Immunohistochemical study of nuclear factor-κB expression in esophageal squamous cell carcinoma: prognostic significance and sensitivity to treatment with 5-FU.

Nuclear factor-κB (NF-κB) is expressed in many types of cancers. It has been suggested that the expression of NF-κB is associated with poor prognosis and resistance to chemoradiation therapies. This study evaluated the relationship between the expression of NF-κB and the prognosis and sensitivity of esophageal squamous cell carcinoma (ESCC) to chemotherapy. One hundred and nine ESCC specimens, from patients who had undergone radical esophagectomy, were divided into two groups depending on the expression of NF-κB. Surgical data and prognosis were compared between the two groups. NF-κB-positive tumors were detected in 61.5% of the cases. In 69 patients with stage II and III disease, 41 patients who were NF-κB-positive showed poor survival. The sensitivity of esophageal squamous cell carcinoma cell lines to 5-fluorouracil (5-FU) was analyzed by their NF-κB expression, and the effect of 5-FU was evaluated on the proliferation and activity of two cell lines of cultured ESCCs expressing NF-κB. ESCCs with activated NF-κB had poor sensitivity to 5-FU. These results suggest that the increased expression of NF-κB is associated with poor prognosis in patients with ESCC. NF-κB may be a target for ESCC therapy because of its selective expression in this type of cancer.

Intramural bronchogenic cyst of the oesophagus.

Bronchogenic cysts are relatively common mediastinal cysts. Here, we report removal of a rare paraoesophageal-type bronchogenic cyst by video-assisted thoracoscopic surgery in a 52-year-old male patient. The defect of the oesophageal wall was successfully reinforced with an intercostal muscle flap.

Benign metastasizing leiomyoma of the lung: potential role of low-grade malignancy.

We report here 2 cases of multiple metastatic lung tumors after hysterectomy for leiomyoma. One patient was diagnosed as having a benign metastasizing leiomyoma (BML), while the other patient simultaneously developed a left pelvic tumor and multiple lung tumors, both of which were finally diagnosed as low-grade endometrial stromal sarcomas (ESSs). The metastatic potential of BML is not completely understood, but previously reported cases of BML may include low-grade ESS, which may play a significant role in the metastasis of benign uterine tumors.

Inverse correlation between NKG2D expression on CD8+ T cells and the frequency of CD4+CD25+ regulatory T cells in patients with esophageal cancer.

Although malignant diseases are known to be associated with immune suppression, detailed mechanisms of this phenomenon are still unknown. NKG2D is an activating cell surface receptor expressed by natural killer (NK) cells and CD8+ T cells, and it has been reported that NKG2D engagement is extremely important for T cell activation. In the current study, NKG2D expression on CD8+ T cells and the frequency of CD4+ CD25+ regulatory T (Treg) cells were determined by multicolor flow cytometry to investigate one of the mechanisms responsible for immune evasion in esophageal cancer patients. NKG2D expression on CD8+ T lymphocytes in esophageal cancer patients was significantly lower than in those of normal controls. NKG2D expression in T3/T4 esophageal cancer was significantly lower than that in T1/T2 esophageal cancer. CD8+ T cells from patients with lymph node metastasis expressed significantly lower NKG2D than those without lymph node metastasis. Moreover, significantly lower NKG2D expression was observed in stage III/IV cancer in comparison with stage I/II. The frequency of CD4+CD25+ Treg cells in esophageal cancer patients was significantly higher than those in normal controls. NKG2D expression on CD8+ T cells was significantly inversely correlated with the frequency of CD4+CD25+ Treg cells in esophageal cancer patients. Our data indicates that decreased NKG2D expression on CD8+ T cells is correlated with disease severity. Decreased NKG2D expression and an increase in Treg cells may be one of the key mechanisms responsible for immune evasion in esophageal cancer.

Abdominal aortic aneurysm in a patient presenting with ischemic colitis.

