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Biomedical subjects

H Handa

Publications and source records attributed to H Handa.

At least 145 records · Page 8Linked to original sources
PubMed1995

Interaction between influenza virus proteins and pine cone antitumor substance that inhibits the virus multiplication.

Fractions obtained from pine cone extract (PCE) of Pinus parviflora Sieb. et Zucc. have been shown to suppress the growth of influenza virus. The inhibitory effects of one of the fractions, Fraction VII, on the formation of RNA-viral protein complex and the viral RNA synthesis were investigated. The formation of M1-RNA or NP-RNA complex was inhibited when M1 or NP was preincubated with the PCE fraction. The in vitro viral RNA synthesis was inhibited by the PCE fraction, while this inhibitory effect was titrated out by the increasing concentration of M1 protein. These results suggest that the major target of the PCE fraction was M1 protein.

Antineoplastic Agents↗
PubMed1995

Replication factor encoded by a putative oncogene, set, associated with myeloid leukemogenesis.

DNA replication of the adenovirus genome complexed with viral core proteins is dependent on the host factor designated template activating factor I (TAF-I) in addition to factors required for replication of the naked genome. Recently, we have purified TAF-I as 39- and 41-kDa polypeptides from HeLa cells. Here we describe the cloning of two human cDNAs encoding TAF-I. Nucleotide sequence analysis revealed that the 39-kDa polypeptide corresponds to the protein encoded by the set gene, which is the part of the putative oncogene associated with acute undifferentiated leukemia when translocated to the can gene. The 41-kDa protein contains the same amino acid sequence as the 39-kDa protein except that short N-terminal regions differ in both proteins. Recombinant proteins, which were purified from extracts of Escherichia coli, expressing the proteins from cloned cDNAs, possessed TAF-I activities in the in vitro replication assay. A particular feature of TAF-I proteins is the presence of a long acidic tail in the C-terminal region, which is thought to be an essential part of the SET-CAN fusion protein. Studies with mutant TAF-I proteins devoid of this acidic region indicated that the acidic region is essential for TAF-I activity.

Amino Acid Sequence↗
PubMed1995

Cell type-specific trans-activation by the B-myb gene product: requirement of the putative cofactor binding to the C-terminal conserved domain.

The myb gene family has three members, c-myb, A-myb and B-myb. We have examined the trans-activating capacity of the B-myb gene product (B-Myb) in various types of cells. B-Myb functions as a transcriptional activator in CV-1 and HeLa cells, but not in NIH3T3 cells, indicating that B-Myb is a cell type-specific transcriptional activator. Deletion analyses of B-Myb have demonstrated that the region conserved between three members of the myb gene family (CR for conserved region) is necessary for trans-activation by B-Myb. An in vivo competition assay suggests that regulatory factor(s) that binds to the CR of B-Myb is required for transactivation. Analyses using an affinity resin show that multiple proteins bind to the CR of B-Myb and that the CR-binding proteins in CV-1 and HeLa cells are different from those in NIH3T3 cells. These results suggest that the CR-binding cofactor(s) is critical for the cell type-specific trans-activation by B-Myb.

3T3 Cells↗
PubMed1995

Silk gland factor-1 involved in the regulation of Bombyx sericin-1 gene contains fork head motif.

Silk gland factor-1 (SGF-1) regulates transcription of the Bombyx sericin-1 gene via interaction with the SA site. In this study, two related SGF-1 polypeptides of apparent molecular masses of 40 and 41 kDa were purified. Specific interaction of these proteins with the SA site was demonstrated by electrophoretic mobility shift and dimethyl sulfate methylation interference assays. The SGF-1 40-kDa protein was partially sequenced and characterized as a new member of the fork head/HNF-3 family. Several full-length cDNAs encoding the SGF-1 40-kDa and possibly also the 41-kDa proteins were cloned and sequenced. SGF-1 mRNA is expressed consistently with the presumed role of the SGF-1 protein product in regulating the sericin-1 gene. The SGF-1 protein contains putative transactivation domains. We conclude that the 40- and 41-kDa SGF-1 proteins affect transcription of the sericin-1 gene via binding to the SA site.

Amino Acid Sequence↗
PubMed1995

Stimulation of DNA transcription by the replication factor from the adenovirus genome in a chromatin-like structure.

Adenovirus (Ad) genome DNA is complexed with viral core proteins in the virus particle and in host cells during the early stages of infection. This DNA protein complex, called Ad core, is thought to be the template for transcription and DNA replication in infected cells. The Ad core functioned as template for DNA replication in the cell-free system consisting of viral replication proteins, uninfected HeLa nuclear extracts, and a novel factor, template activating factor-I (TAF-I) that we have isolated from uninfected HeLa cytoplasmic fractions. The Ad core did not function as an efficient template in the cell-free transcription system with nuclear extracts of uninfected HeLa cells. The addition of TAF-I resulted in the stimulation of transcription from E1A and ML promoters on the Ad core. TAF-I was required, at least, for the formation of preinitiation complexes. These observations suggest that, in addition to factors essential for transcription on naked DNA template, the factor such as TAF-I needed for replication of the Ad core is also required for transcription from the Ad genome in a chromatin-like structure.

