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Biomedical subjects

L Tao

Publications and source records attributed to L Tao.

At least 127 records · Page 7Linked to original sources
PubMed1996

Protective mucosal immunity elicited by targeted iliac lymph node immunization with a subunit SIV envelope and core vaccine in macaques.

Prevention of sexually transmitted HIV infection was investigated in macaques by immunization with a recombinant SIV (simian immunodeficiency virus) envelope gp 120 and core p27 vaccine. In two independent series of experiments, we used the novel targeted iliac lymph node (TILN) route of immunization, aiming close to the iliac lymph nodes draining the genitorectal mucosa. Rectal challenge with the SIVmac 32H J5 molecular clone in two series induced total protection in four out of seven macaques immunized by TILN, compared with infection in 13 of 14 unimmunized macaques or immunized by other routes (P = 0.025). The remaining three macaques showed either a decrease in viral load ( > 90%) or transient viremia, indicating that all seven TILN-immunized macaques showed total or partial protection (P = 0.001). Protection was associated with significant increase in the iliac lymph nodes of IgA antibody-secreting cells to p27 (P < 0.02), CD8-suppressor factor (P < 0.01), and the chemokines RANTES and MIP-1 beta (P < 0.01).

Animals↗
PubMed1996

Purification and characterization of human lymphoblast N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase.

The enzyme N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase (EC 3.1.4.45; uncovering enzyme) catalyzed the removal of N-acetylglucosamine from the N-acetylglucosamine-alpha-phospho-mannose portion of selected lysosomal enzyme oligosaccharide chains, thereby forming the mannose 6-phosphate signal which is responsible for the targeting of these lysosomal enzymes for transport into lysosomes. The uncovering enzyme has been purified approximately 7000-fold to electrophoretic homogeneity from Epstein-Barr virus-transformed human lymphoblast cells. The purification sequence involves solubilizing this membrane-bound enzyme with Tergitol NP-10, affinity chromatography on Lentil lectin-Sepharose 4B, ion-exchange chromatography on DEAE-Sephacel, chromatography on zinc(II)-IDA-Sepharose 6B, and preparative SDS-PAGE electrophoresis. The purified enzyme migrated as a single band of 114 kDa which was coincident with enzyme activity on analytical SDS-PAGE electrophoresis. Characterization studies of the purified enzyme demonstrated that catalytic activity was maximal at pH 6.95 and that the enzyme retained full activity following incubation for 10 min at 60 degrees C. No requirement was found for a divalent cation, but Zn2+, Hg2+, and Cu2+ were found to reduce the enzyme's activity by 30-40%. The highest catalytic efficiency was observed with N-acetylglucosamine-phospho-methylmannoside as a substrate while uridine diphosphate-N-acetylglucosamine, N-acetylglucosamine-phosphomannose-uteroferrin, and N-acetylglucosamine-phosphate were also cleaved by the enzyme with decreasing efficiency. Acetamino-deoxycastanospermine was a potent inhibitor of the human enzyme with a Ki of 0.35 microM, while N-acetylglucosamine phosphate (Ki 1.58 mM) and N-acetylglucosamine (Ki 5.1 mM) inhibited the enzyme to a lesser degree.

Chromatography, Affinity↗
PubMed1996

Analysis of Lactobacillus phages and bacteriocins in American dairy products and characterization of a phage isolated from yogurt.

Yogurt and acidophilus milk that contain Lactobacillus acidophilus could promote human health because L. acidophilus can inhibit enteric and food-borne microbial pathogens. To evaluate the stability of diary L. acidophilus cultures, we studied whether some diary lactobacilli could be inhibited by phages or bacteriocins released by other dairy lactobacilli. From 20 yogurts and two acidophilus milks purchased at local food markets, 38 Lactobacillus strains were isolated. Eight Lactobacillus type strains were used as controls. With mitomycin induction and agar spot assay, phages and bacteriocins were isolated from these strains and their activities were analyzed. Lactobacillus strains from 11 yogurts released phages, while the strains from most of the remaining products released bacteriocins. One phage, designated phi y8, was characterized. It was spontaneously released from its host strain L. acidophilus Y8, at a rate of about 10(4)/ml. This phage lysed nine other dairy Lactobacillus strains tested. It had a burst size of 100, an elongated prolate head of 39 by 130 nm, a long, flexible but noncontractile tail of 300 nm, and a 54.3-kb linear double-stranded DNA. DNA fingerprinting analysis indicated that L. acidophilus phages of nine yogurts in this study belonged to the same type as phi y8. Although they may be sensitive to bacteriocins, all lysogens resisted further phage attacks, whereas most nonlysogens were sensitive to both phages and bacteriocins. Therefore, Lacotbacillus cultures of some American yogurts and acidophilus milks may be unstable or unsafe because they can either be inhibited by phages or bacteriocins or release them to inhibit lactobacilli or other diary products.

