1. Introduction
ell therapy is a subset of restorative medicine. Moreover, stem cells are the first hope for the repair of damaged tissues [3, 4]. Concerning origin, these cells are divided into two main categories of embryonic stem cells and adult stem cells [5]. Due to some limitations in the production and use of embryonic stem cells, in recent years, a new wave of research on adult stem cells has begun, i.e., extensively performed [6]. Mesenchymal Stem Cells (MSCs), as major adult stem cells, can differentiate into cells that are not mesenchymal derivatives [7]. However, due to the unfavorable conditions of the transplant recipient, including hypoxia and the presence of oxygen-free radicals that activate and increase aging factors by causing stress in the cell, eventually leading to apoptosis and cell death, most of the transplanted mesenchymal stem cells are lost in the early days; which in turn, this condition reduces their efficiency [12, 13]. Studies attempted to identify and use factors that prevent oxidative stress in these cells. One of these characteristics included antioxidants [14]. Ginger, with the scientific name of Zingiber officinale, is a medicinal plant with pharmacological properties, such as antioxidant, anti-apoptotic, and anti-inflammatory effects [18]. Therefore, the present study aimed to explore the antioxidant effect of ginger extract on cytotoxicity and the induction of apoptosis due to oxidative stress in human adipose tissue-derived MSCs and rat bone marrow.  
2. Materials and Methods
In this study, human Adipose Tissue Mesenchymal Stem Cells (AD-MSCs) were obtained from the National Center for Genetic and Biological Resources of Iran and rat bone marrow mesenchymal stem cells were extracted from the tibia and femur of rats. Cells in Dulbecco’s Modified Eagle Medium (DMEM) culture medium (Gibco, USA) containing 20% Foetal Bovine Serum (FBS) (Gibco, USA) and 1% Penicillin-Streptomycin (Penstrep) (Gibco, USA) in an incubator (Memmert, Germany) were cultured in flask 75 with 5% CO2 gas pressure, 90% humidity, and at 37°C. The culture medium was changed three times a week and trypsin/EDTA solution was used to harvest the cells. To prepare the ginger extract, 500 gr of the dried ginger plant was prepared and after grinding, the resulting powder was placed in 96% alcohol for 10 days to dissolve its active ingredients in alcohol. The contents of the filter paper and the solution obtained by the rotary apparatus were then concentrated at 50°C and 60 rpm. H2O2 was prepared in liquid form. The assay of human adipose tissue-derived MSCs was evaluated using MTS colorimetric method. The status of apoptosis in rat bone marrow-derived MSCs were also explored by Annexin V-FITC/PI test. 
3. Results
The effects of ginger extract on cytotoxicity induced by oxidative stress in human adipose tissue-derived MSCs. Oxidative stress-induced cytotoxicity in human adipose tissue-derived MSCs treated with different concentrations of ginger extract (50, 100, 200 & 400 mg/mL), after incubation periods (4 & 6 hours with ginger extract & 2 hours with H2O2) were evaluated using MTS test. The obtained results indicated that ginger extract could increase the bioavailability of human adipose tissue-derived MSCs in a dose- and time-dependent manner (P≤0.022*) (Figure 1). The effects of ginger extract on the induction of oxidative stress-induced apoptosis in rat bone marrow-derived MSCs
Annexin V-FITS test was applied to evaluate the induction of apoptosis. The relevant results suggested a dose- and time-dependent reduction in the frequency of death in bone marrow-derived MSCs in the treated experimental groups.
As illustrated in Figure 2, in the 4-hour treatment, the frequency of cells in the early stages of apoptosis increased from 37.60%  at 50 mg/mL to 29.48% at 400. Such a decrease in the frequency of apoptosis in all concentrations was significantly different from that of the negative control group. Furthermore, the frequency of cells in the late stages of apoptosis decreased with the increasing dose of ginger extract, from 13.42% at a concentration of 50 mg/mL to 12.18% at a concentration of 400 mg/mL, i.e., also a significant decrease in all concentrations, compared to the negative control group (P≤0.030) (Figure 2).
Furthermore, in the 6-hour treatment, the frequency of cells in the early stages of apoptosis increased from 23.67% at a concentration of 50 mg/mL to 21.59% at a concentration of 400 mg/mL. This decrease in the frequency of apoptosis in all concentrations significantly differed from that of the negative control group. In addition, the frequency of cells in the late stages of apoptosis decreased with increasing dose and treatment time with ginger extract, from 12.98% at a concentration of 50 mg/mL to 15.9% to 400 mg/mL, i.e., a significant decrease in the frequency of apoptosis in all concentrations, compared to the negative control group (P≤0.016) (Figure 3). 4. Discussion and Conclusion 
During the normal functioning of the cells used in cell therapy, reactive oxygen species are produced that have high reactivity with DNA, proteins, carbohydrates, and lipids; they cause irreparable damage to these macromolecules to enter. Ginger is an essential medicinal plant that has long been used to treat various diseases [19]. The antioxidant effects of ginger have been studied for years. This study explored the antioxidant effects of ginger extract on cytotoxicity and the induction of oxidative stress-induced apoptosis in human adipose tissue-derived MSCs and rat bone marrow. The obtained results indicated that ginger extract reduced the frequency of human adipose tissue-derived MSCs and rat bone marrow by acting as an antioxidant, which further increased the success of using mesenchymal stem cells in the cell treatment. 

