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Biomedical subjects

Y Young

Publications and source records attributed to Y Young.

At least 19 recordsLinked to original sources
PubMed2000

Characterization of Alzheimer's beta -secretase protein BACE. A pepsin family member with unusual properties.

The cerebral deposition of amyloid beta-peptide is an early and critical feature of Alzheimer's disease. Amyloid beta-peptide is released from the amyloid precursor protein by the sequential action of two proteases, beta-secretase and gamma-secretase, and these proteases are prime targets for therapeutic intervention. We have recently cloned a novel aspartic protease, BACE, with all the known properties of beta-secretase. Here we demonstrate that BACE is an N-glycosylated integral membrane protein that undergoes constitutive N-terminal processing in the Golgi apparatus. We have used a secreted Fc fusion-form of BACE (BACE-IgG) that contains the entire ectodomain for a detailed analysis of posttranslational modifications. This molecule starts at Glu(46) and contains four N-glycosylation sites (Asn(153), Asn(172), Asn(223), and Asn(354)). The six Cys residues in the ectodomain form three intramolecular disulfide linkages (Cys(216)-Cys(420), Cys(278)-Cys(443), and Cys(330)-Cys(380)). Despite the conservation of the active site residues and the 30-37% amino acid homology with known aspartic proteases, the disulfide motif is fundamentally different from that of other aspartic proteases. This difference may affect the substrate specificity of the enzyme. Taken together, both the presence of a transmembrane domain and the unusual disulfide bond structure lead us to conclude that BACE is an atypical pepsin family member.

Amino Acid Sequence↗
PubMed2000

A comparison of folding techniques in the chemical synthesis of the epidermal growth factor-like domain in neu differentiation factor alpha/beta.

The 52-residue alpha/beta chimera of the epidermal growth factor-like domain in neu differentiation factor (NDFealpha/beta) has been synthesized and folded to form a three disulfide bridge (Cys182-Cys196, Cys190-Cys210, Cys212-Cys221) containing peptide. We investigated two general strategies for the formation of the intramolecular disulfide bridges including, the single-step approach, which used fully deprotected and reduced peptide, and a sequential approach that relied on orthogonal cysteine protection in which specific pairs are excluded from the first oxidation step. Because there are 15 possible disulfide bridge arrangements in a peptide with six cysteines, the one-step approach may not always provide the desired disulfide pairing. Here, we compare the single-step approach with a systematic evaluation of the sequential approach. We employed the acetamidomethyl group to protect each pair of cysteines involved in disulfide bridges, i.e. Cys182 to Cys196, Cys190 to Cys210 and Cys212 to Cys221. This reduced the number of possible disulfide patterns from 15 to three in the first folding step. We compared the efficiencies of folding for each protected pair using RP-HPLC, mapped the disulfide connectivity of the predominant product and then formed the final disulfide from the partially folded intermediate via 12 oxidation. Only the peptide having the Cys182-Cys196 pair blocked with acetamidomethyl forms the desired disulfide isomer (Cys190-Cys210/Cys212-Cys221) as a single homogeneous product. By optimizing both approaches, as well as other steps in the synthesis, we can now rapidly provide large-scale syntheses of NDFealpha/beta and other novel EGF-like peptides.

Amino Acid Sequence↗
PubMed1999

Disulfide assignment of the C-terminal cysteine knot of agouti-related protein (AGRP) by direct sequencing analysis.

We have assigned the disulfide structure of Md-65 agouti-related protein (Md65-AGRP) using differential reduction and alkylation followed by direct sequencing analysis. The mature human AGRP is a single polypeptide chain of 112 amino acid residues, consisting of an N-terminal acidic region and a unique C-terminal cysteine-rich domain. The C-terminal domain, a 48 amino acid peptide named Md65-AGRP, was expressed in Escherichia coil cells and refolded under different conditions from the mature recombinant protein. The disulfide bonds in the cystine knot structure of Md65-AGRP were partially reduced using tris(2-carboxyethyl) phosphine (TCEP) under acidic conditions, followed by alkylation with N-ethylmaleimide (NEM). The procedure generated several isoforms with varying degrees of NEM alkylation. The multiple forms of Md65-AGRP generated by partial reduction and NEM modification were then completely reduced and carboxymethylated to identify unreactive disulfide bonds. Differentially labeled Md65-AGRP were directly sequenced and analyzed by MALDI mass spectrometry. The results confirmed that Md65-AGRP contained the same disulfide structure as that of Md5-AGRP reported previously [Bures, E. J., Hui, J. O., Young, Y. et al. (1998) Biochemistry 37, 12172-12177].

Agouti-Related Protein↗
PubMed1999

Use of complementary therapies by patients attending musculoskeletal clinics.