Animals↗
PubMed1996

Three-dimensional organization of rat hepatocyte cytoskeleton: relation to the asialoglycoprotein endocytosis pathway.

Analysis by confocal microscopy has revealed features of the microtubule network of rat hepatocytes in culture, establishing the three-dimensional disposition of the microtubule-based cytoskeleton, its relation to the actin-based cytoskeleton and to ligand-containing endosomes during receptor-mediated endocytosis and the alterations in its structure and disposition by the microtubule pertubant, Taxol. By co-localization studies, we have been able to demonstrate that the microtubules have a significant role in receptor-mediated endocytosis of asialoglycoproteins in this cell. Asialoorosomucoid-containing endosomes attach to widely spaced arrays of microtubules running under the baso-lateral surface of the hepatocytes 5-15 minutes after the initiation of endocytosis and then travel along microtubule paths to become concentrated with microtubules near the centrosome and at bile canaliculi after 30-60 minutes of receptor-mediated endocytosis. Receptor-mediated endocytosis is affected, but not abolished by Taxol, which inhibits the rate of asialoorosomucoid degradation at the same concentrations as those that disrupt microtubule and cytoplasmic dynein distribution, and that prevent the concentration of endosomes centrally. The results support suggestions that asialoorosomucoid-containing endosomes are captured by microtubules just below the actin layer at the cell periphery and these are actively transported centrally along microtubules, possibly by cytoplasmic dynein, so that the concentration of endosomes near the centrosome, and the subsequent efficient lysosomal degradation of ligand, are consequences of the confluence of microtubules in this region.

Actin Cytoskeleton↗
PubMed1996

The penetration of chromium-EDTA from blood plasma into various compartments of rat testes as an indicator of function of the blood-testis barrier after exposure of the testes to heat.

The concentration of chromium51-EDTA in blood plasma after an intravenous infusion was found to be about 40 times that present in rete testis fluid and 20 times that in the additional seminiferous tubular fluid resulting from ligation of the efferent ducts. These values indicate the effectiveness of the blood-testis barrier to small water-soluble molecules, like Cr-EDTA. The volume of distribution in microlitres of Cr-EDTA in the parenchyma was about 60% of the volume of the interstitial tissue as determined on frozen sections by morphometry, and was similar, or slightly less, in the ligated testes, compared with the unligated testes. Heating the testes to 43 degrees C for 30 min led to the expected reduction several days later in testis mass, but the volume of distribution of Cr-EDTA was no greater than that in the testes of control rats, and the ratio of Cr-EDTA space to interstitial tissue was not different, while the concentration of Cr-EDTA in the additional seminiferous tubular fluid increased only slightly as testis mass fell. These results indicate that the blood-testis barrier was only slightly less effective, if changed at all, during the period of spermatogenic disruption following local heating of the testis.

Animals↗
PubMed1996

[Effects of enalapril and taurine on left ventricular hypertrophy and arrhythmia in renovascular hypertensive rat].

