1/2 is associated with TPA-differentiated and RA-differentiated HL-60 cells (1417). These previous reports indicated that ERK5 phosphorylation is required to differentiate HL-60 cells into monocytes, while ERK1/2 is required to differentiate HL-60 cells into granulocytes and macrophages.

The MAPK signaling pathways, protein kinase C (PKC) and oxidative stress may also be associated with HL-60 differentiation (1821). The activation of PKC is observed in RA-, vitamin D- and TPA-differentiated HL-60 cells (13,14,22,23). However, oxidative stress can affect HL-60 cell differentiation. A previous study revealed that antioxidants, catalase, superoxide dismutase and N-acetyl cysteine increase the differentiation rate of vitamin D-treated HL-60 cells (24). However, compared with vitamin D-treated HL-60 cells, antioxidant inhibits cell differentiation in TPA-treated HL-60 cells (25). Therefore, oxidative stress exerts a dual role to promote vitamin D-differentiated cells and to inhibit TPA-differentiated cells.

It is well known that RA and vitamin D can induce HL-60 cells to differentiate into granulocytes and monocytes, respectively. As with RA and vitamin D, ascorbic acid is also a type of vitamin. Previous studies have demonstrated that ascorbic acid can activate the ERK signaling pathway to induce progenitor cell differentiation (26,27). Additionally, several studies have demonstrated that high-doses of ascorbic acid (>100 μM) can activate a caspase cascade to promote radiation-induced and etoposide-induced apoptosis in HL-60 cells (28,29). A previous study also demonstrated that high-doses of ascorbic can induce HL-60 cell apoptosis and induce a fraction of HL-60 cells to express the granulocyte marker, CD 66b (30). By contrast, low-doses of ascorbic acid decreases levels of cellular H2O2 and protects HL-60 cells against X ray- and As2O3-induced apoptosis (3134). However, whether ascorbic acid affects TPA-differentiated HL-60 cells remains to be elucidated.

A previous study demonstrated that H2O2 may be a secondary messenger associated with cell differentiation (25). Several studies have demonstrated that ascorbic acid exerts anti-oxidative stress functions (3538) and a previous study demonstrated that ascorbic acid decreases levels of cellular H2O2 (39). Previous studies have also reported that the ERK pathway is required for TPA-differentiated HL-60 cells (13,14).

Therefore, the aim of the present study was to determine the cellular effects of treatment with ascorbic acid on TPA-differentiated HL-60 cells.

Materials and methods

Chemicals

An MTT assay kit was purchased from Bio Basic Inc. (Markham, ON, Canada). TPA, ascorbic acid and luminol were purchased from Sigma-Aldrich (St. Louis, MO, USA). Ac-DEVD-pNA, a Caspase-3-like substrate, Ac-IETD-pNA, a caspase-8 substrate, and Ac-LEHD-pNA, a caspase-9 substrate, were purchased from Anaspec (San Jose, CA, USA). Fetal bovine serum, RPMI-1640 media, non-essential amino acid, L-glutamine and penicillin/streptomycin were purchased from Gibco Life Technologies (Carlsbad, CA, USA).

Cell line and cell culture

The HL-60 cells were purchased from Bioresources Collection and Research Center (Hsin Chu, Taiwan) and were cultured in Dulbecco's modified Eagle's media, containing 10% fetal bovine serum, 2 mM L-glutamine, 100 IU/ml penicillin/streptomycin and 0.1 mM non-essential amino acids. The cells were maintained in a humidified atmosphere containing 5% CO2 at 37°C.

Cell survival rate assay

A total of 3,000 cells were cultured in each well of a 96-well culture dish. The survival rates of the cells in the control group (non-ascorbic acid treated-cells) and the experimental groups (5 μM and 5 mM ascorbic acid-treated cells) were determined for 96 h at 37°C. Every 24 h, the cells were treated using an MTT assay kit, according to the manufacturer's instructions. Following incubation for 3 h, the absorbance (570 nm) was measured using a multi-well enzyme-linked immunosorbent assay reader (SpectraMax Paradigm Multi-Mode Microplate Reader; Molecular Devices, Sunnyvale, CA, USA). The cell survival rate was calculated using the following formula: Absorbanceexperimental group/Absorbancecontrol group × 100%.

