Development of nested PCR based on the ViaB sequence to detect Salmonella typhi.
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Antigens, Bacterial↗
Biomedical subjects
Y Fujinaga
Publications and source records attributed to Y Fujinaga.
Development of nested PCR based on the ViaB sequence to detect Salmonella typhi.
For a rapid diagnosis of typhoid fever, we developed a nested PCR based on the nucleotide sequence encoding the Vi antigen. All Salmonella typhi strains along with a Salmonella paratyphi C strain were PCR positive. This assay was able to detect S. typhi at the single-cell level.
Antigens, Bacterial↗
Molecular construction of Clostridium botulinum type C progenitor toxin and its gene organization.
The 16S progenitor toxin of Clostridium botulinum type C is made up by conjugation of a neurotoxin with nontoxic components designated as nontoxic-nonHA and hemagglutinin (HA). The HA was found to be composed of subcomponents having 53, 33, 22-23, and 17 kDa molecular masses. Since we previously determined the whole nucleotide sequences of the genes for neurotoxin, nontoxic-nonHA, and HA-33, the cloning and nucleotide sequencing of the genes for the remaining HA subcomponents were performed. Two open reading frames coding for 16.7 kDa (HA-17) and 70.6 kDa proteins were identified. The N-terminal amino acid sequences of HA-53 and HA-22-23 indicated that the 70.6 kDa protein is split into 53 and the 22-23 kDa proteins after translation and that the 22-23 kDa protein consists of at least four proteins showing slightly different molecular weights.
Amino Acid Sequence↗
Unusual splice sites in the E1A-E1B cotranscripts synthesized in adenovirus type 40-infected A549 cells.
The adenovirus E1 DNA region consists of two transcription units, E1A and E1B. In this paper we report that the E1A-E1B cotranscripts containing sequences of both the E1A and E1B regions are synthesized during adenovirus type 40 (Ad40) infection of A549 cells. Cytoplasmic RNA was isolated from Ad40-infected A549 cells at 24, 72, and 100 h post infection (p.i.). The complementary (c) DNA was synthesized by reverse transcription using an oligo-dT primer and then amplified by the polymerase chain reaction (PCR) using primers derived from the E1A and E1B regions. The cDNAs thus amplified were sequenced either directly or after cloning into bacteriophage M13 vectors. Analysis of cDNA indicated that the E1A-E1B cotranscripts are synthesized at 72 h p.i., but not at 24 or 100 h p.i. Nucleotide sequences of three cDNAs of the E1A-E1B cotranscripts indicated that the cotranscripts originate from the E1A promoter and lack sequences for both the E1A poly(A) site and E1B cap site. The splices create open reading frames for E1A-E1B fused polypeptides around the E1A-E1B junctions in these mRNAs. Most interestingly, the sequence analysis showed that the 5' and 3' splice junctions in the two E1A-E1B cotranscripts do not conform to the splice consensus GT-AG rule. Our results thus suggest that factor(s) which lead to unusual splicing in the E1 mRNAs are present in Ad40-infected A549 cells.
Adenovirus E1A Proteins↗
Sequence variation of human papillomavirus type 16 E7 in preinvasive and invasive cervical neoplasias.
Variation in the nucleotide sequence of the HPV 16 E7 gene in preinvasive cervical intraepitherial neoplasia (CIN) and invasive cervical carcinoma specimens was analyzed. Direct DNA sequencing of PCR-amplified products with primers different from those used for PCR with 5'-end labeling generated distinct sequence ladders with a low background, even in specimens containing relatively low copy numbers of HPV. Of 14 cervical neoplasias, 11 cases showed sequence diversity from prototype HPV16, and a total of 22 nucleotide exchanges were detected. Nine of these led to single amino acid exchanges: [Thr5] to [Lys5] in one case and [Asn29] to [Ser29] in eight cases. The [Ser29] E7 was distributed uniformly among invasive carcinomas and precancerous legions, and was also found in a normal cervix. The [Lys5] E7 and [Ser29] E7 had transforming potential similar to the prototype E7 assessed by cooperation with the activated ras gene in rat embryo fibroblasts.
Amino Acid Sequence↗
Isolation of a cDNA encoding the adenovirus E1A enhancer binding protein: a new human member of the ets oncogene family.
