The renal clear cell carcinoma immune landscape.

A comprehensive evaluation of the clear cell renal cell carcinoma (ccRCC) immune landscape was found using 584 RNA-sequencing datasets from The Cancer Genome Atlas (TCGA), we identified 17 key dysregulated immune-associated genes in ccRCC based on association with clinical variables and important immune pathways. Of the numerous findings from our analyses, we found that several of the 17 key dysregulated genes are heavily involved in interleukin and NF-kB signaling and that somatic copy number alteration (SCNA) hotspots may be causally associated with gene dysregulation. More importantly, we also found that key immune-associated genes and pathways are strongly upregulated in ccRCC. Our study may lend novel insights into the clinical implications of immune dysregulation in ccRCC and suggests potential immunotherapeutic targets for further evaluation.

Potential for improving fiber digestion in the rumen of cattle (Bos taurus) through microbial inoculation from bison (Bison bison): In situ fiber degradation.

The objective of this experiment was to determine if partial replacement of cattle rumen contents with those from bison would increase in situ ruminal fiber degradation of various forages. The second objective was to examine individual variation among cattle in their ability to degrade forage and their responses to inoculation. In situ degradation of barley straw, canola straw, alfalfa hay, and timothy hay was measured in 16 ruminally cannulated heifers fed a barley straw-based diet before and after inoculation with combined rumen contents from 32 bison (performed twice, 14 d apart). Each feed was incubated in the rumen of each heifer for 0, 4, 8, 12, 24, 48, 96, and 120 h, and the degradation parameters were determined as washout fraction (), potentially degradable fraction (), rate of digestion of fraction (), and total potentially degradable fraction (). The of barley straw decreased ( = 0.04) after inoculation, whereas fraction of NDF increased ( = 0.03) and fraction of NDF and ADF decreased ( ≤ 0.02) by inoculation. In contrast, of alfalfa hay NDF and ADF decreased ( = 0.002) after inoculation, but fraction of NDF and ADF ( ≤ 0.02) increased. There were no major effects ( > 0.06) of inoculation on the fiber degradation of timothy hay or canola straw. The differential response between barley straw and alfalfa hay may have occurred because the cattle were previously adapted to a barley straw diet, whereas the bison were fed barley silage and oats. Some animals consistently ranked higher or lower for or across at least 3 of the 4 feeds incubated, but the rankings changed after inoculation. In conclusion, inoculation of cattle with bison rumen contents failed to improve degradation of fiber from barley straw, canola straw, or timothy hay in cattle well adapted to a barley straw diet, although there were small improvements in the extent of degradation of fiber from alfalfa hay. Cattle varied both in their ability to degrade various forages and in their responses to inoculation with bison rumen contents.

Sentinel lymph node biopsy in early breast cancer surgery--working with the risks of vital blue dye to reap the benefits.

AIMS: Sentinel lymph node biopsy (SLNB) is an important method of staging early breast cancer because of the inherent benefits it confers on patients in terms of arm function and quality of life. Its success depends on a high level of accuracy in detecting the sentinel node. This is achieved by a dual mapping technique that employs a radio-labelled nanocolloid and a vital blue dye. The vital dyes however carry the risk of anaphylaxis, and as more surgeons employ SLNB in their daily practice, a proportionate rise in the number of anaphylactic reactions can be expected. A comprehensive review of risks and benefits associated with using vital blues dyes has not been published and therefore a retrospective review was undertaken of the different levels of anaphylaxis associated with vital dyes as well as their benefits in SLNB. METHODS: An OVID MEDLINE search was performed of the English published literature using appropriate search terms to find published trial data and case series that focused on adverse reactions to vital blue dyes. RESULTS: The risk of severe anaphylaxis (grade 3) can be as low as 0.06%, and up to 0.4% for patients undergoing SLNB when data is analysed from large trials. Furthermore, adverse reactions associated with blue dyes are reversible with appropriate management. CONCLUSIONS: Surgeons should continue to use vital dyes to ensure that SLNB remains a highly sensitive procedure.

Development of an oncolytic herpes simplex virus using a tumor-specific HIF-responsive promoter.

