Biomedical subjects

M Miyahara

Publications and source records attributed to M Miyahara.

At least 91 records · Page 5Linked to original sources

PubMed1995

Detection of reverse transcriptase activity by enzyme-linked immunosorbent assay in human immunodeficiency virus type 1.

An enzyme-linked immunosorbent assay (ELISA) using biotin-labelled oligo-dT primer and digoxigenin (Dig)-dUTP was designed to measure the reverse transcriptase (RT) activity of human immunodeficiency virus type 1 (HIV-1). The ELISA system involves the selective detection step of a newly synthesized cDNA by two specific bindings, biotin-streptavidin binding and alkaline phosphatase (AP)-conjugated anti-Dig-Dig binding, and the enzymatic amplification step to increase coloring generated by AP. This method was used to measure the activity of RT in the culture supernatants of peripheral leukocytes obtained from four anti-HIV-1-positive persons cocultivated with those from four anti-HIV-1-negative persons. RT activity was detected in all of four anti-HIV-1-positive culture supernatants but not in those cultivated with anti-HIV-1-negative supernatants alone. Thus, our improved ELISA for detection of HIV-1 appears to be sensitive enough and useful for routine laboratory work. This non-radioactive method will also be useful for detecting other retroviruses and for screening of RT inhibitors.

Cells, Cultured↗

PubMed1995

Laparoscopic approaches in the management of patients with early gastric carcinomas.

In eight Japanese patients, three different laparoscopic procedures were used to excise an early gastric carcinoma; partial resection in four, intragastric resection of the gastric mucosa in two, and laparoscopic-assisted distal partial gastrectomy with the abdominal wall-elevating method in two patients. Histological examinations revealed that the lesions were completely resected, and there was no evidence of lymphatic metastasis. The operation time ranged from 2 to 4 h. These forms of laparoscopic gastric surgery for patients with early gastric carcinomas may be useful from the standpoint of minimal access, rapid recovery, less pain, and good cosmesis.

Aged↗

PubMed1994

Characterization of the myosin-binding subunit of smooth muscle myosin phosphatase.

A myosin phosphatase was purified from chicken gizzard smooth muscle. The holoenzyme is a trimer and consists of 130,000-, 38,000-, and 20,000-Da subunits (in agreement with the results of Alessi et al.: Alessi, D., MacDougall, L. K., Sola, M. M., Ikebe, M., and Cohen, P. (1992) Eur. J. Biochem. 210, 1023-1035). The catalytic subunit, 38,000 Da, is the type 1 delta isoform, and its derived amino acid sequence is identical to the rat isoform. The larger subunit bound to myosin and also interacted with the catalytic subunit. cDNA clones encoding the large subunit were isolated from chicken gizzard cDNA libraries. Overlapping clones indicated the presence of two isoforms, and open reading frames of 2889 and 3012 bases were obtained. These encoded proteins of 963 and 1004 amino acids, with masses of 106,700 and 111,600 Da, respectively. The insert in the larger isoform is in the center of the molecule, at residues 512-552. The N-terminal third of the molecule is composed of eight repeat sequences, similar to the cdc10/SWI6 or ankyrin repeat. Myosin binding and binding to the catalytic subunit are properties of a 58,000-Da fragment that represents the N-terminal part of the molecule.

Amino Acid Sequence↗

PubMed1994

Activation of PKN, a novel 120-kDa protein kinase with leucine zipper-like sequences, by unsaturated fatty acids and by limited proteolysis.

PKN, a novel protein kinase with catalytic domain homologous to PKC family and unique amino terminal leucine zipper-like sequences, was purified partially from COS7 cells transfected with the cDNA construct encoding human PKN for enzymatic characterization of the enzyme. Using serine containing synthetic peptides based on PKC pseudosubstrate sites as the phosphate acceptors, kinase activities estimated from partially purified PKN were not stimulated by Ca2+/phosphatidylserine/diolein but were activated several-fold to several tens-fold by 40 microM unsaturated fatty acids, such as arachidonic acid, linoleic acid, and oleic acid. Autophosphorylation of the immunoprecipitates using anti-PKN antiserum was also stimulated by various unsaturated fatty acids. Limited proteolysis of PKN with trypsin induced an enhancement of the peptide kinase activity that was almost independent of arachidonic acid.

Amino Acid Sequence↗

PubMed1994

A regulatory subunit of smooth muscle myosin bound phosphatase.