Caspase activity assay

Caspase activities in the cells were determined using a substrate cleavage assay, as previous described (40,41). Briefly, the cells were treated with lysis buffer, containing 50 mM Tris-HCl, 120 mM NaCl, 1 mM EDTA, 1% NP-40 (pH 7.5) and protease inhibitors. The cell pellets were collected by centrifugation at 15,000 × g for 30 min at 4°C and the quantity of protein was determined using a Bradford assay (Bio-Rad Laboratories, Inc., Hercules, CA, USA). Subsequently, 40 μl of the cell lysates (80 μg total protein) were prepared in 158 μl reaction buffer, containing 20% glycerol, 0.5 mM EDTA, 5 mM dithiothreitol, 100 mM HEPES (pH 7.5) and 2 μl fluorogenic caspase substrate (Ac-LEHD-pNA, Ac-DEVD-pNA or Ac-IETD-pNA). The total solution was incubated at 37°C in the dark. Following incubation for 6 h, fluorogenic substrate cleavage was determined at 405 nm using a FLx800™ fluorescence microplate reader (Bio-Tek Instruments, Inc., Winooski, VT, USA). The fold increase of caspase activity was calculated with the following formula: (Absorbanceexperimental group − Absorbancecontrol group) / Absorbancecontrol group.

Measurement of H2O2

The levels of H2O2 in the cells was determined using a lucigenin-amplified method, as described previously (39,42,43). Briefly, the sample (200 μl, containing 104 cells) was added to 0.2 mmol/l luminol solution (100 ml). The mixture was then analized using a chemiluminescence analyzing system (CLA-FSI; Tohoku Electronic Industrial Co., Ltd., Sendal, Japan).

Observation of cell morphology and suspension cell counts

Undifferentiated HL-60 cells grow as suspension cells and TPA-differentiated HL-60 cells (macrophages) grow as attached cells (14,44). The morphologies of the suspension cells and attached cells were observed under a phase-contrast microscope (Olympus CK40, Olypmus Corporation, Tokyo, Japan; magnification, ×200). The suspended cells located in the media were collected using a pipette, whereas the attached cells remained in the bottom of the culture dish. The cells in suspension were counted using a trypan blue exclusion assay (0.4% in PBS), as described previously (45). Briefly, the media containing the suspended cells were mixed with trypan blue and placed in a CBC Customized Logo Hemocytometer Blood Counting Chamber (VIC Science, Xixiang City, China). The number of cells were then counted under a light microscope (Olympus, CK40; Olympus Corporation).

Western blotting

The cells were treated with lysis buffer and centrifuged at 16,000 × g for 10 min at 4°C. The proteins were located in the supernatant layer and were collected, concentrated and determined using a Bradford assay (Bio-Rad Laboratories, Inc.). Equal quantities of protein were separated on a 13.3% SDS-polyacrylamide gel (GHE320 Mini-STD Vertical Gel Electrophoresis Tank; Genepure Technology, Co., Ltd, Taichung, Taiwan). Following separation, the proteins were transferred onto a polyvinylidene fluoride membrane (EMD Millipore, Billerica, MA, USA). The membranes were placed in phosphate-buffered saline (PBS) containing 5% non-fat milk. Following incubation for 2 h at 25°C, the membranes were washed with PBS. The membranes were further incubated in PBS buffer, containing 5% non-fat milk with primary anti-human monoclonal antibodies to ERK (cat. no. 4965) and p-ERK (cat. no. 4370) (1:400; Cell Signaling Technology, Inc., Danvers, MA, USA) for 2 h at 25°C. Following incubation, the membranes were washed with PBS and incubated with secondary mouse anti-human antibodies (1:2,000; cat. no. 10702-MM01E-50; Sino Biological Inc. Beijing, China) for 1 h at 25°C. The proteins were detected using Western Lightning Chemiluminescence Reagent Plus (PerkinElmer, Waltham, MA, USA)

Statistical analysis

The data were calculated from four independent experiments and are presented as the mean ± standard deviation. Student's t-test was used to analyze the statistical differences. P<0.05 was considered to indicate a statistically significant difference.

Results

High-dose ascorbic acid inhibits HL-60 cell growth, whereas a low-dose does not

Previous studies have demonstrated that a high-dose of ascorbic acid can increase radiation-induced and etoposide-induced apoptosis in HL-60 cells (28,29). Similar to these studies, the present study demonstrated that a high-dose (5 μM) of ascorbic acid inhibited cell growth in the HL-60 cells, whereas a low-dose (5 μM) had no affect on the growth of the HL-60 cells (Fig. 1). As shown in Fig. 1, the cell survival rate was <50% in the high-dose ascorbic acid-treated HL-60 cells at 72 h, however, the survival rate was >90% in the low-dose ascorbic acid-treated HL-60 cells at 96 h. Whether ascorbic acid activates caspase death signals in the HL-60 cells was subsequently investigated. The results demonstrated that activation of caspase-8, caspase-9 and caspase-3 occurred in the high-dose ascorbic acid-treated HL-60 cells, but not in low-dose ascorbic acid-treated HL-60 cells (Fig. 2). These findings indicated that a high-dose of ascorbic acid exerted antitumor activities in the HL-60 cells.

