Prevalence of human papillomavirus in oral gargles and tonsillar washings.

OBJECTIVES: Human papillomavirus (HPV) infection drives carcinogenesis in the oropharynx. No standard sampling or HPV detection methods for evaluating oropharyngeal HPV infection exist. The prevalence of oral HPV infection in Japan is unknown. MATERIALS AND METHODS: We examined 435 healthy Japanese individuals to address whether adding tonsillar washing to oral gargling would improve HPV detection. We compared HPV assessment using GENOSEARCH HPV31 versus nested PCR and direct sequencing. Associations between HPV infection and demographic and behavioral characteristics were examined. RESULTS: Most participants who were HPV-positive based on oral gargles were also HPV-positive based on tonsillar washings: 11 (64.7%) of 17 on nested PCR and 12 (70.6%) of 17 on GENOSEARCH HPV31. Although HPV infection was more prevalent in oral gargles followed by tonsillar washings than in oral gargles alone, the difference was not statistically significant (nested PCR, 4.8% vs. 3.9%, P = 0.46; GENOSEARCH HPV31, 5.3% vs. 3.9%, P = 0.33). The overall agreement between nested PCR and GENOSEARCH HPV31 was 98.6%, with 76.0% positive agreement. The overall prevalence of oral HPV infection in Japan was 5.7% (95% confidence interval, 3.9-8.3%). Men had a significantly higher prevalence of oral HPV infection than women (8.3% vs. 2.6%, P = 0.02). Infection increased with number of lifetime sexual partners (P < 0.001 for trend). CONCLUSION: The oropharynx is probably the major source of HPV-infected cells in oral gargles. Oral gargling could be a standard sampling method for evaluating oropharyngeal HPV infection. GENOSEARCH HPV31 could be an option for oral HPV detection.

Metabolic tumor volume of metastatic lymph nodes and survival after total laryngectomy in laryngeal and hypopharyngeal cancer.

OBJECTIVES: The prognostic value of metabolic tumor volume (MTV) in locally advanced laryngeal or hypopharyngeal cancer is established in the setting of chemoradiotherapy, while it remains unknown in the setting of upfront total laryngectomy. MATERIALS AND METHODS: We retrospectively analyzed 88 patients receiving total laryngectomy and neck dissection, using Cox regression models. RESULTS AND CONCLUSION: Variables related to metastatic lymph node were associated with overall survival, whereas those related to primary tumor were not. In multivariable models, MTV of metastatic lymph nodes (N-MTV) as a continuous variable (Akaike's information criterion (AIC), 277.5) was equivalent to pathological nodal status (AIC, 278.2; P = 0.40), and superior to pathological nodal classification as an ordinal variable (AIC, 281.4; P < 0.05) in ability of predicting death. The risk of death was increased by 1.2-fold (95% confidence interval (CI), 1.0-1.4; P = 0.03) every 10-ml increment of N-MTV, while patients with pN+ disease were at a higher risk of death by 2.9-fold (95% CI, 1.0-12.2; P < 0.05) compared with patients with pN0 disease. Using recursive partitioning analysis (RPA), we classified the patients as having a low, intermediate, or high risk of death on the basis of N-MTV and extranodal extension (ENE). This RPA classification system exhibited greater concordance with overall survival than the classification considering pathological nodal status and ENE (AIC, 275.8 versus 281.4; P = 0.02). In the setting of upfront total laryngectomy, N-MTV is a critical predictor of mortality. A staging system in which N-MTV is incorporated may better inform adjuvant treatment decisions.

MicroRNAs and liver function.

