Lasers in Medical Science

, Volume 31, Issue 4, pp 749–757 | Cite as

Combined effects of low-level laser therapy and human bone marrow mesenchymal stem cell conditioned medium on viability of human dermal fibroblasts cultured in a high-glucose medium

Original Article
First Online:
Received:
Accepted:

Abstract

Low-level laser therapy (LLLT) exhibited biostimulatory effects on fibroblasts viability. Secretomes can be administered to culture mediums by using bone marrow mesenchymal stem cells conditioned medium (BM-MSCs CM). This study investigated the combined effects of LLLT and human bone marrow mesenchymal stem cell conditioned medium (hBM-MSCs CM) on the cellular viability of human dermal fibroblasts (HDFs), which was cultured in a high-glucose (HG) concentration medium. The HDFs were cultured either in a concentration of physiologic (normal) glucose (NG; 5.5 mM/l) or in HG media (15 mM/l) for 4 days. LLLT was performed with a continuous-wave helium-neon laser (632.8 nm, power density of 0.00185 W/cm2 and energy densities of 0.5, 1, and 2 J/cm2). About 10 % of hBM-MSCs CM was added to the HG HDF culture medium. The viability of HDFs was evaluated using dimethylthiazol-diphenyltetrazolium bromide (MTT) assay. A significantly higher cell viability was observed when laser of either 0.5 or 1 J/cm2 was used to treat HG HDFs, compared to the control groups. The cellular viability of HG-treated HDFs was significantly lower compared to the LLLT + HG HDFs, hBM-MSCs CM-treated HG HDFs, and LLLT + hBM-MSCs CM-treated HG HDFs. In conclusion, hBM-MSCs CM or LLLT alone increased the survival of HG HDFs cells. However, the combination of hBM-MSCs CM and LLLT improved these results in comparison to the conditioned medium.

Keywords

Human bone marrow-mesenchymal stem cell Conditioned medium Diabetes mellitus Low-level laser therapy Human dermal fibroblasts Hyperglycemia 

Abbreviations

DM

Diabetes mellitus

IGFs I and II

Insulin growth factors 1 and 2

HG

High glucose

IL-6

Interleukin-6

bFGF

Basic fibroblast growth factor

HSFs

Human skin fibroblasts

MTT

Dimethylthiazol-diphenyltetrazolium bromide

ELISA

Enzyme-linked immunosorbent assay

NG

Normal glucose

ASCs

Adipose-derived stem cells

HDFs

Human dermal fibroblasts

hBM-MSCs CM

Human bone marrow mesenchymal stem cells conditioned medium

LLLT

Low-level laser therapy

BM

Bone marrow

PBS

Phosphate-buffered saline

DMEM

Dulbecco’s modified eagle’s medium

FCS

Fetal calf serum

EDTA

Ethylenediaminetetraacetic acid

FITC

Fluorescence isothiocyanate

PE

Phycoerythrin

OD

Optical density

SD

Standard deviation of mean

ANOVA

Analysis of variance

LSD

Least significant difference

NO

Nitric oxide

PCNA

Proliferating cell nuclear antigen

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

Notes

Acknowledgments

This article is financially supported by the “research department of the school of medicine” and Vice Chancellors of Research at the Shahid Beheshti University of Medical Sciences, Tehran, Iran (Grant no 6299).

Compliance with ethical standards

Conflict of interest

No competing financial interests exist. The authors declare no conflict of interest. The authors are solely responsible for the content and writing of this paper.

