PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Isogenic control”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 829 records · Page 46Linked to original sources
PubMed1987

Transferability of immune plasma and pronephric cells in isogeneic, allogeneic and xenogeneic transfer systems in crucian carp.

Transfer of immunity to horse erythrocytes (HRBC) by intravascular injection of immune plasma and pronephric cells was performed using histocompatible and histoincompatible crucian carp to examine "MHC" regulation in adoptive immunity. Clonal ginbuna crucian carp, Carassius gibelio langsdorfii and outbred kinbuna crucian carp, C. carassius buergeri immunized with HRBC were used as donors of plasma and cells for transfer to unimmunized members. Recipients were then evaluated by measuring the antibody titre in the plasma. Transfer of plasma was highly successful in all patterns of transfer systems such as isogeneic, allogeneic and xenogeneic transfer. Adoptive transfer of antibody productivity by pronephric cells was successful in isogeneic and weak H gene-disparate transfer systems, while antibody productivity was not transferable in xenogeneic and strong H gene-disparate transfer systems. In allogeneic fish, however, antibody productivity was transferable by the transfer of cells in some recipients which rejected allograft in an acute fashion. These results suggest that transferability of immune cells is strictly controlled by "MHC" and that the locus of "class I" antigens involved in cell collaboration or cytotoxic lymphocyte effector function is limited in polymorphism in fish.

Animals↗
PubMed2005

Fluoroquinolone treatment of experimental Salmonella enterica serovar Typhimurium DT104 infections in chickens selects for both gyrA mutations and changes in efflux pump gene expression.

OBJECTIVES: To determine the efficacy of enrofloxacin (Baytril) in chickens in eradicating three different resistance phenotypes of Salmonella enterica and to examine the resistance mechanisms of resulting mutants. METHODS: In two separate replicate experiments (I and II), three strains of Salmonella enterica serovar Typhimurium DT104 [strain A, fully antibiotic-sensitive strain; strain B, isogenic multiple antibiotic-resistant (MAR) derivative of A; strain C, veterinary penta-resistant phenotype strain containing GyrA Phe-83], were inoculated into day-old chicks at approximately 10(3) cfu/bird. At day 10, groups of chicks (n =10) were given either enrofloxacin at 50 ppm in their drinking water for 5 days or water alone (control). Caecal contents were monitored for presence of Salmonella and colonies were replica plated to media containing antibiotics or overlaid with cyclohexane to determine the proportion of isolates with reduced susceptibility. The MICs of antibiotics and cyclohexane tolerance were determined for selected isolates from the chicks. Mutations in topoisomerase genes were examined by DHPLC and expression of marA, soxS, acrB, acrD and acrF by RT-PCR. RESULTS: In experiment I, but not II, enrofloxacin significantly reduced the numbers of strain A compared with the untreated control group. In experiment II, but not I, enrofloxacin significantly reduced the numbers of strain B. Shedding of strain C was unaffected by enrofloxacin treatment. Birds infected with strains A and B gave rise to isolates with decreased fluoroquinolone susceptibility. Isolates derived from strain A or B requiring >128 mg/L nalidixic acid for inhibition contained GyrA Asn-82 or Phe-83. Isolates inhibited by 16 mg/L nalidixic acid were also less susceptible to antibiotics of other chemical classes and became cyclohexane-tolerant (e.g. MAR). CONCLUSIONS: These studies demonstrate that recommended enrofloxacin treatment of chicks rapidly selects for strains with reduced fluoroquinolone susceptibility from fully sensitive and MAR strains. It can also select for MAR isolates.

Animals↗
PubMed1993

Formation of linear plasmid multimers promoted by the phage lambda Red-system in lon mutants of Escherichia coli.

