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Biomedical subjects

M Kuwahara

Publications and source records attributed to M Kuwahara.

At least 19 recordsLinked to original sources
PubMed1995

p53 and human papillomavirus DNA in renal pelvic and ureteral carcinoma including dysplastic lesions.

Ninety-eight cases of transitional-cell carcinoma (TCC) of the renal pelvis and ureter, including dysplastic lesions, were studied for tumor incorporation of human papillomavirus (HPV) type-16 and type-18 DNA by in situ hybridization (ISH) with DNA probes for each HPV viral type. Immunohistochemical analysis of p53 expression was also performed. Fresh tumor tissues from 26 patients were also studied for p53 mutations in exons 4 through 9 by direct sequencing and for HPV infection by polymerase chain reaction (PCR). Thirty-two tumors were positive for HPV DNAs, including 6 double-positive cases. Among these tumors, adjacent dysplastic lesions in 21 cases (66%) also revealed identical reactivity. Overexpressed p53 was detected in 26 cases. Expression of p53 was also detected in dysplastic lesions in 19 out of these 26 cases (73%). Three cases were positive for both HPV DNA and p53 antibody. p53 point mutation was detected in 7 of 26 cases, 6 of which were also positive for p53. HPV type-16 DNA was detected in 6 cases by PCR, 4 of which were also ISH-positive. Overexpressed p53 was frequently detected in invasive and non-papillary tumors (p < 0.01) and in high-grade tumors (p < 0.05). HPV infection was more common in non-invasive and papillary tumors (p < 0.05). These findings suggest that HPV infection or overexpression (mutation) of p53 may be an early event and be related to phenotypes of tumor-cell growth patterns and progression.

Adult↗
PubMed1995

Structures of genomic and complementary DNAs coding for Pleurotus ostreatus manganese (II) peroxidase.

To study the mechanism of regulation and structure/function relationship of the Pleurotus ostreatus manganese (II) peroxidase (MnP), we amplified the full-length genomic and complementary DNAs for the major isozyme of the MnP mainly by the cassette-primer PCR technique and then sequenced them. The cDNA contained an open reading frame of 1083 bp encoding for a polypeptide of 361 amino-acid residues, including the suggested signal peptide of 29 amino-acid residues with a prepro structure. The predicted amino-acid sequence of the protein shared several common characteristics with those of fungal lignin and manganese (II) peroxidases. We could find a suggested metal response element and two heat-shock element-like sequences in the 5'-flanking region of the structural gene. The structural gene contained 15 introns, many of which lie identical to those in lignin peroxidase genes rather than to those in the known MnP genes.

Amino Acid Sequence↗
PubMed1995

cAMP-dependent phosphorylation stimulates water permeability of aquaporin-collecting duct water channel protein expressed in Xenopus oocytes.

Among water channel proteins (aquaporins), aquaporin-collecting duct (AQP-CD) is the vasopressin-regulated water channel. Vasopressin causes cAMP production in the renal collecting duct cells, and this is believed to lead to exocytic insertion of water channel into the apical membrane (shuttle hypothesis). AQP-CD contains a consensus sequence for cAMP-dependent protein kinase, residues at positions 253-256 (Arg-Arg-Gln-Ser). To determine the role of this site, Ser-256 was substituted for Ala, Leu, Thr, Asp, or Glu by site-directed mutagenesis. In Xenopus oocytes injected with wild-type or mutated AQP-CD cRNAs, osmotic water permeability (Pf) was 4.8-7.7 times higher than Pf of water-injected oocytes. Incubation with cAMP plus forskolin or direct cAMP injection into the oocytes increased Pf of wild-type, but not mutated, AQP-CD-expressing oocytes, whereas the amounts of AQP-CD expression were similar in wild and mutated types as identified by Western blot analysis. In vitro phosphorylation studies of AQP-CD proteins expressed in oocyte showed that cAMP-dependent protein kinase phosphorylated wild-type, but not mutated, AQP-CD proteins. Phosphoamino acid analysis revealed that this phosphorylation occurred at the serine residue. Moreover, phosphorylation of AQP-CD protein in intact rat kidney medulla tissues was stimulated by incubation with cAMP. Our data suggest that cAMP stimulates water permeability of AQP-CD by phosphorylation. This process may contribute to the vasopressin-regulated water permeability of collecting duct in addition to the apical insertion of AQP-CD by exocytosis.

