Clinical significance of plasma fibrinogen level as a predictive marker for postoperative recurrence of esophageal squamous cell carcinoma in patients receiving neoadjuvant treatment.

Among multidisciplinary therapies developed for advanced esophageal cancer, neoadjuvant chemotherapy and chemoradiotherapy have been established as standard treatments. To deliver cautious follow up and intense treatment for high-risk patients, a simple and instructive biomarker for the postoperative recurrence needs to be identified. Fibrinogen, a common component of hemostasis, has been suggested to not only play an important role in cancer metastasis, but also correlate with tumor recurrence. We aim to clarify the validity of plasma fibrinogen as a marker for predicting the postoperative recurrence of esophageal squamous cell carcinoma patients who received neoadjuvant treatment. We reviewed 72 consecutive patients with esophageal squamous cell carcinoma who received neoadjuvant chemotherapy or chemoradiotherapy, followed by esophagectomy at the Keio University Hospital from 2001 to 2010. Of them, we retrospectively examined 68 patients who underwent plasma fibrinogen examination before and after neoadjuvant treatment and underwent transthoracic radical esophagectomy. We investigated patient characteristics, clinicopathological factors, neoadjuvant treatment effects, postoperative course, and plasma fibrinogen levels. We investigated pretreatment and preoperative (postneoadjuvant treatment) plasma fibrinogen levels, as well as changes in fibrinogen levels before and after neoadjuvant treatment. Patients with preoperative hyperfibrinogenemia (>350 mg/dL) and patients with increased plasma fibrinogen levels during neoadjuvant treatment showed significantly shorter postoperative disease-free survival (DFS) (P = 0.002 and P = 0.037, respectively). Moreover, we classified these patients into three classes on the basis of their preoperative fibrinogen levels and changes in fibrinogen levels during neoadjuvant treatment. Patients who had both high preoperative plasma fibrinogen and increased fibrinogen levels showed significantly shorter DFS than others. In contrast, patients who had normal preoperative plasma fibrinogen and decreased fibrinogen levels showed significantly longer DFS. Based on this fibrinogen classification, we could differentiate between significantly favorable and poor prognosis patients group. Overall, this classification (hazard ratio = 1.812, P = 0.013) and the response to neoadjuvant treatment (hazard ratio = 0.350, P = 0.007) were found to be significant determining factors for postoperative DFS. With the validity of preoperative plasma fibrinogen levels and changes in fibrinogen levels during neoadjuvant treatment, the plasma fibrinogen level was found to be a possible biomarker for postoperative recurrence in advanced esophageal cancer patients who received neoadjuvant treatment. Moreover, plasma fibrinogen classification could be a simple and valuable predictive marker for postoperative follow up.

Clinical utility of preoperative evaluation of bronchial arteries by three-dimensional computed tomographic angiography for esophageal cancer surgery.

An identification of bronchial arteries (BAs) is critical in esophageal cancer surgery to avoid tracheobronchial ischemia and unexpected massive bleeding during surgical procedure particularly in thoracoscopic video-assisted esophagectomy. We describe the efficacy of three-dimensional computed tomographic angiography (3D-CTA) of BAs for preoperative evaluation in esophageal cancer surgery. Sixty-four patients with esophageal cancer who preoperatively underwent multidetector computed tomography examination were included in this study. We evaluated the number, origin, and intraoperative preservation rate of BAs, and we compared the number of thoracic paratracheal lymph nodes harvested between two groups comprising patients who either underwent preoperative 3D-CTA of BAs (3D-CTA group) or did not (non-3D-CTA group). The right and left BAs were preoperatively identified in 62 patients (97%) and 55 patients (86%), respectively, using 3D-CTA. In 34 patients (53%), the right BA originated as a common trunk with the right intercostal artery. In 48 patients (75%), the left BA originated from the descending aorta as a single or double branch. Some anomalies such as the right BA originated from the left subclavian artery were observed. In all patients, either the right or the left BA was preserved. The number of harvested lymph nodes in left side of paratrachea was significantly increased in 3D-CTA group, than those in non-3D-CTA group. 3D-CTA clearly revealed BA anatomy, contributing to BA preservation and safe and precise lymphadenectomy in esophageal cancer surgery. 3D-CTA of BAs is useful for preoperative evaluation in esophageal cancer surgery.

[Clinical application of sentinel node navigation surgery for esophageal cancer].

