Implementing nitrous oxide cracking technology in the labour ward to reduce occupational exposure and environmental emissions: a quality improvement study.

Nitrous oxide, a potent greenhouse gas, is a common labour analgesic. One method which may reduce its carbon footprint is to 'crack' the exhaled gas into nitrogen and oxygen using catalytic destruction. In this quality improvement project, based on environmental monitoring and staff feedback, we assessed the impact of nitrous oxide cracking technology in the maternity setting. Mean ambient nitrous oxide levels were recorded during the final 30 minutes of uncomplicated labour in 36 cases and plotted on a run chart. Interventions were implemented in four stages, comprising: stage 1, baseline (12 cases); stage 2, cracking with nitrous oxide delivered and scavenged via a mouthpiece (eight cases); stage 3, cracking with nitrous oxide via a facemask with an air-filled cushion (eight cases); stage 4, cracking with nitrous oxide via a low-profile facemask, and enhanced coaching on the use of the technology (eight cases). The median ambient nitrous oxide levels were 71% lower than baseline in stage 2 and 81% lower in stage 4. Staff feedback was generally positive, though some found the technology to be cumbersome; successful implementation relies on effective staff engagement. Our results indicate that cracking technology can reduce ambient nitrous oxide levels in the obstetric setting, with potential for reductions in environmental impacts and occupational exposure.

Substantial long-term loss of alpha and gamma diversity of lake invertebrates in a landscape exposed to a drying climate.

Many regions across the globe are shifting to more arid climates. For shallow lakes, decreasing rainfall volume and timing, changing regional wind patterns and increased evaporation rates alter water regimes so that dry periods occur more frequently and for longer. Drier conditions may affect fauna directly and indirectly through altered physicochemical conditions in lakes. Although many studies have predicted negative effects of such changes on aquatic biodiversity, empirical studies demonstrating these effects are rare. Global warming has caused severe climatic drying in southwestern Australia since the 1970s, so we aimed to determine whether lakes in this region showed impacts on lake hydroperiod, water quality, and α, ß and γ diversity of lake invertebrates from 1998 to 2011. Seventeen lakes across a range of salinities were sampled biennially in spring in the Wheatbelt and Great Southern regions of Western Australia. Multivariate analyses were used to identify changes in α, ß and γ diversity and examine patterns in physicochemical data. Salinity and average rainfall partially explained patterns in invertebrate richness and assemblage composition. Climatic drying was associated with significant declines in lake depth, increased frequency of dry periods, and reduced α and γ diversity (γ declined from ~300 to ~100 taxa from 1998 to 2011 in the 17 wetlands). In contrast, ß diversity remained consistently high, because each lake retained a distinct fauna. Mean α diversity per-lake declined both in lakes that dried and lakes that did not dry out, but lakes which retained a greater proportion of their maximum depth retained more α diversity. Accumulated losses in α diversity caused the decline in γ diversity likely through shrinking habitat area, fewer stepping stones for dispersal and loss of specific habitat types. Biodiversity loss is thus likely from lakes in drying regions globally. Management actions will need to sustain water depth in lakes to prevent biodiversity loss.

A potent Chk1 inhibitor is selectively cytotoxic in melanomas with high levels of replicative stress.

There are few effective treatments for metastatic melanoma. Checkpoint kinase 1 (Chk1) inhibitors are being trialled for their efficacy in enhancing conventional chemotherapeutic agents, but their effectiveness as single agents is not known. We have examined the effectiveness of two novel Chk1 selective inhibitors, AR323 and AR678, in a panel of melanoma cell lines and normal cell types. We demonstrate that these drugs display single-agent activity, with IC50s in the low nanomolar range. The drugs produce cytotoxic effects in cell lines that are most sensitive to these drugs, whereas normal cells are only sensitive to these drugs at the higher concentrations where they have cytostatic activity. The cytotoxic effect is the consequence of inhibition of S-phase Chk1, which drives cells prematurely from late S phase into an aberrant mitosis and results in either failure of cytokinesis or cell death through an apoptotic mechanism. The sensitivity to the Chk1 inhibitors was correlated with the level of endogenous DNA damage indicating replicative stress. Chk1 inhibitors are viable single-agent therapies that target melanoma cells with high levels of endogenous DNA damage. This sensitivity suggests that Chk1 is a critical component of an adaptation to replicative stress in these cells. It also suggests that markers of DNA damage may be useful in identifying the melanomas and potentially other tumour types that are more likely to be sensitive to Chk1 inhibitors as single agents.

Multiple melanoma susceptibility factors function in an ultraviolet radiation response pathway in skin.

