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.

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.

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.

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.

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.

Does screen-detected breast cancer have better survival than symptomatic breast cancer?

OBJECTIVES: Evidence obtained from several randomized control trials suggest that mortality from breast cancer could be reduced by mammographic screening. However, a recent meta-analysis questioned the general acceptance that screening for breast cancer is beneficial. The purpose of the study was to analyze prospectively collected data from our unit and produce overall and comparative 5-year survival rates for screen-detected and symptomatic breast cancer. METHODS: Prospectively collected data on all patients diagnosed with invasive breast cancer between January 1993 and December 1994 (24 months), and monitored until the end of 1999, were collated and analyzed. Five-year survival was estimated and broken down by age at diagnosis, tumour size, grade and nodal status. The overall 5-year survival for women with screen-detected cancers was compared with that for women with symptomatically presenting cancers. RESULTS: Between January 1993 and December 1994, 308 patients with invasive breast cancer were referred to the unit (162 via the breast screening programme and 146 presenting symptomatically). The overall 5-year survival was 85.5% (confidence interval [CI], 80.8-89.1). Small tumour size, low grade and negative nodal status were associated with higher survival rates. Five-year survival of the screen-detected cancer patients (91.7%; CI, 85.8-95.2) was higher than that of patients presenting symptomatically (78.6%; CI, 70.6-84.6; p < 0.001). CONCLUSIONS: These findings suggest that patients with screen-detected breast cancer may have better survival compared to those with symptomatically detected breast cancer. The results support the argument in favour of a beneficial impact of breast screening programmes on patients' survival.

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.

Gilbert's syndrome in patients with gallbladder stones.

Patients with Gilbert's syndrome suffer from an abnormality which makes them jaundiced from time to time. A number also develop gallstones and come to cholecystectomy. If this condition has not been recognized these patients may subsequently run the risk of unnecessary operations on their bile ducts from the mistaken assumption that the intermittent episodes of jaundice which are a feature of the syndrome are due to a stone which has been left behind. Such a case history is reported here.In an attempt to determine how frequently these conditions coexist a prospective study was carried out on patients about to undergo cholecystectomy for stones in the gallbladder. Gilbert's disease was found to be present in 2 of 67 males (3.2% +/- 0.8%) but not in 184 females. Hence it seems that about 1 in every 30 males subjected to cholecystectomy may be expected to have this abnormality.It is suggested that this places an obligation on the clinician to have liver function tests done on at least two occasions preoperatively in male patients with cholelithiasis in an attempt to detect this abnormality and avoid this surgical pitfall.

A critical assessment of fluoroscopy in peroperative cholangiography.

In a prospective study of 248 patients undergoing peroperative cholangiography fluoroscopy improved the quality of radiographs by ensuring the use of a volume of contrast medium appropriate to each patient's duct capacity and the correct positioning of the ducts away from obscuring objects. Failure of contrast medium to enter the duodenum is usually considered to be an indication for choledochotomy. Although this occurred in 35 patients, the use of fluoroscopy and the injection of an anticholinergic drug without further contrast medium showed the choledochoduodenal junction to be normal, making choledochotomy unnecessary, in 28. The low negative duct exploration rate (3 of 56 patients) and the virtual elimination of false positive peroperative postexploratory cholangiograms (1 of 50 patients) provide further evidence of the value of fluoroscopy.

Choledochoscopy? Post-exploratory fluorocholangiography? Or both?

Per-operative, post-exploratory fluorocholangiography and choledochoscopy were evaluated prospectively in 59 consecutive patients undergoing bile duct exploration for calculus disease. Fluorocholangiography showed filling defects in 13 cases and was valuable in localisation of stones, assessment of ampullary patency and visualisation of intrahepatic radicles. Choledochoscopy confirmed stones in nine cases, gave immediate confirmation of four false positive X-rays, enabled removal of five residual stones and biopsy of one benign stricture. This study suggests that fluorocholangiography should be performed routinely after duct exploration. Choledochoscopy is a useful adjunct but need only be performed in patients with abnormal X-ray findings; to confirm residual stones and aid in their removal or to permit biopsy of strictures.

A prospective randomised controlled trial of mezlocillin versus netilmicin in biliary surgery.

A prospective randomised controlled trial of mezlocillin versus netilmicin in 133 patients undergoing biliary surgery at a district hospital is reported. Sixty-four patients received mezlocillin and 69 received netilmicin. The two groups of patients were comparable with regard to age, sex, underlying pathology and operative procedures performed. The incidence of infected bile at operation was 14.2% and both antibiotics were equally effective in reducing postoperative bacteraemia (0.75%) and wound infection (4.5%) to acceptable levels. It is concluded that netilmicin is a more cost-effective antibiotic in biliary surgery than mezlocillin.

Duodenal ulcer perforation: the effect of H2 antagonists?