MicroRNAs (miRNAs) are small, noncoding RNA molecules that regulate gene expression post-transcriptionally through complementary base pairing with thousands of messenger RNAs. Although the target genes and precise biological functions of individual miRNAs remain largely unknown, miRNAs are speculated to play important roles in diverse biological processes in both normal and pathological states. The liver is a vital organ that plays major roles in a number of physiological functions. Recent advances in the study of liver miRNAs using gene-modified mice or in vivo nucleic acid delivery to overexpress specific miRNAs or inhibit miRNA functions have revealed the crucial biological roles of individual miRNAs in physiologically essential liver functions in vivo. Because miRNA-based strategies are being applied to clinical therapeutics, the importance of precise knowledge of miRNA functions cannot be underestimated, not only from a scientific point of view, but also from a clinical perspective to make the most of such drugs and avoid unexpected harmful effects. The aims of this review are to describe current knowledge regarding both known and as-yet-undiscovered molecular aspects of the biological roles of miRNAs in the liver, with a special emphasis on lipid, glucose, drug, and iron metabolism as vital functions of the liver as well as important therapeutic targets.

Clinical significance of heparin-binding epidermal growth factor-like growth factor in peritoneal fluid of ovarian cancer.

Epidermal growth factor receptor (EGFR) has been implicated in tumour growth and extension of ovarian cancer. Peritoneal fluid in ovarian cancer patients contains various growth factors that can promote tumour growth and extension. In order to investigate the clinical significance of EGFR ligands as activating factors of ovarian cancer, we examined the cell proliferation-promoting activity and the level of EGFR ligands in peritoneal fluid obtained from 99 patients. Proliferation-promoting activity in peritoneal fluid from 63 ovarian cancer patients (OVCA) was much higher than peritoneal fluid from 18 ovarian cyst patients (OVC) and 18 normal ovary patients (NO), and the activity was suppressed only by antibodies against EGFR or heparin-binding epidermal growth factor (HB-EGF). A large difference was observed in the level of EGFR ligands between HB-EGF and TGF-alpha or amphiregulin. The concentration of HB-EGF in OVCA significantly increased compared to that in OVC or NO (P<0.01). No significant difference in the concentration of TGF-alpha and amphiregulin was found between the OVCA and NO or OVC groups. In peritoneal fluid, HB-EGF is sufficiently elevated to activate cancer cells even at an early stage of OVCA. These results suggested that HB-EGF in peritoneal fluid might play a key role in cell survival and in the proliferation of OVCA.

A role of mast cells for hepatic fibrosis in primary sclerosing cholangitis.

We encountered four patients with overt primary sclerosing cholangitis (PSC) which were histologically classified into stage 2 or 3. We examined the expression of stem cell factor (SCF), a ligand of c-kit, in injured bile ducts by immunohistochemistry, and mast cells were identified by immunohistochemistry using anti-HMCT (human mast cell tryptase) and anti-c-kit antibodies to clarify their relation with portal fibrosis coincident with destroyed bile ducts. SCF was detected in the epithelia of most bile ducts in PSC, and many HMCT- and c-kit-positive mast cells were found in portal tracts. Image analysis showed more significant numbers of c-kit-positive mast cells per area of portal tract in PSC than in chronic hepatitis C, and they might increase from stage 2 to 3. c-Kit-positive cells infiltrated into the portal tracts with SCF-positive destroyed bile ducts, and c-kit mast cells should be investigated in detail to make a role for portal fibrosis in PSC.

Preoperative diagnosis of fallopian tube cancer by imaging.

Primary cancer of the fallopian tube (FTC) is among the most unusual gynecologic malignancies and rarely is diagnosed correctly before surgery. The imaging results of eight patients with FTC and four with benign tubal disease were analyzed. FTCs were small cystic or solid masses that typically were shaped like a sausage, a snail, or a gourd, regardless of clinical stage.

Effect of melatonin on changes in hepatic antioxidant enzyme activities in rats treated with alpha-naphthylisothiocyanate.

