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Biomedical subjects

K Yoshikawa

Publications and source records attributed to K Yoshikawa.

At least 145 records · Page 8Linked to original sources

Analysis of T-cell antigen receptor gamma chain gene rearrangement by polymerase chain reaction in combination with denaturing gradient gel electrophoresis in the differential diagnosis of cutaneous T-lymphoproliferative diseases.

A polymerase chain reaction (PCR)-based strategy has been developed for analysis of clonal rearrangement of the T-cell receptor gamma gene (TCR gamma) and was shown to be useful for detection of clonal T-cell populations. In this study, we performed PCR combined with denaturing gradient gel electrophoresis (DGGE) on fresh frozen biopsy samples from 16 patients with cutaneous T-lymphoproliferative diseases in whom a definite diagnosis was difficult to make on morphological and immunohistochemical grounds alone. Ages of the patients at biopsy ranged from 28 to 81 (median 62) years, and the subjects consisted of 8 men and 8 women. They presented with erythema on the extremities in 5 cases, trunk in 7, buttock in 2, and papules on the trunk and face in one case each. Clonal rearrangement of TCR gamma was observed in 3 of 16 cases. Clinical diagnoses of these three cases were mycosis fungoides, cutaneous invasion of adult T-cell leukemia (ATL), and large granular lymphocytic leukemia (LGL) of T-cell type, respectively, but they were histologically difficult to differentiate from reactive cutaneous T-cell proliferation. The skin lesions of the LGL case worsened, and this patient died two years after biopsy. Another patient with suspected mycosis fungoides in the plaque stage died due to dissemination of tumors 22 months after biopsy. The remaining one patient with ATL survived with cutaneous lesions for over four years. Clonality was not demonstrated in the remaining 13 cases, and their clinical courses were favorable. These findings showed that demonstration of clonal TCR gamma gene rearrangement using the PCR-DGGE method is very helpful for diagnosis of cutaneous T-cell neoplasms.

Adult↗

Double involvement of extramammary Paget's disease in the genitalia and axilla.

We here reported a case of an 82-year-old man with double involvement of extramammary Paget's disease in the genitalia and axilla. Physical examination revealed erythema and reddish tumors on the pubic area and scrotum and irregular-shaped erythema on the left axilla. The skin biopsy sample from the genital area showed Paget's cells in the epidermis and upper dermis. The specimen from the left axilla showed Paget's cells scattered in the epidermis but not in the dermis.

Aged↗

Annexin VII as a novel marker for invasive phenotype of malignant melanoma.

Both F10 and BL6 sublines of B16 mouse melanoma cells are metastatic after intravenous injection, but only BL6 cells are metastatic after subcutaneous injection. While examining the genetic difference between the two sublines, we found a marked reduction of annexin VII expression in BL6 cells. In addition, fusion cell clones of both sublines were as poorly metastatic as F10 cells after subcutaneous injection, and contained the annexin VII message as abundantly as F10 cells. Hence, we examined whether the annexin VII expression was correlated with the less malignant phenotype of clinical cases by immunohistochemistry. Immunoreactivities to anti-annexin VII antibody in melanoma cells were evaluated quantitatively by using skin mast cells as an internal positive control. Eighteen patients with malignant melanoma were divided into two groups: lymph node metastasis-negative and positive groups. The ratio of numbers of patients positive versus negative to the antibody was significantly larger in the former than in the latter group. These results not only indicated that annexin VII serves as a marker for less invasive phenotype of malignant melanoma, but also suggested a possible role of annexin VII in tumor suppression.

Adolescent↗

Production of a single-chain variable fragment antibody recognizing type III mutant epidermal growth factor receptor.