Low-dose ascorbic acid reduces cellular levels of H2O2 in TPA-treated HL-60 cells

H2O2 is important in cell differentiation (25). The present study demonstrated that the levels of H2O2 increased in the TPA-differentiated HL-60 cells, compared with the TPA-treated HL-60 cells (P<0.05; Fig. 3), which suggested that H2O2 may be associated with HL-60 cells differentiation by TPA. Anti-oxidative functions of ascorbic acid have been demonstrated (3538), therefore, the present study further determined whether ascorbic acid reduces the levels of H2O2 in TPA-treated HL-60 cells. The results revealed that a low-dose of ascorbic acid inhibited the increased levels of H2O2 levels the TPA-treated HL-60 cells (Fig. 3).

Pretreatment with ascorbic acid inhibits the differentiation of HL-60 cells into macrophages following TPA treatment

As shown in Fig. 3, a low-dose of ascorbic acid reduced the levels of H2O2 in the TPA-treated HL-60 cells. In addition, a previous study demonstrated that H2O2 may be an important messenger for cell differentiation (25). Therefore, whether low-dose ascorbic acid inhibits the differentiation of HL-60 cells into macrophages treated with TPA was determined. Previous studies have revealed that TPA-differentiated HL-60 cells (macrophages) are attached cells, whereas the undifferentiated HL-60 cells are suspensed (14,44). The present study assessed the morphology of the cells using a phase contrast microscope, and observed that the control HL-60 cells were in suspension (Fig. 4A) and the TPA-treated HL-60 cells were attached (Fig. 4B). These data suggested that TPA induced the HL-60 cells to differentiate into macrophages. In addition, suspended cells were observed in the TPA-treated HL-60 cells pretreated with ascorbic acid (Fig. 4C), whereas attached cells were observed in the TPA-treated HL-60 cells post-treated with ascorbic acid (Fig. 4D). These data indicated that ascorbic acid pretreatment inhibited the TPA-induced differentiation of HL-60 cells into macrophages, however, post-treatment did not inhibit the ability of TPA to induce HL-60 cell differentiation into macrophages. The number of cells in suspension were also quantified in the present study (Fig. 5). There were ~35,000 suspended cells in the control group and the TPA + ascorbic acid pretreatment group, however, very few suspended cells were observed in the TPA-treated group and the TPA + ascorbic acid post-treatment group, compared with the control group (P<0.05; Fig. 5) Taken together, these results suggested that pretreatment with ascorbic acid inhibited the ability of TPA to induce the differentiation of HL-60 cells into macrophages.

Ascorbic acid inhibits TPA-induced HL-60 cell differentiation via ERK phosphorylation

Previous studies have revealed that the induction of HL-60 cells to differentiate into macrophages by TPA requires ERK phosphorylation (13,14). These studies demonstrated that the inhibition of p-ERK inhibits TPA-induced HL-60 cell differentiation. In the present study, as shown in Figs. 4 and 5, pretreatment with ascorbic acid inhibited the differentiation of the HL-60 cells into macrophages following TPA treatment. Whether pretreatment with ascorbic acid inhibited TPA-differentiated HL-60 cells via ERK phosphorylation was subsequently investigated, and western blotting revealed that TPA induced an increase in the protein expression of p-ERK (Fig. 6; lane 2). In addition, pretreatment with ascorbic acid reduced the expression of p-ERK in the TPA-treated HL-60 cells (Fig. 6; lane 3). This data suggested that ascorbic acid inhibited the ability of TPA to induce HL-60 cell differentiation via ERK phosphorylation.

Discussion

Previous studies have demonstrated that TPA induces ERK phosphorylation, which in turn causes HL-60 cells to differentiate into macrophages (13,14). In addition, a previous study indicated that H2O2 accumulation is important for macrophage differentiation following TPA treatment (25). Similar to previous findings, the present study demonstrated that TPA induced an increase in the levels of H2O2 and induced ERK phos-phorylation (Figs. 3 and 6). These results suggested that TPA induced HL-60 cells to differentiate into macrophages via the accumulation of H2O2 and phosphorylation of ERK. However, the association between H2O2 and the phosphorylation of ERK remains to be elucidated. The present study demonstrated that pretreatment with ascorbic acid reduced TPA-induced H2O2 accumulation (Fig. 3) and inhibited TPA-induced HL-60 cell differentiation into macrophages (Figs. 4 and 5). However, the data also revealed that post-treatment with ascorbic acid did not have an inhibitory effect of TPA (Figs. 4 and 5). The results of the present study indicated H2O2 accumulation as an upstream signal, affecting HL-60 cell differentiation by TPA at an early stage. In addition, several previous studies have demonstrated that H2O2 induces the phosphorylation of EKR in various types of cell (4648). Therefore, the present study indicated that TPA induced an increase in the levels of H2O2 initially, and subsequently induced the phosphorylation of ERK, leading to HL-60 cell differentiation. However, pretreatment with ascorbic acid inhibited TPA-induced H2O2 accumulation at an early stage, preventing HL-60 cell differentiation.

