Purchase data: a proxy for safety status.

This study demonstrated the use of purchase data to determine the incidence of sharps injuries in a major tertiary referral hospital in Australia. The incidence rates of injuries per 100,000 items purchased were 2.65 and 12.60 for syringe needles and scalpel blades, respectively. These figures were lower than those reported previously using this method. The incidence rate for injuries with suture needles, which had not been reported previously, was 31.89/100,000 items purchased. Incidence data calculated in this manner may be used in conjunction with purchase cost estimates to inform policy and practices on institutional staff safety measures.

Investigating the relationships between peristaltic contraction and fluid transport in the human colon using Smoothed Particle Hydrodynamics.

Complex relationships exist between gut contractility and the flow of digesta. We propose here a Smoothed Particle Hydrodynamics model coupling the flow of luminal content and wall flexure to help investigate these relationships. The model indicates that a zone of muscular relaxation preceding the contraction is an important element for transport. Low pressures in this zone generate positive thrust for low viscosity content. The viscosity of luminal content controls the localization of the flow and the magnitude of the radial pressure gradient and together with contraction amplitude they control the transport rate. For high viscosity content, high lumen occlusion is required for effective propulsion.

In vitro mechanical evaluation of a limited contact dynamic compression plate and hybrid carpal arthrodesis plate for canine pancarpal arthrodesis.

OBJECTIVE: To compare the mechanical properties of pancarpal arthrodesis (PCA) constructs stabilized at 20° of extension using either a 3.5 mm limited contact dynamic compression plate (LC-DCP) or a 3.5/2.7 mm hybrid plate (HP). METHODS: Seven forelimb pairs were used from dogs of similar size. All soft tissues were removed except for supporting structures of the carpus and proximal metacarpal region. All plates were accurately bent to 20°, and then instrumented with two, 350Ω strain gauges applied at the level of the bend. Constructs were embedded in epoxy moulds then mounted onto a servo-hydraulic testing machine. Specimens were loaded for 10 cycles at 100N, 200N and 300N. Tenth cycle construct compliance (CC), maximum angular deformation (MAD), and peak plate strain (PPS) were compared using two-factor analysis of variance (ANOVA) and Student-Newman-Keuls post-hoc tests (p <0.05). RESULTS: Regardless of load, CC was 29% to 33% smaller in the HP than the LC-DCP group (p <0.03). In each group, the CC significantly increased with increasing loads (p <0.02). Mean MAD was 19% to 22% less in HP than LC-DCP constructs, with significant differences seen at 200N and 300N loads. In both groups, MAD was significantly greater with increasing loads (p <0.02). In addition, PPS was 37% to 43% smaller for HP than LC-DCP. CLINICAL SIGNIFICANCE: The mechanical advantages of the HP over the LC-DCP make it a viable alternative for PCA. Smaller CC, MAD and PSS of the HP may reduce the risk of implant failure and postoperative morbidity following PCA.

Effect of bending direction on the mechanical behaviour of interlocking nail systems.

OBJECTIVES: To compare the mechanical properties of various interlocking nail constructs in medio-lateral (ML) and cranio-caudal (CC) bending. METHODS: Synthetic bone models simulating a severely comminuted tibial fracture were treated with either screwed or bolted, 6 or 8 mm standard interlocking nails (ILN), or an angle-stable ILN (AS-ILN), after which they were then sequentially tested in ML and CC bending. Construct compliance, maximum angular deformation (MaxDef) and slack were statistically compared (p<0.05). RESULTS: The compliance of all constructs was significantly greater in CC than in ML bending. However, due to the presence of a greater slack in the ML plane, standard ILN constructs sustained significantly more deformation in that plane. Maximum deformation of the novel AS-ILN constructs was the smallest of all constructs and consistently occurred without slack regardless of bending direction. CLINICAL SIGNIFICANCE: This study suggested that standard ILN construct overall deformation and acute instability (slack) may be more critical in ML than in CC bending. Conversely, the small MaxDef and the absence of slack in both bending planes seen in novel angle-stable AS-ILN may provide optimal construct stability and in turn may be more conducive to bone healing.

