Identification of the promoter region regulating the transcription of the REV7 gene.

REV7 is involved in various biological processes including DNA repair and mutagenesis, cell cycle regulation, gene transcription, and carcinogenesis. REV7 is highly expressed in adult testicular germ cells as well as several malignant tumors. REV7 expression levels are associated with prognosis in several human cancers, however, the mechanism of REV7 transcriptional regulation has not been elucidated. In this study, we characterized the promoter region of the REV7 gene. A luciferase reporter assay using the human germ cell tumor cell line NEC8 was utilized to examine the upstream genomic region of REV7 for transcriptional activity, and two transcriptional activation regions were identified. We determined a small genomic region important for transcriptional activation using site-directed mutagenesis; this region is shared by several putative binding motifs for transcription factors, including the cAMP-responsive element modulator (CREM), cAMP-response element binding protein (CREB), and B-lymphocyte-induced maturation protein-1 (BLIMP-1). Exogenous CREM and CREB expression had no effect on the transcriptional activity in NEC8 cells or the human embryonic kidney cell line HEK293T. In contrast, exogenous BLIMP-1 expression increased luciferase reporter activity in HEK293T cells but unexpectedly decreased activity in NEC8 cells. Chromatin immunoprecipitation analysis demonstrated that BLIMP-1 binds to the genomic region near the binding motif in the REV7 promoter. Additionally, BLIMP-1 overexpression promoted endogenous REV7 expression in HEK293T cells. These findings suggest that BLIMP-1 may be a putative transcriptional regulator of REV7 in mammalian cells.

Taste Disorders: Effect of Education in Patients With Breast Cancer Receiving Chemotherapy.

BACKGROUND: For patients with breast cancer treated with certain chemotherapy regimens, taste disorders associated with those regimens can negatively affect quality of life. OBJECTIVES: This study evaluated the effects of taste disorder-related education on meal satisfaction and sense of taste in Japanese women with breast cancer undergoing chemotherapy. METHODS: A sample of 53 newly diagnosed women with breast cancer scheduled for chemotherapy treatment were randomly assigned to the control or intervention (nurse-provided education about chemotherapy-associated taste disorders) group. Meal satisfaction and sense of taste were assessed using a visual analog scale. FINDINGS: The proportions of patients with meal dissatisfaction and impaired sense of taste were lower in the intervention group than in the control group. Although meal dissatisfaction and impaired sense of taste recovered in the intervention group two months after protocol completion, they did not recover in the control group. Providing education to women with breast cancer scheduled for chemotherapy treatment can affect patients' experience of treatment-associated taste disorders.

Influence of arm position and respiration technique during liver examinations on the detectability of mammary lesions.

During liver computed tomography (CT), scanning is performed with the raised arm position and an inhalation technique. However, for liver magnetic resonance imaging (MRI), the arms are placed at the sides of the body and an exhalation technique is used. This study was aimed at evaluating the effect that the patient's arm position and respiration technique had on the ability to detect mammary glands in the scan range to discover unexpected mammary lesions during the liver MRI examination. Liver MRI and CT images were compared for 337 female patients. More than half of the mammary glands were included in 97.3% of MRI, but in 4.7% of CT. No mammary lesions were found during CT, whereas seven were found during MRI. The mammary lesions are more likely to be detected when the patient places her arms at the sides of the body and uses the exhalation technique during liver MRI.

Spontaneous malignant myoid thymoma in an aged female Fischer 344 rat.

A whitish mass approximately 30 mm in diameter was noted in the anterior mediastinum of a 67-week-old female Fischer 344 rat. Histopathologically, two types of tumor cells were identified on the basis of morphologic features: epithelial tumor cells with a tubular or cord-like growth pattern and rhabdomyosarcomatous tumor cells characterized by the presence of cross-striations. Immunohistochemically, the epithelial tumor cells reacted positively for cytokeratin AE1/AE3, and some reacted positively for p63, which is expressed in normal thymic epithelial cells. The rhabdomyosarcomatous tumor cells stained positively for desmin, sarcomeric actin, and S-100 protein, which coincides with the stainability of normal thymic myoid cells. Since the tumor was also found to have malignant features such as high proliferative activity, cytologic atypia, and necrotic behavior, it was diagnosed as a malignant myoid thymoma. We believe that this is the first case report of such a tumor in a rodent.