Animals↗
PubMed1995

Reducing interference from heterophilic antibodies in a two-site immunoassay for carcinoembryonic antigen (CEA) by using a human/mouse chimeric antibody to CEA as the tracer.

To reduce heterophilic antibody interference in a two-site immunoassay for carcinoembryonic antigen (CEA), we utilized a human/mouse chimeric antibody to CEA as the tracer. One mouse monoclonal antibody (MAb), F82-61, which reacts with an epitope present on the domain N of CEA, was immobilized on 96-well polystyrene microtiter plates. A human/mouse chimeric antibody (Ch F11-39), which recognizes an epitope present on the domain B3 of CEA, was biotinylated for the tracer (Ch F11-39 system). Another MAb F11-39, the parental MAb of Ch F11-39, was also biotinylated and used as the control tracer (F11-39 system). For a fair comparison, the same 503 serum samples from healthy individuals were simultaneously assayed in the present study. When a tentative common reference limit of 5 ng/ml was used, the false positive rate with the Ch F11-39 system was only 2.8% (14/503) and that with the F11-39 system was 29.0% (146/503). Adding normal mouse serum (NMS; 1%) or a mixture of purified mouse IgG subclasses (heterophilic blocking reagent (HBR, 15 micrograms/test)) to the F11-39 system reduced the false positive rate from 29.0% to 6.2% (31/503) or 4.8% (24/503), respectively, suggesting that heterophilic antibodies reactive with mouse IgG gave rise to the high positive rate in normal populations with the F11-39 system. On the other hand, the false positive rate with the Ch F11-39 system was only slightly reduced from 2.8% to 2.6% (13/503) or to 2.0% (10/503) by adding NMS or HBR to the Ch F11-39 system. The false positive rates with two commercially available assay systems, CEA Roche EIA.DM or Abbott IMx CEA, were 5.4% (27/503) and 5.8% (29/503), respectively, which both corresponded roughly to that with the F11-39 system including NMS or HBR. These results indicate that the application of human/mouse chimeric antibodies in two-site immunoassays is more effective for reducing interference from heterophilic antibodies than the adding of NMS or purified mouse IgG in the assay using conventional MAbs.

Animals↗
PubMed1995

A reference of the GOLD classification of monoclonal antibodies against carcinoembryonic antigen to the domain structure of the carcinoembryonic antigen molecule.

The epitopes of 42 well-characterized monoclonal antibodies (MAbs) against carcinoembryonic antigen (CEA) from 10 different research groups were mapped in terms of domain structure (domains N, A1-B1, A2-B2, and A3-B3) of the CEA molecule on the basis of the reactivities with recombinant CEA proteins expressed in Chinese hamster ovary cells. Thirty-six of the 42 MAbs tested have previously been classified into 5 essentially nonoverlapping epitope groups (GOLD 1-5) by cross-competition assays among MAbs for CEA binding (Hammarström S, et al.: Cancer Res. 1989; 49:4852-4858). The epitopes recognized by GOLD 2 MAbs were all present on domain A2-B2, those for GOLD 5 MAbs were all on domain N, and those for GOLD 4 were mapped around domains A1-B1 and A2-B2. On the other hand, the epitopes for GOLD 1 MAbs were distributed into domains N, A2-B2, and A3-B3, and those for GOLD 3 MAbs were separated into domains N and A3-B3. Although the exact reasons for the dispersed patterns of GOLD 1 and 3 MAbs on the domain structure of the CEA molecule are unclear at present, several factors, such as a spatial relation or a close proximity of epitopes, conformation dependency, and repetitivity of epitopes, may be considered as possible explanations. The epitope mapping reported here helps form the basis for understanding the relation between the chemical structure and antigenic activities of the CEA molecule and may be useful to study the functions of the CEA molecule, especially those of the respective domains.