References

  1. 1.
    American Diabetes Association (2008) Standards of medical care in diabetes–2008. Diabetes Care 31:S12CrossRefGoogle Scholar
  2. 2.
    Goodson WH, Hunt TK (1979) Deficient collagen formation by obese mice in a standard wound model. Am J Surg 138:692–694CrossRefPubMedGoogle Scholar
  3. 3.
    Greenhalgh DG (2003) Wound healing and diabetes mellitus. Clin Plast Surg 30:37–45CrossRefPubMedGoogle Scholar
  4. 4.
    O'Loughlin A, McIntosh C, Dinneen SF, O'Brien T (2010) Review paper: basic concepts to novel therapies: a review of the diabetic foot. Int J Low Extrem Wounds 9:90–102CrossRefPubMedGoogle Scholar
  5. 5.
    Dall TM, Zhang Y, Chen YJ, Quick WW, Yang WG, Fogli J (2010) The economic burden of diabetes. Health Aff 29:297–303CrossRefGoogle Scholar
  6. 6.
    Kern P, Moczar M, Robert L (1979) Biosynthesis of skin collagens in normal and diabetic mice. Biochem J 182:337–345CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Franzén LE, Roberg K (1995) Impaired connective tissue repair in streptozotocin-induced diabetes shows ultrastructural signs of impaired contraction. J Surg Res 58:407–414CrossRefPubMedGoogle Scholar
  8. 8.
    Brown DL, Kane CD, Chernausek SD, Greenhalgh DG (1997) Differential expression and localization of insulin-like growth factors I and II in cutaneous wounds of diabetic and nondiabetic mice. Am J Pathol 151:715PubMedPubMedCentralGoogle Scholar
  9. 9.
    Algenstaedt P, Schaefer C, Biermann T, Hamann A, Schwarzloh B, Greten H (2003) Microvascular alterations in diabetic mice correlate with level of hyperglycemia. Diabetes 52:542–549CrossRefPubMedGoogle Scholar
  10. 10.
    Loots MA, Lamme EN, Mekkes JR, Bos JD, Middelkoop E (1999) Cultured fibroblasts from chronic diabetic wounds on the lower extremity (non-insulin-dependent diabetes mellitus) show disturbed proliferation. Arch Dermatol Res 291:93–99CrossRefPubMedGoogle Scholar
  11. 11.
    Benazzoug Y, Borchiellini C, Labat-Robert J, Robert L, Kern P (1998) Effect of high-glucose concentrations on the expression of collagens and fibronectin by fibroblasts in culture. Exp Gerontol 33:445–455CrossRefPubMedGoogle Scholar
  12. 12.
    Yevdokimova NY (2003) High glucose-induced alterations of extracellular matrix of human skin fibroblasts are not dependent on TSP-1–TGFβ1 pathway. J Diabetes Complicat 17:355–364CrossRefPubMedGoogle Scholar
  13. 13.
    Deveci M, Gilmont R, Dunham W, Mudge B, Smith D, Marcelo C (2005) Glutathione enhances fibroblast collagen contraction and protects keratinocytes from apoptosis in hyperglycaemic culture. Br J Dermatol 152:217–224CrossRefPubMedGoogle Scholar
  14. 14.
    Grossman N, Schneid N, Reuveni H, Halevy S, Lubart R (1998) 780 nm low power diode laser irradiation stimulates proliferation of keratinocyte cultures: involvement of reactive oxygen species. Lasers Surg Med 22:212–218CrossRefPubMedGoogle Scholar
  15. 15.
    Almeida‐Lopes L, Rigau J, Amaro Zângaro R, Guidugli‐Neto J, Marques Jaeger MM (2001) Comparison of the low level laser therapy effects on cultured human gingival fibroblasts proliferation using different irradiance and same fluence. Lasers Surg Med 29:179–184CrossRefPubMedGoogle Scholar
  16. 16.
    Hawkins DH, Abrahamse H (2006) The role of laser fluence in cell viability, proliferation, and membrane integrity of wounded human skin fibroblasts following helium‐neon laser irradiation. Lasers Surg Med 38:74–83CrossRefPubMedGoogle Scholar
  17. 17.
    Evans DH, Abrahamse H (2008) Efficacy of three different laser wavelengths for in vitro wound healing. Photodermatol Photoimmunol Photomed 24:199–210CrossRefPubMedGoogle Scholar
  18. 18.
    Houreld N, Abrahamse H (2007) In vitro exposure of wounded diabetic fibroblast cells to a helium-neon laser at 5 and 16 J/cm2. Photomed Laser Surg 25:78–84CrossRefPubMedGoogle Scholar