An introduction to indigenous health and culture: the first tier of the Three Tiered Plan.

The objective of the present study was to prepare new doctors with an awareness of cultural and health issues to facilitate positive experiences with indigenous patients. The study incorporated the 1998 intern orientation programs in Queensland public hospitals. The study method included tier one of the Three Tiered Plan, which was implemented and audited. Indigenous liaison officers, directors of clinical training and medical education officers were surveyed prior to this implementation to determine whether any or similar initiatives had been carried out in previous years and/or were planned. Post-implementation feedback from interns was obtained by using questionnaires. Follow-up telephone interviews with the directors of clinical training, medical education officers and indigenous hospital liaison officers detailed the format and content of tier one at each hospital. The results indicate that this active intervention improved the implementation rate of tier one from nine of 19 (47%) Queensland public hospitals in 1997 to 17 (90%) in 1998. The 14 indigenous hospital liaison officers (100%) involved in the intervention perceived it as beneficial. Forty-three (67%) of interns who responded to the survey indicated they had encountered an indigenous patient within the last 2-4 months. The level of knowledge of indigenous health and culture self-reported by interns was between the categories 'enough to get by' and 'inadequate'. In conclusion, it appears that tier one has been successful and is to be a formal component of intern orientations in Queensland public hospitals. Further initiatives in indigenous health and culture targeting medical staff (i.e. tier two and tier three), are needed.

Engineering of a glycosidase Family 7 cellobiohydrolase to more alkaline pH optimum: the pH behaviour of Trichoderma reesei Cel7A and its E223S/ A224H/L225V/T226A/D262G mutant.

The crystal structures of Family 7 glycohydrolases suggest that a histidine residue near the acid/base catalyst could account for the higher pH optimum of the Humicola insolens endoglucanase Cel7B, than the corresponding Trichoderma reesei enzymes. Modelling studies indicated that introduction of histidine at the homologous position in T. reesei Cel7A (Ala(224)) required additional changes to accommodate the bulkier histidine side chain. X-ray crystallography of the catalytic domain of the E223S/A224H/L225V/T226A/D262G mutant reveals that major differences from the wild-type are confined to the mutations themselves. The introduced histidine residue is in plane with its counterpart in H. insolens Cel7B, but is 1.0 A (=0.1 nm) closer to the acid/base Glu(217) residue, with a 3.1 A contact between N(epsilon2) and O(epsilon1). The pH variation of k(cat)/K(m) for 3,4-dinitrophenyl lactoside hydrolysis was accurately bell-shaped for both wild-type and mutant, with pK(1) shifting from 2.22+/-0.03 in the wild-type to 3.19+/-0.03 in the mutant, and pK(2) shifting from 5.99+/-0.02 to 6.78+/-0.02. With this poor substrate, the ionizations probably represent those of the free enzyme. The relative k(cat) for 2-chloro-4-nitrophenyl lactoside showed similar behaviour. The shift in the mutant pH optimum was associated with lower k(cat)/K(m) values for both lactosides and cellobiosides, and a marginally lower stability. However, k(cat) values for cellobiosides are higher for the mutant. This we attribute to reduced non-productive binding in the +1 and +2 subsites; inhibition by cellobiose is certainly relieved in the mutant. The weaker binding of cellobiose is due to the loss of two water-mediated hydrogen bonds.

Endogenous sex hormone levels in postmenopausal women with Alzheimer's disease.

A cross-sectional study examined whether there was a difference in endogenous serum sex hormone levels between community-dwelling postmenopausal women with Alzheimer's disease (AD) and healthy controls. Total morning levels of serum estrone, estradiol, androstenedione, testosterone, and cortisol were measured in 52 nondepressed women with AD and 60 postmenopausal women who were neither depressed nor cognitively impaired. Estradiol was undetectable in 35.7% of cases, but detectable hormone was found in 96-100% of cases otherwise. After adjustment for potential confounds, serum levels were significantly higher for estrone (P = 0.0057) and androstenedione (P = 0.02), but not testosterone (P = 0.086) or estradiol (P = 0.59), in subjects with AD. Sex hormone levels did not correlate with cognitive scores in either group. Although the failure to detect estradiol in a third of cases limits the conclusions that can be drawn for this hormone, the possibility that AD is associated with abnormalities in certain serum sex hormone levels should be considered and warrants further research.