The dual role of ascorbic acid in promoting cell death and preventing cell damage have been previously reported. Generally, a high-dose of ascorbic acid induces cell cytotoxicity (28,29), whereas a low-dose of ascorbic acid protects cells against oxidative stress-induced damage (3234). Similar to these studies, the present study demonstrated that a high-dose of ascorbic acid inhibited cell growth and activated the caspase-death pathway in the HL-60 cells (Figs. 1 and 2). However, a low-dose of ascorbic acid reduced TPA-induced increases in H2O2 levels (Fig. 3). Therefore, high-dose and low-dose ascorbic acids exerted different mechanisms to affect cell growth. Previous studies have also reported that ascorbic acid induces ERK phosphorylation in various types of cell, including acute myeloid leukemia cells and human endothelial cells (49,50). By contrast, ascorbic acid inhibits ERK phosphorylation in human dermal fibroblasts (51). The present study demonstrated that a low-dose of ascorbic acid inhibited the TPA-induced phosphorylation of ERK (Fig. 6; lane 3). Therefore, it was hypothesized that ascorbic acid induces different signaling pathways to affect cell growth in a dose-dependent and cell-dependent manner.

Regarding the association between ascorbic acid and cell differentiation, several studies have demonstrated that ascorbic acid can promote cell differentiation in various types of cell, including periodontal ligament progenitor cells, osteoblastic cells and embryonic stem cells (26,27,5256). However, the present study demonstrated that low-doses of ascorbic acid inhibited the HL-60 cells from differentiating into macrophages following TPA treatment. The possible reason may be that TPA-induced cell differentiation requires increases in cellular oxidative stress (25), while ascorbic acid can reduce cellular H2O2 levels to inhibit TPA-treated cells. Another possible reason is that ascorbic acid induces a small fraction of HL-60 cells to express the granulocyte marker, CD66b (30) and induces a small fraction of HL-60 cells to differentiate into granulocytes, therefore, inhibiting the differentiation of HL-60 cells into macrophages, induced by TPA.

In conclusion, the present study demonstrated for the first time, to the best of our knowledge, that low-doses of ascorbic acid inhibited TPA-treated HL-60 cells from differentiating into macrophages by decreasing TPA-induced levels of H2O2 and ERK phosphorylation.

Acknowledgments

This study was supported by the Ministry of Science and Technology (grant. no. MOST103 2320-B-039–052-MY3), the National Health Research Institute (grant. no. N HRI-EX102–10245BI) and Taipei Tzu Chi Hospital (grant. nos. TCRD-TPE-102–26 and TCRD-TPE-103–48).

References

1 

Trayner ID, Bustorff T, Etches AE, Mufti GJ, Foss Y and Farzaneh F: Changes in antigen expression on differentiating HL60 cells treated with dimethylsulphoxide, all-trans retinoic acid, alpha1,25-dihydroxyvitamin D3 or 12-O-tetradecanoyl phorbol-13-acetate. Leuk Res. 22:537–547. 1998. View Article : Google Scholar : PubMed/NCBI

2 

Zylber-Katz E and Glazer RI: Phospholipid- and Ca2+-dependent protein kinase activity and protein phosphorylation patterns in the differentiation of human promyelocytic leukemia cell line HL-60. Cancer Res. 45:5159–5164. 1985.PubMed/NCBI

3 

Congleton J, MacDonald R and Yen A: Src inhibitors, PP2 and dasatinib, increase retinoic acid-induced association of Lyn and c-Raf (S259) and enhance MAPK-dependent differentiation of myeloid leukemia cells. Leukemia. 26:1180–1188. 2012. View Article : Google Scholar :

4 

Ji Y, Kutner A, Verstuyf A, Verlinden L and Studzinski GP: Derivatives of vitamins D2 and D3 activate three MAPK pathways and upregulate pRb expression in differentiating HL60 cells. Cell Cycle. 1:410–415. 2002. View Article : Google Scholar

5 

Wang N, Wang LW, Gou BD, Zhang TL and Wang K: Realgar-induced differentiation is associated with MAPK pathways in HL-60 cells. Cell Biol Int. 32:1497–1505. 2008. View Article : Google Scholar : PubMed/NCBI