Patients with musculoskeletal disorders commonly seek treatment outside orthodox medicine (complementary therapy). In patients attending hospital clinics we investigated the prevalence of such behaviour and the reasons for it. Patients attending rheumatology and orthopaedic clinics who agreed to participate were interviewed on the same day by means of a structured questionnaire in three sections: the first section about demographic characteristics; the second about the nature and duration of the complaint, the length of any treatment and whether the patient was satisfied with conventional treatment; and the third about the use of complementary medicine, the types of therapy that had been considered and the reasoning behind these decisions. The data were examined by univariate and bivariate analysis as well as logistic regression multivariate analysis. 166 patients were interviewed (99% response rate) and the predominant diagnosis was rheumatoid arthritis (22.3%). 109 patients (63%) were satisfied with conventional medical treatment; 63 (38%) had considered the use of complementary therapies, and 47 (28%) had tried such a therapy. 26 of the 47 who had used complementary therapy said they had gained some benefit. Acupuncture, homoeopathy, osteopathy and herbal medicine were the most popular types of treatment to be considered. Patients of female gender (P = 0.009) and patients who had expressed dissatisfaction with current therapies (P = 0.01) were most likely to have considered complementary medicine. These results indicate substantial use of complementary therapy in patients attending musculoskeletal disease clinics. The reasons for dissatisfaction with orthodox treatment deserve further investigation, as does the effectiveness of complementary treatments, which must be demonstrated before they are integrated with orthodox medical practice.

Adolescent↗
PubMed1998

Human leptin receptor. Determination of disulfide structure and N-glycosylation sites of the extracellular domain.

The leptin receptor (OB-R) is a member of the class I cytokine receptor family and mediates the weight regulatory effects of its ligand through interaction with cytoplasmic kinases. The extracellular domain of this receptor is comprised of two immunoglobulin-like and cytokine-receptor homology domains each and type III fibronectin domains. The extracellular domain of human leptin receptor was expressed in and purified from Chinese hamster ovary cells and was found to contain extensive N-glycosylation (approximately 36% of the total protein). The purified protein had a molecular weight of approximately 145,000 and exhibited ligand binding ability as evidenced by formation of ligand-receptor complex, followed by chemical cross-linking. The determined disulfide motif of the soluble leptin receptor contained several distinct cystine knots as well as 10 free cysteines. The N-glycosylation analysis revealed that Asn624 of the WSXWS motif (residues 622-626) within the C-terminal cytokine receptor homology domain was glycosylated, indicating that this region is solvent-exposed. On the other hand, the N-terminal WSXWS motif was not glycosylated.

Amino Acid Sequence↗
PubMed1998

Determination of disulfide structure in agouti-related protein (AGRP) by stepwise reduction and alkylation.

The agouti-related protein gene (Agrp) plays an important role in body weight regulation. The mature human protein is a single polypeptide chain of 112 amino acid residues, consisting of an N-terminal acidic region and a unique C-terminal cysteine-rich domain. The disulfide structure of recombinant human AGRP was determined by chemical methods using partial reduction with tris(2-carboxyethyl)phosphine under acidic conditions, followed by direct alkylation with N-ethylmaleimide or fluorescein-5-maleimide. Partial reduction and alkylation provided several forms of AGRP that were modified in a stepwise fashion. The resulting proteins were characterized by peptide mapping, sequence analysis, and mass spectrometry, showing that AGRP contained a highly reducible disulfide bond, C85-C109, followed by less reactive ones, C90-C97, C74-C88, C67-C82, and C81-C99, respectively. The chemically defined disulfide connectivity of the recombinant human AGRP was homologous to that of omega-agatoxin IVB except for an additional disulfide bond, C85-C109.

Agatoxins↗
PubMed1998

Outcome after hemiarthroplasty for femoral neck fractures in the elderly.

A prospective outcome study was performed on 270 patients, 65 years of age and older, who sustained a femoral neck fracture and underwent hemiarthroplasty. The treatment compared was the use of a noncemented unipolar versus either a cemented or a press fit bipolar prosthesis. The outcome variables assessed included the occurrence of a postoperative complication, length and cost of hospitalization, and function in various quality of life measurements. Patients who underwent bipolar hemiarthroplasty with either a cemented or a press fit prosthesis had better pain relief and function than patients who had a noncemented unipolar prosthesis at a minimum of 24 months after surgery. However, the mean hospitalization cost for patients who had a bipolar prosthesis was $12,290 compared with $8876 for a unipolar prosthesis.

Activities of Daily Living↗
PubMed1997

Propofol neuroprotection in a rat model of ischaemia reperfusion injury.

