During bleb formation, actin and myosin buy Doramapimod filaments slide over each other, resulting in contraction of the cell border toward the center. This process impairs the binding of actin filaments to the cell membrane. The mechanism by which cinnamic acid causes microfilament disorganization is not well understood; however, because taxol does not exhibit direct effects on microfilaments, this suggests interdependency between actin filaments and microtubules [52]. The disorganization of microtubules in cells treated

with cinnamic acid may be directly caused by impairment in the tubulin molecules or indirectly by an alteration in the molecules associated with microtubule polymerization. It is known that the dynamic equilibrium of tubulin may be altered at high concentrations of free cytosolic calcium (higher than 10-7 M), which results in the depolymerization of microtubules [54]. Studies using other natural compounds have shown that the induction of cell death by caffeic acid and curcumin in HL-60 cells [8] and L929 mouse fibroblasts (Thayyllathil at al., 2008), respectively, is associated with mitochondrial disruption, which may be due to an augmented concentration of calcium that results in cytoskeletal disruption. These results are similar to the

observations found in our system. Our results allow us to affirm that microtubule depolymerization, as well as microfilament disorganization, occurred MK-8931 ic50 after exposure to 3.2 mM cinnamic acid. Microtubule disruptions have been previously described as a trigger of the apoptotic pathway, which eventually results in cell death [54]. Our data suggest that there is no relationship between the effects of cinnamic acid on cytoskeletal elements and apoptotic induction. We have demonstrated that M30 staining and microtubule

disorganization are, at least in part, independent events. Caffeic acid, another cinnamamide compound, causes apoptosis in HL-60 ZD1839 cells via mitochondrial dysfunction [8]. Previous studies have shown a relationship between cancer chemotherapeutic agents targeting microtubules and apoptosis [55, 56]. The flow cytometry assay did not show G2/M arrest; however, microtubule disorganization was caused by cinnamic acid treatment. Thus, the apoptotic events observed in our study were not caused by cytoskeletal reorganization. Tseng et al. [57] studied podophylotoxin and suggested that mitotic arrest is not a prerequisite for apoptosis, although they often can occur concomitantly. The present data suggest that microtubule disorganization after cinnamic acid exposure is dependent on the drug concentration. In our system, cytoskeletal disorganization is mainly responsible for the formation of nuclear aberrations. We clearly observed apoptotic HT-144 cells, as assessed by phosphorylated cytokeratin 18. The M30 antibody stains cells in early apoptosis.