6 

Kim SH, Yoo JC and Kim TS: Nargenicin enhances 1,25-dihy-droxyvitamin D (3)- and all-trans retinoic acid-induced leukemia cell differentiation via PKCbetaI/MAPK pathways. Biochem Pharmacol. 77:1694–1701. 2009. View Article : Google Scholar : PubMed/NCBI

7 

Stixová L, Procházková J, Soucek K, Hofmanová J and Kozubík A: 5-Lipoxygenase inhibitors potentiate 1alpha,25-dihy-droxyvitamin D3-induced monocytic differentiation by activating p38 MAPK pathway. Mol Cell Biochem. 330:229–238. 2009. View Article : Google Scholar

8 

Battle TE, Levine RA and Yen A: Retinoic acid-induced blr1 expression promotes ERK2 activation and cell differentiation in HL-60 cells. Exp Cell Res. 254:287–298. 2000. View Article : Google Scholar : PubMed/NCBI

9 

Yen A, Roberson MS and Varvayanis S: Retinoic acid selectively activates the ERK2 but not JNK/SAPK or p38 MAP kinases when inducing myeloid differentiation. In Vitro Cell Dev Biol Anim. 35:527–532. 1999. View Article : Google Scholar : PubMed/NCBI

10 

Yu HN, Lee YR, Noh EM, Lee KS, Song EK, Han MK, Lee YC, Yim CY, Park J, Kim BS, et al: Tumor necrosis factor-alpha enhances DMSO-induced differentiation of HL-60 cells through the activation of ERK/MAPK pathway. Int J Hematol. 87:189–194. 2008. View Article : Google Scholar : PubMed/NCBI

11 

Li C, Yu Y, Wang Y, Liu L, Zhang M, Sugano S, Wang Z and Chang Z: Both ERK and JNK are required for enhancement of MD-2 gene expression during differentiation of HL-60 cells. Biol Cell. 100:365–375. 2008. View Article : Google Scholar : PubMed/NCBI

12 

Zhang J, Harrison JS and Studzinski GP: Isoforms of p38MAPK gamma and delta contribute to differentiation of human AML cells induced by 1,25-dihydroxyvitamin D (3). Exp Cell Res. 317:117–130. 2011. View Article : Google Scholar

13 

Matsumoto E, Hatanaka M, Bohgaki M and Maeda S: PKC pathway and ERK/MAPK pathway are required for induction of cyclin D1 and p21Waf1 during 12-o-tetradecanoylphorbol 13-acetate-induced differentiation of myeloleukemia cells. Kobe J Med Sci. 52:181–194. 2006.

14 

Yiang GT, Yu YL, Hu SC, Chen MH, Wang JJ and Wei CW: PKC and MEK pathways inhibit caspase-9/-3-mediated cytotoxicity in differentiated cells. FEBS Lett. 582:881–885. 2008. View Article : Google Scholar : PubMed/NCBI

15 

Wang X, Pesakhov S, Harrison JS, Danilenko M and Studzinski GP: ERK5 pathway regulates transcription factors important for monocytic differentiation of human myeloid leukemia cells. J Cell Physiol. 229:856–867. 2014. View Article : Google Scholar

16 

Jamshidi F, Zhang J, Harrison JS, Wang X and Studzinski GP: Induction of differentiation of human leukemia cells by combinations of COX inhibitors and 1,25-dihydroxyvitamin D3 involves Raf1 but not Erk 1/2 signaling. Cell Cycle. 7:917–924. 2008. View Article : Google Scholar : PubMed/NCBI

17 

Wang X, Pesakhov S, Weng A, Kafka M, Gocek E, Nguyen M, Harrison JS, Danilenko M and Studzinski GP: ERK 5/MAPK pathway has a major role in 1α,25-(oh) vitamin D3-induced terminal differentiation of myeloid leukemia cells. J Steroid Biochem Mol Biol. 144:223–227. 2013. View Article : Google Scholar

18 

Uruno A, Noguchi N, Matsuda K, Nata K, Yoshikawa T, Chikamatsu Y, Kagechika H, Harigae H, Ito S, Okamoto H, et al: All-trans retinoic acid and a novel synthetic retinoid tami-barotene (Am80) differentially regulate CD38 expression in human leukemia HL-60 cells: possible involvement of protein kinase C-delta. J Leukoc Biol. 90:235–247. 2011. View Article : Google Scholar : PubMed/NCBI

19 

Ju SM, Kang JG, Pae HO, Lee GS, Kim WS, Lyu YS and Jeon BH: Nardostachys chinensis induces the differentiation of human promyelocytic leukemic cells through the activation of the protein kinase C-dependent extracellular signal-regulated kinase signaling pathway. Int J Mol Med. 33:573–580. 2014.

