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PubMed1987

The Blalock-Taussig shunt in infants: standard versus modified.

In recent years, the modified Blalock-Taussig shunt--a polytetrafluoroethylene graft from the subclavian artery to the pulmonary artery--has been preferred over the standard shunt by some surgeons because (1) it requires less dissection and (2) length of native vessels is not critical. From January, 1979, to June, 1985, we operated on 51 infants less than 1 year of age, including 26 less than 1 week of age, to palliate severe complex cyanotic congenital cardiac malformations. Twenty-four modified Blalock-Taussig shunts and 29 standard Blalock-Taussig shunts were created. The groups were concurrent. We reviewed all available cineangiograms and measured branch pulmonary and subclavian arteries. Pulmonary artery index was not different preoperatively in patients given a modified versus a standard Blalock-Taussig shunt (144 +/- 118 and 118 +/- 59 mm2/m2, respectively), but it was greater postoperatively in patients with a modified shunt (431 +/- 188 and 189 +/- 106 mm2/m2) (p = 0.07). Distortion of the pulmonary artery occurred less often after a modified Blalock-Taussig shunt (4/11) than a standard Blalock-Taussig shunt (6/8) (p = 0.06), though none of the distortions was severe. Early and late shunt failure occurred less often with a modified shunt (5/24) than with a standard shunt (15/29) (p less than 0.05). The modified Blalock-Taussig shunt had advantages over the standard Blalock-Taussig shunt in our series: pulmonary artery growth was greater, distortion of pulmonary arteries was less commonly seen, and shunt failure occurred less often. Thus, in infants, we believe the modified Blalock-Taussig shunt should be considered a reasonable alternative to the standard Blalock-Taussig shunt.

Blood Vessel Prosthesis↗

PubMed2003

Autoantibodies against modified low-density lipoproteins in coronary artery disease.

OBJECTIVES: To evaluate the importance of different autoantibodies against modified low-density lipoprotein (LDL) in patients with coronary artery disease (CAD). BACKGROUND: Previous studies of autoantibodies against LDL have shown that patients with CAD have increased titers of autoantibodies against LDL modified by copper and malondialdehyde (MDA), whereas there is a lack of information about autoantibody titers against LDL modified by hypochlorite (HOCl). Studies of autoantibodies in relation to severity of atherosclerosis are few and have reached divergent results. Furthermore, no data exist on the relationship between autoantibody titers and prognosis. METHODS: Titers of autoantibodies against copper-, MDA- and HOCl-modified LDL were determined in serum by ELISA. Autoantibody titers in young male survivors of a first myocardial infarction were compared with those of healthy controls and related to coronary angiographic findings and to prognosis during 11 years of follow-up. RESULTS: Patients had higher titers of autoantibodies against LDL modified by copper and MDA than controls. In contrast, no consistent associations were found between autoantibody titers and global severity of coronary atherosclerosis or number and severity of coronary stenoses and prognosis. CONCLUSIONS: The prognostic value of autoantibodies against modified LDL is limited in young postinfarction patients despite the fact that autoantibody titers against copper- and MDA-modified LDL are raised compared with healthy controls. Furthermore, the results indicate that autoantibodies against modified LDL are not protective in later stages of coronary atherosclerosis.

Adult↗

PubMed2003

Modified reperfusion and ischemia-reperfusion injury in human lung transplantation.