  19. 19.
    Houreld N, Abrahamse H (2007) Irradiation with a 632.8 nm helium-neon laser with 5 J/cm2 stimulates proliferation and expression of interleukin-6 in diabetic wounded fibroblast cells. Diabetes Technol Ther 9:451–459CrossRefPubMedGoogle Scholar
  20. 20.
    Houreld N, Abrahamse H (2008) Laser light influences cellular viability and proliferation in diabetic-wounded fibroblast cells in a dose-and wavelength-dependent manner. Lasers Med Sci 23:11–18CrossRefPubMedGoogle Scholar
  21. 21.
    Esmaeelinejad M, Bayat M, Darbandi H, Bayat M, Mosaffa N (2014) The effects of low-level laser irradiation on cellular viability and proliferation of human skin fibroblasts cultured in high glucose mediums. Lasers Med Sci 29:121–129CrossRefPubMedGoogle Scholar
  22. 22.
    Esmaeelinejad M, Bayat M (2013) Effect of low-level laser therapy on the release of interleukin-6 and basic fibroblast growth factor from cultured human skin fibroblasts in normal and high glucose mediums. J Cosmet Laser Ther 15:310–317CrossRefPubMedGoogle Scholar
  23. 23.
    AlGhamdi KM, Kumar A, Moussa NA (2012) Low-level laser therapy: a useful technique for enhancing the proliferation of various cultured cells. Lasers Med Sci 27:237–49CrossRefPubMedGoogle Scholar
  24. 24.
    Tamama K, Kerpedjieva SS (2012) Acceleration of wound healing by multiple growth factors and cytokines secreted from multipotential stromal cells/mesenchymal stem cells. Adv Wound Care 1:177–182CrossRefGoogle Scholar
  25. 25.
    Chen L, Tredget EE, Wu P, Wu Y, Wu Y (2008) Paracrine factors of mesenchymal stem cells recruit macrophages and endothelial lineage cells and enhance wound healing. PLoS One 3:e1886CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Walter M, Wright KT, Fuller H, MacNeil S, Johnson WEB (2010) Mesenchymal stem cell-conditioned medium accelerates skin wound healing: an in vitro study of fibroblast and keratinocyte scratch assays. Exp Cell Res 316:1271–1281CrossRefPubMedGoogle Scholar
  27. 27.
    Yew T-L, Hung Y-T, Li H-Y, Chen HW, Chen LL, Tsai KS (2011) Enhancement of wound healing by human multipotent stromal cell conditioned medium: the paracrine factors and p38 MAPK activation. Cell Transplant 20:693–706CrossRefPubMedGoogle Scholar
  28. 28.
    Lee SH, Jin SY, Song JS, Seo KK, Cho KH (2012) Paracrine effects of adipose-derived stem cells on keratinocytes and dermal fibroblasts. Ann Dermatol 24:136–143CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Chen L, Xu Y, Zhao J et al (2014) Conditioned medium from hypoxic bone marrow-derived mesenchymal stem cells enhances wound healing in mice. PLoS One 9:e96161CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Ganji R, Piryaei A, Bayat M, RajabiBazl M, Mohsenifar Z, Kheirjoo R (2014) The effect of human bone marrow-mesenchymal stem cells secretoms on diabetic wound healing. Pejouhesh 38:10–18Google Scholar
  31. 31.
    Eslaminejad MB, Nikmahzar A, Piriea A (2006) The structure of human mesenchymal stem cells differentiated into cartilage in micro mass culture system. Yakhteh 8:162–171Google Scholar
  32. 32.
    Piryaei A, Soleimani M, Heidari MH, Saheli M, Rohani R, Almasieh M (2015) Ultrastructural maturation of human bone marrow mesenchymal stem cells‐derived cardiomyocytes under alternative induction of 5‐azacytidine. Cell Biol Int 39:519–530CrossRefPubMedGoogle Scholar
  33. 33.
    Vinck EM, Cagnie BJ, Cornelissen MJ, Declercq HA, Cambier DC (2005) Green light emitting diode irradiation enhances fibroblast growth impaired by high glucose level. Photomed Laser Ther 23:167–171CrossRefGoogle Scholar
  34. 34.
    Lian Z, Yin X, Li H et al (2014) Synergistic effect of bone marrow-derived mesenchymal stem cells and platelet-rich plasma in streptozotocin-induced diabetic rats. Ann Dermatol 26:1–10CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Lee YH, Chang JJ, Chien CT, Yang MC, Chien HF. Antioxidant sol–gel improves cutaneous wound healing in streptozotocin-induced diabetic rats. Experimental Diabetes Research 2012; 2012.Google Scholar
