Skip to main content

Advertisement

SpringerLink
Log in

N-acetyl-l-cysteine increases MnSOD activity and enhances the recruitment of quiescent human fibroblasts to the proliferation cycle during wound healing

  • Original Article
  • Published:
Molecular Biology Reports Aims and scope Submit manuscript

Abstract

The rebuilding of the connective tissue during wound healing requires the recruitment of fibroblasts to the wound area as well as reentry of quiescent fibroblasts to the proliferative cycle. Whether this process can be modulated by a small molecular weight thiol antioxidant N-acetyl-l-cysteine (NAC) was tested in normal human skin fibroblasts (NHFs) using a uni-directional wound healing assay. NAC treated cells demonstrated a decreased migration rate but increased number of proliferating cells recruited into the wound area post wounding. Fifteen day quiescent control and NAC treated NHFs were re-plated at a lower density and cell numbers counted at different days post-plating. Interestingly, NAC treated cells exhibited increased cellular proliferation indicated by both decreased cell population doubling time and increased S phase cells. NAC treated cells demonstrated decreased steady state levels of reactive oxygen species as well as increased protein and activity levels of manganese superoxide dismutase (MnSOD). NAC treatment failed to induce proliferation in quiescent cells lacking MnSOD expression. These results demonstrate that NAC enhanced the recruitment of quiescent NHFs into proliferation cycle during wound healing. Our results also suggest that the wound healing properties of NAC might be due to its ability to induce and enhance MnSOD expression and activity. Altogether, these findings suggest NAC might be potentially developed as a dietary intervention to improve tissue injury in animals and humans.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

BrdU:

Bromodeoxyuridine

Cdk:

Cyclin-dependent kinase

CuZnSOD:

Copper and zinc superoxide dismutase

DHE:

Dihydroethidium

ECM:

Extracellular matrix

EPCs:

Endothelia progenitor cells

GSH:

Reduced glutathione

HBSS:

Hanks buffer salt solution

IL-1β:

Interleukin-1β

MMP-1:

Matrixmetalloproteinase-1

MnSOD:

Manganese superoxide dismutase

NAC:

N-acetyl-l-cysteine

NHFs:

Normal human skin fibroblasts

NO:

Nitric oxide

NOS:

Endothelial NO synthase

PI:

Propidium iodide

PKC:

Protein kinase C

PM:

Protein malnutrition

ROS:

Reactive oxygen species

TNF-α:

Tumor necrosis factor-α

References

  1. Tsai ML, Huang HP, Hsu JD, Lai YR, Hsiao YP, Lu FJ, Chang HR (2014) Topical N-acetylcysteine accelerates wound healing in vitro and in vivo via the PKC/Stat3 pathway. Int J Mol Sci 15(5):7563–7578. doi:10.3390/ijms15057563

    Article  PubMed  PubMed Central  Google Scholar 

  2. Demir EO, Cakmak GK, Bakkal H, Turkcu UO, Kandemir N, Demir AS, Tascilar O (2011) N-acetyl-cysteine improves anastomotic wound healing after radiotherapy in rats. J Investig Surg 24(4):151–158. doi:10.3109/08941939.2011.560237

    Article  Google Scholar 

  3. Aktunc E, Ozacmak VH, Ozacmak HS, Barut F, Buyukates M, Kandemir O, Demircan N (2010) N-acetyl cysteine promotes angiogenesis and clearance of free oxygen radicals, thus improving wound healing in an alloxan-induced diabetic mouse model of incisional wound. Clin Exp Dermatol 35(8):902–909. doi:10.1111/j.1365-2230.2010.03823.x

    Article  PubMed  CAS  Google Scholar 

  4. Lim Y, Levy MA, Bray TM (2006) Dietary supplementation of N-acetylcysteine enhances early inflammatory responses during cutaneous wound healing in protein malnourished mice. J Nutr Biochem 17(5):328–336. doi:10.1016/j.jnutbio.2005.08.004