A spontaneous myoepithelial carcinoma in the mammary gland of an aged female ICR (CD-1) mouse.

We report a female Crlj:CD1(ICR) mouse with a spontaneous mammary gland tumor composed of biphasic tumor cells, i.e., epithelioid and spindle-shaped myoepithelial cells. Macroscopically, a subcutaneous mass, approximately 3 cm in diameter was found in the lumbodorsal region. Histopathologically, the epithelioid cells proliferated in an alveolar or nest-like growth pattern, occasionally forming glandular-like structures. On the other hand, the spindle-shaped cells proliferated in a sarcomatous pattern. Normal mammary gland was observed in the vicinity of the tumor. Both types of tumor cells showed immunoreactivity for cytokeratin (wide spectrum screening), vimentin, S100, and p63. In addition, the epithelioid cells and spindle-shaped cells were immunopositive for glial fibrillary acidic protein and smooth muscle actin, respectively. Moderate atypia, high proliferative activity, massive necrosis, and partial infiltration to the surrounding tissues were also observed. We made a diagnosis of myoepithelial carcinoma, which is extremely rare in ICR mice.

Effects of a warm compress containing menthol on the tear film in healthy subjects and dry eye patients.

Menthol is thought to stimulate lacrimation via activation of cold-sensitive primary afferent neurons in the cornea. We evaluated a warm compress containing menthol as a potential treatment for dry eye by examining its effects on the tear film in healthy subjects (n = 20) and dry eye patients (n = 35). Disposable eyelid-warming steamers that either did (MH) or did not (HO) contain menthol were applied to one eye of each subject either once only for 10 min or repeatedly over 2 weeks. Single application of MH significantly increased tear meniscus volume (P = 8.6 × 10-5, P = 1.3 × 10-5) and tear film breakup time (P = 0.006, P = 0.002) as well as improved meibum condition in healthy subjects and dry eye patients, respectively. Repeated application of MH significantly increased tear meniscus volume (P = 0.004, P = 1.7 × 10-4) and tear film breakup time (P = 0.037, P = 0.010) in healthy subjects and dry eye patients, respectively. Repeated application of MH thus induced persistent increases in tear fluid volume and tear film stability in dry eye patients, suggesting that repeated use of a warm compress containing menthol is a potential novel treatment for dry eye disease.

Late Metabolic Acidosis Caused by Renal Tubular Acidosis in Acute Salicylate Poisoning.

A 16-year-old man was transferred to our emergency department seven hours after ingesting 486 aspirin tablets. His blood salicylate level was 83.7 mg/dL. He was treated with fluid resuscitation and sodium bicarbonate infusion, and his condition gradually improved, with a decline in the blood salicylate level. However, eight days after admission, he again reported nausea, a venous blood gas revealed metabolic acidosis with a normal anion gap. The blood salicylate level was undetectable, and a urinalysis showed glycosuria, proteinuria and elevated beta-2 microglobulin and n-acetyl glucosamine levels, with a normal urinary pH despite the acidosis. We diagnosed him with relapse of metabolic acidosis caused by renal tubular acidosis.

Spontaneous extraskeletal osteosarcoma with various histological growth patterns in the abdominal wall of an ICR mouse.

