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Biomedical subjects

A L Peel

Publications and source records attributed to A L Peel.

At least 19 recordsLinked to original sources

Specific caspase interactions and amplification are involved in selective neuronal vulnerability in Huntington's disease.

Huntington's disease (HD) is an autosomal dominant progressive neurodegenerative disorder resulting in selective neuronal loss and dysfunction in the striatum and cortex. The molecular pathways leading to the selectivity of neuronal cell death in HD are poorly understood. Proteolytic processing of full-length mutant huntingtin (Htt) and subsequent events may play an important role in the selective neuronal cell death found in this disease. Despite the identification of Htt as a substrate for caspases, it is not known which caspase(s) cleaves Htt in vivo or whether regional expression of caspases contribute to selective neuronal cells loss. Here, we evaluate whether specific caspases are involved in cell death induced by mutant Htt and if this correlates with our recent finding that Htt is cleaved in vivo at the caspase consensus site 552. We find that caspase-2 cleaves Htt selectively at amino acid 552. Further, Htt recruits caspase-2 into an apoptosome-like complex. Binding of caspase-2 to Htt is polyglutamine repeat-length dependent, and therefore may serve as a critical initiation step in HD cell death. This hypothesis is supported by the requirement of caspase-2 for the death of mouse primary striatal cells derived from HD transgenic mice expressing full-length Htt (YAC72). Expression of catalytically inactive (dominant-negative) forms of caspase-2, caspase-7, and to some extent caspase-6, reduced the cell death of YAC72 primary striatal cells, while the catalytically inactive forms of caspase-3, -8, and -9 did not. Histological analysis of post-mortem human brain tissue and YAC72 mice revealed activation of caspases and enhanced caspase-2 immunoreactivity in medium spiny neurons of the striatum and the cortical projection neurons when compared to controls. Further, upregulation of caspase-2 correlates directly with decreased levels of brain-derived neurotrophic factor in the cortex and striatum of 3-month YAC72 transgenic mice and therefore suggests that these changes are early events in HD pathogenesis. These data support the involvement of caspase-2 in the selective neuronal cell death associated with HD in the striatum and cortex.

Animals↗

Double-stranded RNA-dependent protein kinase, PKR, binds preferentially to Huntington's disease (HD) transcripts and is activated in HD tissue.

Fourteen neurological diseases have been associated with the expansion of trinucleotide repeat regions. These diseases have been categorized into those that give rise to the translation of toxic polyglutamine proteins and those that are untranslated. Thus far, compelling evidence has not surfaced for the inclusion of a model in which a common mechanism may participate in the pathobiology of both translated and untranslated trinucleotide diseases. In these studies we show that a double-stranded RNA-binding protein, PKR, which has previously been linked to virally-induced and stress-mediated apoptosis, preferentially binds mutant huntingtin RNA transcripts immobilized on streptavidin columns that have been incubated with human brain extracts. These studies also show, by immunodetection in tissue slices, that PKR is present in its activated form in both human Huntington autopsy material and brain tissue derived from Huntington yeast artificial chromosome transgenic mice. The increased immunolocalization of the activated kinase is more pronounced in areas most affected by the disease and, coupled with the RNA binding results, suggests a role for PKR activation in the disease process.

Animals↗

Cochlear function and transgene expression in the guinea pig cochlea, using adenovirus- and adeno-associated virus-directed gene transfer.

