Effect of anaesthesia maintained with sevoflurane and propofol on surgical site infection after elective open gastrointestinal surgery.

Perioperative increase in oxidative activity in surgical patients reportedly prevents postoperative surgical site infection (SSI). Several clinical studies have shown that oxidative activity under sevoflurane anaesthesia was higher than that under propofol anaesthesia. Therefore, we hypothesised that sevoflurane anaesthesia would discourage SSI compared with propofol anaesthesia. To examine the effect of anaesthesia maintained with sevoflurane and propofol on SSI, a total of 265 consecutive adult patients, with American Society of Anesthesiologists physical status 1-3, who underwent elective open gastrointestinal surgery under general anaesthesia, were surveyed for SSI between January 2007 and December 2008. Sevoflurane or propofol was selected to maintain anaesthesia in 95 and 170 patients, respectively. A propensity score was used for pairwise matching of these patients to avoid selection biases between the two methods of anaesthesia. Propensity matching yielded 84 pairs of patients. We compared standardised infection ratios (SIRs), i.e. the quotient of the number of SSI cases observed and the number of SSI cases expected, calculated using data from the National Nosocomial Infection Surveillance, between sevoflurane and propofol anaesthesia. After propensity matching, SIR after sevoflurane anaesthesia was 1.89 [95% confidence interval (CI): 1.46-2.32], which was significantly lower than after propofol anaesthesia (4.78; 95% CI: 4.30-5.27) (P=0.02). This study suggests that sevoflurane tends to suppress SSI after elective open gastrointestinal surgery compared with propofol.

Somatic alterations of the DPC4 and Madr2 genes in colorectal cancers and relationship to metastasis.

The DPC4 and Madr2 genes are located at 18q21, and the LOH on chromosome 18q21 has been shown to occur frequently in colorectal cancers. To investigate the role of these genes in advanced colorectal cancers, we analyzed 29 colorectal specimens for alterations in the DPC4 and Madr2 genes. Twelve (63.2%) of 19 informative primary colorectal cancers showed allelic loss of chromosome 18q21.3 marker. An alteration of the DPC4 gene sequence was identified in 6 (20.7%) of 29 colorectal carcinomas, and the distinct Madr2 gene mobility shifts were present in 3 (10.3%) cancers. Somatic mutations were identified in these tumors by sequencing analysis. DPC4 gene alterations of 4 cases were detected in Mad homology 2 domains. There was no significant correlation between the somatic alteration of Madr2 and clinicopathological findings. However, the frequency of DPC4 mutation was significantly higher in tumors associated with liver metastasis than in those without such metastasis. Our findings suggest that somatic alteration of the DPC4 gene may play a role in tumorigenesis and liver metastasis of human colorectal cancers.

Expression of variant CD44 containing variant exon v8-10 in gallbladder cancer.

We studied the Cd44v8-10 expression in gallbladder cancer immunohistochemically. Eighteen of 37 gallbladder cancer tissues expressed CD44v8-10. There were significant correlations between CD44v8-10 immunoreactivity and perineural invasion, venous invasion, and lymph node metastasis. Patients with CD44v8-10-positive tumors showed poor prognoses, whereas those with CD44v8-10-negative tumors had favorable prognoses. A multivariate analysis using the Cox regression model showed the immunoreactivity of CD44v8-10 to be an independent prognostic indicator of gallbladder cancer. The results suggest that CD44v8-10 expression may be a biologic marker of prognostic significance in gallbladder cancer.

An EGF receptor/Ral-GTPase signaling cascade regulates c-Src activity and substrate specificity.

c-Src is a membrane-associated tyrosine kinase that can be activated by many types of extracellular signals, and can regulate the function of a variety of cellular protein substrates. We demonstrate that epidermal growth factor (EGF) and beta-adrenergic receptors activate c-Src by different mechanisms leading to the phosphorylation of distinct sets of c-Src substrates. In particular, we found that EGF receptors, but not beta(2)-adrenergic receptors, activated c-Src by a Ral-GTPase-dependent mechanism. Also, c-Src activated by EGF treatment or expression of constitutively activated Ral-GTPase led to tyrosine phosphorylation of Stat3 and cortactin, but not Shc or subsequent Erk activation. In contrast, c-Src activated by isoproterenol led to tyrosine phosphorylation of Shc and subsequent Erk activation, but not tyrosine phosphorylation of cortactin or Stat3. These results identify a role for Ral-GTPases in the activation of c-Src by EGF receptors and the coupling of EGF to transcription through Stat3 and the actin cytoskeleton through cortactin. They also show that c-Src kinase activity can be used differently by individual extracellular stimuli, possibly contributing to their ability to generate unique cellular responses.