The relationship between two putative myosin-binding subunits of smooth muscle myosin phosphatase was investigated. A monoclonal antibody (MoAb) to the 58 kD component of smooth muscle myosin-bound phosphatase (MBP) cross-reacted with a 130 kD protein in extracts of fresh chicken gizzards. The MoAb in combination with protein A immunoprecipitated from gizzard extracts a complex of the 130 kD protein plus the 38 kD catalytic subunit of the type 1 delta protein phosphatase. It is proposed that the 130 kD component is a native subunit of MBP and that the 58 kD protein is its proteolytic degradation product. The distribution of the 130 kD component in chicken tissues was screened using the MoAb. An immunoreactive band of appropriate mass was detected in all tissues except liver and skeletal muscle. Higher concentrations of the 130 kD component were evident in the smooth muscle samples.

Animals↗

PubMed1994

Extensive lymphadenectomy for thoracic esophageal carcinoma: a two-stage operation for high-risk patients.

Extensive lymphadenectomy (ELA) for esophageal carcinoma has always been limited to low-risk patients; however, we have developed a two-stage operation which extends the indications for ELA to include certain high-risk patients. Of 70 patients who underwent transthoracic subtotal esophagectomy for thoracic esophageal carcinoma between 1986 and 1991, 48 qualified for ELA which was performed on 45, including 27 low-risk patients (group A) whom underwent the traditional one-stage operation, while 18 high-risk patients underwent the two-stage operation (group B). The number of hospital deaths and postoperative complications was similar between the two groups. The median survival was 26 months in group B and 17 months in group A. The 40-day delay in resection of the neck and abdominal nodes in the two-stage operation did not significantly affect lymph node metastases, postoperative recurrence, or long-term survival. Thus, the two-stage operation is an oncologically meaningful approach to the treatment of high-risk patients who present with thoracic esophageal carcinoma.

Esophageal Neoplasms↗

PubMed1994

c-myc mRNA overexpression is associated with lymph node metastasis in colorectal cancer.

To elucidate the relationship between oncogene overexpression and malignant potential of colorectal cancer, the expression of three genes, c-myc, c-fos and c-Ki-ras was examined. Northern blot hybridisation was performed on fresh tissues excised from 35 patients with primary colorectal cancer. Overexpression of c-myc mRNA was evident in 25 of the patients (71%). Tumours with c-myc overexpression had a significantly higher frequency of lymph node metastasis than did those without (68 versus 20%, P < 0.05). The survival rate tended to be poorer (P = 0.06) in patients with c-myc overexpression (79% for 3 years) than in those without it (100%). We found no significant relationship between overexpression of the other two genes and pathological features. These findings suggest that c-myc overexpression is one parameter which can be used to determine the malignant potential of colorectal cancer.

Adult↗

PubMed1994

Laparoscopic surgical correction of circumcaval ureter.

Laparoscopic transposition and reanastomosis of a circumcaval ureter were performed in a 52-year-old man with right flank pain. A preoperative perfusion pressure study revealed abnormally high intrapelvic pressure. Under laparoscopy, the renal pelvis was divided above the ureteropelvic junction and the ureter was relocated from behind the vena cava. A 5 cm segment of redundant ureter containing the postcaval segment was resected and the ureteral end and renal pelvis were reapproximated with interrupted sutures by intracorporeal knot typing. The postoperative convalescence was uneventful, not necessitating the administration of analgesics. The patient resumed full activities 3 weeks later. The intravenous urogram and renogram obtained 2 months after the operation revealed remarkable improvement in the ureteral obstruction.

Abnormalities, Multiple↗

PubMed1994

Acetylsalicylate (ASA)-induced mitochondrial dysfunction and its potentiation by Ca2+.

Although it has been suggested that acetylsalicylate (ASA)-induced mitochondrial dysfunction plays an important role in the pathogenesis of Reye's syndrome, administration of ASA alone does not cause this syndrome in therapeutic doses. We noted, however, that Ca2+ plays important roles in the regulation of cellular functions. ASA at concentrations of 250 microM or less, which had little effect on succinate-linked respiration, impaired Ca2+ accumulation in liver mitochondria by causing an increase in Ca2+ release. ASA plus Ca2+, which in concentrations of 150 microM or less alone had little effect on mitochondrial respiration, inhibited state 3 respiration and dinitrophenol-induced uncoupling of respiration. In addition, ASA plus Ca2+ increased state 4 respiration and ATPase activity. These results indicate that ASA plus Ca2+ impaired mitochondrial ATP synthesis, and suggest that ASA and ASA-induced Ca2+ increases in cytosol form a vicious circle of effects. Furthermore, oral administration of ASA (150 mg/kg for 5 days running) to rats did not affect mitochondrial structure or liver function, but resulted in aberrations of mitochondrial respiration. These results suggest that even therapeutic doses of ASA may induce alteration in mitochondrial function.

