COVID-19 infection in patients with intestinal failure: UK experience.

BACKGROUND: The direct effect of the coronavirus disease 2019 (COVID-19) pandemic on patients with intestinal failure (IF) has not been described. METHODS: We conducted a nationwide study of UK IF centers to evaluate the infection rates, presentations, and outcomes in patients with types 2 and 3 IF. RESULTS: A total of 45 patients with IF contracted COVID-19 between March and August 2020; this included 26 of 2191 (1.2%) home parenteral nutrition (HPN)-dependent adults and 19 of 298 (6.4%) adults hospitalized with type 2 IF. The proportion of patients receiving nursing care for HPN administration was higher in those with community-acquired COVID-19 (66.7%) than the proportion in the entire HPN cohort (26.1%; P < .01). Two HPN-dependent and 1 hospitalized patient with type 2 IF died as a direct consequence of the virus (6.7% of 45 patients with types 2 or 3 infected). CONCLUSION: This is the first study to describe the outcomes of COVID-19 in a large cohort of patients requiring long-term PN. Methods to reduce hospital and community nosocomial spread would likely be beneficial.

UK practice in the prevention of central venous catheter-associated thrombosis in adults on home parenteral nutrition.

BACKGROUND: Maintaining central access is imperative for the delivery of home parenteral nutrition (HPN) in those with intestinal failure. Methods to reduce central venous catheter infection are well recognised; however, the prevention of line thrombosis is less well studied. METHODS: This paper reviews the current evidence and reports a survey of current practice within the UK. Using an electronic survey, respondents were asked to detail their use of anticoagulation in different patient groups and the type of anticoagulation used. RESULTS: 41 replies were received from 31 centres. Only one responder used low-dose warfarin routinely; 80% however anticoagulated those with a previous line thrombosis and 65% anticoagulated those that had any deep vein thrombosis or pulmonary embolus. The most commonly used anticoagulant was dose-adjusted warfarin aiming for an international normalised ratio of 2-3. CONCLUSIONS: The evidence from the current literature in both HPN and the wider field is that there is no clear evidence that anticoagulation is either beneficial or harmful in the prevention of line thrombosis. This survey suggested that practice is varied across the UK likely reflecting the lack of evidence within the current literature.

Electrochemical control of a DNA Holliday Junction nanoswitch by Mg2+ ions.

The molecular conformation of a synthetic branched, 4-way DNA Holliday junction (HJ) was electrochemically switched between the open and closed (stacked) conformers. Switching was achieved by electrochemically induced quantitative release of Mg(2+) ions from the oxidised poly(N-methylpyrrole) film (PPy), which contained polyacrylate as an immobile counter anion and Mg(2+) ions as charge compensating mobile cations. This increase in the Mg(2+) concentration screened the electrostatic repulsion between the widely separated arms in the open HJ configuration, inducing switching to the closed conformation. Upon electrochemical reduction of PPy, entrapment of Mg(2+) ions back into the PPy film induced the reverse HJ switching from the closed to open state. The conformational transition was monitored using fluorescence resonance energy transfer (FRET) between donor and acceptor dyes each located at the terminus of one of the arms. The demonstrated electrochemical control of the conformation of the used probe-target HJ complex, previously reported as a highly sequence specific nanodevice for detecting of unlabelled target [Buck, A.H., Campbell, C.J., Dickinson, P., Mountford, C.P., Stoquert, H.C., Terry, J.G., Evans, S.A.G., Keane, L., Su, T.J., Mount, A.R., Walton, A.J., Beattie, J.S., Crain, J., Ghazal, P., 2007. Anal. Chem., 79, 4724-4728], allows the development of electronically addressable DNA nanodevices and label-free gene detection assays.

Molecular recognition with DNA nanoswitches: effects of single base mutations on structure.

This paper investigates the properties of a simple DNA-based nanodevice capable of detecting single base mutations in unlabeled nucleic acid target sequences. Detection is achieved by a two-stage process combining first complementary-base hybridization of a target and then a conformational change as molecular recognition criteria. A probe molecule is constructed from a single DNA strand designed to adopt a partial cruciform structure with a pair of exposed (unhybridized) strands. Upon target binding, a switchable cruciform construct (similar to a Holliday junction) is formed which can adopt open and closed junction conformations. Switching between these forms occurs by junction folding in the presence of divalent ions. It has been shown from the steady-state fluorescence of judiciously labeled constructs that there are differences between the fluorescence resonance energy transfer (FRET) efficiencies of closed forms, dependent on the target sequence near the branch point, where the arms of the cruciform cross. This difference in FRET efficiency is attributed to structural variations between these folded junctions with their different branch point sequences arising from the single base mutations. This provides a robust means for the discrimination of single nucleotide mismatches in a specific region of the target. In this paper, these structural differences are analyzed by fitting observed time-resolved donor fluorescence decay data to a Gaussian distribution of donor-acceptor separations. This shows the closest mean separation (approximately 40 A) for the perfectly matched case, whereas larger separations (up to 50 A) are found for the single point mutations. These differences therefore indicate a structural basis for the observed FRET differences in the closed configuration which underpins the operation of these devices as biosensors capable of resolving single base mutations.

