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Biomedical subjects

J M Payne

Publications and source records attributed to J M Payne.

At least 19 recordsLinked to original sources
PubMed1999

Leukemic HRX fusion proteins inhibit GADD34-induced apoptosis and associate with the GADD34 and hSNF5/INI1 proteins.

One of the most common chromosomal abnormalities in acute leukemia is a reciprocal translocation involving the HRX gene (also called MLL, ALL-1, or HTRX) at chromosomal locus 11q23, resulting in the formation of HRX fusion proteins. Using the yeast two-hybrid system and human cell culture coimmunoprecipitation experiments, we show here that HRX proteins interact directly with the GADD34 protein. We have found that transfected cells overexpressing GADD34 display a significant increase in apoptosis after treatment with ionizing radiation, indicating that GADD34 expression not only correlates with apoptosis but also can enhance apoptosis. The amino-terminal third of the GADD34 protein was necessary for this observed increase in apoptosis. Furthermore, coexpression of three different HRX fusion proteins (HRX-ENL, HRX-AF9, and HRX-ELL) had an anti-apoptotic effect, abrogating GADD34-induced apoptosis. In contrast, expression of wild-type HRX gave rise to an increase in apoptosis. The difference observed here between wild-type HRX and the leukemic HRX fusion proteins suggests that inhibition of GADD34-mediated apoptosis may be important to leukemogenesis. We also show here that GADD34 binds the human SNF5/INI1 protein, a member of the SNF/SWI complex that can remodel chromatin and activate transcription. These studies demonstrate, for the first time, a gain of function for leukemic HRX fusion proteins compared to wild-type protein. We propose that the role of HRX fusion proteins as negative regulators of post-DNA-damage-induced apoptosis is important to leukemia progression.

Antigens, Differentiation↗
PubMed1999

Effective laser ablation of bone based on the absorption characteristics of water and proteins.

BACKGROUND: Collateral thermal damage and residual char formation have severely limited the use of conventional lasers in the surgical preparation of bony tissue. Thermal damage from lasers can be minimized by selecting a wavelength that is strongly absorbed and by reducing the laser pulse duration. In contrast to the fixed wavelengths and microsecond pulse intervals of conventional lasers, the Vanderbilt free electron laser (FEL) can be set at wavelengths ranging from 2.1 to 9.8 microm, and the pulse duration can be reduced to a series of 1 to 2 picosecond (ps) micropulses delivered in succession over intervals of 4 microsecond macropulses. The purpose of this study was to compare the morphologic and chemical changes induced in the near-surface region of bone following exposure to the FEL at 3.0, 6.1, and 6.45 microm wavelengths. The selected wavelengths coincide with the vibrational modes of proteins and water within bone. METHODS: Under general anesthesia, laser incisions were made in the tibias of 14 skeletally mature rabbits. Laser parameters included 22.5+/-2.5 mJ/pulse delivered in individual 4 microsecond macropulses at a repetition rate of 30 Hz, focused to 200 microm and 500 microm spot sizes. Laser incisions were made using a computer-assisted surgical program, and control incisions were created with a bone saw. Rabbits were euthanized after the final incision, tibias recovered, and non-decalcified specimens processed for light microscopy. Separate samples were prepared for FTIR (Fourier transform infrared) photoacoustic spectroscopic analysis. RESULTS: The light microscopy sections of the ablation defects created at the differing wavelengths showed similar features, i.e., 2 zones of collateral damage, a zone generally < 10 mm of extensive thermal damage, and a wider zone of empty lacunae. In comparing treated and untreated surfaces, the spectral differences were limited to a relative decrease in intensity of the amide II and III absorption peaks in all laser-treated surfaces. CONCLUSIONS: Spectroscopic and histologic results indicated minimal thermal damage to bone ablated at 3.0, 6.1, and 6.45 microm wavelengths using the FEL (Fourier transform infrared) at the specified parameters. The FTIR photoacoustic spectroscopic results suggest that the char layer is limited to an area less than approximately 6 microm from the surface.

