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Biomedical subjects

Y Furuta

Publications and source records attributed to Y Furuta.

At least 73 records · Page 4Linked to original sources
PubMed1997

Bone morphogenetic proteins (BMPs) as regulators of dorsal forebrain development.

Bone Morphogenetic Proteins (BMPs) play crucial roles in a variety of developmental processes, but their functions during early vertebrate brain development are largely unknown. To investigate this problem, we have compared by in situ hybridization the expression of five Bmp genes belonging to the Drosophila Decapentaplegic (Bmp2 and Bmp4) and 60A subgroups (Bmp5, Bmp6 and Bmp7). Striking co-expression of these Bmps is observed within the dorsomedial telencephalon, coincident with a future site of choroid plexus development. Bmp co-expression overlaps that of Msx1 and Hfh4, and is complementary to that of Bf1. The domain of Bmp co-expression is also associated with limited growth of the neuroectoderm, as revealed by morphological observation, reduced cell proliferation, and increased local programmed cell death. In vitro experiments using explants from the embryonic lateral telencephalic neuroectoderm reveal that exogenous BMP proteins (BMP4 and BMP2) induce expression of Msx1 and inhibit Bf1 expression, a finding consistent with their specific expression patterns in vivo. Moreover, BMP proteins locally inhibit cell proliferation and increase apoptosis in the explants. These results provide evidence that BMPs function during regional morphogenesis of the dorsal telencephalon by regulating specific gene expression, cell proliferation and local cell death.

Animals↗
PubMed1997

Distribution of herpes simplex virus types 1 and 2 genomes in human spinal ganglia studied by PCR and in situ hybridization.

Clinical data indicate that the recurring herpes simplex virus (HSV) from oro-labial lesions is HSV subtype 1 and that the virus from genital lesions is HSV-2. This suggests that HSV-1 and HSV-2 reside in latent forms in the trigeminal ganglia and sacral ganglia, respectively. However, the distribution of latent HSV-1 and HSV-2 infections in human spinal ganglia has not been fully examined. This report concerns the application of polymerase chain reaction (PCR) and in situ hybridization (ISH) to such a study. By using PCR and employing the respective primers, HSV-1 and HSV-2 DNAs were detected in 207 of 524 samples from 262 spinal ganglia (from the cervical to the sacral ganglia) examined on both sides. The percentages of HSV-1 and HSV-2 detected in a given set of ganglia were similar, indicating an absence of site preference. By ISH, few but positive hybridization signals were detected evenly in sacral ganglia sections. The data suggest that regional specificity of recurrent HSV infections is not due to regional distribution of latent virus, but that local host factors may be important for recurrences.

Adult↗
PubMed1997

Detection of varicella-zoster virus DNA in patients with acute peripheral facial palsy by the polymerase chain reaction, and its use for early diagnosis of zoster sine herpete.

Varicella-zoster virus (VZV) reactivation without cutaneous vesicles (zoster sine herpete) has been demonstrated in 8 to 25% of patients with acute peripheral facial palsy (APFP) by serological methods. To make an early diagnosis of zoster sine herpete, VZV DNA in oropharyngeal swabs from patients with APFP was examined by the polymerase chain reaction (PCR). VZV DNA was detected in oropharyngeal swabs from 6 of 36 (17%) patients with APFP by PCR. VZV DNA was detected in the oropharyngeal swabs from the six patients at their initial visit (2 to 4 days after the onset of APFP), while the anti-VZV IgM and IgG antibody titers were not increased significantly. In contrast, VZV DNA was undetectable in the oropharyngeal swabs at the time when the VZV specific antibody response appeared. These results indicate that detection of VZV DNA in oropharyngeal swabs by PCR is more useful than currently available serological assays for the early diagnosis of zoster sine herpete in patients with APFP.

Acute Disease↗
PubMed1996

Impairment of mobility in endodermal cells by FAK deficiency.

