The purpose of the present study was to evaluate the neural protein expression pattern of human multipotent mesenchymal stromal cells (hMSCs) treated with forskolin (free-form/FF). The study investigated forskolin’s capacity to enhance intracellular levels of cyclic adenosine monophosphate (cAMP) by activating adenylate cyclase and probably by inducing neuron-like cells in vitro. In addition, because nanotechnology is a growing field of tissue engineering, we also assessed the action of a new system called the nanostructured-forskolin (NF) to examine the improvement of drug delivery. Afterwards, the cells were submitted to low-level laser irradiation to evaluate possible photobiostimulatory effects. Investigations using the immunofluorescence by confocal microscopy and Western blot methods revealed the expression of the neuronal marker β-tubulin III. Fluorescence intensity quantification analysis using INCell Analyzer System for β-tubulin III was used to examine significant differences. The results showed that after low-level laser irradiation exposure, there was a tendency to increase the β-tubulin III expression in all groups, as expected in the photobiostimulation process. Notably, this process induced for irradiation was more pronounced in irradiated nanoforskolin cells (INF) compared to non-irradiated free-forskolin control cells (NFFC). However, there was also an increase in β-tubulin III protein expression in the groups: irradiated nanocontrol cells (INC) compared to non-irradiated free-forskolin control cells (NFF) and after treatment with non-irradiated free-forskolin (NFF) and non-irradiated nanoforskolin (NNFC). We concluded that the methods using low-level laser irradiation and/or nanoparticles showed an up-regulation of neural-protein expression in hMSCs that could be used to facilitate cellular therapy protocols in the near future.
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Abramovitch-Gottlib, L., Gross, T., Naveh, D., et al. (2005). Low level laser irradiation stimulates osteogenic phenotype of mesenchymal stem cells seeded on a three-dimensional biomatrix. Lasers in Medical Science, 20(3–4), 138–146.
Agostinis, P., Berg, K., Cengel, K. A., et al. (2011). Photodynamic therapy of cancer: an update. CA Cancer Journal for Clinicians., 61(4), 250–281.
Allen, T. M., & Cullis, P. R. (2013). Liposomal drug delivery systems: from concept to clinical applications. Advanced Drug Delivery Reviews, 65(1), 36–48.
Arthur, A., Rychkov, G., Shi, S., Koblar, S. A., & Gronthos, S. (2008). Adult human dental pulp stem cells differentiate toward functionally active neurons under appropriate environmental cues. Stem Cells, 26(7), 1787–1795.
Beavo, J. A., & Brunton, L. L. (2002). Cyclic nucleotide research-still exponding after half a century. Nature Reviews, 3(9), 710–718.
Berry, S. E., Andruszkiewicz, P., Chun, J. L., & Hong, J. (2013). Nestin expression in end-stage disease in dystrophin-deficient heart: implications for regeneration from endogenous cardiac stem cells. Stem Cells Translational Medicine, 2(11), 848–861.
Bose, S., & Tarafder, S. (2012). Calcium phosphate ceramic systems in growth factor and drug delivery for bone tissue engineering: a review. Acta Biomaterialia, 8(4), 1401–1421.
Caddick, J., Kingham, P. J., Gardiner, N. J., Wiberg, M., & Terenghi, G. (2006). Phenotypic and functional characteristic of MSCs differentiated along a Schwann cell lineage. Glia, 54(8), 840–849.
Caplan, A. I., & Bruder, S. P. (2001). Mesenchymal stem cells: building blocks for molecular medicine in the 21st century. Trends Molecular Medicine, 7(6), 259–264.
Carrol, J. D., Milward, M. R., Cooper, P. R., Hadis, M., & Palin, W. M. (2014). Developments in low level light therapy (LLLT) for dentistry. Dental Materials, 30(5), 465–475.
Cavalcanti, M. F., Maria, D. A., de Isla, N., et al. (2015). Evaluation of the proliferative effects induced by low-level laser therapy in bone marrow stem cell culture. Photomedicine Laser Surgery, 33(12), 610–616.
Corseli, M., Chen, C. W., Sun, B., Yap, S., Rubin, J. P., & Péault, B. (2012). The tunica adventitia of human arteries and veins as a source of MSCs. Stem Cells Development, 21(8), 1299–1308.
