title: MGDG extracted from spinach enhances the cytotoxicity of radiation in pancreatic cancer cells.
abstract: BACKGROUND
  In our previous study, monogalactosyl diacylglycerol (MGDG) purified from spinach was found to have cytotoxic effects in human cancer cell lines. This study further assessed whether MGDG can enhance the cytotoxic effects of radiation in human pancreatic cancer cells in vitro and in vivo.
  
  
  METHODS
  Glycoglycerolipids from spinach including MGDG were extracted from dried spinach. The cytotoxicity of MGDG were evaluated by the MTT assay using four human pancreatic cancer cell lines (MIAPaCa-2, AsPC-1, BxPC-3 and PANC-1) and normal human dermal fibroblasts (NHDFs). The effects of radiation and MGDG alone or in combination in MIAPaCa-2 cells was analyzed with the colony forming and apoptosis assays, western blotting and cell cycle and DNA damage analyses (γ-H2AX foci staining and comet assay). The inhibitory effects on tumor growth were assessed in a mouse xenograft tumor model.
  
  
  RESULTS
  MGDG showed dose- and time-dependent cytotoxicity, with half-maximal inhibitory concentrations (IC50) in PANC-1, BxPC-3, MIAPaCa-2 and AsPC-1 cells at 72 h of 25.6 ± 2.5, 26.9 ± 1.3, 18.5 ± 1.7, and 22.7 ± 1.9 μM, respectively. The colony forming assay revealed fewer MIAPaCa-2, BxPC-3 and AsPC-1 cell colonies upon treatment with both MGDG and radiation as compared to irradiation alone (P < 0.05). The combination of MGDG and radiation induced a higher proportion of apoptosis in MIAPaCa-2 cells; this effect was associated with increased mitochondrial release of cytochrome c and activation of cleaved poly (ADP-ribose) polymerase and caspase-3. DNA damage was detected and DNA repair mechanisms were more frequently impaired in cells receiving the combination treatment as compared to either one alone. Tumor growth was inhibited to a greater degree in mice treated by intratumoral injection of MGDG combined with irradiation as compared to either one alone (P < 0.05).
  
  
  CONCLUSIONS
  This is the first report demonstrating that MGDG enhances the cytotoxicity of radiation to induce apoptosis of cancer cells in vitro and in vivo. Our findings indicate that this therapeutic combination can be an effective strategy for the treatment of pancreatic cancer.
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Akasaka
authors.affiliation: Graduate School of Agriculture, Shinshu University, Minamiminowa-mura, Kamiina-gun, Nagano, 399-4598, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Mizushina
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Yoshida
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Ejima
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Naritoshi
authors.initials: N
authors.lastname: Mukumoto
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Tianyuan
authors.initials: T
authors.lastname: Wang
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Sachiko
authors.initials: S
authors.lastname: Inubushi
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Masao
authors.initials: M
authors.lastname: Nakayama
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Wakahara
authors.affiliation: Division of Radiation Oncology, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, Hyogo, 650-0017, Japan. rsasaki@med.kobe-u.ac.jp.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Sasaki
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D015153:Blotting, Western; D045744:Cell Line, Tumor; D059248:Chemoradiotherapy / Q000379:methods*; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects / Q000528:radiation effects; D004260:DNA Repair / Q000187:drug effects / Q000528:radiation effects; D038983:Galactolipids / Q000494:pharmacology*; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010190:Pancreatic Neoplasms / Q000473:pathology*; D010936:Plant Extracts / Q000494:pharmacology*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D018724:Spinacia oleracea; D023041:Xenograft Model Antitumor Assays
chemical_list: D038983:Galactolipids; D010936:Plant Extracts; D011838:Radiation-Sensitizing Agents; C009909:monogalactosyldiacylglycerol
keywords: Apoptosis; Monogalactosyl diacylglycerol (MGDG); Pancreatic cancer; Radiation; Radiosensitizer