Animals↗
PubMed1995

High-energy underwater shock wave treatment for internal iliac muscle metastasis of prostatic cancer: a first clinical trial.

The first clinical trial of high-energy shock wave (SW) combined with chemotherapy to treat metastasis of prostate cancer in the internal iliac muscle was conducted. The patient, a 57-year-old man, diagnosed as having mucin-producing, poorly differentiated adenocarcinoma of the prostate invading the bladder wall, had been treated by total cystoprostatectomy. Five months later, metastatic tumors were found in the left axillar subcutaneous tissue and the right internal iliac muscles. For the axillar metastasis we performed radiation and left subclavicular arterial infusion of cisplatin 70 mg, THP-adriamycin (THP) 50 mg and methotrexate 50 mg. For the right internal muscular metastasis, 10,000 to 20,000 shots of SW and simultaneous intravenous injection of carboplatin 100 mg and THP 10 mg were carried out. Neither of the tumors decreased in size, but on magnetic resonance images, the SW-treated tumor exhibited a central low-intensity area. The SW-treated tumor was resected and central necrosis and a collection of mucin in the central area were observed. Hormone-resistant prostate cancer is well-known to be a multidrug-resistant tumor. It is noteworthy that SW and chemotherapy induced necrosis in such a refractory cancer without any significant side effects.

Adenocarcinoma↗
PubMed1995

Role of intra- and extracellular calcium stores in mesothelial cell response to histamine.

Ca2+ may play an important role in mesothelial cellular responses because Ca2+ acts as an intracellular messenger in a wide variety of cellular responses in different tissues. The present study was designed to clarify the mechanisms of cytosolic Ca2+ mobilization in the mesothelial cells. Rat pleural and pericardial mesothelial cells were maintained in vitro, and the Ca2+ movement was evaluated using fura 2. Histamine (30 microM to 10 mM) induced a biphasic elevation of intracellular levels of Ca2+ concentration ([Ca2+]i) that consisted of a rapid initial transient elevation followed by a sustained elevation. Neither removal of extracellular Ca2+ nor inhibition of Ca2+ influx by 1 microM nifedipine affected the histamine-induced initial transient elevation of [Ca2+]i in mesothelial cells. Nifedipine did not block histamine-induced sustained elevation of [Ca2+]i. KCl (25 and 50 mM) elicited a biphasic elevation of [Ca2+]i. However, this KCl-induced elevation of [Ca2+]i was abolished by nifedipine treatment. Ryanodine (10 microM) induced a transient elevation of [Ca2+]i in Ca(2+)-free solution. The histamine-induced elevation of [Ca2+]i was completely blocked by H1-receptor antagonists. These results suggest that the mesothelial cells have three pathways to increase [Ca2+]i: release from intracellular Ca2+ stores, Ca2+ influx through L-type voltage-dependent Ca2+ channels, and receptor-operated Ca2+ channels.

Actins↗
PubMed1995

Synthetic studies on condensed-azole derivatives. I. Synthesis and anti-asthmatic activities of omega-substituted alkylthioimidazo[1,2-b]pyridazines.

A series of novel omega-substituted alkylthioimidazo[1,2-b]pyridazines was designed and synthesized in an effort to find a novel anti-asthmatic agent. The anti-asthmatic activity of these compounds was evaluated on the basis of their ability to inhibit thromboxane A2 synthetase and platelet activating factor (PAF)-induced bronchoconstriction in guinea pigs. None of these compounds significantly inhibited thromboxane A2 synthetase, though, sulfonamide derivatives potently inhibited PAF-induced bronchoconstriction. Among them, 3-(imidazo[1,2-b]pyridazin-6-yl)thiopropanesulfonamide (5) showed the most potent inhibitory effect. The anti-asthmatic effects of compound 5 in experimental models were superior to those of theophylline.

