Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs.

Only a small proportion of the mouse genome is transcribed into mature messenger RNA transcripts. There is an international collaborative effort to identify all full-length mRNA transcripts from the mouse, and to ensure that each is represented in a physical collection of clones. Here we report the manual annotation of 60,770 full-length mouse complementary DNA sequences. These are clustered into 33,409 'transcriptional units', contributing 90.1% of a newly established mouse transcriptome database. Of these transcriptional units, 4,258 are new protein-coding and 11,665 are new non-coding messages, indicating that non-coding RNA is a major component of the transcriptome. 41% of all transcriptional units showed evidence of alternative splicing. In protein-coding transcripts, 79% of splice variations altered the protein product. Whole-transcriptome analyses resulted in the identification of 2,431 sense-antisense pairs. The present work, completely supported by physical clones, provides the most comprehensive survey of a mammalian transcriptome so far, and is a valuable resource for functional genomics.

Specific detection of circulating tumor cells by reverse transcriptase-polymerase chain reaction of a beta-casein-like protein, preferentially expressed in malignant neoplasms.

Beta-casein-like protein (BCLP) is a putative protein on cervical cancer and exhibits immunological characteristics similar to those of bovine beta-casein. We evaluated if BCLP mRNA detection in the blood is useful in gynecologic malignancies. We examined 30 patients with uterine cancer, nine cultured cancer cell lines and 26 healthy women volunteers. From these study populations and samples, total RNA was obtained. Reverse transcriptase-PCR (RT-PCR) of BCLP was performed on each sample. Eighteen (60.0%) patients and 4 (15.4%) volunteers were positive for BCLP mRNA. The RT-PCR reached sensitivity and specificity of 60.0% and 84.6%, respectively. Of eight patients having diagnosed recurrence, 7 (87.5%) were positive for BCLP mRNA. In patients with recurrence, sensitivity and specificity were 87.5% and 50%, respectively. The expression of mRNA showed a correlation with recurrence, but no correlation with metastasis or histological type. BCLP was specifically expressed in cancer cells and might be an aid in clinical diagnosis.

Computational analysis of full-length mouse cDNAs compared with human genome sequences.

Although the sequencing of the human genome is complete, identification of encoded genes and determination of their structures remain a major challenge. In this report, we introduce a method that effectively uses full-length mouse cDNAs to complement efforts in carrying out these difficult tasks. A total of 61,227 RIKEN mouse cDNAs (21,076 full-length and 40,151 EST sequences containing certain redundancies) were aligned with the draft human sequences. We found 35,141 non-redundant genomic regions that showed a significant alignment with the mouse cDNAs. We analyzed the structures and compositional properties of the regions detected by the full-length cDNAs, including cross-species comparisons, and noted a systematic bias of GENSCAN against exons of small size and/or low GC-content. Of the cDNAs locating the 35,141 genomic regions, 3,217 did not match any sequences of the known human genes or ESTs. Among those 3,217 cDNAs, 1,141 did not show any significant similarity to any protein sequence in the GenBank non-redundant protein database and thus are candidates for novel genes.

[A case of sepsis by Neisseria meningitidis beginning with pneumonia during a trip abroad].

A 62-year-male presented a high fever and a dry cough during a trip to Australia. He was admitted to a hospital as soon as be returned to Japan. The next day after returning to Japan, he was transferred to our hospital with septic shock and loss of consciousness. Neisseria meningitidis was cultured from his blood. N. meningitidis is rare in Japan. However its seems common, in some foreign countries. With these findings, it can be postulated that N. meningitidis might be one of the etiological agents of the imported infectious disease.

Characterization of silica-supported Ni catalysts effective for methane decomposition by Ni K-edge XAFS.

The structural change of Ni species during the methane decomposition into hydrogen and carbon over Ni/SiO2 catalyst was investigated by Ni K-edge XANES/EXAFS. Before the contact of methane with the Ni/SiO2 catalyst, Ni species were present as Ni metal mainly. The structure of the Ni metal did not change appreciably when the Ni/SiO2 was actively decomposing methane. In contrast, the formation of nickel carbide species was observed at the deactivation stage of the catalyst.

