PubMed Health⌕ Search

Biomedical subjects

H Nakabayashi

Publications and source records attributed to H Nakabayashi.

At least 19 recordsLinked to original sources
PubMed1996

Gene therapy for alpha-fetoprotein-producing human hepatoma cells by adenovirus-mediated transfer of the herpes simplex virus thymidine kinase gene.

We have developed a recombinant replication-defective adenovirus containing human alpha-fetoprotein (AFP) promoter/enhancer to direct cell type-specific expression of the herpes simplex virus thymidine kinase (HSVtk) gene to AFP-producing hepatocellular carcinoma (HCC) cells. After an in vitro infection by a recombinant adenovirus carrying the lacZ gene under the control of human AFP promoter/enhancer (AdAFPlacZ), an expression of the lacZ gene was demonstrated efficiently in AFP-producing HuH-7 and HepG2 cell lines, but not in AFP-nonproducing HLE and HLF cell lines, although lacZ gene expression was demonstrated in all these cell lines when infected with adenovirus vector carrying lacZ gene driven by the beta-actin-based promoter. Expression of the HSVtk gene by adenovirus, from AFP promoter/enhancer (AdAFPtk) induced the cells sensitive to ganciclovir (GCV) in the AFP-producing cell line efficiently, but not in AFP-nonproducing HLF hepatoma cells. An in vitro bystander effect was observed when only 10% of the cells were infected with AdAFPtk. These findings suggest that the AFP promoter/enhancer sequence can provide the tumor-specific activity for the therapeutic gene expression, and that the AdAFPtk vector induces the selective growth inhibition by GCV in the adenovirus-infected human hepatoma cells in vitro. Recombinant adenovirus transfer of the HSVtk gene under the control of tumor-specific promoter followed by GCV may have promise as a targeted in situ treatment for solid neoplasms.

Adenoviridae↗
PubMed1996

[Detection of p27/kip1 mRNA in blood cells by nonradioactive ribonuclease protection assay].

Cyclin and cyclin-dependent kinase(cdk) complexes, and their inhibitors (CKIs) play important roles in growth regulation on the cells. p27/kip1 is a CKI associated with G1 arrest induced by cell to cell contact, transforming growth factor-beta and cyclic AMP. The abnormality of p27/Kip1 genes in human tumors usually appears as a steady level defect of expression, since mutations in them is rare. Thus it is important to estimate the expression level of this gene. To detect the change of p27/Kip1 mRNA level in blood cells, we developed the ribonuclease protection assay using nonradioactive riboprobe which was produced by reverse transcriptase-polymerase chain reaction (RT-PCR) with T7 promoter-added antisense primer and the in vitro transcription system. Our assay may be useful for clinical evaluation of the mRNA level.

Base Sequence↗
PubMed1996

[Regulation of tumor marker gene expression].

Tumor marker is a group of proteins that specifically expressed in association with carcinogenic processes. Thus, studies of regulation mechanisms of the tumor marker genes are important for understanding the possible alteration of gene expression during neoplastic transformation. In this article, we described the molecular mechanisms of the regulation of two major tumor marker genes, i.e. human alpha-fetoprotein (AFP) gene and rat glutathione transferase P(GST-P) gene. Positive and negative mechanisms of each gene regulations are discussed.

Animals↗
PubMed1995

Adenovirus-mediated gene therapy of hepatocellular carcinoma using cancer-specific gene expression.

Most patients with hepatocellular carcinoma have an elevated alpha-feto-protein (AFP) level. This high level of AFP expression is transcriptionally controlled by the 5'-flanking sequence of the AFP gene. Using the 5'-flanking sequence as a promoter for the herpes simplex virus thymidine kinase (HSV-TK) gene in an adenoviral vector (Av1AFPTK1), the therapeutic efficacy of adenovirus-mediated HSV-TK gene transduction, followed by ganciclovir (GCV) administration, was studied in tumors in athymic nude mice. Av1AFPTK1 transduction of two cell lines demonstrated HSV-TK enzyme activity only in the AFP-producing cells (HuH7) and not in the AFP nonproducing cells (SK-Hep-1). As expected, only transduced HuH7 cells were killed by GCV treatment. Transduction by an adenoviral vector harboring a Rous sarcoma virus promoter and HSV-TK gene (Av1TK1) showed enzymatic activity and GCV killing in both cell lines. All HuH7 tumors that were transduced with either Av1AFPTK1 or Av1TK1 completely regressed after GCV treatment. On the other hand, there was complete regression of SK-Hep-1 tumors only when treated with Av1TK1 and GCV and not when treated with Av1AFPTK1 and GCV. Thus, cell-specific killing was achieved by adenoviral vector containing AFP promoter for the HSV-TK gene and GCV treatment.

