Study On The Changes Of Nuclear Matrix Proteins During The Apoptosis Of The Immortalized Human Epithelial HaCaT Cells With Curcumin Treatment

In this study, the curcumin(Cur) was used to induce the immortalized human epithelial cell line HaCaT cells into apoptosis, and its effects were investigated by selective extraction-whole mount optic and transmission electron microscopy and techniques of proteomic. The differentially expressed nuclear matrix proteins were analyzed in order to explore the molecular mechanisms in a system level.The results revealed that the HaCaT cells were induced into apoptosis after treated with 7.5μg/mL curcumin as the proliferation of HaCaT cells were inhibited, and the inhibitory rate is 83.03%. The results of flow cytometry analysis showed that curcumin could induce the emergence of the phase of apoptosis, and the rate is 13.1% ,the cell cycle were arrested in G₂/M phase. Agarose gel electrophoresis revealed that cell DNA fragment exhibited characteristic " DNA ladder " . Light microscope and electron microscope showed that the morphology of the cells treated with curcumin appeared shrinked, cell nucleus concerntrated,chromatin agglutination, mitochondria swelling, and apoptosis body forming. Immunocytochemistry revealed that the expression of wt p53, Bax, Fas were upregulated significatntly while the products of Bcl-2 proteins were downregulated significantly in the cells treated with Cur. Selective extraction, whole-mount optic and transmission electron microscopy showed that after treated with 7.5μg/mL curcumin the configuration of nuclear matrix-intermediate filament in HaCaT cells was remarkably changed as the filament was few and scattered, not well distributed and arranged irregularly.and connected to lamina respectively, and the filaments of nuclear matrix,nuclear lamin and intermediatefilment connected not very tightly and the formation but tends to fracture but also relatively more complete network structure.There were 27 spots changed remarkably during the apoptosis induced by curcumin,21 of which were identified. the nine up-regulated proteins including nucleoporin NUP107, six down-regulated proteins including LMNA protein and six new proteins were identified. In conclusion, the configuration and composition of nuclear matrix were changed following the HaCaT cell apoptosis.It is confirmed that specific nuclear matrix proteins take part in the regulation of cell proliferation and apoptosis, and the associated signal transduction pathways are suggested. It provides proofs and a new way to study the machnisms of epithelial cell apoptosis and it is possible to get a system understanding of the molecular mechanisms from the studies of nuclear matrix, as well as to reveal the relationship of a series of life activities, such as DNA replication, gene expression, signal transduction, and cell cycle regulation.