Shoulder disarticulation: a sequel of vascular injury secondary to a proximal humeral fracture.
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Aged↗
Biomedical subjects
H R Williams
Publications and source records attributed to H R Williams.
CCR5, CXCR4, and CD4 are clustered and closely apposed on microvilli of human macrophages and T cells.
The chemokine receptors CCR5 and CXCR4 act synergistically with CD4 in an ordered multistep mechanism to allow the binding and entry of human immunodeficiency virus type 1 (HIV-1). The efficiency of such a coordinated mechanism depends on the spatial distribution of the participating molecules on the cell surface. Immunoelectron microscopy was performed to address the subcellular localization of the chemokine receptors and CD4 at high resolution. Cells were fixed, cryoprocessed, and frozen; 80-nm cryosections were double labeled with combinations of CCR5, CXCR4, and CD4 antibodies and then stained with immunogold. Surprisingly, CCR5, CXCR4, and CD4 were found predominantly on microvilli and appeared to form homogeneous microclusters in all cell types examined, including macrophages and T cells. Further, while mixed microclusters were not observed, homogeneous microclusters of CD4 and the chemokine receptors were frequently separated by distances less than the diameter of an HIV-1 virion. Such distributions are likely to facilitate cooperative interactions with HIV-1 during virus adsorption to and penetration of human leukocytes and have significant implications for development of therapeutically useful inhibitors of the entry process. Although the mechanism underlying clustering is not understood, clusters were observed in small trans-Golgi vesicles, implying that they were organized shortly after synthesis and well before insertion into the cellular membrane. Chemokine receptors normally act as sensors, detecting concentration gradients of their ligands and thus providing directional information for cellular migration during both normal homeostasis and inflammatory responses. Localization of these sensors on the microvilli should enable more precise monitoring of their environment, improving efficiency of the chemotactic process. Moreover, since selectins, some integrins, and actin are also located on or in the microvillus, this organelle has many of the major elements required for chemotaxis.
Animals↗
Substituted 2-aminopyridines as inhibitors of nitric oxide synthases.
A series of substituted 2-aminopyridines was prepared and evaluated as inhibitors of human nitric oxide synthases (NOS). 4,6-Disubstitution enhanced both potency and specificity for the inducible NOS with the most potent compound having an IC50 of 28 nM.
Aminopyridines↗
Complications of tibial shaft soccer fractures.
A retrospective study of AO type 42 tibial diaphyseal fractures that presented to a teaching hospital over a 54 month period was made to identify the proportion sustained whilst playing soccer, determine their characteristics and report treatment and outcome. Sport accounts for 73/329 (22.1%) of these fractures and soccer 58/73 (79. 5%) of these. All patients were male with mean age of 24.3 years (range 8-48). Fifty-four fractures were closed and 93.1% (54/58) were situated in the middle third or at the junction of the middle and distal thirds of the diaphysis. Fifty-six (96.6%) had simple or wedge patterns and 45 (77.6%) were right sided. Forty-four (76.2%) were treated non-operatively in plaster, 12 (20.3%) by intramedullary nails and two (3.4%) with external fixators. Two patients were lost to follow-up and the remaining 56 fractures united at a mean of 6.5 months. There were 21 complications in 19/56 (33.9%) patients which included 8/56 (14.3%) delayed/non-unions requiring surgery. There was a significantly higher complication rate for operated fractures (p<0.005) but no significant link to AO fracture type. Thus we cannot assume that treatment of these common fractures is without risk, especially if they are treated operatively.
Adolescent↗
Castleman's disease of the erector spinae muscle.
Castleman's disease, angio-follicular lymph node hyperplasia, is a relatively rare benign tumour. It typically arises in the mediastinum. We report a unique case arising in the erector spinae muscle. The case demonstrates the importance of radiological investigation and embolisation prior to obtaining a tissue diagnosis and subsequent surgical excision.
Adult↗
Out-patient follow-up after routine surgery: a questionnaire study.