Adenoviridae↗
PubMed1995

Isolation and sequence polymorphism of a rat retinoblastoma (RB) cDNA.

A cDNA of the rat retinoblastoma gene (RB) was prepared from total RNA of rat liver using reverse transcription-polymerase chain reaction (RT-PCR). The 4432-nt sequence isolated contained 2700-nt translated and 1732-nt 3'-untranslated regions (UTR). The isolated cDNA detected poly(A)+RNAs of 5.4 and 3.4 kb in rat liver and kidney by Northern blot hybridization. The nt sequence of the isolated cDNA had 85% homology with that of mouse and 73% with human. The 899-amino-acid (aa) sequence was 95% homologous to that of mouse and 90% to human. The aa sequences of two functional domains of oncoprotein-binding and ten putative phosphorylation sites regulating RB function were conserved in the three species. However, the 3'-UTR were less homologous among the three, and had polymorphism in three portions, even in rats. These polymorphisms were strain-specific and genetically segregated. Thus, the rat RB cDNA and its sequence information may be useful for clarifying the role of the RB protein and genetic linkage analysis in basic biomedical research using rats, especially in experimental carcinogenesis.

Amino Acid Sequence↗
PubMed1995

[Microsatellite polymorphism of muscle glycogen synthase gene and non-insulin dependent diabetes mellitus].

It has been reported recently that A2 allele of muscle glycogen synthase gene determined by Xba I RFLP (restriction fragment length polymorphism) is related to the frequency of NIDDM in Finland. Using PCR-RFLP analysis, we detected no A2 allele in Japanese patients with NIDDM and control. We found a new microsatellite of CA repeat in the human muscle glycogen synthase gene, and the polymorphism of the repeat number was detected by the polymerase chain reaction. Six different alleles were observed, indicating the highly polymorphic nature of this marker. We analyzed the repeat numbers and the genotypes in diabetics and control. The allele frequencies were not significantly different between NIDDM and control. However, the allele frequencies in the patients without high total cholesterolemia was significantly different from those of control. These findings suggest that the muscle glycogen synthase gene or neighboring genes are related to one of the disease genes of diabetes mellitus without high total cholesterolemia.

Adolescent↗
PubMed1995

Reduced dimerization of lipoprotein lipase in post-heparin plasma of a patient with hyperchylomicronemia.

As in post-heparin plasma of control subjects, post-heparin plasma of a patient with hyperchylomicronemia contained lipoprotein lipase (LPL) subunits with M(r) = 57,000. But although the amount of LPL was the same as in post-heparin plasma of controls, no LPL activity was detectable. Nearly all the LPL in post-heparin plasma of controls bound to heparin-Sepharose and this LPL bound was mainly eluted with 1.5 M NaCl in parallel with the activity. In post-heparin plasma of the patient, 58% of the LPL subunits did not bind to heparin-Sepharose and 23% was eluted with 0.6 M NaCl. Studies by sucrose density gradient centrifugation showed that almost all the LPL in post-heparin plasma of controls was recovered in the peak with a sedimentation coefficient of 6.8 S, corresponding to the position of a dimeric form of LPL, in parallel with the activity; little LPL was recovered in the peak with a sedimentation coefficient of 4.0 S, corresponding to the position of a monomeric form of LPL. In post-heparin plasma of the patient, 35% of the LPL subunits was recovered in fractions with larger sedimentation coefficients at the bottom of the centrifuge tube, indicating the presence of an aggregated form(s) of LPL; the amount of the monomeric form of LPL was increased, while that of the dimeric form was decreased. Thus, defect of LPL activity in post-heparin plasma of the patient with hyperchylomicronemia could result from reduced dimerization of LPL subunits.

Adolescent↗
PubMed1995

Somatostatin receptor on the afferent nerve terminals in the rat hepatoportal area.

To determine whether somatostatin receptor (SSR) actually exists on the nerve terminals in the rat hepatoportal area, the area was immunostained by the labeled streptoavidin-biotin complex method using a monoclonal antibody against rat brain SSR. The SSR staining revealed many fiber arborizations with terminal nodular swellings like the afferent nerve endings in the neural body, which was located beneath the endothelium of the large branches of the intrahepatic portal vein. The results indicate that SSR is expressed on the characteristic structure, adding further evidence for our previous observation which showed the hepatic vagal reception for intraportal somatostatin and the somatostatin-binding neural body as a relevant structure.

Afferent Pathways↗
PubMed1995

Proliferative potential of meningiomas evaluated by proliferating cell nuclear antigen expression.

