Evaluation of whole cell fixation methods for the analysis of nanoscale surface features of Yersinia pestis KIM.

Manipulation of viable Yersinia pestis (etiologic agent of plague) in the laboratory usually necessitates elevated biosafety and biocontainment procedures, even with avirulent or vaccine strains. To facilitate downstream biochemical or physical analyses in a Biosafety Level 1 laboratory environment, effective inactivation without affecting its intrinsic properties is critical. Here, we report on the morphological and biochemical changes to Y. pestis surfaces following four different fixation methods that render the cells nonviable. The results, obtained at the single cell level, demonstrate that methanol inactivation is best able to preserve bacterial morphology and bioactivity, enabling subsequent analysis. This nanoscale evaluation of the effects of inactivation on cell morphology and surface bioactivity may provide a crucial preparatory approach to study virulent pathogens in the lab setting using high-resolution microscopic techniques such as atomic force microscopy.

ASSESSMENT OF ADRENOCORTICAL DYSFUNCTION IN PATIENTS WITH STABLE LIVER CIRRHOSIS.

INTRODUCTION: Relative adrenal insufficiency (RAI) is common in the setting of critical illness as well as in hemodynamically instable cirrhotic patients with sepsis. Several studies have also shown that RAI is frequent in patients with stable cirrhosis without sepsis. The aim of this study was to prospectively assess the incidence of RAI in patients with stable cirrhosis. PATIENTS AND METHODS: Forty-seven patients with hemodynamically stable liver cirrhosis without sepsis were prospectively included. RAI, assessed by using low dose-short Synacthen test (LD-SST), was defined as either a basal total cortisol concentration below 3.6 µg/dL or a peak total serum cortisol ≤ 16 µg/dL at 30 min after stimulation. RESULTS: RAI was present in 10 (21.3%) of 47 cirrhotic patients. Peak cortisol level was negatively correlated with the severity of cirrhosis evaluated by Child-Turcotte-Pugh (CTP) (r=-0.46; P=0.001) and Model for End-Stage Liver Disease (MELD) (r=-0.51; P=0.001) scores. The frequency of RAI increased from CTP-A (10%) to CTP-B (30%) to CTP-C (60%). CONCLUSION: RAI diagnosed by LD-SST is frequent in patients with stable cirrhosis and is related to the severity of liver disease. Further studies are needed to define clinical importance of RAI in stable cirrhotic patients.

Clostridium difficile infection in patients with liver disease: a review.

Over the past two decades, there has been a dramatic worldwide increase in both the incidence and severity of Clostridium difficile infection (CDI). Paralleling the increased incidence of CDI in the general population, there has been increased interest in CDI among patients with liver disease, particularly in those with liver cirrhosis and post liver transplantation. MEDLINE and several other electronic databases from January 1995 to December 2014 were searched in order to identify potentially relevant literature. Patients with cirrhosis and liver transplant recipients are at high risk for the development CDI because of antibiotics and proton pump inhibitors use, frequent and prolonged hospitalization, immunosuppressant therapy, and multiple comorbidities. Enzyme immunoassay to detect C. difficile toxins A and B in stool remains the most widely used test for CDI diagnosis, although, more recently, polymerase chain reaction (PCR)-based assays have become the preferred diagnostic test in many laboratories. Metronidazole and vancomycin, given orally, have proved to be effective in the treatment of CDI. Both cirrhotic patients and liver transplant recipients with CDI have longer length of hospital stay, increased mortality, and higher healthcare costs than those without CDI. A rapid diagnosis and adequate therapy of CDI are of paramount importance to improve liver disease patients' outcome. The aim of this review is to provide up-to-date information on the epidemiology, risk factors, pathogenesis, treatment, and outcomes in liver disease patients with CDI.

Clostridium difficile infection and inflammatory bowel disease: what gastroenterologists and surgeons should know.

Over the past two decades there has been a dramatic increase worldwide in both incidence and severity of Clostridium difficile infection (CDI). Paralleling the rising incidence of CDI in the general population, there has been an even higher increase in the incidence of CDI among patients with inflammatory bowel disease (IBD). CDI may mimic a flare of IBD as symptoms and laboratory parameters are often similar, and therefore, screening for CDI is recommended at every flare in such patients. Enzyme immunoassay to detect Clostridium difficile toxin A and B in stool is still the most widely used test for CDI diagnosis despite its low sensitivity. Metronidazole for mild/moderate CDI,and vancomycin for severe CDI are the preferred agents for the treatment of infection. CDI has a negative impact both on short- and long- term IBD outcomes, increasing the need for surgery, as well as the mortality rate and healthcare costs. All gastroenterologists and surgeons should have a high index of suspicion for CDI when evaluating a patient with IBD flare, as prompt diagnosis and adequate treatment of infection improve outcomes. Measures must be taken to prevent spreading of infection in gastroenterology /surgery settings.

Different profile of peripheral antioxidant enzymes and lipid peroxidation in active and non-active inflammatory bowel disease patients.

BACKGROUND: The role of oxidative stress in inflammatory bowel diseases (IBD) has been extended lately from a simple consequence of inflammation to a potential etiological factor, but the data are still controversial. Active disease has been characterized before by an enhanced production of reactive oxygen species and the increased peroxidation of lipids, but patients in remission were generally not considered different from healthy people in terms of oxidative stress. AIMS: We evaluated the antioxidant defense capacity and lipid peroxidation status in the serum of patients with active and non-active disease compared with healthy matched control subjects. METHODS: The study included 20 patients with confirmed IBD in clinical and biological remission, 21 patients with active disease, and 18 controls. We determined the serum levels of two antioxidant enzymes, superoxide dismutase (SOD) and glutathione peroxidase (GPX), and a lipid peroxidation marker, malondialdehyde (MDA). RESULTS: Active disease patients had an increased activity of both SOD and GPX, as well as significant high values of MDA versus controls. Furthermore, patients being in remission had significantly lower values of antioxidant enzymes (SOD and GPX) and increased lipid peroxidation measured by MDA serum levels, as compared with healthy control subjects. CONCLUSIONS: Our study confirmed the presence of high oxidative stress in active IBD. More importantly, we have demonstrated a lower antioxidant capacity of patients in remission versus control group. This may represent a risk factor for the disease and can be an additional argument for the direct implication of oxidative stress in the pathogenesis of IBD.