One hundred and two patients with perforated duodenal ulcers over a 13 year period (1970 to 1982) have been prospectively followed-up at a special gastric clinic. Of the 37 patients with perforation of their acute ulcer, 34 were treated by oversew and three had an initial definitive operation (vagotomy and drainage). The remaining 65 patients presented with perforation of a chronic ulcer and 54 were treated by oversew and 11 underwent definitive surgery--nine had vagotomy and drainage and two had partial gastrectomies. Seven of the 34 patients (20.5%) with acute ulcer perforation treated by simple oversew subsequently required definitive ulcer surgery at a mean 17.5 months after perforation and 31 of the 54 patients (57.4%) with chronic ulcer perforations required definitive surgery at a mean 27.4 months after perforation. The introduction of H2 antagonists in 1977 did not alter the re-operation rate in patients with chronic ulcer perforation managed by oversew. Results of this study provide further evidence in favour of treating patients with perforation of their chronic duodenal ulcer by definitive surgery whenever possible.

Sphincter patency and hepatic BSP uptake after biliary sphincterotomy.

An attempt has been made to assess the patency and function of the choledochoduodenal junction after biliary sphincterotomy by estimating the first arrival time of injected bromsulphalein (BSP) in duodenal aspirate and the effect of morphine on this. Studies on 49 patients 1--12 years after this operation showed the arrival time to be normal in 47, but in 2 cases there was delay, which suggests that stenosis may develop in time in abot 4 per cent of cases. This had not been detected clinically, radiologically, or by liver function testing. Although it did not effect the BSP arrival time in T-tube bile, morphine was found to prolong the duodenal arrival time in control patients and also after sphincterotomy of the type used, indicating that the protective mechanism of the sphincter region may not have been destroyed by this operation. Abnormal BSP retention in plasma has been reported up to 5 years afte supraduodenal exploration of the bile ducts in 30 per cent of cases. Similarly, in the present study after biliary sphincterotomy plasma BSP levels were above normal in 14 of 49 patients, suggesting that this abnormality reflects the original pathological condition rather than the type of operation used to deal with it.

Technique of transduodenal exploration of the common bile duct. Duodenoscopic appearances after biliary sphincterotomy.

A simple technique for performing biliary sphincterotomy is described. From an experience of 150 such operations this has been found to provide satisfactory access for exploring the biliary tree and makes T-tube drainage unnecessary. Duodenoscopic observation up to 9 years after the operation showed that the sphincteric region had healed without stenosis. Sphincteric activity had not been completely abolished.

High fibre diet in symptomatic diverticular disease of the colon.

Over the past decade fibre supplementation has achieved widespread acceptance in the management of symptomatic diverticular disease, although the efficacy of this treatment has been debated. We have conducted a retrospective review of 72 patients admitted to hospital with symptomatic diverticular disease over a ten year period in order to determine whether or not high fibre diet afforded protection against the development of complications, necessity for surgery or persistence of symptoms. Fifty-six patients were treated non-operatively, of these 43 received advice concerning a high fibre diet but only 31 patients complied. The 12 patients who failed to take additional fibre and the 13 patients who never received dietary advice (25 patients) formed the non high fibre group. Those treated with fibre supplementation fared significantly better in developing fewer complications and required less surgery (P less than 0.05). At the time of follow-up review patients on a high fibre diet reported significantly fewer symptoms (P less than 0.05).

The value of HIDA scanning in intestinal fistulae.

Intestinal fistulae have been outlined using the delayed phase of Tc-99m labelled HIDA. The technique involved is simple, safe and easily reproducible. This method has advantages over conventional radiography particularly for the leaking duodenal stump and involves minimal disturbance to the patient.

How should the common bile duct be explored?

An attempt has been made to find which of 3 operations currently in use for exploring the bile ducts gave the best clinical results. For this purpose 3 series of consecutive patients treated at The London Hospital over a 12-year period have been studied retrospectively. The operations used were as follows: (1) conventional supraduodenal choledochotomy; (2) transduodenal biliary sphincterotomy; and (3) a combined approach. The mortality in the supraduodenal series was 4 (4%) of 101 patients and in the transduodenal series 2 (2.4%) of 82 patients. Both routes were used in 26 patients, of whom 2 (7.7%) died. Early complications were commoner after choledochotomy than after biliary sphincterotomy, but when both procedures were combined the incidence was higher still. Late complications were also more frequent after both the supraduodenal and the combined approach, residual or recurrent stones and cholangitis being 6 times more common than after sphincterotomy alone. Postexploratory cholangiography, however, was not used routinly in the supraduodenal series and might conceivably have reduced this factor further, but not below 3. Stenosis occurred in one patient after choledochotomy and in one patient after the combined operation but not after sphincterotomy alone. In this study, therefore, transduodenal biliary sphincterotomy gave the lowest mortality and morbidity. With the combined procedure, however, the mortality and morbidity were much higher than after either method alone.

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.