Infiltrating regulatory T cell numbers is not a factor to predict patient's survival in oesophageal squamous cell carcinoma.

CD4/8 status has been previously reported to be a critical factor in the prognosis of oesophageal squamous cell carcinoma (OSCC). In the current study, we investigated the effect of regulatory T cells (Treg; Foxp3+ lymphocytes) on the status of CD4+ and CD8+ T cells in 122 patients with OSCC. Immunohistochemical analysis of Treg was performed using an antibody against Foxp3. The survival rate for low Foxp3 patients was significantly lower than for high Foxp3 patients (P=0.0028 by log-rank test), but Foxp3 status did not significantly correlate with prognosis in CD4/8(+/+) patients or CD4/8(+/-) or (-/+) patients (P=0.5185 and 0.8479, respectively, by log-rank test). We also found that Foxp3 status correlated with CD4/8 status (P=0.0002 by chi2 test) and that the variance of CD8/CD4 ratio in patients with low Foxp3 was larger than in patients with high Foxp3 (P<0.0001 by F-test). Thus, the results do not support the idea that Treg suppress anti-tumour immunity in patients with OSCC. Rather, the CD8/CD4 ratio and CD4/8 status appear to be critical factors in anti-tumour immunity. Furthermore, Treg numbers correlate with both the CD8/CD4 ratio and the CD4/8 status, suggesting that Treg number is not a factor to predict patient's survival in OSCC.

Molecular epidemiology of rabies from Maranhão and surrounding states in the northeastern region of Brazil.

Although many outbreaks of rabies have been reported in northern Brazil, few epidemiological studies of these outbreaks have been undertaken. In this study, molecular epidemiological analyses were performed using 41 rabies virus samples isolated in the Maranhão (MA), Pará (PA), and Tocantins (TO) states of northeastern Brazil. A 599-bp region of the glycoprotein (G) gene was first amplified from each sample by RT-PCR, then sequenced and subjected to phylogenetic analysis. A phylogenetic tree divided the 41 isolates into two clades: Clade I was associated with terrestrial carnivores and Clade II was associated with vampire bats. The Clade I isolates were further sub-divided into two groups. The first group was closer to carnivore isolates that predominate in central Brazil, whereas the second group more closely resembled wild fox isolates from the northeastern coastal state of Paraíba (PB). MA isolates of Clade II formed an entirely separate group. These results demonstrate that bat- and dog-transmitted rabies occur in northwestern Brazil.

Solitary adrenal metastasis in a patient with sigmoid colon cancer; report of a case.

A 73-year-old man had sigmoidectomy for sigmoid colon cancer in December 2001. Although he was followed regularly with chemotherapy, his serum carcinoembryonic antigen (CEA) increased on August 2002. Abdominal computed tomography and magnetic resonance imaging showed a right adrenal mass and no other abnormality. The preoperative diagnosis was a solitary adrenal metastasis from sigmoid colon cancer; the lesion was removed in September 2002. On pathology, adrenal metastasis was confirmed. Although the patient's serum CEA normalized soon thereafter, 12 months after adrenalectomy, the CEA again increased; the patient had local recurrence of the resected adrenal lesion and liver metastasis. Therefore, the patient was given systemic chemotherapy, but his condition deteriorated, and he died 38 months after adrenalectomy. Adrenal metastasis from colorectal cancer is not unusual; however, a solitary metastasis is rarely found and resected surgically. As surgical treatment of the metastatic lesion could improve patients' prognosis to some extent if it is detected early, the possibility of adrenal metastasis should be kept in mind when colorectal cancer patients are followed.

Downregulation of prostaglandin E receptor subtype EP3 during colon cancer development.