In esophageal cancer, sentinel nodes (SNs) are identified as multiple nodes and widely spread from cervical to abdominal areas. In more than 80% of the cases, at least one SN is located in the 2nd or 3rd compartment of regional lymph nodes which have been considered to be "skip metastases". This characteristic distribution of SNs is attributed to the multi-directional lymphatic drainage routes from the esophagus. Clinical application of SN navigation surgery will be expected to play a key role for intraoperative diagnosis for lymph node metastasis and individualized multimodal therapy in patients with cT1N0 esophageal cancer.

Neutrophil elastase inhibitor improves postoperative clinical courses after thoracic esophagectomy.

Sivelestat sodium hydrate is a selective inhibitor of neutrophil elastase (NE), and is effective in acute lung injury associated with systemic inflammatory response syndrome (SIRS). The effect of Sivelestat for postoperative clinical courses after transthoracic esophagectomy was investigated. Consecutive patients with carcinoma of the thoracic esophagus who underwent transthoracic esophagectomy between 2003 and 2004 were assigned to the Sivelestat-treated group (n = 18), and those between 1998 and 2003 were assigned to the control group (n = 25). The morbidity rate, duration of postoperative SIRS, mechanical ventilation, and intensive care unit (ICU) stay, and the sum of the sequential organ failure assessment scores at all time points after the operation were compared. Serum NE activities and serum concentrations of TNF-alpha, IL-1beta, IL-6, and high mobility group box chromosomal protein 1 (HMGB1) were measured. Postoperative complications developed in three patients in the control group, and one in the Sivelestat-treated group. The durations of SIRS, mechanical ventilation, and ICU stay were significantly shorter in the Sivelestat-treated group. Even in patients without complications, the durations of mechanical ventilation, and ICU stay were also significantly shorter, and the arterial oxygen pressure/fraction of inspired oxygen ratio at postoperative day 1 was significantly higher in the Sivelestat-treated group. Serum NE activities and serum concentrations of IL-1beta, IL-6, and HMGB1 were significantly suppressed in the Sivelestat-treated group. Postoperative Sivelestat treatment after transthoracic esophagectomy improves the condition of SIRS and postoperative clinical courses, even in patients without complications.

Serum concentrations of high-mobility group box chromosomal protein 1 before and after exposure to the surgical stress of thoracic esophagectomy: a predictor of clinical course after surgery?

High-mobility group box chromosomal protein 1 (HMGB-1) has recently been shown as an important late mediator of endotoxin shock, intra-abdominal sepsis, and acute lung injury. However, its role in the systemic inflammatory response syndrome after major surgical stress, which may lead to multiple organ dysfunction syndrome, has not been thoroughly investigated. We hypothesized that serum HMGB-1 participates in the pathogenesis of postoperative organ system dysfunction after exposure to major surgical stress. A prospective clinical study was performed to consecutive patients (n = 24) with carcinoma of the thoracic esophagus who underwent transthoracic esophagectomy with three field lymph node resection between 1998 and 2003 at Keio University Hospital, Japan. Serum HMGB-1 concentrations were measured by enzyme-linked immunosorbent assay. Preoperative serum HMGB-1 levels correlated with postoperative duration of SIRS, mechanical ventilation, and intensive care unit stay. Three of the 24 patients had serious postoperative complications: sepsis in two, and acute lung injury in one. Serum HMGB-1 levels in patients without complications increased within the first 24 h postoperatively, remained high during postoperative days 2-3, and then decreased gradually by postoperative day 7. In patients with serious complications, serum HMGB-1 was significantly higher than that found in patients without postoperative complications at every time point except postoperative day 2. Preoperative serum HMGB-1 concentration seems to be an important predictor of the postoperative clinical course. Transthoracic esophagectomy induces an increase in HMGB-1 in serum even in patients without complications. Postoperative serum HMGB-1 concentrations were higher in patients who developed complications, and may be a predictive marker for complications in this setting.

Expression of a candidate marker for progenitor cells, Musashi-1, in the proliferative regions of human antrum and its decreased expression in intestinal metaplasia.