We exploited the differential activation of hypoxia-inducible factor (HIF)-dependent gene expression in tumors versus normal tissue for the design of a targeted oncolytic herpes simplex virus type-1 (HSV-1). A gene that is essential for viral replication, infected cell polypeptide 4 (ICP4), was placed under the regulation of an HIF-responsive promoter and then introduced into the thymidine kinase locus (U(L)23) of HSV d120, which contains partial deletions in the two endogenous ICP4 genes. Recombinant HIF-HSV was isolated and their derivation from d120 was verified by expression of a truncated, non-functional form of ICP4 protein. Disruption of the U(L)23 locus was confirmed by loss of thymidine kinase expression and resistance to acyclovir. Unexpectedly, HIF-HSV expressed ICP4 and induced tumor cell lysis at similar levels under normoxia and hypoxia. The lack of HIF-dependent ICP4 transgene expression by HIF-HSV was due to two factors that have not previously been reported-reversion of the ICP4 gene region to its wild-type configuration and increased HIF-transcriptional activity under normoxia when cells were infected with any strain of HSV-1. The findings that an oncolytic HSV-1 is genetically unstable and can activate a tumor-related promoter in a non-specific manner have important implications for any proposed use of this virus in cancer therapy.

Development of a method to measure plasma and whole blood choline by liquid chromatography tandem mass spectrometry.

BACKGROUND: Current gold standard markers for myocardial damage are troponins I and T, which are both sensitive and specific for the detection of myocardial infarction, but require up to 6 h to become reliably elevated in serum. Investigation into markers with potential to identify patients with early ischaemic changes is therefore intense. Choline is reported to be prognostic in patients presenting with acute coronary syndromes via its release from ischaemic cell membranes. METHODS: Liquid chromatography tandem mass spectrometry was used to develop a method to quantitate choline in plasma and blood. The method involves addition of a deuterated internal standard to an aliquot of plasma or blood followed by organic solvent addition, which precipitates the proteins in the sample. Preparation was carried out directly into a 96-deep-well plate. Chromatography of choline used a strong cation exchange column and separation used a Waters Atlantis dC18 analytical column positioned directly before the mass spectrometer source, allowing on-line preanalytical clean up of the sample. RESULTS: The lower limit of quantitation was 0.38 micromol/L, linearity was observed up to 754 micromol/L, with a working concentration range of 0.38-224 micromol/L, inter- and intra-assay coefficients of variation were <6% and <4%, respectively. Samples were stable throughout five freeze-thaw cycles and recovery was between 94% and 114%. CONCLUSIONS: The assay was successfully validated in accordance with FDA guidelines and is suitable for quantitation of choline in research and clinical settings.

Successful AV fistula creation does not lead to higher catheter use: the experience by the Northwest Renal Network 16 Vascular Access Quality Improvement Program. Four years follow-up.

BACKGROUND: In 2002, the Center for Medicare and Medicaid Services (CMS) required all 18 Renal Networks to participate in a Vascular Access Quality Improvement Program (QIP). The Northwest Renal Network (NWRN 16) chose to increase arteriovenous fistula (AVF) use. NWRN 16 hypothesized that strategies which targeted the improvement of AVF rate and the reduction of catheter use were the same. In December 2001, 44.2% of hemodialysis (HD) patients in the NWRN 16 received HD using an AVF which met the Dialysis Outcome Quality Initiative (K/DOQI) 40% AVF guideline for prevalent patients. However, 43% of HD facilities (2869 patients) had less than 40% of AVF and higher HD catheter rates than the average Network catheter rates (25.0 vs. 20.3%). To address the needs of underperforming facilities, NWRN 16 provided education and tools for their vascular access decision makers to promote AVF creation and catheter reduction. METHODS: In 2002, NWRN 16 sponsored four regional workshops targeted at nephrologists, vascular surgeons, HD nurses, and interventional radiologists. RESULTS: Percentage of AVFs in use in invited facilities increased from 31.3% pre-intervention to 56.2% at 4 yrs: 78% increase (99% confidence interval: 77.8% to 81.5%). Percentage of catheters increased from 25% to 25.8%: 3.2% change over 4 yrs (99% confidence interval: 2.5% to 4%). CONCLUSION: The success of Network 16's AVF interventions demonstrates the effectiveness of Network education promoting multidisciplinary teamwork, and innovative strategies to increase dramatically AVF use without substantial increase in catheter use.