  36. 36.
    Sharifian Z, Bayat M, Alidoust M, Farahani RM, Bayat M, Rezaie F (2014) Histological and gene expression analysis of the effects of pulsed low-level laser therapy on wound healing of streptozotocin-induced diabetic rats. Lasers Med Sci 29:1227–1235CrossRefPubMedGoogle Scholar
  37. 37.
    Hocking AM, Gibran NS (2010) Mesenchymal stem cells: paracrine signaling and differentiation during cutaneous wound repair. Exp Cell Res 316:2213–2219CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Kiyosaki T, Mitsui N, Suzuki N, Shimizu N (2010) Low-level laser therapy stimulates mineralization via increased Runx2 expression and ERK phosphorylation in osteoblasts. Photomed Laser Surg 28:S-167–S-172CrossRefGoogle Scholar
  39. 39.
    Tsai W-C, Cheng J-W, Chen J-L, Chen CY, Chang HN, Liao YH (2014) Low-level laser irradiation stimulates tenocyte proliferation in association with increased NO synthesis and upregulation of PCNA and cyclins. Lasers Med Sci 29:1377–1384CrossRefPubMedGoogle Scholar
  40. 40.
    Houreld N, Abrahamse H (2010) Low-intensity laser irradiation stimulates wound healing in diabetic wounded fibroblast cells (WS1). Diabetes Technol Ther 12:971–978CrossRefPubMedGoogle Scholar
  41. 41.
    Mirzaei M, Bayat M, Mosafa N, Mohsenifar Z, Piryaei A, Farokhi B (2007) Effect of low-level laser therapy on skin fibroblasts of streptozotocin-diabetic rats. Photomed Laser Surg 25:519–525CrossRefPubMedGoogle Scholar
  42. 42.
    Vasilenko T, Slezák M, Kovác I, Bottková Z, Jakubco J, Kostelníková M (2010) The effect of equal daily dose achieved by different power densities of low-level laser therapy at 635 and 670 nm on wound tensile strength in rats: a short report. Photomed Laser Surg 28:281–3CrossRefPubMedGoogle Scholar
  43. 43.
    Gál P, Mokrý M, Vidinský B, Kilík R, Depta F, Harakalová M et al (2009) Effect of equal daily doses achieved by different power densities of low-level laser therapy at 635 nm on open skin wound healing in normal and corticosteroid-treated rats. Lasers Med Sci 24:539–47CrossRefPubMedGoogle Scholar
  44. 44.
    Vinck EM, Cagnie BJ, Cornelissen MJ, Declercq HA, Cambier DC (2003) Increased fibroblast proliferation induced by light emitting diode and low power laser irradiation. Lasers Med Sci 18(2):95–9CrossRefPubMedGoogle Scholar
  45. 45.
    Pogrel MA, Chen JW, Zhang K (1997) Effects of low-energy gallium-aluminum-arsenide laser irradiation on cultured fibroblasts and keratinocytes. Lasers Surg Med 20(4):426–32CrossRefPubMedGoogle Scholar
  46. 46.
    Hallman HO, Basford JR, O'Brien JF, Cummins LA (1988) Does low-energy helium-neon laser irradiation alter “in vitro” replication of human fibroblasts? Lasers Surg Med 8(2):125–9CrossRefPubMedGoogle Scholar
  47. 47.
    Karu T (1987) Photobiological fundamentals of low-power laser therapy. IEEE J Quantum Electron 23:1703–1717CrossRefGoogle Scholar
  48. 48.
    Ginani F, Soares DM, Barreto MP, Barboza CA (2015) Effect of low-level laser therapy on mesenchymal stem cell proliferation: a systematic review. Lasers Med Sci 30(8):2189–94. doi: 10.1007/s10103-015-1730-9 CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag London 2016

Authors and Affiliations

  1. 1.Department of Biology and Anatomical Sciences, School of MedicineShahid Beheshti University of Medical SciencesTehranIran
  2. 2.Urogenital Stem Cell Research CenterShahid Beheshti University of Medical SciencesTehranIran
  3. 3.Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and TechnologyACECRTehranIran
  4. 4.Department of Immunology, School of MedicineShahid Beheshti University of Medical SciencesTehranIran

Personalised recommendations