Cloning and characterization of a tumor-associated antigen, beta-casein-like protein.

Monoclonal antibody (MAb) 1C5 reacts with 87% of uterine cervical adenocarcinoma and bovine beta -casein, but not with squamous cell carcinoma. To clarify the characteristics of the antigen (beta-casein-like protein; BCLP) recognized by MAb 1C5, molecular cloning was performed using 5' rapid amplification of the cDNA ends (5' RACE) and the oligo-capping method. The protein predicted from the cDNA consisting of 937 nucleotides comprises 222 amino acids. The BCLP gene and deduced amino acid sequences were novel and showed no similarity to known cancer-associated genes in the database. Northern blot analysis showed that a 1.1 kb transcript was ubiquitously expressed in cancer cell lines and was predominantly expressed in uterine cervical adenocarcinoma. To clarify the function of BCLP, BCLP cDNA was transfected into L929 cells, resulting in a significant increase in cell area, a downregulation of cell growth rate and a decrease in cell attachment. We conclude that BCLP might be associated with cell morphology and a regulation of growth pattern of tumor.

Differential expression of major gap junction proteins, connexins 26 and 32, in rat mammary glands during pregnancy and lactation.

We examined the expression pattern of two major gap junction proteins, connexin 26 (Cx26) and connexin 32 (Cx32), in rat mammary glands during pregnancy and lactation. Immunohistochemically the two different Cxs were coexpressed in acinar cells and were independently modulated according to the physiological cell activity. Western blot analysis demonstrated that Cx26 gradually increased from early pregnancy, while Cx32 rapidly and dramatically increased at 16 h after parturition, and that both Cxs reached a maximum early in lactation. Increased expression of both Cxs was confirmed by Northern blot analysis showing that their mRNA transcripts were significantly induced on the day of parturition. We also analyzed double-immunofluorescent staining for Cx26 and Cx32 on a confocal laser scanning microscope, in order to examine colocalization of these Cxs in situ. Cx26 immunoreactivity mostly overlapped with Cx32-positive sites in acinar cells of lactating mammary glands, indicating that both Cxs were colocalized together in the same gap junctional plaques in lactation. These results suggest that upregulation of Cx26 and Cx32 in acinar cells at lactating stages, with colocalization in the same gap junctional plaques, may be important for control of secretion by acinar cells in rat mammary glands.

Relocalization of apoptosis-inducing factor in photoreceptor apoptosis induced by retinal detachment in vivo.

Apoptosis-inducing factor (AIF) is a novel mediator in apoptosis. AIF is a flavoprotein that is normally confined to the mitochondrial intermembrane space, yet translocates to the nucleus in several in vitro models of apoptosis. To investigate the role of AIF in the apoptotic process in vivo, we induced retinal detachment (RD) by subretinal injection of sodium hyaluronate, either in Brown Norway rats or in C3H mice. Apoptotic DNA fragmentation, as determined by terminal nick-end labeling, was most prominent 3 days after RD. The subcellular localization of AIF was examined by immunohistochemistry and immunoelectron microscopy. In normal photoreceptor cells, AIF was present in the mitochondrion-rich inner segment. However, AIF was found in the nucleus after RD. Photoreceptor apoptosis developed similarly in C3H control mice, and in mice bearing the gld or lpr mutations, indicating that cell death occurs independently from the CD95/CD95 ligand system. Both the mitochondrio-nuclear transition of AIF localization and the nuclear DNA fragmentation were inhibited by subretinal application of brain-derived neurotrophic factor. To our knowledge, this is the first description of AIF relocalization occurring in a clinically relevant, in vivo model of apoptosis.

Functional annotation of a full-length mouse cDNA collection.

The RIKEN Mouse Gene Encyclopaedia Project, a systematic approach to determining the full coding potential of the mouse genome, involves collection and sequencing of full-length complementary DNAs and physical mapping of the corresponding genes to the mouse genome. We organized an international functional annotation meeting (FANTOM) to annotate the first 21,076 cDNAs to be analysed in this project. Here we describe the first RIKEN clone collection, which is one of the largest described for any organism. Analysis of these cDNAs extends known gene families and identifies new ones.

