Generic placeholder image

Current Drug Discovery Technologies

Editor-in-Chief

ISSN (Print): 1570-1638
ISSN (Online): 1875-6220

Research Article

Efficacy of β-D-Mannuronic Acid [M2000] on the Pro-Apoptotic Process and Inflammatory-Related Molecules NFκB, IL-8 and Cd49d using Healthy Donor PBMC

Author(s): Atousa Khalatbari, Mehdi Mahdavi, Fahimeh Jafarnezhad, Sanaz Afraei, Farzaneh Tofighi Zavareh, Zahra Aghazadeh, Afshin Ghaderi and Abbas Mirshafiey*

Volume 17 , Issue 2 , 2020

Page: [225 - 232] Pages: 8

DOI: 10.2174/1570163815666181109165837

Price: $65

Abstract

Objective: This investigation evaluates the pro-apoptotic and anti-inflammatory effects of β-D-mannuronic acid [M2000] compared to diclofenac, based on gene expression involved in apoptosis and inflammation process [including Bcl2, NFκB, IL-8 and Cd49d] in Peripheral Blood Mononuclear Cells [PBMCs] of healthy donors under exvivo conditions.

Materials: The venous blood samples of twelve healthy volunteers with aged 25-60 years were collected in heparinized tubes. The healthy volunteers were selected from no smoking group and without using illicit drugs and suffering from diabetes. The PBMCs were separated and divided into untreated and treated groups.

Methods: The PBMCs of each sample were cultured in 5 wells of culture plate, so that the first well consisted of 2×106 cells exposed by LPS-EB [1μg/ml] to stimulate PBMCs and absence of M2000 [untreated well]. The second, third, fourth and fifth wells containing 2×106 cells/well and LPS-EB, after 4 hours incubation at 37ºC, received 5, 25 and 50 μg/well of M2000 and 5 μg/well of diclofenac, respectively as treated group.

Results: The PBMCs were separated and RNAs were then extracted and cDNAs synthesized and gene expression levels were assessed by qRT-PCR. Furthermore, we studied whether M2000 is able to facilitate apoptosis in PBMCs. Our findings represent that the high dose of M2000 could significantly decrease the expression level of NFκB gene compared to untreated group (p < 0.0002). On the other hand, no significant change was observed in treated cells with diclofenac. All doses of M2000 could significantly augment apoptosis compared to untreated group [p < 0.0001]. Additionally, we observed the same apoptotic effects between the medium dose of M2000 and diclofenac. Besides, no significant reduction was shown in expression levels of IL8, Bcl2 and Cd49d genes in all doses of M2000 and diclofenac compared to untreated group. This experiment demonstrates M2000 as a new effective NSAID with immunosuppressive characteristics capable of stimulating apoptosis through lowering expression levels of NFκB gene, which might be probably considered as an appropriate drug for reducing the risk of developing inflammatory diseases and cancer.

Keywords: Mannuronic acid, M2000, pro-apoptotic, immunosuppressive, NSAID, inflammation.