Extraskeletal osteosarcoma is extremely rare in mice. This case report demonstrates a spontaneous murine extraskeletal osteosarcoma that exhibited various histological growth patterns in an ICR mouse. At necropsy, the tumor mass was located in the abdominal wall and was 45 × 30 × 25 mm in size. Histopathologically, the tumor showed the following four growth patterns: a solid pattern of polygonal cells embedded in an osteoid eosinophilic matrix with calcification, an irregular sheet pattern of short spindle cells accompanying some eosinophilic multinucleated cells, a fascicular pattern of spindle cells and a cystic pattern lined by short spindle cells. Immunohistochemically, most of the tumor cells were positive for vimentin, proliferating cell nuclear antigen and osterix. The multinucleated cells mentioned above were desmin positive and were regarded as regenerative striated muscles but not tumor cells. Since no clear continuity with normal bone tissues was observed, the tumor was diagnosed as an "extraskeletal osteosarcoma."

Poorly differentiated salivary gland carcinoma with prominent squamous metaplasia in a pregnant Wistar Hannover rat.

A subcutaneous pale brown-colored mass was observed macroscopically in the ventral neck of a 16-week-old Wistar rat on day 18 of gestation. The mass was well demarcated from the adjacent tissues with partial invasion into connective tissues. Necrosis and hemorrhage were evident throughout the mass. The mass comprised a diffuse sheet and a nest-like structure of epithelial cells with prominent squamous metaplasia. The neoplastic cells tested immunopositive for keratin, vimentin, glial fibrillary acidic protein and p63. A portion of the neoplastic cells exhibited a similar immunoreaction of prominin-1 to the ductal and acinar cells in normal submandibular and parotid glands. Collectively, the tumor was diagnosed as a poorly differentiated carcinoma derived from epithelial/myoepithelial lineages in the submandibular and/or parotid glands.

[A Case of Stage IV Gastric Cancer Successfully Treated Using Salvage Surgery after S-1/CDDP Therapy].

A 75-year-old man admitted for left lateral abdominal pain was found to have advanced poorly differentiated gastric adenocarcinoma with abdominal para-aortic and Virchow's lymph node metastases, which was diagnosed to be clinical Stage IV (T3N3H0M1[LYM]). As curative surgery was not deemed possible, we started chemotherapy administration using S-1 (120mg/day)administered orally for 3 weeks and cisplatin(CDDP 100mg/body)administered intravenously on day 8. After 6 courses of chemotherapy, a CT scan showed that all lymph nodes metastases had disappeared, resulting in downstaging to clinical Stage II (T3[SE]N0H0P0M0). Thus, we performed total gastrectomy, lymph node dissection(D2), and splenectomy. Histological findings showed no residual tumor cells in any of the lymph nodes. However, cancer cells remained in the primary gastric lesion. The pathological response to chemotherapy was judged to be Grade 2. The patient has been recurrence-free for 5 years after surgery.

Inhibition of lipopolysaccharide-induced liver injury in rats treated with a hepatic drug-metabolizing enzyme inducer p,p'-DDT.

Hepatocellular hypertrophy in association with drug-metabolizing enzyme induction is considered to be an adaptive change associated with drug metabolism. To improve our understanding of liver hypertrophy, we determined the effect of a single ip injection of either lipopolysaccharide (LPS) or vehicle in male F344 rats with hepatocellular hypertrophy induced by oral delivery of p,p'-DDT for 2 weeks. The rats were sacrificed 3h or 24h after LPS or vehicle injection. LPS induced a focal hepatocellular necrosis in rats fed the control diet. When rats pre-treated with p,p'-DDT were injected with LPS, necrotic foci surrounded by ballooned hepatocytes were observed in the liver. The change was consistent with reduced LPS-mediated increases in plasma hepatic biomarkers, neutrophil influx, and apoptosis, and also associated with hepatic mRNA levels of TNF-α, CYPs, and NOS2. By contrast, when combined with p,p'-DDT and LPS, faint hepatocellular fatty change was extended, together with a synergistic increase in total blood cholesterol. These results suggest that hepatocytes exposed to p,p'-DDT are protected from the cell-lethal toxic effects of an exogenous stimulus, resulting in cell ballooning rather than necrosis in association with reduced inflammation and apoptosis, but compromised by an adverse effect on lipid metabolism.

Somatic insulin signaling regulates a germline starvation response in Drosophila egg chambers.