The effects of enalpril (Ena, 6 mg.kg-1) and taurine (Tau, 30 mg.kg-1) on left ventricular hypertrophy (LVH) and ventricular arrhythmia were studied in two-kidney, one clip renovascular hypertensive rats (RHR). From the 9th week after operation, Ena and Tau were given per oral daily for 9 weeks. These drugs significantly decreased the systolic arterial pressure and the weight of the left ventricle. Combination of both drugs was found to reduce the blood pressure further than either drug used alone. Arrhythmias induced by trains of electrical stimuli were more frequent in working hearts isolated from RHR than that from normotensive rats. Ena and Tau could decrease the incidence of this arrhythmias in RHR. The calcium content in the myocardial mitochondria in RHR was increased compared with that in normotensive rats. Treatment with Ena and Tau reduced this increase significantly. These results suggest that chronic therapy with Ena and Tau can induce an attenuation of systemic arterial pressure and reduce the propensity of RHR heart to arrhythmogenesis by limiting cardiac hypertrophy and calcium overload of the myocardium.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1995

Construction of a Streptococcus pyogenes recA mutant via insertional inactivation, and cloning and sequencing of the complete recA gene.

To facilitate future genetic studies with Streptococcus pyogenes (Sp), a recA mutant (Rec11) was constructed using a streptococcal integration vector carrying a PCR-derived internal recA fragment. The insertion of the plasmid in the mutant chromosome was identified by Southern hybridization. Resistance to UV and the ability to accept linear DNA transformation by Rec11 were greatly decreased, confirming its RecA phenotype. Using the PCR-derived fragment as a probe, we cloned and sequenced the complete Sp recA gene, which is highly homologous to the recA of S. pneumoniae and Lactococcus lactis.

Amino Acid Sequence↗
PubMed1995

Human hepatoma cell mutant defective in cell surface protein trafficking.

To isolate a mutant liver cell defective in the endocytic pathway, a selection strategy using toxic ligands for two distinct membrane receptors was devised. Ovalbumin-gelonin and asialoorosomucoid (ASOR)-gelonin were incubated with mutagenized HuH-7 cells, and a rare survivor termed trafficking mutant 1 (Trf1) was isolated. Trf1 cells were stably 3-fold more resistant than the parental HuH-7 to both toxic conjugates. The anterograde steps of intracellular endocytic processing of ASOR, including internalization, endosomal acidification, and ligand degradation, were unaltered in Trf1 cells. In contrast, retrograde diacytosis of asialoglycoprotein receptor (ASGR).ASOR complex back to the cell surface was enhanced by about 250%. Selective labeling revealed an approximately 46% reduction in cell surface-associated ASGR in Trf1 cells, although their total cellular ASGR content was essentially equivalent to that in HuH-7. Similar results were obtained with the transferrin receptor. Binding of 125I-ASOR and 125I-transferrin was reduced in Trf1 cells to 49 +/- 2.5% and 30 +/- 2%, respectively, of HuH-7 cells. The methionine transporter was also reduced in Trf1 cells, as revealed by a 2-fold reduction in Vmax with no change in apparent Km. Pretreatment with monensin, sodium azide, or colchicine reduced surface binding of 125I-ASOR in HuH-7 cells by 50% but had no effect on binding to Trf1 cells. This result is predicted for a cell that expresses only State 1 ASGRs, which are resistant to modulation by metabolic and cytoskeletal inhibitors in contrast to State 2, which are responsive to these agents (Weigel, P. H., and Oka, J. A. (1984) J. Biol. Chem. 259, 1150-1154). The Trf1 mutant, having lost the ability to express State 2 receptors, provides genetic evidence for the existence of these two receptor subpopulations and an approach to identifying the biochemical mechanism by which they are generated.

Asialoglycoprotein Receptor↗
PubMed1995

Genital-associated lymphoid tissue in female non-human primates.

We investigated genital-associated lymphoid tissue (GENALT) in non-human primates (macaques), by augmenting vaginal with oral immunization. The vaccine was a recombinant particulate SIV antigen (p27:Ty-VLP), linked to CT-B, and administered into the vagina by a paediatric naso-gastric tube and into the stomach by a gastric tube. Oro-vaginal or vagino-oral sequence of immunization elicited specific CD4+ T cell proliferative responses to p27 antigen in the genital lymph nodes and the spleen but not in unrelated lymph nodes. CD4+ T cells reconstituted with B cells and macrophages from the genital lymph nodes induced specific IgA and to a lesser extent IgG anti-p27 antibodies. However, the corresponding splenic cells induced greater IgG than IgA antibody synthesis. Intramuscular immunization primed splenic but not genital lymph node cells, and induced CD4+ T cell proliferative responses and predominantly B cell IgG antibody synthesis. Finding primed B and T cells in the genital lymph nodes after augmenting vaginal by oral immunization provides experimental evidence for GENALT in non-human primates. This primate model of vaginal immunization suggests 3 levels of specific immunity: (1) secretory IgA (and IgG) in the cervico-vaginal mucosal epithelium; (2) primed CD4+ T cells and B cells in the genital lymph nodes and the spleen; and (3) circulating CD4+ T cells, B cells and IgG and IgA antibodies specific to the immunizing antigen.