A modified ATP benchmark for evaluating the cleaning of some hospital environmental surfaces.

Hospital cleaning continues to attract patient, media and political attention. In the UK it is still primarily assessed via visual inspection, which can be misleading. Calls have therefore been made for a more objective approach to assessing surface cleanliness. To improve the management of hospital cleaning the use of adenosine triphosphate (ATP) in combination with microbiological analysis has been proposed, with a general ATP benchmark value of 500 relative light units (RLU) for one combination of test and equipment. In this study, the same test combination was used to assess cleaning effectiveness in a 1300-bed teaching hospital after routine and modified cleaning protocols. Based upon the ATP results a revised stricter pass/fail benchmark of 250 RLU is proposed for the range of surfaces used in this study. This was routinely achieved using modified best practice cleaning procedures which also gave reduced surface counts with, for example, aerobic colony counts reduced from >100 to <2.5 cfu/cm(2), and counts of Staphylococcus aureus reduced from up to 2.5 to <1 cfu/cm(2) (95% of the time). Benchmarking is linked to incremental quality improvements and both the original suggestion of 500 RLU and the revised figure of 250 RLU can be used by hospitals as part of this process. They can also be used in the assessment of novel cleaning methods, such as steam cleaning and microfibre cloths, which have potential use in the National Health Service.

An oncolytic mutant of herpes simplex virus type-1 in which replication is governed by a promoter/enhancer of human papillomavirus type-16.

Although herpes simplex virus type-1 (HSV-1) can be used as an oncolytic virus it has the undesirable side effect of neurotoxicity. To create a virus with improved specificity for oral cancer we used a fragment of human papillomavirus type-16, which is frequently found in oral and cervical cancers, but not elsewhere. The upstream regulatory region, URR16, was shown to have a high level of transcriptional activity in three of four oral cancer cell lines but low activity in three cell lines derived from brain cancers. URR16 was therefore placed in HSV-1, replacing the promoter of the essential gene ICP4, and the resulting virus was named HSPV-1. When cells were infected with HSPV-1, ICP4 was expressed at levels that were not associated with the level of activity of URR16. The virus replicated in each type of cell to a final titer that showed a correlation with the level of expression of ICP4, but with no correlation to either the tumor of origin or the presence of HPV sequences in the cells. To find if some function of HSV-1 was affecting the activity of URR16, oral cancer cells were transfected with a URR-reporter construct and were then infected with virus. This induced transcription, which was attributed to immediate-early viral genes other than ICP4. A promoter/enhancer from a papillomavirus therefore has the potential to regulate the functions of an oncolytic strain of HSV-1, and is affected by functions of both the host cell and of HSV-1 itself.

Problems associated with traditional hygiene swabbing: the need for in-house standardization.

AIMS: To investigate factors influencing the recovery of micro-organisms from surfaces using traditional swabbing techniques. METHODS AND RESULTS: Stainless steel squares were inoculated with known levels (approx. 2.7x10(2)-2.7x10(4)) of either Escherichia coli or Staphylococcus aureus and sampled using different swab/solution combinations. Overlaying the coupons with agar allowed colonies remaining on the surface to be enumerated. Conventional cultivation was used to determine the ease with which the bacteria were released from the swabs and the viability of the organisms within the solutions over a 24-h period. Minimal bacterial growth occurred when the samples were stored at 4 degrees C. At room temperature, whilst the presence of nutrients significantly increased bacterial numbers over time, the addition of Tween 80 to nutrient depleted environments significantly reduced the viability of Staph. aureus. The percentage of bacteria released from directly inoculated swabs was significantly higher than that recovered from surface swabs, highlighting the importance of effectively removing bacterial contaminants from a surface. Increasing the level of mechanical energy generated during swabbing increased the number of bacteria removed from a wet surface. However, it is hypothesized that cellular damage, perhaps caused by the swabbing action itself, may have reduced recoverability from a dry surface. Nonetheless, an increased ability to effectively remove bacteria from a surface did not necessarily correlate with higher bacterial recovery, implying that an equally important factor in terms of swabbing efficiency is the ability of a swab to effectively release bacteria into a diluent. CONCLUSIONS: Both swab and wetting solution can influence the number of bacteria recovered. Under the experimental conditions described here, the use of swabs coated with a brush-textured nylon flock in combination with a non-growth-enhancing wetting solution appeared the best system to use when sampling a wet surface. However, this combination may not always be ideal and proper consideration must be given to how the sample is to be taken, transported and, if necessary, stored prior to analysis. SIGNIFICANCE AND IMPACT OF THE STUDY: Careful selection of swabbing materials can increase the sensitivity of traditional microbiological analysis. However, any improvements made are likely to be insignificant in relation to the overall poor performance of the swabbing technique.

