[NANC relaxation and potassium channel in rat colon and rectum].
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Acetylcholine↗
Biomedical subjects
M Kishi
Publications and source records attributed to M Kishi.
Nonsense mutations in the vpr gene of HIV-1 during in vitro virus passage and in HIV-1 carrier-derived peripheral blood mononuclear cells.
Long-term, persistent infection by HIV-1 is a prerequisite for the development of AIDS. However, little is known of the determinants required for HIV-1 to cause persistence. We have reported previously that persistent infection of a T cell line by a cytopathogenic strain of HIV-1 became increasingly likely with in vitro serial passage of the virus. DNA sequencing of the persistent strains revealed a nonsense mutation in the vpr gene in all isolates tested. Here, we report the development and use of a semi-quantitative PCR method to detect the vpr nonsense mutation within populations of virus. Our results show that vpr mutants also arise in cells during acute infection and increase progressively with serial passage of the virus. In addition, HIV-1-seropositive individuals were examined and found to carry the same vpr nonsense mutation at high frequency in virus-infected PBMC. These data are consistent with a mechanism of HIV-1 persistence in vivo and in vitro in which virus cytopathogenic potential is lost by the build up of nonsense mutations in vpr.
AIDS-Related Complex↗
FTIR spectral study of intramolecular hydrogen bonding in thromboxane A2 receptor agonist (U-46619), prostaglandin (PG)E2, PGD2, PGF2 alpha, prostacyclin receptor agonist (carbacyclin), and their related compounds in dilute CCl4 solution: structure-activity relationships.
FTIR spectra measurements and full optimization curve analysis of their spectra were done to obtain parameters of the OH and C = O stretching vibration bands for intramolecular hydrogen bondings in thromboxane (TX)A2 receptor partial agonist (CTA2), prostaglandin (PG)E2, PGD2, PGF2 alpha, prostacyclin (PGI2) receptor agonist (carbacyclin), and their related compounds in dilute CCl4 solutions. For CTA2, PGE2, PGD2, and PGF2 alpha, cyclic intramolecular hydrogen bonds involving a 15-membered ring similar to that observed for the TXA2 receptor agonist (U-46619) were found between a carboxyl group of the alpha-side chain and a 15-hydroxyl group of the omega-side chain. The arrangement of these side chains was P-shaped, and the percentage of the intramolecular hydrogen-bonded molecules with the 15-membered ring in CCl4 solution showed a high value of ca. 80% for these compounds. In addition, it was found that the cyclic intramolecular hydrogen bonds involving the 13-, 12-, and 12-membered rings in PGE2, PGD2, and PGF2 alpha, respectively, are formed between the carboxyl group of the alpha-side chain and the 11-, 9-, and 9-hydroxyl groups of a cyclopentane ring, respectively, although the percentages of the intramolecular hydrogen-bonded molecules with these membered rings are very small. It was also found that the hydrogen bond is more easily formed in the order of the 11-, 9-, and 15-hydroxyl groups. For carbacyclin, the cyclic intramolecular hydrogen bond involving the 13-membered ring was found between the carboxyl group of the alpha-side chain and the 11-hydroxyl group. The percentage of the intramolecular hydrogen-bonded molecules showed the value of 58% for carbacyclin. On the basis of information on the side-chain conformations in CCl4, we examined the structure-activity relationships for U-46619 in place of TXA2, PGE2, PGD2, PGF2 alpha, and carbacyclin in place of PGI2.
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
Persistent infection of MT-4 cells by human immunodeficiency virus type 1 becomes increasingly likely with in vitro serial passage of wild-type but not nef mutant virus.