Covas, D. T., Panepucci, R. A., Fontes, A. M., et al. (2008). Multipotent mesenchymal stromal cells obtained from diverse human tissues share functional properties and gene-expression profile with CD146+ perivascular cells and fibroblasts. Experimental Hematology, 36(5), 642–654.
da Silva Meirelles, L., Chagastelles, P. C., & Nardi, N. B. (2006). Mesenchymal stem cells reside in virtually all post-natal organs and tissues. Journal of Cell Science, 119(11), 2204–2213.
Dezawa, M., Kanno, H., Hoshino, M., et al. (2004). Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation. Journal of Clinical Investigation, 113(12), 1701–1710.
Dezawa, M., Takahashi, I., Esaki, M., Takano, M., & Sawada, H. (2001). Sciatic nerve regeneration in rats induced by transplantation of in vitro differentiated bone-marrow stromal cells. European Journal of Neuroscience, 14(11), 1771–1776.
Ginani, F., Soares, D. M., Barreto, M. P., & Barbosa, C. A. (2015). Effect of low-level laser therapy on mesenchymal stem cell proliferation: a systematic review. Lasers in Medical Science, 30(8), 2189–2194.
Jang, S., Cho, H. H., Cho, Y. B., Park, J. S., & Jeong, H. S. (2010). Functional neural differentiation of human adipose tissue-derived stem cells using bFGF and forskolin. Biomed Central Cell Biology, 11(25), 1–13.
Jang, Y. H., Koo, G. B., Kim, J. Y., Kim, Y. S., & Kim, Y. C. (2013). Prolonged Activation of ERK Contributes to the Photorejuvenation Effect in photodynamic therapy in human dermal fibroblasts. Journal of Investigative Dermatology, 133(9), 2265–2275.
Jiang, Y. Q., & Oblinge, M. M. (1992). Differential regulation of βIII and other tubulin genes during peripheral and central neuron development. Journal Cell Science, 103(3), 643–651.
Joo, D., Woo, J. S., Cho, K. H., Han, S. H., Min, T. S., Yang, D. S., Yun, C. H., et al. (2016). Biphasic activation of extracellular signal-regulated kinase (ERK) 1/2 in epidermal growth factor (EGF)-stimulated SW480 colorectal cancer cells. Biochemistry Molecular Biology Reports, 49(4), 220–225.
Karu, T. I., & Kolyakov, S. F. (2005). Exact action spectra for cellular responses relevant to phototherapy. Photomedicine and Laser Surgery, 23(4), 355–361.
Kingham, P. J., Kalbermatten, D. F., Mahay, D., Armstrong, S. J., Wiberg, M., & Terenghi, G. (2007). Adipose-derived stem cells differentiate into a Schwann cell phenotype and promote neurite outgrowth in vitro. Experimental Neurology, 207(2), 267–274.
Lei, X., Liu, B., Huang, Z., & Wu, J. (2015). A clinical study of photodynamic therapy for chronic skin ulcers in lower limbs infected with Pseudomonas aeruginosa. Archives of Dermatological Research, 307(1), 49–55.
Lendahl, U., Zimmerman, L. B., & Mckay, R. D. G. (1990). CNS stem cells express a new class of intermediate filament protein. Cell, 60(4), 585–595.
Lipson, R. L., Baldes, E. J., & Olsen, A. M. (1961). Use of a derivative of hematoporphyrin in tumor detection. Journal of the National Cancer Institute, 26(1), 1–11.
Loreti, E. H., Pascoal, V. L., Nogueira, B. V., Silva, I. V., & Pedrosa, D. F. (2015). Use of laser therapy in the healing process: A literature review. Photomedicine and Laser Surgery, 33(2), 104–116.
Ludueña, R. F. (1997). Multiple forms of tubulin: different gene products and covalent modification. International Review of Cytology, 178, 207–275.
Maltman, D. J., Hardy, S. A., & Przyborski, S. A. (2011). Role of mesenchymal stem cells in neurogenesis and nervous systems repair. Neurochemistry International, 59(3), 347–356.
Mamalis, A. D., Lev-Tov, H., Nguyen, D. H., & Jaqdeo, J. R. (2014). Laser and light-based treatment of Keloids – a review. Journal of the European Academy of Dermatology and Venereology, 28(6), 689–699.
Merino, S., Martín, C., Kostarelos, K., Prato, M., & Vázquez, E. (2015). Nanocomposite Hydrogels: 3D Polymer Nanoparticle Synergies for On-Demand Drug Delivery. American Chemistry Society Nano, 9(5), 4686–4697.