Time-resolved FRET and FLIM of four-way DNA junctions.

Conformational transitions in a 4-way DNA junction when titrated with ionic solutions are studied using time-resolved fluorescence resonance energy transfer. Parameters characterising the transition in terms of critical ion concentration (c1/2) and the Hill coefficient for ion binding are obtained by fitting a simple two-state model using steady-state spectra. Data obtained from a fluorescence lifetime plate reader and analysed by fitting a single exponential to donor fluorescence lifetime decays are shown to be in good agreement with the parameters obtained from steady-state measurements. Fluorescence lifetimes, however, offer advantages, particularly in being independent of fluorophore concentration, output intensity, inhomogeneity in the excitation source and output wavelength. We demonstrate preliminary FRET-FLIM images of DNA junction solutions obtained using a picosecond gated CCD which are in agreement with results from a fluorescence lifetime plate reader. The results suggest that time-resolved FRET-FLIM is sensitive to subtle structural changes and may be useful in assays based on 4-way DNA junctions.

The stability and characteristics of a DNA Holliday junction switch.

A Holliday junction (HJ) consists of four DNA double helices, with a branch point discontinuity at the intersection of the component strands. At low ionic strength, the HJ adopts an open conformation, with four widely spaced arms, primarily due to strong electrostatic repulsion between the phosphate groups on the backbones. At high ionic strength, screening of this repulsion induces a switch to a more compact (closed) junction conformation. Fluorescent labelling with dyes placed on the HJ arms allows this conformational switch to be detected optically using fluorescence resonance energy transfer (FRET), producing a sensitive fluorescent output of the switch state. This paper presents a systematic and quantitative survey of the switch characteristics of such a labelled HJ. A short HJ (arm length 8 bp) is shown to be prone to dissociation at low switching ion concentration, whereas an HJ of arm length 12 bp is shown to be stable over all switching ion concentrations studied. The switching characteristics of this HJ have been systematically and quantitatively studied for a variety of switching ions, by measuring the required ion concentration, the sharpness of the switching transition and the fluorescent output intensity of the open and closed states. This stable HJ is shown to have favourable switch characteristics for a number of inorganic switching ions, making it a promising candidate for use in nanoscale biomolecular switch devices.

Unsupervised feature dimension reduction for classification of MR spectra.

We present an unsupervised feature dimension reduction method for the classification of magnetic resonance spectra. The technique preserves spectral information, important for disease profiling. We propose to use this technique as a preprocessing step for computationally demanding wrapper-based feature subset selection. We show that the classification accuracy on an independent test set can be sustained while achieving considerable feature reduction. Our method is applicable to other classification techniques, such as neural networks, support vector machines, etc.

Diagnosis and prognosis of breast cancer by magnetic resonance spectroscopy of fine-needle aspirates analysed using a statistical classification strategy.

BACKGROUND: The aim was to develop robust classifiers to analyse magnetic resonance spectroscopy (MRS) data of fine-needle aspirates taken from breast tumours. The resulting data could provide computerized, classification-based diagnosis and prognostic indicators. METHODS: Fine-needle aspirate biopsies obtained at the time of surgery for both benign and malignant breast diseases were analysed by one-dimensional proton MRS at 8.5 Tesla. Diagnostic correlation was performed between the spectra and standard pathology reports, including the presence of vascular invasion by the primary cancer and involvement of the excised axillary lymph nodes. RESULTS: Malignant tissue was distinguished from benign lesions with an overall accuracy of 93 per cent. From the same spectra, lymph node involvement was predicted with an overall accuracy of 95 per cent, and tumour vascular invasion with an overall accuracy of 94 per cent. CONCLUSION: The pathology, nodal involvement and tumour vascular invasion were predicted by computerized statistical classification of the proton MRS spectrum from a fine-needle aspirate biopsy taken from the primary breast lesion.

Identification of Enterococcus, Streptococcus, and Staphylococcus by multivariate analysis of proton magnetic resonance spectroscopic data from plate cultures.

A new fingerprinting technique with the potential for rapid identification of bacteria was developed by combining proton magnetic resonance spectroscopy ((1)H MRS) with multivariate statistical analysis. This resulted in an objective identification strategy for common clinical isolates belonging to the bacterial species Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus agalactiae, and the Streptococcus milleri group. Duplicate cultures of 104 different isolates were examined one or more times using (1)H MRS. A total of 312 cultures were examined. An optimized classifier was developed using a bootstrapping process and a seven-group linear discriminant analysis to provide objective classification of the spectra. Identification of isolates was based on consistent high-probability classification of spectra from duplicate cultures and achieved 92% agreement with conventional methods of identification. Fewer than 1% of isolates were identified incorrectly. Identification of the remaining 7% of isolates was defined as indeterminate.

