The aim of this study was to assess clinical and pulmonary thin-section CT findings in patients with acute Pseudomonas aeruginosa (PA) pulmonary infection.
We retrospectively identified 44 patients with acute PA pneumonia who had undergone chest thin-section CT examinations between January 2004 and December 2010. We excluded nine patients with concurrent infections. The final study group comprised 35 patients (21 males, 14 females; age range 30–89 years, mean age 66.9 years) with PA pneumonia. The patients' clinical findings were assessed. Parenchymal abnormalities, enlarged lymph nodes and pleural effusion were evaluated on thin-section CT.
Underlying diseases included malignancy (n=13), a smoking habit (n=11) and cardiac disease (n=8). CT scans of all patients revealed abnormal findings, including ground-glass opacity (n=34), bronchial wall thickening (n=31), consolidation (n=23) and cavities (n=5). Pleural effusion was found in 15 patients.
PA pulmonary infection was observed in patients with underlying diseases such as malignancy or a smoking habit. The CT findings in patients with PA consisted mainly of ground-glass attenuation and bronchial wall thickening.
Advances in knowledge
The CT findings consisted mainly of ground-glass attenuation, bronchial wall thickening and cavities. These findings in patients with an underlying disease such as malignancy or a smoking habit may be suggestive of pneumonia caused by PA infection.
The aim of this study was to compare the pulmonary thin-section CT findings of patients with acute Streptococcus pneumoniae pneumonia with and without concurrent infection.
The study group comprised 86 patients with acute S. pneumoniae pneumonia, 36 patients with S. pneumoniae pneumonia combined with Haemophilus influenzae infection, 26 patients with S. pneumoniae pneumonia combined with Pseudomonas aeruginosa infection and 22 patients with S. pneumoniae pneumonia combined with methicillin-susceptible Staphylococcus aureus (MSSA) infection. We compared the thin-section CT findings among the groups.
Centrilobular nodules and bronchial wall thickening were significantly more frequent in patients with pneumonia caused by concurrent infection (H. influenzae: p<0.001 and p<0.001, P. aeruginosa: p<0.001 and p<0.001, MSSA: p<0.001 and p<0.001, respectively) than in those infected with S. pneumoniae alone. Cavity and bilateral pleural effusions were significantly more frequent in cases of S. pneumoniae pneumonia with concurrent P. aeruginosa infection than in cases of S. pneumoniae pneumonia alone (p<0.001 and p<0.001, respectively) or with concurrent H. influenzae (p<0.05 and p<0.001, respectively) or MSSA infection (p<0.05 and p<0.05, respectively).
When a patient with S. pneumoniae pneumonia has centrilobular nodules, bronchial wall thickening, cavity or bilateral pleural effusions on CT images, concurrent infection should be considered.
The aim of this study was to evaluate the normal anatomy of the thoracic duct and cisterna chyli obtained by axial and multiplanar reformation (MPR) images of 1 mm slice thickness using multidetector row CT (MDCT).
We evaluated the ability of MDCT to examine the normal anatomy of the thoracic duct and cisterna chyli. The axial and coronal images of thoracoabdominal MDCT images obtained in 50 patients (20 females and 30 males; mean age, 63.5 years; range, 32–81 years) were reviewed between January and October 2005. We excluded patients with malignant neoplasms, inflammation or vascular diseases (e.g. aortic aneurysm, aortic dissection) and those with a history of thoracoabdominal surgery. The thoracic duct was divided into three anatomical sections: the upper, middle and lower. We evaluated the degree of visualisation and the maximum size of the thoracic duct. We also evaluated the degree of visualisation, maximum size, configuration and location of the cisterna chyli.
Visualisation of the thoracic duct and cisterna chyli was almost 100% on axial and coronal images. The lower section of the thoracic duct was most clearly visualised among the three sections. There was little difference in the maximum size of the thoracic duct among the three sections. The cisterna chyli was most frequently located at the Th12 or L1 level, and the most common type was the “straight thin tube type”.
Axial and MPR images of 1 mm slice thickness using MDCT can clearly depict the thoracic duct and cisterna chyli.
The purpose of this study was to compare the clinical and thin-section CT findings in patients with meticillin-resistant Staphylococcus aureus (MRSA) and meticillin-susceptible S. aureus (MSSA).