Increased expression of angiotensin-converting enzyme in retinas of diabetic rats.

PURPOSE: The aim of this study was to examine the localization and the changes in the amount of angiotensin-converting enzyme (ACE) and the relationship between the renin-angiotensin (RA) system and vascular endothelial growth factor (VEGF)/VEGF-receptor system in the retinas of diabetic rats. METHODS: Immunohistochemical localization of ACE, VEGF, and VEGF-receptor fetal liver kinase-1 (Flk-1) was examined in cryosections of the retinas of streptozotocin-injected diabetic rats. A semi-quantitative comparison of diabetic rats with age-matched controls was also performed by counting the ACE- or Flk-1-positive vessels per microscopic field. RESULTS: ACE immunoreactivity was localized in the retinal vessel walls, and the percentages of ACE-positive vessels were significantly increased in the retinas of diabetic rats maintained 3 to 5 months. Both VEGF and Flk-1 signals increased simultaneously with the increment of ACE immunoreactivity. CONCLUSIONS: ACE, expressed in the retinal vessel walls, increases simultaneously with the increment of both VEGF and Flk-1 in the retinas of diabetic rats, suggesting that upregulation of ACE might play some role in the progression of diabetic retinopathy through the VEGF/VEGF receptor system.

[A case of lateral origin of the left internal mammary artery].

A 66 years old men, with left anterior descending coronary artery (LAD) stenosis and aortic valve stenosis, underwent coronary artery bypass grafting (CABG) to LAD with left internal mammary artery (LIMA) and aortic valve replacement. His postoperative course was uneventful. But, postoperative angiogram showed that his patent LIMA graft was originated from much lateral side of the left subclavian artery. Internal mammary artery is considered the most ideal graft for CABG, but its' anomality is not well known. We present this rare case with the anomalous origin of LIMA.

Surgical management of genital prolapse: is chain cystourethrography useful for evaluating anatomical corrections and urinary symptoms after surgery?

OBJECTIVE: To evaluate the relationship between chain cystourethrography and surgery for genital prolapse and urinary symptoms. METHODS: A retrospective study of women with genital prolapse was conducted from January 1991 through December 1997. To assess the preoperative and postoperative anatomical situations objectively, chain cystourethrography was performed. RESULTS: Fifty-seven patients underwent surgical repairs for genital prolapse. All of them recovered from the genital prolapse both subjectively and objectively after surgery. Chain cystourethrography is useful for evaluating postoperative anatomical corrections. However, 6 patients (12%) suffered from a recurrent feeling of genital prolapse, and 14 patients (28%) had urinary symptoms postoperatively. CONCLUSIONS: Our treatment showed good anatomical corrections based on chain cystourethrography, and there were no prognostic differences among the surgical procedures. However, anatomical correction does not always mean improvement of urinary symptoms. Further studies are needed to clarify what factors contribute to the prognosis in such cases.

[Anesthetic management for a patient with autoimmune hemolytic anemia].

A 69-year-old woman with autoimmune hemolytic anemia (AIHA) received gastric resection and splenectomy under general anesthesia combined with epidural anesthesia. Her blood type had been determined as type A, Rh type CCDee, with anti-e-antibody and anti-pdl-antibody positive as autoantibodies. We applied the technique of hemodilutional autologous transfusion to supplement the blood loss during the operation to prevent hemolysis or occurrence of new antibody after homologous transfusion. We should pay much attention to a patient with AIHA to find signs of peri-operative hemolysis and to give the treatment for hemolysis in early stage. We consider that the hemodilutional autologous transfusion is a useful technique for the anesthetic management of the patient having a rare blood type.

Complete hydatidiform mole coexisting with a live fetus after gamete intrafallopian transfer. A case report.