OBJECTIVE: Ischemia-reperfusion injury remains a major cause of mortality and morbidity in clinical lung transplantation. Interaction of activated leukocytes with injured graft endothelial cells participates in the development of ischemia-reperfusion injury. We sought to determine if modification of the reperfusate (with depletion of leukocytes and alteration of its composition) would decrease the incidence of ischemia-reperfusion injury in human lung transplantation when compared with whole blood reperfusion in a historical group of patients. METHODS: Between June 1999 and July 2001, 23 adult patients undergoing lung transplantation consented to modified reperfusion. After implantation, a catheter was inserted into the main or individual pulmonary arteries, and modified reperfusate was administered at a pressure less than 20 mm Hg. The modified reperfusate was depleted of leukocytes, supplemented with nitroglycerin, adjusted for pH and calcium level, and enriched with aspartate, glutamate, and dextrose. After 10 minutes of modified reperfusion, the removal of pulmonary artery clamp or weaning of cardiopulmonary bypass was performed per usual protocol. Age- and diagnosis-matched historical patients served as the control group. Ischemia-reperfusion injury was defined as Pao(2)/Fio(2) < 150 with diffuse infiltrate on the radiograph in absence of other causes. RESULTS: There was no difference in donor age or oxygenation indices, recipient age, the number of patients requiring cardiopulmonary bypass, ischemia time, and recipient oxygenation indices between the modified reperfusate group and the control group. However, none of the patients in the modified reperfusate group developed ischemia-reperfusion injury in contrast to 5 patients in the control group (P <.05). The early survival in the modified reperfusate group was 96% versus 81% in the control group (P = NS). CONCLUSION: This study suggests that modification of the reperfusate content decreases the incidence of ischemia-reperfusion injury in human lung transplantation when compared with whole blood reperfusion in a historical group of patients. Modified reperfusate may allow acceptance of marginal lungs and expansion of the donor pool.

Adult↗

PubMed2001

Sorption and cosorption of organic contaminant on surfactant-modified soils.

Three kinds of soils were modified with the cationic surfactants, hexadecyltrimethylammonium (HDTMA) bromide and tetramethylammonium (TMA) bromide to increase their sorptive capabilities. Sorption of chlorobenzene in simulated groundwater by these soils was investigated. HDTMA-modified soil has a higher ability to sorb chlorobenzene from simulated groundwater than unmodified soil. TMA-modified soil did not show the superiority. HDTMA thus can be used to modify soil to improve its sorption capability. Cosorption of chlorobenzene in simulated groundwater in the absence or presence of nitrobenzene and dichloromethane on HDTMA-modified soil was also investigated. Nitrobenzene facilitated sorption of chlorobenzene on all HDTMA-modified soil. Dichloromethane did not influence the sorption of chlorobenzene by HDTMA-modified soil. The results suggest that HDTMA-modified soil is a highly effective sorbent for chlorobenzene and multiple organic compounds did not impede the uptake of chlorobenzene.

Adsorption↗

PubMed2003

Fracture toughness of resin-modified glass ionomer restorative materials: effect of powder/liquid ratio and powder particle size reduction on fracture toughness.

OBJECTIVES: The objectives of the current investigation were two-fold: (1) to examine the effect of different powder/liquid (P/L) ratios on the fracture toughness of commercial resin-modified glass ionomer cement and conventional glass ionomer cement, and (2) to evaluate the effect of powder size reduction on the fracture toughness of experimental resin-modified glass ionomers in order to improve their physical properties. METHODS: The P/L ratios of the glass ionomer and resin-modified glass ionomers were varied from the manufacturer's recommended ratio to 2.0 and 1.0 by weight. The powder particle sizes for the experimental resin-modified glass ionomers tested were 2, 5, 10 and 25 micro m in diameter. Fracture toughness was determined on ring-shaped specimens with a fatigued pre-crack. RESULTS: The fracture toughness of the resin-modified glass ionomers was significantly higher (p<0.005) than that of the glass ionomer and was not greatly influenced by the P/L ratio. For the experimental resin-modified glass ionomers, it was observed that fracture toughness gradually decreased as the powder particle size became finer. SIGNIFICANCE: The resin components in the liquid play an important role in the improvement of the physical properties of the resin-modified glass ionomer. A reduction in the powder particle size of up to 10 micro m, which resulted in a smoother surface, can maintain high fracture toughness. The high fracture toughness values of the resin-modified glass ionomer may be one of the factors contributing to a favorable clinical outcome in high stress-bearing areas.

Analysis of Variance↗

PubMed2001

Balloon angioplasty for obstructed modified systemic-pulmonary artery shunts and pulmonary artery stenoses.