The type III deletion mutant of the epidermal growth factor receptor (EGFR) is a potential target in diagnostic and therapeutic approaches for those glioblastomas characterized by its expression. We previously raised a mouse monoclonal antibody, 3C10 (IgG2b) specifically recognizing this mutant EGFR. In this study, a single-chain variable fragment (scFv) antibody was produced. Partial determination of its N-terminal amino acid sequence and preparation of adequate primers for variable heavy chain (V(H)) and variable light chain (V(L)) genes were performed to allow cloning by means of reverse transcriptase-polymerase chain reaction. The genes cloned were assembled with a linker, (Gly4Ser)3, and ligated into a bacterial expression vector to express the scFv as cytoplasmic inclusion bodies. After appropriate refolding, the antibody activity of the V(H)-V(L) scFv was examined in an enzyme-linked immunosorbent assay. 3C10 scFv showed a selective reactivity with the mutant peptide, similarly to the parental 3C10 antibody. A mouse transfectant expressing the type III mutant EGFR and a glioblastoma with type III deletion-mutant EGFR were positively stained by immunofluorescence. By Biacore analysis, the affinity (K(A)) of the parental 3C10 for the mutant peptide was 9.7 x 10(7) M(-1), while that of 3C10 scFv was 2.45 - 2.48 x 10(7) M(-1), being approximately 4-fold weaker. The results together suggested that the scFv antibody retained the appropriate structure to recognize a conformational epitope of the mutant receptor, similarly to the parental antibody.

3T3 Cells↗

[Gastrointestinal diseases associated with HIV infection].

A clinical studies were carried out on gastrointestinal diseases associated with HIV infection. During the 6 years between January 1993 and December 1998, 71 HIV infected cases visited to Yokohama Municipal Citizen's hospital, and 26 of them developed gastrointestinal complications during the course of their illness. They consisted of 24 males and 2 females, with the mean age of 44.7 years and the medial value of 42.5 years. Of the 26 patients, 21 were Japanese, and the remaining 5 were Southeast Asian. The mean CD4 count was 143/microliter and the medial value was 32/microliter at the time of development of complications. Gastrointestinal complications were esophageal candidiasis in 6 patients, cytomegalovirus (CMV) gastritis and gastric Kaposi's sarcoma in 1 patient each, amebiasis in 8 patients, infectious colitis in 11 patients, and asymptomatic pathogen carriers in 3 patients. Esophageal and gastric complications were common in patients with low count of CD4, and endoscopy was useful for diagnosis. Amebiasis developed even in patients with normal CD4 and was common in males with experience in homosexual contact. It seems that homosexual contact acquire not only HIV infection but also Entamoeba histolytica through sexual contact. Protozoan and acid-fast bacteria were detected at high rate in patients with infectious colitis and asymptomatic pathogen carriers. Besides food-born infections, imported infections were seen in foreign and Japanese patients who had traveled abroad. The gastrointestinal diseases associated with HIV infections for the most part were opportunistic infections or tumors but imported, food-born, and sexually transmitted infections were also observed. It seems necessary to take into consideration of varying background of patients in the treatment of gastrointestinal diseases associated with HIV infections.

AIDS-Related Opportunistic Infections↗

beta-cyanoalanine production by marine bacteria on cyanide-free medium and its specific inhibitory activity toward cyanobacteria.

In screening the culture broth of marine bacteria collected at Yap (Micronesia), Palau (Belau), and Okinawa (the southwest islands of Japan) for antimicroalgal activity, 37 out of 2,594 bacterial isolates tested were found to produce anticyanobacterial substances against Oscillatoria amphibia NIES-361. One strain, C-979, identified as a Vibrio sp., was selected and cultured in 2.4 liters of marine broth 2216 to identify the bioactive compound produced by the strain. The purified very hydrophilic compound (16.4 mg) was determined to be beta-cyano-L-alanine (L-CNAla) by instrumental analyses and the application of the advanced Marfey method. L-CNAla did not inhibit the growth of bacteria, yeast, or eukaryotic microalgae, but some cyanobacteria were found to be sensitive to L-CNAla at a concentration of 0.4 to 25 microg/ml. The effect of L-CNAla on some other environmental organisms, including invertebrates and a macroalgae, is discussed. CNAla production in marine broth was examined by thin-layer chromatography for the 37 bacterial isolates which produced an anticyanobacterial substance. The broth of 36 of these strains contained CNAla, suggesting the wide distribution of CNAla production by marine bacteria. This is the first report on bacteria that produce CNAla without a supply of the cyanide ion in the medium.