We have reported that melatonin protects against alpha-naphthylisothiocyanate (ANIT)-induced acute liver injury in rats by preventing enhanced lipid peroxidation. Herein, we examine the effect of melatonin on hepatic antioxidant enzyme activities in rats with a single i.p. injection of ANIT (75 mg/kg body weight) in order to clarify the protective mechanism of the indoleamine against ANIT-induced acute liver injury. Rats received a single oral administration of melatonin (10 or 100 mg/kg body weight) at 12 hr after ANIT treatment. Hepatic Cu,Zn-superoxide dismutase (Cu,Zn-SOD), Mn-superoxide dismutase (Mn-SOD), catalase (CAT), Se-glutathione peroxidase (Se-GSH-Px), glutathione reductase (GSSG-R), and glucose-6-phosphate dehydrogenase (G-6-PDH) activities and reduced glutathione (GSH) concentration were determined 12 and 24 hr after ANIT treatment. ANIT-treated rats showed decreases in hepatic Cu,Zn-SOD and GSSG-R activities at 24 hr after treatment, transient increases in hepatic CAT and Se-GSH-Px activities at 12 hr, and no changes in hepatic Mn-SOD and G-6-PDH activities at 12 or 24 hr. Only the high dose of melatonin attenuated the decrease in hepatic Cu,Zn-SOD activity, while both doses of the indoleamine almost completely attenuated the decrease in hepatic GSSG-R activity. Neither dose of melatonin affected hepatic CAT, Se-GSH-Px, and G-6-PDH activities. ANIT-treated rats showed an increase in hepatic GSH concentration at 24 hr after treatment. Neither dose of melatonin affected the increase in hepatic GSH concentration. These results indicate that orally administered melatonin prevents decreases in Cu,Zn-SOD and GSSG-R activities in the liver of ANIT-treated rats, and suggest that the indoleamine may protect against ANIT-induced acute liver injury by attenuating the disruption of hepatic antioxidant defense systems.

Effectiveness of a traditional Chinese medicine, Wulingsan, in suppressing the development of nephrocalcinosis induced by a high phosphorus diet in young rats.

The development of nephrocalcinosis in rats fed a high phosphorus diet, and the effectiveness of the Chinese traditional medicine, Wulingsan, and its components (Poria, Alismatis Rhizoma, Atractylodis Rhizoma, Cinnamomi Ramus, Polyporus) in suppressing the development of calcinosis were studied. The rats were fed a high phosphorus diet (1.5% P) supplemented with Wulingsan or its individual components (0.5 g/kg body weight) as separate experimental groups for a 2-week period. Upon histological observation by light microscopy and electron microscopy, signs of nephrocalcinosis were observed in almost all areas of the kidney. Calculi, consisting mainly of needle-shaped crystals of hydroxyapatite, were observed in the proximal tubules, in the collecting ductal lumina, and in the mitochondria of the proximal tubular cells and the interstitial cells. X-ray microanalysis revealed that the calculi were composed of hydroxyapatite (Ca and P). In the group fed the diet supplemented with Wulingsan, the severity of calcinosis in the corticomedullary junction was only slight. In all groups fed individual components of Wulingsan, the severity of calcinosis was almost the same as that in the group fed the high phosphorus diet (1.5% P). Wulingsan suppressed the development of calcinosis in rats fed the high phosphorus diet supplemented with this Chinese medicine, whereas its individual components alone had no effect. The process of calcinosis and the mechanism responsible for the activity of this Chinese medicine in the suppression of calcinosis are discussed.

Ultrastructural study of the alveolar lining and the bronchial mucus layer by block staining with oolong tea extract: the role of various surfactant materials.

To investigate the roles of various surfactant materials in the lung, we examined rat lung fixed with a mixture of 0.2% oolong tea extract (which contains various polyphenols) and 2.5% glutaraldehyde by electron microscopy. We also measured the surface tension of various isolated surfactant fractions, with a Wilhelm balance. A fraction containing lamellar bodies and a fraction containing lattice-like structures were obtained by discontinuous sucrose gradient centrifugation. From the 15 000 g supernatant, a fraction containing electron-dense amorphous materials was obtained as the 105 g precipitate. The fraction containing lamellar bodies and the fraction containing electron-dense amorphous materials displayed surfactant activity, but the fraction containing lattice-like structures did not. The lamellar bodies were found to be gradually secreted from type II epithelial cells while self-decomposition occurred. The alveolar lining layer had the form of a thin film consisting of electron-dense amorphous materials. These electron-dense amorphous materials may be precursors of the phospholipid film, which exhibits surfactant activity, on the alveolar surface. Lattice-like structures and lamellar bodies were found to be located in the interalveolar pores. The interalveolar pores were filled with surfactant, and this indicated that they do not play a role in the collateral ventilation of the alveoli. It may be that the lattice-like structures serve to connect lung epithelial cells in the interalveolar pores, as well as serving as the basis for the formation of the alveolar ducts. The bronchial mucus layer, which consisted of fibrillar mucous materials, was not divided into an epiphase and a hypophase. A surfactant, in the form of an osmiophilic surface film and some trilaminar materials, was found to cover the mucus layer. Thus, it is possible that the lamellar bodies could be transformed into various surfactant materials, which then serve bronchial or alveolar mechanical and physiological functions.

