SADS: A new component of Fas-DISC is the accelerator for cell death signaling and is downregulated in patients with colon carcinoma.

Fas is the death receptor, transducing cell death signaling upon stimulation by Fas ligand. During Fas-initiated cell death signaling, the formation of Fas-death inducing signaling complex (Fas-DISC) is the first step. Here we have identified a new component of Fas-DISC which we call 'small-accelerator for death signaling' (SADS). SADS cDNA encodes a 150 amino acid polypeptide (Mr = 16,700). During Fas-mediated cell death, SADS enhances the interaction of Fas-death domain-interactive factors (FADD) and procaspase-8, and deletion mutant analysis has identified FADD- and caspase-8-interactive domains in SADS. Inhibition or removal of SADS delays Fas-mediated cell death. In addition, we demonstrate the deletion or mutation of SADS in patients with colon carcinoma and that exogenous SADS expression in human colon carcinoma SW480 cells that lack SADS leads to re-acquisition of Fas-mediated cell death. Here, we propose that SADS is one of the cell death-associated factors and enhances Fas-DISC formation, especially FADD and procaspase-8 recruitment.

[Coronary malperfusion of left main trunk due to localized dissection of the ascending aorta].

Case 1. Forty nine years woman was given a diagnosis of acute myocardial infarction. Coronary angiography and trans-esophageal echocardiography showed left main trunk dissection due to local aortic root dissection. We operated surgical repair at left main trunk by pericardium after percutaneous coronary intervention. Case 2. Forty nine years man was given a diagnosis of acute myocardial infarction caused by left main trunk dissection due to traumatic local aortic root dissection. We operated coronary artery bypass grafting after insertion of perfusion catheter to left main trunk for maintain coronary perfusion. Although local dissection of aortic aorta is relatively rare, it is potentially complicated with coronary malperfusion. We describe 2 success a cases of surgical treatment for local acute type A aortic dissection complicated with coronary malperfusion.

Crystallization and preliminary X-ray diffraction studies of Bacillus stearothermophilus farnesyl diphosphate synthase expressed in Escherichia coli.

Thermostable farnesyl diphosphate synthase (EC 2.5.1.10) from Bacillus stearothermophilus, which was overexpressed in Escherichia coli, has been crystallized by the vapor-diffusion procedure. Tetragonal crystals were obtained using ammonium sulfate as a precipitant. The crystals diffracted X-rays to about 3 A resolution. The diffraction pattern indicated that the space group is I4(1)22 with unit-cell dimensions of a = b = 114 A and c = 247 A. It is thought that the asymmetric unit comprises two or three molecules of farnesyl diphosphate synthase.

Development and testing of an endoscopic pseudo-viewpoint alternating system.

PURPOSE: An endoscopic system is needed that presents informative images irrespective of the surgical situation and the number of degrees of freedom in endoscopic manipulation. This goal may be achieved with a virtual reality view for a region of interest from an arbitrary viewpoint. An endoscopic pseudo-viewpoint alternation system for this purpose was developed and tested. METHOD: Surgical experts and trainees from an endoscopic surgery training course at the minimally invasive surgery training center of Kyushu University were enrolled in a trial of a virtual reality system. The initial viewpoint was positioned to approximate the horizontal view often seen in laparoscopic surgery, with [Formula: see text] between the optical axis of the endoscope and the task surface. A right-to-left suturing task with right hand, based on a task from the endoscopic surgery training course, was selected for testing. We compared task outcomes with and without use of a new virtual reality-viewing system. RESULT: There was a 0.37 mm reduction in total error ([Formula: see text]) with use of the proposed system. Error reduction was composed of 0.1 mm reduction on the y-axis and 0.27 mm reduction on the x-axis. Experts benefited more than novices from use of the proposed system. Most subjects worked at a pseudo-viewpoint of around 34[Formula: see text]. DISCUSSION: Suturing performance improved with the new virtual reality endoscopic display system. Viewpoint alternation resulted in an overview that improved depth perception and allowed subjects to better aim the marker. This suggests the proposed method offers users better visualization and control in endoscopic surgery.

Functional cloning of a cDNA encoding Mei2-like protein from Arabidopsis thaliana using a fission yeast pheromone receptor deficient mutant.

To isolate Arabidopsis cDNAs that encode signal transducers and components involved in the regulation of meiosis, a trans-complementation analysis was performed using a Schizosaccharomyces pombe meiosis-defective mutant in which the genes for pheromone receptors were disabled. One cDNA obtained in this screening encodes a polypeptide, named AML1, that shows significant similarity to S. pombe Mei2 protein and has three putative RNA-recognition motifs like as Mei2. Mei2 is involved in the regulation of meiosis in fission yeast. Northern blot analysis showed that the AML1 gene is expressed in each organ. The possible functions of AML1 are discussed.