Animals↗
PubMed1995

Synthetic studies on condensed-azole derivatives. III. Synthesis and anti-asthmatic activities of C-substituted alkyl side chain derivatives of omega-sulfamoylalkylthioimidazo[1,2-b]pyridazines and related compounds.

A series of novel alkylthioimidazo[1,2-b]pyridazines was synthesized and evaluated for ability to inhibit platelet activating factor (PAF)-induced bronchoconstriction in guinea pigs. Among them, 3-(imidazo[1,2-b]pyridazin-6-yl)thio-2,2-dimethylpropanesulfona mide (15) showed the most potent inhibitory effect. The structure-activity relationships in this series of compounds, in particular, the effects of conversion of the imidazopyridazine ring into other heterocyclic rings, introduction of a substituent group at the 2 or 3 position of the imidazopyridazine ring and introduction of a substituent group into the alkyl side chain, are also discussed.

Animals↗
PubMed1995

Mechanisms of regulatory volume decrease in collecting duct cells.

The mechanisms of cell volume regulation upon osmotic cell swelling were examined in the inner stripe of the outer medullary collecting duct (OMCDi). Segments of OMCDi were dissected from rabbit kidney and perfused in vitro. Using an image processing system, the cross-sectional area of tubule cells was monitored as an index of the relative cell volume. In response to a decrease in extracellular osmolality (290 to 190 mOsm), the tubule cells swelled promptly and restored gradually their original cell volume by a mechanism termed regulatory volume decrease (RVD). The initial response of RVD (the rate of the decrease in cell volume during the first 10 min) was inhibited by 79% at a high concentration of basolateral K+ (50 mM). By contrast, the same concentration of luminal K+ did not affect the response. When basolateral Cl- was removed 30 min before the experiment, the initial response of RVD was decreased by 77%, whereas the response was not affected 30 min after removal of luminal Cl-. Addition of basolateral Ba2+ and basolateral anthracene-9-CO2H inhibited the response by 70 and 65%, respectively. RVD response was accompanied by a transient rise in intracellular Ca2+. The Ca2+ rise was abolished when intracellular Ca2+ was chelated by acetoxymethyl ester of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM). In this condition, the initial RVD response was decreased by 68%. Our data suggest that the exit of basolateral K+ and Cl- via conductive pathways mainly mediates the RVD response in rabbit OMCDi cells, and that intracellular Ca2+ is involved in this response.

Animals↗
PubMed1995

[Value of PSA/gamma-Sm ratio (P/S ratio) for diagnosis of prostate cancer in patients with urinary retention].

It is known that serum prostate specific antigen (PSA) in urinary retention due to benign prostatic hypertrophy (BPH) increase. To evaluate prognostic value of the ratio of serum PSA to gamma-seminoprotein (P/S Ratio) for prostate cancer (PC) in patients with urinary retention, we have studied the P/S Ratio at the initial examination in 33 patients with untreated PC (10 with and 23 without urinary retention) and 193 patients with untreated BPH (38 with and 155 without urinary retention) histopathologically diagnosed at our hospital between January, 1992 and December, 1993. The results were as follows: 1) The mean P/S ratio of PC patients was significantly higher than that of BPH patients in both groups with and without urinary retention. 2) When the cut off value of P/S Ratio was determined to be 1.35, the highest efficiency, 59.3% was obtained in the group without urinary retention. The sensitivity and specificity were 65.2% and 91.0%. 3) In the group with urinary retention, the efficiency was also the highest, 80.0% with a cut off value of 1.35. The sensitivity and specificity were 80.0% and 100%. 4) In all patients, the efficiency was 64.6%, the sensitivity was 69.7%, and the specificity was 92.7% with a cut off value of 1.35. 5) Positive rate of serum PSA in BPH patients with urinary retention was 47.4% and that in BPH patients without urinary retention was 17.4%. The mean P/S ratio of the BPH patients with urinary retention was significantly lower than that of BPH patients without urinary retention, which suggested that the serum free PSA increase in the former.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1995

[High flow priapism after perineal trauma, successfully treated by unilateral embolization of the internal pudendal artery: a case report].

