The feasibility of micro-ultrasound as a tool to image peripheral nerves.

Despite widespread use of ultrasound imaging to guide needle placement, the incidence of transient and permanent nerve damage as a complication of regional anaesthesia has not changed over the last decade. In view of the controversy surrounding intraneural injection there is a need to understand the structural changes caused by subepineural and subperineural needle penetration. Clinical ultrasound machines do not provide adequate anatomical resolution, and anaesthetists have difficulty judging the precise location of the needle tip relative to the epineurium. We studied the suitability of micro-ultrasound imaging (which offers anatomical resolution better than 100 µm) as a tool for viewing neural anatomy and deformation caused by needle insertion. The primary objective was to assess micro-ultrasound imaging as a method to view fascicles within nerves resected from fresh and soft-embalmed cadavers. Secondary objectives were to observe any disruption of the neural anatomy caused by anaesthetic needle penetration, and to assess the integrity of fresh and Thiel method soft-embalmed nerves, after handling and during needle insertion using ultrasound images and histology. We imaged nine nerves from the left and right sides of fresh and soft-embalmed cadavers. A regional block needle was inserted into three median nerves. We identified fascicles > 0.4 mm in width using micro-ultrasound. Subepineural needle placement was associated with denting, rotation and elastic deformation of fascicles, whereas subperineural needle insertion split fascicles permanently.

Physical properties and functional alignment of soft-embalmed Thiel human cadaver when used as a simulator for ultrasound-guided regional anaesthesia.

BACKGROUND: We evaluated the physical properties and functional alignment of the soft-embalmed Thiel cadaver as follows: by assessing tissue visibility; by measuring its acoustic, mechanical and elastic properties; by evaluating its durability in response to repeated injection; and by aligning images with humans. METHODS: In four soft-embalmed Thiel cadavers, we conducted three independent studies. We assessed the following factors: (i) soft tissue visibility in a single cadaver for 28 weeks after embalming; (ii) the displacement of tissues in response to 1 and 5 ml interscalene and femoral nerve blocks in a single cadaver; and (iii) the stiffness of nerves and perineural tissue in two cadavers. We aligned our findings with ultrasound images from three patients and one volunteer. Durability was qualified by assessing B-mode images from repetitive injections during supervised training. RESULTS: There was no difference in visibility of nerves between 2 and 28 weeks after embalming {geometric mean ratio 1.13 [95% confidence interval (CI): 0.75-1.68], P=1.0}. Mean tissue displacement was similar for cadaver femoral and interscalene blocks [geometric mean ratio 1.02 (95% CI: 0.59-1.78), P=0.86], and for 1 and 5 ml injection volumes [geometric mean ratio 0.84 (95% CI: 0.70-1.01), P=0.19]. Cadavers had higher intraneural than extraneural stiffness [Young's modulus; geometric mean ratio 3.05 (95% CI: 2.98-3.12), P<0.001] and minimal distortion of anatomy when conducting 934 left-sided interscalene blocks on the same cadaver throughout a 10 day period. CONCLUSIONS: The soft-embalmed Thiel cadaver is a highly durable simulator that has excellent physical and functional properties that allow repeated injection for intensive ultrasound-guided regional anaesthesia training.

Trainee anaesthetist diagnosis of intraneural injection-a study comparing B-mode ultrasound with the fusion of B-mode and elastography in the soft embalmed Thiel cadaver model.