Extracellular ATP-dependent activation of plasma membrane Ca(2+) pump in HEK-293 cells.

1. It is well known that extracellular ATP (ATP(o)) elevates the intracellular Ca(2+) concentration ([Ca(2+)](i)) by inducing Ca(2+) influx or mobilizing Ca(2+) from internal stores via activation of purinoceptors in the plasma membrane. This study shows that ATP(o) also activates the plasma membrane Ca(2+) pumps (PMCPs) to bring the elevated [Ca(2+)](i) back to the resting level in human embryonic kidney-293 (HEK-293) cells. 2. The duration of ATP(o)-induced intracellular Ca(2+) transients was significantly increased by PMCP blockers, La(3+) or orthovanadate. In contrast, replacement of extracellular Na(+) with NMDG(+), a membrane-impermeable cation, had no significant effect on duration, thus suggesting that Na(+)/Ca(2+) exchangers do not participate in the ATP(o)-induced Ca(2+) transient. 3. A rapid and significant decrease in [Ca(2+)](i), which was not dependent on extracellular Na(+), was induced by ATP(o) in cells pretreated with thapsigargin (TG). This decrease was blocked by orthovanadate, indicating that it was caused by PMCPs rather than sarco/endoplasmic reticulum Ca(2+) pumps (SERCPs). 4. UTP and ATPgammaS also caused a decrease in [Ca(2+)](i) in cells pretreated with TG, although they were less effective than ATP. The effect of UTP implies the involvement of both P2Y(1) and P2Y(2) receptors, while the effect of ATPgammaS implies no significant role of ectophosphorylation and agonist hydrolysis in the agonist-induced [Ca(2+)](i) decreases. 5. These results point to a role of PMCPs in shaping the Ca(2+) signal and in restoring the resting [Ca(2+)](i) level to maintain intracellular Ca(2+) homeostasis after agonist stimulation.

Narrow photon beam dosimetry for linear accelerator radiosurgery.

The dosimetric characteristics of linear accelerator radiosurgery for 10-MV X-ray were measured. Measurement of the relative output factor and tissue maximum ratio with a microchamber produced results equivalent to those of measurement with X-ray film. The 80% isodose level width measured with the microchamber was significantly smaller than that measured with the X-ray film. For the measurement of relative output factor and tissue maximum ratio, a microchamber seems to be the more appropriate choice. X-Ray film was found to be suitable for beam profile measurement.

Molecular cloning and nucleotide sequence determination of gene encoding Streptomyces subtilisin inhibitor (SSI).

A gene for Streptomyces subtilisin inhibitor (SSI) from Streptomyces albogriseolus S-3253 was cloned into E. coli plasmid pBR322 using two oligodeoxyribonucleotides corresponding to Asp68 to Pro77 and Asn99 to Gly107 of the protein, respectively. The SSI gene was localized on a 1.8-kbp BglII/SalI fragment. The nucleotide sequence of this 1.8-kbp fragment was determined by the dideoxy sequencing method. The amino acid sequence of the mature SSI coding region derived from the nucleotide sequence determination corresponded exactly to that from protein sequencing analysis. The nucleotide sequence analysis showed the presence of a putative signal peptide comprising 31 amino acids preceding the mature SSI region. The major transcriptional start point was identified to be 60 nucleotides upstream from the putative initiation codon for translation by the primer extension method. The -45 to -25 region upstream from transcriptional start point was quite homologous to that of CTC promoter of Bacillus subtilis. The overall G + C content of this 1.8-kbp fragment was 72%. On the other hand, an extremely high G + C content (96%) was found at the third letter of codons in the SSI coding region.

High-level expression in Streptomyces lividans 66 of a gene encoding Streptomyces subtilisin inhibitor from Streptomyces albogriseolus S-3253.

A secretory expression system for Streptomyces subtilisin inhibitor (SSI) was established in a heterologous host, Streptomyces lividans 66, by introducing the 1.8-kbp BglII/SalI fragment containing SSI gene into the Streptomyces multicopy vector, pIJ 702. The expression of SSI did not depend on the orientation of the 1.8-kbp BglII/SalI fragment or on the promoter for tyrosinase gene (mel) in pIJ 702, which suggested that this fragment also carries the SSI promoter. The expressed SSI in S.lividans 66 was secreted into the culture medium in a large amount, as observed with the original strain, S. albogriseolus S-3253. Amino acid sequence analysis showed that the SSI secreted from S. lividans 66 contained three additional amino acid residues in the NH2-terminal region. The inhibitory activity toward subtilisin BPN' and the antigenic activity of the SSI secreted from S. lividans 66 were found to be identical with those of authentic SSI.