Egg chambers from starved Drosophila females contain large aggregates of processing (P) bodies and cortically enriched microtubules. As this response to starvation is rapidly reversed upon re-feeding females or culturing egg chambers with exogenous bovine insulin, we examined the role of endogenous insulin signaling in mediating the starvation response. We found that systemic Drosophila insulin-like peptides (dILPs) activate the insulin pathway in follicle cells, which then regulate both microtubule and P body organization in the underlying germline cells. This organization is modulated by the motor proteins Dynein and Kinesin. Dynein activity is required for microtubule and P body organization during starvation, while Kinesin activity is required during nutrient-rich conditions. Blocking the ability of egg chambers to form P body aggregates in response to starvation correlated with reduced progeny survival. These data suggest a potential mechanism to maximize fecundity even during periods of poor nutrient availability, by mounting a protective response in immature egg chambers.

Effects of pH and lactate on hydrogen sulfide production by oral Veillonella spp.

Indigenous oral bacteria in the tongue coating such as Veillonella have been identified as the main producers of hydrogen sulfide (H2S), one of the major components of oral malodor. However, there is little information on the physiological properties of H2S production by oral Veillonella such as metabolic activity and oral environmental factors which may affect H2S production. Thus, in the present study, the H2S-producing activity of growing cells, resting cells, and cell extracts of oral Veillonella species and the effects of oral environmental factors, including pH and lactate, were investigated. Type strains of Veillonella atypica, Veillonella dispar, and Veillonella parvula were used. These Veillonella species produced H2S during growth in the presence of l-cysteine. Resting cells of these bacteria produced H2S from l-cysteine, and the cell extracts showed enzymatic activity to convert l-cysteine to H2S. H2S production by resting cells was higher at pH 6 to 7 and lower at pH 5. The presence of lactate markedly increased H2S production by resting cells (4.5- to 23.7-fold), while lactate had no effect on enzymatic activity in cell extracts. In addition to H2S, ammonia was produced in cell extracts of all the strains, indicating that H2S was produced by the catalysis of cystathionine γ-lyase (EC 4.4.1.1). Serine was also produced in cell extracts of V. atypica and V. parvula, suggesting the involvement of cystathionine ß-synthase lyase (EC 4.2.1.22) in these strains. This study indicates that Veillonella produce H2S from l-cysteine and that their H2S production can be regulated by oral environmental factors, namely, pH and lactate.

Serum levels of apoptosis inhibitor of macrophage are associated with hepatic fibrosis in patients with chronic hepatitis C.

BACKGROUND: Apoptosis inhibitor of macrophage (AIM) and adipocytokines are involved in the metabolic syndrome, which has been putatively associated with the progression of chronic hepatitis C (CHC). However, the association between these cytokines and CHC is not fully elucidated. The aim of this study is to test whether serum levels of AIM and adipocytokines are associated with histological features, homeostasis model assessment-insulin resistance index (HOMA-IR), or whole body insulin sensitivity index (WBISI) in CHC patients. METHODS: Serum samples were obtained from 77 patients with biopsy-proven CHC. In 39 patients without overt diabetes mellitus, a 75 g oral glucose tolerance test (OGTT) was performed and HOMA-IR and WBISI were calculated. RESULTS: A serum AIM level of ≥ 1.2 µg/ml was independently associated with advanced hepatic fibrosis (F2 or F3) (odds ratio [OR], 5.612; 95% confidence interval [CI], 1.103-28.563; P = 0.038) based on a multivariate analysis, but there was no significant association between AIM and hepatic steatosis or inflammation. Furthermore, a serum leptin level of ≥ 8.6 ng/ml was independently associated with the presence of hepatic steatosis (≥ 5%) (OR, 6.195; 95% CI, 1.409-27.240; P = 0.016), but not hepatic fibrosis or inflammation. No relationship was observed between levels of adiponectin or resistin and hepatic histological parameters based on a multivariate analysis. Although serum levels of leptin, resistin, and adiponectin were significantly correlated with HOMA-IR and WBISI, there was no significant relationship between serum AIM levels and HOMA-IR or WBISI, respectively. CONCLUSION: High serum levels of AIM in CHC patients are potentially related to advanced hepatic fibrosis. AIM and adipocytokines are possibly associated with pathological changes via a different mechanism.