BACKGROUND: The incidence of intraneural injection during trainee anaesthetist ultrasound guided nerve block varies between 16% in experts and up to 35% in trainees. We hypothesized that elastography, an ultrasound-based technology that presents colour images of tissue strain, had the potential to improve trainee diagnosis of intraneural injection during UGRA, when integrated with B-Mode ultrasound onto a single image. METHODS: We recorded 40 median nerve blocks randomly allocated to 0.25 ml, 0.5 ml, 1 ml volumes to five sites, on both arms of two soft embalmed cadavers, using a dedicated B-Mode ultrasound and elastography transducer. We wrote software to fuse elastogram and B-Mode videos, then asked 20 trainee anaesthetists whether injection was intraneural or extraneural when seeing B-Mode videos, adjacent B-Mode and elastogram videos, fusion elastography videos or repeated B-Mode ultrasound videos. RESULTS: Fusion elastography improved the diagnosis of intraneural injection compared with B-Mode ultrasound, Diagnostic Odds Ratio (DOR) (95%CI) 21.7 (14.5 - 33.3) vs DOR 7.4 (5.2 - 10.6), P < 0.001. Compared with extraneural injection, intraneural injection was identified on fusion elastography as a distinct, brighter translucent image, geometric ratio 0.33 (95%CI: 0.16 - 0.49) P < 0.001. Fusion elastography was associated with greater trainee diagnostic confidence, OR (95%CI) 1.89 (1.69 - 2.11), P < 0.001, and an improvement in reliability, Kappa 0.60 (0.55 - 0.66). CONCLUSIONS: Fusion elastography improved the accuracy, reliability and confidence of trainee anaesthetist diagnosis of intraneural injection.

Translation of sonoelastography from Thiel cadaver to patients for peripheral nerve blocks.

Ultrasound guidance is now common in regional anaesthesia practice, but remains limited by poor visibility of the needle tip and poor quantification of local anaesthetic spread. Sonoelastography based on tissue compression is a technique depicting tissue strain. Hitherto used largely for tumour diagnosis, we used it in both Thiel embalmed cadavers and two patients receiving interscalene and femoral blocks to observe changes in tissue strain during local anaesthetic injection. The primary aim of our study was to measure the area under the curve (weighted for time) of the strain pattern in Thiel perineural tissue when using a range of volumes of embalming fluid (0.25, 0.5, 1, 2.5, 5 and 7.5 ml) for interscalene and femoral blocks using sonoelastography. Our secondary aims were to evaluate static images of anatomy and videos of needle insertion and perineural injection using combined B-Mode ultrasound and sonoelastography. Independent raters assessed the anatomy and spread using a 7-point Likert scale, ranked from extremely poor to extremely good. We performed 83 blocks in cadavers. Concordance between both raters was good, with weighted Kappa (95% CI) 0.66 (0.61-0.71). The characteristics of spread were similar with both interscalene and femoral block; spread increased with injectate volume up to 1 ml. Analysis of variance showed differences in spread between injection volumes (p = 0.009), but not between regional blocks (p = 0.05). Post-hoc analysis showed greater spread with 1 and 2.5 ml volumes compared with 0.25 ml. In patients, visibility of strain during injection was better with sonoelastography than with B-Mode ultrasound and showed a dose response from 1 to 5 ml volumes of local anaesthetic. Colour strain recognition using sonoelastography offers the ability to differentiate between nerve and surrounding tissue during local anaesthetic injection by improving visibility of spread (p = 0.04).

Genetic reprogramming in pathways of colonic cell maturation induced by short chain fatty acids: comparison with trichostatin A, sulindac, and curcumin and implications for chemoprevention of colon cancer.

The short-chain fatty acid butyrate, produced by microbial fermentation of dietary fiber in the large intestine, is a physiological regulator of major pathways of colonic epithelial cell maturation: cell cycle arrest, lineage-specific differentiation, and apoptosis. Microarray analysis of 8,063 sequences demonstrated a complex cascade of reprogramming of SW620 colonic epithelial cells upon treatment with butyrate characterized by the progressive recruitment of gene sets as a function of time. Comparison with the effects of trichostatin A, in conjunction with differences in the kinetics of alteration of histone acetylation induced by butyrate and trichostatin A, identified subsets of induced and repressed genes likely coordinately regulated by altered histone acetylation. The butyrate response was also compared in detail with that of sulindac, a nonsteroidal anti-inflammatory drug with significant chemopreventive activity for colon cancer, and curcumin, a component of mustard and curry structurally and functionally related to sulindac that also has chemopreventive activity. Although gene clusters were identified that showed similar responses to butyrate and sulindac, the data were characterized by the extensive differences in the effects of the two agents. This was striking for functional classes of genes involved in signaling pathways and in cell cycle progression, although butyrate and sulindac induce a similar G0-G1 arrest, elevation of beta-catenin-Tcf signaling, and apoptotic cascade. As regards cell cycle arrest, the underlying mechanism in response to butyrate was most similar to that of the Caco-2 cell line that had spontaneously undergone a G0-G1 arrest and least similar to the G2-M arrest stimulated by curcumin. Thus, high-throughput microarray analysis of gene expression profiles can be used to characterize and distinguish the mechanisms of response of colonic epithelial cells to physiological and pharmacological inducers of cell maturation. This has important implications for characterization of chemopreventive agents and recognition of potential toxicity and synergies. The data bases, gene clusters, and analyses are available at http:// sequence.aecom.yu.edu/genome/.

c-myc/p53 interaction determines sensitivity of human colon carcinoma cells to 5-fluorouracil in vitro and in vivo.