We report here the formation of plasmid linear multimers promoted by the Red-system of phage lambda using a multicopy plasmid comprised of lambda red alpha and red beta genes, under the control of the lambda cI857 repressor. Our observations have revealed that the multimerization of plasmid DNA is dependent on the red beta and recA genes, suggesting a concerted role for these functions in the formation of plasmid multimers. The formation of multimers occurred in a recBCD+ sbcB+ xthA+ lon genetic background at a higher frequency than in the isogenic lon+ host cells. The multimers comprised tandem repeats of monomer plasmid DNA. Treatment of purified plasmid DNA with exonuclease III revealed the presence of free double-chain ends in the molecules. Determination of the size of multimeric DNA, by pulse field gel electrophoresis, revealed that the bulk of the DNA was in the range 50-240 kb, representing approximately 5-24 unit lengths of monomeric plasmid DNA. We provide a conceptual framework for Red-system-promoted formation and enhanced accumulation of plasmid linear multimers in lon mutants of E. coli.

ATP-Dependent Proteases↗
PubMed1993

Expression of gene 19 of the conjugative plasmid R1 is controlled by RNase III.

Specific cleavage of mRNAs by RNase III has been shown to control the expression of several Escherichia coli genes. We show here that the expression of gene 19 of the conjugative resistance plasmid R1 is controlled in its expression by the same endoribonuclease. In vivo studies revealed that a DNA fragment of 150 nucleotides including a perfect 22 nucleotide inverted repeat in the gene 19 coding region is responsible for the low expression of the gene both at the protein and the RNA levels. By using a translational gene 19-lacZ fusion in isogenic RNase III+ and RNase III- strains we could identify RNase III as the key element in the down-regulation of gene 19 expression. The sequencing of in vitro generated and RNase III-digested transcripts confirmed the in vivo studies and revealed the exact positions of the RNase III cleavage sites within the coding part of the gene 19 transcript. The in vitro determined RNase III cleavage of gene 19 mRNA was confirmed by in vivo primer extension analysis. Finally, we could show that an exchange of three nucleotides within the RNase III recognition site abolished RNase III cleavage in vitro.

Amino Acid Sequence↗
PubMed2003

LuxS-based signaling in Streptococcus gordonii: autoinducer 2 controls carbohydrate metabolism and biofilm formation with Porphyromonas gingivalis.

Communication based on autoinducer 2 (AI-2) is widespread among gram-negative and gram-positive bacteria, and the AI-2 pathway can control the expression of genes involved in a variety of metabolic pathways and pathogenic mechanisms. In the present study, we identified luxS, a gene responsible for the synthesis of AI-2, in Streptococcus gordonii, a major component of the dental plaque biofilm. S. gordonii conditioned medium induced bioluminescence in an AI-2 reporter strain of Vibrio harveyi. An isogenic mutant of S. gordonii, generated by insertional inactivation of the luxS gene, was unaffected in growth and in its ability to form biofilms on polystyrene surfaces. In contrast, the mutant strain failed to induce bioluminescence in V. harveyi and was unable to form a mixed species biofilm with a LuxS-null strain of the periodontal pathogen Porphyromonas gingivalis. Complementation of the luxS mutation in S. gordonii restored normal biofilm formation with the luxS-deficient P. gingivalis. Differential display PCR demonstrated that the inactivation of S. gordonii luxS downregulated the expression of a number of genes, including gtfG, encoding glucosyltransferase; fruA, encoding extracellular exo-beta-D-fructosidase; and lacD encoding tagatose 1,6-diphosphate aldolase. However, S. gordonii cell surface expression of SspA and SspB proteins, previously implicated in mediating adhesion between S. gordonii and P. gingivalis, was unaffected by inactivation of luxS. The results suggest that S. gordonii produces an AI-2-like signaling molecule that regulates aspects of carbohydrate metabolism in the organism. Furthermore, LuxS-dependent intercellular communication is essential for biofilm formation between nongrowing cells of P. gingivalis and S. gordonii.

Bacterial Adhesion↗
PubMed2003

Fine mapping of quantitative trait loci for improved fruit characteristics from Lycopersicon chmielewskii chromosome 1.