Hepatitis C virus NS3/4A protease inhibits complement activation by cleaving complement component 4.

BACKGROUND: It has been hypothesized that persistent hepatitis C virus (HCV) infection is mediated in part by viral proteins that abrogate the host immune response, including the complement system, but the precise mechanisms are not well understood. We investigated whether HCV proteins are involved in the fragmentation of complement component 4 (C4), composed of subunits C4α, C4ß, and C4γ, and the role of HCV proteins in complement activation. METHODS: Human C4 was incubated with HCV nonstructural (NS) 3/4A protease, core, or NS5. Samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then subjected to peptide sequencing. The activity of the classical complement pathway was examined using an erythrocyte hemolysis assay. The cleavage pattern of C4 in NS3/4A-expressing and HCV-infected cells, respectively, was also examined. RESULTS: HCV NS3/4A protease cleaved C4γ in a concentration-dependent manner, but viral core and NS5 did not. A specific inhibitor of NS3/4A protease reduced C4γ cleavage. NS3/4A protease-mediated cleavage of C4 inhibited classical pathway activation, which was abrogated by a NS3/4A protease inhibitor. In addition, co-transfection of cells with C4 and wild-type NS3/4A, but not a catalytic-site mutant of NS3/4A, produced cleaved C4γ fragments. Such C4 processing, with a concomitant reduction in levels of full-length C4γ, was also observed in HCV-infected cells expressing C4. CONCLUSIONS: C4 is a novel cellular substrate of the HCV NS3/4A protease. Understanding disturbances in the complement system mediated by NS3/4A protease may provide new insights into the mechanisms underlying persistent HCV infection.

Correlation of serum levels of complement C4a desArg with pathologically estimated severity of glomerular lesions and mesangial hypercellularity scores in patients with IgA nephropathy.

The aim of the present study was to explore serum biomarkers for the pathology of IgA nephropathy using serum proteomics. The subjects were 57 patients with IgA nephropathy who were divided into two groups (group 1, n=25; group 2, n=32) and 14 healthy controls. Serum protein profiles were analyzed using the ProteinChip surface-enhanced laser desorption ionization (SELDI) system. Associations between signal intensities of proteins and histological findings in patients with IgA nephropathy were studied in group 1. Serum levels of a candidate biomarker protein (complement component C4a desArg) for IgA nephropathy were determined by enzyme linked-immunosorbent assay (ELISA) in group 2 and the relationships of these levels with histological findings were evaluated. There were significant differences in 93 protein signals between patients in group 1 and controls. Among these signals, 3 proteins at 8592, 8757 and 8806 m/z were significantly correlated with the severity of glomerular lesions. The protein at 8592 m/z was identified as C4a desArg and the signal intensity of 8592 m/z was strongly correlated with serum C4a levels, including C4a desArg, determined by ELISA. In addition, the serum levels of C4a (mainly C4a desArg) were significantly higher in patients in group 2 compared to controls and were correlated with the severity of glomerular lesions and with mesangial hypercellularity scores. In conclusion, the serum levels of complement C4a desArg are significantly higher in patients with IgA nephropathy compared to healthy controls and are significantly correlated with the severity of glomerular lesions and mesangial hypercellularity scores. Thus, serum C4a desArg is a potential biomarker for the severity of histological findings in patients with IgA nephropathy.

[A case of adenocarcinoma of small intestine surviving for many years].

