BAG-1 suppresses expression of the key regulatory cytokine transforming growth factor ß (TGF-ß1) in colorectal tumour cells.

As colorectal cancer remains the second highest cause of cancer-related deaths in much of the industrialised world, identifying novel strategies to prevent colorectal tumour development remains an important challenge. BAG-1 is a multi-functional protein, the expression of which is up-regulated at relatively early stages in colorectal tumorigenesis. Importantly, BAG-1 is thought to enhance colorectal tumour progression through promoting tumour cell survival. Here, we report for the first time a novel role for BAG-1, establishing it as a suppressor of transforming growth factor ß (TGF-ß1) expression in colorectal tumour cells. Microarray analysis first highlighted the possibility that BAG-1 may regulate TGF-ß1 expression, a key cytokine in normal colonic tissue homoeostasis. Q-RT-PCR and ELISA demonstrated TGFB1 mRNA and protein expression to be significantly increased when BAG1 levels were reduced by small interfering RNA; additionally, induction of BAG-1L caused suppression of TGFB1 mRNA in colorectal tumour cells. Using reporter and chromatin immunoprecipitation assays, a direct association of BAG-1 with the TGFB1 gene regulatory region was identified. Immunohistochemistry and Weiser fraction data indicated that the levels of BAG-1 and TGF-ß1 are inversely correlated in the normal colonic epithelium in vivo, consistent with a role for BAG-1-mediated repression of TGF-ß1 production. In vitro studies showed that the change in TGF-ß1 production following manipulation of BAG-1 is functionally relevant; through induction of anchorage-independent growth in TGF-ß1-dependent normal rat kidney fibroblasts and regulation of SMAD2 phosphorylation in TGF-ß1-sensitive adenoma cells. Taken together, this study identifies the anti-apoptotic protein BAG-1 as a suppressor of the inhibitory growth factor TGF-ß1, suggesting that high expression of BAG-1 can impact on a number of the hallmarks of cancer, of potential importance in promoting the early stages of colorectal tumorigenesis. Establishing BAG-1 as a repressor of TGF-ß1 has important biological implications, and highlights a new role for BAG-1 in colorectal tumorigenesis.

The power of words: sources of anxiety in patients undergoing local anaesthetic plastic surgery.

INTRODUCTION: With local or regional anaesthesia being employed for more as well as more complex surgical procedures, an increasing number of patients remain fully conscious during their operation. This is generally perceived as being advantageous to the patient as less time is spent in hospital and the side effects of general anaesthesia are avoided. However, there is no direct measure of the patient experience during 'awake surgery', in particular of which aspects of the process may be distressing. METHODS: Seventy patients undergoing day case plastic surgery under local anaesthesia were asked to complete a short questionnaire immediately following their operation. This was designed to identify specific factors likely to either increase or reduce anxiety during surgery. The questionnaire was initially validated on a pilot group of ten patients. RESULTS: Unsurprisingly, painful stimuli such as injections were identified as potential stressors. More interestingly, the data highlighted that some commonly used surgical terms such as 'knife' and 'scalpel' provoke considerable anxiety in the conscious patient. This varied according to age and sex with younger and female patients being most vulnerable. Other events identified as potential stressors, such as casual conversations and movements among theatre staff, were actually shown to be non-stressful and, in some cases, stress relieving. CONCLUSIONS: Technical jargon used by surgical staff can elevate anxiety levels among patients who are awake for their operation. Careful consideration of the words we use may reduce this, particularly in female patients.

BAG-1 interacts with the p50-p50 homodimeric NF-κB complex: implications for colorectal carcinogenesis.

Understanding the mechanisms that promote aberrant tumour cell survival is critical for the determination of novel strategies to combat colorectal cancer (CRC). We have recently shown that the anti-apoptotic protein BAG-1, highly expressed in pre-malignant and CRC tissue, can potentiate cell survival through regulating NF-κB transcriptional activity. In this study, we identify a novel complex between BAG-1 and the p50-p50 NF-κB homodimers, implicating BAG-1 as a co-regulator of an atypical NF-κB pathway. Importantly, the BAG-1-p50 complex was detected at gene regulatory sequences including the epidermal growth factor receptor (EGFR) and COX-2 (PTGS2) genes. Suppression of BAG-1 expression using small interfering RNA was shown to increase EGFR and suppress COX-2 expression in CRC cells. Furthermore, mouse embryonic fibroblasts derived from the NF-κB1 (p105/p50) knock-out mouse were used to demonstrate that p50 expression was required for BAG-1 to suppress EGFR expression. This was shown to be functionally relevant as attenuation of BAG-1 expression increased ligand activated phosphorylation of EGFR in CRC cells. In summary, this paper identifies a novel role for BAG-1 in modulating gene expression through interaction with the p50-p50 NF-κB complexes. Data presented led us to propose that BAG-1 can act as a selective regulator of p50-p50 NF-κB responsive genes in colorectal tumour cells, potentially important for the promotion of cell survival in the context of the fluctuating tumour microenvironment. As BAG-1 expression is increased in the developing adenoma through to metastatic lesions, understanding the function of the BAG-1-p50 NF-κB complexes may aid in identifying strategies for both the prevention and treatment of CRC.

