Increased metabolite production by deletion of an HDA1-type histone deacetylase in the phytopathogenic fungi, Magnaporthe oryzae (Pyricularia oryzae) and Fusarium asiaticum.

Histone deacetylases (HDACs) play an important role in the regulation of chromatin structure and gene expression. We found that dark pigmentation of Magnaporthe oryzae (anamorph Pyricularia oryzae) ΔMohda1, a mutant strain in which an orthologue of the yeast HDA1 was disrupted by double cross-over homologous recombination, was significantly stimulated in liquid culture. Analysis of metabolites in a ΔMohda1 mutant culture revealed that the accumulation of shunt products of the 1,8-dihydroxynaphthalene melanin and ergosterol pathways were significantly enhanced compared to the wild-type strain. Northern blot analysis of the ΔMohda1 mutant revealed transcriptional activation of three melanin genes that are dispersed throughout the genome of M. oryzae. The effect of deletion of the yeast HDA1 orthologue was also observed in Fusarium asiaticum from the Fusarium graminearum species complex; the HDF2 deletion mutant produced increased levels of nivalenol-type trichothecenes. These results suggest that histone modification via HDA1-type HDAC regulates the production of natural products in filamentous fungi. SIGNIFICANCE AND IMPACT OF THE STUDY: Natural products of fungi have significant impacts on human welfare, in both detrimental and beneficial ways. Although HDA1-type histone deacetylase is not essential for vegetative growth, deletion of the gene affects the expression of clustered secondary metabolite genes in some fungi. Here, we report that such phenomena are also observed in physically unlinked genes required for melanin biosynthesis in the rice blast fungus. In addition, production of Fusarium trichothecenes, previously reported to be unaffected by HDA1 deletion, was significantly upregulated in another Fusarium species. Thus, the HDA1-inactivation strategy may be regarded as a general approach for overproduction and/or discovery of fungal metabolites.

The feasibility of non-contact ultrasound for medical imaging.

High intensity focused ultrasound in air may provide a means for medical and biological imaging without direct coupling of an ultrasound probe. In this study, an approach based on highly focused ultrasound in air is described and the feasibility of the technique is assessed. The overall method is based on the observations that (1) ultrasound in air has superior focusing ability and stronger nonlinear harmonic generation as compared to tissue propagation and (2) a tightly focused field directed into tissue causes point-like spreading that may be regarded as a source for generalized diffraction tomography. Simulations of a spherically-curved transducer are performed, where the transducer's radiation pattern is directed from air into tissue. It is predicted that a focal pressure of 162 dB (2.5 kPa) is sufficient to direct ultrasound through the body, and provide a small but measurable signal (∼1 mPa) upon exit. Based on the simulations, a 20 cm diameter array consisting of 298 transducers is constructed. For this feasibility study, a 40 kHz resonance frequency is selected based on the commercial availability of such transducers. The array is used to focus through water and acrylic phantoms, and the time history of the exiting signal is evaluated. Sufficient data are acquired to demonstrate a low-resolution tomographic reconstruction. Finally, to demonstrate the feasibility to record a signal in vivo, a 75 mm × 55 mm section of a human hand is imaged in a C-mode configuration.

Expression and translocation of aquaporin-2 in the endolymphatic sac in patients with Meniere's disease.