A questionnaire was sent to all consultant general surgeons and urologists in Wales to assess current practice in out-patient follow-up after surgery for nine commonly performed procedures. A further questionnaire was sent to a random sample of general practitioners in South Glamorgan to assess the possibility of GPs taking on responsibility for post-operative follow-up. This should, therefore, reduce the number of post-operative patients passing through the out-patient department. Of the 58 (77%) consultants who responded, the percentage who routinely followed up patients with at least one post-operative visit was calculated. Of the 33 (66%) GPs who responded the percentage who were prepared to take responsibility for post-operative follow-up was calculated for patients having the same operations. There was agreement and disagreement between consultants and GPs, but not distinct pattern emerged. A larger study must be performed before the out-patient department is rationalized using protocols based on this study.
Aftercare↗
Aggrecan degradation in human cartilage. Evidence for both matrix metalloproteinase and aggrecanase activity in normal, osteoarthritic, and rheumatoid joints.
To examine the activity of matrix metalloproteinases (MMPs) and aggrecanase in control and diseased human articular cartilage, metabolic fragments of aggrecan were detected with monospecific antipeptide antibodies. The distribution and quantity of MMP-generated aggrecan G1 fragments terminating in VDIPEN341 were compared with the distribution of aggrecanase-generated G1 fragments terminating in NITEGE373. Both types of G1 fragments were isolated from osteoarthritic cartilage. The sizes were consistent with a single enzymatic cleavage in the interglobular domain region, with no further proteolytic processing of these fragments. Both neoepitopes were also detected by immunohistochemistry in articular cartilage from patients undergoing joint replacement for osteoarthritis (OA), rheumatoid arthritis (RA), and in cartilage from adults with no known joint disease. In control specimens, the staining intensity for both G1 fragments increased with age, with little staining in cartilage from 22-wk-old fetal samples. There was also an increase with age in the extracted amount of MMP-generated neoepitope in relation to both aggrecan and collagen content, confirming the immunohistochemical results. After the age of 20-30 yr this relationship remained at a steady state. The staining for the MMP-generated epitope was most marked in control cartilage exhibiting histological signs of damage, whereas intense staining for the aggrecanase-generated fragment was often noted in adult cartilage lacking overt histological damage. Intense staining for both neoepitopes appeared in the more severely fibrillated, superficial region of the tissue. Intense immunostaining for both VDIPEN- and NITEGE- neoepitopes was also detected in joint cartilage from patients with OA or RA. Cartilage in these specimens was significantly more degraded and high levels of staining for both epitopes was always seen in areas with extensive cartilage damage. The levels of extracted VDIPEN neoepitope relative to collagen or aggrecan in both OA and RA samples were similar to those seen in age-matched control specimens. Immunostaining for both types of aggrecan fragments was seen surrounding the cells but also further removed in the interterritorial matrix. In some regions of the tissue, both neoepitopes were found while in others only one was detected. Thus, generation and/or turnover of these specific catabolic aggrecan fragments is not necessarily coordinated. Our results are consistent with the presence in both normal and arthritic joint cartilage of proteolytic activity against aggrecan based on both classical MMPs and "aggrecanase."
Adolescent↗
An audit of trauma documentation.
Research into aspects of the care of the injured depends on accurate and complete documentation. Inadequate records make audit almost impossible and are unacceptable medico-legally. An audit was made of the standard of trauma documentation in the Yorkshire region before the introduction of trauma charts in 1992. After it was established that a problem did exist, an audit was made of the standard of documentation before and after introduction of trauma charts in one city teaching hospital. Case notes were inspected for completeness of documentation of respiratory, circulatory and neurological status. Results from 1988-1989 showed that only 39.4 per cent of notes recorded the four parameters under investigation. In 1992-1994 the standard had improved. Without trauma charts documentation increased to 90 per cent, but with trauma charts in primary referrals the result improved to 97 per cent. Only 56 per cent of notes were complete for tertiary referrals when a trauma chart was not used. The standard of documentation in major injuries improved with the use of trauma charts. It is recommended that trauma charts are used routinely for all primary and tertiary referrals of injured patients. Tertiary referral patients should be reassessed fully after hospital transfer and a new trauma chart completed.
Blood Pressure↗
Aggrecanase and metalloproteinase-specific aggrecan neo-epitopes are induced in the articular cartilage of mice with collagen II-induced arthritis.