Identification of Asp-130 as the catalytic nucleophile in the main alpha-galactosidase from Phanerochaete chrysosporium, a family 27 glycosyl hydrolase.

Characterization of the complete gene sequence encoding the alpha-galactosidase from Phanerochaete chrysosporium confirms that this enzyme is a member of glycosyl hydrolase family 27 [Henrissat, B., and Bairoch, A. (1996) Biochem. J. 316, 695-696]. This family, together with the family 36 alpha-galactosidases, forms glycosyl hydrolase clan GH-D, a superfamily of alpha-galactosidases, alpha-N-acetylgalactosaminidases, and isomaltodextranases which are likely to share a common catalytic mechanism and structural topology. Identification of the active site catalytic nucleophile was achieved by labeling with the mechanism-based inactivator 2',4', 6'-trinitrophenyl 2-deoxy-2,2-difluoro-alpha-D-lyxo-hexopyranoside; this inactivator was synthesized by anomeric deprotection of the known 1,3,4,6-tetra-O-acetyl-2-deoxy-2, 2-difluoro-D-lyxo-hexopyranoside [McCarter, J. D., Adam, M. J., Braun, C., Namchuk, M., Tull, D., and Withers, S. G. (1993) Carbohydr. Res. 249, 77-90], picrylation with picryl fluoride and 2, 6-di-tert-butylpyridine, and O-deacetylation with methanolic HCl. Enzyme inactivation is a result of the formation of a stable 2-deoxy-2,2-difluoro-beta-D-lyxo-hexopyranosyl-enzyme intermediate. Following peptic digestion, comparative liquid chromatographic/mass spectrometric analysis of inactivated and control enzyme samples served to identify the covalently modified peptide. After purification of the labeled peptide, benzylamine was shown to successfully replace the 2-deoxy-2,2-difluoro-D-lyxo-hexopyranosyl peptidyl ester by aminolysis. The labeled amino acid was identified as Asp-130 of the mature protein by further tandem mass spectrometric analysis of the native and derivatized peptides in combination with Edman degradation analysis. Asp-130 is found within the sequence YLKYDNC, which is highly conserved in all known family 27 glycosyl hydrolases.

A new affinity ligand for the isolation of a single 'feruloyl esterase' (FAE-III) from Aspergillus niger.

8-Aminooctyl 5'-S-coniferyl-5'-deoxy-thio-alpha-L-arabinofuranoside has been synthesised and shown to be a selective affinity ligand for the feruloyl esterase III of Aspergillus niger. The hydrolyses of methyl 5-O-coumaroyl, feruloyl, or sinapoyl alpha-L-arabinofuranosides by this enzyme proceed at comparable rates.

Substrate specificity in glycoside hydrolase family 10. Tyrosine 87 and leucine 314 play a pivotal role in discriminating between glucose and xylose binding in the proximal active site of Pseudomonas cellulosa xylanase 10A.

The Pseudomonas family 10 xylanase, Xyl10A, hydrolyzes beta1, 4-linked xylans but exhibits very low activity against aryl-beta-cellobiosides. The family 10 enzyme, Cex, from Cellulomonas fimi, hydrolyzes aryl-beta-cellobiosides more efficiently than does Xyl10A, and the movements of two residues in the -1 and -2 subsites are implicated in this relaxed substrate specificity (Notenboom, V., Birsan, C., Warren, R. A. J., Withers, S. G., and Rose, D. R. (1998) Biochemistry 37, 4751-4758). The three-dimensional structure of Xyl10A suggests that Tyr-87 reduces the affinity of the enzyme for glucose-derived substrates by steric hindrance with the C6-OH in the -2 subsite of the enzyme. Furthermore, Leu-314 impedes the movement of Trp-313 that is necessary to accommodate glucose-derived substrates in the -1 subsite. We have evaluated the catalytic activities of the mutants Y87A, Y87F, L314A, L314A/Y87F, and W313A of Xyl10A. Mutations to Tyr-87 increased and decreased the catalytic efficiency against 4-nitrophenyl-beta-cellobioside and 4-nitrophenyl-beta-xylobioside, respectively. The L314A mutation caused a 200-fold decrease in 4-nitrophenyl-beta-xylobioside activity but did not significantly reduce 4-nitrophenyl-beta-cellobioside hydrolysis. The mutation L314A/Y87A gave a 6500-fold improvement in the hydrolysis of glucose-derived substrates compared with xylose-derived equivalents. These data show that substantial improvements in the ability of Xyl10A to accommodate the C6-OH of glucose-derived substrates are achieved when steric hindrance is removed.