A 76-year-old man with an abdominal aortic aneurysm (AAA) initially presented with ischemic colitis, which was improved by conservative treatment. Preoperative assessment by computerized axial tomography scanning and aortography revealed an infrarenal type AAA with mural thrombus, stenoses of the right common iliac artery and the left internal iliac artery. The patient underwent aortoiliac bypass surgery with resection of the stenoses, and reconstruction of the left internal iliac artery. No complications including bowel ischemia, were noted postoperatively. This case emphasized the potential benefits of the extraperitoneal approach to the aorta, reconstruction of both internal iliac arteries, and use of prostaglandin E1.

Unusual remnant thymic tissue in an adult mimicking malignant neoplasm: escape from age-related involution.

Here we report on a 55-year-old man with an abnormal anterior mediastinal shadow and multiple nodules in the thymus, which increased in size over a period of 15 months. He was diagnosed with early prostatic cancer, and treated with chemotherapy. Although no definite preoperative diagnosis was obtained, surgery was performed because of the possibility of malignant neoplasm or metastasis. Extended thymectomy was performed and pathological examination revealed that the nodules were remnant thymic tissue and not malignant lesions. Although the cause of this unusual remnant thymic tissue remains unclear, it may have been related to autoimmune or endocrinological disease.

[Coronary subclavian steal syndrome; report of a case].

A 64-year-old man was admitted to our hospital with chief complaint of chest discomfort. He received coronary artery bypass grafting utilizing the in situ left internal thoracic artery 10 years ago. Coronary and left subclavian artery angiogram revealed coronary subclavian steal syndrome and 90% stenosis in the proximal left subclavin artery. Ultrasonography of neck vessels demonstrated 75% stenosis in the bifurcation of left carotid artery. We performed axilloaxillary artery bypass grafting to avoid brain ischemia. Myocardial thallium scintigraphy on dipyridamole testing after axilloaxillary artery bypass grafting could not detect myocardial ischemia. Axilloaxillary artery bypass grafting was effective for coronary subclavian steal syndrome.

[Hemolytic anemia due to aortic valve regurgitation after mitral valve replacement].

A 50-year-old woman was admitted to our hospital because of heart failure (NYHA III) due to mitral valve regurgitation (MR) with pulmonary hypertension (PH) and tricuspid valve regurgitation (TR). She had a history of chronic renal failure undergoing dialysis (peritoneal dialysis, homodialysis) since 1996. Cardiac catheterization and ultrasonic cardiography showed severe MR (Sellers III), severe TR and PH (mean pressure 33 mmHg). So we performed mitral valve replacement and tricuspid annuloplasty (DeVega). Frequent blood transfusion was needed because severe hemolytic anemia appeared after operation. Ultrasonic cardiography demonstrated moderate aortic valve regurgitation (AR) with no paravalvular prosthetic leakage. We diagnosed hemolytic anemia due to AR. We performed aortic valve replacement. Hemolytic anemia improved soon after second operation. We investigated the mechanical process of the AR. She had a very short subaortic curtain (5.9 mm) compared with the average (8.7 +/- 2.1 mm: mean +/- SD) of cardiac patients. We think that we must be very careful with suture to short subaortic curtain. In addition measurement of subaortic curtain before operation is very useful.

Differential sensitivities of the MRP gene family and gamma-glutamylcysteine synthetase to prooxidants in human colorectal carcinoma cell lines with different p53 status.