AIM: Reliable makers for progenitor cells in the human stomach have not been elucidated. The aim of the present study was to clarify whether Musashi-1 (Msi-1), which has recently been proposed as a stem cell marker in mouse intestine, serves as a marker for progenitor cells in human stomach. METHODS AND RESULTS: Immunohistochemistry revealed that Msi-1+ cells were detected especially in the isthmus/neck region (the putative position of stem cells) of the adult antrum, but were limited to the basal regions of fetal pyloric glands during the early stages of development. These results suggest that Msi-1 expression occurs specifically in the stem cell-containing regions. Msi-1+ cells were intermingled with proliferating cell nuclear antigen (PCNA)+ cells in the isthmus/neck region of the adult antrum, but did not coexpress PCNA or Ki 67. Msi-1 expression overlapped partly with expression of MUC 5 AC and MUC 6, indicating that Msi-1+ cells retain some features of both foveolar and pyloric gland cell differentiation phenotypes. In contrast, Msi-1 expression in gastric glands showing intestinal metaplasia (IM) became weaker than that in the glands without IM. CONCLUSION: The specific expression of Msi-1 within the proliferative regions suggests that Msi-1 is a marker of cells with progenitor characteristics before active proliferation in human antrum.

Anti-tumour effects of nobiletin, a citrus flavonoid, on gastric cancer include: antiproliferative effects, induction of apoptosis and cell cycle deregulation.

AIM: To demonstrate the antitumour effects of nobiletin (5,6,7,8,3',4'-hexamethoxyflavone), a citrus flavonoid extracted from Citrus depressa Hayata, on human gastric cancer cell lines TMK-1, MKN-45, MKN-74 and KATO-III. MATERIALS AND METHODS: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the TdT-mediated dUTP biotin nick-end labelling (TUNEL) method and cell-cycle analysis revealed that nobiletin acted on these cells in several ways, namely by direct cytotoxicity, induction of apoptosis and modulation of cell cycle. The efficacy of combined treatment of nobiletin with a conventional anticancer drug, CDDP, was also examined. Treatment with nobiletin 24 h prior to CDDP administration showed a synergistic effect compared to the control. CONCLUSIONS: Although the effective dose and administration route of nobiletin require further investigation, our study represents a potential successful linking of this compound with the treatment of gastric cancer.

Cyclin D1 antisense oligonucleotide inhibits cell growth stimulated by epidermal growth factor and induces apoptosis of gastric cancer cells.

The cyclin D1 protein is one of the cell cycle regulators required for cell cycle progression through G1 phase to S phase. The cyclin D1-cyclin-dependent kinase (CDK) system is thought to control the cell cycle through mediating extracellular signals from mitogens, such as epidermal growth factor (EGF). In this study, we attempted to examine the therapeutic effect of cyclin D1 antisense oligonucleotides (AS/D1) on cell proliferation and apoptosis of the gastric cancer cell line MKN-74, in the presence and absence of EGF-stimulation. Evaluation of cell survival and DNA synthesis revealed that enhanced cell growth following EGF-stimulation was completely inhibited by a 24 h pre-incubation with 100 nM AD/D1. This inhibition was down to 19.3% compared with maximal DNA synthesis after stimulation with 3 nM EGF alone. Western blotting demonstrated that while EGF-stimulation led to cyclin D1 over-expression, AS/D1 inhibited cyclin D1 protein expression. We also demonstrated the induction of apoptosis in MKN-74 cells by AS/D1. In conclusion, EGF-stimulated MKN-74 cell proliferation was inhibited by AS/D1, which could overcome EGF-induced cyclin D1 over-expression. AS/D1 also affected cell survival by inducing apoptosis through cell cycle arrest following cyclin D1 depletion. Thus, AS/D1 may be a candidate for use as a novel cancer therapy specifically targeted against the over-expression of cyclin D1 enhanced by EGF in malignant cells.

A novel single-nucleotide polymorphism in the 3'-untranslated region of the human dihydrofolate reductase gene with enhanced expression.

A novel single-nucleotide polymorphism (SNP), 829C-->T in the 3'-untranslated region of the human dihydrofolate reductase (DHFR) gene transcript, was identified in the study population of 37 patients with childhood leukemias/lymphomas and 83 healthy Japanese children. Frequencies of the DHFR 829C/C, 829C/T, and 829T/T genotypes were 83.8, 10.8, and 5.4%, respectively, in the cases and 74.7, 19.3, and 6.0% in the controls, showing no significant difference in genotype frequencies between the cases and controls. When determined by real-time quantitative reverse transcription-PCR analysis, the highest expression of the DHFR transcript was demonstrated in the samples with a DHFR 829T/T polymorphism (P < 0.001). Direct association of the presence of the SNP with methotrexate-related adverse events in each patient was not demonstrated in this limited analysis. These data suggest that the novel DHFR 829 polymorphism is associated with a positive role in gene expression and provide evidence of a functional SNP in the 3' regulatory region of the gene.