20 

Ogino T, Ozaki M and Matsukawa A: Oxidative stress enhances granulocytic differentiation in HL 60 cells, an acute promy-elocytic leukemia cell line. Free Radic Res. 44:1328–1337. 2010. View Article : Google Scholar : PubMed/NCBI

21 

Hu XM, Yuan B, Tanaka S, Zhou Q, Onda K, Toyoda H and Hirano T: Involvement of oxidative stress associated with glutathione depletion and p38 mitogen-activated protein kinase activation in arsenic disulfide-induced differentiation in HL-60 cells. Leuk Lymphoma. 55:392–404. 2014. View Article : Google Scholar

22 

Kambhampati S, Li Y, Verma A, Sassano A, Majchrzak B, Deb DK, Parmar S, Giafis N, Kalvakolanu DV, Rahman A, et al: Activation of protein kinase C delta by all-trans-retinoic acid. J Biol Chem. 278:32544–32551. 2003. View Article : Google Scholar : PubMed/NCBI

23 

Kim SH, Kang SN, Kim HJ and Kim TS: Potentiation of 1,25-dihydroxyvitamin D (3)-induced differentiation of human promyelocytic leukemia cells into monocytes by costunolide, a germacranolide sesquiterpene lactone. Biochem Pharmacol. 64:1233–1242. 2002. View Article : Google Scholar : PubMed/NCBI

24 

Bondza-Kibangou P, Millot C, El Khoury V and Millot JM: Antioxidants and doxorubicin supplementation to modulate CD14 expression and oxidative stress induced by vitamin D3 and seocalcitol in HL60 cells. Oncol Rep. 18:1513–1519. 2007.PubMed/NCBI

25 

Yamamoto T, Sakaguchi N, Hachiya M, Nakayama F, Yamakawa M and Akashi M: Role of catalase in monocytic differentiation of U937 cells by TPA: hydrogen peroxide as a second messenger. Leukemia. 23:761–769. 2009. View Article : Google Scholar

26 

Yan Y, Zeng W, Song S, Zhang F, He W, Liang W and Niu Z: Vitamin C induces periodontal ligament progenitor cell differentiation via activation of ERK pathway mediated by PELP1. Protein Cell. 4:620–627. 2013. View Article : Google Scholar : PubMed/NCBI

27 

Mimori K, Komaki M, Iwasaki K and Ishikawa I: Extracellular signal-regulated kinase 1/2 is involved in ascorbic acid-induced osteoblastic differentiation in periodontal ligament cells. J Periodontol. 78:328–334. 2007. View Article : Google Scholar : PubMed/NCBI

28 

Shinozaki K, Hosokawa Y, Hazawa M, Kashiwakura I, Okumura K, Kaku T and Nakayama E: Ascorbic acid enhances radiation-induced apoptosis in an HL60 human leukemia cell line. J Radiat Res. 52:229–237. 2011. View Article : Google Scholar : PubMed/NCBI

29 

Gokhalé P, Patel T, Morrison MJ and Vissers MC: The effect of intracellular ascorbate on the susceptibility of HL60 and Jurkat cells to chemotherapy agents. Apoptosis. 11:1737–1746. 2006. View Article : Google Scholar : PubMed/NCBI

30 

Kang HK, Suh JH, Lee JJ, Yoon SH, Hyun JW, Choi SW, Choi JY, Ryu KH and Chung MH: Induction of the differentiation of HL-60 promyelocytic leukemia cells by L-ascorbic acid. Free Radic Res. 37:773–779. 2003. View Article : Google Scholar : PubMed/NCBI

31 

Witenberg B, Kletter Y, Kalir HH, Raviv Z, Fenig E, Nagler A, Halperin D and Fabian I: Ascorbic acid inhibits apoptosis induced by X irradiation in HL60 myeloid leukemia cells. Radiat Res. 152:468–478. 1999. View Article : Google Scholar : PubMed/NCBI

32 

Karasavvas N, Carcamo JM, Stratis G and Golde DW: Vitamin C protects HL60 and U266 cells from arsenic toxicity. Blood. 105:4004–4012. 2005. View Article : Google Scholar : PubMed/NCBI

33 

Cuddihy SL, Parker A, Harwood DT, Vissers MC and Winterbourn CC: Ascorbate interacts with reduced glutathione to scavenge phenoxyl radicals in HL60 cells. Free Radic Biol Med. 44:1637–1644. 2008. View Article : Google Scholar : PubMed/NCBI