Adenosine Triphosphatases↗

PubMed1994

Differences in the effects of two hexachlorobiphenyls on superoxide generation by polymorphonuclear leucocytes stimulated by N-formyl-methionyl-leucyl-phenylalanine and phorbol myristate acetate.

The effects of hexachlorobiphenyls (HCBs) on superoxide (O2-) generation by guinea pig polymorphonuclear leucocytes were examined. 2,3,6,2',3',6'-HCB by itself had only a weak inductive effect on O2- generation. This compound, however, enhanced O2- generation stimulated by N-formyl-methionyl-leucyl-phenylalanine (FMLP) about twofold, but not the generation induced by phorbol myristate acetate (PMA). On the other hand, 3,4,5,3',4',5'-HCB suppressed O2- generation stimulated by both FMLP and PMA. The inhibitory potency of this compound was far greater with PMA (ID50, 5 microM) than with FMLP (ID50, 40 microM).

Animals↗

PubMed1994

[DNA stemline heterogeneity as preoperative prognostic factor in esophageal cancer].

Flow cytometric nuclear DNA analysis was performed on 36 preoperative endoscopic biopsy specimens and 89 surgically resected specimens of esophageal carcinomas without preoperative radiotherapy. Carcinomas with aneuploid or DNA stem-line heterogeneity had a higher frequency of lymph node metastasis (p < 0.01). The correspondence rate of nuclear DNA ploidy patterns was 97% between biopsy and resected specimens, and that of heterogeneity was 72%. Though only six cases showed heterogeneity in biopsy specimens out of 12 cases which showed heterogeneity in resected specimens, preoperative detection of heterogeneity was supposed to be more accurate by an increase of biopsy specimens. DNA analysis of biopsy specimens may be a possible indicator of the malignant potential of esophageal carcinoma.

Biopsy↗

PubMed1994

[Comparison of several fluorescence HPLC methods for fumonisin analysis].

Fumonisin has no particular cromophore groups in its chemical structure applicable for trace analysis, and therefore requires derivatization prior to measurement. Currently, the analysis of fumonisin B1 (FB1), B2 (FB2) is generally carried out after derivatizing FB1 mainly with o-phthalaldehyde to make a fluorescent compound (OPA-F). The OPA-F is, however, not sufficiently stable for precise analysis, and we have thus examined other fluorescent derivatives including fluorescamine, FMOC, NBD-F and DBD-F. The analytical method using FMOC as fluorescent agent required column chromatography after the derivatizing reaction, while reaction of FB1 with fluorescamine gave two chromatographic peaks. FMOC and fluorescamine are accordingly not desirable as fluorescent agents for analysis of small quantities of FB1 and FB2 in foods. In conclusion, for precise analysis of such trace amounts of FB1 and FB2, NBD-F or DBD-F should be used as fluorescent agents.

4-Chloro-7-nitrobenzofurazan↗

PubMed1993

DNA stem line heterogeneity in esophageal cancer accurately identified by flow cytometric analysis.

BACKGROUND: This study was designed to evaluate the importance of DNA stem line heterogeneity in determining the malignant potential of esophageal cancer. METHODS: Flow cytometric analysis of intratumor heterogeneity of DNA contents was performed on step-sectioned slices of 57 resected esophageal carcinomas. RESULTS: DNA stem line heterogeneity, as assessed by DNA content measurements, was present in 25 (44%) tumors; 6 (11%) were a combination of diploid and aneuploid DNA pattern, and 19 (33%) had two or more aneuploid peaks with different DNA contents (multiploid). Of the remaining 32 homogeneous tumors, 4 showed only a diploid DNA pattern in all samples examined, whereas 28 tumors had only the aneuploid pattern. Tumors with the heterogeneous DNA pattern had a significantly higher frequency of lymph node metastasis than did those with the homogeneous DNA pattern (P < 0.05). CONCLUSIONS: For evaluation of the highly malignant potential of esophageal carcinoma by nuclear DNA contents, it is important to identify accurately intratumoral heterogeneity. Because different DNA stem lines were evident in different areas of the lesion, evaluation of multiple specimens from a wide area of each lesion is needed to determine with accuracy the degree of intratumor DNA stem line heterogeneity.