Hydrolyses of alpha- and beta-cellobiosyl fluorides by Cel6A (cellobiohydrolase II) of Trichoderma reesei and Humicola insolens.

We have measured the hydrolyses of alpha- and beta-cellobiosyl fluorides by the Cel6A [cellobiohydrolase II (CBHII)] enzymes of Humicola insolens and Trichoderma reesei, which have essentially identical crystal structures [Varrot, Hastrup, Schülein and Davies (1999) Biochem. J. 337, 297-304]. The beta-fluoride is hydrolysed according to Michaelis-Menten kinetics by both enzymes. When the approximately 2.0% of beta-fluoride which is an inevitable contaminant in all preparations of the alpha-fluoride is hydrolysed by Cel7A (CBHI) of T. reesei before initial-rate measurements are made, both Cel6A enzymes show a sigmoidal dependence of rate on substrate concentration, as well as activation by cellobiose. These kinetics are consistent with the classic Hehre resynthesis-hydrolysis mechanism for glycosidase-catalysed hydrolysis of the 'wrong' glycosyl fluoride for both enzymes. The Michaelis-Menten kinetics of alpha-cellobiosyl fluoride hydrolysis by the T. reesei enzyme, and its inhibition by cellobiose, previously reported [Konstantinidis, Marsden and Sinnott (1993) Biochem. J. 291, 883-888] are withdrawn. (1)H NMR monitoring of the hydrolysis of alpha-cellobiosyl fluoride by both enzymes reveals that in neither case is alpha-cellobiosyl fluoride released into solution in detectable quantities, but instead it appears to be hydrolysed in the enzyme active site as soon as it is formed.

Combined oestrogen-progestogen replacement therapy does not inhibit low-density lipoprotein oxidation in postmenopausal women.

AIMS: The use of oestrogen containing hormone replacement therapy (HRT) is related to a significantly reduced atherosclerotic cardiovascular risk in postmenopausal women. Oestrogen is thought to be antioxidant and may inhibit low-density lipoprotein (LDL) oxidation in vitro. We investigated the effect of combined oestrogen and progestogen HRT on LDL oxidation in postmenopausal women. METHODS: Eighteen healthy women were given oestrogen/progestogen, and the susceptibility of LDL to oxidation was measured as the level of autoantibody to oxidative modified LDL and the production of conjugated dienes during copper-dependent oxidation after 3 and 6 months HRT. The levels of vitamin E, the major antioxidant in LDL, were also measured. RESULTS: After HRT, the anti-oxidatively modified LDL antibody level remained unchanged [1.58+/-0.16, 0.10 (-0.10, 0.26), and 0.08 (-0.09, 0.19), mean+/-s.d. at baseline, and mean change with 95% confidence intervals for differences at 3 and 6 months, respectively, P>0.05] as did the production of conjugated dienes when determined as lag phase [51.2+/-7.5, -0.3 (-3.9, 3.3), and 1.5 (-3.4, 6.4) min, P>0.05]. The LDL vitamin E content, measured as alpha-tocopherol, was also not altered [2.34+/-0.54, -0.07 (-0.27, 0.13), and -0.07 (-0.33, 0.16) nmol mg(-1) LDL, P>0.05] by treatment. CONCLUSIONS: Combined oestrogen and progestogen therapy for 6 months in postmenopausal women does not protect LDL against oxidation.