Expression of DCC protein in colorectal tumors and its relationship to tumor progression and metastasis.

A loss of heterozygosity (LOH) at the DCC gene locus was detected in colorectal tumors, and this LOH might be related to metastasis. The aim of this study was to determine DCC protein expression in colorectal cancer and to evaluate its prognostic value. Allelic loss of the DCC locus was observed in 16 of the 23 patients (66.7%). In all 16 patients with LOH, DCC expression was decreased in the cancer tissue compared with the adjacent normal mucosa. All 23 colorectal tumors had decreased expression of this protein relative to the adjacent normal colonic mucosa in Western blot analysis. The levels of DCC protein were significantly lower in cancer tissues than in adenoma tissues. Decreased DCC protein expression was also observed by immunohistochemistry in the colorectal cancer cases. There were significant correlations between DCC protein expression and histologic type, venous invasion, and hematogenous metastasis. Patients with DCC-protein-negative tumors had a greater relative risk of recurrence compared with those whose tumors were DCC protein-positive. The 5-year survival rate was 91.0% in patients with DCC-protein-positive tumors, and 58.8% in those with DCC-protein-negative tumors; these differences between the two groups of patients were significant (p < 0.01). In multivariate analysis using the Cox regression model, DCC protein expression emerged as an independent prognostic indicator. These findings suggested that a decrease in DCC expression may have an important role in the progression of colorectal cancers and may be a biologic marker of prognostic significance.

Ral-specific guanine nucleotide exchange factor activity opposes other Ras effectors in PC12 cells by inhibiting neurite outgrowth.

Ras proteins can activate at least three classes of downstream target proteins: Raf kinases, phosphatidylinositol-3 phosphate (PI3) kinase, and Ral-specific guanine nucleotide exchange factors (Ral-GEFs). In NIH 3T3 cells, activated Ral-GEFs contribute to Ras-induced cell proliferation and oncogenic transformation by complementing the activities of Raf and PI3 kinases. In PC12 cells, activated Raf and PI3 kinases mediate Ras-induced cell cycle arrest and differentiation into a neuronal phenotype. Here, we show that in PC12 cells, Ral-GEF activity acts opposite to other Ras effectors. Elevation of Ral-GEF activity induced by transfection of a mutant Ras protein that preferentially activates Ral-GEFs, or by transfection of the catalytic domain of the Ral-GEF Rgr, suppressed cell cycle arrest and neurite outgrowth induced by nerve growth factor (NGF) treatment. In addition, Rgr reduced neurite outgrowth induced by a mutant Ras protein that preferentially activates Raf kinases. Furthermore, inhibition of Ral-GEF activity by expression of a dominant negative Ral mutant accelerated cell cycle arrest and enhanced neurite outgrowth in response to NGF treatment. Ral-GEF activity may function, at least in part, through inhibition of the Rho family GTPases, CDC42 and Rac. In contrast to Ras, which was activated for hours by NGF treatment, Ral was activated for only approximately 20 min. These findings suggest that one function of Ral-GEF signaling induced by NGF is to delay the onset of cell cycle arrest and neurite outgrowth induced by other Ras effectors. They also demonstrate that Ras has the potential to promote both antidifferentiation and prodifferentiation signaling pathways through activation of distinct effector proteins. Thus, in some cell types the ratio of activities among Ras effectors and their temporal regulation may be important determinants for cell fate decisions between proliferation and differentiation.

Clinical significance of combined immunohistochemical detection of CD44v and sialyl LeX expression for colorectal cancer patients undergoing curative resection.

To evaluate their prognostic value, the expressions of CD44v and sialyl LeX (SLX) in colorectal cancers were studied immunohistochemically. Tissue specimens were reacted with monoclonal antibodies (mAb) CD44-1V and CSLEX-1. Of the 145 colorectal cancer patients undergoing curative resection, 59 (40.7%) were positive for mAb CD44-1V, and 40 (27.6%) were positive for mAb CSLEX-1. There was a significant correlation between the combined expression of SLX and CD44v8-10 and lymph node metastasis. The patients with tumors negative for CD44v8-10 and SLX had the most favorable prognoses. Conversely, the patients with tumors positive for both CD44v8-10 and SLX had a high recurrence rate and the poorest prognoses. In a multivariate analysis using the Cox regression model, the combined expression of SLX and CD44v8-10 emerged as an independent prognostic indicator. These results suggested that the combined expression of CD44v8-10 and SLX may be a biologic marker of prognostic significance.

