Identification of genes whose expression is upregulated in lung adenocarcinoma cells in comparison with type II alveolar cells and bronchiolar epithelial cells in vivo

To identify genes whose expression is upregulated in lung adenocarcinoma (AdC) cells in comparison with noncancerous peripheral lung epithelial cells, type II alveolar cells and bronchiolar epithelial cells, as well as AdC cells, were isolated by laser capture microdissection, and subjected to cDNA microarray analysis of 637 human cancer-related genes. Each of the component cells was obtained from several different individuals and analysed independently. As a comparison, two lung AdC cell lines and two primarily cultured normal lung epithelial cell lines were also subjected to cDNA microarray analysis. Four genes, TOP2A, MMP15, MX2 and KOC1, were commonly upregulated in microdissected AdC cells in comparison with microdissected epithelial cells. Hierarchical clustering analysis revealed that differences in gene-expression profiles were more evident between cultured and uncultured cells than between cancerous and noncancerous cells. To further identify the common molecular targets of AdC cells in vivo, quantitative real-time RT-PCR was performed against the four genes upregulated by cDNA microarray analysis. The TOP2A, MMP15, MX2 and KOC1 genes were overexpressed in 10/10 (100%), 8/10 (80%), 5/10 (50%) and 3/10 (30%) microdissected AdC cell samples, respectively, in comparison with any of nine independently microdissected noncancerous epithelial cell samples. The TOP2A gene was commonly overexpressed in lung AdC cells, as previously reported. In addition, the MMP15 and MX2 genes were identified, for the first time, as being commonly overexpressed in lung AdC cells. These results strongly indicate that the MMP15 and MX2 genes could be novel markers for molecular diagnosis and therapy of lung AdC.     

Effect of exogenous MSH6 and POLD1 expression on the mutation rate of the HPRT locus in a human colon cancer cell line with mutator phenotype, DLD-1

The DLD-1 human colon cancer cell line displays an elevated spontaneous mutation rate. Since DLD-1 carries frameshift mutations in both alleles of the MSH6 gene and missense mutations in the POLD1 gene, either or both of these mutations were suggested to be involved in this mutator phenotype. Therefore, we examined the effect of exogenous wild-type MSH6 and POLD1 expression on the spontaneous mutation rate at the HPRT locus in DLD-1 cells. POLD1 genotypes were first determined, since four POLD1 missense mutations were previously reported in DLD-1 cells. Sequencing analyses on the genomic DNA and cDNA of the POLD1 gene revealed that DLD-1 cells are a mixture of two distinct sublines with regard to POLD1 genotypes. Moreover, the wild-type POLD1 allele was not present in either of the two DLD-1 sublines. We next established MSH6- and POLD1-transfected DLD-1 clones from both sublines, respectively. The two DLD-1 sublines exhibited HPRT mutation rates of 4.8 x 10(-6) and 5.4 x 10(-6) mutations/cell/generation. The mutation rates were more than 4-fold decreased in both of the MSH6-transfected DLD-1 clones examined, while they were not significantly decreased in three of four POLD1-transfected DLD-1 clones. Thus, it was indicated that mutations in the MSH6 gene, and not in the POLD1 gene, are primarily responsible for the elevated mutation rates in DLD-1 cells.     

Angiogenesis in colon hyperplastic polyp

Despite the significance of tumor angiogenesis and the extensive knowledge on the molecular basis of blood vessel formation in carcinoma of colorectum, no data exist in hyperplastic polyp. This prompted us to examine angiogenesis in hyperplastic polyp. Eleven small hyperplastic polyps, 13 large hyperplastic polyps and their adjacent normal mucosas were included in this study. Angiogenesis was assessed by immunohistochemistry using monoclonal antibody against CD34. Angiogenic factor, thymidine phosphorylase was also examined by immunohistochemistry. Intra-tumoral microvessel density (IMD) in large hyperplastic polyp was significantly higher than that in small hyperplastic polyp (P<0.01) and that in normal mucosa (P<0.01). DAD in small hyperplastic polyp was also significantly higher than that in normal mucosa (P<0.01). Expression of dThdPase was almost observed in stromal cells in normal, small and large hyperplastic polyp. In addition, the proportion of the stromal cells expressing dThdPase in large hyperplastic polyp was significantly higher than that in small hyperplastic polyp and normal tissue (P<0.01, respectively). The proportion of the stromal cells expressing dThdPase in small hyperplastic polyp was significantly higher than that in normal tissue (P<0.01). The present study provides that angiogenesis may have an important role(s) in the development of hyperplastic polyp and dThdPase in stromal cells may support angiogenesis in hyperplastic polyp. Anti-angiogenic therapy might be available for suppression of hyperplastic polyp.     