Ethical Considerations
Compliance with ethical guidelines
This study was approved by the Ethics Research Committee of Shahrekord Branch, Islamic Azad University (code: IR.IAU.SHK.REC.1397.028).

Funding
This study was extracted from the Msc. thesis of the first author at the Department of Biochemistry, Faculty of Science, Shahrekord Branch, Islamic Azad University. 

Authors' contributions
All authors equally contributed to preparing this article.

Conflicts of interest
The authors declared no conflict of interest.

Acknowledgements
We would like to thank the esteemed Vice Chancellor for Research of Shahrekord Branch of Azad University and everyone who contributed to this research.


Refrences

1. Le Blanc K, Rasmusson I, Sundberg B, Götherström C, Hassan M, Uzunel M, et al. Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells. Lancet. 2004; 363(9419):1439-41. [DOI:10.1016/S0140-6736(04)16104-7][PMID]
2. Kuo TK, Hung SP, Chuang CH, Chen CT, Shih YRV, Fang SCY, et al. Stem cell therapy for liver disease: parameters governing the success of using bone marrow mesenchymal stem cells. Gastroenterology. 2008; 134(7):2111-21. [DOI:10.1053/j.gastro.2008.03.015][PMID][PMCID]
3. Patel AN, Genovese J. Potential clinical application of adult human mesenchymal stem cell (Prochymal) therapy. Stem Cell Cloning. 2011; 4:61-72. [DOI:10.2147/SCCAA.S11991]
4. Nassiri Asl M, Aali E. [Review on the mesenchymal stem cells and their potential application in regenerative medicine (Persian)]. J Qazvin Univ Med Sci. 2018; 21(6):74-89. [DOI:10.29252/qums.21.6.89]
5. Pournasr Khakbaz B, Baharvand H. [Human mesenchymal stem cells and their clinical application (Persian)]. J Iran Anat Sci. 2007; 5(19):157-206. https://www.sid.ir/en/journal/ViewPaper.aspx?id=113590
6. Parson AB. The proteus effect: Stem cells and their promise for medicine. Washington, D.C.: Joseph Henry Press; 2004. [DOI:10.1172/JCI25763]
7. Song L, Tuan RS. Transdifferentiation potential of human mesenchymal stem cells derived from bone marrow. FASEB J. 2004; 18(9):980-2. [DOI:10.1096/fj.03-1100fje][PMID]
8. Oubari F, Amirizade N, Mohammadpour H, Nakhlestani M, Nikougoftar Zarif M. [The important role of FLT3-L in ex vivo expansion of hematopoietic stem cells following co-culture with mesenchymal stem cells (Persian)]. Cell Journal (Yakhteh). 2015; 17(2):201-10. [DOI:10.22074/cellj.2015.3715]
9. Oubari F, Nikougoftar Zarif M, Amirizadeh N, Shaiegan M, Atarodi K, Nakhlestani M, et al. [Isolation and expansion of Mesenchymal Stem cells from placenta (Persian)]. Sci J Iran Blood Transfus Organ. 2013; 10:222-30. http://bloodjournal.ir/article-1-791-en.html
10. Dehghani Fard A, Saki N, Ahmadvand M, Mahmoodinia Maymand M, Mosahebi Mohammadi M, Soleimani M. [Mesenchymal stem cell biology, application and its role in regenerative medicine (Persian)]. Sci J Iran Blood Transfus Organ. 2012; 8(4):306-20. http://bloodjournal.ir/article-1-586-en.html
11. Ryan JM, Barry FP, Murphy JM, Mahon BP. Mesenchymal stem cells avoid allogeneic ejection. J Inflamm (Lond). 2005; 2(8):1-11. [DOI:10.1186/1476-9255-2-8][PMID][PMCID]
12. Nasiri F, Amiri F, Mohammadipour M, Molaei S, Habibi Roudkenar M, Jalili MA. [H₂O₂-preconditioned mesenchymal stem cell regenerative effects on acute liver failure mice (Persian)]. Sci J Iran Blood Transfus Organ. 2015; 12(2):111-24. http://bloodjournal.ir/article-1-916-en.html
13. Baksh D, Song L, Tuan RS. Adult mesenchymal stem cells: Characterization, differentiation and application in cell and gene therapy. J Cell Mol Med. 2004; 8(3):301-16. [DOI:10.1111/j.1582-4934.2004.tb00320.x]
14. Chapel A, Bertho JM Bensidhoum M, Fouillard L, Young RG, Frick J, et al. Mesenchymal stem cells home to injure tissues when coinfused with hematopoietics cell to treat at radiation, inducedmulti, organfailure syndrome. J Gene Med. 2003; 5(12):1028-38. [DOI:10.1002/jgm.452][PMID]
