Chrysin was originally found to be responsible

To determine if the increased expression of these integrin subunits was actually involved in the enhancement of cell adhesion to extracellular matrices, we examined cell adhesion to fibronectin Chrysin and vitronectin in the presence or absence of the synthetic tripeptide RGD. This peptide was originally found to be responsible for the fibronectin or vitronectin cell attachment activity. When endothelial cell suspensions were plated on dishes coated with fibronectin or vitronectin, many of the adherent cells did not spread and remained round. After treatment with baicalein, the number of adherent cells was increased and most of the adherent cells spread with extension of lamellipodia.
As shown in Figure 4a, baicalein enhanced endothelial adhesion to fibronectin and vitronectin was significantly suppressed by RGD peptide, but not by the control peptide SDGRG. In addition, treatment with a blocking antibody to integrin a5b1 almost completely blocked XL880 the baicalein enhanced endothelial cell adhesion to fibronectin but not to vitronectin. However, anti integrin a2 antibody did not affect baicalein induced cell adhesion. These findings suggest that baicalein induced integrin upregulation might play an essential role in the increased adhesion. Within each focal contact, cell surface receptors of the integrin family members interact with ECM molecules outside the cell and with the cytoskeletal system in the cytoplasm.
Thus, we examined the expression of cytoskeletal molecules in baicalein treated cells by western blot analysis, using antiactin, anti a actinin, anti talin, anti paxillin, anti vinculin, anti tubulin, anti FAK, anti phospho FAK and phospho FAK specific antibodies. As shown in Figure 5, baicalein treatment increased the expression level only of vinculin protein, whereas the levels of actin, a actinin, FAK, phospho FAK, phospho FAK, paxillin, talin and tubulin were not affected by this treatment. Reorganization of microfilaments and promotion of focal adhesion contact formation by baicalein To characterize the effects of baicalein on the microfilament organization and focal adhesion contact formation, fluorescence cytostaining using rhodamine labelled phalloidin and immunostaining of integrins and vinculin were investigated by laser scanner confocal microscopy.
As illustrated in Figure 6a, staining with rhodamine labelled phalloidin for cytoskeletal architecture showed extensive stress fibre formation in untreated endothelial cells that was in a random orientation throughout the monolayer, moreover, vinculinspecific immunofluorescence was found in fine fibrillar streaks in untreated cells and associated with the ends of actin microfilaments around the periphery of each cell andat the ends of centrally located stress fibres. However, exposure to baicalein drastically altered the arrangement of F actin within the monolayer, by inducing it to concentrate under the plasma membrane. The streaks of vinculin appeared to be longer and thicker after baicalein treatment. The formation of stress fibres occurred simultaneously with the appearance of vinculin streaks at their terminals, in correspondence with adhesion sites. To quantify changes in the number of intensely stained focal adhesions

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