OBJECTIVE: To analyze the roles of two classes of proteinases, 'aggrecanase', and matrix metalloproteinases (MMPs), in chondrodestruction during murine collagen-induced arthritis (CIA). METHODS: Generation of the 'aggrecanase' neo-epitope (NITEGE373), and the MMP neo-epitope (VDIPEN341) within aggrecan was studied by immunoperoxidase microscopy using specific anti-peptide antibodies in normal and stromelysin-1 (SLN-1) deficient knockout mice with CIA. RESULTS: High levels of NITEGE373 and VDIPEN341 neo-epitopes were observed in foci within CIA paw articular cartilage exhibiting depletion of glycosaminoglycans, in advance of significant cartilage erosion. The highest concentrations of NITEGE373 and VDIPEN341 labeling were observed and often co-distributed in the chondrocyte pericellular matrix, suggesting that stimulated chondrocytes can synthesize and/or activate both enzymes. Other regions of the cartilage frequently exhibited either NITEGE373 or VDIPEN341 labeling, but not both neo-epitopes simultaneously, suggesting that 'aggrecanase' and MMP cleavages of aggrecan may be generated independently. No detectable differences were observed in expression or distribution of either neo-epitope in SLN-1 knockout versus wild-type mice. In addition, in vitro digestion of joint sections with SLN-1 did not alter the expression of cartilage NITEGE373, while markedly increasing VDIPEN341 labeling. Peripheral nerves and brains of naive mice also exhibited intense anti-NITEGE373 labeling. CONCLUSIONS: These data indicate that NITEGE373 and VDIPEN341 aggrecan neo-epitopes are sensitive and specific markers of early joint pathology, and are consistent with the hypothesis that SLN-1 does not have 'aggrecanase' activity, and that 'aggrecanase' is distinct from the MMPs which cleave aggrecan at the MMP site.
Aggrecans↗
Audit of thromboembolic prophylaxis in hip and knee surgery.
An audit of the departmental policy for thromboembolic prophylaxis was undertaken, examining the use of TED stockings, administration of subcutaneous low-dose heparin and inclusion into a multicentre pulmonary embolism prevention (PEP) trial for fractured neck of the femur. The results showed that despite an established unit policy, only 43% of patients undergoing primary hip replacement and 14% undergoing revision replacement received subcutaneous heparin. All patients undergoing primary and revision total knee replacement received subcutaneous heparin, but 75% of these patients received an incorrect dose. Use of TED stockings in patients who had sustained a fractured neck of the femur, ranged from 0% to 70% depending on the type of fixation. Use of subcutaneous heparin in these patients ranged between 0% and 20% and inclusion into the PEP trial from 0% to 20%. The results of this study were presented to the clinicians working in the orthopaedic department and 3 months later the audit cycle was completed by repeating the study. It was found there was a statistically significant improvement in the administration of subcutaneous heparin and in the wearing of TED stockings in the joint arthroplasty group as well as in the inclusion of hip fracture patients into the PEP trial. This study demonstrates that established protocols are of little value unless audited and that completion of the audit cycle is essential. It does not attempt to show that one prophylactic method is better than another.
Anticoagulants↗
Expression and immunoaffinity purification of human inducible nitric-oxide synthase. Inhibition studies with 2-amino-5,6-dihydro-4H-1,3-thiazine.
Recombinant human inducible nitric-oxide synthase (rH-iNOS) was expressed in the baculovirus system and purified by a novel immunoaffinity column. rH-iNOS and its native counterpart from cytokine-stimulated primary hepatocytes exhibited similar molecular mass of 130 kDa on SDS-polyacrylamide gel electrophoresis, recognition by antipeptide antibodies, specific activities, and IC50 values for inhibitors. The active dimeric form exhibited a specific activity range of 114-260 nmol/min/mg at 37 degrees C and contained 1.15 +/- 0.04 mol of calmodulin/monomer. The enzyme exhibited a Soret lambdamax at 396 nm with a shoulder at 460 nm and contained 0. 28-0.64 mol of heme/monomer. Dithionite reduction under CO yielded an absorbance maximum at 446 nm, indicating a P450-type heme. Imidazole induced a type II difference spectrum, reversible by L-Arg. 2-Amino-5,6-dihydro-4H-1,3-thiazine (ADT) was competitive versus L-Arg (Ki = 22.6 +/- 1.9 nM), reversed the type II difference spectrum induced by imidazole (Kd = 17.7 nM), and altered the CO-ferrous absorbance of rH-iNOS. L-Arg did not perturb the CO-ferrous adduct directly, but it partially reversed the ADT-induced absorbance shift, indicating that both bind similarly to the protein but interact differently with the heme.