The near-isogenic line (NIL) TA1150 contains a 56-cM introgression from Lycopersicon chmielewskii chromosome 1 and has several interesting phenotypic characteristics including fruit with orange color, high levels of soluble solids, thick pericarp, small stem scars, and good firmness. A set of overlapping recombinant lines (subNILs) was developed and field tested to fine map the quantitative trait loci (QTL) controlling these traits. The results indicated that the solids, pericarp thickness, and firmness QTL are distinct from the color locus. Several of the QTL mapped in this study, including the soluble-solids QTL, probably correspond to QTL mapped in other wild species of tomato. However, analysis of a set of TA523 subNILs containing complementary introgressions from Lycopesicon hirsutum chromosome 1 suggests that this wild species may contain a different locus for improved soluble solids. Thus, it might be possible to combine the L. chmielewskii and L. hirsutum alleles for these loci in a single line with the potential for extremely highly soluble solids. The TA1150 subNIL TA1688 contains the smallest introgression of the solids locus (approximately 19 cM), as well as the pericarp thickness and firmness QTL, with a yield that was equivalent to two of the three control lines. Isolation of recombinant subNILs from TA1688 should break the linkage between orange color and high solids and provide a small introgressed segment for marker-assisted breeding and genetic improvement of processing tomato.

Alleles↗
PubMed1998

Restricted HIV-1 infection of human astrocytes: potential role of nef in the regulation of virus replication.

A small percentage of astrocytes are consistently infected in vivo by HIV-1 and may contribute to neuropathogenesis despite a non-productive infection. Overexpression of the nef gene product has been associated with their infection both in vivo and in vitro. We examined the role of the nef gene during HIV replication in astrocytes (U251MG cells) following transfection with pNL4-3 proviral plasmid or isogenic strains containing a deletion or point mutation in the nef gene (pNL4-3deltaNef; pNL4-3-nef-stop). We were able to initiate virus replication which peaked at 5 days post-transfection and became non-productive after 21 days. Nef protein expression by wild type pNL4-3 was observed at low levels compared to control HeLa cells at peak virus replication. At later time points after development of a non-productive infection, viral antigen and Nef protein was not detectable however virus was readily recovered by co-culture with CD4+T-cells. Interestingly, virus production was significantly enhanced by a 222 base pair deletion in the nef reading frame. This was not observed with a frame shifting point mutation in nef, indicating a suppressive effect of nef on virus production in astrocytes. The enhanced virus production from nef-deleted pNL4-3 in U251MG cells was not reversed by co-expression of Nef from a second Nef-expressing plasmid, and in fact Nef expression in trans had a further positive effect on virus production. This suggested opposing effects of the Nef protein and elements contained within the nef sequence on virus production in astrocytes. Despite the low expression of Nef by U251MG astrocytes, relatively high amounts of multiply spliced 2 kb mRNA were present compared to HeLa cells. These data demonstrate that an acute low-level infection of astrocytes rapidly becomes a non-productive infection and this process is assisted by sequences in nef. The low level Nef protein expression, despite high levels of mRNA, suggests a block in translation of multiply spliced HIV mRNA in astrocytes, or a translational control mechanism not yet characterised.

AIDS Dementia Complex↗
PubMed1993

Identification and cloning of the CHL4 gene controlling chromosome segregation in yeast.

A collection of chl mutants characterized by decreased fidelity of chromosome transmission and by minichromosome nondisjunction in mitosis was examined for the ability to maintain nonessential dicentric plasmids. In one of the seven mutants analyzed, chl4, dicentric plasmids did not depress cell division. Moreover, nonessential dicentric plasmids were maintained stably without any rearrangements during many generations in the chl4 mutant. The rate of mitotic heteroallelic recombination in the chl4 mutant was not increased compared to that in an isogenic wild-type strain. Analysis of the segregation of a marked chromosome indicated that sister chromatid nondisjunction and sister chromatid loss contributed equally to chromosome malsegregation in the chl4 mutant. A genomic clone of CHL4 was isolated by complementation of the chl4-1 mutation and was physically mapped to the right arm of chromosome IV near the SUP2 gene. Nucleotide sequence analysis of CHL4 clone revealed a 1.4-kb open reading frame coding for a 53-kD predicted protein which does not have homology to published proteins. A strain containing a null allele of CHL4 is viable under standard growth conditions but has a temperature-sensitive phenotype (conditional lethality at 36 degrees). We suggest that the CHL4 gene is required for kinetochore function in the yeast Saccharomyces cerevisiae.