Mester, E., Spiry, T., Szende, B., Spiry, F., & Sacher, A. (1971). Effect of laser rays on wound healing. The American Journal of Surgery, 122(4), 532–535.
Mignani, S., Kazzouli, E., Bousmina, S., Majoral, M., JP (2013). Expand classical drug administration ways by emerging routes using dendrimer drug delivery systems: A concise overview. Advanced Drug Delivery Review, 65(10), 1316–1330.
Minguell, J. J., Fierro, F. A., Epunã, M. J., Erices, A. A., & Sierralta, W. D. (2005). Nonstimulated human uncommitted mesenchymal stem cell express cell markers of mesenchymal and neural lineages. Stem Cells Development, 14(4), 408–414.
Mura, S., Nicolas, J., & Couvreur, P. (2013). Stimuli-responsive nanocarriers for drug delivery. Nature Materials, 12(11), 991–1003.
Nadur-Andrade, N., Barbosa, A. M., Carlos, F. P., Lima, C. J., Coqo, J. C., & Zamuner, S. R. (2012). Effects of photobiostimulation on edema and hemorrhage induced by Bothrops moojeni venom. Lasers in Medical Science, 27(1), 65–70.
Panyam, J., & Labhasetwar, V. (2003). Biodegradable nanoparticles for drug and gene delivery to cells and tissue. Advanced Drug Delivery Review;, 55(3), 329–347.
Passarella, S., & Karu, T. (2014). Absorption of monochromatic and narrow band radiation in the visible and near IR by both mitochondrial and non-mitochondrial photoacceptors results in photobiomodulation. Journal of Photochemistry and Photobiology, 140, 344–358.
Peplow, P. V., Chung, T. Y., & Baxter, G. D. (2012). Photodynamic modulation of wound healing: a review of human and animal studies. Photomedicine and Laser Surgery, 30(3), 118–148.
Phinney, D. G., & Prockop, D. J. (2007). Concise review: mesenchymal stem/multipotent stromal cells: the state of transdifferentiation and modes of tissue repair – current views. Stem Cells, 25(11), 2896–2902.
Pittenger, M. F., Mackay, A. M., Beck, S. C., et al. (1999) Multilineage potential of adult human mesenchymal stem cells. Science, 284(5411), 143 – 47.
Primo, F. L., Siqueira-Moura, M. P., Simioni, A. R., & Tedesco, A. C. (2007). Preparation, characterization and cytotoxicity assays of chloroaluminum phthalocyanine photosensitizer drug loaded in PLGA-nanocapsules. Drugs Future, 32, 74.
Primo, F. L., Reis, M. B. D., Porcionatto, M. A., & Tedesco, A. C. (2011). In Vitro Evaluation of chloroaluminum phthalocyanine nanoemulsion and low-level laser therapy on human skin dermal equivalents and bone marrow mesenchymal stem cells. Current Medicinal Chemistry, 18(22), 3376–3381.
Primo, F. L., Bentley, M. V. L. B., & Tedesco, A. C. (2008). Photophysical studies and in vitro skin permeation/retention of Foscan®/Nanoemulsion (NE) applicable to photodynamic therapy skin cancer treatment. Journal of Nanoscience and Nanotechnology, 8(1), 340–347.
Prockop, D. J. (1997). Marrow stromal cells as stem cells for nonhematopoietic tissues. Science, 276(5309), 71–74.
Rooney, G. E., Howard, L., O`Brien, T., Windebank, A. J., & Barry, F. P. (2009) Elevation of cAMP in mesenchymal stem cells transiently up regulates neural markers rather than inducing neural differentiation. Stem Cells Development, 18(3), 387 – 98.
Sanchez-Ramos, J., Song, S., Cardozo-Pelaez, F., et al. (2000). Adult bone marrow stromal cells differentiate into neural cells in vitro. Experimental Neurology, 164(2), 247–256.
Sartore, R. C., Campos, P. B., Trujillo, C. A., et al. (2011). Retinoic acid-treated pluripotent stem cells undergoing neurogenesis present increased aneuploidy and micronuclei formation. PLoS One, 6(6), 1–10.
Seamon, K. B., Padgett, W., & Daly, J. W. (1981) Forskolin: Unique diterpene activator of adenylate cyclase in membranes and in intact cells. Proceedings of the National Academy of Sciences of United States of America, 78(6), 3363–3367.