BACKGROUND: Exposure to ultraviolet radiation (UVR) and the familial melanoma susceptibility gene p16 (CDKN2A) are among the major risk factors which have been identified to contribute to the development of melanoma, and also significantly contribute to squamous cell carcinoma. We have previously shown that UVR induces p16(CDKN2A) expression in melanoma and keratinocyte cell lines and human skin, but the regulatory mechanisms controlling this expression are unknown. OBJECTIVES: To determine the mechanism by which UVR induces p16(CDKN2A) expression in melanocytes and keratinocytes in the epidermis. METHODS: We have used an in vitro cell lines model of the UVR response in skin to assess the changes in p16(CDKN2A) expression and the signalling pathways regulating these changes, and validated these findings in whole human skin cultures. RESULTS: We show that UVR-induced ERK signalling, mediated by BRAF, regulates p16(CDKN2A) expression at the transcriptional, and possibly translational level. CONCLUSIONS: This study demonstrates the biological connection between the known melanoma genes p16 (CDKN2A) and BRAF in a normal physiological response to UVR in the skin, and highlights the importance of defects in this biological pathway to melanoma and squamous cell carcinoma development.

Nitrite directly vasodilates hypoxic vasculature via nitric oxide-dependent and -independent pathways.

BACKGROUND AND PURPOSE: It is postulated that nitrite requires reduction to nitric oxide in order to exert its relaxant effect upon isolated hypoxic vessels. Herein, we evaluate the relative contribution of nitric oxide and characterize the downstream mechanisms of nitrite-induced vasorelaxation. EXPERIMENTAL APPROACH: Aortic rings were treated with pharmacological agents and exposed to hypoxia (<1% O(2)). Following pre-constriction, nitrite (10 microM final) was added to appropriate baths; isometric tension was recorded throughout. KEY RESULTS: Nitrite (under hypoxic conditions at physiological pH) is capable of exerting physiological effects that cannot be completely inhibited by the inhibitor of soluble guanylate cyclase (sGC), 1H [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one or a nitric oxide scavenger (carboxy-2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide). Simultaneous blockade of both sGC and cyclooxygenase (COX) completely inhibited the response to nitrite. With regard to the nitric oxide-dependent component, we confirm that aldehyde oxidase, but not xanthine oxidase or endothelial nitric oxide synthase, was important for the actions of nitrite in our model. CONCLUSIONS AND IMPLICATIONS: Nitric oxide generated from nitrite is not exclusively responsible for the physiological actions observed in isolated hypoxic vessels. Nitrite operates via different pathways dependent on the presence or absence of endothelium to produce vasorelaxation. In intact vessels, both sGC and COX enzymes appear to be important. Irrespective of this difference in relaxation mechanism, nitrite is capable of producing the same maximum relaxation, regardless of the presence of endothelium. Having investigated possible nitrite reduction sites, we confirm that aldehyde oxidase is important for the actions of nitrite.

Anaesthesia for caesarean section in women with complex cardiac disease: 34 cases using the Braun Spinocath spinal catheter.

BACKGROUND: Cardiac disease in pregnancy is now the leading medical cause of maternal mortality in the UK. Whilst anaesthesia has not been the precipitant of this morbidity, its safety cannot be taken for granted. Spinal catheter anaesthesia, a relatively uncommon choice in obstetric practice, offers the potential of maintaining haemodynamic stability through accurate and gradual titration of neuraxial blockade. METHODS: Thirty-four women with cardiac disease requiring caesarean section were selected for spinal catheter anaesthesia. All received invasive arterial pressure measurement but in only two were central venous catheters sited. After inserting a 24-gauge Braun Spinocath, spinal anaesthesia was induced using diamorphine 300 microg and 0.5% hyperbaric bupivacaine in 0.25-mL increments. Technical problems, block quality and haemodynamic stability were recorded. RESULTS: Successful anaesthesia was achieved in 33 women. Spinal catheterisation proved impossible in one case, but the catheter was successfully used to provide epidural anaesthesia. There were no conversions to general anaesthesia. Eight women (24%) received supplementation with intravenous alfentanil, but all reported high satisfaction. Mild, transient hypotension occurred in six women (18%), and there was one case of vasovagal syncope induced by rapid exteriorisation of the uterus. Three patients (8.8%) experienced post dural puncture headache requiring a blood patch; two had received repeat dural puncture during catheter insertion. CONCLUSIONS: Incremental spinal catheter anaesthesia offers effective anaesthesia with excellent haemodynamic control. Post dural puncture headache is of concern, and whilst it may be addressed by product modification, it currently limits widespread use of the Braun Spinocath in obstetric practice.

Dose response to intrathecal diamorphine for elective caesarean section and compliance with a national audit standard.

