Photodynamic action of methylene blue in osteosarcoma cells in vitro
Introduction
Osteosarcoma is one of the commonest malignant bone tumors threatening the life of young people worldwide [1]. The survival of patients with osteosarcoma has been enhanced by advanced modern remedies [2], [3], however, the long-term efficacy of the current therapeutic modalities is very limited due to drug-resistance and cancer recurrence [4], [5]. Therefore, novel and more effective strategies are in urgent need for eradicating osteosarcoma cells.
Photodynamic therapy (PDT) is an alternative for combating malignant tumors [6], [7], [8] via photochemical action-induced cytotoxic reactive oxygen species (ROS) from photosensitizers retaining in over-growing tumor cells activated by appropriate light [9]. In PDT photosensitizer and light source are two key components. In order to explore photodynamic therapy as a new therapeutic technique, we have successfully set up new light sources using light-emitting diodes (LED). Our previous study showed that red light from this LED light source could effectively activate pyropheophorbide-α methyl ester (MPPa) and pheophorbide a (Pa) to kill various tumor cells in vitro [10], [11]. Blue light from this LED light source could remarkably activate hypocrellin B to deactivate ovarian and breast cancer cells [12], [13]. Methylene blue (MB) is a second-generation photosensitizer from phenothiazine dyes with remarkable photocytotoxicity of tumor cells. Emerging studies have shown that MB was retained in the malignant tissues [14]. Recently, Matsubara et al. reported that MB under the irradiation of light from a 500 W xenon lamp source had a strong photocytotoxic effect on mouse osteosarcoma (LM8) cells through affecting cytoplasmic ballooning, and inducing necrosis and apoptosis [15]. In this present study, we used red light from a new LED light source to activate MB for observing the killing efficacy of LED-activated MB on osteosarcoma-derived UMR106 cells in vitro and explore apoptotic cell death and mitochondrial membrane potential (MMP) of UMR106 cells induced by photodynamic action of MB.
Section snippets
Photosensitizer and light source
Methylene blue (MB) (C16H18ClN3S, M.W. 319.90, ≥82%) was purchased from Sigma–Aldrich Co. (St. Louis, MO, USA). A stock solution (3 M) was made in phosphate buffer saline (PBS) and stored in the dark at −20 °C until used.
The light source was from a red LED with the wavelength of 630 nm. Its highest power density is 106.5 mW/cm2 with a uniform distribution over an area of 78.5 cm2. The samples were exposed to the LED light at the energy density of 0, 2, 4, 8, 16 and 32 J/cm2, respectively.
Cell culture
Rat
Photocytotoxicity of MB in UMR 106 cells
The photocytotoxicity of UMR106 cells by LED-activated MB was measured using SRB assay and observed under a light microscopy. The cells were incubated in different MB concentrations (0–100 μM) for 1 h and exposed to red light from LED light source with various light energies (0–32 J/cm2). No significant dark cytotoxicity of MB was observed in the range of 0–100 μM. MB under the light irradiation caused a concentration-dependent cytotoxicity in UMR106 cells, and photocytotoxicity of MB in UMR106
Discussion
About 30% of osteosarcomas are found to be multidrug-resistant [16]. However, surgical resection is very difficult to fully remove osteosarcoma cells. The residual malignant cells are a main cause of osteosarcoma recurrence. Gong's study demonstrated that benzochloroporphyrin derivative-17-mediated PDT exhibited a cytotoxic effect on cultured LM-8 osteosarcoma cells, and the rate of local recurrence and the surgical margins were reduced by PDT after marginal resection of tumors [17]. Therefore,
Acknowledgements
This work was supported by the GRF grant from Hong Kong RGC (476912) and Innovation and Technology Fund of Shenzhen (CXZZ20120619150627260), and the Direct Grant from the Chinese University of Hong Kong (2030446, 4053026). We express our sincere thanks to Miss Winnie Lee for her helpful assistance.
References (32)
- et al.
Adjuvant and neoadjuvant chemotherapy for osteosarcoma of the extremities: 27 year experience at Rizzoli Institute, Italy
European Journal of Cancer
(2005) - et al.
Survival from high-grade localised extremity osteosarcoma: combined results and prognostic factors from three European Osteosarcoma Intergroup randomised controlled trials
Annals of Oncology
(2012) - et al.
Inhibition of cell growth induced by photosensitizer PP(Arg)2-mediated photodynamic therapy in human breast and prostate cell lines. Part I
Photodiagnosis and Photodynamic Therapy
(2011) - et al.
Low dose hypericin-PDT induces complete tumor regression in BALB/c mice bearing CT26 colon carcinoma
Photodiagnosis and Photodynamic Therapy
(2011) - et al.
Photodiagnosis and photodynamic therapy of peritoneal metastasis of ovarian cancer
Photodiagnosis and Photodynamic Therapy
(2012) - et al.
Apoptosis of pancreatic adenocarcinoma cells induced by LED-activated pheophorbide a
Laser Physics Letters
(2011) - et al.
Apoptosis of breast cancer cells induced by hypocrellin B under light-emitting diode irradiation
Photodiagnosis and Photodynamic Therapy
(2012) - et al.
Methylene blue reverts multidrug resistance: sensitivity of multidrug resistant cells to this dye and its photodynamic action
Cancer Letters
(2000) - et al.
Treatment of melanoma lesions using methylene blue and RL50 light source
Photodiagnosis and Photodynamic Therapy
(2004) - et al.
Intraluminal treatment of inoperable oesophageal tumours by intralesional photodynamic therapy with methylene-blue
Lancet
(1995)
Methylene blue and the photodynamic therapy of superficial bladder cancer
Journal of Photochemistry and Photobiology B: Biology
Apoptosis of ovarian cancer cells induced by methylene blue-mediated sonodynamic action
Ultrasonics
Targeting autophagy potentiates chemotherapy-induced apoptosis and proliferation inhibition in hepatocarcinoma cells
Cancer Letters
Hypocrellin B-mediated sonodynamic action induces apoptosis of hepatocellular carcinoma cells
Ultrasonics
Apoptosis induced by the sonomechanical effects of low intensity pulsed ultrasound in a human leukemia cell line
Cancer Letters
Hematoporphyrin monomethyl ether enhances the killing of ultrasound on osteosarcoma cells involving intracellular reactive oxygen species and calcium ion elevation
Integrative Cancer Therapies
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