This study was designed to test the hypothesis that propofol, which possesses antioxidant properties, would produce greater protection than isoflurane in cerebral ischaemia reperfusion injury, at dose levels that produced similar affects on brain electrical activity. Twenty Sprague-Dawley rats were anaesthetized using isoflurane 1.5% in air/oxygen, and mechanically ventilated to a PaCO2 of 4.5 kPa. Following 90 min of middle cerebral artery occlusion using an intraluminal filament, rats were given, in random order, either propofol 1 mg kg-1 min-1 or isoflurane 3% (both of which have been shown to reliably produce EEG burst suppression). After a further 30 min, reperfusion was induced by withdrawing the filament. Animals were killed following 240 min of reperfusion. Rats in the propofol group showed a 21% reduction in mean hemispheric infarct volume when compared with the isoflurane group (P < 0.0001). These data suggest that propofol may act by mechanisms in addition to CMRO2 depression, and provide a basis for further studies of propofol neuroprotection.

Anesthesia↗
PubMed1997

Living related small bowel transplantation: anaesthesia and peri-operative care.

The peri-operative management of the only surviving case, to date, of living-related small bowel transplantation is described. The anaesthetic technique was chosen to optimize hepato-splanchnic blood flow. Peri-operative splanchnic blood flow was measured and alterations in flow with changes in inotropic agents and volume loading monitored. There appears to be a role for the use of dopexamine and aggressive volume loading.

Adult↗
PubMed1997

Effects of desflurane and isoflurane on splanchnic microcirculation during major surgery.

Isoflurane has been considered the agent of choice where preservation of splanchnic blood flow is required. Liver blood flow and the hepatic artery buffer response are maintained better in the presence of isoflurane than with other volatile anaesthetic agents. The effects of desflurane have not been assessed in humans. Therefore, we have compared the effects of isoflurane and desflurane anaesthesia on small bowel and hepatic microcirculatory flow during major surgery using laser Doppler flowmetry in a prospective, randomized, single-blind, crossover study. Patients were allocated randomly to receive desflurane or isoflurane (1 MAC) in oxygen-enriched air. Steady-state jejunal and liver blood flow in segment III were assessed by laser Doppler flowmetry. Volatile anaesthetics were then interchanged, and measurements repeated at steady state. Desflurane anaesthesia at 1 MAC was associated with significantly greater gut blood flow than 1 MAC of isoflurane. These differences could not be explained by systemic haemodynamic differences. The similarity in total hepatic flow between groups implies an intact hepatic artery buffer response with desflurane and isoflurane.

Abdomen↗
PubMed1997

Continuing medical education: the question of evaluation.

This paper examines the question of evaluation which has been largely neglected in the credit-based systems of continuing medical education adopted by the Medical Royal Colleges. These systems are seen to encourage a training model of continuing education and a scientific model of evaluation as measurement. By contrast, humanistic evaluation is interpretative and differs not only in its criteria and methods, but also in its underpinning curricular ideologies and values. This model has closer links with concepts of education and professional practice associated with continuing professional development. Decisions about who should conduct the evaluation, what is to be evaluated, how it should be carried out, and about the goals and purposes of evaluation are outlined, noting that they presuppose an ideological view of the relationship of professional knowledge, values and practice. In a concluding discussion of evaluation and the control of professional knowledge, it is argued that the narrow, professional control of evaluation, buttressed by the quality assurance and monitoring mechanisms of the Colleges, is inappropriate, given the increasingly diverse accountabilities which affect medical professionals.

Education, Medical, Continuing↗
PubMed1997

A longitudinal examination of functional recovery among older people with subcapital hip fractures.

OBJECTIVE: Few studies have examined the time-dependent change in functional recovery along with the factors that affect the change among older hip fracture patients. The focus of this study is to examine the predictors of functional recovery in community-dwelling older people with subcapital fractures using longitudinal data analysis methods. DESIGN: A 1-year prospective study. Information was obtained through structured interviews following surgery and at 2, 6, and 12 months after hospital discharge and from medical chart review. SETTING AND PATIENTS: The sample consisted of 312 community-dwelling older adults, admitted to one of the seven Baltimore area hospitals with a subcapital fracture, who received either internal fixation or hemiarthroplasty. RESULTS: The longitudinal data analysis using mixed-effects regression indicates that time and prefracture physical activities of daily living (PADL) had a significant association with PADL functional recovery over the course of 1 year. Among those 85 years and older, those who were disoriented after surgery had poorer PADL functional recovery over time than those who were not disoriented. Also, recovery in instrumental activities of daily living (IADL) was associated significantly with time, prefracture IADL function, unsteady gait prefracture, hospital length of stay, and discharge to an institution. The type of surgical procedure performed was not associated significantly with either PADL or IADL functional recovery. CONCLUSIONS: The factors that are related to functional recovery in this study shed light on the complexity of the recovery process in hip fracture patients. The rate of recovery in postsurgical PADL and IADL function is not constant over time; postsurgical IADL function among disoriented patients (without dementia) continues to deteriorate over time compared with the nondisoriented group; this difference in trends of deterioration on IADL function is most profound in the oldest-old aged group, those 85 and older. The types of surgical procedure performed was not significantly associated with postsurgical functional recovery.