Focal adhesion kinase (FAK) is a novel nonreceptor protein tyrosine kinase that localizes in focal adhesions. It is expressed in a variety of cell types, and we reported earlier that is deficiency causes a decrease of mobility in mesodermal cells with enhanced formation of focal adhesions. With embryoid bodies generated from embryonic stem cells, we also observed a decrease of mobility in FAK-deficient endodermal cells with enhanced focal adhesion formation.

Animals↗
PubMed1996

Hepatitis B virus carriers without precore mutations in hepatitis B e antigen-negative stage show more severe liver damage.

Hepatitis B e antigen (HBeAg) is considered to be a major target for the immune response in chronic hepatitis B. The G-->A mutation at nucleotide 1896 may mediate viral escape by creating a TAG stop codon in the precore region, thus preventing HBeAg production. This mutation frequently evolves during HBe seroconversion if thymine, but rarely if cytosine, is present in position 1858. Applying a combination of polymerase chain reaction (PCR) and restriction enzyme action, we have studied the relation of the TAG mutation and the nucleotide (nt) 1858 variants to liver damage assessed by histology activity index (HAI) scoring in 175 chronic hepatitis B virus (HBV) carriers. A TAG mutation was found in 68 of 71 (96%) of HBeAg-negative carriers infected with a T-1858 strain, but not in any of 33 carriers infected exclusively with a C-1858 strain. Four patients showed a mutation of the precore start codon, and 2 had a TAA stop mutation at codon 2. HBeAg-positive infection with a mixture of wild-type and TAG mutant virus indicated active liver damage, because 8 of 9 (89%) of such patients had a Knodell HAI greater than or equal to 8. In HBeAg-negative stage, both inflammation and fibrosis were more pronounced in carriers infected with wild-type HBV compared with precore mutants. C-1858 strains were associated with more inflammation and fibrosis compared with T-1858 strains. C-1858 strains were found in 71% of northern European, 17% of southern European, 31% of African, 2% of Middle Eastern, and 10% of Far Eastern carriers. Analysis of these variants and mutants may prove useful for clinical evaluation and choice of therapy, and may be facilitated by the methods described.

Adolescent↗
PubMed1996

Localization of annexin I (lipocortin I, p35) mRNA in normal and diseased human skin by in situ hybridization.

Annexin I is a calcium- and phospholipid-binding protein that is involved in the regulation of cellular differentiation. The aim of the present study was to determine the localization of annexin I mRNA expression in normal and diseased human skin. In situ hybridization with a specific digoxigenin-labelled RNA probe was used throughout. We detected no annexin I mRNA signals in basal and suprabasal cells of normal epidermis, but positive signals were evident in the sudoriferous ducts. Annexin I mRNA expression was detected in the keratinizing squamous cells in keratotic type seborrhoeic keratosis and in keratinocytes at the periphery of the horn pearl in well-differentiated squamous cell carcinoma. Positive signals were also seen at the border between involved and noninvolved skin in psoriasis vulgaris and in dyskeratotic epidermal keratinocytes in keratosis follicularis Darier. By contrast, no annexin I mRNA signals were detected in tumour cells in basal cell carcinoma. The present results suggest that annexin I expression is related to, and may play a role in, keratinization disorders.

Annexin A1↗
PubMed1996

Pharmacokinetic studies of human urinary kininogenase in healthy volunteers and animals.

