Inhibition of human embryonic palatal mesenchymal cell cycle by secalonic acid D: a probable mechanism of its cleft palate induction.
OBJECTIVE
To assess the mechanism(s) of cleft palate induction by secalonic acid D (SAD) in human embryonic palatal mesenchymal (HEPM) cells and compare them with those evaluated in the murine embryonic palate.


METHODS
Effect of SAD on HEPM cell proliferation was studied by obtaining dose response curves for cell numbers, uptake of 3H-thymidine and the expression of proliferating cell nuclear antigen (PCNA). Effects of SAD on cell cycle were assessed by flowcytometry. Cell-labeling with 3H-glucosamine and immunoblot analysis were conducted to study SAD effects on the synthesis of glycosaminogycans (GAG) and the expression of fibronectin and tenascin, respectively.


RESULTS
SAD induced a concentration-dependent decrease in HEPM cell number and 3H-thymidine uptake beginning at 0.1 microg of SAD/ml. Expression of PCNA and progression of cell cycle from G1 to S phase were inhibited following SAD exposure. Cell viability was significantly reduced only at 7.5 microg/ml of SAD or higher indicating that the reduction in cell numbers by SAD at lower concentrations is likely due to reduced proliferation and at higher concentrations due to both reduced proliferation and cell death. Synthesis of extra cellular matrix components (GAGs, fibronectin or tenascin) by HEPM cells, however, was not inhibited by SAD.


CONCLUSIONS
The results of these studies confirmed those of our previous studies with mice and the MEPM cells that SAD may induce cleft palate by reducing numbers of palatal mesenchymal cells by inhibition of their proliferation thereby leading to a reduction in the size of the developing palate shelves.
Dhulipala
V C
VC
Department of Veterinary Biomedical Sciences, University of Missouri, Columbia, MO 65211, USA.
Welshons
W V
WV
Reddy
C S
CS
D000818:Animals; D002452:Cell Count; D002453:Cell Cycle / Q000187:drug effects; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002972:Cleft Palate / Q000139:chemically induced*; D004305:Dose-Response Relationship, Drug; D005353:Fibronectins / Q000187:drug effects; D006025:Glycosaminoglycans / Q000032:analysis; D006801:Humans; D008648:Mesoderm / Q000187:drug effects*; D051379:Mice; D009183:Mycotoxins / Q000008:administration & dosage / Q000009:adverse effects*; D010159:Palate / Q000187:drug effects* / Q000196:embryology; D018809:Proliferating Cell Nuclear Antigen / Q000187:drug effects; D019063:Tenascin / Q000187:drug effects; D013936:Thymidine / Q000378:metabolism; D044004:Xanthones / Q000008:administration & dosage / Q000009:adverse effects*
D005353:Fibronectins; D006025:Glycosaminoglycans; D009183:Mycotoxins; D018809:Proliferating Cell Nuclear Antigen; D019063:Tenascin; D044004:Xanthones; C009218:secalonic acid; D013936:Thymidine