Our previous studies have shown that human immunodeficiency virus type 1 (HIV-1), with mutations in accessory genes such as vif, vpr or vpu, can generate persistent infection of MT-4 cells, whereas infection by wild-type or nef mutant HIV-1 causes extensive cell death. The possibility of generating a naturally attenuated form of HIV-1 with reduced cytopathogenicity in MT-4 cells was examined by in vitro serial passage of the wild-type and a nef mutant form of HIV-1, each derived from the infectious molecular clone pNL432. The ability to cause persistent infection was observed after four passages of wild-type HIV-1 with the frequency of persistence becoming progressively higher with serial passage. In contrast, persistent infection was not observed even after 50 passages of the nef mutant virus. Sequence analysis of the accessory gene loci in genomes recovered from the persistent infections caused by passaged virus revealed mutations in vif and vpr, but not in vpu. The processing of the Env precursor to mature forms was not modified in any of the passages of either wild-type or nef mutant HIV-1. However, when compared with acute infections caused by similarly passaged virus of both wild-type and nef mutant HIV-1, persistent infections by passaged wild-type HIV-1 showed a significant decrease in the cell surface expression and function of Env. Cell surface CD4 was only partially down-regulated on cells acutely infected with the passaged viruses, whereas on cells persistently infected with passaged wild-type HIV-1 it was completely down-regulated. These results suggest that, during serial passage of HIV-1, mutations accumulate at least in the accessory genes vif and vpr in parallel with a lesser interaction between cell surface Env and CD4 molecules, and lead to the generation of less cytopathogenic viruses capable of persistent infection. Our results also suggest an important role for the nef gene product in the generation of HIV-1 strains that are less cytopathogenic.
Amino Acid Sequence↗
[Molecular basis for HIV infection: implication for the therapy].
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AIDS Vaccines↗
Persistently human immunodeficiency virus type 1-infected T cell clone expressing only doubly spliced mRNA exhibits reduced cell surface CD4 expression.
Several cell clones possessing the human immunodeficiency virus type 1 (HIV-1) genome, consisting of an almost full-length DNA sequence, were isolated by limiting dilution of the clonal cell line M10 derived from MT-4 that survived infection with HIV-1 vpr mutant (M10/vpr-). One of the isolated clones (termed Vpr-1) expressed only doubly spliced mRNA, but not unspliced or singly spliced mRNA. Western blots of Vpr-1 revealed the presence of the nef translation product, although no expression of major structural genes such as gag, pol, and env was detected by indirect immunofluorescence and assay of reverse transcriptase activity. These HIV-1 phenotypes differed greatly from those of the original M10/vpr-, most of which expressed major structural HIV-1 proteins. Despite undetectable levels of env expression in Vpr-1, CD4 antigens were greatly down-modulated on the surface without alteration of steady-state levels of CD4 mRNA expression, similar to M10/vpr-. These HIV-1 phenotypes in Vpr-1 did not change after the treatment of the cells with both phorbol 12-myristate 13-acetate and phytohemagglutinin. Therefore, the abnormal HIV-1 life cycle in Vpr-1 seems to be due to some viral factor(s), as well as cellular factors. Thus, Vpr-1 could be a useful model for understanding one HIV-1 latent form.
Base Sequence↗
Finite element model of facial soft tissue. Effects of thickness and stiffness on changes following simulation of orthognathic surgery.
A study was conducted to investigate the influence of the thickness and stiffness of soft tissue on changes following orthognathic surgery. The finite element method was used to simulate surgery. Three finite element models with normal thickness and stiffness, twice the thickness and twice the stiffness were made in preparation for the simulation. The thickness of soft tissue had more effect than stiffness on posterior and vertical changes.
Cephalometry↗
Xanthoquinodins, new anticoccidial agents produced by Humicola sp. Production, isolation and physico-chemical and biological properties.
Humicola sp. FO-888, a soil isolate, was found to produce a series of new anticoccidial compounds. Five active compounds, designated xanthoquinodins A1, A2, A3, B1 and B2 were isolated from the fermentation broth of the producing strain by solvent extraction and HPLC. Xanthoquinodins inhibited the growth of Eimeria tenella in an in vitro assay system using BHK-21 cells as a host. No schizont in the cells was observed at concentrations of 0.02-0.2 micrograms/ml for xanthoquinodins A1, A3, B1 and B2 and at 0.02 micrograms/ml for xanthoquinodin A2.