Graphical Abstract
[1]
Rock KL, Latz E, Ontiveros F, Kono H. The sterile inflammatory response. Annu Rev Immunol 2010; 28: 321-42.
[http://dx.doi.org/10.1146/annurev-immunol-030409-101311] [PMID: 20307211]
[2]
Mantovani A, Allavena P, Sica A, Balkwill F. Cancer-related inflammation. Nature 2008; 454(7203): 436-44.
[http://dx.doi.org/10.1038/nature07205] [PMID: 18650914]
[3]
Grivennikov SI, Greten FR, Karin M. Immunity, inflammation, and cancer. Cell 2010; 140(6): 883-99.
[http://dx.doi.org/10.1016/j.cell.2010.01.025] [PMID: 20303878]
[4]
Johnstone RW, Ruefli AA, Lowe SW. Apoptosis: a link between cancer genetics and chemotherapy. Cell 2002; 108(2): 153-64.
[http://dx.doi.org/10.1016/S0092-8674(02)00625-6] [PMID: 11832206]
[5]
Igney FH, Krammer PH. Death and anti-death: tumour resistance to apoptosis. Nat Rev Cancer 2002; 2(4): 277-88.
[http://dx.doi.org/10.1038/nrc776] [PMID: 12001989]
[6]
Reed JC. Bcl-2 and the regulation of programmed cell death. J Cell Biol 1994; 124(1-2): 1-6.
[http://dx.doi.org/10.1083/jcb.124.1.1] [PMID: 8294493]
[7]
Korsmeyer SJ. BCL-2 gene family and the regulation of programmed cell death. Cancer Res 1999; 59(7)(Suppl.): 1693s-700s.
[PMID: 10197582]
[8]
Oeckinghaus A, Ghosh S. The NF-kappaB family of transcription factors and its regulation. Cold Spring Harb Perspect Biol 2009; 1(4)a000034
[http://dx.doi.org/10.1101/cshperspect.a000034] [PMID: 20066092]
[9]
Tak PP, Firestein GS. NF-kappaB: a key role in inflammatory diseases. J Clin Invest 2001; 107(1): 7-11.
[http://dx.doi.org/10.1172/JCI11830] [PMID: 11134171]
[10]
Makarov SS. NF-kappaB as a therapeutic target in chronic inflammation: recent advances. Mol Med Today 2000; 6(11): 441-8.
[http://dx.doi.org/10.1016/S1357-4310(00)01814-1] [PMID: 11074370]
[11]
Karin M, Lawrence T, Nizet V. Innate immunity gone awry: linking microbial infections to chronic inflammation and cancer. Cell 2006; 124(4): 823-35.
[http://dx.doi.org/10.1016/j.cell.2006.02.016] [PMID: 16497591]
[12]
Harada A, Sekido N, Akahoshi T, Wada T, Mukaida N, Matsushima K. Essential involvement of interleukin-8 (IL-8) in acute inflammation. J Leukoc Biol 1994; 56(5): 559-64.
[http://dx.doi.org/10.1002/jlb.56.5.559] [PMID: 7964163]
[13]
Koch AE, Polverini PJ, Kunkel SL, et al. Interleukin-8 as a macrophage-derived mediator of angiogenesis. Science 1992; 258(5089): 1798-801.
[http://dx.doi.org/10.1126/science.1281554] [PMID: 1281554]
[14]
Ridger VC, Wagner BE, Wallace WA, Hellewell PG. Differential effects of CD18, CD29, and CD49 integrin subunit inhibition on neutrophil migration in pulmonary inflammation. J Immunol 2001; 166(5): 3484-90.
[http://dx.doi.org/10.4049/jimmunol.166.5.3484] [PMID: 11207307]
[15]
Muraro PA, Leist T, Bielekova B, McFarland HF. VLA-4/CD49d downregulated on primed T lymphocytes during interferon-β therapy in multiple sclerosis. J Neuroimmunol 2000; 111(1-2): 186-94.
[http://dx.doi.org/10.1016/S0165-5728(00)00362-3] [PMID: 11063837]
[16]
Gilroy DW, Colville-Nash PR, Willis D, Chivers J, Paul-Clark MJ, Willoughby DA. Inducible cyclooxygenase may have anti-inflammatory properties. Nat Med 1999; 5(6): 698-701.
[http://dx.doi.org/10.1038/9550] [PMID: 10371510]
[17]