We retrospectively identified 201 patients with acute MRSA pneumonia and 164 patients with acute MSSA pneumonia who had undergone chest thin-section CT examinations between January 2004 and March 2009. Patients with concurrent infectious disease were excluded from our study. Consequently, our study group comprised 68 patients with MRSA pneumonia (37 male, 31 female) and 83 patients with MSSA pneumonia (32 male, 51 female). Clinical findings in the patients were assessed. Parenchymal abnormalities, lymph node enlargement and pleural effusion were assessed.
Underlying diseases such as cardiovascular were significantly more frequent in the patients with MRSA pneumonia than in those with MSSA pneumonia. CT findings of centrilobular nodules, centrilobular nodules with a tree-in-bud pattern, and bronchial wall thickening were significantly more frequent in the patients with MSSA pneumonia than those with MRSA pneumonia (p=0.038, p=0.007 and p=0.039, respectively). In the group with MRSA, parenchymal abnormalities were observed to be mainly peripherally distributed and the frequency was significantly higher than in the MSSA group (p=0.028). Pleural effusion was significantly more frequent in the patients with MRSA pneumonia than those with MSSA pneumonia (p=0.002).
Findings from the evaluation of thin-section CT manifestations of pneumonia may be useful to distinguish between patients with acute MRSA pneumonia and those with MSSA pneumonia.
We have previously reported that intrarenal angiotensin II (Ang II) levels are increased long before diabetes becomes apparent in obese Otsuka-Long-Evans-Tokushima-Fatty (OLETF) rats, a model of type 2 diabetes. In this study, we examined the changes in intrarenal renin-angiotensin system (RAS) activity in the developing kidneys of OLETF rats. Ang II contents and mRNA levels of RAS components were measured in male OLETF and control Long-Evans Tokushima (LETO) rats at postnatal days (PND) 1, 5, and 15, and at 4–30 weeks of age. In both LETO and OLETF rats, kidney Ang II levels peaked at PND 1, then decreased during the pre- and post-weaning periods. However, Ang II levels and gene expression of RAS components, including angiotensinogen (AGT), renin, and angiotensin-converting enzyme (ACE), were not significantly different between LETO and OLETF rats. Intrarenal Ang II contents further decreased during puberty (from 7 to 11 weeks of age) in LETO rats, bur not in OLETF rats. At 11 weeks of age, kidney Ang II levels, urinary AGT excretion, and mRNA levels of AGT and renin were higher in OLETF rats than in LETO rats, while blood glucose levels were not significantly different between these groups of rats. These data indicate that continued intrarenal expression of Ang II during pubescence contributes to the increases in intrarenal Ang II levels in prediabetic OLETF rats, and is associated with increased intrarenal AGT and renin expression. Inappropriate activation of the intrarenal RAS in the prediabetic stage may facilitate the onset and development of diabetic nephropathy in later life.
angiotensin II; developing kidney; diabetes; angiotensinogen
The aim of this study was to assess pulmonary thin-section CT findings in patients with acute Haemophilus influenzae pulmonary infection.
Thin-section CT scans obtained between January 2004 and March 2009 from 434 patients with acute H. influenzae pulmonary infection were retrospectively evaluated. Patients with concurrent infection diseases, including Streptococcus pneumoniae (n=76), Staphylococcus aureus (n=58) or multiple pathogens (n=89) were excluded from this study. Thus, our study group comprised 211 patients (106 men, 105 women; age range, 16–91 years, mean, 63.9 years). Underlying diseases included cardiac disease (n=35), pulmonary emphysema (n=23), post-operative status for malignancy (n=20) and bronchial asthma (n=15). Frequencies of CT patterns and disease distribution of parenchymal abnormalities, lymph node enlargement and pleural effusion were assessed by thin-section CT.
The CT findings in patients with H. influenzae pulmonary infection consisted mainly of ground-glass opacity (n=185), bronchial wall thickening (n=181), centrilobular nodules (n=137) and consolidation (n=112). These abnormalities were predominantly seen in the peripheral lung parenchyma (n=108). Pleural effusion was found in 22 patients. Two patients had mediastinal lymph node enlargement.
These findings in elderly patients with smoking habits or cardiac disease may be characteristic CT findings of H. influenzae pulmonary infection.
Moraxella catarrhalis is an important pathogen in the exacerbation of chronic obstructive pulmonary disease. The aim of this study was to assess the clinical and pulmonary thin-section CT findings in patients with acute M. catarrhalis pulmonary infection.