Animals↗
[A tuberculosis epidemic in one kindergarten].
In a routine PPD skin test among 1st grade pupils in 4 elementary schools, Fukushima prefecture in April 1989, the number of pupils that showed strongly positive was unusually high. Moreover, 2 tuberculosis cases were detected among them. Since those children graduated the same kindergarten, and one of 6 kindergarten teachers had been complaining cough and hoarseness for about 3 to 5 months before being diagnosed as a smear positive patient in the contact survey, she was suspected to be the source of infection. Meanwhile, she had been treated for common cold and chronic bronchitis in several clinics, therefore, the main cause of this epidemic was due to the doctor's delay in detecting the case. With the discovery of 2 additional cases in the contact survey of the kindergarten, there were 5 cases altogether including the index cases. Three of 4 children's cases had been vaccinated with BCG. If the children who showed strong reaction to tuberculin and eventually treated prophylactically with isoniazid were assumed the infected, the rate of development of disease among the infected was 13.6 to 27.3%. Usually, lymph node enlargement of the hilum is said not to be observed among vaccinated, though slightly, it was evident among the cases in this epidemic.
Adult↗
Functioning survival rate of the fixture and superstructure of osseointegrated implants (first report).
Osseointegrated implant bridges (OIB) have shown excellent progressing in the majority of cases. Since 1983, OIBs has been used to treat about 110 cases in Tokyo Dental College Hospital with more than 500 fixtures. The survival rate of OIB, especially the functioning survival rate of these fixtures, is described, herein. The survival rate of the superstructure of OIB has been almost 100% in both maxilla and mandible. However, the functioning survival rate of OIB fixtures was 88% in maxillary cases and 96% in mandibular cases at less than 1 year. Very few fixtures had to be removed after more than 1 year in either maxilla or the mandible. It appears that the functioning survival rate of OIB fixture is generally decided during the period just between connecting these components and 1 year. A few fixtures may need to be removed later.
Adolescent↗
Association of functional alteration in intracellular signal transduction systems with the occurrence of up-regulation of muscarinic receptors in primary cultured neurons.
1. Alterations in intracellular biosignaling systems associated with the up-regulated muscarinic receptor were investigated using primary cultured neurons exposured to 10 nM atropine for 5 days. 2. In neurons treated with atropine, the response of PI turnover to muscarinic stimulation decreased in comparison with that in non-treated neurons. 3. The GTP gamma S-stimulated PI turnover also reduced in neurons possessing up-regulated muscarinic receptor, although phospholipase C activity was not different in these two types of neurons. 4. In addition, the long-term exposure (5 days) to atropine induced the increase in GTPase activity and [3H]GppNHp binding. 5. These results suggest that the decreased response of PI turnover to muscarinic stimulation may be attributed to the functional deterioration of G-protein itself and/or coupling between G-protein and phospholipase C. 6. The present results also strongly suggest that the accentuation of the function of G protein may occur in association with the up-regulation of muscarinic receptor.
Animals↗
Cells surviving infection by human immunodeficiency virus type 1: vif or vpu mutants produce non-infectious or markedly less cytopathic viruses.