Liu B, Hong J-S. Role of microglia in inflammation-mediated neurodegenerative diseases: mechanisms and strategies for therapeutic intervention. J Pharmacol Exp Ther 2003; 304(1): 1-7.
[http://dx.doi.org/10.1124/jpet.102.035048] [PMID: 12490568]
[18]
Whelton A. Nephrotoxicity of nonsteroidal anti-inflammatory drugs: physiologic foundations and clinical implications. Am J Med 1999; 106(5B): 13S-24S.
[http://dx.doi.org/10.1016/S0002-9343(99)00113-8] [PMID: 10390124]
[19]
Shacter E, Weitzman SA. Chronic inflammation and cancer. Oncology (Williston Park) 2002; 16(2): 217-26.
[PMID: 11866137]
[20]
Dolcet X, Llobet D, Pallares J, Matias-Guiu X. NF-kB in development and progression of human cancer. Virchows Arch 2005; 446(5): 475-82.
[http://dx.doi.org/10.1007/s00428-005-1264-9] [PMID: 15856292]
[21]
Matés JM, Sánchez-Jiménez FM. Role of reactive oxygen species in apoptosis: implications for cancer therapy. Int J Biochem Cell Biol 2000; 32(2): 157-70.
[http://dx.doi.org/10.1016/S1357-2725(99)00088-6] [PMID: 10687951]
[22]
Baeuerle PA, Henkel T. Function and activation of NF-kappa B in the immune system. Annu Rev Immunol 1994; 12(1): 141-79.
[http://dx.doi.org/10.1146/annurev.iy.12.040194.001041] [PMID: 8011280]
[23]
McCool KW, Miyamoto S. DNA damage-dependent NF-κB activation: NEMO turns nuclear signaling inside out. Immunol Rev 2012; 246(1): 311-26.
[http://dx.doi.org/10.1111/j.1600-065X.2012.01101.x] [PMID: 22435563]
[24]
Sica A, Larghi P, Mancino A, Rubino L, Porta C, Totaro MG, Eds. Macrophage polarization in tumour progression Seminars in cancer biology. Elsevier 2008.
[25]
Galdiero MR, Garlanda C, Jaillon S, Marone G, Mantovani A. Tumor associated macrophages and neutrophils in tumor progression. J Cell Physiol 2013; 228(7): 1404-12.
[http://dx.doi.org/10.1002/jcp.24260] [PMID: 23065796]
[26]
Chen J, Wang F-L, Chen W-D. Modulation of apoptosis-related cell signalling pathways by curcumin as a strategy to inhibit tumor progression. Mol Biol Rep 2014; 41(7): 4583-94.
[http://dx.doi.org/10.1007/s11033-014-3329-9] [PMID: 24604727]
[27]
Haslett C. Granulocyte apoptosis and its role in the resolution and control of lung inflammation. American journal of respiratory and critical care medicine 1999; 160(supplement_1): S5-S11.
[http://dx.doi.org/10.1164/ajrccm.160.supplement_1.4]
[28]
Fulda S, Meyer E, Debatin K-M. Inhibition of TRAIL-induced apoptosis by Bcl-2 overexpression. Oncogene 2002; 21(15): 2283-94.
[http://dx.doi.org/10.1038/sj.onc.1205258] [PMID: 11948412]
[29]
Fattahi MJ, Jamshidi AR, Mahmoudi M, et al. Evaluation of the efficacy and safety of β-d-mannuronic acid in patients with ankylosing spondylitis: A 12-week randomized, placebo-controlled, phase I/II clinical trial. Int Immunopharmacol 2018; 54: 112-7.
[http://dx.doi.org/10.1016/j.intimp.2017.11.003] [PMID: 29127910]
[30]
Roozbehkia M, Mahmoudi M, Aletaha S, et al. The potent suppressive effect of β-d-mannuronic acid (M2000) on molecular expression of the TLR/NF-kB Signaling Pathway in ankylosing spondylitis patients. Int Immunopharmacol 2017; 52: 191-6.
[http://dx.doi.org/10.1016/j.intimp.2017.08.018] [PMID: 28938189]
[31]
Jafarnezhad-Ansariha F, Yekaninejad MS, Jamshidi AR, et al. The effects of β-D-mannuronic acid (M2000), as a novel NSAID, on COX1 and COX2 activities and gene expression in ankylosing spondylitis patients and the murine monocyte/macrophage, J774 cell line. Inflammopharmacology 2018; 26(2): 375-84.
