The role of canonical and non-canonical Hedgehog signaling in tumor progression in a mouse model of small cell lung cancer.

Hedgehog (Hh) signaling regulates cell fate and self-renewal in development and cancer. Canonical Hh signaling is mediated by Hh ligand binding to the receptor Patched (Ptch), which in turn activates Gli-mediated transcription through Smoothened (Smo), the molecular target of the Hh pathway inhibitors used as cancer therapeutics. Small cell lung cancer (SCLC) is a common, aggressive malignancy with universally poor prognosis. Although preclinical studies have shown that Hh inhibitors block the self-renewal capacity of SCLC cells, the lack of activating pathway mutations have cast doubt over the significance of these observations. In particular, the existence of autocrine, ligand-dependent Hh signaling in SCLC has been disputed. In a conditional Tp53;Rb1 mutant mouse model of SCLC, we now demonstrate a requirement for the Hh ligand Sonic Hedgehog (Shh) for the progression of SCLC. Conversely, we show that conditional Shh overexpression activates canonical Hh signaling in SCLC cells, and markedly accelerates tumor progression. When compared to mouse SCLC tumors expressing an activating, ligand-independent Smo mutant, tumors overexpressing Shh exhibited marked chromosomal instability and Smoothened-independent upregulation of Cyclin B1, a putative non-canonical arm of the Hh pathway. In turn, we show that overexpression of Cyclin B1 induces chromosomal instability in mouse embryonic fibroblasts lacking both Tp53 and Rb1. These results provide strong support for an autocrine, ligand-dependent model of Hh signaling in SCLC pathogenesis, and reveal a novel role for non-canonical Hh signaling through the induction of chromosomal instability.

Role of endoplasmic reticulum stress induction by the plant toxin, persin, in overcoming resistance to the apoptotic effects of tamoxifen in human breast cancer cells.

BACKGROUND: Persin is a plant toxin that displays synergistic cytotoxicity with tamoxifen in human breast cancer cell lines. Here, we examined the ability of persin to circumvent tamoxifen resistance and delineated the intracellular signalling pathways involved. METHODS: The induction of apoptosis in tamoxifen-resistant and -sensitive breast cancer cells was measured by flow cytometry following treatment with persin±tamoxifen. Markers of endoplasmic reticulum stress (ERS) were analysed following treatment, and their causal role in mediating persin-induced apoptosis was determined using chemical inhibitors and RNA interference. RESULTS: Cells that were resistant to an apoptotic concentration of tamoxifen maintained an apoptotic response to persin. Persin-induced apoptosis was associated with an increase in markers of ERS, that is, CHOP expression and XBP-1 splicing and was decreased by CHOP siRNA. The CASP-4 inhibitor Z-YVAD-FMK markedly inhibited persin-induced apoptosis in both tamoxifen-sensitive and -resistant cells. CONCLUSION: The cytotoxic effects of persin are CASP-4 dependent and mediated by CHOP-dependent and -independent ERS signalling cascades. Increased ERS signalling contributes to persin-induced reversal of tamoxifen resistance.

Identification of PUMA as an estrogen target gene that mediates the apoptotic response to tamoxifen in human breast cancer cells and predicts patient outcome and tamoxifen responsiveness in breast cancer.

Recognition of the pivotal role of estrogen in the aetiology of breast cancer has led to the development of antiestrogens (AE), such as tamoxifen (TAM) as effective therapies for the treatment and prevention of this disease. However, despite their widespread clinical efficacy, response to AEs is often short-lived, and acquired or innate therapeutic resistance remains a major obstacle in the successful treatment of breast cancer. Thus, delineating the intracellular pathways that mediate the cellular response to estrogen could potentially lead to new, more effective approaches to the treatment of breast cancer, particularly endocrine-resistant disease. Here, we have identified the BCL-2 homology 3 (BH3)-only, pro-apoptotic regulator, PUMA (p53 upregulated modulator of apoptosis) as an estrogen target gene that is acutely downregulated in response to estrogen in breast cancer cell lines, independently of their p53 status. PUMA is transcriptionally upregulated following treatment with TAM, and knock down of PUMA expression in these cells attenuates the apoptotic response to TAM. Furthermore, low PUMA expression in breast carcinomas is significantly associated with breast cancer-specific death (P=0.0014 and P=0.0115, for mRNA and protein, respectively), and worse outcome in TAM-treated patients (mRNA, P=1.49e-05). These findings suggest that the dysregulation of apoptotic signaling pathways such as those executed through PUMA, can significantly impact on both the progression and therapeutic responsiveness of breast cancer. Moreover, they provide a convincing rationale for exploring new therapeutic approaches involving endocrine and non-endocrine therapies that target apoptotic pathways as an effective strategy for tackling endocrine refractory disease.