OBJECTIVES: The results of percutaneous balloon angioplasty for obstructed modified Blalock-Taussig (BT) or central shunts and pulmonary artery (PA) stenoses were studied to assess its role as an alternative to second shunt and surgical PA angioplasty. BACKGROUND: Obstruction of a modified shunt and PA stenosis related to the shunt or ductus are not infrequent. A second shunt with or without PA angioplastv is required if the PA size, morphology or age of the patient is suboptimal for definitive surgery. METHODS: From June 1994 to May 1999, balloon angioplasty for obstructed systemic-to-PA shunts was performed in 46 patients, with ages ranging from 1 month to 7.4 years (2.2 +/- 1.9 years). Among the 46 patients, 32 had modified BT shunts, 5 had bilateral shunts, 7 had modified central shunts, and 2 had both modified BT and central shunts. Stenoses were seen in 27 main branch PAs, and interruption was present in three. A concurrent balloon angioplasty was attempted in 28 main branch PAs, but it was performed in only 25 vessels. RESULTS: Balloon dilation for obstructed modified shunts was considered to be effective in 42 patients (91%), while angioplasty for PA stenosis was effective in 14 vessels and not effective in 11 vessels. After balloon dilation angioplastv, oxygen saturation in the aorta increased from 74.4 +/- 4.3% to 80.8 +/- 3.6% (p < 0.01) in these 46 patients. One patient died of pneumonia. Eight patients required an additional modified BT shunt soon after the procedure because of severe stenosis or interruption at main branch PA. After a mean follow-up period of 11.6 +/- 5.4 months, 29 patients underwent a repeated imaging study to evaluate the morphology and size of the PAs. Of these 29 patients, 26 underwent open-heart surgery, with two mortalities. CONCLUSIONS: When a second shunt is under consideration because of obstruction of the modified shunt, balloon angioplasty is a possible alternative procedure. Pulmonary artery stenosis, if present, can be simultaneously dilated.

Angioplasty, Balloon↗

PubMed1995

Mixed-mode retention of peptides on phosphate-modified polybutadiene-coated zirconia.

Zirconia HPLC packing materials were found to be potentially advantageous for large-scale protein separations due to their excellent pH stability and mechanical stability. However, Lewis acid sites on zirconia's surface cause irreversible adsorption of proteins due to their interactions with hard Lewis bases such as the carboxyl groups in proteins. Although the Lewis acid sites can be effectively blocked by adsorbing phosphate ions onto zirconia's surface, proteins and peptides cannot be eluted using a typical reversed-phase mobile phase. In this work, we found that the separation of peptides on a phosphate-modified polybutadiene-coated zirconia (PBD-ZrO2) can be brought about by using a mobile phase containing both an organic modifier and a high concentration of sodium perchlorate. The salt is needed to cancel the Coulombic interactions between the negatively charged stationary phase and the positively charged proteins. To understand the retention mechanism of proteins and peptides on phosphate-modified PBD-ZrO2, this work was aimed at the study of the surface characteristics of the phosphate-modified PBD-ZrO2. We found that the phosphate-modified PBD-ZrO2 phase has both reversed-phase and cation-exchange characteristics under the acidic mobile-phase conditions used for protein and peptide separations. The PBD coating provides hydrophobic moieties, and the phosphate ions adsorbed on zirconia's surface provide cation-exchange sites. Reversed-phase separation of a peptide standard mixture and cation-exchange separation of a cationic peptide standard mixture on the same phosphate-modified PBD-ZrO2 column shows excellent column resolution in both modes. Although mixed-mode stationary phases provide unique selectivity, the secondary equilibrium on phosphate-modified PBD-ZrO2 can cause peak broadening. Applications of the phosphate-modified PBD-ZrO2 to peptide separations are demonstrated here.

Amino Acid Sequence↗

PubMed2002

Low-energy collision-induced dissociation fragmentation analysis of cysteinyl-modified peptides.

The development of methods to chemically modify and isolate cysteinyl-residue-containing peptides (Cys-peptides) for LC-MS/MS analysis has generated considerable interest in the field of proteomics. Methods using isotope-coded affinity tags (ICAT) and (+)-biotinyl-iodoacetamidyl-3,6-dioxaoctanediamine (iodoacetyl-PEO-biotin) employ similar Cys-modifying reagents that contain a thiolate-specific biotin group to modify and isolate Cys-containing peptides in conjunction with immobilized avidin. For these strategies to be effective on a proteome-wide level, the presence of the ICAT or acetyl-PEO-biotin tag should not interfere with the efficiency of induced dissociation in MS/MS experiments or with the identification of the modified Cys-peptides by automated database searching algorithms. We have compared the collision-induced dissociation (CID) fragmentation patterns of peptides labeled with iodoacetyl-PEO-biotin and the ICAT reagent to those of the unmodified peptides. CID of Cys-peptides modified with either reagent resulted in the formation of ions attributed to the modified Cys-peptides as well as those unique to the labeling reagent. As demonstrated by analyzing acetyl-PEO-biotin labeled peptides from ribonuclease A and the ICAT-labeled proteome of Deinococcus radiodurans, the presence of these label-specific product ions provides a useful identifier to discern whether a peptide has been modified with the Cys-specific reagent, especially when a number of peptides analyzed using these methods do not contain a modified Cys residue, and to differentiate identical Cys-peptides labeled with either ICAT-d0 or ICAT-d8.