Chromatography, High Pressure Liquid↗
Cell-mediated catabolism of aggrecan. Evidence that cleavage at the "aggrecanase" site (Glu373-Ala374) is a primary event in proteolysis of the interglobular domain.
A rat chondrosarcoma cell line and primary bovine chondrocytes have been used to study cell-mediated aggrecan catabolism. Addition of 1 microM retinoic acid to chondrosarcoma cultures resulted in aggrecan proteolysis with the release of greater than 90% of the cell layer aggrecan into the medium within 4 days. NH2-terminal sequencing of chondroitin sulfate-substituted catabolic products gave a single major NH2-terminal sequence of ARGNVILTXK, initiating at Ala374. This showed that the proteinase, commonly referred to as "aggrecanase," which cleaves the Glu373-Ala374 bond of the interglobular domain of aggrecan (Sandy, J. D., Neame, P. J., Boynton, R. E., and Flannery, C. R. (1990) J. Biol. Chem. 266, 8683-8685), is active in this cell system. Aggrecan G1 domain, generated by cleavage of the interglobular domain, was also liberated during catabolism and this was characterized with three antipeptide antisera. Anti-CDAGWL was used as a general probe for G1 domain. Anti-FVDIPEN was used to specifically detect G1 domain with COOH terminus of Asn341, the form which is readily generated by cleavage of aggrecan by a wide range of matrix metalloproteinases. Anti-NITEGE antiserum was used to specifically detect G1 domain with COOH terminus of Gln373, the form which is the expected product of "aggrecanase"-mediated cleavage of aggrecan. Western blot analysis indicated that a single form of G1 domain of about 60 kDa was formed. G1 domain of this size reacted with both anti-CDAGWL and anti-NITEGE but not with anti-FVDIPEN. Similar experiments with primary bovine chondrocyte cultures, treated with either retinoic acid or interleukin 1, showed that two forms of catabolic G1 domain, of about 62 and 66 kDa, were formed. Both of these forms reacted on Western blots with anti-CDAGWL and also with anti-NITEGE. It is suggested that cell-mediated catabolism of the aggrecan interglobular domain in these culture systems, whether promoted by retinoic acid or interleukin 1, primarily involves cleavage of the Glu373-Ala374 bond by aggrecanase. The accumulation of G1 domain with a COOH-terminal of Glu373 shows that such aggrecanase-mediated cleavage can occur independent of the cleavage of the Asn341-Phe342 bond by matrix metalloproteinases.
Aggrecans↗
VDIPEN, a metalloproteinase-generated neoepitope, is induced and immunolocalized in articular cartilage during inflammatory arthritis.
The destruction of articular cartilage in immune inflammatory arthritic disease involves the proteolytic degradation of its extracellular matrix. The role of activated matrix metalloproteinases (MMPs) in the chondrodestructive process was studied by identifying a selective cleavage product of aggrecan in murine arthritis models initiated by immunization with either type II collagen or proteoglycan. We conducted semiquantitative immunocytochemical studies of VDIPEN341 using a monospecific polyclonal antibody requiring the free COOH group of the COOH-terminal Asn for epitope detection. This antibody recognizes the aggrecan G1 domain fragment generated by MMP [i.e., stromelysin (SLN) or gelatinase A] cleavage of aggrecan between Asn341-Phe342 but does not recognize intact aggrecan. VDIPEN was undetectable in normal mouse cartilage but was observed in the articular cartilage (AC) of mice with collagen-induced arthritis 10 d after immunization, without histological damage and clinical symptoms. This aggrecan neoepitope was colocalized with high levels of glycosaminoglycans (GAGs) in pericellular matrices of AC chondrocytes but was not seen at the articular surface at this early time. Digestion of normal (VDIPEN negative) mouse paw cryosections with SLN also produced heavy pericellular VDIPEN labeling. Computer-based image analysis showed that the amount of VDIPEN expression increased dramatically by 20 d (70% of the SLN maximum) and was correlated with GAG depletion. Both infiltration of inflammatory cells into the synovial cavity and early AC erosion were also very prominent at this time. Analysis of adjacent sections showed that both induction of VDIPEN and GAG depletion were strikingly codistributed within sites of articular cartilage damage. Similar results occurred in proteoglycan-induced arthritis, a more progressive and chronic model of inflammatory arthritis. These studies demonstrate for the first time the MMP-dependent catabolism of aggrecan at sites of chondrodestruction during inflammatory arthritis.