Adult↗

PubMed1993

Autopsy report of acute necrotizing opticomyelopathy associated with thyroid cancer.

We report an autopsied case of paraneoplastic necrotizing myelopathy. The patient had bilateral blindness, quadriplegia, and dyspnea of acute onset and died without remission 7 weeks later. The severe tissue necrosis and demyelination were found in the optic chiasm and from the medulla oblongata throughout the whole length of spinal cord. A papillary carcinoma was found in the thyroid gland at autopsy. In the present case IgG, myelin basic protein and activated helper T cells were increased in the CSF at onset, suggesting a mechanism of autoimmune demyelination for the condition.

Autoimmune Diseases↗

PubMed1993

Isolation and characterization of the StyD4I restriction endonuclease, a neoschizomer of ScrFI, from Escherichia coli K-12 carrying a small multicopy Hsd plasmid of Salmonella typhi origin.

A restriction endonuclease, designated StyD4I, a neoschizomer of ScrFI, has been isolated from Escherichia coli K-12 carrying a small multicopy host specificity for DNA (Hsd) plasmid of Salmonella typhi D4 origin. In the presence of 10 mM Mg2+, StyD4I cleaves the sequence 5'-/CCNGG-3' and generates a 5-nucleotide cohesive end. StyD4I should be useful for recombinant DNA technology, because of the stability and ease in handling the producer cells.

Deoxyribonucleases, Type II Site-Specific↗

PubMed1993

Swelling-induced O2- generation in guinea-pig neutrophils.

Without the addition of any exogenous stimuli, neutrophils generated O2- and then ceased in a reversible manner that correlated with cellular swelling and contraction. The nature of the possible mechanism responsible for this O2- generation was studied and compared with that observed in the triggering of stimulant-dependent O2- generation (respiratory burst). The swelling-induced O2- generation was inhibited by diphenyliodonium, and was independent of the functional distortion of mitochondrial and/or microsomal electron transport and xanthine oxidase. This suggested that such generation was involved in respiratory-burst oxidase activation; however, this generation was not accompanied by any new phosphorylation of the 47-kDa protein or of tyrosine proteins. Dihydrocytochalasin B potentiated the O2- generation. The cellular swelling produced a priming effect on the triggering of respiratory burst with different stimuli. Cellular contraction, conversely, suppressed the respiratory burst. The structural specificity of the swelling-induced plasma membrane modulation for the O2- generation was suggested by the finding that modulation of plasma membrane structures by various non-ionic detergents per se inhibited O2- generation. Lipophilic and positively-charged agents inhibited the generation and this inhibition was abrogated by negatively-charged, but not by non-ionic agents. Negatively-charged agents potentiated the O2- generation. These results suggest that both the interaction of the plasma membrane with the cytoskeleton and an increase in net negative charges at the plasma membrane play important role in evoking O2- generation; this is discussed and compared with the signal transduction reported previously for respiratory burst.

Animals↗

PubMed1993

Prediction of operative mortality based on impairment of host defense systems in patients with esophageal cancer.

To determine whether operative risk based on impaired defense systems can be predicted, we examined 21 markers in 32 patients with esophageal cancer seen between 1983 and 1985 and related them to postoperative deaths. A discriminant analysis proved useful in determining whether the patient would be at risk of operative mortality, based on impaired defense systems. This model, the host defense index (HDI), was then used clinically to evaluate findings in 64 patients seen between 1986 and 1991. During this period, a change in policy for performing transthoracic esophagectomy on and perioperative care for patients with impaired defense systems was associated with a decrease in operative mortality. Thus, the HDI is beneficial to predict the risk of operative mortality based on impaired defense systems in patients with esophageal cancer.

Aged↗

PubMed1993

Sacral chordoma developing two years after low anterior resection for rectal cancer.

A 43-year-old male with sacral chordoma associated with rectal cancer is herein reported. A presacral tumor with extensive destruction of S4 and S5 was found 2.5 years after a low anterior resection for advanced rectal mucinous carcinoma. Under the preoperative diagnosis of a solitary sacral metastasis of rectal cancer, the lower sacral segments together with the tumor were removed by amputation at S3. Histologically, the tumor was a chordoma composed of polyhedral cells with an abundant eosinophilic cytoplasm mixed with typical vacuolated physaliferous cells within a myxoid matrix. Colorectal cancer associated with an extracolic primary malignant neoplasm is not uncommon; however, this is only the second case of colorectal cancer associated with chordoma to the best of our knowledge.

Adenocarcinoma, Mucinous↗