34 

Parker A, Cuddihy SL, Son TG, Vissers MC and Winterbourn CC: Roles of superoxide and myeloperoxidase in ascorbate oxidation in stimulated neutrophils and H2O2-treated HL60 cells. Free Radic Biol Med. 51:1399–1405. 2011. View Article : Google Scholar : PubMed/NCBI

35 

Taniguchi M, A rai N, Kohno K, Ushio S and Fukuda S: Anti-oxidative and anti-aging activities of 2-O-alpha-glucopyranosyl-L-ascorbic acid on human dermal fibroblasts. Eur J Pharmacol. 674:126–131. 2012. View Article : Google Scholar

36 

Naziroglu M, Akkus S, Soyupek F, Yalman K, Çelik Ö, Eriş S and Uslusoy GA: Vitamins C and E treatment combined with exercise modulates oxidative stress markers in blood of patients with fibromyalgia: A controlled clinical pilot study. Stress. 13:498–505. 2010.PubMed/NCBI

37 

Yokoo S, Furumoto K, Hiyama E and Miwa N: Slow-down of age-dependent telomere shortening is executed in human skin keratinocytes by hormesis-like-effects of trace hydrogen peroxide or by anti-oxidative effects of pro-vitamin C in common concurrently with reduction of intracellular oxidative stress. J Cell Biochem. 93:588–597. 2004. View Article : Google Scholar : PubMed/NCBI

38 

Kim YH, Kim CH, Cho MK, Kim KM, Lee SY, Ahn BW, Yang SY, Kim SM and Song TB: Total peroxyl radical-trapping ability and anti-oxidant vitamins of the umbilical venous plasma and the placenta in pre-eclampsia. J Obstet Gynaecol Res. 32:32–41. 2006. View Article : Google Scholar : PubMed/NCBI

39 

Yiang GT, Chou PL, Hung YT, Chen JN, Chang WJ, Yu YL and Wei CW: Vitamin C enhances anticancer activity in methotrexa-tetreated Hep3B hepatocellular carcinoma cells. Oncol Rep. 32:1057–1063. 2014.PubMed/NCBI

40 

Yu YL, Yiang GT, Chou PL, Tseng HH, Wu TK, Hung YT, Lin PS, Lin SY, Liu HC, Chang WJ, et al: Dual role of acetaminophen in promoting hepatoma cell apoptosis and kidney fibroblast proliferation. Mol Med Rep. 9:2077–2084. 2014.PubMed/NCBI

41 

Yiang GT, Chen YH, Chou PL, Chang WJ, Wei CW and Yu YL: The NS3 protease and helicase domains of Japanese encephalitis virus trigger cell death via caspasedependent and independent pathways. Mol Med Rep. 7:826–830. 2013.PubMed/NCBI

42 

Chen KH, Li PC, Lin WH, Chien CT and Low BH: Depression by a green tea extract of alcohol-induced oxidative stress and lipogenesis in rat liver. Biosci Biotechnol Biochem. 75:1668–1676. 2011. View Article : Google Scholar : PubMed/NCBI

43 

Lin BR, Yu CJ, Chen WC, Lee HS, Chang HM, Lee YC, Chien CT and Chen CF: Green tea extract supplement reduces D-galactosamine-induced acute liver injury by inhibition of apoptotic and proinflammatory signaling. J Biomed Sci. 16:352009. View Article : Google Scholar : PubMed/NCBI

44 

Das D, Pintucci G and Stern A: MAPK-dependent expression of p21 (WAF) and p27 (kip1) in PMA-induced differentiation of HL60 cells. FEBS Lett. 472:50–52. 2000. View Article : Google Scholar : PubMed/NCBI

45 

Wei CW, Hu CC, Tang CH, Lee MC and Wang JJ: Induction of differentiation rescues HL-60 cells from Rana catesbeiana ribonuclease-induced cell death. FEBS Lett. 531:421–426. 2002. View Article : Google Scholar : PubMed/NCBI

46 

Siebel A, Cubillos-Rojas M, Santos RC, Schneider T, Bonan CD, Bartrons R, Ventura F, Rodrigues de Oliveira J and Rosa JL: Contribution of S6K1/MAPK signaling pathways in the response to oxidative stress: Activation of RSK and MSK by hydrogen peroxide. PLoS One. 8:e755232013. View Article : Google Scholar : PubMed/NCBI

47 

Moslehi M, Meshkini A and Yazdanparast R: Flavonoid baicalein modulates H2O2-induced mitogen-activated protein kinases activation and cell death in SK-N-MC cells. Cell Mol Neurobiol. 32:549–560. 2012. View Article : Google Scholar : PubMed/NCBI

