Trends in C-Reactive Protein Levels Are Associated with Neurological Change Twenty-Four Hours after Thrombolysis for Acute Ischemic Stroke.

BACKGROUND: Elevated inflammatory markers such as C-reactive protein (CRP) are associated with worse outcomes in patients thrombolysed for acute ischemic stroke (AIS). AIMS: To investigate whether changes in CRP levels are associated with neurological change after thrombolysis for AIS. METHODS: Retrospective analysis of a single-center database of consecutive thrombolysis cases for AIS from October 18, 2011, to June 15, 2015, inclusive. Multivariate regression analysis was used to investigate the relationship between change in CRP 12-24 hours after thrombolysis and change in NIHSS (National Institutes of Health Stroke Scale) score 24 hours after thrombolysis. The other potentially confounding predictor variables included in the model were CRP on admission and NIHSS score before thrombolysis. RESULTS: Complete data were available for 108 out of possible 435 eligible patients. Increases in CRP levels 12-24 hours after thrombolysis were negatively associated with reduction in NIHSS score 24 hours after thrombolysis (coefficient .08, 95% confidence interval .031-.129, P = .002). Thus, on average, for every 12.5 mg/L additional increase in CRP 12-24 hours after thrombolysis, NIHSS score at 24 hours improved by 1 point less. CONCLUSION: While it was previously known that elevated CRP levels are associated with worse outcomes in patients thrombolysed for AIS, the current work demonstrates that changes in CRP levels after thrombolysis also relate to neurological change, and thus may have scope for use as prognostic markers.

A Fall in Systolic Blood Pressure 24 Hours after Thrombolysis for Acute Ischemic Stroke Is Associated with Early Neurological Recovery.

BACKGROUND: Outcomes are worse in patients who underwent thrombolysis for acute ischemic stroke (AIS) with persistent hypertension. The objective of this study is to investigate whether fall in systolic blood pressure (SBP) has any relationship with neurological outcome 24 hours after thrombolysis, after adjusting for potentially confounding factors. METHODS: Retrospective analysis of a single-center database of consecutive thrombolysis cases for AIS. Multivariate regression analysis was used to explore the relationship between fall in SBP and reduction in National Institutes of Health Stroke Scale (NIHSS) score 24 hours after thrombolysis. Other potentially confounding predictor variables used in the model were SBP on thrombolysis, blood glucose level on thrombolysis, NIHSS score on thrombolysis, administration of antihypertensive medications, and the time to thrombolysis after symptom onset. RESULTS: A fall in SBP 24 hours after thrombolysis is independently associated with greater improvement in NIHSS score 24 hours after thrombolysis (coefficient .051, 95% confidence interval .023-.078, P < .001). Thus, a reduction of 10 mmHg in SBP after 24 hours is associated with a .51 point reduction in the NIHSS score. CONCLUSIONS: Restoration of SBP toward normal limits after thrombolysis for AIS is associated with greater early neurological improvement.

The theory of reference values: an unfinished symphony.