Development of a viral vector that can infect hair cells of the cochlea without producing viral-associated ototoxic effects is crucial for utilizing gene replacement therapy as a treatment for certain forms of hereditary deafness. In the present study, cochlear function was monitored using distortion-product otoacoustic emissions (DPOAEs) in guinea pigs that received infusions of either (E1(-), E3(-)) adenovirus, or adeno-associated virus (AAV), directly into the scala tympani. Replication-deficient (E1(-), E3(-)) adenovirus-directed gene transfer, using the cytomegalovirus (CMV) promoter, drove transgene expression to inner hair cells and pillar cells of the cochlea. AAV transduction was tested with several promoters, such as platelet-derived growth factor (PDGF), neuron-specific enolase (NSE), and elongation factor 1alpha (EF-1alpha) promoters; which drove transgene expression to cochlear blood vessels, nerve fibers, and certain spiral limbus cells, respectively. AAV transgene expression was visualized by green fluorescent protein immunostaining. Immunocytochemistry to heparan sulfate confirmed the absence of proteoglycans in guinea pig hair cells, indicating that the receptor for AAV was not present on these cells. However, the heparan sulfate proteoglycan expression pattern mimicked the AAV transduction pattern. An overall finding was that cochlear function was not altered throughout the infection period using AAV titers as high as 5 x 10(8) IP/infused cochlea. In contrast, cochlear function was severely compromised by 8 days postinfection with adenoviral titers of 5 x 10(8) PFU/infused cochlea, and outer hair cells were eliminated. Thus, cochlear hair cells are amenable to in vivo gene transfer using a replication-deficient (E1(-), E3(-)) adenovirus. However, replication-defective or gutted adenovirus vectors must be employed to overcome the ototoxic effects of (E1(-), E3(-)) adenovirus vectors.

Adenoviridae↗

Surgery in the United Kingdom.

Surgery in the United Kingdom has been practiced for nearly 2000 years. It has evolved as a result of the experiences of warfare and the introduction of the scientific basis of surgery. The influence of the 4 surgical royal colleges in setting standards for training and examinations has ensured that new surgeons are equipped for independent practice as consultants. Responsibility for the National Health Service rests with the government, which determines the number of trainee surgeons in the various surgical specialties. Conflicts between service provision and training are highlighted, as are the pressures on academic institutions to meet the demands of clinical surgery. The government's National Health Service plan for England promises a major expansion in undergraduate places and an increase of 7500 consultants in all specialties by 2004. Time will tell if these changes lead to an improvement in surgical services and a reduction in waiting times.

Education, Medical↗

Adeno-associated virus vectors: activity and applications in the CNS.

Transgenic strategies are useful for functional studies and they may also lead to novel therapies. Controlling transgene expression in defined cell populations over time is increasingly important for both functional and gene therapy experiments. The adeno-associated virus (AAV) vector may provide sufficient spatio-temporal control of gene expression for these purposes. This paper reviews in vivo somatic gene transfer methodology using AAV. Advantageous features of this system include neuronal gene expression that is: (1) efficient; (2) long-lived; and (3) non-toxic. Thus, AAV-mediated gene transfer is a good method for functional genomic research. From characterizing vector activity in the brain using different combinations of promoters and transgenes in the mid to late 1990s, researchers continue to discover novel uses of AAV for both basic and clinical neuroscience.

Animals↗

Long-term actions of vector-derived nerve growth factor or brain-derived neurotrophic factor on choline acetyltransferase and Trk receptor levels in the adult rat basal forebrain.

Trophic factor gene therapy may provide a rational treatment strategy for neurodegenerative disease. Recombinant adeno-associated virus vectors, incorporating a neuron-specific promoter driving bicistronic expression of green fluorescent protein and either nerve growth factor or brain-derived neurotrophic factor, transduced 10,000-15,000 neurons in the medial septum for periods of at least six months. Both cholinergic and non-cholinergic neurons expressed green fluorescent protein. Nerve growth factor and brain-derived neurotrophic factor vectors produced up to 50% increases in immunohistochemical detection of the acetylcholine-synthesizing enzyme in septal neurons ipsilateral to the injection. Increased levels of this enzyme, choline acetyltransferase, persisted for six months with the brain-derived neurotrophic factor vector. The nerve growth factor vector increased Trk receptor immunoreactivity in a volume of brain exceeding that of the transduced cells. Counterstaining for the neuronal marker, NeuN, or Nissl substance did not reveal any vector toxicity at any time-point. It therefore appears that the lasting effects of vector-mediated trophic factor gene transfer will offer a new approach for modulating septal cholinergic transmission and Trk receptor activity.

