title: Atractylodes macrocephala Koidz stimulates intestinal epithelial cell migration through a polyamine dependent mechanism.
abstract: BACKGROUND
  Atractylodes macrocephala Koidz (AMK), a valuable traditional Chinese herbal medicine, has been widely used in clinical practice for treating patients with disorders of the digestive system. AMK has shown noteworthy promoting effect on improving gastrointestinal function and immunity, which might represent a promising candidate for the treatment of intestinal mucosa injury. The aim of this study was to investigate the efficacy of AMK on intestinal mucosal restitution and the underlying mechanisms via intestinal epithelial (IEC-6) cell migration model.
  
  
  METHODS
  A cell migration model of IEC-6 cells was induced by a single-edge razor blade along the diameter of the cell layers in six-well polystyrene plates. After wounding, the cells were grown in control cultures and in cultures containing spermidine (5μM, SPD, reference drug), alpha-difluoromethylornithine (2.5mM, DFMO, polyamine inhibitor), AMK (50, 100, and 200mg/L), DFMO plus SPD and DFMO plus AMK for 12h. The polyamines content was detected by high-performance liquid chromatography (HPLC) with pre-column derivatization. The Rho mRNAs expression levels were assessed by Q-RT-PCR. The Rho and non-muscle myosin II proteins expression levels were analyzed by Western blot. The formation and distribution of non-muscle myosin II stress fibers were monitored with immunostaining techniques using specific antibodies and observed by confocal microscopy. Cell migration assay was carried out using inverted microscope and the Image-Pro Plus software. All of these indexes were used to evaluate the effectiveness of AMK.
  
  
  RESULTS
  (1) Treatment with AMK caused significant increases in cellular polyamines content and Rho mRNAs and proteins expression levels, as compared to control group. Furthermore, AMK exposure increased non-muscle myosin II protein expression levels and formation of non-muscle myosin II stress fibers, and resulted in an acceleration of cell migration in IEC-6 cells. (2) Depletion of cellular polyamines by DFMO resulted in a decrease of cellular polyamines levels, Rho mRNAs and proteins expression, non-muscle myosin II protein formation and distribution, thereby inhibiting IEC-6 cell migration. AMK not only reversed the inhibitory effects of DFMO on the polyamines content, Rho mRNAs and proteins expression, non-muscle myosin II protein formation and distribution, but also restored cell migration to control levels.
  
  
  CONCLUSIONS
  The results obtained from this study revealed that AMK significantly stimulates the migration of IEC-6 cells through a polyamine dependent mechanism, which could accelerate the healing of intestinal injury. These findings suggest the potential value of AMK in curing intestinal diseases characterized by injury and ineffective repair of the intestinal mucosa in clinical practice.
authors.affiliation: Institute of TCM Diagnostics, Hunan University of Chinese Medicine, 300 Xueshi Road, Yuelu District, Changsha 410208, PR China; Spleen and Stomach Institute, Guangzhou University of Chinese Medicine, 12 Airport Road, Baiyun District, Guangzhou 510405, PR China. Electronic address: pansyy-2008@163.com.
authors.forename: Hou-Pan
authors.initials: HP
authors.lastname: Song
authors.affiliation: Spleen and Stomach Institute, Guangzhou University of Chinese Medicine, 12 Airport Road, Baiyun District, Guangzhou 510405, PR China. Electronic address: lrl@gzucm.edu.cn.
authors.forename: Ru-Liu
authors.initials: RL
authors.lastname: Li
authors.affiliation: The First Affiliated Hospital of Guangzhou University of Chinese Medicine, 16 Airport Road, Baiyun District, Guangzhou 510405, PR China.
authors.forename: Chi
authors.initials: C
authors.lastname: Zhou
authors.affiliation: Institute of TCM Diagnostics, Hunan University of Chinese Medicine, 300 Xueshi Road, Yuelu District, Changsha 410208, PR China.
authors.forename: Xiong
authors.initials: X
authors.lastname: Cai
authors.affiliation: Institute of TCM Diagnostics, Hunan University of Chinese Medicine, 300 Xueshi Road, Yuelu District, Changsha 410208, PR China.
authors.forename: Hui-Yong
authors.initials: HY
authors.lastname: Huang
mesh_terms: D000818:Animals; D031174:Atractylodes; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects*; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D008516:Medicine, Chinese Traditional; D024462:Myosin Type II / Q000378:metabolism; D010936:Plant Extracts / Q000494:pharmacology*; D011073:Polyamines / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D014945:Wound Healing; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D010936:Plant Extracts; D011073:Polyamines; D012333:RNA, Messenger; D024462:Myosin Type II; D020741:rho GTP-Binding Proteins
keywords: Atractylodes macrocephala Koidz; Cell migration; Intestinal epithelial cells; Non-muscle myosin II; Polyamines; Rho GTPases