The cDNA encoding adenovirus E1A enhancer-binding protein E1A-F was isolated by screening a HeLa cell lambda gt11 expression library for E1A-F site-specific DNA binding. One cDNA clone produced recombinant E1A-F protein with the same DNA binding specificity as that endogenous to HeLa cells. Sequence analysis of the cDNA showed homology with the ETS-domain, a region required for sequence-specific DNA binding and common to all ets oncogene members. Analysis of the longest cDNA revealed about a 94% identity in amino acids between human E1A-F and mouse PEA3 (polyomavirus enhancer activator 3), a recently characterized ets oncogene member. E1A-F was encoded by a 2.5kb mRNA in HeLa cells, which was found to increase during the early period of adenovirus infection. In contrast, ets-2 mRNA was significantly reduced in infected HeLa cells. The results indicate that E1A enhancer binding protein E1A-F is a member of the ets oncogene family and is probably a human homologue of mouse PEA3.
Adenovirus E1A Proteins↗
[Catheter-induced thrombosis of the inferior vena cava--case report].
A 47-year-old woman was admitted for bleeding from gastric ulcer for which a central venous catheter was placed through the right femoral vein. Seven days later, abdominal echograms unexpectedly revealed a large thrombus along the catheter in the inferior vena cava. Because the echograms also disclosed the thrombus immobile even in postural change, the catheter was pulled out. Within two weeks following the catheter removal, the thrombus spontaneously disappeared with no evidence of pulmonary embolism. Although thrombolysis therapy may be the treatment of choice for catheter-induced thrombosis, simple removal of the catheter is effective in case that the thrombus is immobile.
Catheterization, Central Venous↗
Biological and biochemical activity of E7 genes of the cutaneous human papillomavirus type 5 and 8.
In contrast to the observed activity of the E7 genes of the genital high-risk human papillomavirus (HPV)16 and HPV18, E7s of the cutaneous high-risk HPV5 and HPV8 show no in vitro transforming activity in established rodent cells. We recently reported that the HPV8 E7 driven by the SV40 enhancer/promoter oncogenically transforms primary rat embryo fibroblast (REF) cells collaboratively with the EJras oncogene (Jpn. J. Cancer Res., 82, 1340-1343, 1991). To study the functional differences between cutaneous HPV5 and HPV8 E7s and genital HPV16 E7, we cloned each of the E7 open reading frames and tested their immortalizing and transforming activities, the binding ability of their products with retinoblastoma protein (RB) and their complementation activity of a RB-nonbinding adenovirus E1A mutant. In contrast to results with HPV16 E7, transfection of HPV5 and HPV8 E7s did not produce any G418-resistant colonies in primary baby rat kidney (BRK) cells. However, they induced morphological transformation of primary BRK cells as well as of primary REF cells when cotransfected with the EJras oncogene. The ras-cooperating activity of HPV8 E7 appears to be extremely low, since, unlike the case of HPV5 and HPV16 E7s, transformed BRK colonies induced by HPV8 E7 plus ras have had a very low survival rate. The in vitro RB binding experiment showed that HPV5 and 8 E7s are able to form complexes with RB protein with reduced affinities of about one fourth and one nineteenth that of HPV16 E7, respectively. Moreover, not only HPV16 E7 but also HPV5 and 8 E7s complemented a nontransforming adenovirus 5 E1A mutant (dl922/947) incapable of binding to RB in inducing E1A-specific transformed foci on primary BRK cells. Since both the activities, the ras-collaborative transformation and complementation of the inert E1A mutant by E7s, all correlate with in vitro RB binding affinity (HPV16 E7 > HPV5 E7 > HPV8 E7), it is likely that RB binding of HPV5 and HPV8 E7s is an integral part of the biological activities of these proteins.
Adenovirus E1A Proteins↗
Clinicopathologic evaluation of recurrence in early gastric cancer.
Five hundred ninety-two patients with early gastric cancer underwent surgical resection from 1970 to 1986 in our hospital, and 13 died from a recurrence of their disease. A careful analysis of these 13 patients suggests that carcinomas which invaded to the submucosa tend to recur more often than those confined to the mucosa. Well differentiated and papillary adenocarcinomas characterized by protruded or elevated lesions tend to recur earlier than poorly differentiated or signet-ring cell carcinomas characterized by depressed or excavated lesions. However, both types recurred from hematogenous metastases, with the liver being the most common site. Therefore, the macroscopic and histological features presently used to characterize early gastric cancer do not provide sufficient information to accurately predict which patients are at most risk for recurrence.