Cryptococcomas distinguished from gliomas with MR spectroscopy: an experimental rat and cell culture study.

PURPOSE: To use magnetic resonance (MR) spectroscopy to characterize clinical isolates of Cryptococcus neoformans and a glioma cell line in culture and in experimental rats. MATERIALS AND METHODS: One- and two-dimensional hydrogen 1 MR spectra were acquired from fungi cultured in vitro (16 isolates of C neoformans, three of Candida albicans, three of Aspergillus fumigatus, three of Saccharomyces cerevisiae) and a C6 glioma cell line. Cerebral biopsy specimens were obtained from healthy rats and animals with experimental infections or gliomas (19 healthy brains, 20 cryptococcomas, and 19 gliomas). Unequivocal signal assignment was performed for cell suspensions and tissue samples by using homo- and heteronuclear two-dimensional correlation spectra. RESULTS: MR spectra of C neoformans and cerebral cryptococcomas--but not of other fungi, healthy brains, or gliomas--were dominated by resonances from the cytosolic disaccharide alpha,alpha-trehalose. This spectral pattern was different from that of gliomas, which was dominated by lipids and an increased choline-creatine ratio, and that of healthy brain. CONCLUSION: A remarkably high concentration of alpha,alpha-trehalose in relation to other metabolites that are visible with MR spectroscopy is diagnostic of C neoformans. Cerebral cryptococcomas are an uncommon but serious manifestation of cryptococcosis in humans. Application of these results to the noninvasive diagnosis of cerebral cryptococcomas would help reduce the risk and expense of unnecessary surgery or biopsy and expedite patient treatment.

Fine-needle biopsy specimens of benign breast lesions distinguished from invasive cancer ex vivo with proton MR spectroscopy.

PURPOSE: To determine whether invasive breast cancer can be distinguished from benign lesions with proton magnetic resonance (MR) spectroscopy ex vivo on the basis of altered cellular chemistry. MATERIALS AND METHODS: Two hundred eighteen fine-needle biopsy specimens were obtained in 191 patients undergoing surgery and were analyzed with proton MR spectroscopy. MR spectroscopic and histopathologic findings were compared. RESULTS: Invasive carcinoma produced increased signal at 3.25 ppm, attributable to choline-containing metabolites. Discrimination between invasive carcinoma (n = 82), benign lesions (n = 106), or carcinoma in situ (n = 17) was based on the resonance intensity at 3.25 ppm standardized to the resonance at 3.05 ppm (P < .001). The ratio of peak height intensities of resonances at 3.25 to those at 3.05 ppm was less than 1.7 in 102 of the 106 normal or benign lesions. All carcinoma in situ specimens with comedonecrosis or a microinvasive component (n = 6) were categorized at MR spectroscopy with invasive carcinoma, while others with in situ disease alone were categorized with benign lesions (n = 11). The sensitivity and specificity of MR spectroscopy in fine-needle biopsy specimens in distinguishing benign lesions from invasive cancer were 95% and 96%, respectively. CONCLUSION: Proton MR spectroscopy of fine-needle biopsy specimens provides objective diagnostic information that complements findings of conventional preoperative investigations of breast lesions.

Cancer pathology in the year 2000.

The last one hundred and fifty years has produced the mature and sophisticated discipline of histopathology, yet still leaves the diagnosis of human cancer, by the best available technique, as more art than science. Proton magnetic resonance spectroscopy (1H MRS) ex vivo identifies the chemical markers of established pathobiological disorders within excised biopsies and fine needle aspirates, in particular, those associated with the development and progression of malignant disease. Alterations to cellular chemistry monitored by 1H MRS allows distinction between invasive and pre-invasive lesions of the uterine cervix, and separate truly benign follicular neoplasms from follicular carcinomas on analysis of fine needle aspirates containing as few as 10(6) cells. 1H chemical shift imaging (CSI) determines the spatial location of these chemical changes and provides insight into the chemistry of neoplastic transformation. It is our hypothesis that, by the year 2000, CSI will aid image guided biopsy techniques and that correlation of biopsy histology with in vivo localised 1H MRS data will: (a) lead to improved assessment of the extent of malignant disease and (b) establish the sensitivity and specificity of in vivo 1H MRS for the simultaneous determination of the size, location and neoplastic potential of a tumour mass.

A proton magnetic resonance spectroscopy study of aging and transformed human fibroblasts.