Lignocellulose degradation by Phanerochaete chrysosporium: purification and characterization of the main alpha-galactosidase.

The main alpha-galactosidase was purified to homogeneity, in 30% yield, from a solid culture of Phanerochaete chrysosporium on 1 part wheat bran/2 parts thermomechanical softwood pulp. It is a glycosylated tetramer of 50 kDa peptide chains, which gives the N-terminal sequence ADNGLAITPQMG(?W)NT(?W)NHFG(?W)DIS(?W)DTI. It is remarkably stable, with crude extracts losing no activity over 3 h at 80 degrees C, and the purified enzyme retaining its activity over several months at 4 degrees C. The kinetics of hydrolysis at 25 degrees C of various substrates by this retaining enzyme were measured, absolute parameters being obtained by active-site titration with 2',4',6'-trinitrophenyl 2-deoxy-2, 2-difluoro-alpha-D-galactopyranoside. The variation of kcat/Km for 1-naphthyl-alpha-D-galactopyranoside with pH is bell-shaped, with pK1=1.91 and pK2=5.54. The alphaD(V/K) value for p-nitrophenyl-alpha-D-glucopyranoside is 1.031+/-0.007 at the optimal pH of 3.75 and 1.114+/-0.006 at pH7.00, indicating masking of the intrinsic effect at optimal pH. There is no alpha-2H effect on binding galactose [alphaD(Ki)=0.994+/-0.013]. The enzyme hydrolyses p-nitrophenyl beta-L-arabinopyranoside approximately 510 times slower than the galactoside, but has no detectable activity on the alpha-D-glucopyranoside or alpha-D-mannopyranoside. Hydrolysis of alpha-galactosides with poor leaving groups is Michaelian, but that of substrates with good leaving groups exhibits pronounced apparent substrate inhibition, with Kis values similar to Km values. We attribute this to the binding of the second substrate molecule to a beta-galactopyranosyl-enzyme intermediate, forming an E.betaGal. alphaGalX complex which turns over slowly, if at all. 1-Fluoro-alpha-D-galactopyranosyl fluoride, unlike alpha-D-galactopyranosyl fluoride, is a Michaelian substrate, indicating that the effect of 1-fluorine substitution is greater on the first than on the second step of the enzyme reaction.

Implantable centrifugal pump with hybrid magnetic bearings.

Test methods and results of in vitro assessment of a centrifugal pump with a magnetically suspended impeller are provided. In vitro blood tests have been completed with a resulting normalized milligram index of hemolysis (NmIH) of 12.4 +/- 4.1, indicating that hemolysis is not a problem. Hydraulic characterization of the system with water has shown that a nominal pumping condition of 6 L/min at 100 mmHg was met at 2,200 rpm. Maximum clinically usable cardiac output is predicted be 10 L/min. The magnetic bearing supported impeller did not contact the housing and was shown to be stable under a variety of pumping conditions. The driving motor efficiency is 75% at the nominal condition. Finally, a description of the clinical version of the pump under development is provided.

Definition of the substrate specificity of the 'sensing' xylanase of Streptomyces cyaneus using xylooligosaccharide and cellooligosaccharide glycosides of 3,4-dinitrophenol.

The title compounds, (Xylp beta (1-->4))nXylp beta-3,4-DNP (n = 0-4) have been made by selective anomeric deprotection of peracetylated xylose oligosaccharides with hydrazine, followed by formation of the trichloroacetimidate, uncatalysed reaction with 3,4-dinitrophenol, and Zemplén deacetylation. The values of k(cat)/K(m) for 3,4-dinitrophenol release from these substrates by xylanase III of Streptomyces cyaneus, expressed in Escherichia coli, increase with increasing n up to n = 2 and then slightly decrease. Since it is known from previous work that in its normal host, the enzyme is produced constitutively at low levels and excreted, these results suggest that the biological function of the enzyme may be to produce small molecule inducers, predominantly xylotriose, from the non-reducing end of the xylan. Activity on cellooligosaccharide glycosides (Glcp beta (1-->4))nGlcp beta-3,4-DNP (n = 0-3) was detected, at a rate about two-and-a-half orders of magnitude less than that observed on the corresponding xylooligosaccharides, indicating that the enzyme is a true xylanase.