A procedure-specific systematic review and consensus recommendations for postoperative analgesia following laparoscopic cholecystectomy.

BACKGROUND: Laparoscopic cholecystectomy has advantages over the open procedure for postoperative pain. However, a systematic review of postoperative pain management in this procedure has not been conducted. METHODS: A systematic review was conducted according to the guidelines of the Cochrane Collaboration. Randomized studies examining the effect of medical or surgical interventions on linear pain scores in patients undergoing laparoscopic cholecystectomy were included. Qualitative and quantitative analyses were performed. Recommendations for patient care were derived from review of these data, evidence from other relevant procedures, and clinical practice observations collated by the Delphi method among the authors. RESULTS: Sixty-nine randomized trials were included and 77 reports were excluded. Recommendations are provided for preoperative analgesia, anesthetic and operative techniques, and intraoperative and postoperative analgesia. CONCLUSIONS: A step-up approach to the management of postoperative pain following laparoscopic cholecystectomy is recommended. This approach has been designed to provide adequate analgesia while minimizing exposure to adverse events.

The classification of oesophageal 24 h pH measurements using a Kohonen self-organizing feature map.

Analysis of 24 h oesophageal pH studies can be problematic with many patients asymptomatic during the investigation, despite observations of reflux. The aim of this study was to carry out a cluster analysis of ambulatory pH studies to determine any underlying patterns and classes within the data. The results of 900 24 h pH studies were investigated using the Kohonen self-organizing feature map (SOFM), a neural network that can be used to identify clusters within multidimensional data. The clinical features were presented to the network and the main classes identified. The SOFM-based analysis showed that patients clinically assessed as having symptomatic reflux during the study could be described by four major classifications. The results also showed that the probability of identifying a correlation between symptoms and reflux during an investigation varies from 0.49 to 0.78 for the classes identified. The developed network may be a useful tool in the classification of pH data. The cluster-based technique may offer an alternative to standard statistical techniques for high-dimensional gastrointestinal data and form the basis of an expert system for the automated analysis of pH data.

Navigating and visualizing three-dimensional data sets.

This paper provides an overview of the main techniques being used for three-dimensional (3D) visualization of medical data sets and highlights some of the clinical benefits that can be obtained. One of the major advantages of using a 3D representation is that all of the slice data produced by the latest multislice CT and high gradient MR scanners can be utilized, and then presented to the clinician in an intuitive format. Continued advances in technology mean that high resolution 3D representations of patient specific anatomy can now be routinely obtained and so provide valuable input to diagnosis, planning and navigation tasks. Examples from these areas are presented and illustrated below. Future developments and possibilities are also discussed.

A pH-mucosa area unit of measure to consider morphology of the oesophagus when evaluating oesophagitis.

Gastro-oesophageal pH measurements are routinely carried out to quantify and determine if levels of acid reflux are responsible for symptoms. Although considered the 'gold standard', evidence suggests that pH measurements do not correlate well with the degree of oesophagitis seen during endoscopy. In this study the current measure of pH was critically examined taking into account both the effects of changes in luminal diameter and endoscopy observations. The oesophageal lumen diameter was investigated using a barium swallow for 25 patients presenting with oesophageal disorders. For each subject the widest luminal diameter was measured for a series of five controlled swallows. The results showed that the lumen diameter varied widely from 0.9 to 3.8 cm. An alternative approach to the current measurement of pH was explored. In this approach the exposure not only included the luminal pH and time exposed but also the area of mucosa exposed as a result of differing luminal diameters. Although it is currently not possible to assess the diameter or morphology of the oesophageal lumen during a pH study, the analysis highlighted that the current measure of pH exposure time does not include the area of mucosa exposed. These results may explain, to some extent, the poor correlation between pH measurements and degree of oesophagitis seen during endoscopy.

Intravenous n-acetylcysteine, ascorbic acid and selenium-based anti-oxidant therapy in severe acute pancreatitis.