Adenoviridae↗
PubMed1995

Characterization of the mitochondrial orfB gene and its derivative, orf224, a chimeric open reading frame specific to one mitochondrial genome of the "Polima" male-sterile cytoplasm in rapeseed (Brassica napus L.).

orf224 is a novel reading frame present upstream of the atp6 gene in the mitochondria of "Polima" cms cytoplasm of rapeseed. In order to determine the origin of orf224, the sequences homologous to orf224 were isolated and characterized. Sequence analysis indicated that orf224 originated by recombination events involving the 5'-flanking region and the amino-terminal segment of the coding region of orf158 (well-known as orfB in other plants), part of exon 1 of the ribosomal protein S3 (rps3) gene, and an unidentified sequence. Transcripts of the orf158 gene were found to be edited at three positions, one of which induces an amino-acid change, while orf224 transcripts have only one RNA editing site within the region homologous to the rps3 gene. This editing site is also present in the proper rps3 transcripts. This result indicates that editing of orf224 occurred because of the sequence homology to rps3. Polyclonal antibodies prepared against a rapeseed ORF158 fusion protein specifically recognize a 18-kDa protein in the membrane fractions of mitochondria from both normal and cms rapeseed.

Amino Acid Sequence↗
PubMed1995

Immortalisation of human oesophageal epithelial cells by a recombinant SV40 adenovirus vector.

We introduced the origin-defective SV40 early gene into cultured human oesophageal epithelial cells by infection of a recombinant SV40 adenovirus vector. The virus-infected cells formed colonies 3-4 weeks after infection in medium containing fetal calf serum. When the cells derived from 'serum-resistant' colonies were then maintained in the serum-free medium with a low calcium ion concentration, some of them passed the cell crisis and kept growing for over 12 months. These cells, regarded as immortalised cells, resembled the primarily cultured oesophageal epithelial cells in morphology and had some of their original characteristics. Treatment of the cells with a high calcium concentration induced phenotypic changes. These cells still responded to transforming growth factor beta. When the immortalised cells were injected into severe combined immunodeficient mice, they transiently formed epithelial cysts, although the typical differentiation pattern of the oesophageal epithelium was not observed. These cysts regressed within 2 months without development into tumours. The results indicated that human oesophageal epithelial cells were reproducibly immortalised by infection with a recombinant SV40 adenovirus vector at relatively high efficiency. The immortalised cells should be useful in studies on oesophageal carcinogenesis and in assessing the cooperative effects with other oncogene products or carcinogens.

Animals↗
PubMed1995

A nuclear factor (NF2d9) that binds to the male-specific P450 (Cyp 2d-9) gene in mouse liver.

Expression of the Cyp 2d-9 (steroid 16 alpha-hydroxylase) gene in mouse liver is male specific in such Mus musculus domesticus strains as FVB/N, whereas the corresponding P450 genes in the wild mouse species Mus spretus are not sex specific in their expression. These parental differences in the gene expressions were independently inherited in F1 offspring from crosses of FVB/N and M. spretus. A 5' flanking sequence (-110CTC CTCCCTATTCCGGGCC-92) was defined as a regulatory element (named SDI-A1) for the domestic Cyp 2d-9 promoter. The nucleotide which corresponds to T at position -99 within SDI-A1 was found to be substituted with C in the wild mouse P450 genes. The placing of C at position -99 abolished the transcriptional activity of SDI-A1 in HepG2 cells as well as the binding of SDI-A1 to a nuclear factor. This factor (designated NF2d9) was purified from mouse nuclear extracts, and its cDNA cloned. The purified NF2d9 bound to SDI-A1 but not to the mutated SDI-A1 with C at position -99. The deduced amino acid sequence revealed that NF2d9 is 72 and 94% identical to mouse CP2 and human LBP-1a, respectively. NF2d9 thus belongs to the CP2 family and is the mouse homolog of human LBP-1a, which modulates human immunodeficiency virus type 1 transcription. Anti-NF2d9, which was raised against the bacterially expressed protein, supershifted the SDI-A1 complex with the liver nuclear extract. Both the bacterially expressed and in vitro-translated NF2d9 inhibited SDI-A1 complex formation, although they did not bind to SDI-A1 directly. The results, therefore, indicate that the domestic Cyp 2d-9 gene can be regulated through a specific association of NF2d9 with SDI-A1.