Adenocarcinoma↗
[The usefulness of dipyridamole stress myocardial scintigram as a preoperative test for detecting ischemic heart disease before vascular surgery].
We evaluated the usefulness of dipyridamole 201Tl myocardial scintigraphy (DS) for detecting myocardial ischemia in 18 candidate for vascular surgery. DS indicated significant thallium redistribution in 6 patients. Coronary angiography was performed in 5 of these 6 patients and revealed significant coronary artery disease in all patients. As a result, operation was cancelled in 4 patients with severe coronary artery disease. One of these cases died of acute myocardial infarction after being discharged from hospital. One other patient was operated after percutaneous transluminal coronary angioplasty. Another patient was operated knowing that he had significant but mild coronary artery disease. Subsequently, operations were performed in 14 patients. No serious cardiac event occurred in these 14 patients except one case with T wave inversion without myocardial enzyme elevation. Thus, we concluded that DS was a useful method for detecting ischemic heart disease and prevent cardiac event associated with vascular surgery.
Aged↗
Simultaneous detection and typing of genital human papillomavirus DNA using the polymerase chain reaction.
A simple method has been developed for detecting a broad range of genital human papillomavirus (HPV) types using the polymerase chain reaction (PCR). We utilized two consensus sequence primer pairs within the E6 and E7 open reading frames to amplify HPV DNA; malignant HPV DNA (from HPV-16, -18, -31, -33, -52b and -58) was amplified using the pU-1M/pU-2R primer pair whereas benign HPV DNA (from HPV-6 and -11) was amplified using the pU-31B/pU-2R primer pair. Identification of the amplification product was confirmed by restriction enzyme digestion. In this study, a pU-1M/pU-2R-mediated PCR was successfully applied to 39 cervical carcinoma specimens; HPV-16 was detected in 19 cases, HPV-18 in five cases, HPV-31 in two cases, HPV-33 in two cases, HPV-52b in one case, HPV-58 in three cases, and an unknown type(s) was detected in four cases. Overall, the prevalence of HPV was 84.6%. The results indicate that this detection system is useful for the detection of HPVs not only of known types but also of new types.
Base Sequence↗
Peripheral pulmonary adenoid cystic carcinoma with substantial submucosal extension to the proximal bronchus.
A rare case of a peripheral pulmonary adenoid cystic carcinoma is presented. Histological examination after lobectomy showed pronounced submucosal invasion of the carcinoma to proximal bronchi.
Aged↗
[Analysis of human papillomavirus genome by PCR].
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Animals↗
[Cataract--clinic and pathology].
Clinicopathological studies were performed on 156 lenses of human senile cataract obtained by cataract operations between 1970 and 1988. It became clear that the aging influences the functional destruction of the equatorial region, the pathological changes of the bow area, and changes of the extralens environment. After operation for the atrophic type of the posterior subcapsular cataract, aftercataract easily develops on the intraocular lens and this requires treatment. Long-term observations were carried out in 180 Wistar male rats under the same laboratory condition and histological studies were performed. The similarities between the senile Wistar rat cataract and the human senile cataract indicate that the Wistar rat cataract is useful as a model for studying the human senile cataract. These rats were initially classified into six groups (control, vitamin E diet, EPC eye drops, catalin eye drops and reduced catalin eye drops). To study the effects of the agents (vitamin E, ARI, EPC, catalin, reduced catalin) on the cataract in senile Wistar rats the mean cell density of lens epithelia were measured at 2 or 3-month intervals. There were no statistically significant differences in treated groups and the control group. The results suggest that these agents affect another factor of lens apart from the proliferative activity of lens epithelial cell. Effects of anti-cataract agents were investigated using cultured lens epithelial cells. When cultured rat lens epithelial cells were incubated in medium containing selenite, super-oxide dismutase (SOD) activity and GSH in the cells markedly decreased, and GSSG was markedly increased. When cultured rabbit lens epithelial cells were incubated in medium contained selenite and glutathione, SOD activity was maintained normal level. When cultured lens epithelial cells were incubated in medium contained selenite and pirenoxin, SOD activity also maintained a normal level. These results suggest that both glutathione and pirenoxin are effective as anti-cataract agents. Cataracts in spontaneously hypertensive rats (SHR) was investigated on male of Wistar-Kyoto rats (WKY), stroke resistant SHR (SHRSR) and stroke-prone SHR (SHRSP) rats aged 3 to 9 months. Cataracts in these rats were classified as follows: Type 0: no opaciiy, Type 1: nuclear opacity, Type 2: posterior subcapsular opacity, Type 3: nuclear opacity associated posterior subcapsular opacity and Type 4: complete opacity in both lenses. Incidence of cataract in WKY was 2.6%, SHRSR, 76.8% ant SHRSP, 88.2%. Incidence of nuclear opacity was remarkably higher in SHRSP (48.5%). In SHR aged from 3 to 5 months, nuclear opacity was ahead of the appearance of posterior subcapsular opacity which was increased during aging.(ABSTRACT TRUNCATED AT 400 WORDS)
Aged↗
A pertussis toxin-sensitive GTP-binding protein plays a role in the G0-G1 transition of rat hepatocytes following establishment in primary culture.