BACKGROUND AND AIMS: Involvement of prostaglandin E(2) (PGE(2)) receptors EP(1), EP(2), and EP(4) in the formation of aberrant crypt foci (ACF) and/or intestinal polyps has been suggested. In contrast, EP(3) appears to have no influence on the early stages of colon carcinogenesis. In the present study, we examined expression of PGE(2) receptor subtypes EP(1), EP(2), EP(3), and EP(4) in normal colon mucosa and colon cancers, and assessed the contribution of EP(3) to colon cancer development. METHODS: mRNA expression of PGE(2) receptor subtypes EP(1), EP(2), EP(3), and EP(4) in normal colon mucosa and colon cancers in azoxymethane (AOM) treated mice and rats, and in humans, were examined by reverse transcription-polymerase chain reaction (RT-PCR), quantitative real time RT-PCR, and immunohistochemical analyses. Evaluation of the role of EP(3) was performed by intraperitoneal injection of AOM, using EP(3) receptor knockout mice. Effects of EP(3) receptor activation on cell growth of human colon cancer cell lines were examined using ONO-AE-248, an EP(3) selective agonist. Moreover, EP(3) expression in colon cancer cell lines was analysed with or without 5-aza-2'-deoxycytidine (5-aza-dC) treatment. RESULTS: Expression levels of EP(1) and EP(2) mRNA were increased in cancer tissues. EP(4) mRNA was constantly expressed in normal mucosa and cancers. In contrast, expression of EP(3) mRNA was markedly decreased in colon cancer tissues, being 5% in mice, 9% in rats, and 28% in humans compared with normal colon mucosa, analysed by quantitative real time RT-PCR. Immunohistochemical staining demonstrated the rat EP(3) receptor protein to be expressed in epithelial cells of normal mucosa and some parts of small carcinomas but hardly detectable in large carcinomas of the colon. Colon cancer development induced by AOM in EP(3) receptor knockout mice was enhanced compared with wild-type mice, with a higher incidence of colon tumours (78% v 57%) and mean number of tumours per mouse (2.17 (0.51) v 0.75 (0.15); p<0.05). Expression of EP(3) mRNA was detected in only one of 11 human colon cancer cell lines tested. Treatment with 5 microM of an EP(3) selective agonist, ONO-AE-248, resulted in a 30% decrease in viable cell numbers in the HCA-7 human colon cancer cell line in which EP(3) was expressed. Treatment with 5-aza-dC restored EP(3) expression in CACO-2, CW-2, and DLD-1 cells but not in WiDr cells, suggesting involvement of hypermethylation in the downregulation of EP(3) to some extent. CONCLUSION: The PGE(2) receptor subtype EP(3) plays an important role in suppression of cell growth and its downregulation enhances colon carcinogenesis at a later stage. Hypermethylation of the EP(3) receptor gene could occur and may contribute towards downregulating EP(3) expression to some extent in colon cancers.

Molecular cloning and functional characterization of bottlenose dolphin (Tursiops truncatus) tumor necrosis factor alpha.

Bottlenose dolphin tumor necrosis factor alpha (doTNF-alpha) cDNA was cloned by reverse transcription polymerase chain reaction (RT-PCR) and the nucleic and deduced amino acid sequences were determined. The sequence of the cDNA clones shows that doTNF-alpha has an open reading frame of 699bp encoding 233 amino acids. The nucleic acid sequence of doTNF-alpha indicates 90, 88, 87, and 79% similarity with the cattle, pig, human, and mouse TNF-alpha gene, respectively. Based on the analysis of human and mouse TNF-alpha molecules, doTNF-alpha is processed to a mature protein with 157 amino acids. The 233 amino acids precursor has a hydrophobic region that could serve as a transmembrane domain. The recombinant doTNF-alpha expressed in Escherichia coli as a glutathione S-transferase fusion protein reacted with anti-human TNF-alpha antibody and exerted cytotoxity to the TNF-alpha sensitive murine cell line L929.

Apoptosis induction by epigallocatechin gallate involves its binding to Fas.

Epigallocatechin gallate (EGCG) is known to induce apoptosis in various types of tumor cells, but the precise mechanism by which EGCG induces apoptosis remains to be elucidated. The Fas-Fas ligand system is one of the major pathways operating in the apoptotic cascade. The aim of this study was to examine the possibility that EGCG-binding to Fas triggers the Fas-mediated apoptosis. The EGCG treatment of human monocytic leukemia U937 cells resulted in elevation of caspase 8 activity and fragmentation of caspase 8. The DNA ladder formation caused by the EGCG treatment was inhibited by the caspase 8 inhibitor. These findings suggested the involvement of the Fas-mediated cascade in the EGCG-induced apoptosis in U937 cells. Affinity chromatography revealed the binding between EGCG and Fas. Thus, the results suggest that EGCG-binding to Fas, presumably on the cell surface, triggers the Fas-mediated apoptosis in U937 cells.