Thin-section CT scans obtained between January 2004 and March 2009 from 292 patients with acute M. catarrhalis pulmonary infection were retrospectively evaluated. Clinical and pulmonary CT findings in the patients were assessed. Patients with concurrent infection including Streptococcus pneumoniae (n = 72), Haemophilus influenzae (n = 61) or multiple pathogens were excluded from this study.
The study group comprised 109 patients (66 male, 43 female; age range 28–102 years; mean age 74.9 years). Among the 109 patients, 34 had community-acquired and 75 had nosocomial infections. Underlying diseases included pulmonary emphysema (n = 74), cardiovascular disease (n = 44) or malignant disease (n = 41). Abnormal findings were seen on CT scans in all patients and included ground-glass opacity (n = 99), bronchial wall thickening (n = 85) and centrilobular nodules (n = 79). These abnormalities were predominantly seen in the peripheral lung parenchyma (n = 99). Pleural effusion was found in eight patients. No patients had mediastinal and/or hilar lymph node enlargement.
M. catarrhalis pulmonary infection was observed in elderly patients, often in combination with pulmonary emphysema. CT manifestations of infection were mainly ground-glass opacity, bronchial wall thickening and centilobular nodules.
This study aimed to compare thin-section CT images from sarcoidosis patients who had either normal or elevated serum KL-6 levels.
101 patients with sarcoidosis who underwent thin-section CT examinations of the chest and serum KL-6 measurements between December 2003 and November 2008 were retrospectively identified. The study group comprised 75 sarcoidosis patients (23 male, 52 female; aged 19–82 years, mean 54.1 years) with normal KL-6 levels (152–499 U ml–1, mean 305.7 U ml–1) and 26 sarcoidosis patients (7 male, 19 female; aged 19–75 years, mean 54.3 years) with elevated KL-6 levels (541–2940 U ml–1, mean 802.4 U ml–1). Two chest radiologists, unaware of KL-6 levels, retrospectively and independently interpreted CT images for parenchymal abnormalities, enlarged lymph nodes and pleural effusion.
CT findings in sarcoidosis patients consisted mainly of lymph node enlargement (70/75 with normal KL-6 levels and 21/26 with elevated KL-6 levels), followed by nodules (50 and 25 with normal and elevated levels, respectively) and bronchial wall thickening (25 and 21 with normal and elevated levels, respectively). Ground-glass opacity, nodules, interlobular septal thickening, traction bronchiectasis, architectural distortion and bronchial wall thickening were significantly more frequent in patients with elevated KL-6 levels than those with normal levels (p<0.001, p<0.005, p<0.001, p<0.001, p<0.001 and p<0.001, respectively). By comparison, there was no significant difference in frequency of lymph node enlargement between the two groups.
These results suggest that serum KL-6 levels may be a useful marker for indicating the severity of parenchymal sarcoidosis.
Giant cell carcinoma of the lung is a very rare primary malignant tumour and localised right upper-lobe pulmonary oedema is also unusual. We report a case of giant cell carcinoma, which invaded the left atrium through the left pulmonary vein and caused localised right upper-lobe pulmonary oedema.
Human herpesvirus 6 (HHV-6)-associated encephalitis or pneumonitis has been reported in immunocompetent and immunosuppressed individuals. Several MRI studies in patients with HHV-6-associated encephalitis have been presented. However, to the best of our knowledge, no studies describing thin-section CT imaging in patients with HHV-6-associated pneumonitis have been reported. Here we describe a case of HHV-6-associated encephalitis and pneumonitis that developed after bone marrow transplantation. Thin-section CT images of the chest revealed ground-glass attenuation, consolidation and centrilobular nodules in both lungs.