Expression of nm23-H1 gene and Sialyl Lewis X antigen in breast cancer.

The nm23 gene has been proposed as a candidate tumor metastasis suppressor in some human cancers. Sialyl Lewis X (sLex) has been demonstrated to play an important role in the adhesion of human cancer cells to human vascular endothelium, inducing metastasis. Little information has been reported about the correlation between the expression of nm23 and sialylated carbohydrate antigens. In the present study, 102 surgically resected primary breast cancer tissues were sectioned and stained with antibody against nm23-H1 and sLex. Of the 102 cases, 39 (38.2%) cases with a reduced expression of nm23-H1 were observed, and the numbers of sLex-positive cases were 61 (59.8%), respectively. The reduced expression of nm23-H1 and the positive expression of sLex were significantly associated with lymph node involvement. Among the 100 patients who underwent curative surgery, the disease-free survival rate was significantly correlated to both the nm23-H1 and sLex expressions. No interrelated expressions were found between nm23-H1 and sLex. In multivariate analysis using Cox regression model, combination assay of nm23-H1 and sLex expression emerged as independent significant prognostic factors. These results suggest that nm23-H 1 gene and sLex may be involved in different steps of the metastatic process in human breast cancer, and immunohistochemical detection of the combination of sLex and nm23-H1 may be a biologic marker of prognostic significance.

Clinical significance of serum levels of CD44 variant exons 8-10 protein in colorectal cancer.

We examined serum levels of a CD44 splice variant that contained variant exons 8-10 (CD44v8-10) as a tumor marker in colorectal cancer patients. We performed enzyme-linked immunosorbent assays in 81 sera obtained from 71 colorectal cancer patients and 10 healthy controls. Serum CD44v8-10 levels were significantly higher in the colorectal cancer patients than in the healthy controls (0.209 +/- 0.098 versus 0.114 +/- 0.019 OD; P < 0.01). There was a close correlation between immunohistochemical expression and serum CD44v8-10 levels. Surgical resection of the tumors resulted in a reduction of serum CD44v8-10 levels. There was no significant correlation between serum CD44v8-10 level and serosal invasion or histologic type. However, a significant correlation was observed between serum CD44v8-10 level and lymphatic or venous invasion. In addition, serum CD44v8-10 levels were significantly higher in carcinomas associated with lymph node or liver metastasis than in those without metastasis. These findings suggest the usefulness of serum CD44v8-10 level in the prediction of colorectal cancer metastasis.

[Telomerase activity in colorectal carcinoma and its relationship to clinicopathological findings].

Telomerase is thought to be responsible for cell immortality. The telomerase activity in carcinomas has been remarked since 1995. We examined telomerase activity in colorectal carcinoma by TRAP (Telomeric repeat amplification protocol) assay, and investigated its relationship to clinicopathological findings. We could analyse telomerase activities in 33 cases (66%) of 50 colorectal carcinomas, whereas the activity was not found in all 13 cases of noncancerous colorectal mucosa. There was no relation between the telomerase activity and the clinicopathological findings or metastatic status. We confirmed telomerase activities in much of colorectal carcinomas in spite of their progression. The carcinoma cells might be immortal from their early stage of progression by means of telomerase activity.

Reduced expression of deleted colorectal carcinoma (DCC) protein in established colon cancers.

Using a bacterial fusion protein, a deleted colorectal carcinoma (DCC)-specific monoclonal antibody (MAb) 127-22 was established. Although MAb 127-22 reacted with almost all normal tissues, it did not react or only weakly reacted with many cancer cell lines, including colonic cancer lines, in flow cytometry. In Western immunoblots, the MAb reacted with a single 190-kDa molecule in a myeloma line Ara-10 extract. This component was scarcely detected in colonic cancer cell lines. Immunoblots of samples from 25 pairs of colonic cancers and adjacent normal tissues and from five adenoma tissues revealed that all normal colonic and adenoma tissues significantly expressed the DCC protein, whereas colonic cancer tissues showed poor expression. These results indicate not only deletion of and lowered mRNA expression of the DCC gene, but also marked reduction of DCC protein occurred in colonic cancer tissues. In addition, colonic cancer patients with liver metastasis expressed significantly lower levels of DCC than those without, suggesting the prognostic value of DCC expression.

