title: Apoptosis of U937 cells induced by hematoporphyrin monomethyl ether-mediated sonodynamic action.
abstract: OBJECTIVE
  The present study aims to investigate apoptosis of U937 cells induced by hematoporphyrin monomethyl ether (HMME)-mediated sonodynamic therapy (SDT).
  
  
  METHODS
  HMME concentration was kept constant at 10 μg/mL. Tumor cells suspended in serum-free RPM1640 were exposed to ultrasound at 1.1 MHz for up to 60 seconds with an intensity of 1 W/cm(2) in the presence and absence of HMME. The viability of cells was determined by the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltertrazolium bromide tetrazolium (MTT) test. Apoptosis was analyzed using a flow cytometer with Annexin V-PE/7-ADD staining as well as fluorescence microscopy with 4'-6-diamidino-2-phenylindole (DAPI) staining. The DNA damage of U937 cells, intracellular reactive oxygen species (ROS), and mitochondria membrane potential (MMP) were also analyzed by a flow cytometer after exposures. Western blotting and reverse transcriptase-polymerase chain reaction were used to analyze the protein and mRNA expression level of caspase-3 and poly(ADP-ribose) polymerase (PARP).
  
  
  RESULTS
  Fluorescent imaging revealed that HMME mainly localized in the mitochondria. MTT assay showed 55.6% of cell survival at 4 hours post-SDT. Flow cytometric analysis displayed a significant increase in the early- and late-apoptotic cell populations (35.6%) of U937 cells by HMME-mediated SDT. Compared with the control, ultrasound-alone, and HMME-alone groups, the intracellular ROS and the MMP loss were greatly increased in the combined SDT group. Obvious nuclear condensation was also found with DAPI staining, and the DNA fragment increased to 33.9% at 2 hours post-SDT treatment. Immunofluorescent staining indicated obvious Bax translocation after SDT. Western blot showed visible enhancement of caspase-3 and PARP cleavage. In addition, caspase-3 and PARP mRNA expression of U937 cells increased remarkably after SDT treatment.
  
  
  CONCLUSIONS
  The findings demonstrated that HMME-mediated sonodynamic action (HMME-SDT) significantly induced apoptosis of U937 cells, suggesting that HMME may be a good sonosensitizer, and HMME-SDT might be a potential therapeutic strategy for cancer treatment.
authors.affiliation: Key Laboratory of Medicinal Resources and Natural Pharmaceutical Chemistry, Ministry of Education, National Engineering Laboratory for Resource Developing of Endangered Chinese Crude Drugs in Northwest of China, College of Life Sciences, Shaanxi Normal University, Xi'an, Shaanxi, China.
authors.forename: Xiaomin
authors.initials: X
authors.lastname: Su
authors.forename: Pan
authors.initials: P
authors.lastname: Wang
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Wang
authors.forename: Bing
authors.initials: B
authors.lastname: Cao
authors.forename: Long
authors.initials: L
authors.lastname: Li
authors.forename: Quanhong
authors.initials: Q
authors.lastname: Liu
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D002470:Cell Survival; D006415:Hematoporphyrins / Q000493:pharmacokinetics / Q000494:pharmacology*; D006801:Humans; D007150:Immunohistochemistry; D010778:Photochemotherapy / Q000379:methods*; D020298:U937 Cells; D014464:Ultrasonic Therapy / Q000379:methods*
chemical_list: D006415:Hematoporphyrins; C494379:hematoporphyrin monomethyl ether