The purpose of this study was to identify the clinical and thin-section CT findings in patients with acute Klebsiella pneumoniae pneumonia (KPP) alone and with concurrent infection. We retrospectively identified 160 patients with acute KPP who underwent chest thin-section CT examinations between August 1998 and August 2008 at our institution. The study group comprised 80 patients (54 male, 26 female; age range 18–97 years, mean age 61.5) with acute KPP alone, 55 (43 male, 12 female; age range 46–92 years, mean age 76.0) with KPP combined with methicillin-resistant Staphylococcus aureus (MRSA) and 25 (23 male, 2 female; age range 56–91 years, mean age 72.7) with KPP combined with Pseudomonas aeruginosa (PA). Underlying diseases in patients with each type of pneumonia were assessed. Parenchymal abnormalities were evaluated along with enlarged lymph nodes and pleural effusion. In patients with concurrent pneumonia, underlying conditions such as cardiac diseases, diabetes mellitus and malignancy were significantly more frequent than in patients with KPP alone. The mortality rate in patients with KPP combined with MRSA or PA was significantly higher than in those with KPP alone. In concurrent KPP, CT findings of centrilobular nodules, bronchial wall thickening, cavity, bronchiectasis, nodules and pleural effusion were significantly more frequent with concurrent pneumonia than in those with KPP alone.
The aim of this study was to evaluate the anatomy of and normal variations in the craniocervical junction veins. We retrospectively reviewed 50 patients who underwent contrast-enhanced CT with a multidetector scanner. Axial and reconstructed images were evaluated by two neuroradiologists with special attention being paid to the existence and size of veins and their relationships with other venous branches around the craniocervical junction. The venous structures contributing to craniocervical junction venous drainage, including the inferior petrosal sinus (IPS), transverse-sigmoid sinus, jugular vein, condylar vein, marginal sinus and suboccipital cavernous sinus were well depicted in all cases. The occipital sinus (OS) was identified in 18 cases, including 4 cases of prominent-type OS. The IPS showed variations in drainage to the jugular vein through the jugular foramen or intraosseous course of occipital bone via the petroclival fissure. In all cases, the anterior condylar veins connected the anterior condylar confluence to the marginal sinus; however, a number of cases with asymmetry and agenesis in the posterior and lateral condylar veins were seen. The posterior condylar vein connected the suboccipital cavernous sinus to the sigmoid sinus or anterior condylar confluence. The posterior condylar canal in the occipital bone showed some differences, which were accompanied by variations in the posterior condylar veins. In conclusion, there are some anatomical variations in the venous structures of the craniocervical junction; knowledge of these differences is important for the diagnosis and treatment of skull base diseases. Contrast-enhanced CT using a multidetector scanner is useful for evaluating venous structures in the craniocervical junction.
RECK is a novel tumour suppressor gene that negatively regulates matrix metalloproteinases (MMPs) and inhibits tumour invasion, angiogenesis and metastasis. In the present study, we investigated the effects of epigallocatechin-3-gallate (EGCG), a major polyphenol in green tea, on the methylation status of the RECK gene and cancer invasion in oral squamous cell carcinoma cell lines. Our results showed that treatment of oral cancer cells with EGCG partially reversed the hypermethylation status of the RECK gene and significantly enhanced the expression level of RECK mRNA. Inhibition of MMP-2 and MMP-9 levels was also observed in these cells after treatment with EGCG. Interestingly, EGCG significantly suppressed cancer cell-invasive ability by decreasing the number of invasive foci (P<0.0001) as well as invasion depth (P<0.005) in three-dimensional collagen invasion model. Although further investigation is required to assess the extent of contribution of RECK on MMPs to the suppression of invasive behaviour, these results support the conclusion that EGCG plays a key role in suppressing cell invasion through multiple mechanisms, possibly by demethylation effect on MMP inhibitors such as RECK.
green tea polyphenol; EGCG; RECK hypermethylation; oral cancer invasion
The hypoglossal canal contains a venous plexus that connects the inferior petrous sinus, condylar vein, jugular vein and paravertebral plexus. The venous plexus is one of the venous drainage routes of the posterior skull base. Only a few cases of dural arteriovenous fistulas (AVFs) involving the hypoglossal canal have been reported.
We describe three cases (a 62-year-old female, a 52-year-old male, and an 83-year-old male) of dural AVFs involving the hypoglossal canal. Symptoms were pulse-synchronous bruit in two cases and proptosis/chemosis in one. All dural AVFs were mainly fed by the ipsilateral ascending pharyngeal artery. Two of three dural AVFs involving the hypoglossal canal mainly drained through the anterior condylar confluence into the inferior petrosal sinus retrogradely with antegrade drainage through the lateral condylar vein. The other one drained through the lateral and posterior condylar veins into the suboccipital cavernous sinus. All dural AVFs were completely occluded by selective transvenous embolization without any complications, and the symptoms disappeared within one week in all cases.
