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At least 199 records · Page 11Linked to original sources
PubMed1993

Non-radioactive PCR method for quantification of HIV-infected peripheral blood mononuclear cells in HIV-positive subjects.

A nested PCR method for quantification of HIV-infected peripheral blood mononuclear cells (PBMC) which does not use radioactivity nor plasmids is described. Quantification was achieved by means of limiting dilutions and an ACH2 cell standard. The method was used to study 23 HIV-infected patients. A significant correlation was seen between the stage of HIV disease, as classified by CDC criteria, and the number of infected PBMC. Although the CD4+ cell count also correlated well with the stage of HIV disease, there was only weak correlation between the result obtained by the PCR method and the CD4+ cell count.

Base Sequence↗
PubMed1983

The complexity of the overlap method for sequencing biopolymers.

The problem of trying to reconstruct the sequence of a biopolymer by using overlapping fragments obtained from cleaving agents is shown to be computationally intractable. This strongly suggests that any computer program for overlap sequencing, even though it may work well for a limited number of inputs, will not work sufficiently for all inputs. However, if the problem is restricted so that certain crucial fragments are known, called prime strings, a sequence can be found efficiently in all cases. Graph theory techniques for doing so can also be used to count the number of sequences consistent with the fragment data to determine whether a unique sequence has been obtained.

Amino Acid Sequence↗
PubMed1990

Partial amino acid sequence of erythrocyte carbonic anhydrase from tiger shark.

1. A partial primary structure (197 residues) of carbonic anhydrase from tiger shark (Galeocerdo cuvieri) erythrocytes has been determined. 2. The amino acid sequence of the enzyme is identical to those of human cytoplasmic carbonic anhydrases (CA I-III) by as much as 52-60%. 3. It is shown that tiger shark CA most closely resembles the CA II isoenzyme of amniotes. 4. Isoelectric focusing and inhibition studies on carbonic anhydrase from dogfish (Squalus acanthias) blood and muscle indicate the presence of the same isoenzyme in shark blood and muscle.

Alkylation↗
PubMed1994

Effect of chronic feline immunodeficiency virus infection on experimental feline calicivirus-induced disease.

Acute feline calicivirus (FCV) infection caused a more severe disease in chronically feline immunodeficiency virus (FIV) infected than in non-FIV infected cats. FIV infected cats shed significantly higher amounts of FCV through their saliva after FCV challenge than the non-FIV infected cats. However, there was no difference in the duration of FCV shedding. None of the cats exposed to FCV developed chronic FCV carriage. Both groups of cats mounted similar titers of neutralizing antibodies to FCV. Although FIV infected cats started out with significantly lower total lymphocyte and neutrophil numbers than the non-FIV infected cats, the transient lymphopenia and neutrophilia attributable to the FCV infection was of similar intensity in both groups of animals. There was no evidence that the underlying FIV-related disease or viremia was influenced by acute FCV infection. Acute FCV infection did not significantly alter the CD4+/CD8+ T lymphocyte ratio in FIV infected compared to non-FIV infected cats. The ongoing humoral IgG response to FIV was not affected by the FCV infection. There was no significant change in the proportion of FIV infected peripheral blood mononuclear cells during 8 subsequent weeks after FCV challenge as determined by polymerase chain reaction.

Animals↗
PubMed2002

Small molecule modulators of HIV Rev/Rev response element interaction identified by random screening.

A high throughput scintillation proximity assay with biotinylated human immunodeficiency virus (HIV) Rev protein and tritiated Rev response element RNA was used to screen over 500,000 small molecules. Several chemical classes of inhibitors and two chemical classes of enhancers of binding were identified, with the molecular weight range being 400-600. The most common structural motif of inhibitor was an acidic moiety at the end of a linear aromatic system. Most of these modulators had EC(50) values in the 1-10 microM potency range, with several below 1 microM. Several classes displayed structure-activity relationships suggesting specific molecular interactions between small molecule and macromolecule. Several molecules were confirmed as inhibitors in a gel shift assay and by surface plasmon resonance analysis. Furthermore, one inhibitor was shown to bind the Rev protein with a binding constant equal to its IC(50) value, consistent with the mechanism of inhibition being binding Rev. Thus, small molecules can modulate this macromolecular protein-RNA interaction in vitro. However, no compound demonstrated HIV antiviral activity in a relevant cell-based assay.