BACKGROUND: This double-blind randomised controlled trial investigated the most appropriate dose of intrathecal diamorphine to use with high-dose diclofenac as part of a multimodal analgesic regimen for caesarean section under subarachnoid block. We also wished to establish whether it was possible to satisfy the Royal College of Anaesthetists postoperative pain audit recommendation for this patient group. METHODS: One hundred and twenty patients presenting for elective caesarean section under subarachnoid block were recruited and divided into four groups. Treatment was standard except that patients were given either placebo or one of three different doses of intrathecal diamorphine (100 microg, 200 microg or 300 microg). All patients were given regular paracetamol, high-dose diclofenac and an hourly subcutaneous diamorphine regimen for breakthrough pain. RESULTS: There was a dose-dependent improvement in analgesia with intrathecal diamorphine. Only 37.9% of patients given 300 microg of intrathecal diamorphine had a visual analogue pain score of 3/10 or less throughout the study. There was a dose-dependent increase in the incidence of itching with intrathecal diamorphine although the incidence of nausea and vomiting was similar between groups. CONCLUSIONS: We found that for elective caesarean section under subarachnoid block with high dose diclofenac, analgesia was optimal with 300 microg of intrathecal diamorphine. Even the highest dose of intrathecal diamorphine did not achieve the Royal College of Anaesthetists postoperative audit target that 90% of patients should have a pain score of no more than 3/10. We believe that this target is too arduous.

Meningococcal meningitis after combined spinal-epidural analgesia.

We present a case of bacterial meningitis in a 32-year-old parturient following combined spinal-epidural analgesia for labour. The patient made a full recovery with no residual neurological sequelae, but important lessons were learnt. Firstly, investigating obstetricians and physicians were unaware that a combined spinal-epidural technique included an intrathecal component, so did not consider treating organisms that might be acquired by this route. Anaesthetists, on the other hand, in the absence of an isolated organism, saw this as a likely combined spinal-epidural complication. Infectious disease experts eventually diagnosed community-acquired meningococcal meningitis by analysing bacterial deoxyribonucleic acid (DNA) fragments using polymerase chain reaction studies. This test and the management of suspected meningitis in the post-partum period are discussed.

Haemodynamic changes caused by oxytocin during caesarean section under spinal anaesthesia.

The haemodynamic effects of oxytocin receive scant attention in pharmacology texts, but may be clinically significant in vulnerable patients. Despite prescriber information recommending a dose of 5 international units by slow i.v. injection, it is the authors' experience that it is very common practice in the UK to give 10 units as a rapid injection. We therefore conducted a randomised, double-blind study of the haemodynamic changes induced by rapid bolus of 5 or 10 units of oxytocin in 34 healthy term parturients at caesarean section under spinal anaesthesia. There was a small but statistically significant (P < 0.05) reduction in mean arterial pressure from baseline 30 s after a 10-unit bolus. However, large, statistically significant increases in heart rate and cardiac output occurred 1 min after 5 units and 2 min after 10 units. These changes peaked 1 min after oxytocin administration and were greater in the 10-unit group (P < 0.05). The importance of these findings is that some women with hypovolaemia or cardiac disease may be unable to mount these compensatory responses and are therefore at risk of haemodynamic collapse after oxytocin boluses. This has been illustrated by a maternal death reported to the Confidential Enquiries into Maternal Deaths in the United Kingdom. The need to adhere to a dose regimen of 5 units by slow injection needs re-emphasis, but no evidence exists to claim that even this will be haemodynamically inert. We therefore recommend that oxytocin boluses be avoided in women with hypovolaemia or cardiac disease.

Apoptosis can be detected in attached colonic adenocarcinoma HT29 cells using annexin V binding, but not by TUNEL assay or sub-G0 DNA content.

BACKGROUND: Induction of apoptosis in adherent cell lines is associated with cell loss from the substratum. In this study the adenocarcinoma cell line, HT29, treated with indomethacin (400microM) has been employed as a model system to demonstrate how flow cytometric analysis can be used to quantify the changes that occur during this process. METHODS: Adherent and floating cell populations have been analyzed independently for effects on cell number, cell cycle characteristics and apoptosis using TUNEL assay and Annexin V binding. In addition apoptosis has been assessed using DNA laddering and morphology. RESULTS: Apoptosis was detected in adherent cells treated with indomethacin using Annexin V binding but not by other techniques employed in this study. In contrast, analysis of "floating" cells revealed the presence of apoptotic cells both in control and indomethacin treated cells using all the techniques employed. However quantification by flow cytometry showed that a significantly higher proportion of control "floaters" were late apoptotic/necrotic rather than apoptotic. DISCUSSION: The data here illustrate the need to interpret measures of apoptosis in adherent cell lines with care and the value of using flow cytometric techniques in the quantitative evaluation of the process.

n-3 polyunsaturated fatty acids inhibit the antigen-presenting function of human monocytes.