A child with adrenocortical adenoma accompanied by congenital hemihypertrophy: report of a case.

We report herein the findings of a 7-year-old male child with a ruptured adrenocortical adenoma and congenital hemihypertrophy which was incidentally detected after suffering a trauma. A review of 21 pediatric cases of adrenocortical neoplasms in the literature was made. The patient showed precocious puberty such as pubis and advanced bone age, but an endocrinological examination revealed no definite abnormalities. The right adrenal tumor with hematoma was resected after these evaluations. Adrenocortical adenoma is considered to occur more frequently in female children. However, the incidence of adrenocortical tumors accompanied by congenital hemihypertrophy does not differ between males and females. The outcomes were relatively good, although the observation periods were short in some patients. A large number of patients presented with a tumor and hemihypertrophy on the same side. This finding is of interest when considering the possible association between hemihypertrophy of the organs and tumor proliferation. However, their association in terms of development was unclear. It is necessary for patients with hemihypertrophy to have regular examinations for the possible development of malignant tumors, especially in the kidney, adrenal gland, and liver.

Specific role of T and Tn tumor-associated antigens in adhesion between a human breast carcinoma cell line and a normal human breast epithelial cell line.

The possibility that tumor-associated antigens T and Tn act as adhesion molecules between normal and malignant breast epithelial cells at the early stages of recognition in the metastatic pathway was examined in vitro. The adhesive specificity of the antigens was assessed by means of in vitro adhesion tests between a carcinomatous breast cancer cell line (ZR75-30) and a normal epithelial breast cell line (HLB100) using both monoclonal antibodies and lectins specific as well as nonspecific for each antigen. Adhesion assay was performed using monolayers of the normal cell line prepared on plastic culture plates and the tumor cell line labeled with a fluorescent dye as a probe. The adhesion between the two cell types occurred with significant specificity via T and Tn antigens (P<0.001), and was temperature-dependent. The results suggest that at the early stages of recognition by tumor cells in the metastatic process, T and Tn antigens play a role as adhesion molecules between the tumor cells and adjacent normal cells.

Effects of oral repetitive loading of disopyramide on acute-onset atrial fibrillation with concurrent monitoring of serum drug concentration.

We investigated the efficacy and safety of oral repetitive loading of disopyramide, for the termination of acute-onset (i.e., therapy started within 2 days after the onset of palpitations) atrial fibrillation (AF) in 96 consecutive patients, with concurrent monitoring of the serum concentration of this agent in fifteen of the patients. Outpatients with AF verified by standard electrocardiogram (ECG) were hospitalized and received disopyramide (200 mg) every 4-6 h, with a maximal dose of 800 mg daily, until the termination of AF under ECG monitoring was obtained. Conversion to sinus rhythm occurred within the first day of treatment in 88 patients (92%), on the second day of treatment in six patients (6%), and on the third and fifth days of treatment in the remaining two patients. No major adverse effects, such as hypotension, congestive heart failure, proarrythmic events or systemic embolism were noted. The serum levels of disopyramide evaluated in fifteen of the enrolled patients were found to be maintained within the therapeutic range throughout the treatment period. In spite of the absence of any placebo-controlled group in this study, these findings suggest that repetitive oral loading of disopyramide (200 mg) with an interval of 4-6 h is effective and safe for the termination of acute-onset AF under a stable therapeutic serum drug concentration, hence offering the possibility of self-medication for patients with episodic AF.