[http://dx.doi.org/10.1007/s10787-017-0386-4] [PMID: 28819782]
[32]
Mirshafiey A, Taeb M, Mortazavi-Jahromi SS, et al. Introduction of β-d-mannuronic acid (M2000) as a novel NSAID with immunosuppressive property based on COX-1/COX-2 activity and gene expression. Pharmacol Rep 2017; 69(5): 1067-72.
[http://dx.doi.org/10.1016/j.pharep.2017.04.015] [PMID: 28951072]
[33]
Ahmadi H, Jamshidi AR, Gharibdoost F, et al. A phase I/II randomized, controlled, clinical trial for assessment of the efficacy and safety of β-D-mannuronic acid in rheumatoid arthritis patients. Inflammopharmacology 2018; 26(3): 737-45.
[http://dx.doi.org/10.1007/s10787-018-0475-z] [PMID: 29696564]
[34]
Ahmadi H, Mahmoudi M, Gharibdoost F, et al. Targeting of circulating Th17 cells by β-D-mannuronic acid (M2000) as a novel medication in patients with rheumatoid arthritis. Inflammopharmacology 2018; 26(1): 57-65.
[http://dx.doi.org/10.1007/s10787-017-0410-8] [PMID: 29063487]
[35]
Ahmadi H, Jamshidi AR, Mahmoudi M, et al. Hematological improvement of patients with active rheumatoid arthritis by β-d-mannuronic acid (M2000) as a novel NSAID with immunosuppressive property. Iran J Allergy Asthma Immunol 2017; 16(5): 433-42.
[PMID: 29149783]
[36]
Barati A, Jamshidi AR, Ahmadi H, Aghazadeh Z, Mirshafiey A. Effects of β-d-mannuronic acid, as a novel non-steroidal anti-inflammatory medication within immunosuppressive properties, on IL17, RORγt, IL4 and GATA3 gene expressions in rheumatoid arthritis patients. Drug Des Devel Ther 2017; 11: 1027-33.
[http://dx.doi.org/10.2147/DDDT.S129419] [PMID: 28408801]
[37]
Mirshafiey A, Rehm B, Sotoude M, Razavi A, Abhari RS, Borzooy Z. Therapeutic approach by a novel designed anti-inflammatory drug, M2000, in experimental immune complex glomerulonephritis. Immunopharmacol Immunotoxicol 2007; 29(1): 49-61.
[http://dx.doi.org/10.1080/08923970701282387] [PMID: 17464766]
[38]
Mirshafiey A, Rehm B, Abhari RS, Borzooy Z, Sotoude M, Razavi A. Production of M2000 (β-d-mannuronic acid) and its therapeutic effect on experimental nephritis. Environ Toxicol Pharmacol 2007; 24(1): 60-6.
[http://dx.doi.org/10.1016/j.etap.2007.02.002] [PMID: 21783790]
[39]
Fattahi MJ, Abdollahi M, Agha Mohammadi A, et al. Preclinical assessment of β-d-mannuronic acid (M2000) as a non-steroidal anti-inflammatory drug. Immunopharmacol Immunotoxicol 2015; 37(6): 535-40.
[http://dx.doi.org/10.3109/08923973.2015.1113296] [PMID: 26584020]
[40]
Aletaha S, Haddad L, Roozbehkia M, et al. M2000 (β-D-Mannuronic Acid) as a Novel antagonist for blocking the TLR2 and TLR4 downstream signalling pathway. Scand J Immunol 2017; 85(2): 122-9.
[http://dx.doi.org/10.1111/sji.12519] [PMID: 27943385]
[41]
Hosseini S, Abdollahi M, Azizi G, et al. Anti-aging effects of M2000 (β-D-mannuronic acid) as a novel immunosuppressive drug on the enzymatic and non-enzymatic oxidative stress parameters in an experimental model. J Basic Clin Physiol Pharmacol 2017; 28(3): 249-55.
[http://dx.doi.org/10.1515/jbcpp-2016-0092] [PMID: 28207414]

Rights & Permissions Print Export Cite as
© 2022 Bentham Science Publishers | Privacy Policy