Colon carcinoma cells overexpress c-myc due to defective Wnt signaling, but only patients whose tumors have an amplified c-myc gene show improved disease-free and overall survival in response to 5-fluoruracil (5FU). Here we show that in two colon carcinoma cell lines that do not have an amplified c-myc gene but differ in their p53 status, high c-myc levels can be further elevated by introducing a c-myc expression vector. Whereas sensitivity to low serum-induced apoptosis was imposed on the parental lines independent of p53 status and was unaffected by further elevation of c-myc, sensitivity to 5FU-induced apoptosis was dependent on both the higher c-myc levels due to the expression vector and wild-type p53 function. The elevated c-myc levels led to higher c-myc transactivation activity in the p53 wild-type cell line, but not in the mutant p53 cell line. The requirement for both elevated c-myc and p53 for 5FU sensitivity was confirmed using antisense c-myc and pifithrin-alpha, a specific inhibitor of p53. Finally, the in vitro data predicted that only patients with both amplified c-myc and wild-type p53 in their primary tumors would be responsive to 5FU-based therapy, which was borne out by analysis of tumors from 135 patients entered into a Phase III clinical trial of 5FU-based adjuvant therapy. The data provide significant insight into mechanisms that establish colon tumor cell sensitivity to 5FU, clearly demonstrate the necessity of exercising caution in considering combining novel strategies that target elevated c-myc with standard 5FU-based therapy, and suggest alternative therapeutic strategies that target c-myc and/or p53 mutations in the treatment of colon cancer.

An assessment of the performance of a glass bead sterilizer.

The performance of a simple glass bead sterilizer designed for use with hand held instruments such as in chiropody surgeries was studied and found to be generally within specification. However, we recommend certain precautions during use.

Malfunction of medical equipment as a result of mains borne interference.

Medical equipment has become more intelligent as the manufacturers have incorporated the latest microprocessor based technology. Equipment malfunction can be caused at any time by inherent errors in the control program but it is particularly important that this is designed to cope with the effects of electrical interference which, in addition, may cause corruption of the software. We have considered interference found in the mains supply in the hospital environment. Using a test protocol with appropriate interference simulators, a wide range of medical equipment was removed temporarily from use and its immunity to electrical mains borne interference tested. Battery operated mains rechargeable devices were unaffected by mains voltage variations including drop-outs and sags whereas mains powered devices were affected to varying degrees of severity. In particular, repetitive drop-outs caused loss of power due to fuse blowing in some life support equipment. Impulses affected 25% and pulse bursts 50% of the equipment tested with some evidence that the more recent designs coped better. The EEC Directive on electro-medical compatibility compliance may cause the design of equipment to be improved but hospitals will have to cope with the above problems in their existing equipment for many years to come.

Infection control when servicing medical and laboratory equipment.

The risks of infection in servicing medical and hospital laboratory equipment are reviewed. Recommended procedures and precautions for dealing with these risks are detailed. These are based upon the practice adopted in the authors' own department. A 'permit to work' scheme is not felt to be universally applicable and the guiding criteria to judge any applied procedure are set down.

Automated performance assessment of ultrasound systems using a dynamic phantom.