Animals↗

PubMed2005

Thermodynamic studies on the solvent effects in chromatography on molecularly imprinted polymers. 1. Nature of the organic modifier.

Molecularly imprinted polymers (MIPs) are used as highly enantioselective stationary phases in liquid chromatography. To optimize the binding performance of MIPs, different types of polar modifiers are frequently used. Previous studies have shown that the hydrogen-bonding donor parameter (HBD) of the modifier has a large influence on the binding performance of MIPs in chiral separations. This possibility is addressed in a detailed thermodynamic study of a Fmoc-L-tryptophan (Fmoc-L-Trp) imprinted polymer, eluted with four different polar modifiers, i.e., THF, propan-2-ol, methanol, and acetic acid, which have different HBDs (0.00, 0.33, 0.43, and 0.61, respectively). Adsorption isotherm data for each enantiomer in each of these organic modifiers were acquired by frontal analysis over a 20 000 dynamic concentration range. Nonlinear regression of the isotherm data, along with independent calculation of the affinity energy distributions, identified four different types of binding sites coexisting for the enantiomers on the MIP. The exception was acetic acid, which has the highest HBD. In this case, three types of binding sites only coexist on the MIP. The isotherm parameters obtained from these data show the following: (1) The association energies of the two enantiomers with a given type of sites have a similar magnitude; however, the density of the sites is higher for the template than for its antipode. (2) The nature of the organic modifier has a larger influence on the density of high-energy sites than on the association constant of these sites. (3) The molecular size of the organic modifier has a larger influence on the site density (especially for Fmoc-D-Trp) than does HBD. (4) Using an organic modifier with a higher HBD reduces the enantioselectivity on each site. (5) High-energy sites are more enantioselective than low-energy ones. (6) Using an organic modifier with a high HBD causes a larger reduction in the density of high-energy sites approached by the template molecules.

Acetic Acid↗

PubMed1997

Temperature-responsive liquid chromatography. 2. Effects of hydrophobic groups in N-isopropylacrylamide copolymer-modified silica.

We recently reported the new concept of temperature-responsive liquid chromatography using temperature-responsive poly(N-isopropylacrylamide)-modified surfaces as high-performance liquid chromatography media with aqueous mobile phases. Incorporation of hydrophobic sites is an important factor to improve the efficacy (selectivity and retention) of temperature-responsive chromatography. Toward this goal, we have synthesized semitelechelic copolymers of N-isopropylacrylamide (IPAAm) and butyl methacrylate (BMA) having reactive terminal functional groups using telomerization. The lower critical solution temperatures of the copolymers shift to lower temperatures with increasing hydrophobic BMA content in the poly(IPAAm-co-BMA) relative to that of the IPAAm homopolymer. This temperature-responsive semitelechelic copolymer was grafted to the surface of (aminopropyl)silica through the reaction of activated ester-amine coupling. The polymer-modified silica was used as a column packing material. Separation of a mixture of five steroids having various hydrophobicities was investigated. Retention of steroids on poly(IPAAm-co-BMA)-modified columns is increased with an increase in column temperature. The capacity factors for steroids on the copolymer-modified silica beads was much larger than that on homopolymer PIPAAm-modified columns. The capacity factor for testosterone at 50 degrees C was 33.8 for poly(IPAAm-co-BMA) containing 5 mol% BMA, while that for the PIPAAm homopolymer was 15.0 at the same temperature. The influence of column temperature on steroid retention behavior on copolymer-modified stationary phases was significant compared with the case of homopolymer-modified columns. Furthermore, retention times for steroids increased remarkably with increasing BMA composition. The temperature-responsive elution behavior for the steroids was strongly affected by the hydrophobicity of the grafted polymer chains on silica surfaces. Possible protein separation in temperature-responsive liquid chromatography was explored using insulin chains A and B, and beta-endorphin fragment 1-27. On IBc-3.2-modified silica column, these three peptides were successfully separated at 30 degrees C with 0.5 M NaCl aqueous solution (pH 2.1) as mobile phase. The retention times of these peptides were related to the number of hydrophobic amino acid residues in the peptides. In the proposed chromatography system, elution of target substances is controlled only by a small change in column temperature without any further modification of the aqueous mobile phase.