Activities of Daily Living↗
PubMed1997

Continuing education for medical professionals: a reflective model.

The Royal Colleges and their Faculties have moved continuing professional development up the agenda of doctors in the UK. The low educational value and failure to change professional practice of much continuing medical education has led to criticism of its emphasis on formal, didactic teaching and academic knowledge. The ubiquitous scientific or technical bias in medical education makes questionable assumptions about the nature of professional knowledge, how professionals learn, and the linkage of theory and practice in professional work. Given its narrow conception of professional knowledge, it is hardly surprising that the effectiveness of continuing medical education has proven difficult to evaluate. These points of criticism suggest that a more systematic and coherent approach to continuing education is required. The adoption of the concept of continuing professional development, which draws on learning by reflective practice, marks an important step in this direction. Continuing professional development emphasises self-directed learning, professional self-awareness, learning developed in context, multidisciplinary and multilevel collaboration, the learning needs of individuals and their organisations, and an inquiry-based concept of professionalism. It also involves a widening of accountability to patients, the community, managers and policymakers, and a form of evaluation which is internal, participatory and collaborative rather than external and scientific in character.

Clinical Competence↗
PubMed1996

Glial cell line-derived neurotrophic factor: selective reduction of the intermolecular disulfide linkage and characterization of its disulfide structure.

Glial cell line-derived neurotrophic factor is a protein known to enhance the survival of dopaminergic neurons against several neurotoxins. It has been shown to have therapeutic potential in the treatment of Parkinson's disease and other neurodegenerative diseases. We have determined the inter- and intramolecular disulfide linkages of the dimeric molecule by a combination of direct peptide analysis and peptide analysis after either partial reduction or partial oxidation of the protein. Under an acidic condition, the interchain disulfide bond was selectively cleaved with tris(2-carboxyethyl)phosphine, revealing that Cys101 was involved in the intermolecular disulfide linkage. Three other disulfides, Cys68-Cys131, Cys72-Cys133, and Cys41-Cys102, were identified as intramolecular linkages. The determined disulfide structure is highly homologous to that of transforming growth factor beta 2. Since one intramolecular disulfide points through a ring consisting of eight amino acid residues based on the similarity with transforming growth factor beta 2, the disulfide-linked peptides were not purified by conventional methods. Only the peptides from an N-terminal region (residues -1 to 37) were liberated by proteolytic treatment with trypsin or endoproteinase Lys-C, resulting in a stable cystine-knot protein.

Amino Acid Sequence↗
PubMed1996

Prevention of falls in the elderly.

The purpose of these symposium presentations was to examine the role of physical activity as a means of preventing hip fractures through the prevention of falls. Risk factor identification is necessary to develop preventive strategies. Risk factors related to physical activity and other risk factors for falls were identified. Intervention studies aimed at reducing, preventing or delaying falls were identified and discussed. A literature search from 1976-1994, identified 52 studies examining risk factors for falls, recurrent falls and/or falls resulting in injury. Nine intervention studies were identified with the primary outcome of falls. Physical activity-related risk factors for falls include limitations in general functioning, such as ambulation and mobility problems, difficulty or dependence in activities of daily living, and exposures to the risks of falling as indicated by the nature and frequency of daily activities. Impairments in gait and balance as well as neuromuscular and musculoskeletal impairments frequently underlie changes in physical activity in old age. Reduced activity level may occur as a result of these impairments, leading to further declines in physical functioning and an increased risk of falls. A relatively high level of activity in old age is also associated with risk of falls. Other risk factors for falls, such as cognitive impairment, visual deficits and medication use, may combine with physical activity-related risk factors to increase the risk of falls. Intervention studies directed at nursing home populations did not prevent falls but had other statistically and clinically significant outcomes. Studies among the community dwelling that targeted potential or current risk factors and included an exercise component reported a significant reduction in falls, prevented the onset of new disabilities and reduced baseline risk factors. Prevention of falls and subsequent injuries in the institutionalized population remains a challenge. Further development of interventions for community-dwelling elders that facilitate maintenance of physical activity without unduly increasing the risk of falls is also critical. The potential for maintenance of benefits gained from all fall interventions needs further examination.

Accidental Falls↗