BACKGROUND: Molar pregnancy with a coexisting live fetus is a rare occurrence. We report the only known case with a surviving coexistent fetus after gamete intrafallopian transfer (GIFT). CASE: After GIFT, a 28-year-old primary infertility patient was diagnosed as having a complete hydatidiform mole coexisting with a live fetus at 13 weeks of gestation. At 36 weeks of gestation, a cesarean section was performed due to elevated serum human chorionic gonadotropin (hCG) levels, and a male infant with a normal appearance and weighing 2,688 g was delivered. CONCLUSION: If the patient desires to try to carry the fetus to viability after counseling on the possible associated risks of malignancy, it is possible to achieve fetal viability if (1) there is decline in the serum hCG level after it peaks before the second trimester, (2) ultrasound reveals degeneration of the molar part, and (3) there are no complications of pregnancy.

[Anesthetic management in a patient with severe acute pancreatitis during pregnancy].

Continuous epidural anesthesia was used in a 34 year-old pregnant woman with acute pancreatitis related to hypertriglyceridemia. She underwent an emergency cesarean section due to severe pancreatitis under spinal anesthesia. After delivery, extended incision was made to examine the pancreas and to perform drainage. Epidural infusion using 1% mepivacaine and buprenorphine was started to reduce pain and improve microcirculation. After starting epidural infusion with other therapies, clinical feature and data improved. This case suggests that reduction of severe pain and improvement of microcirculation are important in therapies of severe pancreatitis.

Characterization of subclonal cell lines showing high motility derived from an undifferentiated human endometrial cancer cell line.

To examine the relationship between the morphology and the motility of cancer cells, we studied the difference between human endometrial undifferentiated cancer cell lines with different motilities, using light and electron microscopy, immunocytochemistry and RT-PCR. In a transplanted tumor of a cell line with high motility, many connective tissues and microvessels were observed and there were few intercellular spaces, which were ultrastructurally full of extracellular matrix. In the high-motility cell line only cytokeratin 19 was observed, and the level of cytokeratin 19 mRNA was elevated. Thus, motility is suggested to be related to the induction of connective tissues and cytoskeleton.

Utility of colorimetric end-tidal carbon dioxide detector for monitoring during prehospital cardiopulmonary resuscitation.

The purpose of this study was to evaluate a colorimetric end-tidal CO2 (ETCO2) detector (EASY CAP) as a monitor during prehospital cardiopulmonary resuscitation (CPR) without tracheal intubation. This detector was used for 121 patients during CPR with a laryngeal mask airway or face mask by authorized emergency lifesaving technicians. At 7 to 15 minutes after the initiation of CPR, ETCO was <0.5% in 30 cases (group A), 0.5% to 2.0% in 46 cases (group B) and >2.0% in 45 cases (group C). The rate of return of spontaneous circulation was 17% in group A, 24% in group B, and 48% in group C (groups A v C, P < .01). There was a significant difference in the rate of hospital admission between groups A and C. The ETCO2 value may be useful for monitoring during prehospital CPR with a laryngeal mask airway or face mask.

[Right phrenic nerve palsy in a patient with ventricular septal perforation after acute myocardial infarction].

A 77-year-old female had acute myocardial infarction and ventricular septal perforation (VSP). An operation was performed 14 days after VSP. During cardiopulmonary bypass, a patch was sutured in place on the left side of the defect under a mild hypothermia with topical cardiac cooling procedure. In the postoperative period, mechanical ventilation was continued with use of pulmonary monitor (CP-100, BICORE). When the patient was weaned of mechanical ventilation, work of breathing increased (WOBp: 1.67 J.l-1) without the change of SaO2, PaO2, PaCO2 and the respiratory state. After extubation, the chest X-ray showed elevated right diaphragmatic level and a diagnosis of unilateral diaphragmatic paralysis was made. Oral intake was started two days after extubation. SpO2 decreased after oral intake, and it was significantly improved by taking a left lateral position. The symptoms disappeared 47 days after the operation. It should be noticed that the rise of endoceliac pressure in the patient who had a unilateral phrenic nerve palsy, affected the SpO2.