Amino Acid Sequence↗
Purification and characterization by fast-atom-bombardment mass spectrometry of the polymorphonuclear-leucocyte-elastase-generated A alpha (1-21) fragment of fibrinogen from human blood after incubation with calcium ionophore A23187.
The stimulation of human blood with a Ca2+ ionophore, A23187, leads to activation of polymorphonuclear leucocytes (PMN) with release of small amounts of catalyticaly active elastase, as demonstrated by the formation of a characteristic product, the N-terminal A alpha (1-21) peptide of the Aa subunit of fibrinogen. The identity of the peptide was initially established by radioimmunoassay (r.i.a.) with an antibody raised to A alpha (1-21). We now provide independent confirmation of the formation of A alpha (1-21) by fast-atom-bombardment-m.s. analysis of the fractions separated chromatographically after spiking of plasma samples with peptide labelled with [2H8]Phe at position 8. Identity of the peptides was established on the basis of their chromatographic retention time and by the distinct peaks in the mass spectra of these fractions. The relative intensities of the molecular ions of natural and labelled peptides were measured. On the basis of a comparison of the peaks of similar intensities, the concentration of the natural peptide at the time of spiking was close (79%) to the amount obtained by r.i.a. An additional peptide, des-alanyl-A alpha (2-21), was also seen. The total amount of material measured by r.i.a. could be accounted for by the sum of these two provides. The addition of label and assay by m.s. has provided an independent physical-chemical method for identifying A alpha (1-21) as a characteristic product of PMN elastase release in whole blood, but which is absent in freshly drawn blood.
Amino Acid Sequence↗
Programmed electrical stimulation of the ventricle: an efficient, sensitive, and specific protocol.
A relatively simple and efficient ventricular programmed electrical stimulation (PES) protocol was developed, capable of achieving high degrees of sensitivity and specificity. In a series of 481 subjects, 1, 2, and 3 extrastimuli (ES) were used successively during sinus rhythm and ventricular pacing at two drive cycle lengths, at one or more ventricular sites, together with rapid ventricular pacing, and other maneuvers such as isoproterenol infusion. Three ES were used immediately after two ES at each drive rate, rather than returning after completion of the protocol with two ES. Using the protocol, appropriate arrhythmias could be induced in 88% of all patients with ventricular fibrillation, 84% of all patients with sustained ventricular tachycardia (91% with underlying coronary disease), and 58% of patients with severe nonsustained ventricular tachycardia. There were significant differences in inducibility between patients whose ventricular arrhythmias were due to coronary artery disease and other causes. In contrast, sustained ventricular arrhythmias (all ventricular fibrillation) could be induced in only 5% of a control group of control patients, for a specificity of 95%. The protocol described is simpler and more efficient than those that use exhaustive testing of two ES before going to three ES. Three ES during sinus rhythm proved to be the most productive step, with a higher yield ratio (true: false-positives) than two ES or three ES during pacing, especially at faster rates. Greater efficiency is also achieved by leaving the timing of an extrastimulus just beyond its effective refractory period when an additional extrastimulus is to be added, compared to protocols in which the extrastimulus is moved later in the cycle and then decremented in tandem with the additional extrastimulus. Coupling intervals less than 200 msec produced some false-positives, but fewer overall than intervals greater than or equal to 200 msec, and with yield ratios comparable to other protocol steps. The protocol described meets NASPE standards for ventricular programmed stimulation protocols, and with its demonstrated specificity and relative simplicity and efficiency may be useful as a model for groups not yet committed to an alternative protocol.