Selection of human ovarian carcinoma cells with high dissemination potential by repeated passage of the cells in vivo into nude mice, and involvement of Le(x)-determinant in the dissemination potential.

Cells of the human tumor cell line RMG-1, derived from a clear-cell adenocarcinoma of the ovary, were injected intraperitoneally into nude mice, and the cells obtained from the tumor nodules in the mesenterium were found to form a larger number of, and larger-sized, tumor nodules than the original RMG-1 cells. The RMG-1-h cells, transferred into culture from the tumor nodules after a 4th in vivo passage, showed a dissemination potential as high as that of cells disseminating directly from the tissues, and exceedingly higher than that of RMG-1 cells. To assess the molecular bases of the different biological properties of RMG-1 and RMG-1-h cells, we compared the content and expression of various carbohydrate antigens in both cells. The chromosomal profile of RMG-1-h cells revealed their human origin and was identical to that of the original RMG-1 cells. In contrast to the broad histogram for the Le(x)-bearing cells among RMG-1 cells in flow cytometry, the weakly and moderately positive cells toward anti-Le(x) antibody were found to be eliminated from the histogram for the RMG-1-h cells, resulting in the enrichment of cells strongly expressing Le(x), which may account for the high dissemination potential. In addition, the adhesion of RMG-1 cells to mesothelial cells was found to be significantly inhibited by pretreatment of the cells with anti-Le(x) antibody, indicating Le(x)-mediated cell-to-cell interaction between ovarian cancer cells and mesothelial cells. By TLC-immunostaining, two Le(x)-glycolipids, III3Fuc alpha-nLc4Cer and V3Fuc alpha-nLc6Cer were detected in both RMG-1 and RMG-1-h cells, and their total concentrations were not significantly different from each other. However, the hydrophobic moieties of Le(x)-glycolipids in RMG-1-h cells were different from those in RMG-1 cells, suggesting that a difference in the structure of the hydrophobic moieties of Le(x) is partly involved in the enhanced reactivity of RMG-1-h cells toward anti-Le(x) antibody. Thus, the high dissemination potential of ovarian cancer cells was shown to be mediated by the Le(x)-determinant and the Le(x)-bearing cells are enriched by repeated in vivo passage of the cells into nude mice.

A role of SH-compounds in maturation of melanosomes in cells cultured from nevus Ota.

A role of SH-compounds such as cysteine and glutathione in melanogenesis in dermal melanocytes cultured from Ota's nevus tissue was demonstrated in relation to another substrate, dihydroxyphenylalanine (DOPA). Chemical analysis of eumelanin and pheomelanin was performed in addition to the conventional electron microscopic observation. Supplements of the culture medium with each of these compounds separately for two weeks gave rise to the formation of pre-pheomelanosomes and secondary lysosomes or myelinosome-like inclusions. When DOPA and glutathione were added to the medium together, the maturation of melanosomes was promoted. This was proven by the increase in electron-density of pre-melanosomes observed as well as by the content of pheomelanin and eumelanin. However, mature melanosomes were not formed when each of these chemicals was added to the medium individually for the same periods. The melanosome maturation seemed to occur via a process involving secondary lysosomes or myelinosomes, in which more electron-dense particles accumulated in the presence of both reagents. The pheomelanosomal process was also observed, but typical eumelanosome-related processes were not observed in this culture system.

Characterization of melanosomes and melanogenesis in cells cultured from Ota's nevus.

Cells from dermis and epidermis of Ota's nevus lesions were cultured to obtain the pigment producing cells that are specific epidermal and dermal melanocytes of nevus Ota. The following media were used: Eagle's minimum essential medium, fortified or not fortified with cholera toxin and phorbol ester. Both epidermal melanocytes and dermal melanocytes grew in the Eagle's minimum essential medium fortified with cholera toxin and phorbol ester. The melanocytes were characterized by 14C-tyrosine uptake and by identification of melanosomes and status of melanogenesis in comparison with those reported in dysplastic nevi and melanomas, as well as those found in Ota's nevus tissue in situ. Eumelanosomes, pheomelanosomes and secondary lysosomal melanosomes appeared in epidermal melanocytes. Pheomelanosomes and melanolysosomes, or myelinosiderosomes appeared predominantly in the dermal melanocytes. These melanosomes seemed to be related to the peculiar skin color of nevus Ota. Some of the abnormal types of melanosomes were very similar to those found in dysplastic nevi and melanomas.