Meniere's disease, characterised by episodic vertigo, fluctuating hearing loss and tinnitus, can occur under conditions of stress. Its pathology was first revealed to be inner ear hydrops through temporal bone studies in 1938. Although its pathogenesis has been proposed to be a disorder of water transport in the inner ear, subsequently, it remains unsolved, until now. A recent study revealed that both plasma stress hormone, vasopressin (pAVP) and its receptor, V2 (V2R) expression in the inner ear endolymphatic sac were significantly higher in Meniere's patients. In the present study, to link V2R-related molecules and inner ear hydrops, we examined V2R-linked water channel molecule, aquaporin-2 (AQP2) expression and translocation in human endolymphatic sac. AQP2 mRNA expression in the endolymphatic sac was significantly higher in Meniere's patients by using real-time polymerase chain reaction, as further confirmed by western blotting. AQP2-like immunoreactivity (-LIR) was translocated from luminal to basolateral side with endosomal trapping in the endolymphatic sac at the time of AVP exposure in human endolymphatic sac tissue culture. The similar AQP2-LIR translocation was also demonstrated by forskolin and blocked by vasopressin/V2R specific antagonist, OPC31260 and protein kinase A (PKA) specific antagonists, H-89 and KT-5720. We concluded that in the pathogenesis of inner ear hydrops resulting in Meniere's attacks, pAVP elevation as a result of stress and subsequent V2R-cAMP-PKA-AQP2 activation and endosomal trapping of AQP2 in the endolymphatic sac, might be important as a basis of this disease. Further experimental and clinical studies are needed to better clarify the neuroscientific relationship between stress and Meniere's disease.

Dowling-Degos disease associated with squamous cell carcinomas on the dappled pigmentation.

We report the first case of Dowling-Degos disease associated with squamous cell carcinomas (SCCs) in the pigmented area of Dowling-Degos disease. A 64-year-old Japanese man manifested dappled pigmentation unusually localized to the buttocks, and two pigmented adenoid SCCs had developed on his left pigmented buttock. The other findings of Dowling-Degos disease were comedone-like lesions on the face and back, a finger-like fibroma in the right popliteal fossa, dystrophic fingernails, and a large number of seborrhoeic keratosis-like lesions predominantly on the flexural areas. Another unique clinical feature was the lack of vellus hair on the whole body surface. In addition to thin branching and elongation of rete ridges with basal hyperpigmentation, immature hair follicles surrounded by fibrosis and a lace-like pattern of the hair follicle epithelia were observed histologically. These epithelial hamartomatous features were consistent with Dowling-Degos disease. We speculate that the SCCs developed in relation to an underlying naevoid anomaly in pilosebaceous epithelia of Dowling-Degos disease.

[Usefulness of epidural infusion of ketamine for relief of localized superficial pain].

Three patients with localized superficial pain had their pain alleviated by single epidural infusion with low dose ketamine. The patients are as follows: a 62-year-old female with herpetic neuralgia on her right sixth thoracic nerve area; a 52-year-old male whose left shoulder, anterior chest and abdomen had been burned by acetylene gas; and a 49-year-old male whose bilateral hands suffering from frostbite by propane gas. Epidural tube insertion to administer a single dose of 10 mg ketamine with lidocaine or bupibacaine was performed in all the three patients. They were administered single epidural infusion of 10 mg ketamine with lidocaine or bupibacaine everyday and they continued to receive epidural block with lidocaine or bupivacaine including buprenorphine or morphine. Therefore, we suspect that single epidural infusion of ketamine, an antagonist for N-methyl-D-aspartic acid receptor, could be an effective and useful alternative treatment in patients with various refractory localized superficial pain of either acute or chronic nature.

[Treatment with stellate ganglion block, continuous epidural block and ulnar nerve block of a patient with postherpetic neuralgia who developed complex regional pain syndrome (CRPS)].

We present a case of a 46-year-old female patient with systemic lupus erythematosus who developed herpes zoster of the right eighth cervical nerve. Her whole right forearm, hand and the first through fifth fingers were coated with some gel and protected against pain. She had been suffering from continuous and spasmodic burning pain, hyperalgesia, allodynia, drop in skin temperature, sudmotor disturbance, edema, constructure of the joints, muscle atrophy and bone atrophy of her right upper extremity probably due to postherpetic neuralgia (PHN) associated with complex regional pain syndrome (CRPS). She received right stellate ganglion block (SGB), continuous cervical epidural block and right ulnar nerve block. Reduction of pain and edema as well as improvement in mobility of each joint of her right upper extremity was observed. We suspect that SGB, continuous cervical epidural block and ulnar nerve block are effective and useful alternative treatments in a patient with PHN associated with CRPS of the eighth cervical nerve.