Administration, Intravaginal↗
PubMed1995

Alternative splicing in human retinal mRNA transcripts of an opsin-related protein.

An opsin-related gene encodes a putative RPE-retinal G-protein-coupled receptor (RGR) that is most homologous to the visual pigments and invertebrate retinochrome. A splice variant of human RGR mRNA can be demonstrated by the sequence of isolated cDNA clones and by the amplification and analysis of human retinal mRNA. The shortened transcript contains a deletion of 114 nucleotides that correspond exactly to the sequence of exon 6 in the human rgr gene. The predicted RGR variant lacks the putative sixth transmembrane domain and has a calculated molecular weight of 27,726. Variable amounts of a 28-kDa protein were found in the retinas of some individuals by immunoblot assay. Since a similar shortened RGR transcript was not detected in bovine retina or RPE, the RGR variant is not essential for vertebrate vision. Analysis of the structure of the rgr gene and of the sequences of cDNA clones indicates that the truncated mRNA may be produced through alternative splicing of pre-mRNA from which a cassette exon is removed and the predicted RGR variant is radically altered in primary structure.

Base Sequence↗
PubMed1995

Generation of diversity in the hierarchy of T-cell epitope responses following different routes of immunization with simian immunodeficiency virus protein.

OBJECTIVES: To examine whether the route of immunization determines the hierarchy of T-cell epitope proliferative responses in macaques. DESIGN: Macaques were immunized with a recombinant simian immunodeficiency virus (SIV) p27 core protein by the intramuscular, male and female genital or rectal route, each of which was augmented by oral immunization, and by the novel targeted lymph-node immunization route. Overlapping peptides were used to identify the proliferative T-cell epitopes and to determine their hierarchy in the circulation, spleen and lymph nodes. METHODS: T-cell epitope mapping of the proliferative responses was studied in short-term cell lines. Dendritic cells and macrophages were enriched by metrizamide gradient and adherence to plastic, respectively. RESULTS: Intramuscular immunization elicited in the circulating T cells a hierarchy of T-cell epitopes within four peptides in the following descending order of frequency: peptides 121-140 (57.9%), 41-60 (28.9%), 61-80 (18.9%) and 101-120 (5.4%). The hierarchy of these four T-cell epitope responses differed significantly with each of the five routes of immunization, when circulating (P < 0.001), splenic (P < 0.02-< 0.001) or iliac lymph-node cells (P < 0.001) were analysed. The effect of antigen-presenting cells was then investigated and enriched dendritic cells were more effective than macrophages in processing and presenting the p27 antigen and the immunodominant (121-140) and 61-80 T-cell epitopes. CONCLUSIONS: The route of immunization may determine the hierarchy of T-cell epitopes in the lymph nodes draining the mucosa in the circulating and splenic lymphocytes. The diversity of T-cell epitopes may affect the control of HIV at different anatomical sites, the administration route of the vaccine, and selection of polypeptides or recombinant antigens for immunization.

Animals↗
PubMed1995

Insertional inactivation of the Streptococcus mutans dexA (dextranase) gene results in altered adherence and dextran catabolism.