Amino Acid Sequence↗
PubMed1998

Prevention of ischemically induced neointimal hyperplasia using ex vivo antisense oligodeoxynucleotides.

BACKGROUND: Chronic graft vascular disease in cardiac allografts results from coronary artery neointimal formation. Vascular ischemia has been shown to provoke the development of neointimal hyperplasia through endothelial damage. We used a rodent model of neointimal formation to test the in vivo effects of antisense oligodeoxynucleotides (AS ODN) specifically designed to block this process. METHODS: Aortas from ACI rats were mock transfected or transfected with 18 base pair AS ODNs against the 5' start codon region of both CDC2 kinase, and proliferating cell nuclear antigen (PCNA) mRNA. Transfection was accomplished by placing the aorta in ODN solution (transfected group, 40 micromol/L of each sequence) or saline solution alone (mock-transfected group) and exposing to hydrostatic pressure (2 atm) for 24 hours at 4 degrees C. Vessels were then interposition-grafted into the abdominal aorta of untreated isogenic recipients and procured on postoperative days (POD) 1, 6, and 14. RESULTS: Nuclear localization of fluorescein isothiocyanate ODN was observed in 81%+/-3% of medial smooth muscle cells at 24 hours after interposition grafting and reperfusion. Efficacy of AS ODNs at blocking CDC2 kinase and PCNA was verified on POD 6 by enzyme-linked immunosorbent assay. This blockade significantly reduced ischemically induced vascular narrowing on POD 14 as assessed by use of computerized image analysis (3.85%+/-2.45% luminal narrowing for transfected vs 7.11%+/-2.03% for control subjects, p < 0.03). CONCLUSIONS: Our data show the efficacy of AS ODN at blocking rat PCNA and CDC2 kinase up-regulation provoked by ischemia. This ex vivo approach had beneficial effects against vascular narrowing in a rodent model of ischemically induced neointimal hyperplasia, an antigen-independent factor important in the development of subsequent chronic graft vascular disease.

Animals↗
PubMed1978

Effect of adult thymectomy on tumour immunity in mice.

The effect of adult thymectomy in DBA/2J mice on the in vitro response to syngeneic tumour cells was investigated. Spleen cells from adult mice which had been thymectomized 8 weeks previously demonstrated a severely impaired primary cytotoxic response to P815 tumour cells, whereas their cytotoxic responses to allogeneic cells (C57BL/6) and to non-H-2 antigens (BALB/c), and their ability to form a primary antibody response to sheep red blood cells was unimpaired. Suppressor T cells, specific for P815 cells, appeared early in the thymuses of animals inoculated with P815 cells (between 4 and 8 days after tumour-cell injection). No differences in tumour growth between animals thymectomized as adults and sham-operated controls were observed, and thymectomized tumour-bearing animals had levels of specific suppressor cells in their lymph nodes equivalent to the levels found in untreated controls. Severely thymocyte-deprived animals which had been thymectomized, irradiated and reconstituted with either marrow or spleen cells 8 weeks before tumour implantation succumbed more rapidly to metastatic tumour than did control animals.

Animals↗
PubMed1990

Swimming causes myosin adaptations in the rat cardiac isograft.

To investigate the contributions of humoral and hemodynamic factors to cardiac adaptations associated with chronic exercise, female Fischer 344 rats were subjected to chronic swimming, infrarenal cardiac transplantation, or both. Swimming resulted in hypertrophy (11-12%) of the in situ hearts in both the unoperated and operated animals compared with the matched sedentary controls. The cardiac isograft exhibited atrophy (32-35%), which was not attenuated by swimming. The cardiac isograft was also associated with a decrease in the percent of V1 myosin isoenzyme, which was attenuated by swimming (45 +/- 5% versus 66 +/- 6%). Swimming also increased the percent of this isomyosin in the in situ hearts of operated rats. These data suggest that hemodynamic load and/or neural innervation are necessary for hypertrophy associated with chronic conditioning by swimming, whereas myosin isoenzyme control is significantly mediated by humoral factors.