Meningiomas are principally benign in nature. Some meningiomas, however, grow fast or recur even after total removal. The biological behavior of meningiomas often can not be predicted from conventional histopathological studies. A monoclonal antibody against proliferating cell nuclear antigen (PCNA) was used to investigate the usefulness of the PCNA index as a parameter to estimate the proliferative activity of meningiomas. Fifty-two meningiomas were examined. The mean PCNA index of recurrent meningiomas (3.37 +/- 0.92%) was significantly higher than that of non-recurrent meningiomas (1.12 +/- 0.51%) (p < 0.005). The PCNA indices of recurrent cases were all higher than 2.0%. A semilog linear regression analysis between tumor doubling time and PCNA index showed a significant correlation (r = 0.90, p < 0.05). An inverse linear correlation between PCNA index and interval to recurrence was observed (r = 0.62, p < 0.05). A good linear correlation was also shown between PCNA index and BUdR labeling index (r = 0.88, p < 0.01). The results of this study suggest that, providing the methods of tissue processing, immunostaining and counting of positive nuclei are unified, the PCNA index is a useful parameter for estimating the biological behavior of meningiomas.

Adult↗
PubMed1995

Defective copper binding to apo-ceruloplasmin in a rat model and patients with Wilson's disease.

To examine the mechanism of decrease in serum ceruloplasmin (Cp) in Long-Evans Cinnamon (LEC) rats, a proposed model of Wilson's disease, we analyzed Cp products at the stages of transcription and translation. Northern blot analysis and immunoblot analysis showed that the level and the molecular size of Cp mRNA and protein in LEC rats were similar to those in control Long-Evans-Agouti (LEA) rats. However, the ferroxidase activity of Cp was significantly decreased in LEC rats. We separated serum Cp into two forms by native polyacrylamide gel electrophoresis with pH modification: one was a holo-Cp with copper and ferroxidase activity, and the other was an inactive apo-Cp without copper. Holo-Cp was the predominant form in LEA rats and normal humans, whereas apo-Cp was the major form in LEC rats and patients with Wilson's disease. The cosegregation of apo-Cp predominance with the disease in LEC rats was analyzed using backcross rats. Apo-Cp was dominant in 8 of 11 offspring with disease but in none of 19 normal offspring. These results indicate that a genetic disturbance of copper binding to apo-Cp may be closely associated with the pathogenesis in LEC rats, and probably in Wilson's disease.

Animals↗
PubMed1995

A silencer element for the lipoprotein lipase gene promoter and cognate double- and single-stranded DNA-binding proteins.

Transfection experiments with constructs containing various 5'-deleted fragments of the human lipoprotein lipase (LPL) promoter and the chloramphenicol acetyltransferase reporter gene revealed an LPL silencer element (LSE) in the region of nucleotides -225 to -81 of the LPL gene that functioned in Chinese hamster ovary (CHO) and HeLa cells. Gel retardation competition analysis showed the presence of a nuclear factor(s) capable of binding to the sequence of nucleotides -169 to -152 of LSE (LSE-6) in a single-stranded (opposite-strand) and double-stranded specific fashion, the binding affinity being almost the same in the two binding forms. Site-directed mutagenesis indicated that almost the entire sequence of LSE-6 was necessary to form the complexes and also critical for silencing activity in CHO cells. The amounts of this binding factor(s) in CHO and HeLa cells were closely associated with transcriptional silencing activity. Photochemical cross-linking experiments indicated that the single- and double-stranded elements recognized the same binding factor(s) with molecular masses of 54 to 63 kDa and 109 to 124 kDa. The 109- to 124-kDa DNA binding factor(s) was found to be a doublet of that of the 54- to 63-kDa factor by isoelectric focusing or by increasing the time of exposure to UV irradiation. When inserted upstream of another gene such as that of the simian virus 40 enhancer/promoter of pSV2CAT, the sequence of nucleotides -190 to -143 (LSE-1) also suppressed transcription of the reporter gene in CHO cells. These results strongly suggest that the LSE plays a role in regulation of LPL gene expression by suppressing its transcription.

Animals↗
PubMed1995

[A case of multilocular thymic cyst with myasthenia gravis].

We reported a case of 53-year-old male who was admitted to our hospital for ptosis and difficulty in chewing. Because of positive for Tensilon test and high dose anti-acetylcoline receptor antibody, we established diagnosis for myasthenia gravis. Additionally he was pointed out a cystic lesion at anterior superior mediastinum on CT and MRI. As a result we diagnosed him as thymic cyst with myastenia gravis. The extended thymectomy was performed. Histological examination revealed that the mass was a multilocular thymic cyst which is reported by Suster, who suggested that the multilocular thymic cyst arises from processes of reactions to an acquired inflammatory change. There was no report for cases of multilocular thymic cyst with myastenia gravis. MTC-like changes are sometimes presented in association with thymic Hodgkin's disease or thymic seminoma. Malignant transformation of thymic cyst were also reported. So careful examination and evaluation for cystic lesion within thymus are required.

Humans↗
PubMed1995

[Clinical significance of measurement of a novel basement membrane-associated collagen levels in serum].