Amino Acid Sequence↗
PubMed2001

Eimeria telekii n.sp. (Apicomplexa: Coccidia) from Lemniscomys striatus (Rodentia: Muridae): morphology, pathology and phylogeny.

Using a combination of morphological, life-cycle and molecular data, we describe a new apicomplexan parasite Eimeria telekii n.sp. from a striped grass mouse Lemniscomys striatus captured in Kenya. Oocysts are oval to spherical or ellipsoidal, 20.4 x 15.7 (15.5-25.0 x 12.0-20.0) microm with a colourless, smooth and bilayered wall. Sporocysts are ellipsoidal, 11.2 x 7.8 (10.0-12.0 x 7.0-9.0) microm with a small Stieda body and granular sporocyst residuum and contain 2 elongated, banana-shaped sporozoites with a single refractile body. Life-cycle, pathogenicity and host specificity of this parasite were studied in laboratory-bred Lemniscomys barbarus and BALB/c mice. Two asexual stages and the sexual phase took place within the enterocytes of the caecum and colon of L. barbarus but not in inoculated BALB/c mice. An infectious dose of 5000 oocysts caused severe clinical illness and mortality in 2/2 (100%) L. barbarus. Phylogenetic analysis of the small subunit rRNA gene of E. telekii and members of the genera Eimeria, Cyclospora and Isospora placed E. telekii within the eimerian rodent clade.

Animals↗
PubMed2004

A transcriptional coactivator, AtGIF1, is involved in regulating leaf growth and morphology in Arabidopsis.

Previously, we described the AtGRF [Arabidopsis thaliana growth-regulating factor (GRF)] gene family, which encodes putative transcription factors that play a regulatory role in growth and development of leaves and cotyledons. We demonstrate here that the C-terminal region of GRF proteins has transactivation activity. In search of partner proteins for GRF1, we identified another gene family, GRF-interacting factor (GIF), which comprises three members. Sequence and molecular analysis showed that GIF1 is a functional homolog of the human SYT transcription coactivator. We found that the N-terminal region of GIF1 protein was involved in the interaction with GRF1. To understand the biological function of GIF1, we isolated a loss-of-function mutant of GIF1 and prepared transgenic plants subject to GIF1-specific RNA interference. Like grf mutants, the gif1 mutant and transgenic plants developed narrower leaves and petals than did wild-type plants, and combinations of gif1 and grf mutations showed a cooperative effect. The narrow leaf phenotype of gif1, as well as that of the grf triple mutant, was caused by a reduction in cell numbers along the leaf-width axis. We propose that GRF1 and GIF1 act as transcription activator and coactivator, respectively, and that they are part of a complex involved in regulating the growth and shape of leaves and petals.

Arabidopsis↗
PubMed1993

Alterations in spliced and unspliced HIV-1-specific RNA detection in peripheral blood mononuclear cells of individuals with varying CD4-positive lymphocyte counts.

Reverse transcriptase-polymerase chain amplification reactions (RT-PCR) were used to identify transcripts for HIV-1 structural and regulatory proteins in peripheral blood mononuclear cells of a cohort of 48 patients. At least one set of PCR primers was capable of detecting HIV-1 transcripts in 94% of patients. Unspliced gag-pol transcripts were detected with gag or pol primer sets in 60 and 63% of samples, respectively. A significant inverse correlation was noted between transcript identification with the gag primer set and the number of CD4-positive lymphocytes in the blood sample and the clinical stage of infection. Single-spliced env transcripts were identified in 44% of individuals. Multiple-spliced tat or nef transcripts were detected in 6.2 and 53% of individuals, respectively. These findings indicate that viral transcripts are expressed throughout the course of HIV-1 infection.