Adaptation, Physiological↗
PubMed1987

Methylation of the alpha-fetoprotein gene in isogenic rat hepatoma and liver cell lines.

We have found that DNA methylation is inversely correlated with alpha-fetoprotein (AFP) gene expression in a series of isogenic rat hepatoma cell lines. The 5' end of the gene is extensively demethylated in AFP-producing cells and is highly methylated in cell lines which do not produce AFP. Glucocorticoid affects markedly the synthesis of AFP in the hepatoma cells. However, methylation patterns of cell lines which were treated with dexamethasone were not different from those of control cells, indicating that glucocorticoid action on AFP gene expression does not alter DNA methylation in this region of the gene.

Animals↗
PubMed1996

Differential expression of Leishmania major beta-tubulin genes during the acquisition of promastigote infectivity.

Tubulin expression has been analysed as the insect stage of the protozoan parasite Leishmania major differentiates from a non-infective to an infective form. This transformation of the promastigote stage occurs in vitro and analysis of beta-tubulin mRNA expression in axenically grown promastigotes showed that a 2200 nt transcript is predominately expressed in non-infective promastigotes. The message contains a motif associated with mRNA intracellular localisation and its level is reduced by an order of magnitude in infective promastigotes through a mechanism involving RNA stability. A 3200 nt RNA, the major beta-tubulin transcript in the infective stage, is encoded by a single copy gene at the 3' end of the array that encodes the 2200 nt RNA. These RNAs, as well as a gene encoding a beta-tubulin transcript highly up-regulated in the mammalian stage of the parasite, encode polypeptides that are apparently functionally equivalent but have highly diverged 3' untranslated regions. This differential regulation of the dispersed isogenes may reflect the involvement of a mechanism altering tubulin synthesis during the Leishmania life cycle. The analysis of alpha-tubulin RNA levels revealed the abundance of this message falls as promastigotes differentiate into an infectious stage and the transcript is destabilised in infective promastigotes. These data demonstrate that the regulation of mRNA half-life contributes to controlling gene expression as promastigotes differentiate into an infectious form.

Amino Acid Sequence↗
PubMed2001

TRAIL (APO-2L) induces apoptosis in human prostate cancer cells that is inhibitable by Bcl-2.

To determine if TRAIL-induced apoptosis in human prostate tumor cells was suppressed by bcl-2, we compared the levels of apoptosis induced by recombinant human TRAIL in pairs of isogenic cell lines that do or do not express bcl-2. Three human prostate tumor cell lines (PC3, DU145 and LNCaP) and their bcl-2-expressing counterparts were tested for their susceptibility to TRAIL. Cells were exposed to TRAIL in the presence of cycloheximide which acted as a sensitizer. Apoptosis was induced rapidly in PC3 and DU145 neo-control transfected cells, whereas induction in LNCaP required 24 h. All three cell line variants expressing bcl-2 were resistant to the apoptotic effects of TRAIL. Caspase 3 and 8 activation was also detected in the neo control cells after treatment with TRAIL, demonstrating the rapid activation of the caspase cascade similar to that seen with other death receptors. Bcl-2 overexpression in these cells blocked activation of these caspases, suggesting that bcl-2 expression of human cancer cells may be a critical factor in the therapeutic efficacy of TRAIL.

Antineoplastic Agents↗
PubMed2000

Virulence factors of Helicobacter pylori responsible for gastric diseases in Mongolian gerbil.

Helicobacter pylori infection induces various gastroduodenal diseases. We examined the role of two genes, vacA and cagE, in the gastric pathogenesis induced by H. pylori using a long-term (62 wk) animal model. Reportedly, both genes are associated with the virulence of H. pylori: vacA encodes vacuolating cytotoxin, and cagE, with other genes in the cag pathogenicity islands, encodes a type IV secretion system. Mongolian gerbils were challenged in this study by a wild-type TN2 strain and its isogenic mutants of cagE or vacA. The wild-type and vacA mutants induced severe gastritis, whereas cagE mutants induced far milder changes. Gastric ulcer was induced at the highest rate (22/23) by the wild-type TN2, followed by the vacA mutant (19/28). No ulcer was found in the gerbils infected with the cagE mutant (0/27) or in controls (0/27). Intestinal metaplasia was also found in the gerbils infected with the wild-type (14/23) or vacA mutant (15/28). Gastric cancer developed in one gerbil with wild-type infection and in one with vacA mutant infection. In conclusion, the knocking out of the cagE gene deprived wild-type H. pylori of the pathogenicity for gastritis and gastric ulcer, suggesting that the secretion system encoded by cag pathogenicity island genes plays an essential role.