Streptococcus mutans is able to synthesize extracellular glucans from sucrose which contribute to adherence of these bacteria. Extracellular dextranase can partially degrade the glucans, and may therefore affect virulence of S. mutans. In order to isolate mutants unable to produce dextranase, a DNA library was constructed by inserting random Sau3AI-digested fragments of chromosomal DNA from S. mutans into the BamHI site of the streptococcal integration vector pVA891, which is able to replicate in Escherichia coli but does not possess a streptococcal origin of replication. The resultant plasmids were introduced into S. mutans LT11, allowing insertional inactivation through homologous recombination. Two transformants were identified which did not possess dextranase activity. Integration of a single copy of the plasmid into the chromosome of these transformants was confirmed by Southern hybridization analysis. Chromosomal DNA fragments flanking the plasmid were recovered using a marker rescue technique, and sequenced. Comparison with known sequences using the BLASTX program showed 56% homology at the amino acid level between the sequenced gene fragment and dextranase from Streptococcus sobrinus, strongly suggesting that the S. mutans dextranase gene (dexA) had been inactivated. The colony morphology of the dextranase mutants when grown on Todd-Hewitt agar containing sucrose was altered compared to the parent strain, with an apparent build-up of extracellular polymer. The mutants were also more adherent to a smooth surface than LT11 but there was no apparent difference in sucrose-dependent cell-cell aggregation.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

The three-dimensional point spread functions of a microscope objective in image and object space.

The three-dimensional point spread function (3-D PSF) of an optical system in image space is distinguished from the 3-D PSF in object space and the relation between the two 3-D PSFs is derived. By using this relation one 3-D PSF can be easily obtained from the other. The 3-D PSFs are given in a single integral expression, which can be computed numerically. The results of this study can be used in 3-D image processing for microscopy and have been applied to the analysis of the diffusion of fluorescent molecules in a 3-D porous medium.

Algorithms↗
PubMed1995

Extracellular potassium, volume fraction, and tortuosity in rat hippocampal CA1, CA3, and cortical slices during ischemia.

1. An in vitro slice model of ischemia was used to study changes in extracellular potassium concentration and diffusion properties in the stratum pyramidale of CA1 and CA3 regions of the hippocampus and in the cortex of the rat. Slices were submerged in artificial cerebrospinal fluid, and ischemia was induced by removing oxygen and glucose until anoxic depolarization occurred. 2. Extracellular potassium concentration was measured with a valinomycin-based ion-selective microelectrode. The bathing medium contained 5 mM potassium, and in vitro ischemia caused the potassium concentration to rise to 45 mM in CA1, 12 mM in CA3, and 32 mM in cortex. 3. Extracellular volume fraction and tortuosity were determined during normoxic conditions and in vitro ischemia by measuring the diffusion of tetramethylammonium. This cation was iontophoretically released into the extracellular space and its concentration as a function of time determined with an ion-selective microelectrode approximately 100 microns away from the source. 4. During normoxia the volume fraction was 0.14, 0.20, and 0.18, and tortuosity was 1.50, 1.57, and 1.62 in CA1, CA3, and cortex, respectively. These data confirm that the volume fraction of CA1 is smaller than in the two other regions. 5. During ischemia the volume fraction decreased to 0.05, 0.17, and 0.09 in CA1, CA3, and cortex, respectively. Only in CA3 did the tortuosity change significantly by increasing to 1.75. Because of limitations in the time resolution of the diffusion method, the changes in volume fraction and tortuosity during the anoxic depolarization phase of ischemia may have been underestimated.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Induction of IgA and IgG antibodies in vaginal fluid, serum and saliva following immunization of genital and gut associated lymphoid tissue.

Vaginal immunization with the Simian immunodeficiency virus (SIV) was investigated in macaques in order to study genital mucosal antibodies. A combined route of genital- and gut-associated lymphoid tissues was used to stimulate IgA and IgG antibodies in vaginal fluid, serum and saliva. Macaques were immunized with a recombinant, particulate SIV antigen (SIV gag P27), covalently linked to the mucosal adjuvant cholera toxin B subunit (CTB). The animals were immunized sequentially as follows: vaginal (x2) followed by oral (x3), or the reverse sequence of immunization. The results show that both vaginal followed by oral immunization or the reverse sequence induces specific p27 IgA and IgG antibodies in the vaginal fluid and serum. IgA antibodies were also detected in saliva. Vaginal IgA antibodies have the secretory component and J chain suggesting that they are of secretory origin.

Adjuvants, Immunologic↗