We reported a case of 45-year-old patient with high flow priapism secondary to arteriovenous fistula produced by perineal trauma. Diagnosis was based on the results of gasometry in cavernous blood, cavernography and pudental arteriography. Although conservative treatment had been tried, complete resolution of priapism was not obtained. We could successfully treat the priapsim by percutaneous temporary embolization of the right internal pudendal artery with Gelatin. Erection and sexual function, after 2 months of treatment, was normal. The rationale for the use of this embolization in the treatment of high flow priapism and its etiology was discussed.

Embolization, Therapeutic↗
PubMed1995

[Evaluation of the efficacy of recombinant human erythropoietin (rHuEPO) administration on penile erection in males undergoing hemodialysis and effect on pituitary-gonadal function].

Recombinant human erythropoietin (rHuEPO) was administered to males undergoing hemodialysis, and its effects on penile erection and hypothalamus-pituitary-gonadal hormone levels were studied. The subject consisted of 18 males undergoing hemodialysis ranging in age from 22 to 58 years (mean 45.3 years). Chronic glomerulonephritis was present in 16, and diabetic nephropathy in 2, as underlying disease. rHuEPO was administered intravenously at 1,500 U 3 times a week with a target to increase the Ht value to 25% or above. Penile erection was evaluated subjectively by a questionnaire based on a visual analogue scale and objectively by semi quantitative measurement of nocturnal penile tumescence (NPT) using an erectometer. Of the 18 patients, subjective improvements in penile erection were observed in 13 (72%), and objective improvements in NPT were observed in 10 (56%). The administration of rHuEPO may alleviate hyperprolactinemia but was found to have no effect on the FSH, LH, Zn, or HS-PTH level. rHuEPO was suggested to be fairly effective for the treatment of sexual disorders.

Adult↗
PubMed1995

[Biosynthesis of hormones in renal tubular and interstitial cells].

Renal tubular and interstitial cells produce various hormones. Renin is synthesized in juxtaglomerular cells which are derived from the media of the afferent arteriole. Endothelin-1 is formed in mesangial cells of glomeruli and inner medullary collecting duct. In contrast, endothelin-3 synthesis is observed in glomeruli and all the tubule segments. 1 alpha-hydroxylase is present in proximal tubule and catalyzes production of 1,25(OH)2D. PGE2 is synthesized in glomeruli, all the tubule segments, and interstitial cells. The major production sites of PGF2 alpha and 6-keto-PGF1 alpha are glomeruli and medullary collecting duct. Kallikreins are mainly produced in connecting tubule. The cells responsible for erythropoietin synthesis are thought to be interstitial cells which are localized around the proximal tubule.

Animals↗
PubMed1995

Bile duct-specific lectins, Dolichos biflorus agglutinin and peanut agglutinin, as probes in mouse hepatocarcinogenesis.