Animals↗

Diagnostic laparoscopy for suspected appendicitis: only useful in young women.

The clinical diagnosis of acute appendicitis is inaccurate and can be associated with a negative appendicectomy rate in excess of 40%. The latter has an attendant, albeit low, potential for increased morbidity and mortality, particularly related to infection. In this two year prospective audit of 282 patients presenting with the symptoms and signs of acute appendicitis, preliminary diagnostic laparoscopy (PDL) was performed on 145 patients with the remainder (137) proceeding directly to open appendicectomy. In the PDL group, appendicectomy was undertaken when there was clear clinical evidence of inflammation or when the appendix was not visualised. The negative appendicectomy rate was similar in both groups (21.6% vs. 21.2%) with a male predominance in the PDL group. Whereas women of childbearing age obtained most benefit from PDL with avoidance of unnecessary open operation, the results of our audit showed little benefit for men.

Adolescent↗

Neuron-specific transduction in the rat septohippocampal or nigrostriatal pathway by recombinant adeno-associated virus vectors.

Viral vector-mediated gene transfer in brain can provide a means for gene therapy and functional studies. However, robust and persistent transgene expression in specific populations of the adult brain has been difficult to achieve. In an attempt to produce localized and persistent transduction in rat brain, we compared recombinant adeno-associated virus (rAAV) vectors incorporating either the immediate early cytomegalovirus (CMV) promoter or the neuron-specific enolase (NSE) promoter. Transduction in hippocampus resulting from the NSE promoter-containing construct was more efficient and persistent than that resulting from the CMV promoter-containing construct. Most hippocampal cells transduced with the NSE promoter had multipolar neuron morphology. Neurons with glutamatergic morphology were transduced weakly. In order to produce a local supply of neurotrophic factor to cells that degenerate under certain disease and experimental conditions, the NSE promoter was utilized to drive expression of brain-derived neurotrophic factor (BDNF) in medial septum or substantia nigra. In this construct, the NSE promoter drives dicistronic expression of BDNF and an enhanced version of green fluorescent protein (GFP). We estimated 3000-15,000 GFP-positive cells per injection of rAAV into septum or substantia nigra, a transduction ratio of 5-20 infectious virus particles per transduced cell. This frequency may be sufficient for trophic factor gene therapy as well as for investigating specific protein function in "topical (i.e., localized) transgenic" animals produced by rAAV.

Animals↗

Efficient transduction of green fluorescent protein in spinal cord neurons using adeno-associated virus vectors containing cell type-specific promoters.

In this study, we have evaluated the capacity of recombinant adeno-associated virus (rAAV) vectors, containing cell type-specific promoters, to transduce neurons in vivo in the normal adult rat spinal cord. The neuron-specific enolase (NSE) promoter and the platelet-derived growth factor (PDGF) B-chain promoter were used to direct expression of a 'humanized' form of the gene for green fluorescent protein (GFP). Neuron-specific rAAVs were injected into the mid-cervical regions of adult rat spinal cords. At 10-14 days, expression was detected in all animals and persisted for up to 15 weeks. Immunocytochemical and morphological profiles of transduced cells were consistently neuronal, and there was no evidence of transgene expression in glial elements. Transduction efficiencies for the NSE and PDGF rAAVs were estimated at 15 and 45 infectious particles per GFP-positive neuron, respectively, in the absence of detectable adenovirus. This study strongly supports a role for rAAV vectors in CNS gene therapy and lays the groundwork for delivery of more functional genes to spinal cord neurons as a possible way to enhance spinal cord repair following injury.

Animals↗

Differentiation of ionic currents in CNS progenitor cells: dependence upon substrate attachment and epidermal growth factor.