Under conditions in which a clonal cell line (M10) isolated from a human T cell lymphotrophic virus type I-transformed MT-4 cell line was completely killed by infection with wild-type human immunodeficiency virus type 1 (HIV-1), equivalent M10 cells survived infection with HIV-1 vif, vpr or vpu mutant virus after transient cytopathic effects. Several cell clones, which were isolated from the proliferating M10 cells after infection with vif and vpu mutant viruses (M10/vif- and M10/vpu-), had heterogeneous HIV-1 phenotypes in terms of HIV-1 antigen expression, their syncytium forming capacity, reverse transcriptase activity and the infectivity of HIV-1 particles produced. When the replication kinetics of the HIV-1 particles produced were assayed in M10 cells, the clones could be classified into three types, i.e. type I producing non-infectious HIV-1, type II producing infectious HIV-1 with low replicative ability and type III producing infectious HIV-1 with a replicative ability similar to that of wild-type HIV-1. HIV-1 major viral cell proteins and virus particle fractions were almost typical in types II and III but not in type I. Electron microscopic examination of particles released by I, II and III clones revealed rare defective, predominantly defective and essentially normal virions, respectively. Northern and Southern blot analyses revealed no apparent deletion in the proviral DNA and mRNA prepared from these clones, except in the case of type I and II clones isolated from M10/vpu- which contained large deletions in the mRNAs for gag and gag-pol proteins. Thus, M10 cells surviving infection with HIV-1 vif or vpu mutants are heterogeneous, persistently expressing HIV-1 antigens and producing non-infectious or less cytopathic virus.
Blotting, Northern↗
Human immunodeficiency virus type 1 (HIV-1) superinfection of a cell clone converting it from production of defective to infectious HIV-1 is mediated predominantly by CD4 regions other than the major binding site for HIV-1 glycoproteins.
A cell clone, L-2, which produces non-infectious doughnut-shaped human immunodeficiency virus type 1 (HIV-1) particles, was permissive for HIV-1 superinfection, which resulted in the production of infectious particles. The superinfection showed slow kinetics compared with primary HIV-1 infection of M10 cells, the parent of the L-2 cell clone. Inhibition studies on the superinfection of L-2 cells using several CD4-related reagents showed that the CD4 molecule was an essential component of the receptor for superinfection. Strong inhibitory effects were obtained using CD4 peptides such as CD4(68-130), which includes a portion homologous to the immunoglobulin third complementarity-determining region (CDR3), as well as recombinant soluble CD4. In contrast, a CD4(45-60) peptide, which includes most of the CDR2-related region, was not effective, although the Leu-3a monoclonal antibody (MAb), which recognizes a site near the CDR2-related region, did slightly, but significantly, delay the superinfection kinetics. Comparative flow cytometry of L-2 and M10 cells revealed that the cell surface of L-2 cells despite expressing HIV-1 env protein, reacted slightly with OKT4 or anti-CD4(68-130) MAb, but not with Leu-3a or OKT4A MAb. In contrast, no reaction was detected with any of these anti-CD4 MAbs on the surface of another HIV-1 superinfection-resistant cell clone, MOLT-#8IIIB-14, which expresses HIV-1 env proteins but does not produce infectious HIV-1 particles. These results strongly suggest that expression of the CD4 major receptor site for primary HIV-1 infection is preferentially decreased on the surface of L-2 cells, but that the OKT4 epitope and the nearby region corresponding to immunoglobulin CDR3 remain exposed on the cell surface. Consequently, the CD4 CDR3-related region could play a major role as the receptor for the superinfection reported here.
Binding Sites↗
[Experimental urinary tract infection caused by Escherichia coli in diabetic mouse--uroepithelial colonization and polymorphonuclear leukocyte function].
Relationship between polymorphonuclear leukocyte (PMN) function and susceptibility to infections in diabetic mice was studied. In the experimental urinary tract infections, 6 strains of Escherichia coli with different fimbrial expression and hemolysin production were used. In contrast with non-fimbriated strains, fimbriated strains were more susceptible to phagocytosis by PMN. MS+.MR+ strains, however, showed resistance to intracellular killing by PMN. OEC-709, OEC-716 (MS+.MR+ strains), and OEC-734 (MS+.MR- strains), also showed resistance to the bactericidal activity of the serum. The killing activities of PMN derived from diabetic mice against these 3 strains were significantly lower than those of PMN from normal mice. In normal mice, MS+.MR+ strains showed the strongest colonization ability to the bladder and kidney, whereas MS-.MR- strains showed the weakest ability among the strains. In diabetic mice, the number of bacteria colonizing to the bladder and kidney was higher than that in normal mice, except for one MS-.MR- strain. Significant differences were observed in the strains (OEC-709, 716, 734), in which PMN was concerned strongly in the bacterial killing. Finally, these findings suggest that deterioration of the bactericidal activity of PMN in a diabetic state is involved in increased susceptibility of diabetic mice.
