The Effect Of Serum Starvation On Autophagy In Human Keratinocyte Cell Line, HaCaT Cells

The cell homeostasis is mainiained by a precisely regulated balance between synthesis and degradation of cellular components. The constitutive degradation of proteins by the proteasome. However, this process is limited to eytoplasmic short-lived proteins, and the majority of cellular components are long lived. Therefore, there must be another route for large-scale macromolecular recycling. This route is autophagy. This proeess has been observed in all eukaryotic cells, and play an important role in the maintenance of homeostasis. Autophagy pathways can be classified into there different types:macroautophagy, microautophagy and chaperone-mediated autophagy. Macroautophagy is the major lysosomal route for the turnover of cytoplasmic components and will hereafter be referred as autophagy.Objective This project is designed to investigate the effect of serum starvation on autophagy in human keratinocyte cell line, HaCaT cells, and to analyze the molecule mechanism of regulation, Then through the regulation of cell control and treatment of skin diseases and lay the foundation of autophagy.Method The cultured HaCaT cells were treated by serum starvation. To investigate the induction of autophagy, autophagosome was observed through transmission electron microscopy, and expression of LC3II, a special marker of autophagy, was assayed by western-blot. The product of phospho-mTOR(ser2448) was semi-quantitative analyzed by western-blot, for interpreting preliminarily the molecule mechanism involved in induction of autophagy.Result In serum starvation-treating HaCaT cells, formation of autophagosome was seen and conversion of LC3â…¡ was up-regulated versus control. Moreover, the product of phosphor-mTOR(ser2448) decreased induced by serum starvation.Conclusion Autophagy of HaCaT cells can be induced by serum starvation, and induction of autophagy may be involved in decrease of phospho-mTOR(ser2448), a key regulatory element.