Acrylamides↗

PubMedDate not supplied

Streptavidin in antibody pretargeting. 3. Comparison of biotin binding and tissue localization of 1,2-cyclohexanedione and succinic anhydride modified recombinant streptavidin.

Recombinant streptavidin (rSAv) is of interest as a carrier of alpha-emitting radionuclides in pretargeting protocols for cancer therapy. Due to the inherently high kidney localization of rSAv, modification of this protein is required before it can be useful in pretargeting. Previous studies (Wilbur, D. S., Hamlin, D. K. et al. (1998) Bioconjugate Chem. 9, 322-330) have shown that succinylation of rSAv using succinic anhydride decreases the kidney localization appreciably. In continuing studies, the biotin binding characteristics and biodistribution in mice of rSAv modified by reaction with succinic anhydride (amine modification) or 1,2-cyclohexanedione (arginine modification) have been compared. Modification of rSAv was conducted using 5-50 mol equiv of succinic anhydride and 60-200 mol equiv of 1,2-cyclohexanedione. Most studies were conducted using rSAv modified with the highest quantities of reagents. Succinylation of rSAv did not alter binding with biotin derivatives, but a small increase in the biotin derivative dissociation rate was noted for arginine-modified rSAv. Amino acid analysis of 1,2-cyclohexanedione-treated rSAv indicated about 40% of the arginine residues, or an average of 1.6 residues per subunit, were modified, whereas none of the lysine residues were modified. IEF analyses showed that the pI of the arginine-modified rSAv was 5.3-6, whereas the pI for the succinylated rSAv was approximately 4. Electrospray mass spectral analyses indicated that one to three conjugates of 1,2-cyclohexanedione, and two to three conjugates of succinic anhydride, were obtained per subunit. Both modification reactions resulted in greatly decreasing the kidney localization of rSAv (normally 20-25% ID/g at 4, 24, and 48 h pi). However, the kidney concentration for the succinylated rSAv continued to decrease (5% ID/g to 1.5% ID/g) from 4 to 48 h pi, whereas the concentration (5% ID/g) remained constant over that period of time for the arginine-modified rSAv. In contrast to this, the liver concentration appeared to be slightly higher (3% ID/g vs 2% ID/g) at the later time points for the succinylated rSAv. When less than 50 mol equiv of succinic anhydride were employed in the modification of rSAv, a correlation between increasing kidney localization with decreasing equivalents reacted was observed. Although the differences in the two modified rSAv are not substantial, succinylated rSAv appears to have more favorable properties for pretargeting studies.

Animals↗

PubMed2002

Heterodimerization of dye-modified cyclodextrins with native cyclodextrins.