Alanine↗

Cellular and subcellular localization of necdin in fetal and adult mouse brain.

Necdin is a 325-amino-acid residue protein encoded by a cDNA clone isolated from neurally differentiated embryonal carcinoma cells. Ectopic expression of necdin induces growth arrest of proliferative cells. Necdin binds to major transcription factors E2F1 and p53, suggesting that necdin exerts its functions through the interactions with these cell-cycle-regulating factors. However, information about precise localization of endogenous necdin protein is currently lacking. A rabbit polyclonal antibody was raised against a bacterially expressed recombinant protein of necdin (amino acids 83-325). Immunoblot analysis revealed that necdin protein was expressed almost exclusively in the brain of adult mice. A relative molecular mass of endogenous necdin was estimated at approximately 43,000. In developing mouse brain, necdin was most abundant during fetal and neonatal periods. Necdin was highly enriched in the cytoplasm of hypothalamic neurons in fetal and adult mice. The subcellular fractionation analysis revealed that necdin was concentrated in the cytosol fraction of brain cells. These results suggest that endogenous necdin protein is localized predominantly in the cytoplasm of differentiated neurons and moves into the nucleus under specific conditions.

Age Factors↗

A role for heterologous gap junctions between melanoma and endothelial cells in metastasis.

F10 and BL6 sublines of B16 mouse melanoma cells are metastatic after intravenous injection, but only BL6 cells are metastatic after subcutaneous injection. We found that connexin (Cx) 26 is upregulated in BL6 cells. To examine gap junction formation, we devised a coculture system, in which an opened vein segment was placed at the bottom of a culture dish and then dye-labeled melanoma cells were seeded onto it. Immunohistochemistry indicated that the vein segment preserved the integrity of the endothelial monolayer. In this system, BL6 cells could transfer dye into endothelial cells but F10 cells could not. Transfection with wild-type Cx26 rendered F10 cells competent for coupling with endothelial cells and as spontaneously metastatic as BL6 cells. Conversely, transfection with a dominant-negative form of Cx26 rendered BL6 cells deficient in coupling and less metastatic. In human melanoma lesions, the level of Cx26 expression was low in melanoma cells residing in the basal layer, but significantly upregulated in melanoma cells invading the dermis. The results suggested that Cx26 plays a role in intravasation and extravasation of tumor cells through heterologous gap junction formation with endothelial cells.

Animals↗

Antisweet natural products. XV. Structures of Jegosaponins A-D from Styrax japonica Sieb. et Zucc.

From the fresh fruits of Strrax japonica Sieb. et Zucc., four new triterpenoid glycosides, named jegosaponins A-D (1-4), were isolated. Their structures were determined on the basis of spectroscopic data and chemical evidence. Compounds 1-4 are 3-O-tetraglycosides of barringtogenol C having an acetyl and a tigloyl or a (2Z)-hexenoyl groups at C-21, 22 and 28. The acylated saponins, 1-4, all showed antisweet activity.

Carbohydrate Sequence↗

Purification and characterization of an endo-polygalacturonase. From Aspergillus awamori.

An extracellular endo-polygalacturonase (PGase) produced by Aspergillus awamori IFO 4033 was isolated from the culture filtrate. The enzyme was purified to a homogeneous preparation with cation-exchange and size-exclusion chromatographies. Its properties were investigated, comparing them with that of recombinant pgx2 gene product, a PGase having protopectinase activity. This enzyme was a monomeric protein of 41 kDa, with an isoelectric point of pH 6.1. The characteristics of this PGase substantially coincide, with that of recombinant pgx2 gene product, and the PGase is assumed to be native pgx2 gene product. The production of PGase-X2 was confirmed to be regulated by ambient pHs.

Amino Acid Sequence↗

Purification and characterization of thermostable pectate lyase with protopectinase activity from thermophilic Bacillus sp. TS 47.