Effect of halothane on neuronal excitation in the superficial dorsal horn of rat spinal cord slices: evidence for a presynaptic action

The action of the volatile anaesthetic halothane on optically recorded neuronal excitation in juvenile rat spinal cord slices was investigated. Prolonged neuronal excitation lasting approximately 100 ms was evoked in the superficial dorsal horn after single-pulse dorsal root stimulation that activated both A- and C-fibres. Halothane depressed the neuronal excitation in a concentration-dependent manner (IC(50) 0.21 mm, I(max) 28%). In Ca(2+)-free solution, dorsal root stimulation induced excitation with a short duration of several tens of milliseconds, in which the excitation of the postsynaptic component was largely eliminated. Under these conditions, halothane also depressed the excitation concentration-dependently (IC(50) 0.46 mm, I(max) 60%). Most of the suppression occurred within 5 min of halothane application, and the effect of halothane was fully reversible upon washout of the anaesthetic. Application of bicuculline and strychnine or picrotoxin, or reduction of extracellular Cl(-) concentration ([Cl(-)](o)), had no effect on halothane inhibition. Applications of K(+) channel blockers tetraethyl ammonium, 4-aminopyridine, Cs(+) or Ba(2+) either had no effect or augmented the inhibitory effect of halothane. On the other hand, the degree of inhibition by halothane was found to be dependent on [K(+)](o); the higher [K(+)](o), the larger the depression. In addition, decreases in [Na+]o and [Mg(2+)](o) reduced the excitation similar to that of halothane treatment, and the degree of halothane inhibition became larger with lower [Mg(2+)](o). These results lead to a hypothesis that halothane suppresses the excitation of presynaptic elements by inhibiting presynaptic Na(+) channels by shifting the steady-state inactivation curve in the hyperpolarizing direction.     

Recent indications and methods of surgery for inflammatory bowel disease

Inflammatory bowel disease, notably ulcerative colitis (UC) or Crohn disease (CD), is basically benign, but sometimes develops into serious or fatal cancer. While the primary therapies are medical, such as pharmacotherapy and dietetic modification, intractable, serious, and cancerous cases can require surgical intervention. Surgery represents only one of the treatment options, but prediction of whether UC and CD are likely to progress to serious conditions and determination of when to undertake surgery is essential. Various surgical procedures have been developed over time, and the postoperative results are now generally good. Regarding laparoscopic surgeries, relatively few cases have been accumulated, and addressing the indications and limitations is premature at this point. However, this procedure is likely to fulfill a central role in surgical treatment strategies and represent a major benefit to patients. This paper discusses surgical treatment indications and methods for UC and CD, and explains the practical aspects of laparoscopic surgery, which has made remarkable progress in recent years, for such cases.     

No mutations of the Smad2 gene in human sporadic gastric carcinomas

BACKGROUND: The majority of cancer cells escape from TGF-beta-mediated growth control. However, the mechanism of resistance to the growth inhibitory effects by TGF-beta is not clear. TGF-beta signaling is initiated when the type I receptor phosphorylates the SMAD proteins, Smad2 and Smad3. Recently, mutations of Smad2 have been detected in human colon and lung cancers. Mutation of coding sequences of Smad2 in gastric carcinomas has not yet been elucidated adequately. METHODS: PCR-SSCP analysis of the entire coding region of Smad2 in 35 human sporadic gastric cancers and eight gastric cancer cell lines was performed using 11 sets of intron-based primers. RESULTS: No mutations of Smad2 were detected in any tumor or cell line. CONCLUSIONS: The results suggest that mutation of Smad2 does not play a key role in human stomach carcinogenesis.     

Infrequent mutations of the transforming growth factor beta-type II receptor gene at chromosome 3p22 in human lung cancers with chromosome 3p deletions

Mutations in the transforming growth factor beta-type II receptor (TGFbeta RII) gene have been detected in several types of human cancers that represent the phenotype of genomic instability. The TGFbeta RII gene has been mapped to chromosome 3p, on which loss of heterozygosity (LOH) was frequently detected in both small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC). To investigate whether the TGFbeta RII gene on 3p22 is inactivated in lung cancers, we examined 35 sporadic lung cancers (15 SCLC and 20 NSCLC) with LOH on 3p for mutations of the TGFbeta RII gene. We previously produced eight intron based primer pairs for mutational analysis of the entire coding region of the TGFbeta RII gene. Using these primers, we screened for mutations of the TGFbeta RII gene by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis. A mutation was detected in a case of SCLC: one base insertion in the polyadenine tract of exon 3. This tumor showed the replication error (RER) phenotype. There were no mutations in exons 1, 2, 4, 5, 6 and 7. These results indicate that the polyadenine tract is a mutational hot spot in the TGFbeta RII gene in RER positive tumors, and that TGFbeta RII mutations occur rarely in lung cancers with LOH on chromosome 3p.