Plasma concentrations of human urinary kininogenase (HUK) were determined in healthy volunteers during and after intravenous (iv) infusion by enzyme immunoassay (EIA). Plasma kinin concentrations were also determined by radioimmunoassay (RIA), and related to HUK concentrations. When HUK was infused [at 0.04, 0.075, 0.15, and 0.3 p-nitroaniline unit (PNAU)/body] over 30 min, plasma HUK concentration rapidly increased and reached a maximum at the end of dosing. Then, the concentration of HUK in plasma decreased biexponentially, and the elimination half-life of the terminal phase was found to be approximately 170 min. The area under the curve of concentration versus time from 0 to 180 min (AUC0-180min) and the maximum concentration (Cmax) increased in proportion to the dose, whereas the pharmacokinetic parameters [mean residense time (MRTinf) = 200-270 min, plasma clearance (CLp) = 2.5-3.3 mL/min/kg, volume of distribution at steady state (Vdss) = 470-730 mL/kg] did not very significantly within the dose range of the present study. On the other hand, when HUK was infused (at 0.15 PNAU/body), plasma kinin concentrations reached approximately 2 ng of bradykinin eq/mL 15 min after the onset of administration. This concentration was maintained during the dosing period, after which kinin was rapidly eliminated, and its concentration returned to baseline at 10 min after dose withdrawal. Plasma kinin concentrations at 15 to 30 min after the onset of dosing (at 0.075, 0.15, and 0.3 PNAU/body) increased in proportion to the dose. The pharmacokinetic parameters of HUK (MRTinf, CLp, Vdss) were compared with those of rats, rabbits, and dogs (log-log plots of body weight versus MRTinf, CLp, and Vdss). The Vdss value showed a good correlation (r = 0.996 for n = 4) with the body weight of respective animal species, the correlation with CLp was weak (r = 0.911), and MRTinf did not exhibit any correlation.

Adult↗
PubMed1996

Failure of ventral body wall closure in mouse embryos lacking a procollagen C-proteinase encoded by Bmp1, a mammalian gene related to Drosophila tolloid.

The mouse bone morphogenetic protein1 (Bmp1) gene encodes a secreted astacin metalloprotease that cleaves the COOH-propeptide of procollagen I, II and III. BMP-1 is also related to the product of the Drosophila patterning gene, tolloid (tld), which enhances the activity of the TGFbeta-related growth factor Decapentaplegic and promotes development of the dorsalmost amnioserosa. We have disrupted the mouse Bmp1 gene by deleting DNA sequences encoding the active site of the astacin-like protease domain common to all splice variants. Homozygous mutant embryos appear to have a normal skeleton, apart from reduced ossification of certain skull bones. However, they have a persistent herniation of the gut in the umbilical region and do not survive beyond birth. Analysis of the amnion of homozygous mutant embryos reveals the absence of the fold that normally tightly encloses the physiological hernia of the gut. At the electron microscopic level, the extracellular matrix of the amnion contains collagen fibrils with an abnormal morphology, consistent with the incorporation of partially processed procollagen molecules. Metabolical labelling and immunofluorescence studies also reveal abnormal processing and deposition of procollagen by homozygous mutant fibroblasts in culture.

Amnion↗
PubMed1995

Mesodermal defect in late phase of gastrulation by a targeted mutation of focal adhesion kinase, FAK.

FAK is a unique non-receptor protein tyrosine kinase that was found in cellular focal adhesions. An increasing number of in vitro observations has suggested that FAK mediates signaling through integrins brought about by interactions with extracellular matrix (ECM). It is highly tyrosine-phosphorylated in v-src-transformed cells and during embryogenesis. To clarify the function of FAK in cell-ECM interactions, embryonic phenotype of its mutant was analysed. FAK-deficient embryos could implant and initiate gastrulation normally, but showed abnormalities in subsequent development. The abnormalities were characterized as a general deficiency in mesoderm, and the phenotype was quite similar to that caused by fibronectin-deficiency. The results suggest that FAK mediates fibronectin-integrin interactions uniquely at this stage of development, thereby playing an essential role in development of mesodermal cell lineages.

Animals↗
PubMed1995

Reduced cell motility and enhanced focal adhesion contact formation in cells from FAK-deficient mice.