Adenocarcinoma↗
PubMed2002

Regulatory interactions between the Pho and sigma(B)-dependent general stress regulons of Bacillus subtilis.

When Bacillus subtilis is subjected to phosphate starvation, the Pho and sigma(B)-dependent general stress regulons are activated to elicit, respectively, specific and non-specific responses to this nutrient-limitation stress. A set of isogenic mutants, with a beta-galactosidase reporter gene transcriptionally fused to the inactivated target gene, was used to identify genes of unknown function that are induced or repressed under phosphate limitation. Nine phosphate-starvation-induced (psi) genes were identified: yhaX, yhbH, ykoL and yttP were regulated by the PhoP-PhoR two-component system responsible for controlling the expression of genes in the Pho regulon, while ywmG (renamed csbD), yheK, ykzA, ysnF and yvgO were dependent on the alternative sigma factor sigma(B), which controls the expression of the general stress genes. Genes yhaX and yhbH are unique members of the Pho regulon, since they are phosphate-starvation induced via PhoP-PhoR from a sporulation-specific sigma(E) promoter or a promoter that requires the product of a sigma(E)-dependent gene. Null mutations in key regulatory genes phoR and sigB showed that the Pho and sigma(B)-dependent general stress regulons of Bacillus subtilis interact to modulate the levels at which each are activated.

Bacillus subtilis↗
PubMed2005

A stereological analysis of fibrosis and inflammatory reaction induced by four different synthetic slings.

OBJECTIVES: To analyse quantitatively, using stereological methods, the density of the collagen fibres induced by four types of sling materials, and verify by a histopathological analysis the corresponding inflammatory reaction, as fibrosis secondary to sling implantation is considered responsible for restoring urethral support and re-establishing continence in women with stress urinary incontinence, and new synthetic materials that promote adequate fibrosis with the least intensity and duration have been proposed to substitute the aponeurotic sling. MATERIALS AND METHODS: The study comprised 70 isogenic white Wistar rats divided into three groups: group A (30 rats) had 8 x 4 mm strips of silicone and porcine small intestine submucosa (SIS) implanted in the abdominal subcutaneous tissues; group B (30 rats) had 8 x 4 mm strips of polycaprolactone and polylactic acid copolymers and monofilament polypropylene (PLP) implanted the abdominal subcutaneous layer; while a control group of 10 rats had dissection and suturing with 5/0 Nylon in the abdominal subcutaneous layer, as used to fix the strips in the other rats. Picro-Sirius staining was used to assess collagen fibres, and haematoxylin-eosin for the histopathological study. At 7, 30 and 90 days after surgery, 10 rats from each group were killed and assessed. RESULTS: After 7 days all the materials induced a moderate inflammatory reaction that did not differ from that in the control group. At 30 days there was no difference between the control and polycaprolactone and polylactic acid copolymers, having the least inflammatory reaction. PLP and silicone produced a moderate inflammatory reaction, while the porcine SIS induced a more intense reaction. At 90 days there was a more intense inflammatory reaction in polycaprolactone and polylactic acid copolymers than before but it was less than with PLP and silicone, which again were no different. During this period the inflammatory reaction induced by SIS was greater. The stereological analysis indicated that collagen fibres induced by polycaprolactone and polylactic acid copolymers and PLP were less dense (61% and 65%, respectively), and significantly less than with silicone (85%) and SIS (86%). CONCLUSION: PLP was the best nonabsorbable material as it induced a less intense inflammatory reaction than the other tested materials. As porcine SIS was completely absorbed the intense fibrosis induced is useful, as it is exclusively responsible for the urethral support later after surgery.

Animals↗