    Article  PubMed  CAS  Google Scholar 

  5. Millea PJ (2009) N-acetylcysteine: multiple clinical applications. Am Fam Physician 80(3):265–269

    PubMed  Google Scholar 

  6. Aruoma OI, Halliwell B, Hoey BM, Butler J (1989) The antioxidant action of N-acetylcysteine: its reaction with hydrogen peroxide, hydroxyl radical, superoxide, and hypochlorous acid. Free Radic Biol Med 6(6):593–597

    Article  PubMed  CAS  Google Scholar 

  7. Fabiani R, De Bartolomeo A, Rosignoli P, Servili M, Montedoro GF, Morozzi G (2002) Cancer chemoprevention by hydroxytyrosol isolated from virgin olive oil through G1 cell cycle arrest and apoptosis. Eur J Cancer Prev 11(4):351–358

    Article  PubMed  CAS  Google Scholar 

  8. Menon SG, Sarsour EH, Kalen AL, Venkataraman S, Hitchler MJ, Domann FE, Oberley LW, Goswami PC (2007) Superoxide signaling mediates N-acetyl-L-cysteine-induced G1 arrest: regulatory role of cyclin D1 and manganese superoxide dismutase. Cancer Res 67(13):6392–6399. doi:10.1158/0008-5472.CAN-07-0225

    Article  PubMed  CAS  Google Scholar 

  9. Kelly GS (1998) Clinical applications of N-acetylcysteine. Altern Med Rev 3(2):114–127

    PubMed  CAS  Google Scholar 

  10. Sarsour EH, Agarwal M, Pandita TK, Oberley LW, Goswami PC (2005) Manganese superoxide dismutase protects the proliferative capacity of confluent normal human fibroblasts. J Biol Chem 280(18):18033–18041. doi:10.1074/jbc.M501939200

    Article  PubMed  CAS  Google Scholar 

  11. Rodriguez LG, Wu X, Guan JL (2005) Wound-healing assay. Methods Mol Biol 294:23–29

    PubMed  Google Scholar 

  12. Sarsour EH, Kumar MG, Kalen AL, Goswami M, Buettner GR, Goswami PC (2012) MnSOD activity regulates hydroxytyrosol-induced extension of chronological lifespan. Age (Dordr) 34(1):95–109. doi:10.1007/s11357-011-9223-7

    Article  CAS  Google Scholar 

  13. Beauchamp C, Fridovich I (1971) Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Anal Biochem 44(1):276–287

    Article  PubMed  CAS  Google Scholar 

  14. Visner GA, Dougall WC, Wilson JM, Burr IA, Nick HS (1990) Regulation of manganese superoxide dismutase by lipopolysaccharide, interleukin-1, and tumor necrosis factor. Role in the acute inflammatory response. J Biol Chem 265(5):2856–2864

    PubMed  CAS  Google Scholar 

  15. Eastgate J, Moreb J, Nick HS, Suzuki K, Taniguchi N, Zucali JR (1993) A role for manganese superoxide dismutase in radioprotection of hematopoietic stem cells by interleukin-1. Blood 81(3):639–646

    PubMed  CAS  Google Scholar 

  16. Xiong L, Sun J, Hirche C, Yang J, Yang Y, Xia Y, Lehnhardt M, Wang R, Fu X (2012) In vitro N-acetyl-L-cysteine promotes proliferation and suppresses interleukin-8 expression in adipose-derived stem cells. Aesthet Plast Surg 36(5):1260–1265. doi:10.1007/s00266-012-9960-8

    Article  Google Scholar 

  17. Sarsour EH, Kalen AL, Goswami PC (2014) Manganese superoxide dismutase regulates a redox cycle within the cell cycle. Antioxid Redox Signal 20(10):1618–1627. doi:10.1089/ars.2013.5303

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  18. Nagata K, Iwasaki Y, Yamada T, Yuba T, Kono K, Hosogi S, Ohsugi S, Kuwahara H, Marunaka Y (2007) Overexpression of manganese superoxide dismutase by N-acetylcysteine in hyperoxic lung injury. Respir Med 101(4):800–807. doi:10.1016/j.rmed.2006.07.017