Genetic and environmental influences on self-esteem in a Japanese twin sample.

The purpose of the present study is to clarify the mechanism of Japanese self-esteem (SE) in genetic and environmental influences using twin methodology. Eighty-one pairs of adolescent twins, including 50 pairs of monozygotic (MZ) twins and 31 pairs of dizygotic (DZ) twins, participated in this study. Self-esteem was assessed using the Rosenberg Self-Esteem Scale (RSES), translated into Japanese. As a result of using univariate twin analyses, model comparisons using the Akaike Information Criterion (AIC) indicated that the AE model was the best fit (AIC = -5.35). In the best-fitting AE model, the heritability (a2) of SE was revealed to be moderate, accounting for 49% of the variance; environmental influences (individual-specific environmental factors) explained 51% of the variance. These results are consistent with the findings of some behavioral genetics studies of SE in the West and show that there is no difference between Western and Japanese populations in the mechanism of SE considering genetic and environmental influences. The results also suggest the importance of considering both genetic and environmental factors in studies of Japanese SE.

Model equation for strongly focused finite-amplitude sound beams

A model equation that describes the propagation of sound beams in a fluid is developed using the oblate spheroidal coordinate system. This spheroidal beam equation (SBE) is a parabolic equation and has a specific application to a theoretical prediction on focused, high-frequency beams from a circular aperture. The aperture angle does not have to be small. The theoretical background is basically along the same analytical lines as the composite method (CM) reported previously [B. Ystad and J. Berntsen, Acustica 82, 698-706 (1996)]. Numerical examples are displayed for the amplitudes of sound pressure along and across the beam axis when sinusoidal waves are radiated from the source with uniform amplitude distribution. The primitive approach to linear field analysis is readily extended to the case where harmonic generation in finite-amplitude sound beams becomes significant due to the inherent nonlinearity of the medium. The theory provides the propagation and beam pattern profiles that differ from the CM solution for each harmonic component.

cDNA Subtractive Cloning of Genes Expressed during Early Stage of Appressorium Formation by Magnaporthe grisea.

The conidial germ tube of the fungus Magnaporthe grisea differentiates an infection-specific structure, an appressorium, for penetration into the host plant. Formation of the appressorium is also observed on synthetic solid substrata such as polycarbonate. We found that a plant lectin, concanavalin A, specifically suppressed the appressorium formation without affecting the germling adhesion if it was applied within 2-3 hours after germination. Standing on the result, we constructed a cDNA library that represents the early stage of germ tube development and/or appressorium formation from the 2.5-hour-old germ tubes using a cDNA subtraction strategy by the combination of the biotin labeled driver method and adapter-primed PCR method. Out of 686 colonies of the library, 158 distinct clones' nucleotide sequences were partially analyzed. Some clones' expression patterns were detected by RT-PCR and from those results, our library seemed to well represent the objective developmental stage of M. grisea.

Inactivation of blasticidin S by Bacillus cereus. VI. Structure and comparison of the bsr gene from a blasticidin S-resistant Bacillus cereus.

Two types of recombinant plasmids containing 600 bp Nde I fragments that coded the bsr gene in opposite directions were obtained. Nucleotide sequencing shows that the bsr encodes a 140 amino acid protein with a putative molecular weight of 15560, the same as that of purified blasticidin S (BS)-deaminase (BSR), on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (15500). Upstream of the open reading frame, a Shine-Dalgarno (SD) sequence, frequent inverted repeats, and the sigmaA and sigmaB promoter sequences are observed. The transcriptional start point was determined to be the A located 7 bases downstream from the putative sigmaA promoter (91TTGATC and 113TAAAAT) by the primer extension method and site directed mutagenesis at the -10 or -35 promoter region. A comparison of the amino acid sequence of BSR with that of BS-deaminase from Aspergillus terreus (BSD) showed 27.2% homology. Low degrees of homology were also observed with cytidine deaminase and deoxy cytidine monophosphate (CMP) deaminase. Four conserved amino acid motifs were observed, VGAx6G, C(orH)AEx6A, SPCGxCR, and Gx8ELIP (x(n) indicates a nonspecific residue and its position). It is possible that the three Cys residues and the Glu in the conserved motifs comprise the active center. Site-directed mutagenesis of the Cys residues supports this possibility.