Amino Acid Sequence↗
PubMed1995

Ventral spinal subdural hematoma--case report.

A 26-year-old male presented with a very rare spinal subdural hematoma which developed 3 days after minor trauma. Magnetic resonance imaging showed the very large hematoma, extending from the C-1 to the T-11 levels, ventral to the spinal cord. An attempt to remove the hematoma via a posterior approach on the next day failed. On the 3rd hospital day, the hematoma was partially removed at the C7-T1 intervertebral level through an anterior approach. Neurological signs of paraplegia, and urinary and bowel disturbances did not improve postoperatively.

Adolescent↗
PubMed1995

Effect of carbachol on phospholipase C-mediated phosphatidylinositol 4,5-bisphosphate hydrolysis, and its modulation by isoproterenol in rabbit corneal epithelial cells.

The effects of carbachol (CCh) on phospholipase C(PLC)-mediated phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis and its modulation by isoproterenol were investigated in SV40-adenovirus transformed rabbit corneal epithelial cells (RCEC). When examined under light microscope, these cells exhibited a cobblestone-like appearance typical of the corneal epithelial cells grown in primary culture. Addition of CCh (0.1 mM) for 30 min to RCEC, prelabeled with 32Pi, decreased the radioactivity in phosphatidylinositol 4-phosphate and PIP2 by 15 and 27%, respectively, and concomitantly increased the radioactivity in phosphatidylinositol and phosphatidic acid by 14 and 38%, respectively. When the concentration of CCh was increased to 1 mM, the changes in radioactivity were even more pronounced. Addition of CCh (0.1 mM) to the cells, prelabeled with myo[3H]inositol, increased the accumulation of [3H]inositol 1,4,5-trisphosphate ([3H]InsP3) by 115%, indicating stimulation of PLC-mediated PIP2 hydrolysis. Similar increases were also observed in [3H]InsP1 and [3H]InsP2. The effects of CCh on inositol phosphate accumulation were time- and dose-dependent, and were inhibited by atropine (10 microM), suggesting that the observed effects of CCh were mediated by activation of muscarinic cholinergic receptors. The effects of CCh were antagonized more potently by 4-diphenylacetoxy N-methyl-piperidine than by pirenzepine, indicating that the muscarinic receptors involved in PLC activation are probably of M3 type. By Western immunoblotting analysis with various anti-PLC antibodies, the RCEC were shown to contain PLC gamma 1 and PLC delta 1 in the soluble fraction and PLC beta 1 in the microsomal fraction. Addition of isoproterenol to RCEC, increased cAMP both in a time- and dose-dependent manner.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic beta-Agonists↗
PubMed1995

Beneficial effects of enteral fat administration on liver dysfunction, liver lipid accumulation, and protein metabolism in septic rats.

The purpose of this study was to examine the effect of different amounts of fat in enteral diets on liver function, liver lipid accumulation, and protein metabolism in septic rats. Sepsis was induced in Wistar rats by cecal ligation and puncture. The rats were divided into four groups and were fed enterally 0% (FO, n = 7), 10% (F10, n = 7), 20% (F20, n = 8), or 30% (F30, n = 9) of total calories as fat. The liquid diet consisted of medium-chain and long-chain triglyceride mixtures as the fat sources, casein oligopeptide, and dextrin (100 kcal/100 ml). Intraduodenum feeding was ended on the 6th day. Serum glutamic oxaloacetic transaminase and glutamic pyruvic transaminase activities, indices of liver dysfunction, were highest in the FO group, and triglycerides accumulated in the livers of that group, possibly because of the large proportion of carbohydrate in the diet. Value of nitrogen balance was highest in the F 10 group, and serum total protein and albumin concentration were higher in the F10 and F20 groups than in the FO and F30 groups. The liver protein content in the F10 and F20 groups was higher than in the FO and F30 groups. Serum triglyceride in the F30 group was about 2 times higher than in the F10 and F20 groups. These results indicate that enteral fat administration in septic rats as 30% of total calories reduced liver dysfunction and liver triglyceride accumulation, but might have been excessive for optimal protein metabolism. Therefore, the preferable amount may range from 10% to 20% of total calories.

Alanine Transaminase↗
PubMed1995

An SV40-immortalized human corneal epithelial cell line and its characterization.