    Article  PubMed  Google Scholar 

  19. Yang YY, Lee KC, Huang YT, Wang YW, Hou MC, Lee FY, Lin HC, Lee SD (2008) Effects of N-acetylcysteine administration in hepatic microcirculation of rats with biliary cirrhosis. J Hepatol 49(1):25–33. doi:10.1016/j.jhep.2008.02.012

    Article  PubMed  CAS  Google Scholar 

  20. Chaudhuri L, Nicholson AM, Kalen AL, Goswami PC (2012) Preferential selection of MnSOD transcripts in proliferating normal and cancer cells. Oncogene 31(10):1207–1216. doi:10.1038/onc.2011.325

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  21. Sarsour EH, Kalen AL, Xiao Z, Veenstra TD, Chaudhuri L, Venkataraman S, Reigan P, Buettner GR, Goswami PC (2012) Manganese superoxide dismutase regulates a metabolic switch during the mammalian cell cycle. Cancer Res 72(15):3807–3816. doi:10.1158/0008-5472.CAN-11-1063

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  22. Xu Y, Kiningham KK, Devalaraja MN, Yeh CC, Majima H, Kasarskis EJ, St Clair DK (1999) An intronic NF-kappaB element is essential for induction of the human manganese superoxide dismutase gene by tumor necrosis factor-alpha and interleukin-1beta. DNA Cell Biol 18(9):709–722. doi:10.1089/104454999314999

    Article  PubMed  CAS  Google Scholar 

  23. Dhar SK, Xu Y, St Clair DK (2010) Nuclear factor kappaB- and specificity protein 1-dependent p53-mediated bi-directional regulation of the human manganese superoxide dismutase gene. J Biol Chem 285(13):9835–9846. doi:10.1074/jbc.M109.060715

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  24. Luo JD, Wang YY, Fu WL, Wu J, Chen AF (2004) Gene therapy of endothelial nitric oxide synthase and manganese superoxide dismutase restores delayed wound healing in type 1 diabetic mice. Circulation 110(16):2484–2493. doi:10.1161/01.CIR.0000137969.87365.05

    Article  PubMed  CAS  Google Scholar 

  25. Marrotte EJ, Chen DD, Hakim JS, Chen AF (2010) Manganese superoxide dismutase expression in endothelial progenitor cells accelerates wound healing in diabetic mice. J Clin Invest 120(12):4207–4219. doi:10.1172/JCI36858

    Article  PubMed  CAS  PubMed Central  Google Scholar 

Download references

Acknowledgments

We thank Mr. Jian Shao from the Central Microscopy facility for assisting in the light and confocal imaging. We also thank Mr. John Lafin for critical reading of the manuscript. This work was supported by NIH 2R01 CA111365 and McCord research foundation. The authors’ contributions are as follows: G. M. and E. H. S. drafted the manuscript; G. M., M. G., A. L. K., and E. H. S. performed the experiments; P. C. G. and E. H. S. provided advice on experimental design and supervised the study. All authors read and approved the final version of the manuscript. The authors have no financial or personal conflicts of interest to declare.

Author information

Authors and Affiliations

  1. B180 Med Labs, Free Radical and Radiation Biology Division, Department of Radiation Oncology, University of Iowa, Iowa City, IA, 52242, USA

    Gaowei Mao, Monali Goswami, Amanda L. Kalen, Prabhat C. Goswami & Ehab H. Sarsour

Authors
  1. Gaowei Mao
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Monali Goswami
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Amanda L. Kalen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Prabhat C. Goswami
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ehab H. Sarsour
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ehab H. Sarsour.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Mao, G., Goswami, M., Kalen, A.L. et al. N-acetyl-l-cysteine increases MnSOD activity and enhances the recruitment of quiescent human fibroblasts to the proliferation cycle during wound healing. Mol Biol Rep 43, 31–39 (2016). https://doi.org/10.1007/s11033-015-3935-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11033-015-3935-1

Keywords

Search

Navigation