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Biomedical subjects

M O'Dwyer

Publications and source records attributed to M O'Dwyer.

11 recordsLinked to original sources
PubMed2001

Role of double-stranded RNA-dependent protein kinase in mediating hypersensitivity of Fanconi anemia complementation group C cells to interferon gamma, tumor necrosis factor-alpha, and double-stranded RNA.

Hematopoietic cells bearing inactivating mutations of Fanconi anemia group C (FANCC) are excessively apoptotic and demonstrate hypersensitivity not only to cross-linking agents but also to interferon gamma (IFN-gamma) and tumor necrosis factor-alpha. Seeking essential signaling pathways for this phenotype, this study quantified constitutive and induced RNA-dependent protein kinase (PKR) activation in Fanconi anemia cells of the C complementation group (FA-C). PKR was constitutively phosphorylated and exhibited an increased binding affinity for double-stranded RNA (dsRNA) in FANCC(-/-) cells. FANCC(-/-) cells were hypersensitive to both dsRNA and the combination of dsRNA and IFN-gamma in that these agents induced a higher fraction of apoptosis in FANCC(-/-) cells than in normal cells. Overexpression of wild-type PKR-sensitized FANCC(-/-) cells to apoptosis induced by IFN-gamma and dsRNA. Conversely, inhibition of PKR function by enforced expression of a dominant-negative inhibitory mutant of PKR (PKRDelta6) substantially reduced the IFN and dsRNA hypersensitivity of FANCC(-/-) cells. Two PKR target molecules, IkappaB-alpha and IRF-1, were not differentially activated in FANCC(-/-) cells, but enforced expression of a nonphosphorylatable form of eukaryotic translation initiation factor-2alpha reversed the PKR-mediated block of messenger RNA translation and partially abrogated the PKR-mediated apoptosis in FANCC(-/-) cells. Because no evidence was found of a PKR/FANCC complex in normal cells, it was concluded that an essential function of FANCC is to suppress, indirectly, the activity of PKR and that FANCC inactivation results in IFN hypersensitivity, at least in part, because this function of FANCC is abrogated.

Animals↗
PubMed2000

Interferon-gamma-induced apoptotic responses of Fanconi anemia group C hematopoietic progenitor cells involve caspase 8-dependent activation of caspase 3 family members.

Hematopoietic progenitor cells (HPC) from mice nullizygous at the Fanconi anemia (FA) group C locus and children with Fanconi anemia group C (FA-C) are hypersensitive to interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha. This hypersensitivity results, in part, from the capacity of these cytokines to prime the fas pathway. Because fas-mediated programmed cell death in many cells involves sequential activation of specific caspases, we tested the hypothesis that programmed cell death in FA HPC involves the ordered activation of specific caspase molecules. Lysates from lymphoblasts treated with both agonistic anti-fas antibody and IFN-gamma contained activated caspase 3 family members (caspases 3, 6, and 7), as well as caspase 8, whereas activation of caspases 1, 2, 4, 9, and 10 was not detected. The apoptotic effects of fas agonists in IFN-gamma-treated human and murine FA-C cells were blocked when pretreated with inhibitors (ac-DEVD-cho, CP-DEVD-cho, Z-DEVD-FMK) of the caspase 3 protease. Inhibitors (ac-YVAD-cho, CP-YVAD-cho, Z-YVAD-FMK) of caspase 1 did not block apoptosis or caspase 3 activation. Treatment of FA cells with the fluoromethyl ketone tetrapeptide caspase 8 inhibitor (ac-IETD-FMK) did suppress caspase 3 activation. A 4-fold greater fraction of IFN-induced FA-C cells expressed caspase 3 than FA-C cells complemented by retroviral-mediated transfer of FANCC. Therefore fas-induced apoptosis in Fanconi anemia cells of the C type involves the activation of caspase 8, which controls activation of caspase 3 family members and one direct or indirect function of the FANCC protein is to suppress apoptotic responses to IFN-gamma upstream of caspase 3 activation. (Blood. 2000;96:4204-4211)

Animals↗
PubMed2000

Analysis of resveratrol in wine by capillary electrophoresis.

Capillary electrophoresis (CE) is a new analytical technique that has recently been reported as a method for analysis of resveratrol in wine. Several different separation approaches have been taken in these reports. In comparison with high-performance liquid chromatography (HPLC), CE methods have similar sensitivity and can discriminate between trans- and cis-isomers of resveratrol. CE methods also show promise for analysis of other flavonoid antioxidants (glycosides and aglycones) in wine.

Electrophoresis, Capillary↗
PubMed1998

Direct Analysis of Resveratrol in Wine by Micellar Electrokinetic Capillary Electrophoresis.

Resveratrol is an antioxidant found in grapes and other plants. It has been reported to have health benefits including anticarcinogenic activity and protection against coronary heart disease. Previous methods for its quantification in wines have relied on HPLC and GC. However, an alternative method utilizing capillary electrophoresis (CE) has been developed in our laboratory. The CE procedure resolves cis- and trans-resveratrol isomers, using a micellar mode of separation. The detection limit for resveratrol was 1.25 µM using UV detection at 310 nm. This procedure was employed for the direct analysis of trans-resveratrol in wines. The direct analysis method used was advantageous because the analysis time was greatly reduced and sample preparation procedures that might result in resveratrol isomerization were avoided. The CE procedure was used to determine the level of trans-resveratrol in several California red wines, and the values obtained were in close agreement with those reported using other methods.

Journal Article↗
PubMed1995

Children discharged following nutritional rehabilitation: a follow-up study.

The results of a 1-year follow up of children discharged from a nutritional rehabilitation centre (NRC) in eastern Nepal are presented. One hundred and one children were visited at home on discharge from the NRC, at 6 months and 1 year later. Data were collected concurrently on one sibling and two neighbouring children matched for age and sex. Anthropometry was carried out at each visit and information collected on socio-economic variables and child feeding practices. A significantly higher death rate was observed in index children as compared with controls. Index children demonstrated catch-up growth with both greater weight gain and faster increases in height than either control group post-discharge. At 6 months, mean weight-for-height was similar in all three groups but at one year mean height-for-age was still lower in the index children as compared with controls. The prognosis for female children appeared to be poorer than for boys.

Case-Control Studies↗
PubMedDate not supplied

Capillary electrophoretic analysis of cathepsin D action on hemoglobin.

The action of purified cathepsin D on hemoglobin was examined using micellar electrokinetic chromatographic separation of peptide products. Purified cathepsin D was incubated with hemoglobin in 40 mM Na-formate pH 3.1 at 37 degrees C for varying lengths of time. The reaction was stopped by the addition of the inhibitor pepstatin, and the peptide products were isolated from the reaction mixture by ultrafiltration with a 10,000 molecular weight cut-off (MWCO) microfuge type filter. Filtered samples were then separated in 100 mM Tris-Cl pH 8.5 containing sodium dodecyl sulfate (SDS) or Na-deoxycholate at micellar concentrations using a 50-microns (i.d.) fused-silica capillary. Detection was performed at 214 nm. It was found that Na-deoxycholate containing separations were superior in resolution and required less time. This technique was used to determine initial velocity (expressed as peak area per unit time) for nine peptides. Several peptides resulted after very short incubation times (< 10 min). This suggests that this approach may be useful for the determination of cathepsin D activity.

Animals↗