Faculty Bibliography

Ceramide, ceramide-1-phosphate (C1P) sphingosine (SPH) and sphingosine-1-phosphate (S1P) effects on proliferation and extracellular-signal regulated kinases, ERKs (also known as MAPKs), activation were investigated in human and rat osteoblastic cells. MAPK activation was sphingolipid-specific in cells from both species. In human osteoblastic cells, S1P and C1P markedly stimulated ERK2 phosphorylation with a slight increase in phosphorylation of ERK1. SPH nor ceramide induced phosphorylation of either ERK isoform. In rat osteoblastic cells, SIP, ceramide and SPH stimulated phosphorylation of both isoforms. C1P did not induce phosphorylation of ERK1 but produced a mild increase in phosphorylation of ERK2. In human cells, only S1P significantly (P<0.05) increased osteoblastic cell proliferation, while in the rat cells all four sphingolipids significantly (P<0.05) induced proliferation. The calcium channel blocker verapamil blocked (P<0.05) these effects in both cell types. The MAPK inhibitor, PD98059, inhibited (P<0.05) the mitogenic effect of SIP in human cells. In rat cells, PD98059 effects were less substantial but significant for S1P and C1P. This study demonstrates that sphingolipids are mitogens for both human and rat osteoblastic cells with the MAPK pathway and calcium mediating in part these effects in a species specific manner.

The study of signal transduction pathways for mechanisms of apoptosis and proliferation has significantly advanced our understanding of human cancer, subsequently leading to more effective treatments. Discoveries of growth factors and oncogenes, especially those that function through phosphorylation on tyrosine residues, have greatly benefited our appreciation of the biology of cancer. The regulation of proliferation and apoptosis through phosphorylation via tyrosine kinases and phosphatases is discussed, as well as the contributions of other systems, such as serine and threonine kinases and phosphatases. Receptors with seven-transmembrane domains, steroid hormones, genes, and 'death domains' will also be discussed. This review attempts to compare the regulation of the growth of normal tissues and cancers with an effort to highlight the current knowledge of these factors in the growth regulation of oral/oropharyngeal cancers. Despite the strides made in our understanding of growth regulation in human cancers, the study of oral/oropharyngeal cancer specifically lags behind. More research must be done to further our understanding of oral cancer biology, if we are to develop better, more effective treatment protocols

The ascending Method of Limits, used for the determination of pressure pain thresholds (PPT), is not a psychophysically robust method. The present study sought to determine if the examiner's expectancy, based on whether the measurement site was clinically 'painful' or 'non-painful', would bias the obtained PPT values. Twenty-eight patients with facial or temporal area pain served as subjects, and in each subject, a pain site and a control site were identified and marked. According to a randomization schedule, the pain and control sites were correctly marked in half of the subjects and were mis-labeled in the other half, thereby controlling the examiner's knowledge of a site and thus the examiner's expectancy of what the PPT should be. Two examiners, shown to be reliable with each other in both pre-clinical and post-clinical reliability studies, were blind to the true purpose of the study and to the marking procedures. Each examiner made one PPT measurement at each marked site in a counterbalanced measurement order. Manipulating the examiner's prior knowledge of the measurement site's characteristics significantly lowered the obtained PPT values for control sites but did not significantly alter the PPT at the clinically painful sites. Nevertheless, the pain sites still had significantly lower PPTs than did control sites. We conclude that: (i) PPTs at pain sites are robust to a major source of measurement bias associated with the ascending Method of Limits; (ii) measurement order and knowledge of measurement site characteristics can influence obtained PPT; and (iii) the common protocol in which the examiner monitors the amount of pressure during PPT measurement in order to control the force application rate may serve as a mechanism that can bias the obtained values.

Our goal was to examine the effects of ethanol on cell death using rat pheochromocytoma (PC12) cells as a neuronal model. Withdrawal of serum for 24 hr increased the number of PC12 cells labeled with ethidium homodimer indicating an increase in cell death. Inclusion of 50 mM ethanol during the serum deprivation further increased the amount of ethidium fluorescence by 39%. Although reducing the serum concentration from the usual 15 to 4% did not alter cellular viability, a significant increase in the amount of ethidium fluorescence was observed in PC12 cells incubated for 24 hr in the presence of 4% serum and 150 mM ethanol. No change in viability was observed in cells exposed to either 150 mM ethanol in the presence of 15% serum or 50 mM ethanol in the presence of 4% serum. Inclusion of ethanol during serum deprivation increased the amount of soluble DNA found in the 15,000 x g supernatant. Similarly, using the terminal deoxynucleotidyl transferase dUTP nick-end labeling method to visualize DNA fragmentation in situ, ethanol caused a 213% increase in the number of PC12 cells labeled during serum withdrawal. Agarose gel electrophoresis of the DNA isolated from cells maintained in the absence of serum yielded the classical DNA laddering pattern of 180 to 200 bp fragments suggestive of apoptosis. Ethanol caused a concentration-dependent increase in the amount of DNA laddering in cells deprived of serum. Furthermore, ethanol significantly potentiated the DNA laddering of cells maintained in 4% serum. In contrast to the ethanol-induced increase in cell death when serum factors were reduced or withdrawn, 150 mM ethanol lowered by 34% the number of ethidium-labeled PC12 cells observed after a 30-min exposure to 2 mM H2O2. Similarly, agarose gel electrophoresis of the DNA from H2O2-treated cells did not display DNA laddering. This study demonstrates that: 1) ethanol antagonizes the trophic action of serum factors; 2) pharmacologically relevant ethanol concentrations significantly potentiate apoptosis after serum withdrawal and 3) this enhancement appears specific for apoptosis

Protein phosphorylation/dephosphorylation on tyrosine residues are regulated by tyrosine kinases/phosphatases. The tyrosine phosphatase SH-PTP2 (PTP1D, PTP2C) interacts physically through its SH2 domain with phosphorylated epidermal growth factor receptor (EGFR). In KB cells, an oral epidermoid carcinoma, high epidermal growth factor (EGF) concentrations (10-9, 10-8 and 10-7 M) downregulate its receptor for the duration of the incubation with EGF. Thus, it was hypothesized that in KB cells, SH-PTP2 expression would also be downregulated by high EGF concentrations. This hypothesis was investigated by incubating the KB cells with increasing concentrations of EGF (0, 10-11, 10-10, 10-9, 10-8, 10-7 M) and by evaluating the expression of SH-PTP2 protein under these conditions. This study showed that EGF 10-7 and 10-8 M significantly decreased SH-PTP2 protein expression compared to controls. EGF 10-10 and 10-11 M did not change the expression of SH-PTP2 protein. We conclude that high EGF concentrations downregulate the expression of SH-PTP2 protein