BACKGROUND: To observe outcome in a cohort of patients with severe acute pancreatitis receiving multiple anti-oxidant therapy. METHODS: An observational study was carried out in 46 consecutive patients with acute pancreatitis fulfilling current Atlanta consensus criteria for severe disease. All patients received multiple anti-oxidant therapy based on intravenous selenium, N-acetylcysteine and ascorbic acid plus beta-carotene and alpha-tocopherol delivered via nasogastric tube. Principal outcomes were the effect of anti-oxidant supplementation on anti-oxidant levels, morbidity and mortality in patients on anti-oxidant therapy, case-control analysis of observed survival compared to predicted survival derived from logistic organ dysfunction score (LODS), logistic regression analysis of factors influencing outcome and side effect profile of anti-oxidant therapy. RESULTS: Paired baseline and post-supplementation data were available for 25 patients and revealed that anti-oxidant supplementation restored vitamin C (P = 0.003) and selenium (P = 0.028) toward normal. In univariate survival analysis, patient survival to discharge was best predicted by admission APACHE-II score with relative risk of death increasing 12.6% for each unit increase (95% CI 6.0% to 19.6%). The mean LODS calculated on admission to hospital was 3.7 (standard error of the mean 4.1) giving a predicted mortality for the cohort of 21%. The observed in-hospital mortality was 43%. CONCLUSIONS: Case-control analyses do not appear to demonstrate any benefit from the multiple anti-oxidant combination of selenium, N-acetylcysteine and ascorbic acid in severe acute pancreatitis.

Effects of a moderate dose of alcohol on simulated laparoscopic surgical performance.

BACKGROUND: In medicine, there is no professional regulation of the drinking of alcohol, nor a body of experimental evidence on which such regulation might be based. Here we report the acute and longer-term ("hangover") effects of a moderate dose of alcohol on performance, as assessed objectively on a laparoscopic surgical simulator. METHODS: In a single-blind, experimental study, medical student subjects were assigned randomly to an alcohol (1.05 mg/kg) or a placebo condition (n = 14 in each). The effects of alcohol on performance on the MIST Virtual Reality surgical simulator were examined 60-90 min and 600-630 min (after a night's sleep) following its ingestion. Measures of the number of errors, time taken, hand movement economy, and excessive use of diathermy were recorded. RESULTS: On each measure, performance was significantly impaired 60-90 min following alcohol ingestion, but there was no hangover effect 600-630 min later, following a night's sleep. This impairment could not be attributed to between-group differences in either predrink performance, expertise or estimated sleep duration during the night preceding the experimental session. CONCLUSIONS: Simulated surgical performance is impaired severely when estimated blood alcohol concentration (BAC) is just above the UK legal limit for driving. These results contribute new, objective and quantitative evidence to the current debate about the use and misuse of alcohol within the medical profession.

Counterfactual thinking about controllable events.

When people think about what might have been, they mentally undo controllable rather than uncontrollable events. We report the results of two experiments in which we examined this controllability effect in counterfactual thinking. The experiments show that the mutability of controllable events is influenced by the perceived appropriateness or inappropriateness of the events. The first experiment shows that people change inappropriate controllable actions more than appropriate controllable ones. The second experiment shows that people mutate inappropriate controllable events whether the outcome is exceptional or normal with respect to intrapersonal habitual norms, and whether the outcome is positive or negative. We discuss the implications for alternative theories of counterfactual thinking.

Learning rate for laparoscopic surgical skills on MIST VR, a virtual reality simulator: quality of human-computer interface.

Acquiring laparoscopic surgical skills involves initial learning of cognitive and motor skills followed by refinement of those skills. The successful use of a virtual reality simulator depends on the quality of the interface for the human-computer interaction and this can be determined by the initial learning rate. MIST VR, a part-task virtual reality laparoscopic simulator, provides objective assessment of psychomotor skills and can generate an overall score for performance, based upon errors made and time taken for six different tasks. This study analysed the rate of early task/instrument/computer familiarization on consecutive scores achieved by surgically experienced and naive individuals. Eleven surgeons, 18 medical students and seven non-medical personnel were tested on the simulator up to ten consecutive times, within a 2-week period. Performance data from every task and repetition were analysed to obtain individual scores of task performance. The calculation of overall score penalized errors far more heavily than total time taken, with high scores indicating poor performance. The surgeon-computer interface generated a rapid and significant early familiarization curve up to the third session on the simulator, with significant reductions in both time taken and total contact errors made. These results suggest that MIST VR represents a high quality interface. Surgeons scored consistently and significantly better than other subjects on all tasks. For surgically naive individuals, it was possible to predict the level of laparoscopic skills performance that would be attained after overcoming initial simulator learning curve, by studying their initial score. Overall scores reflected surgical experience and suggest that the simulator is measuring surgically relevant parameters. MIST VR provides a validated and much needed method for objective assessment of laparoscopic skills, for a variety of surgical disciplines.