Structural evidence of genomic exon-deletion mediated by Alu-Alu recombination in a human case with heme oxygenase-1 deficiency.

We previously reported a family affected by heme oxygenase-1 (HO-1) deficiency [Yachie et al., 1999]. The proband was a compound heterozygote for a complete loss of exon 2 (the maternal allele) and a two-nucleotide deletion within exon 3 (the paternal allele). In this report, we describe a large genomic deletion (1730 bp) including entire exon 2 in this family as a specific mechanism generating exon-2 absence observed in the HO-1 mRNA. Analysis of the deletion junction demonstrated fusion of a 5' portion of Alu-Sx element with a 3' portion of Alu-Sq element. The junction contained sequences with high homology to the recombinogenic Alu "core" sequence. These structural features of the HO-1 gene suggest homologous recombination associated with Alu element. This study presents the initial characterization of the HO-1 gene defect causing a human case of HO-1 deficiency and provides the molecular basis for understanding this genetic disease.

Jejunal limb obstruction caused by a cholesterol stone 15 years after a total gastrectomy and 20 years after a cholecystectomy: report of a case.

We present herein the rare case of a 74-year-old woman found to have jejunal limb obstruction caused by a cholesterol stone 15 years after a total gastrectomy with Roux-en-Y anastomosis, and 20 years after a cholecystectomy. The patient complained of repeated episodes of upper abdominal distress on three separate occasions over a period of 20 months, and jejunal limb obstruction was diagnosed by abdominal computed tomography scanning and (99m)Tc scintigraphy. Surgery revealed a stone incarcerated in the jejunal limb, where the anastomosis had become slightly stenotic. The removed stone was 3.5 cm in diameter and was subsequently demonstrated to be a cholesterol stone by chemical analysis. This report is thought to be the first to describe jejunal limb obstruction caused by a gallstone incarcerated in the jejunal limb after a total gastrectomy in a patient with a history of cholecystectomy.

Absolute configuration and tautomeric structure of xylindein, a blue-green pigment of Chlorociboria species.

The S absolute configuration of both chiral centers of xylindein was assigned using X-ray crystallographic heavy atom analysis after its conversion to a synthetic derivative. Crystallographic analysis of xylindein crystallized with phenols revealed that the proposed structure is the proper tautomer in the crystals.

Microsatellite analysis of childhood leukemia: correlation of 9p and 12p chromosome abnormalities with expression of related genes.

BACKGROUND: Loss of heterozygosity (LOH) and DNA replication error (RER) have been thought to be involved in carcinogenesis, but have not been investigated in childhood leukemia and lymphoma. METHODS: Eighty samples from 65 patients with childhood leukemia and lymphoma were examined using seven different microsatellite markers for RER analysis. Additionally, LOH in two chromosome regions (9p and 12p) was investigated. Furthermore, expression of the TEL, TEL/AML1 and p27(KIP1) genes on 12p and the p16 gene on 9p were detected by reverse transcriptase polymerase chain reaction. RESULTS: Replication errors were detected in 5/65 patients (7.7%). Most (4/5 patients) RER were preferentially located in the 9p and 12p regions. There were two patients who had DNA abnormalities in both 9p and 12p, one with common acute lymphoblastic leukemia (ALL) showed 9p LOH and the TEL/AML1 fusion gene on 12p and the other with common ALL and 12p RER had diminished expression of both the p27(KIP1) gene on 12p and the p16 gene on 9p. CONCLUSIONS: Combined DNA alterations on 9p and 12p, involving LOH, RER and/or gene mutation and chromosomal translocation, were found in childhood acute leukemia, especially in common ALL.

Increasing prevalence of ampicillin- resistant, non-beta-lactamase-producing strains of Haemophilus influenzae in children in Japan.

Among Haemophilus influenzae isolated from children with respiratory tract infections, the evolution of ampicillin resistance was investigated during 1996 and 1997 in Japan. beta-Lactamase production was assessed and minimum inhibitory concentrations (MICs) of eight antimicrobial agents were determined using a broth microdilution method in Mueller-Hinton-lysed horse blood medium. Of 74 H. influenzae, 11 strains (14.9%) produce beta-lactamase and were thus highly resistant to ampicillin (MIC of >4.0 microgram/ ml). In addition, moderate resistance to ampicillin, defined as an MIC of >==1.0 microgram/ml, was noted in 44.4% of all beta-lactamase-negative isolates. These beta-lactamase-negative ampicillin-resistant (BLNAR) organisms were resistant to other cephalosporins such as cefpodoxime and cefdinir, while beta-lactamase-producing strains were susceptible to them. Cefditoren, cefteram, and minocycline were active against all strains studied, whereas cefaclor and clarithromycin were inactive against all H. influenzae isolates in this study. Results indicate that BLNAR strains have emerged among children with respiratory tract infections in Japan.