BACKGROUND: It is well established that alterations in the expression of cell surface glycoproteins occur during the course of tumorigenesis and can be detected immunohistochemically. However, no consistent markers of malignancy in mouse hepatocellular tumors have yet been identified. EXPERIMENTAL DESIGN: Lectin histochemistry, using three bile duct-specific lectins, Dolichos biflorus agglutinin (DBA), peanut agglutinin (PNA) and soybean agglutinin (SBA), and anti-epidermal keratin immunohistochemistry, was conducted on formalin-fixed, paraffin-embedded tissues of a spectrum of benign and malignant hepatocellular proliferative lesions of mice, including hepatocholangiocarcinomas. DBA- and PNA-binding glycoproteins in normal livers and in bile and liver tumors of mice were verified by SDS-PAGE and Western blot analysis. RESULTS: Normal bile duct cells stained strongly with DBA but minimally to moderately with PNA and SBA. DBA-positive tumor cells were present in 96% of hepatocholangiocarcinomas, 89% of hepatocellular carcinomas, and 35% of hepatocellular adenomas. In comparison, 43% of hepatocholangiocarcinomas, 37% of hepatocellular carcinomas, and 24% of hepatocellular adenomas exhibited PNA staining. SBA did not specifically stain tumor cells. Normal hepatocytes and those in altered foci were consistently negative for these three lectins. Keratin-positive staining was found only in normal bile ductular cells and ductal elements in 70% of hepatocholangiocarcinomas. Electrophoresis and Western blot analysis demonstrated that, in normal livers, DBA and PNA bound to the 13- to 16-kDa and 27- to 30-kDa glycoproteins believed to be of bile duct cell origin and commonly present in hepatocellular adenomas, hepatocellular carcinomas, and hepatocholangiocarcinomas, with strongest expression in the last. In addition, hepatocholangiocarcinomas had the same high molecular mass glycoprotein (> 200 kDa) labeled with DBA as detected in bile. CONCLUSIONS: Our results suggest that some malignant hepatocytes, especially in mouse hepatocholangiocarcinomas, have the potential of biliary differentiation. DBA is a sensitive marker for malignant hepatocytes in mice.

Adenoma↗
PubMed1995

[High flow priapism following a straddle-injury-induced arteriocavernosal fistula: a case report].

A 26-year-old man with high flow priapism after blunt perineal trauma, is described herein. Patient evaluation included intracavernal blood-gasometry, cavernography, color flow Doppler sonography. The blood-gasometry showed pH 7.413, pO2 77.9 mmHg, pCO2 41.0 mmHg, HCO2- 26.1 mmol/L, BE 2.0 mmol/L. By direct cavernosography, pooling of contrast agent was seen at the root of the penis. Color flow Doppler sonography revealed pulsatile, turbulent flow within left corpus cavernosum. Our case was diagnosed as high flow priapism from these findings. Detumescence was not achieved by an alpha-adrenergic agent. Superselective embolization of the deep artery of the penis with autologous blood clot was performed with good results. Our case demonstrates that this procedure is a safe and effective therapy for high flow priapism and that erectile function can return to normal.

Adult↗
PubMed1994

Molecular cloning and expression of a member of the aquaporin family with permeability to glycerol and urea in addition to water expressed at the basolateral membrane of kidney collecting duct cells.

Water transport in highly water-permeable membranes is conducted by water-selective pores--namely, water channels. The recent cloning of water channels revealed the water-selective characteristics of these proteins when expressed in Xenopus oocytes or reconstituted in liposomes. Currently, it is assumed that the function of water channels is to transport only water. We now report the cloning of a member of the water channel that also transports nonionic small molecules such as urea and glycerol. We named this channel aquaporin 3 (AQP3) for its predominant water permeability. AQP3 has amino acid sequence identity with major intrinsic protein (MIP) family proteins including AQP-channel-forming integral membrane protein, AQP-collecting duct, MIP, AQP-gamma tonoplast intrinsic protein, nodulin 26, and glycerol facilitator (33-42%). Thus, AQP3 is an additional member of the MIP family. Osmotic water permeability of Xenopus oocytes measured by videomicroscopy was 10-fold higher in oocytes injected with AQP3 transcript than with water-injected oocytes. The increase in osmotic water permeability was inhibited by HgCl2, and this effect was reversed by a reducing agent, 2-mercaptoethanol. Although to a smaller degree, AQP3 also facilitated the transport of nonionic small solutes such as urea and glycerol, while the previously cloned water channels are permeable only to water when expressed in Xenopus oocytes. AQP3 mRNA was expressed abundantly in kidney medulla and colon. In kidney, it was exclusively immunolocalized at the basolateral membrane of collecting duct cells. AQP3 may function as a water and urea exit mechanism in antidiuresis in collecting duct cells.

Amino Acid Sequence↗