A fibronectin-binding protein from rice bran with cell adhesion activity for animal tumor cells.

A rice bran 57-kDa protein was isolated by affinity chromatography with fibronectin immobilized on agarose. This fibronectin-binding protein designated as RB-57 had an amino-terminal amino acid sequence identical with that of a putative mature form of rice hydroxyproline-rich glycoprotein. A distinct feature of the amino acid composition of RB-57 was the high contents of hydroxyproline and proline representing about 45% of the total amino acids. The sugar analysis indicated that arabinose represented 46.8% of the total carbohydrates. RB-57 showed cell adhesion activity for murine Lewis lung carcinoma cells. The result suggests that RB-57 may play a role in plant cell adhesion, although cell adhesion-promoting activity for plant cells remains to be tested.

Decreased expression of c-myc family genes in thymuses from myasthenia gravis patients.

The thymus is a critical organ for the elimination of autoreactive T cells by apoptosis. We studied the expression of apoptosis-associated genes, bcl-xL, bad, caspase-3, and c-myc family genes in myasthenia gravis (MG) thymuses. We observed that the mRNA levels of myc family genes, c-myc and max, were markedly reduced in MG thymuses. These results indicate that c-myc-mediated signaling is abnormal in MG thymuses. The levels of molecules whose expressions are associated with myc, such as STAM, prothymosin-alpha, and NFkappaB, were also analyzed.

Spontaneous rupture of the thoracic aorta.

BACKGROUND: Spontaneous rupture of the thoracic aorta without trauma, aneurysm, or dissection is an extremely rare but catastrophic disorder. Two cases of spontaneous aortic rupture are presented, both treated surgically with satisfactory results. METHODS: A review of the English literature found 16 patients with the diagnosis of spontaneous rupture of the thoracic aorta from 1961 through 1998. Eighteen reported cases, including the 2 cases presented herein, are reviewed. RESULTS: The representative clinical picture is one of a middle-aged hypertensive patient with acute chest pain and collapse, with imaging modalities demonstrating hemopericardium, hemomediastinum, or hemothorax. According to the reported experiences, aortography was accurate for identifying the rupture site although the findings were sometimes subtle. Misdiagnosis or nonsurgical management resulted in the patient's death. All 8 patients who did not undergo aortic repair died within 3 weeks after the onset, whereas 9 of 10 patients who underwent surgical aortic repair survived. CONCLUSIONS: For patients with a definitive or possible diagnosis of spontaneous rupture of the thoracic aorta, prompt operation is imperative through an optimal surgical approach to identify and repair the rupture site with appropriate circulatory support.

Prolonged activation of the hypothalamus-pituitary-gonadal axis in a child with X-linked adrenal hypoplasia congenita.

X-linked adrenal hypoplasia congenita (AHC) is a rare developmental disorder of the human adrenal cortex that is caused by a mutation of the DAX-1 gene, a member of the nuclear hormone receptor superfamily. Hypogonadotrophic hypogonadism is frequently associated with this disease and the DAX-1 mutation is known to impair gonadotrophin production by acting at both the hypothalamic and pituitary levels. However, three recent studies reported that the hypothalamic-pituitary-gonadal axis was active in six infants with AHC, suggesting that a difference exists in the central regulation of hypothalamic-pituitary-gonadal activity between infant boys and pubertal boys. To determine the effect of the DAX-1 gene mutation on the axis in early childhood, we measured testosterone, LH, and FSH and performed LH-releasing hormone tests on a boy with AHC from birth to 3 years of age. Surprisingly, our findings showed that the axis was active from the infantile period to 3 years of age. This delayed initiation of the prepubertal pause, or prolonged activation of the axis, indicates that the DAX-1 gene is related to the control mechanism of the prepubertal restraint of gonadotrophin secretion.