The intracellular protein tyrosine kinase FAK (focal adhesion kinase) was originally identified gy its high level of tyrosine phosphorylation in v-src-transformed cells. FAK is also highly phosphorylated during early development. In cultured cells it is localized to focal adhesion contacts and becomes phosphorylated and activated in response to integrin-mediated binding of cells to the extracellular matrix, suggesting an important role in cell adhesion and/or migration. We have generated FAK-deficient mice by gene targeting to examine the role of FAK during development. Mutant embryos displayed a general defect of mesoderm development, and cells from these embryos had reduced mobility in vitro. Surprisingly, the number of focal adhesions was increased in FAK-deficient cells, suggesting that FAK may be involved in the turnover of focal adhesion contacts during cell migration.

Animals↗
PubMed1995

Focal adhesion kinase is not essential for in vitro and in vivo differentiation of ES cells.

Focal adhesion kinase, FAK, is a unique protein tyrosine kinase found in cellular focal adhesions. It is widely expressed and highly phosphorylated during embryogenesis. To examine the function of FAK in cell differentiation, we made FAK-deficient embryonic stem (ES) cells by homologous recombination. However, FAK-deficiency did not interfere with differentiation of the ES cells into cells of three germ layers when implanted subcutaneously into nude mice or when treated with retinoic acid in vitro, nor was there any evidence of defects in hematopoiesis in vitro.

Animals↗
PubMed1995

Frontal cingulotomy reconsidered from a WGA-HRP and c-Fos study in cat.

A recent positron emission tomography (PET) study demonstrated that the anterior cingulate cortex (area 24), in addition to SI and SII cortices, was activated by painful stimuli. In order to elucidate the participation of relay nuclei in the ascending pain pathway to area 24, we performed a regrograde labelling study with WGA-HRP injection into area 24 in cats. Area 24 was found to receive pain-related thalamic inputs from the intralaminar nuclei including the central medial nucleus, midline nuclei, modiodorsal nucleus and possibly the submedial nucleus. We then examined the expression of Fos protein in CNS induced by formalin injection into the face in cats. Fos positive neurons were demonstrated in areas 23 and 24, the anterior limbic area, insular cortex, midline and paraventricular nuclei in the thalamus, paraventricular nucleus and other areas in the hypothalamus, and in many nuclei in the brainstem in both the formalin-injected group and the control group (anesthesia only). Labelled regions appeared to correspond to stress-related sites. The sole difference from the control group was the expression of Fos in the coronal gyrus and in the trigeminal caudalis nucleus in the experimental group. Although more Fos positive cells were observed in area 24 in experimental than in control cats, the difference was not significant. Our findings suggest that the demonstrated response of area 24 on PET scan represents stress- and emotion-related events rather than pain. Surgical intervention into the anterior cingulate cortex including cingulotomy thus appears to relieve stress and emotion associated with chronic pain, but not pain itself.

Animals↗
PubMed1995

A new strategy of gene trapping in ES cells using 3'RACE.

"Gene trapping" in embryonic stem (ES) cells is a novel approach to identify a series of genes in mammals concomitant with the production of the corresponding mutant mice. However, this approach is currently unable to identify genes that are not expressed in ES cells. Here we describe a strategy to identify gene trapping clones which is not based on expression of a reporter gene. It uses the neor gene which lacks a polyadenylation signal and has a splice donor signal. Expression of the neor gene as fusion transcripts with the 3' end containing the polyadenylation signal of tagged genes allows the identification of these clones by 3' rapid amplification of the cDNA end in undifferentiated ES cells, even if the genes are not expressed in ES cells. Amplification was observed in about 25% of G418-resistant clones. Sequence analyses suggested the amplifications represent gene trapping events. The feasibility of this approach was further assessed by analysing one clone, PAT-12, in detail.

Amino Acid Sequence↗
PubMed1995

Detection of hepatitis B virus precore TAG mutant by an amplification-created restriction site method.