Dural AVFs involving the hypoglossal canal can be successfully treated by selective transvenous embolization with critical evaluation of venous anatomy in each case.
dural AVShunt, posterior fossa
In the safety stenting, it is important to get to know the characteristics of a plaque. In petrous carotid artery stenosis, it is difficult to know the characteristics of the plaque. We paid our attention to the MPRAGE (Magnetization Prepared Rapid Acquisition with Gradient Echo) method on high resolving power MRI.
By the MPRAGE method, low intensity was observed in these lesions of all cases. This result suggested that the plaque in petrous portion was a fibrous plaque. This method is useful to get to know the characteristics of a plaque in petrous portion before endovascular treatment.
Petrous carotid stenosis, plaque characterization, MPRAGE
Background: Production of N-alpha-methyl-histamine (NAMH), a histamine H3 receptor (H3R) agonist, is reportedly promoted in Helicobacter pylori infected human gastric mucosa. NAMH was suggested to act directly on histamine H2 receptors (H2Rs) in animals to stimulate acid secretion and to be a H2R agonist. As H2Rs and H3Rs play different roles in gastric acid secretion, it is very important to verify that NAMH is a H2R agonist.
Aims: To determine whether NAMH is a H2R agonist, as well as a H3R agonist.
Methods: We used a Chinese hamster ovary (CHO) cell line expressing human H2Rs (CHO-H2R) and control CHO cells. Expression of human H2Rs was confirmed by tiotidine binding. cAMP production in CHO-H2R and control cells in response to histamine or NAMH was measured. cAMP production in response to 10−7 M NAMH was also measured in the presence or absence of the H2R antagonist famotidine and the H3R antagonist thioperamide.
Results: NAMH dose dependently stimulated cAMP productions in CHO-H2R cells. This production was inhibited by famotidine but not by thioperamide. Control CHO cells were unresponsive to either histamine or NAMH. In addition, the effect of NAMH, in terms of cAMP production in CHO-H2R cells, was more potent than that of histamine—that is, with a lower EC50 concentration and higher maximal cAMP production. Both NAMH and histamine, but not R-alpha-methyl-histamine, effectively inhibited [3H] tiotidine binding to CHO-H2R cells.
Conclusions: NAMH, which is produced in the gastric mucosa by H pylori, is a potent H2R agonist as well as a H3R agonist.
Chinese hamster ovary cells; histamine H2 receptors; histamine H3 receptors; N-alpha-methyl-histamine
Cerebellar long-term depression (LTD) at the parallel fiber-Purkinje cell synapses has been proposed to be a neural substrate for classical eyeblink conditioning. Mutant mice lacking the glutamate receptor subunit δ2 (GluRδ2), in which the cerebellar LTD is disrupted, exhibited a severe impairment in the delay eyeblink conditioning with a temporal overlap of CS and US. However, they learned normally trace and delay conditioning without CS-US overlap, suggesting a learning mechanism which does not require the cerebellar LTD.In the present study, we tested possible involvement of the hippocampus in this cerebellar LTD-independent learning. We examined effects of scopolamine and hippocampal lesion on the delay conditioning without CS-US overlap. TheGluRδ2 mutant mice that received scopolamine or aspiration of the dorsalhippocampus together with its overlying cortex exhibited a severe impairment in learning, while the control mutant mice that received saline or aspiration of the overlying cortex learned normally. In contrast, wild-type mice that received either treatment learned as normally as the control wild-type mice. These results suggest that the hippocampus is essential in the cerebellar LTD-independent learning in the GluRδ2 mutant mice, indicating a newrole of hippocampus in the paradigm with a short trace interval.
cerebellar LTD; eyeblink conditioning; gene-knockout mice; glutamate receptor subunit δ2; hippocampus; synaptic plasticity
A 44 year old man with Brugada syndrome and ventricular fibrillation had an autonomic disorder, shown by spectral analysis of heart rate variability and 123I-MIBG myocardial scintigraphy. Periodic variation of the ST segments was detected by Holter ECG. Increased high frequency power (0.15-0.40 Hz), an index of parasympathetic nerve activity, was observed just before ST segment elevation. In addition, local dysfunction of sympathetic nerves in the left ventricle was detected by 123I-MIBG myocardial scintigraphy. Unbalanced autonomic nerve function plays an important role in inducing Brugada-type ECG signs.
