Protein expression profiles in human breast ductal carcinoma and histologically normal tissue

Reference two-dimensional (2-D) gels are presented for human breast ductal carcinoma and histologically normal tissue. Whole biopsy fragments were analyzed, including epithelial and nonepithelial components. Thirty-five spots have been assigned by gel matching to the human liver SWISS-2DPAGE reference map and/or to the human primary keratinocyte IPG map from the Danish Center for Human Genome. N-terminal microsequencing was applied to confirm randomly chosen matching assignments and to identify six new spots. Protein expression profiles in ductal carcinoma and in normal breast tissue appeared to be similar, except for a pattern consisting of 32 spots, which were highly expressed in all carcinoma specimens, and less intense and occasionally undetectable in normal tissue. This difference was statistically significant. Assignment has been obtained for several spots, namely GRP94, GRP78, GRP75, mitochondrial HSP60, calreticulin, protein disulfide isomerase, peptidyl-prolyl cis-trans isomerase, collagen-binding protein 2, fructose bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase, thioredoxin, cytochrome c oxidase VA subunit, tubulin beta isoform and macrophage migration inhibitory factor (MIF). The cancer- and tissue-specificity of the described pattern was assessed by matching to the Swiss-2DPAGE human liver, hepatoma, lymphoma, erythroleukemia reference maps. The pattern of 32 spots was found to be indicative of epithelial neoplasia.     

Molecular modelling of mammalian CYP2B isoforms and their interaction with substrates, inhibitors and redox partners

1. The construction of three-dimensional models of CYP2B isozymes from rat (CYP2B1), rabbit (CYP2B4) and man (CYP2B6), based on a multiple sequence alignment with CYP102, a unique eukaryotic-like bacterial P450 (in terms of possessing an NADPH-dependent FAD- and FMN-containing oxidoreductase redox partner) of known crystal structure, is reported. 2. The enzyme models described are shown to be consistent with experimental evidence from site-directed mutagenesis studies, antibody recognition sites and amino acid residues identified as being associated with redox partner interactions, together with the location of a key serine residue (Ser-128) likely to be involved in protein kinaseA-mediated phosphorylation. 3. A substantial number of known substrates and inhibitors of CYP2B isozymes are shown to fit the putative active sites of the enzyme models in agreement with their reported position of metabolism or mode of inhibition respectively. In particular, there is complementarity between the characteristic non-planar geometries of CYP2B substrates and key groups in the enzymes' active sites. 4. Molecular modelling of CYP2B isozymes appears to rationalize a number of the reported findings from quantitative structure-activity relationship investigations on series of CYP2B substrates and inhibitors.     

Symptomatic recurrent carotid stenosis and aneurysmal degeneration after endarterectomy

BACKGROUND: Aneurysmal degeneration of a carotid reconstruction was not recognized until the patient, who was known to have recurrent carotid artery stenosis, had a thromboembolic stroke. This sequelae of carotid endarterectomy is a serious complication, associated with a high morbidity and mortality rate. This review was conducted to establish the risk of transient ischemic attack and stroke for patients found to have recurrent carotid stenosis associated with aneurysmal degeneration of the carotid artery after endarterectomy. METHODS: A case is reported, and 100 literature references of aneurysmal degeneration of the carotid artery after endarterectomy were reviewed. RESULTS: False aneurysm from anastomotic disruption was the most common presentation identified in the cases reviewed. Nineteen of the patients had a significant neurologic event; however, three (50%) of six patients with aneurysm and recurrent carotid artery stenosis had a transient ischemic attack or stroke. CONCLUSIONS: The incidence of neurologic symptoms is markedly increased when recurrent carotid artery stenosis is associated with carotid aneurysm. During postoperative surveillance after endarterectomy, the identification of recurrent carotid artery stenosis requires evaluation for aneurysmal degeneration of the carotid artery with duplex scanning. These patients are at significant risk for transient ischemic attack and stroke. This rare complication merits operative repair.     