1Recent molecular cloning studies have identified six members in the multidrug-resistance protein (MRP) gene family. However, the regulation of expression of these genes is largely unknown. We previously reported that expression of MRP1, encoding multidrug-resistance associated protein, and gamma-GCSh, which encodes the heavy subunit of gamma-glutamylcysteine synthetase (gamma-GCS), could be up-regulated by prooxidants [Yamane et al., J Biol Chem 1998;273:31075-85]. In the present study, we investigated whether different members of the MRP family exhibit different responses to induction by prooxidants, and whether p53 status influences the levels of induction. A panel of colorectal cancer cell lines with different p53 status, i.e. HCT116 containing wild-type p53, and HT29, SW480, and Caco2 containing mutant p53, was treated with tert-butylhydroquinone (t-BHQ) and pyrrolidinedithiocarbamate (PDTC). MRP1 and gamma-GCSh mRNA levels were determined by the RNase protection assay, using gene-specific probes. We report here that induction of MRP1 and gamma-GCSh expression by these prooxidants varied among the different cell lines, and p53 mutations were not always associated with elevated levels of induction. These results suggest that the effects of p53 on the induced expression of MRP1 and gamma-GCSh depend on the environment of the cell and/or nature of p53 mutations. In an isogenic HCT116 cell line containing p53(-/-) alleles, we demonstrated that, as for MRP1, expression of MRP2 and MRP3 was induced by the prooxidants, whereas expression of MRP4 and MRP5 was not. MRP6 mRNA was not detectable. Induction of MRP2 expression by prooxidants seemed to be independent of p53 status. Our results demonstrated the differential regulation of the MRP gene family by p53 mutation under oxidative stress.

Induction of MRP1 and gamma-glutamylcysteine synthetase gene expression by interleukin 1beta is mediated by nitric oxide-related signalings in human colorectal cancer cells.

Treatment of human colorectal cancer cells HT29 with interleukin 1beta (IL-1beta) induces expression of the multidrug resistance protein (MRP1) gene encoding the ATP-dependent glutathione S-conjugate export (GS-X) pump and the gamma-glutamylcysteine synthetase (gamma-GCSh) gene encoding heavy (catalytic) subunit of gamma-glutamylcysteine synthetase, the rate-limiting enzyme for the biosynthesis of glutathione (GSH). The induction can be suppressed by N(G)-methyl-L-arginine, a specific inhibitor of nitric oxide synthase (NOS). These results suggest that IL-1beta-mediated MRP1 and gamma-GCSh induction involve nitric oxide (NO) -related signaling. Further supports to the involvement of NO in the induction of MRP1 and gamma-GCSh expression are made by the following observations. (i) Expression of MRP1 and gamma-GCSh genes were induced by treating the cells with NO donors, i.e., S-nitro-N-acetyl-D,L-penicillamide (SNAP) and S-nitroso-L-glutathione, in a concentration-dependent manner. (ii) Ectopic expression of inducible NOS (iNOS) activity by transfecting expressible recombinant iNOS cDNA encoding functional iNOS but not the nonfunctional version resulted in elevated expression of MRP1 and gamma-GCSh. We also demonstrated that HT-29 cells treated with either 1L-1beta or SNAP induced ceramide production, and addition of C2 or C6 ceramides into cultured HT-29 cells resulted in induction of gamma-GCSh but not MRP1 expression. Collectively, our results demonstrate that induction of MRP1 and gamma-GCSh by IL-1beta is regulated, at least in part, by an NO-related signaling, and induction of gamma-GCSh is by NO-related ceramide signaling.

Adenovirus-mediated transfer of wild-type p53 gene results in apoptosis or growth arrest in human cultured gastric carcinoma cells.

We examined the susceptibility of six human gastric carcinoma cell lines to infection with recombinant p53 adenovirus vector (AxCA-p53). AxCA-p53 infection at a muliplicity of infection (MOI) of 50 resulted in apoptotic cell death (MKN-1 cells), growth arrest (MKN-45, MKN-74 and KATO-III cells), or non-effectiveness (TMK-1 and OCUM-2M cells). Western blot analysis revealed increasing expression levels of p21/WAF1 protein after infection with AxCA-p53 in all the cell lines. After infection with AxCA-p53, the expression levels of bax or bcl-XL protein changed in MKN-1, but not in the other cell lines. These results suggest that the apoptotic pathway (dependence on the expressions of bcl-2 family proteins) dominates the growth arrest pathway (dependence on the expressions of p21/WAF1 protein) after infection with AxCA-p53. Thus, the bcl-2 family might play a crucial role in p53-mediated growth arrest and apoptosis in human gastric carcinoma cells.

Adenoviral transduction efficiency partly correlates with expression levels of integrin alphavbeta5, but not alphavbeta3 in human gastric carcinoma cells.