Absorption↗
PubMed1997

Safety-modified episomal vectors for human gene therapy.

The effectiveness of ongoing gene therapy trials may be limited by the expression characteristics of viral and plasmid-based vectors. To enhance levels of heterologous gene expression, we have developed a safety-modified episomal expression vector that replicates extrachromosomally in human cells. This vector system employs a simian virus 40 (SV40) large T antigen mutant (107/402-T) that is deficient in binding to human tumor suppressor gene products, including p53, retinoblastoma, and p107, yet retains replication competence. These SV40-based episomes replicate to thousands of copies by 2-4 days after gene transfer in multiple types of human cell lines, with lower activity in hamster cells, and no detectable activity in dog, rat, and murine cell lines. Importantly, 107/402-T has enhanced replication activity compared with wild-type T antigen; this finding may be due, in part, to the inability of p53 and retinoblastoma to inactivate 107/402-T function. We demonstrate that the level and duration of 107/402-T expression regulates the observed episomal copy number per cell. Compared with standard plasmid constructs, episomes encoding 107/402-T yield approximately 10- to 100-fold enhanced levels of gene expression in unselected populations of transient transfectants. To determine if 107/402-T-based episomes replicate extrachromosomally in vivo, tumor explants in nude mice were directly injected with liposome/DNA complexes. Using a PCR-based assay, we demonstrate that SV40-based episomes replicate in human cells after direct in vivo gene transfer. These data suggest that safety-modified SV40-based episomes will be effective for cancer gene therapy because high level expression of therapeutic genes in transient transfectants should yield enhanced tumor elimination.

Animals↗
PubMed1996

Migration of human gingival fibroblasts over guided tissue regeneration barrier materials.

The purpose of this investigation was to determine the ability of human gingival fibroblasts, in vitro, to migrate along a chemotactic gradient over 3 different guided tissue regeneration barrier materials; i.e., polytetrafluoroethylene, polylactic acid, and sterile calcium sulfate. Forty petri dishes were divided into 4 equal groups. In each group of 10 dishes, a different barrier material served as the fibroblast substrate with the polystyrene floor of one group of Petri dishes serving as the control. The under agarose technique of measuring cell migration was employed using platelet derived growth factor-BB homodimer as the chemoattractant and Hanks balanced salt solution to test random migration. In addition, fibroblasts were directly cultured on triplicate sets of barrier materials and the control surface for 24 hours and examined by scanning electron microscopy. Comparative analysis of the fibroblast migration data showed the mean migration distance (adjusted for random migration) for controls to be significantly greater than any of the three barrier materials. Further, mean migration distance over calcium sulfate was significantly greater when compared to that of the polylactic acid barrier group. All other comparisons between groups were not statistically significant. Scanning electron microscopic examination fibroblasts cultured directly on barrier membranes and compared to controls indicated that the calcium sulfate substrate appeared to facilitate cell attachment and spreading whereas cells on polytetrafluoroethylene and polylactic acid barriers exhibited a morphology not conducive to migration or, in many cases, cell health. Based on these limited in vitro results and, given the 3 barrier materials considered, it would appear that calcium sulfate offers the greater potential for guided tissue regeneration in surgical sites where primary wound closure cannot be obtained.

Becaplermin↗
PubMed1993

Partial purification and characterization of two distinct protein kinases that differentially phosphorylate the carboxyl-terminal domain of RNA polymerase subunit IIa.