A strain of thermophilic bacterium, Bacillus sp., with pectolytic activity has been isolated. It produced an extracellular endo-polygalacturonate trans-eliminase (PL, EC 4.2.2.1) when grown at 60 degrees C on a medium containing polygalacturonate (PGA). The PL was purified by hydrophobic, cation exchange, and size exclusion column chromatographies. The molecular mass of the enzyme was 50 kDa by SDS-PAGE. The isoelectric point of the enzyme was pH 5.3. The enzyme had a half-life of 13 and 1 h at 65 and 70 degrees C, respectively, and showed optimal activity around at 70 degrees C and pH 8.0. It had protopectinase activity, besides PL activity, on lemon protopectin and cotton fibers. The first 20 amino acids sequence of the enzyme had significant similarity with that of PL from methophilic Bacillus subtilis, with 50% identity.

Amino Acid Sequence↗

A case of Penicillium marneffei infection in an AIDS patient: the first case in Japan.

A 38-year-old Japanese AIDS patient developed papular lesions which rapidly increased in number, eroded and crusted, and spread over not only skin but also the mucosal surface. High fever, sore throat, malaise and hepatosplenomegaly were also noted, and he died despite 2 months of intensive treatment. An autopsy revealed numerous histiocytes infected with Penicillium marneffei in the lymph nodes, liver, spleen, bone marrow, skin, and mucosal surface of the oral cavity to the pharynx. This case is thought to be the first Japanese case of penicilliosis marneffei.

AIDS-Related Opportunistic Infections↗

Usefulness of PET with 11C-methionine for the detection of hilar and mediastinal lymph node metastasis in lung cancer.

UNLABELLED: We retrospectively evaluated the usefulness of PET with 11C-methionine (methionine PET) for the diagnosis of lymph node metastases in patients with lung cancer. METHODS: Methionine PET and CT were performed before surgical intervention in 41 patients with primary lung cancer. We evaluated individual lymph nodes by methionine PET along with corresponding CT images. The 11C-methionine accumulation of lymph nodes was assessed semiquantitatively by analysis of the tumor-to-muscle ratio (TMR) and was compared with CT and histological diagnoses. RESULTS: A total of 126 lymph nodes, 36 of which were histologically diagnosed as metastatic, were assessed by CT and methionine PET. The TMR in metastatic lymph nodes (n = 36) was 5.15+/-1.69, whereas that of nonmetastatic lymph nodes (n = 90) was 2.91+/-0.76; this difference was significant (P < 0.0001). The most adequate TMR cutoff value for diagnosis of metastasis based on the results of receiver operating characteristic curve analysis was 4.1. The positive and negative predictive values, sensitivity, specificity, and accuracy of methionine PET were 79.5%, 94.3%, 86.1%, 91.1%, and 89.7%, respectively, and were superior to those of CT (57.6%, P = 0.04; 81.7%, P = 0.008; 52.8%, P = 0.002; 84.4%, NS; and 75.4%, P = 0.002, respectively). All positive nodes that were shown to be true-positive by CT, and 12 of 17 false-negatives on CT were correctly diagnosed by PET. Ten of 14 lymph nodes that were false-positive on CT were also correctly diagnosed by PET. CONCLUSION: Methionine PET appears to be superior to CT for the diagnosis of lymph node metastasis in lung cancer patients. The high negative predictive value of methionine PET suggests that cases in which lymph nodes are enlarged on CT with negative PET analysis may be diagnosed as negative for metastasis.

Carbon Radioisotopes↗

Activation of the MN/CA9 gene is associated with hypomethylation in human renal cell carcinoma cell lines.