In adenocarcinoma of the small intestine, delays in diagnosis are frequent. The majority of patients present with advancedstage disease, and have either lymph node involvement or distant metastatic disease. Surgical resection is a mainstay in treatment of this disease, and the effectiveness of chemotherapy for advanced-stage or metastatic disease has been reported. We report a case of adenocarcinoma of the small intestine surviving for many years after surgical resection and chemotherapy. A 47-year-old woman underwent a small intestine resection, because she had a small intestinal tumor with obstruction. Histopathological examination revealed moderately-differentiated adenocarcinoma with lymph node metastasis. Adjuvant chemotherapy with S-1 was administered for a year, but in March 2006, a recurrent lesion at the right ovary was detected, and she underwent right adnexectomy. Because the ascites cytology revealed class V, chemotherapy was administered. In December 2008, CA19-9 elevated and magnetic resonance imaging showed a tumor behind the uterus, which was diagnosed as a recurrent disease. Because the tumor invaded the rectum, she received a low anterior resection, hysterectomy, and left adnexectomy. After surgical resection, UFT/UZEL was administered for half a year. In July 2010, computed tomography showed multiple lung metastases, and chemotherapy was performed again. However, the regimen was changed because her tumor marker elevated. She is being treated using a combination of cisplatin and irinotecan.

Intercellular protein movement in syncytial Drosophila follicle cells.

Ring canals connecting Drosophila germline, follicle and imaginal disc cells provide direct contact of cytoplasm between cells. To date, little is known about the formation, structure, or function of the somatic ring canals present in follicle and imaginal disc cells. Here, we show by confocal and electron microscopy that Pavarotti kinesin-like protein and Visgun are stable components of somatic ring canals. Using live-cell confocal microscopy, we show that somatic ring canals form from the stabilization of mitotic cleavage furrows. In contrast to germline cells, syncytial follicle cells do not divide synchronously, are not maximally branched and their ring canals do not increase in size during egg chamber development. We show for the first time that somatic ring canals permit exchange of cytoplasmic proteins between follicle cells. These results provide insight into the composition and function of ring canals in somatic cells, implying a broader functional significance for syncytial organization of cells outside the germline.

Reversible response of protein localization and microtubule organization to nutrient stress during Drosophila early oogenesis.

The maturation of animal oocytes is highly sensitive to nutrient availability. During Drosophila oogenesis, a prominent metabolic checkpoint occurs at the onset of yolk uptake (vitellogenesis): under nutrient stress, egg chambers degenerate by apoptosis. To investigate additional responses to nutrient deprivation, we studied the intercellular transport of cytoplasmic components between nurse cells and the oocyte during previtellogenic stages. Using GFP protein-traps, we showed that Ypsilon Schachtel (Yps), a putative RNA binding protein, moved into the oocyte by both microtubule (MT)-dependent and -independent mechanisms, and was retained in the oocyte in a MT-dependent manner. These data suggest that oocyte enrichment is accomplished by a combination of MT-dependent polarized transport and MT-independent flow coupled with MT-dependent trapping within the oocyte. Under nutrient stress, Yps and other components of the oskar ribonucleoprotein complex accumulated in large processing bodies in nurse cells, accompanied by MT reorganization. This response was detected as early as 2h after starvation, suggesting that young egg chambers rapidly respond to nutrient stress. Moreover, both Yps aggregation and MT reorganization were reversed with re-feeding of females or the addition of exogenous insulin to cultured egg chambers. Our results suggest that egg chambers rapidly mount a stress response by altering intercellular transport upon starvation. This response implies a mechanism for preserving young egg chambers so that egg production can rapidly resume when nutrient availability improves.

Use of a Gluconobacter frateurii mutant to prevent dihydroxyacetone accumulation during glyceric acid production from glycerol.

To prevent dihydroxyacetone (DHA) by-production during glyceric acid (GA) production from glycerol using Gluconobacter frateurii, we used a G. frateurii THD32 mutant, ΔsldA, in which the glycerol dehydrogenase subunit-encoding gene (sldA) was disrupted, but ΔsldA grew much more slowly than the wild type, growth starting after a lag of 3 d under the same culture conditions. The addition of 1% w/v D-sorbitol to the medium improved both the growth and the GA productivity of the mutant, and ΔsldA produced 89.1 g/l GA during 4 d of incubation without DHA accumulation.