Base Sequence↗
PubMed1997

Structural analysis of Arabidopsis thaliana chromosome 5. I. Sequence features of the 1.6 Mb regions covered by twenty physically assigned P1 clones.

A total of 20 P1 clones with an average insert size of 80 kb and each containing a marker(s) specifically mapped on chromosome 5 were isolated from a P1 library of the Arabidopsis thaliana genome, and their nucleotide sequences were determined according to a shotgun-based strategy and precisely located on the physical map of chromosome 5 separately constructed. The total length of the sequenced regions were summed up to 1,621,245 bp. By comparison with the sequences in protein and EST databases and analysis with computer programs for gene modeling, a total of 347 potential protein-coding genes and/or gene segments with known or predicted functions were identified. The positions of exons which do not exhibit any similarity to known genes were also predicted. An average density of the genes and/or gene segments assigned so far as 1 gene/4,672 bp. Introns were identified in approximately 78% of the potential genes, and the average number and length of the introns per gene were 3.7 and 161 bp. The transcription level of the predicted genes was roughly monitored by counting the numbers of identified Arabidopsis ESTs. The sequence data and gene information are available through the World Wide Web at http:/(/)www.kazusa.or.jp/arabi/.

Arabidopsis↗
PubMed1997

Structural analysis of Arabidopsis thaliana chromosome 5. II. Sequence features of the regions of 1,044,062 bp covered by thirteen physically assigned P1 clones.

A total of 13 P1 clones, each containing a marker(s) specifically mapped on chromosome 5, were isolated from a P1 library of the Arabidopsis thaliana Columbia genome, and their nucleotide sequences were determined according to the shot gun based strategy and precisely located on the physical map of chromosome 5. The total length of the sequenced regions was 1,044,062 bp. Since we have previously reported the sequence of 1,621,245 bp by analysis of 20 non-redundant P1 clones, the total length of the sequences of chromosome 5 determined so far reached 2,665,307 bp. The regions sequenced in this study were analysed by comparison with the sequences in protein and EST databases and analysis with computer programs for gene modeling; a total of 225 potential protein-coding genes and/or gene segments with known or predicted functions were identified. The positions of exons which do not exhibit similarity to known genes were also predicted by computer-aided analysis. An average density of the genes and/or gene was 1 gene/4,640 bp. Introns were identified in approximately 84% of the potential genes, and the average number and length of the introns per gene were 5.3 and 184 bp, respectively. These sequence features are essentially identical to those for the previously sequenced regions. The transcription level of the predicted genes has been roughly monitored by counting the numbers of matched Arabidopsis ESTs. The sequence data and gene information are available through the World Wide Web at http:@www.kazusa.or.jp/arabi/.

Arabidopsis↗
PubMed2000

Deletion and expression analysis of AZFa genes on the human Y chromosome revealed a major role for DBY in male infertility.

Three distinct regions, designated AZFa, b and c from proximal to distal Yq, are required for normal spermato-genesis in humans. Deletions involving AZFa (deletion interval 5C/D) seem to occur less frequently in infertile men and to be associated with a more severe testicular phenotype, with almost complete absence of germ cells. AZFa contains three genes, named USP9Y, DBY and UTY, and presents high homology with the mouse Delta Sxr (b) interval, deletion of which causes a severe spermatogenic impairment. However, the specific role of these genes in human spermatogenesis is still unknown and it is not clear which of them is responsible for the AZFa phenotype. Here we describe a complete sequence map of the AZFa region, the genomic structure of AZFa genes and their deletion analysis in a large number of infertile men characterized by well-defined spermatogenic alterations. Both USP9Y and DBY may cause severe testiculopathies, but DBY appears to be the major AZFa candidate. DBY is frequently deleted in infertile patients and its absence produces severe spermatogenic damage leading to a significant reduction of germ cells or even to their complete absence. Expression analysis of AZFa genes and their X-homologues revealed ubiquitous expression for all of them except DBY; this gene produces a long transcript which is ubiquitously expressed in addition to a shorter transcript which is only expressed in the testis, suggesting a specific role for DBY in the spermatogenic process. This hypothesis is further supported by the high similarity of DBY to other DEAD box proteins belonging to the PL10 subclass.