Indicative value of carcinoembryonic antigen (CEA) for liver recurrence following curative resection of stage II and III gastric cancer.

BACKGROUND/AIMS: We investigated the role of preoperative and serial postoperative serum CEA levels in advanced gastric cancer patients with curative operation. MATERIALS AND METHODS: Preoperative and serum postoperative CEA levels were measured in 115 patients with Stage II and III gastric cancer and who underwent curative gastric resection. RESULTS: The 5-year survival rates of patients with high preoperative CEA levels (> 5 ng/ml) were poorer than in case of low preoperative CEA levels (< 5 ng/ml) in both stage II (P < 0.05) and stage III (P < 0.01) gastric cancer. When the site of the recurrence was diagnosed in 47 patients with less than a 5-year survival, a high level of preoperative CEA was more likely to be associated with a liver metastasis (13/18, 72.2%) than with peritoneal dissemination (4/15, 26.7%) (P < 0.05). In the course of serial measurements of postoperative levels of CEA, CEA increased in patients with liver metastasis, and CEA levels began to elevate 3.2 months before clinical detection, while, little change of postoperative serum CEA levels was noted in patients with peritoneal dissemination. CONCLUSIONS: We propose that the preoperative and serial postoperative assays of serum CEA level are predictive for liver metastasis in patients with stage II and III gastric cancer and treated by curative gastric resection.

Poor prognosis of lower quadrant breast carcinoma. Nishi Nippon Study Group on Adjuvant Chemo-endocrine Therapy for Breast Cancer.

Although some lymphatic plexuses exist in lower quadrants of the breast, there have been no investigations of whether or not carcinoma located in this region is a prognostic factor for breast cancer. Of 914 patients with carcinoma of the breast who underwent curative resection following chemo-endocrine therapy between 1982 and 1985, 149 patients had disease of the lower quadrants. The recurrence-free survival rate was lower in patients with the lower quadrants carcinoma than in those with carcinoma of other breast regions. Multivariate analysis showed that a lower quadrant tumor location was a significant prognostic factor for recurrence, especially soft tissue and visceral recurrence. The worse prognosis of patients with lower quadrant carcinoma of the breast suggests the possible existence of residual or occult tumor cells after surgical resection.

Prognostic value of combination assays for CEA and CA 19-9 in gastric cancer.

Preoperative serum CEA and CA 19-9 levels in 158 patients with gastric cancer were analyzed with respect to prognostic factors, using univariate and multivariate analysis. The incidence of high preoperative levels of both CEA and CA 19-9 was 10.1% (16/158). 13.9% (22/158) showed high CEA levels and normal CA 19-9 levels, whereas the reverse was true in 16.5% (26/158). Neither marker showed a high level in 59.5% (94/158). The multivariate analysis showed that in addition to tumor stage, the depth of invasion, liver metastasis and peritoneal dissemination, combination assays of preoperative serum CEA and CA 19-9 levels were an independent prognostic factor. Combination assays of preoperative serum CEA and CA 19-9 will allow us to conduct a more careful postoperative follow-up of high-risk patients, and also help determine the optimum adjuvant chemotherapy.

Influence of blood transfusion on postoperative long-term liver function in patients with hepatocellular carcinoma.