Proton magnetic resonance spectroscopy (1H MRS) has been used to monitor changes occurring during aging and transformation in human lung fibroblasts. Aging was studied in MRC-5 cells from nonsenescent (early passage) to presenescent (late passage) and senescence. Nonsenescent cells infected with SV40 virus (pretransformed) were monitored through crisis and subsequent immortalization. Aging changes were observed with one- and two-dimensional MR spectra. Cholesterol and lipid resonances were significantly increased from nonsenescent cultures to senescence. These changes could be caused by chemical or structural changes in the plasma membrane or in intracellular lipid pools. In contrast, choline levels rose from nonsenescent to presenescent cells but at senescence dropped to that of nonsenescent cells. Increased choline levels are often associated with increased cellular proliferation. After SV40 infection of MRC-5 cells there was an increase of cholesterol and lipid levels that peaked at crisis. Newly immortalized cells exhibited a drop in cholesterol and lipid to nonsenescent cell levels, but these rose again in established immortalized cells. In contrast to presensescent cultures, the levels of choline gradually increased from pretransformed to crisis phase but still continued to rise after immortalization. Thus, 1H MRS illustrates similarities in lipid behavior at senescence and crisis, whereas the choline levels are different.

Two-dimensional proton magnetic resonance spectroscopy for tissue characterization of thyroid neoplasms.

We have previously demonstrated that one dimensional (1D) proton (1H) magnetic resonance spectroscopy (MRS) can distinguish normal thyroid tissue from thyroid carcinoma using a spectral ratio of peak intensity at 1.7 ppm/0.9 ppm. Two dimensional (2D) 1H-MRS allows identification of specific molecules that have overlapping peaks in the 1D-MR spectrum. Specimens from 93 consecutive thyroid nodules were examined using 2D 1H-MRS on a Bruker AM-360 wide-bore spectrometer. There was a progressive increase in lipid cross peaks assigned to di-/triglycerides when comparing colloid/hyperplastic nodules to follicular adenoma, and adenoma to carcinoma. A specific cross peak attributable to cholesterol/cholesteryl esters was commonly seen in carcinomas. In contrast, two unassigned cross peaks unique to the thyroid were more prevalent in benign lesions. There was an overall increase in cross peaks attributable to cell surface fucosylation in carcinoma when compared to benign lesions, although the fucose spectral pattern was not specific for cancer. On this basis, a spectral ratio of peak intensity at 2.05 ppm/0.9 ppm more clearly distinguished benign follicular adenoma from carcinoma. 2D 1H-MRS thus identifies chemical changes that allow more specific tissue characterization of thyroid neoplasms.

Tetraphenylphosphonium chloride induced MR-visible lipid accumulation in a malignant human breast cell line.

The effect of the cationic lipophilic phosphonium salt tetraphenylphosphonium chloride (TPP) on a human malignant breast cell line, DU4475, was monitored with proton nuclear magnetic resonance (1H MRS). TPP caused a dose- and time- dependent increase in resonances arising from MR-visible lipid as measured by the CH2/CH3 ratio in the 1-dimensional 1H MR spectrum. Two-dimensional MRS identified increases in the glycerophosphocholine/lysine cross-peak ratio and corresponding decreases in the phosphocholine/lysine ratio in a dose- dependent fashion in TPP-treated cells. Lipid metabolic changes are discussed in the light of other MR experiments, and the data indicate that accumulation of MR-visible lipids may arise from the rearrangement of phospholipids accompanying mitochondrial destruction or from the catabolism of phospholipids associated with early events in the cytotoxic process.

Chemical shift imaging of human colorectal tissue (ex vivo).

The spatial location of MR visible lipid in the wall of the normal human colon, and in carcinomatous colonic tissue has been documented using proton chemical shift imaging, one- and two-dimensional magnetic resonance spectroscopy and histochemical staining. Following dissection of the mucosal and submucosal layers of normal colon, these techniques showed high levels of neutral lipid distributed in the submucosal layer. Relatively less lipid was observed in the mucosal layer. Histochemical staining confirmed that the majority of the neutral lipid was in the submucosa, extracellular, and in the lymphatic channels. Carcinomatous tissue gave a variable lipid signal which histochemical staining identified as being from tumour stroma, necrotic and degenerate tumour cells and macrophages.

Human cancers detected by proton MRS and chemical shift imaging ex vivo.

Proton magnetic resonance spectroscopy (1H MRS) has the potential to become a diagnostic adjunct for the detection and grading of human neoplastic disease. This paper describes the use of proton MRS to document changes arising in the lipid chemistry of biopsies arising from the human uterine cervix, thyroid and colon and demonstrates the diagnostic power of ex vivo spectroscopy. Proton chemical shift imaging (CSI) is further used to determine the spatial location of lipid changes in ex vivo human biopsy specimens and provides insight into the chemistry of neoplastic transformation.