Thermostable farnesyl diphosphate synthase of Bacillus stearothermophilus: molecular cloning, sequence determination, overproduction, and purification.

The structural gene for thermostable farnesyl diphosphate synthase from Bacillus stearothermophilus was cloned, sequenced, and overexpressed in Escherichia coli cells. A 1,260-nucleotide sequence of the cloned fragment was determined. This sequence specifies an open reading frame of 891 nucleotides for farnesyl diphosphate synthase. The deduced amino acid sequence shows a 42% similarity with that of E. coli FPP synthase [Fujisaki et al. (1990) J. Biochem. 108, 995-1000]. Comparison with prenyltransferases from a wide range of organisms, from bacteria to human, revealed the presence of seven highly conserved regions. In contrast to thermolabile prenyltransferases, which have four to six cysteine residues, the thermostable farnesyl diphosphate synthase carries only two cysteine residues. This enzyme is also unique in that some of the amino acids that are fully conserved in equivalents from other sources are replaced by functionally different amino acids. Construction of an overproducing strain provided a sufficient supply of this enzyme and it was purified to homogeneity. The purified recombinant enzyme is immunochemically identical with the native B. stearothermophilus enzyme, and it is not inactivated even after treatment at 65 degrees C for 70 min.

Thermostable farnesyl diphosphate synthase of Bacillus stearothermophilus: crystallization and site-directed mutagenesis.

The gene for thermostable farnesyl diphosphate synthase from Bacillus stearothermophilus was cloned, sequenced, and overexpressed in Escherichia coli. The synthase was purified to homogeneity and crystallized. The enzyme carried only two cysteine residues in contrast to its counterparts from other sources, which have four to six cysteine residues. Either or both of the cysteine residues can be replaced with serine without causing a loss of the catalytic activity. The conserved arginine residue that occupies the third position from the C-terminus was also replaced with valine without significant loss of activity, but the valine mutant showed a weakened affinity for isopentenyl diphosphate.

Spontaneous submucosal dissection of the esophagus.

A 41-year-old woman was found to have had spontaneous submucosal dissection of the esophagus by esophagoscopy. Food ingestion was postulated to have been an initiating factor in this instance. Conservative therapy was sufficient in this case. Patients who complain of sore throat with retrosternal pain and an obstructed sensation on swallowing or mild hematemesis should be examined by esophagoscopy for evidence of this disorder.

[An epidemiological study for fungus isolation during the twenty-five year periods from 1976 to 2000 in Kitasato University Hospital].

We investigated an epidemiological study for fungus isolation in our hospital from 1976 to 2000. For 25 years, the total sample number of fungus examination were 64,296, and after 1988, the total sample number increased suddenly. As a whole, the positive ratio was constantly about 40%. When our hospital opened, the obstetrical and gynecological samples showed 38.8% for fungus examination, but recently, samples of the respiratory organ has increased. Ratio of isolation for yeast, Candida albicans was 53.8%, and another yeasts such as Candida glabrata, Candida tropicalis, Candida parapsilosis were 12.5%, 5.3%, and 3.4%, respectively. Recently, isolation of Candida glabrata showed a tendency to increase. For genus Aspergillus, Aspergillus fumigatus was isolated, 48.1%, and Aspergillus nigar, Aspergillus terreus were isolated, 31.4% and 7.5%, respectively. For dermatophytes, Trichophyton rubrum was isolated, 63.6% indermatophytes, and another dermatophytes were Microsporum canis (17.9%), and Trichophyton mentagrophytes (15.9%), respectively. For dermatophytes, isolation of Microsporum canis showed a tendency to increase. Recently, the plural number of species showed a tendency to increase in the samples. Compared with the number of samples at the beginning in our hospital, the plural number of species in the samples increased about six times.

The alcohol tests for drunken criminals using psychological tests.