RNA polymerase II is a multisubunit enzyme composed of two large subunits of molecular weight in excess of 100,000 and a collection of 8-10 smaller subunits. The largest subunit, designated IIa, contains at its carboxyl terminus a highly repetitive domain consisting of tandem repeats of the consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. Extensive phosphorylation within this COOH-terminal domain (CTD) gives rise to subunit IIo which has a markedly reduced mobility in SDS-polyacrylamide gel electrophoresis (PAGE) relative to subunit IIa. Recent evidence suggests that RNA polymerase IIA, containing an unphosphorylated CTD, is involved in preinitiation complex assembly, whereas RNA polymerase IIO is involved in elongation. Consequently, CTD phosphorylation is thought to occur after RNA polymerase II has bound to the promoter by a protein kinase that stably associates with the preinitiation complex. We present here the partial purification and characterization of two distinct CTD kinases from a HeLa cell transcription extract. These CTD kinases, designated CTDK1 and CTDK2, are fractionated by chromatography on Mono Q. CTDK1 catalyzes the incorporation of approximately 33 pmol of phosphate/pmol of calf thymus RNA polymerase subunit IIa, almost exclusively on serine. CTDK2 catalyzes the incorporation of approximately 50 pmol of phosphate/pmol of calf thymus subunit IIa, predominantly on serine; appreciable phosphate transfer onto threonine is also observed. Phosphorylation by CTDK2, but not CTDK1, results in a complete mobility shift in SDS-PAGE of subunit IIa to the position of IIo. CTDK1 can utilize ATP, dATP, or GTP as phosphate donor, whereas CTDK2 can utilize only ATP or dATP. The apparent Km for ATP is 30 microM for CTDK1 and 60 microM for CTDK2. CTDK1 and CTDK2 also differ in their protein substrate specificity. CTDK1 phosphorylates casein whereas CTDK2 does not. Neither kinase phosphorylates phosvitin or histone H1 to an appreciable extent. CTDK1 and CTDK2 do not appear to be related to cdc2 kinases as determined by their inability to phosphorylate H1 and their failure to react with antibodies directed against the cdc2 kinase. These results establish that a partially fractionated HeLa transcription extract contains two distinct CTD kinases that differ in their nucleotide requirements and in their patterns of CTD phosphorylation.

Amino Acid Sequence↗
PubMed1991

Pneumonia and empyema infection associated with a Bacillus species that resembles B. alvei.

An organism resembling Bacillus alvei was isolated from the lung and pleural fluid of an immunocompetent patient. The isolate differed from the type strain of B. alvei in its ability to reduce nitrate and its inability to produce dihydroxyacetone and acetylmethylcarbinol. The isolate was resistant to ciprofloxacin and showed intermediate susceptibility to vancomycin.

Bacillaceae Infections↗
PubMed1989

The transition of RNA polymerase II from initiation to elongation is associated with phosphorylation of the carboxyl-terminal domain of subunit IIa.

Mammalian cells contain two forms of RNA polymerase II, designated IIO and IIA, that differ in the extent of phosphorylation within the C-terminal domain of their largest subunit. Phosphorylation of this domain, which results in the conversion of RNA polymerase IIA to IIO, may play an important role in the transition from the initiation to the elongation phase of transcription. A third form of the enzyme, RNA polymerase IIB, is found in vitro and lacks the repetitive C-terminal domain. Purified calf thymus RNA polymerase IIA was labeled selectively with casein kinase II in the presence of [gamma-32P]ATP and used as substrate for the identification and partial purification of factors that catalyze the conversion of RNA polymerase IIA to IIO. HeLa cell S-100 transcription extracts contain such an activity that cofractionates with factors essential for promoter-dependent transcription through heparin-Sepharose, DEAE-5PW, and DE52 chromatography. The activity is dependent on either ATP, GTP, or dATP, requires a hydrolyzable beta,gamma-phosphoanhydride bond, and cannot utilize pyrimidine nucleoside triphosphates. This observation supports the idea that the conversion activity is a protein kinase. Transcription of the major late promoter of adenovirus-2 was carried out in the presence of a reconstituted transcription extract containing purified RNA polymerases IIO, IIA, or IIB, and the nature of the elongating enzyme was determined by photoaffinity labeling. When the reaction was initiated with RNA polymerase IIO or IIB, nascent transcripts were found cross-linked to subunit IIo or IIb, respectively. However, when the reaction was initiated with RNA polymerase IIA, nascent transcripts were cross-linked to subunit IIo. Consequently, phosphorylation of the C-terminal domain of subunit IIa must have occurred prior to elongation. The copurification of RNA polymerase IIA to IIO conversion activity with factors essential for promoter-dependent transcription and the observation that RNA polymerase II containing an unphosphorylated C-terminal domain is phosphorylated prior to elongation suggest that protein kinases that phosphorylate the C-terminal domain of subunit IIa may play an essential role in transcription.

