Allelotypic characteristics of thorotrast-induced intrahepatic cholangiocarcinoma: comparison to liver cancers not associated with thorotrast

To elucidate the genetic alterations that are specific to Thorotrast-induced liver cancers and their possible roles in tumorigenesis, we analyzed loss of heterozygosity (LOH) at 37 loci. Our previous study of liver cancers that were not associated with Thorotrast found LOH at 9 of these loci to be characteristic of intrahepatic cholangiocarcinoma (ICC), at 19 to be characteristic of hepatocellular carcinoma (HCC), and at 9 to be common to both ICC and HCC. LOH analysis was also performed in tissues of cholangiolocellular carcinoma, which is thought to originate from a common stem cell progenitor of hepatocytes and bile duct epithelial cells. We found frequent LOH at D4S1538, D16S2624 and D17S1303 to be common to all the subtypes of liver cancers, independent of the specific carcinogenic agent. In contrast, LOH at D4S1652 generally was not observed in Thorotrast-induced ICC. LOH analysis revealed that Thorotrast-induced ICC shares some LOH features with both ICC and HCC that were not induced by Thorotrast; however, it is more similar to ICC than to HCC in terms of genetic changes. This study could narrow down the crucial chromosomal loci whose deletions are relevant to hepatobiliary carcinogenesis irrespective of the carcinogenic agent. The study of LOH at loci other the those crucial ones may help us understand how the phenotype of liver cancers is determined.     

Deformation of lipid droplets in fixed samples

Nile red, Sudan III, and oil red O have been used to stain lipid droplets (LDs) for fluorescence microscopy. We noticed that LDs labeled by Nile red are different in appearance from those stained by the latter two dyes. To understand the cause of the difference, we used sequential labeling procedures (first LD stain-photography-quenching-second LD stain-photography), and examined the effect of several factors. Immunofluorescence labeling for adipose differentiation-related protein (ADRP), an LD marker, was also observed comparatively with the lipid stains. As a result, we found that ethanol and isopropanol used for Sudan III and oil red O staining, respectively, and glycerol used for mounting, cause fusion of adjacent LDs even in glutaraldehyde-fixed samples. By the same treatment, immunofluorescence labeling for ADRP was dislocated to the rim of large LDs that were formed as a result of the artifactual fusion. The result indicates that the LD structure can be better observed with Nile red than with Sudan III or oil red O.     

Afrotherian phylogeny as inferred from complete mitochondrial genomes

Afrotheria is a huge assemblage of various mammals encompassing six orders that were once classified as distantly related groups. This superordinal relationship may have resulted from the break-up of Gondowanaland followed by the isolation of the African continent between 105 and 40 million years ago. Although the monophyly of Afrotheria is well supported by recent molecular studies, the interrelationships within afrotherian mammals remain unclarified. In this study, we determined the sequence of the complete mitochondrial genomes of hyrax, golden mole, and elephant shrew. These sequences were compared with those of other eutherians to analyze the phylogenetic relationships among afrotherians and, in particular, those among paenungulates. Our mitochondrial genome analysis supports the monophyly of Tethytheria.     

α-Actinin-4 Enhances Colorectal Cancer Cell Invasion by Suppressing Focal Adhesion Maturation

α-Actinins (ACTNs) are known to crosslink actin filaments at focal adhesions in migrating cells. Among the four isoforms of mammalian ACTNs, ACTN1 and ACTN4 are ubiquitously expressed. Recently, ACTN4 was reported to enhance cancer cell motility, invasion, and metastasis. However, the mechanism by which ACTN4 drives these malignant phenotypes remains unclear. Here, we show that ACTN4, but not ACTN1, induces the formation of immature focal adhesions in DLD-1 cells, leading to the rapid turnover of focal adhesions. Interestingly, zyxin (ZYX) assembly to focal adhesions was markedly decreased in ACTN4-expressing DLD-1 cells, while the recruitment of paxillin (PAX) occurred normally. On the other hand, in ACTN1-expressing DLD-1 cells, PAX and ZYX were normally recruited to focal adhesions, suggesting that ACTN4 specifically impairs focal adhesion maturation by inhibiting the recruitment of ZYX to focal complexes. Using purified recombinant proteins, we found that ZYX binding to ACTN4 was defective under conditions where ZYX binding to ACTN1 was observed. Furthermore, Matrigel invasion of SW480 cells that express high endogenous levels of ACTN4 protein was inhibited by ectopic expression of ACTN1. Altogether, our results suggest that ZYX defective binding to ACTN4, which occupies focal adhesions instead of ACTN1, induces the formation of immature focal adhesions, resulting in the enhancement of cell motility and invasion.     

Lysophosphatidylcholine Acyltransferase1 Overexpression Promotes Oral Squamous Cell Carcinoma Progression via Enhanced Biosynthesis of Platelet-Activating Factor

Background

The relevance of lysophosphatidylcholine acyltransferase1 (LPCAT1), a cytosolic enzyme in the remodeling pathway of phosphatidylcholine metabolism, in oral squamous cell carcinoma (OSCC) is unknown. We investigated LPCAT1 expression and its functional mechanism in OSCCs.

Methods

We analyzed LPCAT1 mRNA and protein expression levels in OSCC-derived cell lines. Immunohistochemistry was performed to identify correlations between LPCAT1 expression levels and primary OSCCs clinicopathological status. We established LPCAT1 knockdown models of the OSCC-derived cell lines (SAS, Ca9-22) for functional analysis and examined the association between LPCAT1 expression and the platelet-activating factor (PAF) concentration and PAF-receptor (PAFR) expression.

Results

LPCAT1 mRNA and protein were up-regulated significantly (p<0.05) in OSCC-derived cell lines compared with human normal oral keratinocytes. Immunohistochemistry showed significantly (p<0.05) elevated LPCAT1 expression in primary OSCCs compared with normal counterparts and a strong correlation between LPCAT1-positive OSCCs and tumoral size and regional lymph node metastasis. In LPCAT1 knockdown cells, cellular proliferation and invasiveness decreased significantly (p<0.05); cellular migration was inhibited compared with control cells. Down-regulation of LPCAT1 resulted in a decreased intercellular PAF concentration and PAFR expression.

Conclusion

LPCAT1 was overexpressed in OSCCs and correlated with cellular invasiveness and migration. LPCAT1 may contribute to tumoral growth and metastasis in oral cancer.