Multipotential progenitor cells grown from central nervous system (CNS) tissues in defined media supplemented with epidermal growth factor (EGF), when attached to a suitable substratum, differentiate to express neural and glial histochemical markers and morphologies. To assess the functional characteristics of such cells, expression of voltage-gated Na+ and K+ currents (INa, IK) was studied by whole-cell patch clamp methods in progenitors raised from postnatal rat forebrain. Undifferentiated cells were acutely dissociated from proliferative "spheres," and differentiated cells were studied 1-25 days after plating spheres onto polylysine/laminin-treated coverslips. INa and IK were detected together in 58%, INa alone in 11%, and IK alone in 19% of differentiated cells recorded with K(+)-containing pipettes. With internal Cs+ (to isolate INa), INa up to 45 pA/pF was observed in some cells within 1 day after plating. I Na ranged up to 150 pA/pF subsequently. Overall, 84% of cells expressed I Na, with an average of 38 pA/pF. INa had fast kinetics, as in neurons, but steadystate inactivation curves were strongly negative, resembling those of glial INa. Inward tail currents sensitive to [K+]out were observed upon repolarization after the 10-ms test pulse with internal Cs+, indicating the expression of K+ channels in 82% of cells. In contrast to the substantial currents observed in differentiating cells, little or no INa or Ik-tail currents were detected in recordings from cells acutely dissociated from spheres. Thus, in the presence of EGF, ionic currents develop early during differentiation induced by attachment to an appropriate substratum. Cells switched from EGF to basic fibroblast growth factor (bFGF) when plated onto coverslips showed greatly reduced proliferation and developed less neuron-like morphologies than cells plated in the presence of EGF. INa was observed in only 53% of bFGF-treated cells, with an average of 9 pA/pF. Thus, in contrast to reports that bFGF promotes neuronal differentiation in some CNS progenitor populations, our EGF-generated postnatal rat CNS progenitors do not develop neuronal characteristics when switched to medium containing bFGF. Thus, differentiated CNS progenitors can express a mix of neuronal and glial molecular, morphological, and electrophysiological properties that can be modified by culture conditions.

Animals↗

Management of non-palpable breast lesions detected mammographically.

A total of 212 consecutive fine-wire localization breast biopsies in 202 patients were performed on impalpable mammographically suspicious lesions. Preoperative fine-needle aspiration cytology was performed on 159 lesions and was valuable in planning the extent of the fine-wire localization biopsy. All 212 target lesions were accurately biopsied during the initial surgery. Overall, 134 lesions were malignant and 78 benign (benign:malignant ratio 1:1.72), with fewer biopsies for benign lesions performed in screened patients (benign:malignant ratio 1:2.43) than in those with symptoms (ratio 1:1). Breast conservation was achieved in 72 patients (80 per cent) with screen-detected in situ or invasive carcinoma and in 19 of 37 presenting via a symptomatic clinic. In 160 of 202 patients (79 per cent) the initial fine-wire localization biopsy was diagnostic and therapeutic.

Biopsy, Needle↗

Serosal imprint cytology in colonic cancer: a simple staging technique.

Accurate staging of colonic cancer is essential in defining the rational use of adjuvant treatments. Recent studies have shown that prognostic accuracy can be significantly improved by recognition of primary tumour extension to the free serosal surface. This study compares the technical results of serosal imprint cytology with the results of histology in assessing serosal involvement. When analysed in terms of the modified Dukes' staging the results of cytology imprints taken from the peritoneum overlying the colonic primary were positive for tumour cells in 4/13 Dukes' B, 7/14 Dukes' C, and 5/9 metastatic cancers. Imprint cytology was positive in 6/7 Dukes' B and C cases with histological serosal invasion and was suspicious in the remaining case. However, a further 5/20 cases without identified invasion on routine histology also had positive cytology. Imprint cytology is an adjunct to routine histology which is easily performed and allows more precise staging of serosal involvement in Dukes' B and C colonic cancers. Final evaluation of this technique requires long-term follow-up of patients.

Colon↗

Gastric cancer: the case for a more selective policy in surgical management.