Diets rich in n-3 polyunsaturated fatty acids (PUFAs) are associated with suppression of cell-mediated immune responses, but the mechanisms are unclear. We hypothesized that n-3 PUFAs can inhibit the function of human antigen-presenting cells. A prerequisite for this role of blood monocytes is the cell surface expression of major histocompatibility complex (MHC) class II molecules [human leukocyte antigen (HLA)-DR, -DP, and -DQ], aided by the presence of intercellular adhesion molecule-1 (ICAM-1) and leukocyte function associated antigens 1 and 3. We showed previously that the n-3 PUFA eicosapentaenoic acid (EPA) inhibits the expression of HLA-DR on unstimulated human monocytes in vitro, but that docosahexaenoic acid (DHA) enhances its expression. However, both n-3 PUFAs suppress the expression of HLA-DR, HLA-DP, and ICAM-1 on interferon-gamma-activated monocytes. We also established that dietary fish-oil supplementation can inhibit the expression of these surface molecules on circulating human monocytes. We subsequently showed that when EPA and DHA were combined in the same ratio as is commonly found in fish-oil-supplement capsules (3:2), there was no significant effect in vitro on the expression of HLA-DR on unstimulated monocytes, but the expression on activated monocytes remained significantly inhibited. In the same in vitro system, the ability of activated monocytes to present antigen to autologous lymphocytes was significantly reduced after culture with the combined n-3 PUFAs. These findings provide one potential mechanism for the beneficial effect of fish oil in the treatment of rheumatoid arthritis, a disorder associated with elevated expression of MHC class II and adhesion molecules on monocytes present within affected joints.

Does tomato consumption effectively increase the resistance of lymphocyte DNA to oxidative damage?

BACKGROUND: Lycopene, the main carotenoid in tomato, has been shown to be a potent antioxidant in vitro. However, there is no significant evidence of its antioxidant action in vivo. OBJECTIVE: We evaluated the effect of tomato intake on plasma carotenoid concentrations and lymphocyte resistance to oxidative stress. DESIGN: Ten healthy women (divided into 2 groups of 5 subjects each) ate a diet containing tomato puree (providing 16.5 mg lycopene) and a tomato-free diet for 21 d each in a crossover design. Before and after each diet period, plasma carotenoid concentrations and primary lymphocyte resistance to oxidative stress (evaluated by means of single-cell gel electrophoresis) were analyzed. RESULTS: After the first 21-d experimental period, total plasma lycopene concentrations increased by 0.5 micromol/L (95% CI: 0.14, 0.87) in the group that consumed the tomato diet and decreased by 0.2 micromol/L (95% CI: -0.11, -0.30) in the group that consumed the tomato-free diet (P < 0.001). Tomato consumption also had an effect on cellular antioxidant capacity: lymphocyte DNA damage after ex vivo treatment with hydrogen peroxide decreased by 33% (95% CI: 0.8%, 61%; P < 0.05) and by 42% (95% CI: 5.1%, 78%; P < 0.05) in the 2 groups of subjects after consumption of the tomato diet. CONCLUSION: The consumption of tomato products may reduce the susceptibility of lymphocyte DNA to oxidative damage.

Effect of eicosapentaenoic acid on the proliferation and incidence of apoptosis in the colorectal cell line HT29.

Fish oil has been shown to reduce the induction of colorectal cancer in animal models by a mechanism which may involve suppression of mitosis, increased apoptosis, or both. We used the human colonic adenocarcinoma cell line HT29 to explore the effects of the long-chain n-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA) on cell proliferation and death in vitro. Cells were cultured in media containing EPA at 5, 10, and 15 microg/mL. Cell number and thymidine incorporation were used to quantify proliferation, and cell cycle effects were studied using flow cytometry. Gel electrophoresis, annexin-V binding, and morphological criteria were used to characterize apoptosis. Adherent cells and freely floating detached cells were treated as two distinct populations. In the presence of EPA at 10 and 15 microg/mL there was a marked reduction in the growth rate of adherent HT29 colonies, owing to an increased detachment of adherent cells. After treatment with 10 or 15 microg/mL EPA the proportion of adherent cells in S-phase increased, indicating either a block in late S-phase or early G2. Floating cells showed evidence of extensive DNA cleavage, but the proportion of floating cells with sub GO DNA content declined on treatment with 10 or 15 microg/mL EPA even though the number of floating cells increased. We conclude that EPA does not inhibit mitosis of adherent cells, but increases the rate at which they become detached from the substrate, probably at an early stage in the initiation of apoptosis. This mechanism may be analogous to "anoikis," or induction of apoptosis in response to loss of cell contact, and may contribute to the anticarcinogenic effects of fish oil in vivo.