Urinary point-of-care test for smoking in the pre-operative assessment of patients undergoing elective plastic surgery.

Self-reported information about smoking habit and cigarette consumption can be inaccurate and subject to bias in the clinical setting. Accurate assessment of a given smoking history at point-of-care is valuable. We describe the use of a comprehensive smoking questionnaire and the use of a disposable biomarker test to verify and quantify the exposure to tobacco smoke. This point-of-care test (SmokeScreen) is a 6-min, easy-to-use urine test that measures nicotine and its breakdown products. One hundred consecutive patients attending plastic surgery pre-assessment clinic filled in the questionnaire and gave a consented urine sample. Qualitative and semi-quantitative assessment of tobacco consumption was observed by a simple sample colour change set against a standardised colorimetric chart for nicotine metabolite containing urine. The questionnaire self-reported smoking prevalence was 30% with 98% test specificity. The cotinine validated smoking prevalence was 54% with a 26% self-denial rate. Half the patients (n = 15) who admitted smoking on the questionnaire underreported the amount they smoked daily, as quantified by biochemical measurement. Objective biochemical assessment shows that 26% of self-reporting non-smokers via self-completed questionnaire studies are actual smokers attending this pre-assessment clinic. When patients did report smoking there was consistent underreporting of cigarette consumption.

Conflicts in the treatment of chronic ulcers in drug addicts--case series and discussion.

Vascular access in intravenous drug abusers may become compromised by the repeated injection of toxic substances. In such circumstances abusers are driven by their addiction to seek alternative routes of drug delivery. We report a series of individuals with chronic ulcers, which were cultivated and maintained for the administration of heroin. We advise that practitioners should be wary of granulation tissue being promoted in this way and suggest that the patients' desire for wound healing may be overridden by their addiction and that successful treatment of these wounds is reliant upon cessation of drug abuse and patient compliance. In our experience this is most easily achieved in a multidisciplinary environment.

Hot-air ballooning injuries in the United Kingdom (January 1976-January 2004).

OBJECTIVES: Injuries due to hot-air ballooning accidents are uncommon but may be severe. In this study we examined the factors contributing to ballooning accidents in the UK and the type of injuries sustained. METHODS: The post-investigation reports of ballooning accidents from 1976 to 2004 were reviewed. They were analysed to determine type and severity of injuries, phase of flight at time of accident and causative or contributory factors. RESULTS: Sixty-one people were seriously injured in 98 ballooning accidents, with 2 fatalities. The majority of these accidents occurred during the approach and landing phase of flight. Ground collisions and crashes with power lines accounted for the majority of accidents. Adverse weather conditions were present in a significant number of crashes. Equipment failure was an uncommon cause of accidents. Fractures and burns were the most common form of injuries sustained. CONCLUSIONS: Changes in design and materials used, as well as adherence to specific recommended safety protocols, may aid to decrease the risk of severe burns in ballooning accidents. We discuss these factors illustrated by our experience of the worst recorded ballooning accident in Britain.

Induction of tetrasomy by human papillomavirus type 16 E7 protein is independent of pRb binding and disruption of differentiation.

We have demonstrated previously that high-risk human papillomaviruses (HPVs) induce tetrasomy in low-grade squamous intraepithelial lesions of the cervix. In this study we show that the E6 and E7 genes of high-risk HPV-16, but not those of low-risk HPV-6, are independently able to induce tetrasomy when constitutively expressed in proliferating monolayer cultures of primary human keratinocytes. Of seven HPV-16 E7 mutants analysed (H2P, Delta6-10, Delta21-24, C24G, S31G/S32G, A50S and S71I), five were severely impaired in their ability to induce tetrasomy in monolayer and raft culture. Only mutant C24G induced tetrasomy to levels comparable with wild-type E7 in monolayer and raft culture. This mutant shows strongly reduced binding to the retinoblastoma gene product pRb. The casein kinase II phosphorylation defective mutant S31G/S32G induced tetrasomy to levels comparable with wild-type E7 in raft culture, but not in monolayer culture, and induction of tetrasomy did not correlate with raft morphology. These results indicate that pRb protein binding is not required for HPV-16 E7 associated tetrasomy and that tetrasomy is not directly related to the ability of this protein to disrupt keratinocyte differentiation.