The expression patterns of standard and variant CD44 molecules in normal uterine cervix and cervical cancer.

CD44 is a cell-surface glycoprotein postulated to play a role in tumor-cell metastasis. We have examined the expression of the standard CD44 (CD44s), and alternative spliced variants of CD44 containing variant exons v6, v9, and v1O (CD44v6, CD44v9, and CD44v10 respectively) in 9 samples of normal cervix, 6 samples of cervical intraepithelial neoplasia (CIN), and 11 samples with invasive cervical carcinomas. RT/PCR demonstrated the presence of CD44s in all samples of normal cervix and those with invasive carcinomas. CD44v6 was also found in all normal cervical samples and in 9 tissue samples of invasive carcinomas. The results also suggested that some tumor specimens had several higher molecular transcripts containing exon v6 compared to specimens of normal cervix. Immunohistochemistry detected the presence of CD44s and the absence of CD44v10 in both epithelial and stromal cells in all specimens. In contrast, CD44v6 and CD44v9 were stained positive in epithelial cells but were absent in stromal cells. The intensity of CD44v6 and CD44v9 staining was strongest in normal cervical epithelium followed by CIN, invasive squamous cell carcinoma, and adenocarcinoma. In the malignant samples, heterogeneity in staining intensity among different clusters of tumor cells was observed. Furthermore, poorly differentiated and undifferentiated carcinomas from patients having poor prognosis did not stain at all. This study suggests that variant CD44 molecules may serve an important function in the cell contact of cervical epithelial cells, and that cervical epithelium acquires heterogeneity in the expression of CD44 adhesion molecules during carcinogenesis, which may be related to tumor metastasis.

An Eps homology (EH) domain protein that binds to the Ral-GTPase target, RalBP1.

Ral proteins constitute a family of small GTPases that can be activated by Ras in cells. In the GTP-bound state, Ral proteins bind to RalBP1, a GTPase-activating protein for CDC42 and Rac GTPases. We have used the two-hybrid system in yeast to clone a cDNA for a novel approximately 85-kDa protein that can bind to an additional site on RalBP1. This newly identified protein contains an Eps homology (EH) domain, which was first detected in the epidermal growth factor (EGF) receptor substrate Eps15. Recently, the EH domain of Eps15 has been shown to bind to proteins containing an asparagine-proline-phenylalanine motif. Moreover, EH domains have been found in proteins involved in endocytosis and/or actin cytoskeleton regulation. The RalBP1 associated Eps-homology domain protein, Reps1, is tyrosine-phosphorylated in response to EGF stimulation of cells. In addition, Reps1 has the capacity to form a complex with the SH3 domains of the adapter proteins Crk and Grb2, which may link Reps1 to an EGF-responsive tyrosine kinase. Thus, Reps1 may coordinate the cellular actions of activated EGF receptors and Ral-GTPases.

Inhibition of liver metastasis formation by anti-CD44 variant exon 9 monoclonal antibody.

The overexpression of variants of the glycoprotein CD44 is thought to be associated with the tumorigenesis and progression of human cancers. We examined the role of the variant CD44v8-10 in the metastasis of the human colon cancer cell line HT29 using a monoclonal antibody (mAb 44-1V) reactive with the v9 product. After immunization with mAb 44-1V, the growth of HT29m cells in vitro was not retarded. Six-to 8-week-old mice were divided into 4 groups for liver metastasis assay. All animals in control groups injected with intrasplenic HT29m developed metastases. In contrast, only one of the animals injected with HT29m that reacted with mAb 44-1V developed a metastatic tumor in the liver. The intravenous administration of mAb 44-1V after intrasplenic HT29m injection did not inhibit the formation of liver metastasis. In addition, the adhesiveness of the HT29m cells to the basement membrane matrix was decreased by treatment with the anti-CD44v9 mAb. These findings indicated that a CD44 variant containing the products of variants of exons v8-10 may play an important role in adhesion of tumor cells to the capillaries of distant organs in the metastatic process.