The effectiveness of existing and modified cleaning regimens in a Welsh hospital.

Hospital cleaning currently has a high media profile. The effectiveness of an existing ward-cleaning regimen was assessed at selected sites over a 14 day period and shown to be highly variable. The cleaning regimen was subsequently modified in two stages, both changes involving a rinse stage and substituting cloths with disposable paper towels. One modification continued using the existing detergent; the other replaced detergent with a quaternary ammonium sanitiser. Both modifications yielded significantly lower and more consistent bacterial counts. Assessment of residual organic soil using adenosine triphosphate (ATP) detection demonstrated that failure rates (measurements exceeding benchmark clean value of 500 relative light units (RLU)) fell from 86-100% after existing cleaning methods, to 0-14% after modified cleaning. Maximum ATP readings fell from 163,870 to 2289 RLU. Incorporating a quaternary ammonium sanitiser into the cleaning regimen produced a further slight, but not significant, improvement in cleaning efficacy. These findings suggest that simple improvements can be made to existing cleaning regimens to increase their efficacy.

Creation of secondary AV fistulas from failed hemodialysis grafts: the role of routine vein mapping.

Pre-existing forearm grafts lead to dilatation of upper arm veins of the ipsilateral arm that greatly facilitates the creation of secondary arteriovenous fistulas (AVF). In this retrospective review of 18 patients, the routine and periodical revision of failed grafts were discontinued. Vein mapping by physical examination or Duplex studies was performed prior to graft failure and a secondary AVF was created when the graft failed. Using this management strategy, an AVF creation success rate of 95% was achieved despite high incidence of co-morbidity factors within the patient population. This success rate demonstrates the value of vein selection by vascular mapping prior to graft failure and early vascular access planning in the successful creation of secondary AVF.

Successful multidisciplinary interventions for arterio-venous fistula creation by the Pacific Northwest Renal Network 16 vascular access quality improvement program.

BACKGROUND: In December 2001, 44.2% of hemodialysis (HD) patients in the Northwest Renal Network (NWRN 16) received dialysis using an arteriovenous fistula (AVF). Substantial differences were noted in percentages of patients with AVF, ranging from 5% to 90% of the facility population, suggesting wide variation in physician practice patterns within the Network. To address the needs of facilities having < 40% AVF, NWRN 16 provided education and tools for their vascular access decision-makers to promote AVF creation. METHODS: In 2002, the Network sponsored 4 regional workshops targeted to nephrologists, vascular surgeons, dialysis nurses, and interventional radiologists. RESULTS: 46 facilities (43% of all Network facilities) had <40% AVF in use in December, 2001, dialyzing 2940 patients (Invited Units). Percent AVF in use in all the Invited Facilities increased from 31.3% pre-intervention to 39.8% at 1 year (p<0.001 vs pre) to 56.2% at four years: 79.8% increase in the prevalent AVF rate over a four-year period (95% confidence interval: 77.8% to 81.7%). CONCLUSION: Low prevalent AVF rates in many NWRN 16 facilities may have resulted from differences in physician practice patterns. The success of Network 16 AVF Intervention demonstrates the effectiveness of Network education promoting multidisciplinary teamwork, innovative strategies to increase AVF rates among dialysis patients.

An evaluation of different methods for the recovery of meticillin-resistant Staphylococcus aureus from environmental surfaces.