The heterodimerization behavior of dye-modified beta-cyclodextrins (1-6) with native cyclodextrins (CDs) was investigated by means of absorption and induced circular dichroism spectroscopy in an aqueous solution. Three types of azo dye-modified beta-CDs (1-3) show different association behaviors, depending on the positional difference and the electronic character of substituent connected to the CD unit in the dye moiety. p-Methyl red-modified beta-CD (1), which has a 4-(dimethylamino)azobenzene moiety connected to the CD unit at the 4' position by an amido linkage, forms an intramolecular self-complex, inserting the dye moiety in its beta-CD cavity. It also associates with the native alpha-CD by inserting the moiety of 1 into the alpha-CD cavity. The association constants for such heterodimerization are 198 M(-1) at pH 1.00 and 305 M(-1) at pH 6.59, which are larger than the association constant of 1 for beta-CD (43 M(-1) at pH 1.00). Methyl red-modified 2, which has the same dye moiety as that for 1 although its substituent position is different from that of 1, does not associate even with alpha-CD due to the stable self-intramolecular complex, in which the dye moiety is deeply included in its own cavity of beta-CD. Alizarin yellow-modified CD (3), which has an azo dye moiety different from that of 1 and 2, caused a slight spectral variation upon addition of alpha-CD, suggesting that the interaction between 3 and alpha-CD is weak. On the other hand, phenolphthalein-modified beta-CD (4), which forms an intermolecular association complex in its higher concentrations, binds with beta-CD with an association constant of 787 M(-1) at pH 10.80, where 4 exists as the dianion monomer in the absence of beta-CD. p-Nitorophenol-modified beta-CDs (5 and 6), each having p-nitorophenol moieties with a different connecting part with an amido and amidophenyl group, respectively, associated with alpha-CD with association constants of 66 and 16 M(-1) for 5 and 6, respectively. The phenyl unit in the connecting part of 6 may prevent the smooth binding with alpha-CD. All these results suggest that the dye-modified CDs, in which the dye part is not tightly included in its CD cavity, associate with the native CD to form heterodimer composed of two different CD units by inserting the dye moiety into the native CD unit. The resulting heterodimers have a cavity that can bind another appending moiety of host molecules. On this basis, more ordered molecular arrays or the supramolecular hereropolymers can be constructed.

Journal Article↗

PubMed2004

Covalent attachment of phospholipid analogous polymers to modify a polymeric membrane surface: a novel approach.

A novel method for the surface modification of a microporous polypropylene membrane by tethering phospholipid analogous polymers (PAPs) is given, which includes the photoinduced graft polymerization of N,N-dimethylaminoethyl methacrylate (DMAEMA) and the ring-opening reaction of grafted poly-(DMAEMA) with 2-alkyloxy-2-oxo-1,3,2-dioxaphospholanes. Five 2-alkyloxy-2-oxo-1,3,2-dioxaphospholanes, containing octyloxy, dodecyloxy, tetradecyloxy, hexadecyloxy, and octadecyloxy groups in the molecular structure, were used to fabricate the PAP-modified polypropylene membranes. The attenuated total reflectance FT-IR spectra of the original, poly(DMAEMA)-grafted, and PAP-modified membranes confirmed the chemical changes on the membrane surface. Scanning electron microscope pictures showed that, compared with the original membrane, the surface porosities ofpoly(DMAEMA)-grafted and PAP-modified membranes were somewhat reduced. Water contact angles measured by the sessile drop method on PAP-modified membranes were slightly lower than that on the original polypropylene membrane, but higher than those on poly(DMAEMA)-grafted membranes with the exception of octyloxy-containing PAP-modified membranes. However, BSA adsorption experiments indicated that the five PAP-modified membranes had a much better protein-resistant property than the original polypropylene membrane and the poly(DMAEMA)-grafted membranes. For hexadecyloxy- and octadecyloxy-containing PAP-modified membranes, almost no protein adsorption was observed when the grafting degree was above 6 wt %. It was also found that the platelet adhesion was remarkably suppressed on the PAP-modified membranes. All these results demonstrate that the described approach is an effective way to improve the surface biocompatibility for polymeric membranes.

Adsorption↗

PubMedDate not supplied

Tagging-via-substrate strategy for probing O-GlcNAc modified proteins.

Identification of proteins bearing a specific post-translational modification would imply functions of the modification. Proteomic analysis of post-translationally modified proteins is usually challenging due to high complexity and wide dynamic range, as well as unavailability of efficient methods to enrich the proteins of interest. Here, we report a strategy for the detection, isolation, and profiling of O-linked N-acetylglucosamine (O-GlcNAc) modified proteins, which involves three steps: metabolic labeling of cells with an unnatural GlcNAc analogue, peracetylated azido-GlcNAc; chemoselective conjugation of azido-GlcNAc modified proteins via the Staudinger ligation, which is specific between phosphine and azide, using a biotinylated phosphine capture reagent; and detection and affinity purification of the resulting conjugated O-GlcNAc modified proteins. Since the approach relies on a tag (azide) in the substrate, we designated it the tagging-via-substrate (TAS) strategy. A similar strategy was used previously for protein farnesylation, phosphorylation, and sumoylation. Using this approach, we were able to specifically label and subsequently detect azido-GlcNAc modified proteins from the cytosolic lysates of HeLa, 3T3, COS-1, and S2 cell lines, suggesting the azido-substrate could be tolerated by the enzymatic systems among these cells from diverse biological species. We isolated azido-GlcNAc modified proteins from the cytosolic extract of S2 cells and identified 10 previously reported and 41 putative O-GlcNAc modified proteins, by nano-HPLC-MS/MS. Our study demonstrates that the TAS approach is a useful tool for the detection and proteomic analysis of O-GlcNAc modified proteins.