Adenovirus has attracted much attention as a vector for gene therapy. Integrin alphavbeta3 and alphavbeta5 mediate adenovirus internalization into cells. We examined adenoviral transduction efficiency and expression of integrin alphavbeta3 alphavbeta5 in 9 human gastric carcinoma cell lines. The percentage of -gal-positive cells was more than 85% 3 days after infection with recombinant adenovirus carrying the bacterial LacZ gene (AxCALacZ) at a dose of 25 MOI in 7 cell lines and the transduction efficiency was 32 and 42% in HSC-39 and MKN-28 cells, respectively. Adenoviral transduction efficiency did not correlate with the histological cell types. Flow cytometric analysis revealed relatively high expression of integrin alphavbeta5 in MKN-28 and OCUM-2M cells, followed by MKN-1, MKN-7, MKN-45, MKN-74, TMK-1 and KATO-III cells. HSC-39 cells minimally expressed integrin alphavbeta5. On the other hand, integrin alphavbeta3 was expressed only in MKN-1 cells. These results suggest that the adenovirus vector might be useful for gene transduction into human gastric carcinoma cells and the transduction efficiency partly correlates with the expression levels of integrin alphavbeta5 but not integrin alphavbeta3.

[A new device for allergen skin testing].

We have made a new device for allergen skin testing, which can be used easily. It consists of two parts. One, several gears are fixed on a board in a row, and when one gear goes round, other gears also go round simultaneously. A disposable needle is attached to each gear. A semi-spherical hole is made in the lower surface of a disposable needle, and a sharp needle is fixed in, but not at the centre of the hole. The other, small tanks are arranged in a row at the same distance as the gears. When the disposable needles are inserted to the tanks filled with allergen extract, it is hold in a semi-spherical hole by surface tension. The allergen extracts go intradermally, when the needles move semi-circular on a human skin. We performed skin tests to 23 patients with this device, and got satisfactory results as screening tests.

Clinical significance of dendritic cell infiltration in esophageal squamous cell carcinoma.

We investigated the clinicopathological significance of dendritic cell infiltration (DCsI) in esophageal squamous cell carcinoma and in regional lymph nodes of 88 patients. The expression of mutated p53 protein and the degree of positive cancer cells of proliferating cell nuclear antigen (PCNA labeling index) in tumors were analyzed as biological markers. These factors were compared with the degree of DCsI in tumors and in lymph nodes. The number of dendritic cells (DCs) were counted and scored as per mm2 in each case. The degree of DCsI of tumors with expression of p53 (19/mm2, n=50) was significantly lower than that of DCsI in 38 tumors without expression of p53 (27/mm2, P=0.0411). However, no significant correlation was detected between the PCNA labeling index and the degree of DCsI in 88 primary tumors (P=0.1273). The degree of DCsI in 53 metastatic lymph nodes (30/mm2) was significantly lower than that of DCsI in 264 cancer-free regional lymph nodes (48/mm2, P=0. 02). Although the degree of DCsI in tumors was not an independent prognostic factor for the 78 surviving patients (P=0.2647), the 3-year survival rate of patients in stage III and IV who underwent curative operation and who had tumors with high DCsI (>9/mm2, n=16, 72%) was significantly higher than that of the 24 patients who had tumors with low DCsI (< or = 9/mm2, 21%, P=0.008). These findings indicate that DCs infiltrated in and around the esophageal cancer may play a defensive role of the hosts against the tumors. This immune defense of the hosts might be an important prognostic factor for patients with advanced esophageal cancer. However, cancer cells which express a mutated p53 protein might regulate the function or activity of DCs.

Apoptosis and cellular proliferation in human epidermal squamous cell neoplasia.