Animals↗
PubMed1993

Variable relationship between proviral DNA load and infectious virus titre in the peripheral blood mononuclear cells of HIV-1-infected individuals.

OBJECTIVES: To determine the relationship between infectious virus titre and proviral copy number in peripheral blood mononuclear cells (PBMC) of infected subjects and to ascertain which, if either, is most closely related to CD4+ cell loss and disease progression. DESIGN AND METHODS: Cellular HIV-1 viraemia was quantified in 45 infected subjects who had not received antiretroviral therapy using limiting dilution tissue culture infective dose (PBMC TCID) and quantitative polymerase chain reaction (PCR) techniques. RESULTS: Proviral DNA was detected in 44 (98%) and infectious virus in 38 (82%) of the 45 subjects. Viraemia as measured by both culture and PCR was inversely correlated with patient CD4+ cell count and associated with disease status. Measurement using both techniques correlated with each other (Spearman's rank correlation rho = 0.52; P = 0.0006). The ratio of proviral copies to PBMC TCID ranged from 1:1 1000:1. to > 1000:1. CONCLUSIONS: The ratio of provirus:PBMC TCID was highest when the PBMC TCID was low, and approached unity when PBMC TCID was high. This ratio could be influenced by a variety of factors but did not correlate significantly with patient disease status or CD4+ cell count.

Base Sequence↗
PubMed2003

123I-Interleukin-2: biochemical characterization and in vivo use for imaging autoimmune diseases.

We describe in detail the labelling of interleukin-2 with I ( I-IL2), its biochemical characterization, the binding assay and its use for the detection of tissues infiltrated with mononuclear cells. Human recombinant IL2 was labelled using an enzymatic method and its biochemical characterization was performed using high performance liquid chromatography (HPLC) analysis of cyanogen bromide-cleaved protein. biological and binding assays were performed on CTLL-2 cell line and on activated peripheral blood lymphocytes. studies were performed 1 h after administration of 2-3 mCi of I-IL2 in 10 newly diagnosed type 1 diabetes patients, five pre-diabetic patients, 10 Hashimoto's thyroiditis patients, 10 coeliac disease patients and 10 normal volunteers. I-IL2 scintigraphy allowed the detection and quantification of activated mononuclear cells in several affected tissues. In detail, I-IL2 accumulation was detected in the thyroid of all patients affected by Hashimoto's thyroiditis, in the bowel of all coeliac disease patients and in the pancreas of all pre-type 1 diabetic patients. By contrast, in newly diagnosed type 1 diabetics, I-IL2 scan was positive in five of the 10 studied patients. I-IL2 scintigraphy may be useful for studying autoimmune phenomena and in diagnostic protocols to evaluate the presence of other tissue involvement in patients with an organ-specific autoimmune disease.

Adolescent↗
PubMed1994

Marked basophilia in acute promyelocytic leukaemia treated with all-trans retinoic acid: molecular analysis of the cell origin of the basophils.

We report a patient with acute promyelocytic leukaemia who developed marked basophilia during all-trans retinoic acid treatment. We studied genomic DNA and RNA extracted from the patient's peripheral leucocytes in order to determine the origin of the basophils. The RAR alpha rearranged band in the Southern blot analysis and a chimaeric product of PML-RAR alpha by polymerase chain reaction were strongly visible before ATRA treatment, but at the time of maximal basophilia both of them were markedly diminished. These findings suggest that the basophils which appeared during the ATRA treatment are reactive in nature rather than a leukaemic clone.

Base Sequence↗
PubMed2006

Globin gene expression in Hb Lepore-BAC transgenic mice.