15. Marquez-Curtis LA, Janowska-Wieczork A, McGann LE, Elliott JAW. Mesenchymal stromal cells derived from various tissues: Biological, clinical and cryopreservation aspects. Cryobiology. 2015; 71:181-97. [DOI:10.1016/j.cryobiol.2015.07.003][PMID]
16. Nasir GA, Mohsin S, Khan M, Shams S, Ali G, Khan SN, et al. Mesenchymal stem cells and Interleukin-6 attenuate liver fibrosis in mice. J Transl Med. 2013; 11(3):78-97. [DOI:10.1186/1479-5876-11-78][PMID][PMCID]
17. Bahmani M, Saki K, Shahsavari S, Rafieian-Kopaei M, Sepahvand R, Adineh A. Identification of medicinal plants effective in infectious diseases in Urmia, northwest of Iran. Asi Paci J Trop Biomed. 2015; 5(10):858-64. [DOI:10.1016/j.apjtb.2015.06.004]
18. Dadfar F, Hosseini S. E, Bahaoddini A. [A review of phytochemical, pharmacological and physiological properties of ginger (zingiber officinale) (Persian)]. Clin Excell. 2014; 3(1):72-86. http://ce.mazums.ac.ir/article-1-133-en.html
19. Haksar A, Sharma A, Chawla R, Kumar R, Arora R, Singh S, et al. Zingiber officinale exhibits behavioral radioprotection against radiation. Pharmacol Biochem Behav. 2006; 84(2):179-88. [DOI:10.1016/j.pbb.2006.04.008][PMID]
20. Stoilova I, Krastanov A, Stoyanova A, Denev P, Gargova S. Antioxidant activity of a ginger extract (Zingiber officinale). Food Chem 2007; 102(3):764-70. [DOI:10.1016/j.foodchem.2006.06.023]
21. Mirazi N, Karami Z. [The protective effect of hydroalcoholic extract from rhizome of Zingiber officinale L. on carbon tetrachloride-induced hepatic injury in male rat (Persian)]. J Kashan Univ Med Sci (FEYZ). 2016; 20(4):297-305.‏ http://feyz.kaums.ac.ir/article-1-3125-en.html
22. Johari H, Sharifi E, Delirnasab F, Hemayatkhah V, Kargar H, Nikpoor M. [The effect of hydro-alcoholic extracts of ginger on lead detoxification of kidney in the immature wistar rats (Persian)]. J Rafsanjan Uni Med Sci. 2013; 12(6):417-24. http://journal.rums.ac.ir/article-1-5290-en.html
23. Khoshtabiat L, Mahdavi M. [The role of oxidative stress in proliferation and cell death (Persian)]. J Mazandaran Univ Med Sci. 2015; 25(127):130-45.‏ http://jmums.mazums.ac.ir/article-1-5946-en.html
24. Halliwell B. Free radicals and antioxidants - quo vadis? Trends Pharmacol Sci. 2011; 32(3):125-30. [DOI:10.1016/j.tips.2010.12.002]
25. Aeschbach R, Löliger J, Scott BC, Murcia A, Butler J, Halliwell B, et al. Antioxidant actions of thymol, carvacrol, 6-gingerol, zingerone and hydroxytyrosol. Food Chem Toxicol. 1994; 32(1):31-6.‏ [DOI:10.1016/0278-6915(84)90033-4][PMID]
26. Kim JK, Kim Y, Na KM, Surh AJ, Kim TY. Gingerol prevents UVB induced Ros production and cox-2 expression invitro and invivo. Free Rad Res. 2007; 41(5):603-14. [DOI:10.1080/10715760701209896][PMID]
27. Dugasani S, Pichika MR, Nadarajah VD, Balijepalli MK, Tandra S, Korlakunta JN. Comparative antioxidant and anti-inflammatory effects of [6]-gingerol,[8]-gingerol,[10]-gingerol and [6]-shogaol. J Ethnopharmacol. 2010; 127(2):515-20.‏ [DOI:10.1016/j.jep.2009.10.004][PMID]
28. Ajayi BO, Adedara IA, Farombi EO. Pharmacological activity of 6-gingerol in dextran sulphate sodium-induced ulcerative colitis in BALB/c mice. Phytother Res. 2015; 29(4):566-72.‏ [DOI:10.1002/ptr.5286][PMID]
29. Abolaji AO, Ojo M, Afolabi TT, Arowoogun MD, Nwawolor D, Farombi EO. Protective properties of 6-gingerol-rich fraction from Zingiber officinale (Ginger) on chlorpyrifos-induced oxidative damage and inflammation in the brain, ovary and uterus of rats. Chem Biol Interact. 2017; 270:15-23.‏ [DOI:10.1016/j.cbi.2017.03.017]
30. Ajayi B. O, Adedara I. A. 6-Gingerol abates benzo [a] pyrene-induced colonic injury via suppression of oxido-inflammatory stress responses in BALB/c mice. Chemico-biological interactions 2019; 307:1-7.‏ [DOI:10.1016/j.cbi.2019.04.026]