Bcl-2 protein expression in breast cancer and its relationship to prognosis.

The expression of bcl-2 protein was studied in invasive breast cancer by immunohistochemistry. Fourty-six (56.8%) bcl-2 protein-positive tumors were found in 81 breast cancers. There was no significant correlation between the bcl-2 protein immunoreactivity and histologic type, primary tumor status, or lymph node metastasis. However, a strong positive relationship was demonstrated between bcl-2 immunoreactivity and estrogen receptor status. The 5-year survival rate and disease-free survival rates were 73.7% and 71.1% of patients with bcl-2-positive tumors, and 62.5% and 58.0% of those with bcl-2-negative tumors; these differences between the two groups of patients wete significant (p<0.05). In multivariate analysis using Cox regression model, bcl-2 immunoreactivity emerged as an independent prognostic indicator in breast cancer patients.

Lectin-histochemical reactivity of sialic acid in breast cancer and its relationship to prognosis using limulus polyphemus agglutinin.

Studies of circulating sialic acid have revealed its relationship with a variety of malignant tumors. It is not vet clear whether sialic acid could be used as a prognostic marker of breast cancer, and few studies have examined sialic acid expression in the cell membrane and cytoplasm of breast cancer cells by means of the lectin-histochemical technique. In the present study, we used biotinylated limulus polyphemus agglutinin (LPA), a special binding lectin of sialic acid, to stain sialic acid in breast cancer cells. Of the 104 cases of breast cancer examined, 59 (56.7%) positive cases were observed. There was a significant correlation between the LPA staining and the clinicopathologic features of all patients, including pathological stage and lymph node metastasis. Among the 100 patients who underwent curative operation, the mean disease-free survival rate of the 45 patients who were LPA-negative was significantly higher than that of the 55 LPA-positive patients (p<0.05). These results suggest that the positive expression of sialic acid in breast cancer could be used as a marker of malignancy potential, as well as a poor survival factor, and the biotinylated LPA assay may provide a convenient and useful method to predict the prognosis of breast cancer.

Significance of tumor marker SLX in colorectal cancer.

We examined serum SLX for its significance as a tumor marker in 109 colorectal cancer patients. There a close correlation with immunohistochemical expression of SLX and serum SLX level. Serum SLX was positive in 16.5% of 109 patients with colorectal cancers. There was no significant correlation between serum SLX level and histologic type or primary tumor status. There were significant correlations between serum SLX positive rates and both lymph node and hematogenous metastasis. In 7 SLX positive cases who underwent curative resection, 4 patients had already recurrence in the liver. Our findings suggest that serum SLX values may be a biologic marker of metastasis.

Expression of a CD44 variant containing exons 8 to 10 is a useful independent factor for the prediction of prognosis in colorectal cancer patients.

PURPOSE: To determine the expression of the CD44 variant containing a variant exon 8 to 10 product (CD44v8-10) in colorectal cancer and to evaluate its prognostic value. MATERIALS AND METHODS: CD44v8-10 was studied in resected tumors and normal mucosae obtained from 215 colorectal cancer patients (118 colon cancer and 97 rectal cancer). The expression of CD44v8-10 was analyzed immunohistochemically using the anti-CD44v8-10 monoclonal antibody (mAb) 44-1V. RESULTS: One hundred of 215 cancer tissues expressed CD44v8-10. Positive staining was intense mainly on the cell membranes. There was no significant correlation between expression of CD44v8-10 and histologic type, primary tumor, lymphatic invasion, venous invasion, or peritoneal invasion. There were significant correlations between CD44v8-10 immunoreactivity and both lymph node and hematogenous metastasis. Patients with CD44v8-10-positive tumors had a greater relative risk of death compared with those whose tumors were CD44v8-10-negative. Among 169 patients who underwent curative resection, CD44v8-10 expression correlated with a high recurrence rate. The 5- and 10-year survival rates were 90.3% of patients with CD44v8-10-negative tumors, and 72.1% and 58.0% of those with CD44v8-10-positive tumors, respectively; these differences between the two groups of patients were significant (P < .01). In multivariate analysis using the Cox regression model, CD44v8-10 expression emerged as an independent prognostic indicator. CONCLUSION: The results suggest that CD44v8-10 plays a role in metastasis of colorectal cancer, and tha CD44v8-10 expression may be a biologic marker of prognostic significance.