Curcumin down-regulates PCNA,cyclin D1 and Bcl-X_L expression in human keratinocyte cell lines

Objective：To evaluate the effects of curcumin on regulating the proliferation,cell cycle distribution,apoptosis and relevant mechanisms in keratinocyte cell lines.Methods：The human immortalized human keratinocyte lines（HaCaT cells） were treated with different doses of curcumin.The effects of curcumin on cell viability were measured by MTT assay,and the cell cycle distribution and apoptosis determined by flow cytometry.The mRNA expression changes of proliferating cell nuclear antigen（PCNA）,cyclin D1 and Bcl-xL were from real-time PCR analysis and the protein levels were detected by Western blotting.Results：Data obtained in the study showed that curcumin could cause significantly inhibitory effect on proliferation in HaCaT cells in a time- and dose-dependent manner.Cell arrest at G1/S phase and significant apoptosis were observed after being treated with curcumin for 24 h.In association with these,the expression of PCNA,cyclin D1 and Bcl-xL were decreased both at mRNA and protein levels for the same treatment.Conclusion：Curcumin can inhibit proliferation,induce cell arrest at G1/S phase and cause apoptosis in HaCaT cells.The decreased expression of PCNA,cyclin D1 and Bcl-xL induced by curcumin contributes to the above effects in vitro.

The presence of phosphorylation form of D1 protein in its cross-linked aggregates in high light treated spinach leaves in vivo

In the present study, using specific antibody against D1 protein, we detected four ag- gregates of D1 protein in thylakoid membranes from spinach leaves illuminated at high light (800―2500 μmol photons·m?2·s?1) for 3 h. Their accumulations were dependent on the light intensity to which the leaves had been subjected. Further immunoblot analysis indicated that 70 kD aggregate was a prod- uct of D1 protein cross-linked with CP43, 65 and 60 kD aggregate were two cross-linked products be- tween D1 and D2 proteins, and 41 kD aggregate was one cross-linked D1 with α-subunit of cytochrome b559 (Cyt b559). This result provided the evidence for the existence of the aggregation of the D1 protein in vivo. The maximal level of D1/Cyt b559 aggregate occurred at 1000 μmol photons·m?2·s?1 but drastically decreased with further increasing light intensity. Im- munoblot analysis with phosphothreonine (Thr (P)) antibody indicated that D1/CP43 and D1/Cyt b559 aggregates contained the phosphorylated protein(s). In vitro dephosphorylation experiment also showed that D1/Cyt b559 and D1/CP43 aggregates lost the immunoreactivity with Thr (P) antibody after the phosphatase treatment of the membranes from high-light-illuminated leaves. Our results demon- strated that strong illumination of spinach leaves in- duced cross-linked products of D1 protein with its nearby polypeptides of PS Ⅱ , some of which con- tained the phosphorylated D1 protein.