Selectively induced high MRP gene expression in multidrug-resistant human HL60 leukemia cells.

A subclone HL60/DOX was selected from a human leukemic HL60 cell line for resistance to doxorubicin (DOX) by exposure to stepwise increasing concentrations of the drug and coexposure to a potential P-glycoprotein (P-gp) inhibitor, cepharanthine (a biscoclaurine alkaloid). Compared with the parent HL60 cells, the HL60/DOX cells were 13.0-fold more resistant to DOX and showed multidrug-resistant (MDR) phenotype characterized by 4.6-fold, 2.3-fold, and 5.7-fold cross-resistance to vincristine, pirarubicin, and etoposide, respectively, but no cross-resistance to alkylating agent, cisplatin. Immunocytochemical analyses using the specific monoclonal antibody, MRPr1, and quantitative analyses using a competitive reverse transcription-polymerase chain reaction (CRT-PCR) confirmed overexpression of MRP gene products (about 8-fold determined by CRT-PCR) in this resistant clone. The P-gp expression was not detectable by the monoclonal antibody, C219, in the HL60/DOX cells, and that was consistent with extremely low levels of mdr1 mRNA expression determined by CRT-PCR in this clone. Drug accumulation and efflux studies demonstrated the significantly increased efflux rate of DOX compared to the parent HL60 cells. This enhancement of DOX efflux was reversed by the addition of 10 microM verapamil. To investigate the additional underlying mechanisms contributing to MDR phenotype in the HL60/DOX cells, the levels of DNA topoisomerases (Topo) including Topo I, Topo IIalpha, and Topo IIbeta, and gamma-glutamylcystein synthetase (y-GCS) expression were determined using CRT-PCR techniques. Normal expression of each enzyme at the transcriptional level was demonstrated in this resistant clone. Southern blot analysis of the gene organization in the HL60/DOX cells revealed the amplification of MRP gene. These results indicate that alteration of the drug accumulation from enhanced efflux appears to be a major mechanism(s) of MDR phenotype and attributable to high levels of MRP expression in the HL60/DOX cells. Overexpression of MRP in this clone is regulated by the genomic amplification of DNA and increased levels of the MRP mRNA, independently with the normal expression of Topo I, Topo IIalpha, Topo IIbeta, or gamma-GCS.

Membrane-type matrix metalloproteinase-1 expression and activation of gelatinase A as prognostic markers in advanced pediatric neuroblastoma.

BACKGROUND: Recently a novel membrane-type matrix metalloproteinase-1 (MT-MMP-1) was discovered to be a specific activator of progelatinase A, and was correlated with tumor invasion. To the authors' knowledge, no information regarding the expression of MT-MMP-1 has been reported in childhood malignancies. In this study, the authors attempted to elucidate the specific mechanisms that underlay the invasive behavior of neuroblastoma (NB) cells with respect to the expression of MT-MMP-1 and its determined prognostic value, especially in pediatric patients with advanced Evans' Stage IV NB. METHODS: Thirty specimens from surgically excised NB (mainly Stage IV) were collected retrospectively. The total levels of progelatinase A (68 kilodaltons [kD]) and its activated form (62 kD) in the tumor lysates were quantified by gelatin zymography. The expression of MT-MMP-1 was estimated by immunostaining with a monoclonal antibody (113-5B7). RESULTS: Progelatinase A and the activated form were detected in each of the 30 specimens. The gelatinase A activation ratio, 62 kD/(62 kD + 68 kD), strongly correlated with the high levels of MT-MMP-1 expression found in specimens of advanced tumor stage. In the patients with advanced Stage IV NB, the activation ratio was strongly associated with unfavorable clinical outcome; the 5-year survival was 88.9% in the patients with a low activation ratio (< or = 26%) versus only 21.2% in the patients with a high activation ratio (>26%). CONCLUSIONS: Gelatinase A activation correlates with high expression of MT-MMP-1 on NB cells and is associated strongly with advanced stage and poor clinical outcome. These results are consistent with the notion that MT-MMP-1 expression is an important prognostic determinant of the biologic behavior of NB.