Isolation of a 41-kDa protein with cell adhesion activity for animal tumor cells from the mushroom Hypsizigus marmoreus by affinity chromatography with type IV collagen immobilized on agarose.

A type IV collagen-binding protein of 41 kDa was isolated from the mushroom Hypsizigus marmoreus and the protein was designated as HM41. The Western blotting analysis with anti-HM41 antibodies demonstrated that HM41 was unrelated to HM23, which had been shown to have an affinity for type IV collagen. The microsequence analysis of the membrane-blotted peptides generated by fragmentation with cyanogen bromide showed no homologous proteins reported. HM41 had cell adhesion-promoting activity for murine Lewis lung carcinoma LL2 cells and human fibrosarcoma HT1080 cells. These results indicate that HM41 is a hitherto undescribed fungus protein that can interact both with animal extracellular matrix protein type IV collagen and with animal tumor cells.

Quantification of telomerase activity in sporadic colorectal carcinoma: association with tumor growth and venous invasion.

BACKGROUND: Activation of telomerase, a ribonucleoprotein enzyme complex that synthesizes telomere repeats, is associated with acquisition of unlimited cellular proliferation and is commonly detected in human cancer. Measurement of telomerase activity (TA) may provide important information as a diagnostic marker or a prognostic indicator. The authors studied the quantification of TA and assessed its utility as a prognostic marker in sporadic colorectal carcinoma. METHODS: Sixty surgical specimens, including 30 specimens of cancer tissue and 30 specimens of corresponding normal colorectal mucosa, were examined. TA was measured by a fluorescence-based telomeric repeat amplification protocol assay. The authors determined the telomerase index (TI = log (A-B), where A represented TA of cancer tissues and B represented TA of normal mucosa) and examined the relation between TI and clinicopathologic factors using the Student t test, analysis of variance, the Chi-square test, and the Fisher PLSD as a post hoc test. RESULTS: TA of cancer and corresponding normal mucosa was 51.87+/-27.38 and 7.14+/-9.85, respectively (P<0.0001). The cutoff value was determined to be 26 in a receiver operating characteristic study, with 90% sensitivity, 96.7% specificity, and 96.4% positive predictive value. TI was closely correlated with depth of invasion (P = 0.0129) but not with age, gender, histologic type, location, lymph node metastasis, lymphatic infiltration, or Dukes stage. There was a significant difference in TI between tumors with and without venous invasion (P = 0.0003). Four of five tumors with synchronous liver metastasis showed high TI (1.555

[Diagnosis of ulcerative colitis and the biopsy method].

The biopsy specimen should be oriented flat on a ground glass slide or on a piece of filter paper so that the submucosa is on the slide and the mucosa uppermost to allow proper histopathological interpretation. It is important to perform the biopsy based on a full knowledge of features of inflammatory diseases of the large intestine. Biopsies should be taken from apparently normal and abnormal mucosa. It is necessary to observe the mucosa of the colon carefully for any mass lesions to find dysplasia or cancer complicating the ulcerative colitis. Any suspicious areas should be liberally biopsied. Biopsies should also be performed at random, because of possible existence of dysplasia or cancer which is not visible at endoscopy. This occurs especially in the rectum; biopsies can also be performed after inversion of the endoscope.

Current trends in restorative proctocolectomy: introduction of an ultrasonically activated scalpel.

PURPOSE: We evaluated the usefulness of an ultrasonically activated scalpel (Harmonic Scalpel) for mucosal proctocolectomy and ileal J-pouch-anal anastomosis. METHODS: Seventy-four patients with ulcerative colitis (70 patients) and familial adenomatous polyposis (4 patients) underwent mucosectomy using the Harmonic Scalpel since 1997. We compared the clinical and functional results with those of the monopolar electrocoagulator (forceps coagulation technique, 86 patients with colitis and 7 with polyposis). RESULTS: We performed graduated mucosal proctectomy by using the Harmonic Scalpel. The operative time (Harmonic Scalpel, 42 minutes vs. forceps coagulation technique, 85 minutes) and blood loss (Harmonic Scalpel, 33 ml vs. forceps coagulation technique, 86.8 ml) were significantly reduced by this method. The Harmonic Scalpel enabled restorative proctocolectomy by the synchronous approach within three hours. The functional results and complications were not significantly different between the two groups. CONCLUSION: The Harmonic Scalpel shortened the operative time, decreased blood loss, and was useful for restorative proctocolectomy in our study.