We generated five lines of transgenic mice carrying 1-3 copies of the Hb Lepore deltabeta fusion gene, in the context of a Bacterial Artificial Chromosome containing the whole human beta globin gene cluster. Normal developmental regulation of human genes occurred at levels approximating to those of endogenous genes. Deltabeta transgene expression became detectable during fetal life and rose to a mean level of 13.0% in adults, similar to that of humans. Low levels of human gamma chains were detectable as F cells in adult mice, but numbers did not increase after treatment with drugs that raise F cells in human subjects, even on a thalassaemic background.

Animals↗
PubMed2003

Chemoattractant receptor-homologous molecule expressed on Th2 cells activation in vivo increases blood leukocyte counts and its blockade abrogates 13,14-dihydro-15-keto-prostaglandin D2-induced eosinophilia in rats.

We cloned, expressed, and characterized in vitro and in vivo the gene encoding the rat ortholog of chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2), a G protein-coupled receptor for prostaglandin D2 (PGD2). Quantitative reverse transcription-polymerase chain reaction analysis demonstrated highest CRTH2 expression in the lung, brain, ovary, and spleen. Pharmacologically, rat CRTH2 stably transfected in mouse preB lymphoma L1.2 cells behaved very similar compared with the mouse and human orthologs, showing a binding affinity for PGD2 of 11 nM, a functional calcium mobilization when exposed to agonist, and similar sensitivity to agonists and antagonists. In vivo, selective activation of CRTH2 by 13,14-dihydro-15-keto (DK)-PGD2 injection into rats led to a dose- and time-dependent increase of the number of leukocytes in the peripheral blood. Specifically, eosinophils, lymphocytes, and neutrophils were recruited with maximum effects seen 60 min after the injection of 300 microg of DK-PGD2 per rat. Pretreatment of the animals with the CRTH2/thromboxane A2 receptor antagonist, ramatroban, completely abrogated DK-PGD2-induced eosinophilia, suggesting that CRTH2 might have a physiological and/or pathophysiological role in controlling leukocyte migration.

Amino Acid Sequence↗
PubMed1994

Homozygous human TAP peptide transporter mutation in HLA class I deficiency.

Human lymphocyte antigen (HLA) class I proteins of the major histocompatibility complex are largely dependent for expression on small peptides supplied to them by transporter associated with antigen processing (TAP) protein. An inherited human deficiency in the TAP transporter was identified in two siblings suffering from recurrent respiratory bacterial infections. The expression on the cell surface of class I proteins was very low, whereas that of CD1a was normal, and the cytotoxicity of natural killer cells was affected. In addition, CD8+ alpha beta T cells were present in low but significant numbers and were cytotoxic in the most severely affected sibling, who also showed an increase in CD4+CD8+ T cells and gamma delta T cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2005

Enhanced survival of Salmonella enterica in vesicles released by a soilborne Tetrahymena species.

Nondestructive ingestion by soilborne protozoa may enhance the environmental resiliency of important bacterial pathogens and may model how such bacteria evade destruction in human macrophages. Here, the interaction of Salmonella enterica serovar Thompson with a soilborne Tetrahymena sp. isolate was examined using serovar Thompson cells labeled with the green fluorescent protein. The bacteria were mixed in solution with cells of Tetrahymena at several ratios. During incubation with serovar Thompson, Tetrahymena cells released a large number of vesicles containing green fluorescent serovar Thompson cells. In comparison, grazing on Listeria monocytogenes cells resulted in their digestion and thus the infrequent release of this pathogen in vesicles. The number of serovar Thompson cells per vesicle increased significantly as the initial ratio of serovar Thompson to Tetrahymena cells increased from 500:1 to 5,000:1. The density of serovar Thompson was as high as 50 cells per vesicle. Staining with propidium iodide revealed that a significantly higher proportion of serovar Thompson cells remained viable when enclosed in vesicles than when free in solution. Enhanced survival rates were observed in vesicles that were secreted by both starved (F = 28.3, P < 0.001) and unstarved (F = 14.09, P < 0.005) Tetrahymena cells. Sequestration in vesicles also provided greater protection from low concentrations of calcium hypochlorite. Thus, the release of this human pathogen from Tetrahymena cells in high-density clusters enclosed in a membrane may have important implications for public health.

Animals↗