Acetylglucosamine↗

PubMed1990

Genetic variation in a modifier system affecting the expression of bare mutant of Drosophila subobscura.

Genetic variation affecting the expression of Bare (Ba), a morphological mutant of Drosophila subobscura that reduces the number of bristles and is located on the O chromosome of this species, is reported. Our results show that O chromosomes extracted from a natural population (El Pedroso, Santiago de Compostela, Spain) show considerable genetic variation in modifier effect upon Ba expression. The amount of modifier variability is dependent on the chromosomal arrangement (OST and O3 + 4 + 7), since modifier variation is higher in OST than in O3 + 4 + 7 chromosomes. Investigations of the genetic architecture of this modifier system on the O chromosome carried out using biometrical methods indicate that a relatively small number of genetic "factors" can explain the differences in modifier effect between a wild O chromosome of high modifier effect and a marker chromosome of low score. In addition, the modifier effects show a non-uniform distribution along the O chromosome and some indication of clustering of modifier "factors" around the major locus (Ba) is also observed.

Analysis of Variance↗

PubMed2002

Immunohistochemical detection of hypochlorite-modified proteins in glomeruli of human membranous glomerulonephritis.

A proposed analogy between atherosclerosis and glomerulosclerosis suggests that factors that contribute to the development of atherosclerosis, ie, oxidatively modified (lipo)proteins, may also participate in glomerular injury. Although the nature of the in vivo oxidants has not been clearly identified, increasing evidence suggested the myeloperoxidase (MPO)-H(2)O(2)-halide system to be responsible for the damage observed in leukocyte-dependent glomerulonephritis. MPO, a heme protein secreted by activated phagocytes, may generate modified/oxidized proteins in vivo via intermediate formation of hypochlorous acid (HOCl)/hypochlorite. HOCl, a reactive oxygen species and powerful oxidant, can convert (lipo)proteins into atherogenic forms in vitro and in vivo. Here we demonstrate the presence of HOCl-modified proteins in glomeruli of patients with membranous glomerulonephritis using monoclonal antibodies that do not cross-react with other oxidative modifications. Immunostaining for HOCl-modified epitopes in human minimal change glomerulopathy revealed glomeruli that were unreactive, although the number of MPO-positive cells/glomerulus was slightly increased in comparison to controls. In contrast to minimal change glomerulopathy, a pronounced infiltration of mononuclear cells/glomerulus in membranoproliferative glomerulonephritis is in line with pronounced staining for HOCl-modified epitopes. Immunostaining was detected in intracapillary cells and immune complex deposits within the glomerular basement membrane. In human membranous glomerulonephritis (Stages I to III), staining for HOCl-modified proteins was localized at the basement membrane and podocytes. Staining of serial sections revealed colocalization of HOCl-modified epitopes and MPO in glomerular peripheral basement membranes. Subsequently, tubulointerstitial staining for HOCl-modified epitopes was observed in foam cells at the border of the cytoplasm and in damaged tubular epithelia in focal advanced chronic lesions. Our results indicate that oxidative modification of the basement membrane structure by phagocyte-derived HOCl may be of importance for glomerular defects. The observed colocalization of HOCl-modified proteins and MPO in podocytes and adjacent basement membranes strengthens the assumption that the MPO-H(2)O(2)-halide system contributes to glomerular dysfunction in patients with membranous glomerulonephritis.

Antibodies, Monoclonal↗

PubMed1974

A kinetic and fluorimetric investigation of papain modified at tryptophan-69 and -177 by N-bromosuccinimide.