BACKGROUND/AIMS: To research for possible adverse effects of blood transfusion on liver function at a relatively long term after hepatic resection, liver function tests at one and two years after the surgery were studied in patients with hepatocellular carcinoma. MATERIALS AND METHODS: Seventy-one patients with no evidence of cancer recurrence during two years after the surgery were selected for this retrospective analysis. Patients were divided into two groups depending on perioperative blood transfusion: those who received blood transfusion (BT(+), n = 38), and those without blood transfusion (BT(-), n = 33). RESULTS: There were no statistical difference in the preoperative clinical features including liver function tests except decreased prothrombin time index (P < 0.01) and larger amount of intraoperative blood loss (P < 0.01) in BT(+) group. Platelet counts at 2 year after the surgery were significantly higher in the BT(-) group (P < 0.05). Two-way ANOVA showed no significant difference in liver function tests between the two groups. In multiple regression analysis, blood transfusion significantly correlated with serum total bilirubin at 1 year, but did not at 2 years after the surgery. CONCLUSIONS: The influence of blood transfusion in the patients with hepatocellular carcinoma was considered to be minimum within 2 years after hepatectomy.

Chromogranin positive cells in colorectal carcinoma and transitional mucosa.

AIMS: Immunostaining of chromogranin identifies gastrointestinal mucosal endocrine cells. The detailed distribution and significance of chromogranin positive cells in colorectal carcinomas and in transitional mucosa remain unclear. The aim of this study was to clarify these aspects. METHODS: The distribution of chromogranin positive cells was studied by immunohistochemical methods in normal epithelium remote from carcinoma, in transitional mucosa, and in carcinomas of the colorectum. In selected cases northern or western blot analyses were performed. RESULTS: Chromogranin positive cells were seen in the lower third of the normal crypts and less frequently in transitional mucosa. Thirty five per cent (n = 38) of colorectal carcinomas showed immunohistochemically positive carcinoma cells in the tumour tissue. Northern and western blot analyses showed similar results. There was no difference in clinicopathological factors, including prognosis, between chromogranin positive cases of colorectal carcinoma (n = 38) and chromogranin negative cases (n = 70). CONCLUSIONS: Neuroendocrine cell differentiation is controlled in transitional mucosa and the presence of chromogranin positive cells in carcinoma tissue does not influence the patient's prognosis.

Improvements in diagnosis have changed the incidence of histological types in advanced gastric cancer.

The data on 912 patients with early cancer and 1245 with advanced cancer who were seen between 1971 and 1990 were compared. The incidence of undifferentiated-type cancer increased significantly in patients with advanced gastric cancer, but not in patients with early gastric cancer. When the histological types were compared with regard to sex, age and location in patients with early gastric cancer the undifferentiated type was found to increase only in males, while in patients with advanced gastric cancer the undifferentiated type increased in both sexes as well as in younger patients and in both the upper and middle third of the stomach. These differences in the trends between early and advanced cancers are probably due to the different degrees of diagnostic accuracy for the early detection of histological types.

The value of prophylactic R2 lymph node dissection for gastric cancer.

Three hundred and eleven patients with an R2 lymph node dissection for gastric cancer were reviewed in order to examine the efficacy of prophylactic R2 lymph node dissection. In 246 patients, who were all macroscopically determined to have either N0 or N1 lymph node metastasis at the time of operation, 30 showed histologic lymph node metastasis in the N2 group. When the 5-year survival rates in the 61 patients with histologic N2 lymph node metastasis were compared according to the macroscopic evaluation of lymph node metastasis at the time of operation, 30 patients who were determined to have either N0 or N1 lymph node metastasis showed a better prognosis than the 31 patients who were determined to have N2 lymph node metastasis (p < 0.01). Therefore, prophylactic R2 lymph node dissection contributes to an improved prognosis for gastric cancer patients who are found to macroscopically have either N0 or N1 lymph node metastasis on the basis of an intra-operative evaluation.

Endoscopic injection sclerotherapy for esophageal varices associated with concomitant portal venous thrombus of hepatocellular carcinoma.