Chronic pancreatitis at Manchester, UK. Focus on antioxidant therapy.

The Manchester 'oxidant stress' hypothesis for the development of pancreatitis accommodates published information on both chronic pancreatitis and acute pancreatitis. Oxidant stress, mainly from reactive xenobiotic metabolites, is perceived as the pivotal pre-morbid problem in chronic pancreatitis and, by depleting glutathione, targets the exocytosis mechanism of the pancreatic acinar cell. Inhalation exposure to petrochemical products is identified as an independent risk factor in patients at Manchester Royal Infirmary, where some 50% of patients referred have non-alcoholic disease. This paper describes the development of antioxidant therapy, using supplements of methionine, vitamin C and selenium, and its validation in a placebo-controlled trial, followed by a retrospective cross-sectional study in 94 consecutive patients for an average of 30 months. Antioxidant therapy emerges as a safe and effective medical alternative to surgery for painful chronic pancreatitis.

An investigation into the spatial relationship between complexity and motility within the oesophagus.

Nonlinear analysis techniques have recently been used in the characterization of complex physiological signals seen in pathological disorders such as epilepsy and cardiac fibrillation. In this study a series of controlled swallows from an asymptomatic demonstration group was investigated using oesophageal manometry. The nonlinear measure of complexity, largest Lyapunov exponents and phase portraits were then used to explore the complexity of motility patterns at different points within the oesophagus. Results indicate greater complexity within the region of the striated muscle in the upper oesophagus than that observed within the region of smooth muscle in the lower oesophagus. Phase portraits showed that manometry patterns within the asymptomatic demonstration group could be quite different, highlighting the problems in clinical diagnosis. The characterization of motility disorders associated with complex manometry patterns such as diffuse oesophageal spasm (DOS) and nonspecific motility disorder (NOMD) still represents a diagnostic challenge. The use of nonlinear techniques enabling the quantitative and qualitative measurement of oesophageal complexity is considered in the classification of such disorders.

Technical report: The introduction of a new synchronized oesophageal manometry and digital video-fluoroscopy (fluoromanometry) system into the radiology suite.

Following the development of a new fluoromanometry system enabling synchronous oesophageal manometry and barium swallow video-fluoroscopy, both the equipment and examination method have been successfully introduced into the radiology suite. The application of the system which uses a PC with video capture and a portable manometry recorder is described together with details of its implementation and the examination technique used.

Natural history of adhesional small bowel obstruction: counting the cost.

BACKGROUND: The aim of this study was to identify patients admitted with adhesional obstruction to determine if there was an identifiable pattern to the type of initial operation, the type of treatment used for the obstructive episode and the subsequent need for further treatment. METHODS: Patients with adhesional obstruction were identified retrospectively in a cross-sectional study using ICD codes relating to admissions in the years 1990 to 1996. The case notes were used to assess their outcome. RESULTS: Fifty-nine case notes from a total of 175 identified initially satisfied the inclusion criteria. These patients had a mean age at presentation of 51 (range 16-88) years and had undergone a total of 122 operations. Thirty-one patients (53 per cent) had a single previous operation with a median time to presentation with obstruction of 5.5 years (range 11 days to 34.7 years); 33 patients (56 per cent) were treated conservatively on their first admission. There was no statistically significant difference in the outcome in patients who received either conservative or surgical treatment. The length of stay in patients treated surgically (median 11 (range 2-47) days) was significantly longer than that for those treated conservatively (median 6 (range 1-39) days) (P< 0.001). A flow chart was constructed demonstrating the eventual outcome of the patients in the study, enabling the cost of adhesional obstruction to be calculated. CONCLUSION: This type of approach could be used to assess the potential effect of different treatment strategies for adhesional obstruction.

Progress with cholecystectomy: improving results in England and Wales.