Quality assurance of medical ultrasound imaging systems is limited by repeatability, difficulty in quantifying results, and the time involved. A particularly interesting approach is demonstrated in the Edinburgh pipe phantom which, with an accompanying mathematical transformation, produces a single figure of merit for image quality from individual measurements of resolution over a range of depths. However, the Edinburgh pipe phantom still requires time-consuming manual scanning, mitigating against its routine use. This paper presents a means to overcome this limitation with a new device, termed the Dundee dynamic phantom, allowing rapid set-up and automated operation. The Dundee dynamic phantom is based on imaging two filamentary targets, positioned by computer control at different depths in a tank of 9.4% ethanol-water solution. The images are analysed in real time to assess if the targets are resolved, with individual measurements at different depths again used to calculate a single figure of merit, in this case for lateral resolution only. Test results are presented for a total of 18 scanners in clinical use for different applications. As a qualitative indication of viability, the figure of merit produced by the Dundee dynamic phantom is shown to differentiate between scanners operating at different frequencies and between a relatively new, higher quality system and an older, lower quality system.

Selectron: rectal dose-rate measurements and modified applicators.

A procedure for estimating the rectal dose-rate in patients treated by a Selectron remote-afterloading machine is described. This entails dose-rate measurements using low-activity test sources whilst the patient is still in theatre. The standard applicators were modified to enable the rectal dose to be reduced.

Molecular mechanisms of action and prediction of response to oxaliplatin in colorectal cancer cells.

The platinum compound oxaliplatin has been shown to be an effective chemotherapeutic agent for the treatment of colorectal cancer. In this study, we investigate the molecular mechanisms of action of oxaliplatin to identify means of predicting response to this agent. Exposure of colon cancer cells to oxaliplatin resulted in G2/M arrest and apoptosis. Immunofluorescent staining demonstrated that the apoptotic cascade initiated by oxaliplatin is characterised by translocation of Bax to the mitochondria and cytochrome c release into the cytosol. Oxaliplatin treatment resulted in caspase 3 activation and oxaliplatin-induced apoptosis was abrogated by inhibition of caspase activity with z-VAD-fmk, but was independent of Fas/FasL association. Targeted inactivation of Bax or p53 in HCT116 cells resulted in significantly increased resistance to oxaliplatin. However, the mutational status of p53 was unable to predict response to oxaliplatin in a panel of 30 different colorectal cancer cell lines. In contrast, the expression profile of these 30 cell lines, assessed using a 9216-sequence cDNA microarray, successfully predicted the apoptotic response to oxaliplatin. A leave-one-out cross-validation approach was used to demonstrate a significant correlation between experimentally observed and expression profile predicted apoptosis in response to clinically achievable doses of oxaliplatin (R=0.53; P=0.002). In addition, these microarray experiments identified several genes involved in control of apoptosis and DNA damage repair that were significantly correlated with response to oxaliplatin.

c-Myc overexpression sensitises colon cancer cells to camptothecin-induced apoptosis.

The proto-oncogene c-Myc is overexpressed in 70% of colorectal tumours and can modulate proliferation and apoptosis after cytotoxic insult. Using an isogenic cell system, we demonstrate that c-Myc overexpression in colon carcinoma LoVo cells resulted in sensitisation to camptothecin-induced apoptosis, thus identifying c-Myc as a potential marker predicting response of colorectal tumour cells to camptothecin. Both camptothecin exposure and c-Myc overexpression in LoVo cells resulted in elevation of p53 protein levels, suggesting a role of p53 in the c-Myc-imposed sensitisation to the apoptotic effects of camptothecin. This was confirmed by the ability of PFT-alpha, a specific inhibitor of p53, to attenuate camptothecin-induced apoptosis. p53 can induce the expression of p21(Waf1/Cip1), an antiproliferative protein that can facilitate DNA repair and drug resistance. Importantly, although camptothecin treatment markedly increased p21(Waf1/Cip1) levels in parental LoVo cells, this effect was abrogated in c-Myc-overexpressing derivatives. Targeted inactivation of p21(Waf1/Cip1) in HCT116 colon cancer cells resulted in significantly increased levels of apoptosis following treatment with camptothecin, demonstrating the importance of p21(Waf1/Cip1) in the response to this agent. Finally, cDNA microarray analysis was used to identify genes that are modulated in expression by c-Myc upregulation that could serve as additional markers predicting response to camptothecin. Thirty-four sequences were altered in expression over four-fold in two isogenic c-Myc-overexpressing clones compared to parental LoVo cells. Moreover, the expression of 10 of these genes was confirmed to be significantly correlated with response to camptothecin in a panel of 30 colorectal cancer cell lines.