48 

Kang KA, Lee KH, Zhang R, Piao MJ, Kang MY, Kwak YS, Yoo BS, You HJ and Hyun JW: Protective effects of Castanopsis cuspidate through activation of ERK and NF-kappaB on oxidative cell death induced by hydrogen peroxide. J Toxicol Environ Health A. 70:1319–1328. 2007. View Article : Google Scholar : PubMed/NCBI

49 

Ulrich-Merzenich G, Zeitler H, Panek D, Bokemeyer D and Vetter H: Vitamin C promotes human endothelial cell growth via the ERK-signaling pathway. Eur J Nutr. 46:87–94. 2007. View Article : Google Scholar : PubMed/NCBI

50 

Park S, Park CH, Hahm ER, Kim K, Kimler BF, Lee SJ, Park HK, Lee SH, Kim WS, Jung CW, et al: Activation of Raf1 and the ERK pathway in response to l-ascorbic acid in acute myeloid leukemia cells. Cell Signal. 17:111–119. 2005. View Article : Google Scholar

51 

Park HJ, Ock SM, Kim HJ, Park HJ, Lee YB, Choi JM, Cho CS, Lee JY, Cho BK and Cho DH: Vitamin C attenuates ERK signalling to inhibit the regulation of collagen production by LL-37 in human dermal fibroblasts. Exp Dermatol. 19:e258–e264. 2010. View Article : Google Scholar : PubMed/NCBI

52 

Cuaranta-Monroy I, Simandi Z, Kolostyak Z, Doan-Xuan QM, Poliska S, Horvath A, Nagy G, Bacso Z and Nagy L: Highly efficient differentiation of embryonic stem cells into adipocytes by ascorbic acid. Stem Cell Res. 13:88–97. 2014. View Article : Google Scholar : PubMed/NCBI

53 

Yu J, Tu YK, Tang YB and Cheng NC: Stemness and transdif-ferentiation of adipose-derived stem cells using L-ascorbic acid 2-phosphate-induced cell sheet formation. Biomaterials. 35:3516–3526. 2014. View Article : Google Scholar : PubMed/NCBI

54 

Valenti MT, Zanatta M, Donatelli L, Viviano G, Cavallini C, Scupoli MT and Dalle Carbonare L: Ascorbic acid induces either differentiation or apoptosis in MG-63 osteosarcoma lineage. Anticancer Res. 34:1617–1627. 2014.PubMed/NCBI

55 

Langenbach F and Handschel J: Effects of dexamethasone, ascorbic acid and beta-glycerophosphate on the osteogenic differentiation of stem cells in vitro. Stem Cell Res Ther. 4:1172013. View Article : Google Scholar

56 

Hadzir SN, Ibrahim SN, Abdul Wahab RM, Zaino Abidin IZ, Senafi S, Ariffin ZZ, Abdul Razak M and Zainal Ariffin SH: Ascorbic acid induces osteoblast differentiation of human suspension mononuclear cells. Cytotherapy. 16:674–682. 2014. View Article : Google Scholar

Related Articles

Journal Cover

October-2015
Volume 12 Issue 4

Print ISSN: 1791-2997
Online ISSN:1791-3004

Sign up for eToc alerts

Recommend to Library

Copy and paste a formatted citation
x
Spandidos Publications style
Yiang GT, Chen JN, Wu TK, Wang HF, Hung YT, Chang WJ, Chen C, Wei CW and Yu YL: Ascorbic acid inhibits TPA-induced HL‑60 cell differentiation by decreasing cellular H2O2 and ERK phosphorylation. Mol Med Rep 12: 5501-5507, 2015.
APA
Yiang, G., Chen, J., Wu, T., Wang, H., Hung, Y., Chang, W. ... Yu, Y. (2015). Ascorbic acid inhibits TPA-induced HL‑60 cell differentiation by decreasing cellular H2O2 and ERK phosphorylation. Molecular Medicine Reports, 12, 5501-5507. https://doi.org/10.3892/mmr.2015.4091
MLA
Yiang, G., Chen, J., Wu, T., Wang, H., Hung, Y., Chang, W., Chen, C., Wei, C., Yu, Y."Ascorbic acid inhibits TPA-induced HL‑60 cell differentiation by decreasing cellular H2O2 and ERK phosphorylation". Molecular Medicine Reports 12.4 (2015): 5501-5507.
Chicago
Yiang, G., Chen, J., Wu, T., Wang, H., Hung, Y., Chang, W., Chen, C., Wei, C., Yu, Y."Ascorbic acid inhibits TPA-induced HL‑60 cell differentiation by decreasing cellular H2O2 and ERK phosphorylation". Molecular Medicine Reports 12, no. 4 (2015): 5501-5507. https://doi.org/10.3892/mmr.2015.4091