BACKGROUND: The Comparative Audit Service of the Royal College of Surgeons of England studied laparoscopic cholecystectomy in England and Wales during 1990-1991. The follow-on study undertaken during 1994 provides data to assess progress. METHODS: Pro formas were sent to consultant surgeons, requesting data on open and laparoscopic cholecystectomies performed in their units during 1994 with data on mean stay, mortality, complications, and the use of peroperative cholangiography and bile duct exploration. The identity of the consultants was treated confidentially. RESULTS: Data were provided by 110 surgeons on 4823 cholecystectomies (1019 open and 3804 laparoscopic) and outcome was compared with that of 3319 attempted laparoscopic and 8035 open cholecystectomies carried out during 1990-1991. The proportion of cases attempted laparoscopically rose from 27.2 per cent in 1990-1991 to 78.9 per cent in 1994, and conversion to open cholecystectomy rose from 5.3 to 6.7 per cent respectively. During 1994 peroperative cholangiography was undertaken in 22.9 per cent of laparoscopic and 44.6 per cent of open cases. Complication rates were similar in the two study periods, except the number of reported haemorrhagic complications was reduced by 40 per cent and bile duct injuries by fivefold (from 0.33 to 0.07 per cent). CONCLUSION: During 1994 the audit sampled approximately 10 per cent of all cholecystectomies performed in England and Wales. The results suggest progress in surgical techniques compared with findings in 1990-1991.

Synchronous recording and review of oesophageal manometry and video fluoroscopy using a portable manometry recorder and PC with integrated digital video acquisition.

We present the technical details of a new system for the synchronous recording and review of a combined oesophageal manometry and video fluoroscopic barium swallow examination. The system developed uses a portable manometry recorder and personal computer (PC) with an integrated digital video acquisition system. These are controlled using software to enable the real time capture of digital video and manometric data throughout the combined examination. The recorded pressure waveforms can then be synchronously displayed on a screen with the recorded digital video of the fluoroscopic barium swallow. This new tool enables both comparative measurement and detailed analysis of the relationship between visualized bolus transport and pressure measurements. It provides for a deeper understanding and improved clinical assessment of complex motility disorders over those obtained when these two modalities are applied separately. The system is easily incorporated into a clinical radiology suite and it is both user and patient friendly. It uses readily available computer hardware together with multimedia software and is a comparatively economical addition to the radiology suite with the manometry analysis available fulfilling the criteria laid down by the Clinical Associates Group of the British Society of Gastroenterology.

A pilot study of antioxidant intake in patients with cholesterol gallstones.

Whereas macronutrient intake has been extensively investigated in an attempt to unravel the pathogenesis of human cholesterol gallstones, theoretical considerations and animal models suggest that deficits in micronutrient antioxidants may be more relevant. We report a pilot study of this aspect. The plan was to obtain 7-d weighed food inventories over a 6-mo period from equal numbers of patients who had not consciously changed their diets, patients who were on low-fat diets and age- and gender-matched controls. Food tables would be used to derive daily intakes of 16 known antioxidants, essential amino acids, and essential fatty acids. Under-reporting of food intake, a recognized drawback of this dietary method, would be sought retrospectively by reference to a key publication giving minimum cut-off limits for ratios of energy intakes to basal metabolic rates. There were 18 pairs for study. Analysis of data for the 9 pairs involving patients on their normal diets showed no differences in the intakes of energy macronutrients, and cholesterol, but the patients ingested lower amounts of 10 among 16 antioxidants (P < 0.05 for methionine, alpha-tocopherol, manganese, and vitamin D; 0.05 < P < 0.10 for cysteine, beta-carotene, vitamin C, selenium, zinc, and phosphorus). Both subsets of patients ingested lower amounts of linoleic acid (diet unchanged P = 0.009, changed P = 0.026) and several essential amino acids than did matched controls. Institution of a low-fat diet caused the expected fall in intakes of energy and saturated fatty acids such that the deficit in alpha-tocopherol was amplified, but substitution of fruit and vegetables by the patients resulted in a fortuitous increase in vitamin C, beta-carotene, and manganese intake. Retrospective analysis confirmed under-reporting of food intake by all four subsets of subjects but there was no significant difference in the mean ratio of energy intake to estimated basal metabolic rate in the subset of patients who had not consciously altered their diets and the subset of matched controls. Furthermore, the lower daily intake of alpha-tocopherol and linoleic acid by these patients persisted when results were expressed relative to total fat consumption. The results support the hypothesis that insufficiency of dietary antioxidants, particularly alpha-tocopherol, may be germane to human gallstone disease; they also suggest that low intakes of linoleic acid and essential amino acids may be relevant. Because of the small sample sizes, however, these deductions should be regarded as tentative, pending confirmation by biochemical analysis of blood and especially of hepatic bile.