Abstract
The cell death mechanisms of necrosis and apoptosis generate biochemical and morphological changes in different manners. However, the changes that occur in cell adhesion and nuclear envelope (NE) topography, during necrosis and apoptosis, are not yet fully understood. Here, we show the different alterations in cell adhesion function, as well as the topographical changes occurring to the NE, during the necrotic and apoptotic cell death process, using the xCELLigence system and atomic force microscopy (AFM). Studies using xCELLigence technology and AFM have shown that necrotic cell death induced the expansion of the cell adhesion area, but did not affect the speed of cell adhesion. Necrotic nuclei showed a round shape and presence of nuclear pore complexes (NPCs). Moreover, we found that the process of necrosis in combination with apoptosis (termed nepoptosis here) resulted in the reduction of the cell adhesion area and cell adhesion speed through the activation of caspases. Our findings showed, for the first time, a successful characterization of NE topography and cell adhesion during necrosis and apoptosis, which may be of importance for the understanding of cell death and might aid the design of future drug delivery methods for anti-cancer therapies.
Highlights
During apoptotic cell death, two characteristic stages, the first consisting of fragmentation of focal adhesion, the second involving nuclear envelope (NE) destruction, as well as nuclear DNA and protein fragmentation, occurring through the activation of the caspase-dependent pathway, are generally observed[8]
DOX-treated cells exhibited a higher percentage of propidium iodide (PI)-positive cells than the control, and this increase was almost independent of caspase-3/7 activity and Annexin V-positive cells (Supplementary Fig. S1B, C)
Cell adhesion plays an important role in cell migration, growth, differentiation, and morphology[9], it is associated with various human diseases, such as ischemia, asthma, diabetes, bacterial infections, and others[37,38]
Summary
Two characteristic stages, the first consisting of fragmentation of focal adhesion, the second involving nuclear envelope (NE) destruction, as well as nuclear DNA and protein fragmentation, occurring through the activation of the caspase-dependent pathway, are generally observed[8]. Studies of cell death have shown that the activation of caspase-3 triggers the cleavage of the key factors of focal adhesion proteins, which are important for the control of cell behavior[13], such as Crk-associated substrate (CAS) and focal adhesion kinase (FAK). Previous studies have shown that DNA damage-inducing chemicals generated caspase activation and triggered cleavage of NPC proteins (e.g., NUP 93, 153, and 214) and INM proteins (e.g., lamina-associated polypeptide 2), during apoptosis[25,26,27]. We introduce a novel and direct application of the xCELLigence cell analysis system and carbon nanotube-attached AFM (CNT/AFM) probes, for cell adhesion and NE topography analyses
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