The history of the theory of reference values can be written as an unfinished symphony. The first movement, allegro con fuoco, played from 1960 to 1980: a mix of themes devoted to the study of biological variability (intra-, inter-individual, short- and long-term), preanalytical conditions, standardization of analytical methods, quality control, statistical tools for deriving reference limits, all of them complex variations developed on a central melody: the new concept of reference values that would replace the notion of normality whose definition was unclear. Additional contributions (multivariate reference values, use of reference limits from broad sets of patient data, drug interferences) conclude the movement on the variability of laboratory tests. The second movement, adagio, from 1980 to 2000, slowly develops and implements initial works. International and national recommendations were published by the IFCC-LM (International Federation of Clinical Chemistry and Laboratory Medicine) and scientific societies [French (SFBC), Spanish (SEQC), Scandinavian societies…]. Reference values are now topics of many textbooks and of several congresses, workshops, and round tables that are organized all over the world. Nowadays, reference values are part of current practice in all clinical laboratories, but not without difficulties, particularly for some laboratories to produce their own reference values and the unsuitability of the concept with respect to new technologies such as HPLC, GCMS, and PCR assays. Clinicians through consensus groups and practice guidelines have introduced their own tools, the decision limits, likelihood ratios and Reference Change Value (RCV), creating confusion among laboratorians and clinicians in substituting reference values and decision limits in laboratory reports. The rapid development of personalized medicine will eventually call for the use of individual reference values. The beginning of the second millennium is played allegro ma non-troppo from 2000 to 2012: the theory of reference values is back into fashion. The need to revise the concept is emerging. The manufacturers make a friendly pressure to facilitate the integration of Reference Intervals (RIs) in their technical documentation. Laboratorians are anxiously awaiting the solutions for what to do. The IFCC-LM creates Reference Intervals and Decision Limits Committee (C-RIDL) in 2005. Simultaneously, a joint working group IFCC-CLSI is created on the same topic. In 2008 the initial recommendations of IFCC-LM are revised and new guidelines are published by the Clinical and Laboratory Standards Institute (CLSI C28-A3). Fundamentals of the theory of reference values are not changed, but new avenues are explored: RIs transference, multicenter reference intervals, and a robust method for deriving RIs from small number of subjects. Concomitantly, other statistical methods are published such as bootstraps calculation and partitioning procedures. An alternative to recruiting healthy subjects proposes the use of biobanks conditional to the availability of controlled preanalytical conditions and of bioclinical data. The scope is also widening to include veterinary biology! During the early 2000s, several groups proposed the concept of 'Universal RIs' or 'Global RIs'. Still controversial, their applications await further investigations. The fourth movement, finale: beyond the methodological issues (statistical and analytical essentially), important questions remain unanswered. Do RIs intervene appropriately in medical decision-making? Are RIs really useful to the clinicians? Are evidence-based decision limits more appropriate? It should be appreciated that many laboratory tests represent a continuum that weakens the relevance of RIs. In addition, the boundaries between healthy and pathological states are shady areas influenced by many biological factors. In such a case the use of a single threshold is questionable. Wherever it will apply, individual reference values and reference change values have their place. A variation on an old theme! It is strange that in the period of personalized medicine (that is more stratified medicine), the concept of reference values which is based on stratification of homogeneous subgroups of healthy people could not be discussed and developed in conjunction with the stratification of sick patients. That is our message for the celebration of the 50th anniversary of Clinical Chemistry and Laboratory Medicine. Prospects are broad, enthusiasm is not lacking: much remains to be done, good luck for the new generations!

Cardiovascular disease, statins and vitamin D.

Cardiovascular disease is the number one killer in the UK, causing more than 50 000 premature deaths per year, at a cost of over £30 billion to the economy (British Heart Foundation, 2010). Reducing this burden is a priority of the Government and health professionals (Department of Health, 2000). The aim of this paper is to inform and update nurses on four aspects. Firstly, to examine a more accurate test than cholesterol to predict cardiovascular risk that used C-reactive protein (CRP), an inflammatory marker, called the Reynold's Risk Score. Use of this score reclassified almost half of women, and one-fifth of men, into lower or higher risk categories, more accurately compared to conventional tools (Ridker et al, 2007; Ridker et al, 2008b). Secondly, to highlight a potential change to the indications for statin-therapy; an indication for those who ordinarily would not receive a statin: healthy middle-aged adults with normal or low cholesterol, but elevated CRP. This includes discussion of the JUPITER trial and its sub-analyses. This large, multicentre, double-blind randomized, placebo-controlled trial in apparently healthy men over the age of 50 years and women over 60 years, with normal or low cholesterol but elevated CRP, demonstrated significant benefit. Rosuvastatin 20 mg per day compared to a placebo reduced myocardial infarction and stroke by half, and reduced venous thromboembolism by almost half (Ridker et al, 2008a; Glynn et al. 2009; Everett, et al, 2010). Thirdly, to discuss the pleiotropic effects of statins, which include reduction of CRP (Ridker et al, 2008a), increases in endothelial repair cells (Spiel, et al, 2008), alteration of clotting factors (Glynn, et al, 2009), and enhancement of vitamin D metabolism (Yavuz et al, 2009). Fourthly, to discuss evidence that the pleiotropic effects of statins may be attributable to the vitamin D effect. This paper therefore evaluates the evidence suggesting that vitamin D supplementation may attain the same benefit as statins (Grimes, 2006), in addition to reducing statinassociated side-effects when co-administered (Ahmed et al, 2009).

Reflective practice: a learning tool for student nurses.

Reflection is a vital skill in contemporary nursing with student nurses expected to engage in reflective learning from the very beginning of the nurse educational programme. This article demonstrates the meaningful learning that resulted as a consequence of using critical reflection on practice. Gibbs' (1988) cycle aided the process highlighting the practical application of this cyclical framework to the author - a first-year student nurse. Matters concerning gender issues in nursing and professional conduct emerged from the analysis and were inherently explored. The article concludes by demonstrating the personal benefits of using Gibbs' (1988) cycle to varying situations and thus promoting its excellence as a learning tool for student nurses worldwide as a consequence.

Micropillar array chip for integrated white blood cell isolation and PCR.