The MN/CA9 (G250) gene expressed in the normal alimentary tract in a tissue-specific manner is often activated in renal cell carcinomas. To cast light on the activation mechanism, we examined the methylation status of this gene in seven human renal cell carcinoma cell lines (SKRC-01, -06, -10, -12, -14, -44, and -59) and three normal kidney tissue samples by using the bisulfite genomic sequencing protocol. CpG methylation was measured at seven locations in the MN/CA9 5' region. MN/CA9 transcripts were detected by reverse transcription-polymerase chain reaction in five of the renal cell carcinoma cell lines (SKRC-01, -06, -10, -44, and -59). These MN/CA9 positive cell lines showed hypomethylation, whereas the remaining two cell lines (SKRC-12, and -14), and three normal kidney tissue samples without transcripts demonstrated hypermethylation. Treatment with the demethylating agent 5-aza-2'-deoxycytidine resulted in activation of the MN/CA9 gene in the negative cell lines (SKRC-12 and -14). These data suggest that hypomethylation in the 5' region may have a major role in expression of the MN/CA9 gene in renal cell carcinoma cells.

Antigens, Neoplasm↗

Attenuation correction in evaluating renal function in children and adults by a camera-based method.

UNLABELLED: Correction for soft-tissue attenuation is required to evaluate absolute renal function by a camera-based method, and an estimate of renal depth and an attenuation coefficient are commonly used for attenuation correction. The first goal of this study was to develop formulas for the calculation of renal depth in both children and adults. The second goal was to optimize the attenuation coefficient for the estimation of renal accumulation of a 99mTc-labeled agent. METHODS: Renal depth was measured by CT in 74 children and 232 adults and compared with the depth calculated using previously published equations. Multiple stepwise linear regression analysis was conducted using data from children and adults together, and new formulas to calculate renal depth were derived. Using the resulting equations, percentage renal uptake at 2-2.5 min was computed from 99mTc-diethylenetriamine pentaacetic acid (DTPA) renography in 40 children and 92 adults. Percentage renal uptake was assessed using various values of an attenuation coefficient, and an optimized attenuation coefficient was determined to maximize the correlation coefficient between percentage renal uptake and glomerular filtration rate (GFR) measured from 2 blood samples. RESULTS: Although the previously published equations appeared to be acceptable in predicting adult renal depth, they substantially underestimated pediatric renal depth. Renal depth (D, cm) was shown by stepwise regression analysis to depend on the ratio of body weight (W, kg) to body height (H, cm) and was successfully calculated in both children and adults using the derived equations (right: D = 16.778 x W/H + 0.752; left: D = 16.825 x W/H + 0.397). The correlation coefficient between percentage renal uptake of 99mTc-DTPA and measured GFR varied substantially according to the attenuation coefficient used and was the highest (0.947) with an attenuation coefficient of 0.087/cm. CONCLUSION: The equations presented here enabled estimation of renal depth irrespective of the patient's age. Attenuation correction using these equations and the optimized attenuation coefficient appears to aid in evaluating renal accumulation and, consequently, renal function in both children and adults.

Adolescent↗

[Extended segmentectomy for small lung tumors].

UNLABELLED: To identify the benefits of limited surgery on small-sized lung tumors, the following subjects are discussed: 1) an updated report from a prospective group study (January 1992-December 1994) for extended segmentectomy for small lung tumors (n = 55); 2) a histopathological study of resected adenocarcinoma specimens less than 2 cm in size (n = 94), proposed by Noguchi; and 3) lung function after limited surgery and the quality of video-assisted thoracoscopic surgery. RESULTS: 1) Among 10 deaths, 4 patients died of their disease including one with local recurrence. The survival rate at 5 years was 91%, not including 6 unrelated deaths. 2) Of 94 patients, twelve with localized bronchioalveolar tumors (type A and B), and 23 of 57 patients with active fibroblastic proliferation (type C), underwent extended segmentectomy. Those 35 patients are all free of disease. The remaining 59 patients had a 70% 5-year survival rate. 3) Forced vital capacity was maintained at 92% of the preoperative level, which was much better than 81% for patients undergoing lobectomy. Card-sized thoracotomy using a thoracoscope was carried out in 92 patients, including 21 patients who underwent segmentectomy, in a series of 175 consecutive lung cancer operations. This approach resulted in less bleeding, the same operating time, and better preservation of vital capacity. CONCLUSIONS: Extended segmentectomy for small lung tumors did not affect the prognosis, and was associated with a better quality of life postoperatively.

Adenocarcinoma↗