Acute spontaneous c-myc gene expression and sustained increase of a GTP-binding protein(s) (G-protein) which is sensitive to islet-activating protein (IAP), pertussis toxin, occurred early during primary culture of adult rat hepatocytes. Following these earlier events, DNA synthesis was demonstrated in response to EGF and insulin. Addition of IAP immediately after plating of primary cultures inhibited c-myc expression and the hormone-induced DNA synthesis. Addition at 24 h or later following cell inoculation, however, produced only weak effects on DNA synthesis, even though the IAP-sensitive G-proteins were completely inactivated. We conclude that the IAP-sensitive G-protein(s) plays a role in the earlier process(es) of the G0-G1 transition, which is essential for the initiation of growth factor-dependent DNA synthesis.
Adenosine Diphosphate Ribose↗
A clinicopathological study of synchronous multiple gastric cancer.
In a 7 1/2-year period (from 1 January 1980 to 30 June 1987), 997 patients with gastric cancer underwent gastric resection at the Department of Surgery, Matsuyama Red Cross Hospital, Japan. We studied clinicopathologically 83 patients (182 lesions) with synchronous multiple gastric cancer and compared them with patients who had solitary cancers only in the same period. We found that elderly men had a relatively high incidence of multiple gastric cancer, and that early cancer of the elevated well differentiated type was the most common. Of the 182 lesions, as many as 42 (23.1 per cent) were missed in the preoperative examination. Eighteen of these 42 lesions were found only with the aid of a microscope in sections almost incidentally excised. Small flat lesions tended to be missed. We should always be conscious that other gastric lesions may be present when treating patients with gastric cancer.
Female↗
Amyloid enhancing factor (AEF) in the aging mouse.
Amyloid enhancing factor (AEF) activity was tested in spleen homogenates from 52, 28, and 9-week-old untreated mice. Strong AEF activity was present in the supernatant of spleen homogenate from the 52-week-old mice. Only weak activity was observed in the supernatant from the 28-week-old mice, and none was seen in that from the 9-week-old mice. Crude spleen homogenates from the 52-week-old mice were subjected to Sephacryl S-300 gel filtration. AEF activity existed in the first gel filtration peak. These findings suggest that AEF increases in the spleens of aged mice and that AEF may be a substance of high molecular weight.
Aging↗
Immunocytochemical evidence of amyloid-enhancing factor (AEF) in polymorphonuclear leukocytes.
Amyloid-enhancing factor (AEF) was extracted from spleens of mice that had received amyloidogenic stimulation. Sephacryl S-300 gel filtration of the crude AEF yielded five fractions, among which strong AEF activity was present in the first peak (F1), and confirmed by an amyloid induction experiment. An anti-AEF antiserum was obtained from a rabbit by immunization with F1. This antibody reacted strongly with splenic polymorphonuclear leukocytes (PML) from mice given amyloidogenic stimulation, and weakly with those from normal untreated mice. Isoelectric focusing (IEF) analysis of both F1 and sera from mice given amyloidogenic stimulation was performed. A single band was observed on IEF analysis of F1, whereas many bands were seen on IEF analysis of the sera. After the substances in the gel had been transferred to nitrocellulose membranes by capillary blotting, the membranes were made to react with the anti-AEF antiserum. The results suggested that AEF is a high-molecular-weight substance derived from PML and increases in the serum at the time of, or shortly prior to, amyloid deposition in the spleen.
Amyloid↗