Sharma, S. K., Kharkwal, G. B., & Sago, M. (2011). Dose response effects of 810 nm laser light on mouse primary cortical neurons. Laser Surgery Medicine, 43(8), 851–859.
Stein, A., Benayahu, D., Maltz, L., & Oron, U. (2005). Low-Level Laser Irradiation Promotes Proliferation and Differentiation of Human Osteoblasts in Vitro. Photomedicine and Laser Surgery, 23(2), 161–166.
Stein, E., Koehn, J., Sutter, W., et al. (2009). Phenothiazine chloride and soft laser light have a biostimulatory effect on human osteoblastic cells. Photomedicine and Laser Surgery, 27(1), 71–77.
Tondreau, T., Lagneaux, L., Dejeneffe, M., et al. (2004). Bone-marrow-derived mesenchymal stem cells already express specific neural proteins before any differentiation. Differentiation, 72(7), 319–326.
Wong-Riley, M. T., Liang, H. L., Eells, J. T., et al. (2005). Photobiomodulation directly benefits primary neurons functionally inactivated by toxins: role of cytochrome c oxidase. The Journal of Biological Chemistry, 280(6), 4761–4771.
Woodbury, D., Schwarz, E. J., Prockop, D. J., & Black, I. B. (2000). Adult rat and human bone marrow stromal cells differentiate into neurons. Journal of Neuroscience Research, 61(4), 364–370.
Wu, Q., Xuan, W., Ando, T., et al. (2012). Low-level laser therapy for closed-head traumatic brain injury in mice: effect of different wavelengths. Lasers in Surgery and Medicine, 44(3), 218–226.
Yu, W., Naim, J. O., & Lanzafame, R. J. (1994). The effect of laser irradiation on the release of bFGF from 3T3 fibroblasts. Photochemistry and Photobiology, 59(2), 167–170.
Zancanela, D. C., Primo, F. L., Rosa, A. L., Ciacanglini, P., & Tedesco, A. C. (2011). The Effect of Photosensitizer Drugs and Light Stimulation on Osteoblast Growth. Photomedicine and Laser Surgery, 29(10), 699–705.
Zhang, G., Zeng, X., & Li, P. (2013). Nanomaterials in cancer-therapy drug delivery systems. Journal of Biomedical Nanotechnology, 9(5), 741–750.
Zhang, L., & Webster, T. J. (2009). Nanotechnology and nanomaterials: promises for improved tissue regeneration. Nanotoday, 4(1), 66–80.
Zhang, Y., Song, S. P., Fong, C. C., Tsang, C. H., Yang, Z., & Yang, M. (2003). cDNA microarray analysis of gene expression profiles in human fibroblast cells irradiated with red light. Journal of Investigative Dermatology, 120(5), 849–857.
Zuk, P. A., Zhu, M., Ashjian, P., et al. (2002) Human adipose tissue is a source of multipotent stem cells. Molecular Biology of the Cell, 13(12), 4279–4295.
The authors are grateful to Hospital Amaral de Carvalho (Jau, São Paulo) and are especially grateful to Drs Antônio Cesarino Mota and Vergílio Rensi Colturato for providing bone marrow samples from donors as well as Nayara Rezende for help with the nanoforskolin and Sandra Navarro Brescian for artwork. I want to thank Fernanda Udinal for language advice and Priscilla Carnavale Gomes Ferreira for reviewing the manuscript. Valéria Ferreira-Silva is grateful for a postdoctoral fellowship from Coordenacão de Aperfeicoamento de Pessoal de Nível Superior (CAPES). This work was supported by Financiadora de Estudos e Projetos (FINEP 01.10.0758.01), CEFAP-USP (São Paulo), Instituto Nacional de Ciência e Tecnologia, Célula-Tronco e Terapia Celular (INCTC), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP# 2013/50181-1), Brazil.
The authors have no potential conflicts of interest to report.
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Ferreira-Silva, V., Primo, F.L., Baqui, M.M. et al. Beneficial Role of Low-Intensity Laser Irradiation on Neural β-tubulin III Protein Expression in Human Bone Marrow Multipotent Mesenchymal Stromal Cells. Stem Cell Rev and Rep 14, 585–598 (2018). https://doi.org/10.1007/s12015-017-9796-3
- Low-intensity laser irradiation
- Nano-drug delivery
- β-Tubulin III
- Multipotent mesenchymal stromal cells