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Since an attenuated response to stress is a characteristic of senescence, a cellular senescence model was used to examine the mechanism of resistance against oxidative stress using human diploid fibroblasts (HDF). With increasing passage, the HDF showed increased production of reactive oxygen species (ROS). Late passage HDF were resistant to the lethal effects of oxidative stress, showing less cleavage of pro-caspase-3 and PARP than those of early ones. Since heat shock proteins (Hsps) are not only cytoprotective but also interfere with the apoptotic cascade, the expression patterns of Hsps during cellular senescence were next examined. Oxidative stress induced a decrease in the mitochondrial Hsp60 levels with a concomitant increase in the cytosolic Hsp60 levels in the early passage HDF, but not in late ones. To show that the resistance to oxidative stress is a specific effect of Hsp60, the levels of Hsp60 were knocked down by siRNA. As expected the Hsp60 knock-down cells were more resistant to oxidative stress. These findings show that Hsp60 is a key player in the resistance mechanism against oxidative stress and aging.

Paired box gene 9 (Pax9) and c-myb are transcription factors that regulate the expression of the genes involved in mediating cell proliferation, resistance to apoptosis, and migration. However, the function of Pax9 in oral squamous cell carcinoma (OSCC) is virtually unknown. This study examined the anti-apoptotic roles of Pax9 and c-myb, and clarified interaction between the two genes in KB cells. Inhibition of Pax9 caused the induction of apoptosis with enhanced cleavage of caspase-3 and PARP, accelerated Bax, and reduced Bcl-2 expression. Transducing c-myb cells with adenovirus c-myb (Ad/c-myb) were induced cell growth and inhibited apoptosis, but dominant-negative myb cells (Ad/DN-myb) were not affected. Pax9 was upregulated in the Ad/c-myb cells with simultaneous decrease in the Ad/DN-myb infection. However, c-myb remained unaffected in the Pax9 small interfering RNA (siRNA) transfected cells. Moreover, the Pax9 siRNA transfected cells and Ad/DN-myb infected cells were able to arrest the cell cycle at the G(0) phase. This suggests that Pax9 and c-myb expression in KB cells is essential for cell growth, and survival is enhanced by c-myb. Disrupting the function of c-myb and Pax9 could be a potential target for cancer treatment.

The stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) pathway is a well-known senescence-related stress activated protein kinase. Multiple environmental stresses induce programmed cell death, such as apoptosis. Normal human diploid fibroblast (HDF) cells have a limited life span in vitro, halting proliferation after a fixed number of cell divisions. Aged passage HDF showed resistance to oxidative stress involving heat shock proteins (Hsp60) through a mechanism involving the translocation of Hsp60 from the mitochondria to the cytosol. The present study showed that the translocation of Hsp60 from the mitochondria to the cytosol followed by high levels of p-SAPK/JNK activation as a result of oxidative stress was observed in the young cells only. The inhibition of SAPK/JNK activation by SP600125 under oxidative stress almost completely blocked the translocation of Hsp60 in both young and aged cells. This suggests that aged HDF cells are resistant to oxidative stress by blocking the translocation of Hsp60 from the mitochondria to the cytosol followed by SAPK/JNK inhibition. Overall, the mechanism of resistance by oxidative stress in aged cells is induced by blocked of the translocation of Hsp60 followed by SAPK/JNK inactivation.