Cardiac Pacing, Artificial↗
Laparoscopic cholecystectomy: the first 155 patients.
Laparoscopic cholecystectomy has been attempted in 155 patients. Eight patients required conversion to open cholecystectomy (5%): three owing to lack of clear anatomical detail, two procedures were abandoned because of instrument failure, two had a dilated common bile duct on cholangiography, and one owing to a very adherent mucocele of the gallbladder. Cholangiography was performed selectively; successfully completed in 45 (22% of all patients). Common bile duct stones were found in two patients. All patients were drained, with two bile leaks, one requiring a laparotomy. There was one ductal injury. Most patients (85%) were discharged within 48 h. This initial experience suggests laparoscopic cholecystectomy is the procedure of choice for most patients with symptomatic cholelithiasis.
Adult↗
Direct current shock ablation: quantitative assessment of proarrhythmic effects.
Catheter ablation using direct current (DC) shock has proved invaluable in the management of a variety of tachycardias. However, sporadic reports of fatal arrhythmias following ablation have raised the question of the proarrhythmic potential of DC shock ablation. The present study was undertaken in 45 patients to assess prospectively any proarrhythmia related to DC shock ablation, using matched pre- and postablation Holter monitors and programmed electrical stimulation (PES). Nineteen of these patients had Holter monitors for three successive postablation days to observe trends. There was unmatched data in 11 additional patients. All 56 patients provided prospective follow-up for clinical events. There was no immediate sustained VT/VF at the time of the ablation. Four patients had sustained VT in the first 72 hours after ablation; three episodes were similar to the preablation clinical arrhythmias; one patient had torsades de pointes interrupting bradycardia. Twelve patients met Holter, PES, or clinical criteria for proarrhythmia; none were treated on the basis of these findings. On Holter monitoring, there were significant increases in VPCs/hour and couplets/hour in patients undergoing atrial or atrioventricular junctional ablations; and an increase in couplets after accessory pathway ablations. Increases in these categories were not significant for VT patients; nor were increases in episodes of VT/hour or atrial arrhythmias significant in any group. Patients were followed for 44 +/- 33 months, with an actuarial survival of 95% at 1 year, 88% at 3 years, and 85% at 4 years. There were six deaths during follow-up. Two patients had sudden death: one at 2 months had early evidence of proarrhythmia; the other at 32 months may have represented later myocardia deterioration. One patient died of heart failure at 77 months; and there were three noncardiac deaths. DC shock ablation in humans is much less proarrhythmic than in dogs. The low incidence of clinical proarrhythmic events during prolonged follow-up after discharge resulted in low sensitivity, specificity, and positive predictive values for Holter and PES, although the negative predictive values of these tests were greater than 90%. Only one of 12 patients who met criteria for proarrhythmia in the days immediately following ablation had subsequent clinical events consistent with proarrhythmia. These results may be useful as standards for comparison with results of radiofrequency or other ablation modalities.
Arrhythmias, Cardiac↗
Structure of human neutrophil elastase in complex with a peptide chloromethyl ketone inhibitor at 1.84-A resolution.
Human neutrophil elastase (HNE) has been implicated as a major contributor to tissue destruction in various disease states, including emphysema. The structure of HNE, at neutral pH, in complex with methoxysuccinyl-Ala-Ala-Pro-Ala chloromethyl ketone (MSACK), has been solved and refined to an R factor of 16.4% at 1.84-A resolution. Results are consistent with the currently accepted mechanism of peptide chloromethyl ketone inhibition of serine proteases, in that MSACK cross-links the catalytic residues His-57 and Ser-195. The structure of the HNE-MSACK complex is compared with that of porcine pancreatic elastase in complex with L-647,957, a beta-lactam inhibitor of both elastases. The distribution of positively charged residues on HNE is highly asymmetric and may play a role in its specific association with the underlying negatively charged proteoglycan matrix of the neutrophil granules in which the enzyme is stored.
Amino Acid Chloromethyl Ketones↗