Animals↗
Human immunodeficiency virus type 1 vif, vpr, and vpu mutants can produce persistently infected cells.
A series of human immunodeficiency virus type 1 (HIV-1) mutants in vif, vpr, vpu, and nef were constructed from an infectious plasmid (pNL 432) containing the full-length HIV-1 DNA by frameshift mutations. The capacities for replication and cell killing of these mutant viruses were examined in a clonal cell line (M 10) isolated from HTLV-I-transformed MT-4 cells. In all cases, the mutant viruses replicated, expressed HIV-1 antigens, and induced drastic cytopathic effects. However, some M 10 cells survived infection with vif, vpr, and vpu mutant viruses and became persistently HIV-1-infected, whereas no cells survived infection with the nef mutant as well as the wild-type virus. The HIV-1 particles produced from the surviving cells after infection with the vif, vpr, or vpu mutant viruses were fully replicative in M 10 cells without apparent cytopathic effects.
Cell Death↗
Ontogeny of beta-adrenergic receptor-mediated cyclic AMP generating system in primary cultured neurons.
The developmental changes in the beta-adrenergic receptor/cyclic AMP generating system were examined using mouse cerebral cortical neurons in primary culture. During neuronal growth in vitro, the number of binding sites for [3H]dihydroalprenolol (DHA) showed a tendency to increase (Bmax), while the affinity (Kd) for [3H]DHA did not show any noticeable changes. Basal and isoproterenol-stimulated adenylate cyclase activities as well as the activation of adenylate cyclase by 5'-guanylylimidodiphosphate (GppNHp), NaF and forskolin showed progressive and parallel increases during neuronal growth on a polylysine-coated surface. The treatment of primary cultured neurons with islet-activating protein (IAP), one of the pertussis toxins, attenuated the inhibitory effect of carbachol, a muscarinic agonist, on isoproterenol-induced activation of adenylate cyclase activity. These results indicate that primary cultured neurons possess a cyclic AMP generating system coupled with beta-adrenergic and muscarinic receptors, which is regulated via stimulatory and inhibitory GTP-binding proteins, respectively. The results described above also suggest that the beta-adrenergic receptor, stimulatory and inhibitory types of GTP-binding proteins and adenylate cyclase may develop in a parallel fashion during neuronal growth on a polylysine-coated surface.
Adenylate Cyclase Toxin↗
Inverted repeat regions of Marek's disease virus DNA possess a structure similar to that of the a sequence of herpes simplex virus DNA and contain host cell telomere sequences.
The genomic structure of Marek's disease virus (MDV) is similar to those of the alphaherpesviruses herpes simplex virus (HSV) types 1 and 2. Sequence analysis of the junction region between the long component (L) and the short component (S) revealed the existence of an a-like sequence, similar in structure to the a sequence of HSV-1. Further study revealed that the MDV genome contains five copies of the a-like sequence within the long terminal repeat region as well as in the short terminal repeat region. The junction between the L and S components was found to contain 10 copies of the a-like sequence. Within the a-like sequence, a structure homologous to the DR2 of HSV was found to contain 17 copies of the telomeric sequence, GGGGTTA. There appears to be little to no sequence homology between the HSV a sequence and the MDV a-like sequence; however, the strong physical homology to its counterpart in HSV-1 suggests that the MDV a-like sequence may have the same functional homology (the domain for cleavage/packaging of the DNA into the viral capsids and for genomic inversion) as well.
Animals↗