Animals↗
PubMed1989

Amnesia of a passive avoidance task due to the beta 2-adrenoceptor antagonist ICI 118,551.

The selective beta 2-adrenoceptor antagonist ICI 118,551 induces amnesia in the domestic chick when given systemically, 10 min after a one-trial PAL task. Young chicks will spontaneously peck at a small bright bead. If the bead has been coated with a distasteful substance, the chicks will learn in a single trial not to peck at a similar bead on subsequent presentation. Administration of ICI 118,551 prevented retention of this task. Vehicle-injected chicks which learnt the task, avoided a similar bead to the training bead in the retention test, but did not avoid a bead of a different colour. The effect of ICI 118,551 is unlikely to be a direct effect on performance since amnesic chicks pecked both beads freely and equally in the test.

Adrenergic beta-Antagonists↗
PubMed1987

Isolation of Stomatococcus mucilaginosus from drug user with endocarditis.

Stomatococcus mucilaginosus was isolated from the blood of a patient with endocarditis and a past history of drug abuse and aortic valve replacement. At autopsy, Gram stain of the aortic valve revealed gram-positive cocci. Our isolate was atypical for S. mucilaginosus in that colonies were nonmucoid and nonadherent to agar surfaces. Cellular capsules were demonstrated by light and electron microscopy. Phenotypic characteristics identified by conventional methods as well as profile numbers obtained by using two commercial identification systems for staphylococci, the API Staph-Ident and the dms Staph Trac, are presented. Practical tests that differentiate S. mucilaginosus from the genera Micrococcus and Staphylococcus include growth on nutrient agar containing salt and lysostaphin susceptibility. Additional tests that helped differentiate our isolate from group D streptococci included hydrolysis of L-pyrrolidonyl-beta-naphthylamide and streptococcal serogrouping.

Adult↗
PubMed1978

Observations of the first service non-return rates of the hypoglycaemic, concentrate-fed dairy herds.

Fourteen dairy herds which had been subjected to the Compton Metabolic Profile test on a total of 29 occasions were selected from the records of the institute for Research on Animal Diseases, United Kingdom, because they had a wide range of mean blood-glucose concentrations. Two herds had to be rejected from the study because of inadequate breeding records, leaving 12 herds (26 profile tests) to be included. The first service non-return rate of each herd was calculated from the breeding records of 10 cows mated closest to the date of each profile test. These rates were positively related to the mean blood-glucose concentrations of the lactating cows (P less than 0.01).

Animals↗
PubMed1975

The influence of dietary protein upon blood composition in dairy cows.

Diets of different protein content were fed to dairy cows in two experiments of seven months and 12 months duration. Significant differences in the mean concentrations of serum urea, albumin and copper and of blood haemoglobin and packed cell volume were observed between cows receiving the various diets. The greatest differences occurred when cows were in mid-lactation. It was concluded that concentrations of urea in the serum of the cows were reflecting dietary protein intake and that concentrations of albumin, haemoglobin and PCV during lactation were affected by long-term protein status. The possible significance of the results is discussed in relation to previous reports indicating interrelationships between protein and copper metabolism, and in relation to the recommended protein content of diets for cows.

Animal Feed↗