Keywords: Brugada syndrome; autonomic disorder; 123I-MIBG myocardial scintigraphy
The present study was designed to investigate the effect of Fas-mediated liver cell apoptosis, induced by a hamster monoclonal antibody against mouse Fas antigen, on diethylnitrosamine (DEN)-induced hepatocarcinogenesis in mice. DEN (10 μg g–1, intraperitoneally (i.p.)) was given to 15-day-old male C3H/HeJ mice. Three weeks after DEN treatment, Fas-mediated liver cell apoptosis induced by anti-Fas antibody resulted in a biphasic effect on induction of liver cell tumours, depending on dosage and time of antibody administration. Single or multiple treatment with high dose anti-Fas antibody (5 μg animal−1), induced gross liver cell damage and decreased the incidence of liver cell tumours in DEN-treated mice. In contrast, five treatments with low dose anti-Fas antibody (2 μg animal−1), induced dispersed localized liver cell damage and promoted the number of large-sized liver cell adenomas and hepatocellular carcinomas. These findings suggest that high dose anti-Fas antibody has a marked effect on the clearance of DEN-initiated liver cells, whereas repeated administration of low dose anti-Fas antibody promotes hepatocarcinogenesis. It is concluded that Fas-mediated liver cell apoptosis has a biphasic effect on hepatocarcinogenesis. © 2000 Cancer Research Campaign
Fas; apoptosis; diethylnitrosamine; mouse; hepatocarcinogenesis
Proline 3-hydroxylase was purified from Streptomyces sp. strain TH1, and its structural gene was cloned. The purified enzyme hydroxylated free L-proline to cis-3-hydroxy-L-proline and showed properties of a 2-oxoglutarate-dependent dioxygenase (H. Mori, T. Shibasaki, Y. Uosaki, K. Ochiai, and A. Ozaki, Appl. Environ. Microbiol, 62:1903-1907, 1996). The molecular mass of the purified enzyme was 35 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric point of the enzyme was 4.3. The optimal pH and temperature were 7.0 and 35 degrees C, respectively. The K(m) values were 0.56 and 0.11 mM for L-proline and 2-oxoglutarate, respectively. The Kcat value of hydroxylation was 3.2 s-1. Determined N-terminal and internal amino acid sequences of the purified protein were not found in the SwissProt protein database. A DNA fragment of 74 bp was amplified by PCR with degenerate primers based on the determined N-terminal amino acid sequence. With this fragment as a template, a digoxigenin-labeled N-terminal probe was synthesized by PCR. A 6.5-kbp chromosome fragment was cloned by colony hybridization with the labeled probe. The determined DNA sequence of the cloned fragment revealed a 870-bp open reading frame (ORF 3), encoding a protein of 290 amino acids with a calculated molecular weight of 33,158. No sequence homolog was found in EMBL, GenBank, and DDBJ databases. ORF 3 was expressed in Escherichia coli DH1. Recombinants showed hydroxylating activity five times higher than that of the original bacterium, Streptomyces sp. strain TH1. It was concluded that the ORF 3 encodes functional proline 3-hydroxylase.
During the screening of microbial proline hydroxylases, novel proline 3-hydroxylase activities, which hydroxylate free l-proline to free cis-3-hydroxy-l-proline, were detected in whole cells of Streptomyces sp. strain TH1 and Bacillus sp. strains TH2 and TH3 from 3,000 strains isolated from soil. The reaction product was purified from a reaction mixture of Streptomyces sp. strain TH1, and its chemical structure was identified as cis-3-hydroxy-l-proline by instrumental analyses. Proline 3-hydroxylase activity was also detected in Streptomyces canus ATCC 12647 which produces the 3-hydroxyproline-containing peptide antibiotic telomycin. Bacillus sp. strains TH2 and TH3 were found to accumulate cis-3-hydroxy-l-proline in culture media at 426 and 352 (mu)M, respectively. It was suggested that hydroxylation occurred in a highly regio- and stereospecific manner at position 3 of l-proline because no hydroxylation product other than cis-3-hydroxy-l-proline was observed. Proline 3-hydroxylases of these strains were first characterized on crude enzyme preparations. Since 2-oxoglutarate and ferrous ion were required for hydroxylation of l-proline, these 3-hydroxylases were thought to belong to a family of 2-oxoglutarate-related dioxygenases. The reaction was inhibited by Co(sup2+), Zn(sup2+), and Cu(sup2+). l-Ascorbic acid accelerated the reaction. The optimum pH and temperature were 7.5 and 35(deg)C, respectively.