We report the fabrication of silicon chips containing a row of 667 pillars, 10 by 20 microm in cross-section, etched to a depth of 80 microm with adjacent pillars being separated by 3.5 microm. The chips were used to separate white blood cells from whole blood in less than 2 min and for subsequent PCR of a genomic target (eNOS). Chip fluid dynamics were validated experimentally using CoventorWare microfluidic simulation software. The amplicon concentrations were determined using microchip capillary electrophoresis and were >40% of that observed in conventional PCR tubes for chips with and without pillars. Reproducible on-chip PCR was achieved using white blood cell preparations isolated from whole human blood pumped through the chip.

Mutation detection using ligase chain reaction in passivated silicon-glass microchips and microchip capillary electrophoresis.

The ligase chain reaction (LCR) following PCR is one of the most sensitive and specific methods for detecting mutations, especially single nucleotide polymorphisms (SNPs). Performing LCR in microchips remains a challenge because of the inhibitory effect of the internal surfaces of silicon-glass microchips. We have tested a dynamic polymer-based surface passivation method for LCR conducted in oxide-coated silicon-glass microchips. The combination of polyvinylpyrrolidone 40 (PVP-40) at 0.75% (w/v) with an excess of the ligase produced successful LCR in the silicon-glass microchips, with yields of ligated primers comparable to reactions performed in conventional reaction tubes. Ligated primers were detected and quantified simply and conveniently using microchip capillary electrophoresis.

Surface effects on PCR reactions in multichip microfluidic platforms.

We evaluated the compatibility of several common plastics, commercially available plastic tubing and disposable syringes which might be useful in the construction of microfluidic platforms with respect to the polymerase chain reaction (PCR). A simple and inexpensive plastic test module was constructed in order to evaluate some of the construction plastics. We also investigated the effect of addition of PEG 8000 to PCR reaction mixtures on the compatibility of materials. These studies identified several common plastics, plastic tubing, and disposable syringes which were compatible with the PCR reaction.

Increased amplification efficiency of microchip-based PCR by dynamic surface passivation.

Surface passivation is critical for effective PCR using silicon-glass chips. We tested a dynamic polymer-based surface passivation method. Polyethylene glycol 8000 (PEG 8000) or polyvinylpyrrolidone 40 (PVP-40) applied at 0.75% (w/v) in the reaction mixture produced significant surface passivation effects using either native or SiO2-precoated silicon-glass chips. PCR amplification was achieved from human genomic DNA as a template as well as from human lymphocytes. The dynamic surface passivation effect of PEG 8000 remained similar under both conditions. Dynamic surface passivation offers a simple and cost-effective method to make microfabricated silicon-glass chips PCR friendly. It can also be used in combination with static passivation (silicon oxide surface layer) to further improve PCR performance using silicon-glass PCR chips.

Genotyping on a thermal gradient DNA chip.

Silicon-based chips with discrete, independently temperature-controlled islands have been developed for use in DNA microarray hybridization studies. Each island, containing a heater made of a diffusion layer and a temperature sensor based on a p-n junction, is created on a silicon dioxide/nitride surface by anisotropic etching. Different reactive groups are subsequently added to the surface of the islands, and allele-specific oligonucleotide probes are attached to discrete spots on the chip. Hybridization is performed with Cy5-tagged single-stranded targets derived by PCR from genomic DNA. Results are assessed by measuring fluorescence of bound dye-tagged targets after hybridization and washing. Temperatures at each island can be set at different values to obtain optimal distinction between perfect matches and mismatches. This approach facilitates definition of optimal temperatures for probe/target annealing and for distinction between perfectly matched versus mismatched solution-phase targets. The thermal gradient DNA chips were then tested for genotyping, and the results for four different loci in two genes are presented. Unambiguous typing was achieved for clinically relevant loci within the factor VII and hemochromatosis genes.

Fabrication of plastic microchips by hot embossing.

Plastic microchips with microchannels (100 microm wide, 40 microm deep) of varying designs have been fabricated in polymethylmethacrylate by a hot embossing process using an electroform tool produced starting with silicon chip masters. Hot-embossed chips were capped with a polymethylmethacrylate top using a proprietary solvent bonding process. Holes were drilled through the top of the chip to allow access to the channels. The chips were tested with fluid and shown to fill easily. The seal between the top of the chip and the hot embossed base was effective, and there was no leakage from the channels when fluid was pumped through the microchannels. The chips were also tested with a semen sample and the plastic chip performed identically to the previous silicon-glass and glass versions of the chip. This microfabrication technique offers a viable and potentially high-volume low cost production method for fabricating transparent microchips for analytical applications.