Over a 5-year period 114 patients with gastric cancer were evaluated. Seventy-seven (68%) of these underwent laparotomy, of which 5A (47%) had a resection performed though only 22 (19%) of these were considered curative. Thirty-seven patients (32%) were not offered surgery because they were aged, had poor cardiorespiratory function, or were thought to have advanced disease based on a combination of clinical (fixed epigastric mass, hepatomegaly, jaundice or ascites), radiological and endoscopic features. Overall 5-year survival was 10.9%, with the patients who had curative and palliative resections having 5-year survivals of 24.4% and 18.2% respectively. Eight of the 12 patients who had palliative gastroenterostomy were not satisfactorily palliated, and 9 patients who had 'open and close' laparotomy fared badly with an operative mortality of 44%; mean survival in these two groups was 3.8 and 3 months respectively. Mean survival in patients treated without operation was 5 months. Unit policy in the management of patients with carcinoma of the stomach has been to resect for cure and palliation whenever possible. However, because so many patients present with advanced disease, the avoidance of inappropriate surgery has been an equal priority. In this context, the wider use of ultrasonography, laparoscopy and perhaps computed tomography (CT) may be of help. In this paper, the experience of these 114 patients is reviewed.

Adenocarcinoma↗

Proposed definitions for the audit of postoperative infection: a discussion paper. Surgical Infection Study Group.

Infection is an important arbiter of success or failure of surgical practice and the incidence of infection is incorporated into all surgical audit systems. If audit is to be of value the outcome end points of clinical practice must be defined. We have attempted to derive clinical definitions suitable for bedside diagnosis of infection and believe that these should bs supported by, but not be dependent upon, the results of laboratory or imaging techniques. The proposed definitions are intended to form the basis for clinical audit and to allow meaningful comparisons to be made on clear clinical criteria.

Bacterial Infections↗

Single dose cefotaxime plus metronidazole versus three dose cefuroxime plus metronidazole as prophylaxis against wound infection in colorectal surgery: multicentre prospective randomised study.

OBJECTIVE: To establish whether a single preoperative dose of cefotaxime plus metronidazole was as effective as a standard three dose regimen of cefuroxime plus metronidazole in preventing wound infection after colorectal surgery. DESIGN: Prospective randomised allocation to one of two prophylactic antibiotic regimens in a parallel group trial. Group sequential analyses of each 250 patients were performed. SETTING: 14 District general and teaching hospitals. PATIENTS: 1018 Adults having colorectal operations were randomised, of whom 943 were evaluated. Demographic features, conditions requiring surgery, and operative procedures were similar in the two groups. Most patients had surgery for carcinoma of the colon or rectum. INTERVENTIONS: Group 1 received cefotaxime 1 g intravenously plus metronidazole 500 mg intravenously preoperatively. Group 2 received cefuroxime 1.5 g intravenously plus metronidazole 500 mg intravenously preoperatively, followed by cefuroxime 750 mg intravenously plus metronidazole 500 mg intravenously eight hours and 16 hours postoperatively. MAIN OUTCOME MEASURES: Development of surgical wound infection (as evidenced by the presence of pus), death, or discharge from hospital. RESULTS: Wound condition was scored on a five point scale on alternate days until discharge or for up to 20 days postoperatively. Wound infection rates were: group 1, 32/453 (7.1%; 95% confidence interval 4.7% to 9.4%); group 2, 33/454 (7.3%; 95% confidence interval 4.9% to 9.6%). Death rates (group 1: 26/470 (5.5%); group 2: 31/471 (6.6%], the incidence of postoperative complications, the median duration of hospital stay (12 days), and antibiotic tolerance were all similar in the two groups. Pooled data from groups 1 and 2 showed that wound infections were more frequent when minor faecal contamination had occurred at operation and when the duration of operation exceeded 90 minutes (greater than 90 min 11.2% of cases; less than 90 min 4.8%) and were associated with an extended hospital stay. CONCLUSIONS: A single preoperative dose of cefotaxime plus metronidazole is an efficacious as a three dose regimen of cefuroxime plus metronidazole in preventing wound infection after colorectal surgery and has practical advantages in eliminating the need for postoperative antibiotics.

Adolescent↗