We examined cell loss (apoptosis) and proliferation in a histopathological spectrum of epidermal squamous cell neoplasia, including 11 cases of solar keratosis (SK), 18 Bowen's diseases (BD) and 19 invasive squamous cell carcinomas (SCC). Apoptotic and proliferative cells were determined by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end labeling (TUNEL) and by the detection of nuclear antigen Ki-67, respectively. Few apoptotic cells were observed in normal epidermis, while TUNEL index (TI; percentage of TUNEL-positive cells) was highest for SCCs, followed by BDs and SKs, in the order given. Although the mean Ki-67 index did not differ between SCCs and BDs, both disease types showed a significantly higher index than the SKs. Of SCCs, both TI and Ki-67 index values were significantly higher in poorly than in well differentiated carcinomas. TI was significantly higher in SCCs without P53 immunohistochemical expression than in SCCs with P53 expression, while TI and Ki-67 indices did not correlate with P53 expression in the SKs and BDs. These results suggest that apoptosis reflects not only cell loss, but also proliferative activity in the epidermal neoplastic lesions.

Anti-Fas antibody-induced apoptosis in human colorectal carcinoma cell lines: role of the p53 gene.

The cell surface Fas antigen transducts an apoptotic signal by its crosslinking with Fas ligand or anti-Fas antibody in a variety of human cultured cells. In this study, we examined the expression of Fas antigen and its mediation of apoptosis in six human colorectal carcinoma cell lines. A flow cytometric analysis revealed that LoVo, DLD-1, WiDr and SW837 cell lines showed higher expression levels of Fas antigen, in contrast to lower expression in COLO201 and COLO320DM. Interferon-gamma enhanced the expression of Fas antigen in all of the cell lines examined. Both Fas ligand and Fas-associated phosphatase-1 (FAP-1) were expressed only in COLO320DM. Anti-Fas antibody induced apoptosis in LoVo carrying wild-type p53 gene, but not in the other five cell lines carrying mutated p53 gene. The transfection of wild-type p53 gene using an adenovirus vector upregulated P53 protein in WiDr and SW837 cells, both of which showed, however, no increase in apoptotic cells by anti-Fas antibody treatment. These results indicated that (1) Fas antigen was variably expressed, regardless of the p53 gene status and (2) the susceptibility to anti-Fas antibody-mediated apoptosis did not correlate to Fas, Fas ligand or FAP-1 expression levels. Therefore, we conclude that wild-type P53 expression might not necessarily be essential for Fas-mediated apoptosis in human colorectal carcinoma cell lines.

[Development of a blood cardioplegia delivery system for children].

We have developed a blood cardioplegia delivery system for children. Essential points of a delivery system in pediatric cardiac surgery are (1) a small amount of priming volume of a delivery system, and (2) slow, steady infusion of a cardioplegic solution. We changed a heat exchanger to a smaller one for reduction of priming volume, and changed a roller pump tube to a smaller one for slow, steady infusion. Thus, priming volume of a delivery system has reduced from 180 to 100 ml, and we can infuse a cardioplegic solution at a steady rate less than 10 ml/min. Our clinical experience with this system suggests that this blood cardioplegia delivery system is useful for pediatric cardiac surgery.

Expression of p21 (waf1/cip1/sdi1), but not p53 protein, is a factor in the survival of patients with advanced gastric carcinoma.

BACKGROUND: The authors examined whether expression of p21 (waf1/cip1/sdi1) and p53 protein was related to survival, rates in patients with advanced gastric carcinoma. METHODS: The expression of p21 and p53 protein was analyzed by immunohistochemistry in 93 patients with advanced gastric carcinoma with serosal invasion and lymph node metastasis. All patients underwent curative surgery. The probability of survival was calculated by the Kaplan-Meier method and compared by the generalized Wilcoxon test. RESULTS: Various levels of p21 and p53 immunoreactivities in carcinoma cells were detected in 30 (32%) and 60 (65%), respectively, of 93 samples. There was no correlation between p21 and p53 expression. The 5-year survival rate of patients with p21 expression was 69.4%, which was significantly better than that of patients without p21 expression (38.1%; P < 0.05). However, p53 protein expression did not correlate with patient survival. CONCLUSIONS: Expression of p21 protein may be a better prognostic factor than p53 protein expression in patients with advanced gastric carcinoma.