PURPOSE: The authors attempted to immortalize human corneal epithelial cells; it is difficult to propagate primary human corneal epithelial cells because of scarcity of available tissue. However, cell immortalization by virus is always accompanied by shedding of free virus. The current study was performed to establish a cell line that produces no free viral particle. METHODS: Primary cultured human corneal epithelial cells were infected with a recombinant sv40-adenovirus vector and were cloned three times to obtain a continuously growing cell line. Morphologic, cytologic, and biochemical characteristics of this cell line were analyzed. RESULTS: This cell line continued to grow for more than 400 generations, exhibiting a cobblestone-like appearance similar to normal corneal epithelial cells in culture. Transmission electron microscopy showed the evidence for the characteristic features of epithelial cells, including desmosome formation and development of microvilli. It expressed cornea-specific, 64-kD cytokeratin in addition to five major insoluble proteins. By enzymatic analysis using NADP as a coenzyme and a gas chromatograph mass spectrometer, this cell line was found to possess 8.71 IU/mg protein of aldehydedehydrogenase activity. When this cell line was grown at air-liquid interface on collagen type I gel, it differentiated in a multilayered fashion. CONCLUSIONS: The authors have established an SV40-immortalized human corneal epithelial cell line with properties similar to normal corneal epithelial cells.

Aldehyde Dehydrogenase↗
PubMed1995

[Outpatient chemotherapy with continuous infusion of 5-fluorouracil (CI 5-FU) and intravenous bolus leucovorin (IVB LV) in advanced gastrointestinal cancer: the second report].

This study was designed to evaluate the efficacy, toxicity, and quality of life (QOL) of outpatient chemotherapy with 5-FU and LV in advanced gastrointestinal cancer. Treatment consisted of CI 5-FU 450 mg/body/day, days 1-28, IVB LV 30 mg/body q wk (schedule, sch. A) and CI 5-FU 200 mg/m2/day, days 1-28, IVB LV 20 mg/m2 q wk (sch. B). 5-FU was administered with the Baxter infusor (0.5 ml/hr). Oral UFT (400-600 mg/day) was administered after these chemotherapy. Sixteen patients (pts) (sch. A 9 pts, sch. B 7 pts) with advanced gastrointestinal cancer have been treated to maintain the efficacy of prior inpatient chemotherapy. Twenty pts treated as adjuvant chemotherapy were added to evaluate toxicity and QOL. The median time to progression in sch. A was 3.0 months and sch. B was 2.4 months. Grade 3 or 4 toxicities of mucositis were seen 40% in sch. A and 0% in sch. B. Grade 1 or 2 skin toxicities were seen 100% in sch. A and 52% in sch. B. Mean score of QOL in sch. A was 78.0 +/- 11.5, sch. B was 89.5 +/- 7.8 ranked between inpatient chemotherapy (5-FU + CDDP 59.0 +/- 13.8) and UFT (91.8 +/- 6.1). In conclusion, these schedule of 5-FU and LV combination offers a high patient QOL and is suitable for advanced gastrointestinal cancer treatment.

Administration, Oral↗
PubMed1994

Regulation of influenza virus RNA polymerase activity by cellular and viral factors.

An in vitro RNA synthesis system mimicking replication of genomic influenza virus RNA was developed with nuclear extracts prepared from influenza virus-infected HeLa cells using exogenously added RNA templates. The RNA synthesizing activity was divided into two complementing fractions, i.e. the ribonucleoprotein (RNP) complexes and the fraction free of RNP, which could be replaced with RNP cores isolated from virions and nuclear extracts from uninfected cells, respectively. When nuclear extracts from uninfected cells were fractionated by phosphocellulose column chromatography, the stimulatory activity for RNA synthesis was further separated into two distinct fractions. One of them, tentatively designated RAF (RNA polymerase activating factor), stimulated RNA synthesis with either RNP cores or RNA polymerase and nucleocapsid protein purified from RNP cores as the enzyme source. In contrast, the other, designated PRF (polymerase regulating factor), functioned as an activator only when RNP cores were used as the enzyme source. Biochemical analyses revealed that PRF facilitates dissociation of RNA polymerase from RNP cores. Of interest is that virus-coded non-structural protein 1 (NS1), which has been thought to be involved in regulation of replication, counteracted PRF function. Roles of cellular factors and viral proteins, NS1 in particular, are discussed in terms of regulation of influenza virus RNA genome replication.

Biological Factors↗
PubMed1994

Multiple nuclear localization signals of the B-myb gene product.

Nuclear entry of the B-myb gene product (B-Myb) is dependent on multiple nuclear localization signals (NLS's). Mutagenesis of the putative NLS's of B-Myb has identified two separate NLS's, NLS1 and NLS2. Each of the two NLS's is essential for efficient nuclear targeting. NLS2 contains two interdependent basic domains separated by 8 intervening spacer amino acids, and both basic domains are required for nuclear entry. Thus, NLS2 belongs to a class of bipartite NLS's. Like the NLS's in yeast transcription factor SW15, NLS2 contains a putative cdc2 kinase site. However, unlike the case of SW15, phosphorylation at this site did not affect the nuclear targeting of B-Myb.

Amino Acid Sequence↗