We measured serum levels of a basement membrane-associated collagen(BMC) by enzyme linked immunosorbent assay using two recently produced monoclonal antibodies (named JK-199 and JK-135) to evaluate that's clinical significance. JK-199 reacted with alpha 1 (IV) chain in triple helical region of type IV collagen and JK-132 is considered to react with an unidentified alpha (IV) related collagen chain. BMC was determined in sera of patients with chronic liver diseases (n = 27), healthy volunteers (n = 30) and in random serum samples of in- or outpatients of our hospital (n = 1125). The degree of fibrosis of the chronic liver diseases was estimated by computed image analysis for the biopsy specimens. BMC levels in sera of healthy volunteers as 0-40ng/ml. Serum BMC levels were significantly higher than that in healthy volunteers in chronic active hepatitis 2A (56.9 +/- 27.9ng/ml; mean +/- SD), 2B (80.9 +/- 39.1ng/ml) and liver cirrhosis (114.5 +/- 26.6ng/ml). Serum BMC levels showed positive correlation with fibrosis area/total area(%) in biopsy specimens of chronic liver diseases (r = 0.678; p < 0.001). Total 149 samples (from 127 patients) of 1125 random serum samples (13.2%) showed increased BMC and 70 of 127 patients with abnormal high BMC (55.1%) had liver diseases. These results suggest that serum BMC increases mainly in patients with liver diseases, especially with progression of chronic liver diseases.

Adult↗
PubMed1995

[A case of pulmonary tumorlet with tuberculoma misdiagnosed as small cell lung carcinoma by transbronchial lung biopsy].

A 59-year-old woman was picked up by chest X-ray findings. Her CT scan film showed a tumor shadow in the upper lobe of the left lung, and by transbronchial lung biopsy, she was diagnosed as a small cell lung carcinoma. Lobectomy was performed, but the lesion was diagnosed as tuberculoma by frozen section. Furthermore, on postoperative histological examination, a minute lesion of tumorlet, which was histologically identical to that preoperatively detected by transbronchial lung biopsy was found in the adjacent tissues of the tuberculoma. Pulmonary tumorlet is clinically a rare lesion, but the diagnosis and treatment for small-sized tumor or tumor-like lesion may be carefully done, taking into account the existence or co-existence of tumorlet lesion.

Biopsy↗
PubMed1994

Sequence of the 5'-flanking region of the gene encoding human muscle glycogen synthase.

The 5'-flanking region of the gene encoding human muscle glycogen synthase was isolated from a human placental genomic library and sequenced. The sequence is TATA-less and G+C-rich, and putative transcription-controlling sequences were identified. Furthermore, a simple (dC-dA)n sequence repeat was identified about 4 kb upstream from the start codon. This sequence was highly polymorphic and five alleles were typed in the Japanese population using the polymerase chain reaction.

Animals↗
PubMed1994

A unique receptor-mediated mechanism in vagal chemoreception of somatostatin in the hepatoportal area.

Our previous results have indicated that the hepatic vagus is receptive to intraportal appearance of somatostatin-14 (SS), a gastroenteropancreatic peptide hormone, in rat. To examine the actual involvement of the SS receptor mechanism in the neural reception, the receptor function was modified with monoclonal antibody against rat neural SS receptor. Our monoclonal antibody of IgG1(kappa) isotype bound to the SS binding site of the receptor in competition with the ligand. Intraportal SS injection at a physiological dose (3.05 pmol) significantly facilitated the afferent firing discharge rate of the hepatic vagus in rats anesthetized with urethane and chloralose. The antibody ascites, but not the control ascites, administered intraportally 5 min prior to the SS injection abolished completely the SS-induced increase in discharge rate. However, when the antibody was administered more than 15 min after the SS injection, the SS-induced increase in discharge rate did appear, implying an unique post-receptor mechanism in reception. The present results indicate that the SS receptor mechanism operates in the neurochemoreception of SS, suggesting the existence of a hepatic vagal, receptor-mediated system for monitoring the hormone secreted from the splanchnic organs.

Afferent Pathways↗
PubMed1994

Marked improvement of diabetic diarrhea with the somatostatin analogue octreotide.

Somatostatin and its long-acting analogue octreotide have been used in various diarrheal disorders, including neoplastic and nonneoplastic diseases of the gastrointestinal tract. In two insulin-dependent diabetic patients with autonomic neuropathy and chronic steatorrheic diarrhea refractory to conventional medications, subcutaneous administration of octreotide markedly improved the volume and frequency of stools in both patients. This change was accompanied by a clear improvement in their rapid gastrointestinal tract transit times. The treatment also greatly improved their orthostatic hypotension. No adverse effects of octreotide were observed after treatment for 7 months in one patient and 2 months in the other.

Adult↗