Sudden death in Whipple's disease.

Despite antibiotic therapy, some patients with uncomplicated Whipple's disease die suddenly and inexplicably. We describe one such patient who died following unexplained cardiorespiratory arrest and was found to have chronic active myocarditis related to the causative organism. We postulate myocarditis as a cause of sudden death.

Larger increases in sensitivity to paracatalytic inactivation than in catalytic competence during experimental evolution of the second beta-galactosidase of Escherichia coli.

Second-order rate constants (M-1.s-1) at 25 degrees C and pH 7.5 for inactivation of first-generation (ebga and ebgb), second-generation (ebgab and ebgabcd) and third-generation (ebgabcde) experimental evolvants of the title enzyme by 2',4'-dinitrophenyl 2-deoxy-2-fluoro-beta-D-galactopyranoside are 0.042, 0.30, 10, 24 and 57 respectively. Only partial inactivation is observed, except for ebgabcde. At a single high inactivator concentration, inactivation of the wild-type ebgo is also seen. The changes in sensitivity to the paracatalytic inactivator (over a range of 10(3.3)) are larger than changes in kcat/Km for lactose (over a range of 10(2.7)) or nitrophenyl galactosides (over a range of only 10(1.3)), or changes in degalactosylation rate (over a range of 10(1.7)). These data raise the possibility that evolution in the reverse sense, towards insensitivity to a paracatalytic inactivator with a proportionally lower effect on transformation of substrate, may become a mechanism for the development of bacterial resistance to antibiotics that act by paracatalytic enzyme inactivation.

Catamenial variations in erythrocyte sodium-lithium countertransport and blood pressure.

1. We undertook a temporal study of external sodium-stimulated lithium efflux (sodium-lithium countertransport) in erythrocytes and blood pressure by measuring these two parameters in three phases of the menstrual cycle (menstrual, midcycle and luteal phases) in 22 healthy, non-medicated females with regular menstrual cycles. Plasma oestradiol and progesterone levels were also determined. 2. Sodium-lithium countertransport activity (activity in 140 mmol/l external NaCl) in the midcycle phase (0.176 +/- 0.017 mmol h-1 l-1 of cells) was lower than in the menstrual (0.192 +/- 0.016 mmol h-1 l-1 of cells, P < 0.030) and luteal (0.203 +/- 0.018 mmol h-1 l-1 of cells, P < 0.030) phases. The Vmax of the transporter changed similarly but the K(m) was unaltered. 3. The plasma oestradiol level was 628.9 +/- 39.1 pmol/l in the midcycle phase, higher than in the menstrual (232 +/- 18.5 pmol/l, P < 0.001) and luteal (372.5 +/- 28.1 pmol/l, P < 0.001) phases. The progesterone level was 28.6 +/- 2.1 nmol/l in the luteal phase, and values were lower in the menstrual (2.5 +/- 0.3 nmol/l, P < 0.001) and midcycle (2.8 +/- 0.4 nmol/l, P < 0.001) phases. 4. There was no correlation between plasma oestradiol and sodium-lithium countertransport activity or Vmax during the menstrual cycle, but plasma progesterone was positively correlated with sodium-lithium countertransport activity (r = 0.478, P < 0.025, n = 22) and Vmax (r = 0.551, P < 0.045, n = 14) in the luteal phase. 5. Systolic blood pressure did not change significantly during the menstrual cycle. However, the diastolic pressure showed variation similar to that in sodium-lithium countertransport activity/Vmax, its midcycle value of 66.6 +/- 1.4 mmHg being lower than that in the luteal (71.6 +/- 1.3 mmHg, P < 0.025) and menstrual (70.6 +/- 1.4 mmHg, P < 0.025) phases. 6. We conclude that sodium-lithium countertransport activity exhibits catamenial variation. Therefore we suggest, given this observation, that blood sampling for the assessment of the state of activity of the transport system be standardized in relation to a phase of the menstrual cycle in future studies involving females.