Expression of epithelial-mesenchymal transition markers at the invasive front of oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is one of the most common malignances. In epithelial-mesenchymal transition (EMT), epithelial cells switch to mesenchymal-like cells exhibiting high mobility. This migratory phenotype is significant during tumor invasion and metastasis. Objective : The aim of this study is to evaluate the expression of the EMT markers E-cadherin, N-cadherin and vimentin in OSCC. Material and Methods : Immunohistochemical detection of E-cadherin, N-cadherin and vimentin was performed on 20 OSCC samples. Differences in the expression of each protein at the invasive front (IF) and in the central/superficial areas (CSA) of the tumor were assessed. Differences in the expression of each protein at the IF of both histologically high- and low-invasive OSCCs were evaluated. Associations among expression of proteins at the IF were assessed. Correlations between the expression levels of each protein at the IF and the tumor stage and clinical nodal status were also evaluated. Results : Reduced expression of E-cadherin was detected in 15 samples (75%). E-cadherin expression was reduced at the IF when compared to the CSA and in high-invasive tumors when compared to low-invasive tumors. All samples were negative for N-cadherin, even though one sample showed an inconspicuous expression. Positive expression of vimentin was observed in 6 samples (30%). Nevertheless, there was no difference in vimentin expression between the IF and the CSA regions or between the low- and high-invasive tumors. Furthermore, no association was observed among protein expression levels at the IF. Finally, no correlations were observed between each protein’s expression levels and tumor stage or clinical nodal status. Conclusions : Reduced E-cadherin expression at the IF and its association with histological invasiveness suggest that this protein is a noteworthy EMT marker in OSCC. Although vimentin was also detected as an EMT marker, its expression was ...

Identification and characterization of lipids from endosomes purified by electromagnetic chromatography.

Traditional separation techniques do not yield endolysosomes of sufficient purity to permit detailed biochemical characterization of this important class of intracellular vesicles. Here, we have used a magnetic chromatography technique to isolate the endosomes from rat peritoneal macrophages and studied their lipid composition. Electromagnetic isolation works by retention of colloidal iron containing vesicles on magnetic column. The data suggested that both early and late endosomes were rich in cholesterol, whereas sphingomyelin (SM) and specific phospholipids like phosphatidylcholine. phosphatidylethanolamine, phosphatidylglycerol and phosphatidylserine are enriched in the late compartments. Our results also indicated that the purified fractions are enriched in raft lipids like SM, but not in cholesterol. The endosomal purification method described here yields pure endosomes with little or no contamination from mitochondria and hence could be used for further biochemical and marker analysis, giving insight into mechanisms of endocytic traffic.

Heat shock protein 70 alters the endosome-lysosomal localization of huntingtin

Huntington's disease is caused by CAG trinucleotide expansions in the gene encoding huntingtin. N- terminal fragments of huntingtin with polyglutamine produce aggregates in the endosome-lysosomal system, where proteolytic fragments of huntingtin is generated. Heat shock protein 70 (HSP70) prevents the formation of protein aggregates, but the effect of HSP70 on the huntingtin in the endosome-lysosomal system is unknown. This study was to determine whether HSP70 alters the distribution of huntingtin in endosome-lysosomal system. HSP70 expressing stable cells (NIH/3T3 or cerebral hybrid cell line A1) were generated, and mutant [(CAG)100] huntingtin was transiently overexpressed. Analysis of subcellular distribution by immnuocytochemistry or proteolysis cleavage by Western blotting was performed. 18 CAG repeat wild type [WT; (CAG)18] huntingtin was used as a control. Cells with huntingtin showed patterns of endosome- lysosomal accumulation, from a 'dispersed vacuole (DV)' type into a coalescent 'perinuclear vacuole (PV)' type over time. In WT huntingtin, HSP70 increased the cells with the PV types that enhanced the proteolytic cleavage of huntingtin. However, HSP70 reduced cells of the DV and PV types expressing mutant huntingtin, that result in less proteolysis than that of control. In addition, intranuclear inclusions were formed only in mutant cells, which was not affected by HSP70. These results suggest that HSP70 alters the distribution of huntingtin in the endosome-lysosomal system, and that this contributes to huntingtin proteolysis.