In the alcohol test for drunken criminals, we introduced Bender-Gestalt test and Rorschach test for the assessment, and examined their usefulness for the evaluation of intoxication patterns according to Binder's classification. The subjects were 24 drunken criminals who were examined by Mental Hygiene Group, Tsukuba University, for psychiatric evidences. The subjects were divided into the ordinary intoxication group (OI group) and the complicated intoxication group (CI group) on the basis of the behavioral assessment, and the psychological tests were performed before and after drinking. The following results were obtained. 1) Alcohol intoxication induced decrease in R1T, W and VIII + IX + X/R and increase of BGT scores and P%, which indicates that subjects become unable to make comprehensive and delicate responses to the external stimuli. 2) When we classified subjects into increasing and decreasing type on the pattern of changes in the BGT score from before to immediately after drinking in each subject, we found the ratio of increasing type in complicated intoxication is more than in ordinary intoxication significantly. And we found significant group x drinking interaction in F+% and At% of Rorschach test. The F+% significantly decreased only in CI group. The At% in CI group tended upward, but downward in OI group. These findings indicated that complicated intoxication reduced the subject's reality testing, while not in ordinary intoxication. 3) Comparing the effects of personality and intoxication factors in complicated intoxication, intoxication factors were considered to play primary roles. 4) We found association between high BAL and reduction of ego function and imagination, which is represented as significant peak of BALx drinking interaction in the BGT scores, M and FM + m. These observations suggest that the psychological tests as part of the alcohol tests are useful for the evaluation and research of intoxication.

[Plasma concentration of polymorphonuclear leukocyte elastase-alpha 1-proteinase inhibitor complex in patients with gastric ulcer].

To test the possible participation of activated leukocytes in the pathogenesis of gastric ulcer, plasma levels of polymorphonuclear leukocyte-elastase-alpha 1-proteinase inhibitor complex (PMN-EC), a discerning indicator for the activation of leukocytes, were determined in patients with gastric ulcer. Plasma levels of PMN-EC in patients with gastric ulcer (185.2 +/- 21.6 micrograms/l, mean +/- S.E.M.) were significantly higher than those in normal healthy subjects (67.6 +/- 4.4 micrograms/l, p less than 0.01). On the other hand, plasma levels of C-reactive protein and sialic acid increased in only 17.8% and 7.1% of patients with gastric ulcer, respectively. No significant correlation was observed between PMN-EC and the plasma levels of CRP or sialic acid. Plasma levels of PMN-EC decreased with the healing of the ulcerative lesion as judged by gastro-endoscopic observation. These findings suggested that activation of leukocytes might deeply be correlated with the pathologic mechanism of gastric ulceration and the measurement of PMN-EC might be useful for monitoring the patients with gastric ulcer.

[Isospora belli infection in a patient with adult T-cell leukemia].

An adult T cell leukemia (ATL) accompanied with Isospora belli infection was described. A 65-year-old male was admitted to our hospital because of a two month history of watery diarrhea. On admission, physical examination showed slight pallor but no detectable superficial lymphadenopathies. Hepatosplenomegaly was not observed. Laboratory examination revealed a leukocyte count 5,500/microliters with 10% abnormal lymphoid cells. A majority of the abnormal lymphoid cells expressed both CD 4 and CD 8 antigens. The patient was diagnosed as chronic ATL, since anti-HTLV-1 antibody in his serum and monoclonal integration of HTLV-1 proviral DNA in his peripheral mononuclear cells were detected. Isospora belli was found in his feces thereafter, and trimethoprim/sulfamethoxazole was effective for diarrhea. In Japan, there have been only 9 reported cases of lymphoproliferative disorders (including five ATL patients) accompanied with Isospora belli infection. From the descriptions in those reports, these 9 cases might all be ATL patients.

[Bronchial arterial infusion of lymphokine-activated killer cells stimulated by autologous tumor cells].

Bronchial arterial infusion (BAI) of Lymphokine activated killer (LAK) cells, stimulated in vitro by autologous tumor cells, was performed for a primary lung cancer patient, a 47-year-old male patient with primary squamous cell carcinoma of the lung who underwent probe thoracotomy and had part of the tumor tissue removed. Peripheral blood lymphocytes were cultured in 500 U/ml of recombinant IL-2(Shionogi Pharm. S6820) for 9 days after in vitro stimulation with mitomycin C-treated primary tumor cells for 3 days. These cells (designated St-LAK cells) were inoculated from the bronchial artery of patients who received 60 Gy irradiation of the primary site and mediastinal lymph nodes. The tumor regressed significantly from 8 X 7 to 3 X 2.5 cm in diameter. The advantages of LAK-BAI using St-LAK cells with irradiation were discussed.

Alteration of the specificity of the Streptomyces subtilisin inhibitor by gene engineering.

We have altered the amino acid at the center of the reactive site (methionine 73) of Streptomyces subtilisin inhibitor (SSI) by site-directed and cassette mutagenesis. Replacement by lysine or arginine resulted in trypsin inhibitory activity, replacement only by lysine gave inhibition of lysyl endopeptidase, and replacement by tyrosine or tryptophan resulted in inhibition of alpha-chymotrypsin. The four mutant SSIs retained their native activity against subtilisin BPN'. Thus by altering only one amino acid residue at the reactive site of SSI to the substrate specificity of the respective protease we could successfully change its inhibitory profile.