A systematic study of the modification of papain (its thiol group protected as a disulphide with mercaptoethanol) by N-bromosuccinimide, showed that 2 molar equiv. modified tryptophan-69 and 4 molar equiv. modified tryptophan-69 and -177. The Michaelis parameters for the catalysed hydrolysis of N-benzyloxycarbonylglycine p-nitrophenyl ester by these modified enzymes were determined. The enzymic activity of the modified enzymes was not seriously impaired, but modification of tryptophan-177 raised the apparent pK(a) of the acidic limb of the pH profile by more than 1 pH unit for both k(cat.) and k(cat.)/K(m). The fluorescence spectra (excitation at 288nm) of the modified enzymes showed that tryptophan-69 contributed about 8% to the fluorescence intensity, whereas tryptophan-177 contributed about 46% at neutral pH. However, the contribution of tryptophan-177 was quenched at low pH and its fluorescence intensity showed sigmoidal pH-dependence, with an apparent pK(a) of 4.2. Histidine-159, which is in close contact with tryptophan-177, is considered to be the residue responsible for the fluorescence quenching. When tryptophan-177 was modified, presumably generating a less hydrophobic micro-environment, the apparent pK(a) determined kinetically was raised to about 5.4. By comparing the Michaelis parameters of native papain, papain modified at tryptophan-69 and papain modified at tryptophan-69 and -177 with N-benzyloxycarbonylglycylglycine amide and N-benzyloxycarbonylglycyltryptophan amide, tryptophan-69 and tryptophan-177 were shown to be structural features of the S(2) and S(1)' subsites respectively.

Amino Acid Sequence↗

PubMed2001

A prospective randomized study comparing volume-standardized modified and conventional ultrafiltration in pediatric cardiac surgery.

BACKGROUND: Modified ultrafiltration has been touted as superior to conventional ultrafiltration for attenuating the consequences of hemodilution after cardiac surgery with cardiopulmonary bypass in children. We conducted a prospective randomized study to test the hypothesis that modified and conventional ultrafiltration have similar clinical effects when a standardized volume of fluid is removed. METHODS: From October 1998 to September 1999, 110 children weighing 15 kg or less (median weight 6.1 kg, median age 6.3 months) undergoing surgery with cardiopulmonary bypass for functionally biventricular congenital heart disease were randomized to conventional (n = 67) or arteriovenous modified ultrafiltration (n = 43) for hemoconcentration. The volume of fluid removed with both methods was standardized as a percentage of effective fluid balance (the sum of prime volume and volume added during cardiopulmonary bypass minus urine output): in patients weighing less than 10 kg, 50% of effective fluid balance was removed, whereas 60% was removed in patients weighing 10 to 15 kg. Hematocrit, hemodynamics, ventricular function, transfusion of blood products, and postoperative resource use were compared between groups. RESULTS: There were no significant differences between groups in age, weight, or duration of cardiopulmonary bypass. The total volume of fluid added in the prime and during bypass was greater in patients undergoing conventional ultrafiltration than in those receiving modified ultrafiltration (205 +/- 123 vs 162 +/- 74 mL/kg; P =.05), although the difference was due primarily to a greater indexed priming volume in patients having conventional ultrafiltration. There was no difference in the percentage of effective fluid balance that was removed in the 2 groups. Accordingly, the volume of ultrafiltrate was greater in patients receiving conventional than modified ultrafiltration (95 +/- 63 vs 68 +/- 28 mL/kg; P =.01). Preoperative and postoperative hematocrit levels were 35.6% +/- 6.6% and 36.3% +/- 5.6% in patients having conventional ultrafiltration and 34.4% +/- 6.7% and 38.7% +/- 7.5% in those having modified ultrafiltration. By repeated-measures analysis of variance, patients receiving modified and conventional ultrafiltration did not differ with respect to hematocrit value (P =.87), mean arterial pressure (P =.85), heart rate (P =.43), or left ventricular shortening fraction (P =.21) from baseline to the postbypass measurements. There were no differences between groups in duration of mechanical ventilation, stay in the intensive care unit, or hospitalization. CONCLUSIONS: When a standardized volume of fluid is removed, hematocrit, hemodynamics, ventricular function, requirement for blood products, and postoperative resource use do not differ between pediatric patients receiving conventional and modified ultrafiltration for hemoconcentration after cardiac surgery.

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