Inhibitory effects of tetragalloylglucose and digalloylhamamelose on adhesion and in vitro invasion of mouse lung carcinoma cells.

Tetragalloylglucose (TgG) and digalloylhamamelose (DgH) were found to inhibit adhesion to and invasion through Matrigel of mouse Lewis lung carcinoma LL2-Lu3 cells, which are highly metastatic. TgG inhibited matrix metalloproteinases (MMPs) from the tumor cells like (-)-epigallocatechin gallate, whereas DgH did not. These results suggest that TgG and DgH inhibit tumor cell invasion by inhibiting MMPs and/or cell adhesion of the tumor cells.

Transanal mucosectomy using an ultrasonically activated scalpel for ulcerative colitis.

We describe herein our technique of using an ultrasonically activated scalpel (Harmonic Scalpel [HS]) to simplify the procedure of anorectal mucosectomy for ulcerative colitis (UC). This technique was successfully employed to perform restorative proctocolectomy and ileoanal anastomosis (IAA) in ten patients with UC. By using the HS, thermal injury of the internal anal sphincter was avoided during mucosectomy, and we were able to dissect the inflamed mucosa sharply without causing bleeding. HS reduced the operative time and blood loss compared with the traditional forceps-coagulation technique. These results indicate that the introduction of this new scalpel will facilitate an increase in the number of cases of mucosectomy for IAA.

Cell adhesion activity for murine carcinoma cells of a wheat germ 55-kDa protein with binding affinity for animal extracellular matrix proteins.

A wheat germ 55-kDa protein was isolated by affinity chromatography with Matrigel immobilized on agarose, followed by preparative gel electrophoresis. This Matrigel-binding protein designated as WG-55 had an amino-terminal amino acid sequence which is identical to that of a putative mature form of wheat storage protein Gbl 1. WG-55 reacted with concanavalin A, indicating its glycoprotein nature as expected from the amino acid sequence of Gbl 1. As expected, similarly, WG-55 exhibited RGD-dependent cell adhesion activity for murine carcinoma cells. These data suggest that WG-55 or mature Gbl 1 protein may play a role in plant cell adhesion.

Presynaptic inhibition of GABA(B)-mediated synaptic potentials in the ventral tegmental area during morphine withdrawal.

Opioids increase the firing of dopamine cells in the ventral tegmental area by presynaptic inhibition of GABA release. This report describes an acute presynaptic inhibition of GABAB-mediated IPSPs by mu- and kappa-opioid receptors and the effects of withdrawal from chronic morphine treatment on the release of GABA at this synapse. In slices taken from morphine-treated guinea pigs after washing out the morphine (withdrawn slices), a low concentration of a mu receptor agonist increased, rather than decreased, the amplitude of the GABAB IPSP. In withdrawn slices, after blocking A1-adenosine receptors with 8-cyclopentyl-1, 3-dipropylxantine, mu-opioid receptor activation inhibited the IPSP at all concentrations and increased the maximal inhibition. In addition, during withdrawal, there was a tonic increase in adenosine tone that was further increased by forskolin or D1-dopamine receptor activation, suggesting that metabolism of cAMP was the source of adenosine. The results indicate that during acute morphine withdrawal, there was an upregulation of the basal level of an opioid-sensitive adenylyl cyclase. Inhibition of this basal activity by opioids had two effects. First, a decrease in the formation of cAMP that decreased adenosine tone. This effect predominated at low mu receptor occupancy and increased the amplitude of the IPSP. Higher agonist concentrations inhibited transmitter release by both kinase-dependent and -independent pathways. This study indicates that the consequences of the morphine-induced upregulation of the cAMP cascade on synaptic transmission are dependent on the makeup of receptors and second messenger pathways present on any given terminal.