Although the association between environmental surfaces contaminated with meticillin-resistant Staphylococcus aureus (MRSA) and hospital infection rates is not fully understood, monitoring programmes can provide an objective starting point for the development and assessment of infection control strategies incorporating improved cleaning. There is, however, no universally accepted method for the recovery of MRSA from environmental surfaces, and the aim of this study was to evaluate a selection of currently available methods. Using five clinical isolates of MRSA and seven protocols, known numbers of bacteria were inoculated on to a stainless steel surface and either recovered immediately (without drying or adsorption) or recovered after 30min (with drying at room temperature and adsorption of cells to the surface). Surfaces were either swabbed or sampled directly by contact methods, and four nutritive media (blood, tryptone soya, oxacillin and meticillin-resistant agars) were tested. Relative sampling efficiencies were determined and the sensitivity of each method per 100cm(2) was calculated. Wide variation in the ability to recover MRSA was found between the different protocols. In the recovery of dried (adsorbed) cells, direct contact methods demonstrated higher sampling efficiency than swabs. The sensitivity of all methods was lower in recovering adsorbed cells from surfaces than unadsorbed cells. Sampling methods consistently proved to be more important than the choice of medium. Dipslides coated with selective agar are recommended for recovering MRSA from flat environmental surfaces.

Brain volume measurements in patients with human T-cell lymphotropic virus-1-associated tropical spastic paraparesis.

Human T-cell lymphotropic virus (HTLV)-1 is associated with a chronic progressive neurologic disease known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) that affects 0.2% to 3% of HTLV-1-infected people. The authors aimed at exploring, in vivo, whether brain volume reduction occurs in patients with HAM/TSP through the use of magnetic resonance imaging (MRI). T1 pre/postcontrast spin echo-weighted images (WIs) and T2WIs of the brain were obtained in 19 HAM/TSP patients and 14 age-and sex-matched healthy volunteers. Both patients and healthy individuals were imaged at a 1.5-Tesla magnet by employing a conventional head coil. Focal T1 and T2 abnormalities were calculated and two measurements of brain parenchyma fraction (BPF) were obtained by using SIENAx (Structural Image Evaluation,using Normalisation, of Atrophy; University of Oxford, Oxford, UK) and MIPAV (Medical Image Processing, Analysis, and Visualization; National Institutes of Health, Bethesda, USA) from T1WIs. No significant differences in BPF were found between patients and healthy subjects when using either SIENAx or MIPAV. Analysis of individual patients detected that BPF was lower by 1 standard deviation (SD) relative to patients' average BPF in one patient. The authors conclude that reductions in BPF do not occur frequently in patients with HAM/TSP. However, the authors believe that one individual case of significant brain atrophy raises the question as to whether atrophy selectively targets the spinal cord of HAM/TSP patients or may involve the brain as well. A larger patient population analyzing regional brain volume changes could be helpful in determining whether brain atrophy is a marker of disease in patients with HAM/TSP.

A laboratory evaluation of the decontamination properties of microfibre cloths.

Standards of cleanliness in health care continue to attract attention. Effective cleaning requires the input of energy, and microfibre cloths may help in the physical removal of soil. The ability of these cloths to remove organic soil (measured by ATP) and bacteria was compared with paper towel and a conventional cloth in controlled wet and dry conditions. When used wet on a dry surface, the cleaning ability of six different microfibre cloths was variable, and in most cases, not significantly better than paper towel or a conventional cloth. One type of microfibre cloth did perform significantly better than the others and paper towel in reducing both organic soil and microbial load. When used dry on a dry surface, there was no significant difference between the cloths, and none of the cloths reduced microbial and organic bioburden effectively. The ability of the cloths to recontaminate the surface was also tested, and some of the microfibre cloths transferred significantly less organic debris and micro-organisms back to the surface than other cloths. Different makes of microfibre cloths have different characteristics, and the name 'microfibre' should not imply superior cleaning efficacy.

Evidence for a polar ethane cloud on Titan.

Spectra from Cassini's Visual and Infrared Mapping Spectrometer reveal the presence of a vast tropospheric cloud on Titan at latitudes 51 degrees to 68 degrees north and all longitudes observed (10 degrees to 190 degrees west). The derived characteristics indicate that this cloud is composed of ethane and forms as a result of stratospheric subsidence and the particularly cool conditions near the moon's north pole. Preferential condensation of ethane, perhaps as ice, at Titan's poles during the winters may partially explain the lack of liquid ethane oceans on Titan's surface at middle and lower latitudes.