A new method for detecting the hepatitis B virus (HBV) precore 1896 G-->A mutation is described. This mutation prevents hepatitis B e antigen production by introducing a TAG stop codon and has been associated with severe chronic and fulminant hepatitis. The method is based on a polymerase chain reaction (PCR) that creates a restriction enzyme (Bsu36I) cleavage site if the mutation is present. After incubation of the PCR product with Bsu36I and a subsequent agarose gel electrophoresis, the presence of the TAG mutant is revealed by an altered position of the DNA band. The method was compared with direct sequencing on 36 serum samples and correctly identified all samples containing mutant HBV. The TAG mutant was present in 17 cases (as mixed wild type and mutant virus in 4). Twelve of 18 patients with advanced liver disease confirmed by biopsy carried mutant HBV. This method of detecting HBV precore 1896 G-->A should be useful for evaluation and follow-up of patients and for prevalence studies.

Adolescent↗
PubMed1995

Emergence of precore TAG mutation during hepatitis B e seroconversion and its dependence on pregenomic base pairing between nucleotides 1858 of 1896.

The G-->A mutation of nucleotide (nt) 1896 in the precore region of hepatitis B virus (HBV) prevents production of hepatitis B e antigen (HBeAg) by creating a TAG stop codon. This mutation is often found in anti-HBe-positive patients with productive HBV infection and has been associated with severe chronic and fulminant hepatitis in some geographic areas. Emergence of the TAG mutation during HBe seroconversion was studied as was its relationship with nt 1858, which forms a base pair with nt 1896 in the pregenomic RNA loop. A TAG mutant evolved in 18 (72%) of 25 patients with a T-1858 strain but in only 1 (8%) of 13 with a C-1858 strain. Viremia with C-1858 strains was, despite their limited ability to develop the TAG mutation, as persistent as with T-1858 strains. Generally, T-1858-infected patients in whom a TAG mutant did not emerge were HBV DNA-negative by polymerase chain reaction on follow-up, whereas patients who developed the TAG mutation had prolonged viremia.

Adolescent↗
PubMed1995

Ovarian teratomas in mice lacking the protooncogene c-mos.

Parthenogenesis has been suggested to be tightly coupled with development of ovarian teratomas. Indeed, ovarian tumors developed in c-mos-deficient female mice, which are characterized by the parthenogenetic activation of oocytes. The tumors appeared at a frequency of 30% between 4 and 8 months of age, and did not develop in younger or older mice. Most of the tumors were benign and consisted of multi-focal cysts most notably with mature ectodermal components, but also with mesodermal and endodermal components. One among 17 tumors observed consisted of extra-embryonic tissues alone, and two bore malignant components with metastasis to peritoneal organs. The results strongly suggest the involvement of c-mos mutations in human germ cell tumors.

Age Factors↗
PubMed1994

Infection of vaginal and colonic epithelial cells by the human immunodeficiency virus type 1 is neutralized by antibodies raised against conserved epitopes in the envelope glycoprotein gp120.

The rectal and genital tract mucosae are considered to be major sites of entry for the human immunodeficiency virus (HIV) during sexual contact. We now demonstrate that vaginal epithelial cells can be infected by HIV type 1 (HIV-1) via a mechanism similar to that described for neuroglial cells and, more recently, for colorectal epithelial cells, involving initial interaction of the HIV-1 envelope glycoprotein gp120 with a cell-surface glycosphingolipid (sulfated lactosylceramide). A hyperimmune serum against gp120 was able to neutralize HIV-1 infection of vaginal epithelial cells. Site-directed immunization was employed to identify sites on gp120 recognized by antibodies neutralizing HIV-1 infection of vaginal and colonic epithelial cells. Hyperimmune sera were raised in monkeys against a series of 40 overlapping synthetic peptides covering the entire sequence of HIV-1 (HTLV-IIIB) gp120. Antisera raised against five synthetic peptides, corresponding to three relatively conserved regions and to the hypervariable region (V3 loop), efficiently neutralized HIV-1 infection of human vaginal epithelial cells in vitro. Similar results were obtained with the colonic cells. Hyperimmune sera to all five peptides have been shown earlier to neutralize HIV-1 infectivity in CD4+ T cells. These results have obvious implications for the design of mucosal subunit vaccines against sexually transmitted HIV-1 infections.

Base Sequence↗