A beta-mannanase (EC 220.127.116.11) from Vibrio sp. strain MA-138 was purified by ammonium sulfate precipitation and several chromatographic procedures including gel filtration, adsorption, and ion-exchange chromatographies. The final ion-exchange chromatography Mono Q yielded one major active fraction and three minor active fractions. The major active fraction was purified to homogeneity on the basis of native polyacrylamide gel electrophoresis (PAGE). This purified enzyme was identified as a glycoprotein by periodic acid-Schiff staining and a monomeric protein with a molecular mass of 49 kDa by sodium dodecyl sulfate-PAGE. The pI of the enzyme was 3.8. The purified enzyme exhibited maximal activity at pH 6.5 and 40 degrees C and hydrolyzed at random the internal beta-1,4-mannosidic linkages in beta-mannan to give various sizes of oligosaccharides. The first 20 N-terminal amino acid sequence of the purified enzyme showed high homology with the N-terminal region of beta-mannanase from Streptomyces lividans 66.
We attempted to clone an inosine kinase gene of Escherichia coli. A mutant strain which grows slowly with inosine as the sole purine source was used as a host for cloning. A cloned 2.8-kbp DNA fragment can accelerate the growth of the mutant with inosine. The fragment was sequenced, and one protein of 434 amino acids long was found. This protein was overexpressed. The overexpressed protein was purified and characterized. The enzyme had both inosine and guanosine kinase activity. The Vmaxs for guanosine and inosine were 2.9 and 4.9 mumol/min/mg of protein, respectively. The Kms for guanosine and inosine were 6.1 microM and 2.1 mM, respectively. This enzyme accepted ATP and dATP as a phosphate donor but not p-nitrophenyl phosphate. These results show clearly that this enzyme is not a phosphotransferase but a guanosine kinase having low (Vmax/Km) activity with inosine. The sequence of the gene we have cloned is almost identical to that of the gsk gene (K.W. Harlow, P. Nygaard, and B. Hove-Jensen, J. Bacteriol. 177:2236-2240, 1995).
The gene encoding an extracellular isomalto-dextranase, designated imd, was isolated from the chromosomal DNA of Arthrobacter globiformis T6 and cloned and expressed in Escherichia coli. A single open reading frame consisting of 1,926 base pairs that encoded a polypeptide composed of a signal peptide of 39 amino acids and a mature protein of 602 amino acids (M(r), 65,900) was found. The primary structure had no significant homology with the structures of any other reported carbohydrases, including two other dextranases. Transformed E. coli cells carrying the 2.3-kb fragment overproduced isomalto-dextranase into the periplasmic space under control of the promoter of the imd gene itself.
Inactivation of the tumour-suppressor gene p53 has been demonstrated in a variety of human tumours. We extracted DNA from paraffin-embedded tissues of 67 ovarian carcinoma samples (54 primary tumours, seven metastases and six tumours obtained after chemotherapy), and analysed allelic losses and mutations of the p53 gene using single-strand conformation polymorphism (SSCP) analysis of DNA fragments amplified by a polymerase chain reaction (PCR). Allelic loss was observed in 24 of 32 informative cases. The mutation was detected in 14 of 54 primary ovarian carcinomas: eight serous cystadenocarcinomas (SCA), 42%), five endometrioid adenocarcinomas (EA, 42%) and one mucinous cystadenocarcinoma (14%). The incidence of the alteration was higher in SCA and EA than in other histological types, but the difference was not statistically significant. The incidence of p53 gene abnormalities in ovarian carcinomas tended to be increased in patients with disease advanced (over FIGO stage II). Mutations were found in exons 5 and 7 only and consisted mainly of single nucleotide substitutions [9 or 14 (64%) in exon 7; 4 of 14 (29%) in exon 5]. In 13 of 14 cases, p53 gene mutations occurred concomitantly with losses of the normal allele. The status of the p53 gene in metastases and the tumours obtained after chemotherapy was identical to that in the primary tumours. The presence of p53 gene mutation did not correlate with histological grade, response to primary therapy and survival. These findings suggest that mutational alterations of the p53 gene are involved in the development of a significant proportion of some ovarian carcinomas (SCAs or EAs), especially in advanced stages. However, they may not be a marker predicting the biological behaviour or the outcome of the disease.