Femorodistal vein grafts: the utility of graft surveillance criteria

PURPOSE: This retrospective review of femorodistal vein grafts was analyzed to determine the usefulness of various graft surveillance criteria. METHOD: The surveillance schedule involved evaluations at 1 month, every 3 months the first year, and then every 6 months. Salvage intervention or graft occlusion occurring within the next follow-up interval defined surveillance end points. One hundred two grafts (329 surveillance visits) had an ankle/brachial index (ABI). A duplex scanning-determined midgraft peak systolic flow velocity (PSFV) was available for 81 grafts (262 visits). Forty-eight grafts (137 visits) had both a PSFV and entire graft duplex scanning (EGDS) to determine stenosis greater than 50%, whereas 40 grafts (91 visits) had simultaneous ABI and EGDS. RESULTS: When a greater than 15% decrease in ABI denoted an abnormal surveillance study result, a positive predictive value (PPV) of 24.3% and negative predictive value of 94.5% were noted. Similarly, a PSFV cutoff of less than 35 cm/sec demonstrated values of 26.3% and 94.2%, respectively. When an EGDS of greater than 50% stenosis or a PSFV of less than 35 cm/sec were the cutoff criteria, the PPV was 36.7% and negative predictive value 99.1%, whereas characterizing abnormal results further with ABI (> 15% decreases) increased the PPV to 83.3%. CONCLUSION: The combination of an EGDS, midgraft PSFV, and ABI provides optimal follow-up for our patients with a femorodistal vein graft.     

DNA recognition by the EcoK methyltransferase. The influence of DNA methylation and the cofactor S-adenosyl-L-methionine

The methyltransferase of the EcoK type I restriction/modification system is trimeric, M2S1, where the S subunit determines the sequence specificity of the enzyme. The methyltransferase has a strong preference for hemimethylated substrate DNA and, therefore, we have investigated the effect of the methylation state of DNA on binding by the enzyme, together with the effects on binding of the cofactor S-adenosyl-L-methionine. Our results indicate that the methyltransferase has two non-interacting S-adenosyl-L-methionine binding sites, each with a dissociation constant of 3.60 (+/- 0.42) microM determined by equilibrium dialysis, or 2.21 (+/- 0.29) microM determined by the displacement of a fluorescent probe. Ultraviolet light-induced crosslinking showed that S-adenosyl-L-methionine binds strongly only to the modification (M) subunits. Changes in the sedimentation velocity of the methyltransferase imply a protein conformational change due to S-adenosyl-L-methionine binding. Gel retardation results show that the binding of S-adenosyl-L-methionine to the methyltransferase enhances binding to both specific and non-specific DNAs, but the enhancement is greater for the specific DNA. Differences in binding affinities contribute to the recognition of the specific nucleotide sequence AAC(N)6GTGC by the methyltransferase in preference to a non-specific sequence. In contrast, although the complexes of unmodified and hemimethylated DNAs with the methyltransferase have different mobilities in non-denaturing gels, there appears to be no contribution of binding affinity to the distinction between these two substrates. Therefore, the preference for a hemimethylated substrate must be due to a difference in catalysis.     

Recall of Descriptive Information: The Roles of Presentation Format, Annotation Strategy, and Individual Differences

The benefits of using a comprehensive annotation strategy (employing underlining/circling, making connections, asking questions, and making comments) with knowledge maps (spatial/verbal arrays) and traditional, linear text to improve free recall scores for learners with individual differences in vocabulary and comprehension ability were examined. Types and frequencies of annotations generated were also examined for each stimulus format condition. Multiple regression analyses indicate that the frequency of use of two component annotation strategies, asking questions and making connections, were significant predictors of recall scores, while frequency of underlining/circling and generating elaborations failed to predict recall scores. Text users generated more underlining/circling, while knowledge map users generated more connections between ideas, suggesting that knowledge maps may facilitate the application of more productive annotation strategies. Also examined were the interrelationships between vocabulary ability, comprehension ability, and free recall scores. Copyright 1997Academic Press