Between 1983 and 1994, we treated 51 patients with esophageal varices and portal trunk and main branch invasion of hepatocellular carcinoma, using endoscopic injection sclerotherapy. Variceal bleeding was controlled in 28 of 29 patients (96.6%), esophageal varices were completely eradicated in 28 (54.9%), and only 2 of 28 (7.1%) bled from small, dilated, venous vessels after eradication. The cumulative nonbleeding rate at 3 years was 87.5%. Death caused by hepatocellular carcinoma accounted for 89.4% of the patients, whereas the rate of bleeding from esophageal varices was 4.3%. Variables significantly associated with the duration of survival were Okuda's clinical stage, alpha-fetoprotein, eradication of esophageal varices by sclerotherapy, and treatment of hepatocellular carcinoma, as determined in a univariate analysis. Multivariate analysis showed that eradication of esophageal varices by sclerotherapy, Okuda's clinical stage, and age were independent factors which significantly influenced survival time. We propose that complete eradication of esophageal varices and close follow-up using endoscopy may lead to a reduction in bleeding from esophageal varices, and hence may reduce mortality rates related to this bleeding.

A simple classification of lymph node level in gastric carcinoma.

BACKGROUND: Because insufficient lymph node examination can cause erroneous listing of earlier-nodal-stage gastric carcinoma (stage migration phenomenon), surgical results must be evaluated based on a highly accurate examination of the dissected lymph nodes. To establish a simple and useful classification of lymph node level, we analyzed the frequency and distribution of lymph node metastasis by using curatively treated node-positive gastric carcinoma. PATIENTS AND METHODS: Various clinicopathologic data were analyzed with reference to the degree of lymph node metastasis by using 240 patients with curatively resected node-positive gastric carcinoma. The cases were divided into the following three groups: 142 with positive level I (perigastric) nodes, 71 with positive level II (intermediate) nodes, and 27 with positive level III (distant) nodes, irrespective of the location of tumors. RESULTS: The level of lymph node metastasis clearly correlated with the survival of patients, with the 5-year survival rates for level I, II, and III cases being 67%, 35%, and 26%, respectively (P < 0.01). The degree of lymph node metastasis was determined by the number of positive nodes (P < 0.01), the depth of invasion (P < 0.01), the size of tumors (P < 0.01), and the location of tumors (P < 0.05). CONCLUSION: This simple classification of lymph node level (level I, II, and III) is useful for the evaluation and prediction of surgical results in gastric carcinoma.

Immunohistochemical expression of sialyl Tn and sialyl Lewis(a) antigens in stromal tissue correlates with peritoneal dissemination in stage IV human gastric cancer.

Immunohistochemical expression of sialyl Tn antigen (TKH2 and B72.3), sialyl Lewis(a) antigen (CA19-9) and CEA was examined in 52 cases with Stage IV gastric cancer, the objective being to examine whether or not these tumour-associated antigens show different expression between peritoneal dissemination and liver metastasis. Stromal type for TKH2, B72.3, CA19-9 and CEA were seen in 14 (50%), 13 (46%), 18 (64%) and nine (32%) cases with peritoneal dissemination, respectively, and in four (17%), three (13%), four (17%) and four (17%) cases with liver metastasis, respectively. The stromal type correlated to peritoneal dissemination in TKH2 (P < 0.05) B72.3 (P < 0.05) and CA19-9 (P < 0.05). In other clinicopathological factors, the stromal type for TKH2, B72.3 and CA19-9 correlated to the undifferentiated type. However, even in the undifferentiated type group, the stromal type correlated to peritoneal dissemination in TKH2 (P < 0.05), B72.3 (P < 0.05) and CA19-9 (P < 0.05). Based on these results, the spread of sialyl-rich tumour-associated antigens into the surrounding stroma in the undifferentiated type of gastric cancer may be associated with peritoneal dissemination.