Phase II study of a new combined primary chemotherapy regimen, intravenous methotrexate and vincristine and intraarterial adriamycin and cisplatin, for locally advanced urinary bladder cancer: preliminary results.

A phase II study of a new combination therapy was performed using intraarterial (i.a.) cisplatin and Adriamycin in combination with i.v. methotrexate and vincristine for 27 patients with invasive urinary bladder carcinoma of stages T2-3NOMO, and the therapeutic effects were assessed. Methotrexate (20 mg/m2) was given i.v. on days 1,15, and 22, and vincristine (0.7 mg/m2) was injected i.v. on day 2 before i.a. infusion therapy and on days 15 and 22. The i.a. chemotherapy was performed after both superior gluteal arteries had been embolized using 3- or 5-mm stainless-steel coils. A mixture of cisplatin (50-70 mg/m2) and Adriamycin (20 mg/m2) was infused i.a. via both internal iliac arteries over a period of 20-30 min. Angiotensin II (mean dose, 21 micrograms) was simultaneously infused i.a. in 15 of 27 patients. In 24 of the 27 patients, at least 2 cycles of full-dose chemotherapy were completed. The dose was decreased in the remaining 3 patients because of their poor health status and advanced age. Among the 27 patients, 9 and 14 had complete (CR) and partial responses (PR), respectively; 3 manifested no change (NC), and 1 had progressive disease (PD). The objective response rate (CR+PR) was 85.2%. Among the 27 patients staged T2-3 NOMO, 6 (CR, 1; PR, 5) underwent total cystectomies and 18 (CR, 8; PR, 8; NC, 2) had transurethral resection of a bladder tumor (TUR-Bt) or partial resections following chemotherapy. The remaining 3 diminished-dose patients had no surgery. Of the 27 patients, 22 were alive after a median follow-up period of 21+ (range, 7-48+) months. No significant side effect was observed except for lower extremity paresthesias in 5 patients (18.5%). These results point to the effectiveness of this therapy and to the possibility of urinary bladder preservation in patients with invasive, advanced urinary bladder cancers.

Two distinct patterns of peritoneal involvement shown by in vitro and in vivo ovarian cancer dissemination models.

We established an in vitro peritoneal dissemination model using six ovarian cancer cell lines and cultured mesothelial cells. Ovarian cancer cells were classified into two types, invasive or adhesive, on the basis of their interaction with the mesothelial cell monolayer. The ovarian cancer cell lines derived from mucinous cystadenocarcinoma, poorly differentiated adenocarcinoma and undifferentiated carcinoma, which belonged to the invasive type, began to invade beneath the mesothelial monolayer from several hours after seeding in vitro, expelling the mesothelial cells at the periphery and forming colonies directly on the dish surface. On the other hand, cancer cell lines of clear cell carcinoma, which belonged to the adhesive type, showed colony formation with adhesion on the mesothelial monolayer even 18 h after seeding. Invasive-type cell lines invaded into the mesothelial monolayer at various rates in vitro, and the degree of invasiveness showed good correlation with the degree of peritoneal dissemination in vivo after intraperitoneal injection of cancer cells into nude mice. Adhesive-type cells showed rather higher dissemination rates in vivo. Microscopic observation of in vivo peritoneal dissemination at one day after inoculation also revealed two patterns of peritoneal involvement similar to those in vitro. In the in vitro model, anti-integrin alpha 2- and beta 1-antibodies inhibited the infiltration of invasive-type cells into the mesothelial monolayer, but did not affect colony formation by adhesive-type cells on the monolayer, indicating that invasion by both cell types was mediated by different molecules. This in vitro model is thought to be useful for analysis of the molecular mechanisms of peritoneal dissemination.