Nanoscale spectroscopic imaging of organic semiconductor films by plasmon-polariton coupling.

Tip-enhanced near-field optical images and correlated topographic images of an organic semiconductor film (diindenoperylene, DIP) on Si have been recorded with high optical contrast and high spatial resolution (17 nm) using a parabolic mirror with a high numerical aperture for tip illumination and signal collection. The DIP molecular domain boundaries being one to four molecular layers (1.5-6 nm) high are resolved topographically by a shear-force scanning tip and optically by simultaneously recording the 6x10{5} times enhanced photoluminescence (PL). The excitation is 4x10{4} times enhanced and the intrinsically weak PL-yield of the DIP-film is 15-fold enhanced by the tip. The Raman spectra indicate an upright orientation of the DIP molecules. The enhanced PL contrast results from the local film morphology via stronger coupling between the tip plasmon and the exciton-polariton in the DIP film.

Tailoring gold nanostructures for near-field optical applications.

A method of combined thin-film deposition, electron beam lithography, and ion milling is presented for the fabrication of gold and silver nanostructures. The flexibility of lithographical processes for the variation of geometric parameters is combined with three-dimensional control over the surface evolution. Depending on the etching angle, different shapes ranging from cones over rods to cups can be achieved. These size- and shape-tunable structures present a toolbox for nano-optical investigations. As an example, optical properties of systematically varying structures are examined in a parabolic mirror confocal microscope.

High NA particle- and tip-enhanced nanoscale Raman spectroscopy with a parabolic-mirror microscope.

Detecting efficiently the plasmon-enhanced Raman signal of molecules created in the nanometre-sized gap between a metal nanoparticle or the apex of a sharp tip and a metal surface is the key problem in particle- or tip-enhanced local surface spectroscopy (Pettinger et al., 2004; Roth et al., 2006). The optical excitation field has to be polarized along the gap, and the field emerging from the gap has to be observed from the side. These geometrical restrictions usually limit the numerical aperture of the lens used for exciting the gap and collecting the scattered photons created in the gap. We present a novel method to overcome this problem. The solution is based on a confocal optical microscope with a high numerical aperture parabolic mirror for excitation and detection. Localized plasmons can be efficiently excited parallel to the surface normal by illuminating the parabolic mirror with a radially polarized doughnut mode and the field emerging sidewise from the gap can be efficiently collected by the rim of the parabolic mirror and directed to the detection system. First results on particle- and tip-enhanced Raman spectroscopic measurements of benzotriazole molecules adsorbed on gold films are presented.

All-optical magnetic recording with circularly polarized light.

We experimentally demonstrate that the magnetization can be reversed in a reproducible manner by a single 40 femtosecond circularly polarized laser pulse, without any applied magnetic field. This optically induced ultrafast magnetization reversal previously believed impossible is the combined result of femtosecond laser heating of the magnetic system to just below the Curie point and circularly polarized light simultaneously acting as a magnetic field. The direction of this opto-magnetic switching is determined only by the helicity of light. This finding reveals an ultrafast and efficient pathway for writing magnetic bits at record-breaking speeds.

CORTICOSTEROID TREATMENT IN THE SETTING OF DECOMPENSATED LIVER CIRRHOSIS WITH RELATIVE ADRENAL INSUFFICIENCY: A CASE REPORT AND A BRIEF REVIEW OF THE LITERATURE.

Relative adrenal insufficiency (RAI) is the term used to describe inadequate production or action of glucocorticoids with respect to the severity of the illness. RAI is frequently found in critically ill patients particularly with septic complications and it is also present in both critically ill and stable patients with liver cirrhosis. In the following study a case report of a patient with decompensated cirrhosis and RAI is presented followed by a brief review of the literature. A 65-year-old male with liver cirrhosis of alcoholic etiology was admitted to hospital with bilateral leg edema, ascites, and marked weakness. At admission, his blood pressure was 82/52 mmHg and he had sinus tachycardia of 130/min. Laboratory analysis revealed hyponatremia (122 mmol/L), while ascites fluid analysis showed no infection. During the first 48 hours of hospitalization the patient remained persistently hypotensive despite adequate vascular filling and the addition of noradrenaline. A standard-dose short synacthen test was performed which revealed a poor cortisol response, which is a compatible criterion for the diagnosis of RAI. Intravenous hydrocortisone therapy was initiated, which resulted in a rapid improvement in patient's general condition, and increase in blood pressure. As the patient became hemodynamically stable without the need of noradrenaline, the hydrocortisone dose was weaned progressively, and he was discharged after 18 days of hospitalization in a stable condition.

NONSELECTIVE BETA-BLOCKERS IN PATIENTS WITH CIRRHOSIS: "THE THERAPEUTIC WINDOW".

For over 30 years, nonselective beta-blockers (NSBB) have been successfully used for preventing variceal bleeding in patients with cirrhosis and portal hypertension. Nevertheless, recent studies suggest that NSBB may be effective only within a particular "therapeutic window" in patients with advanced liver disease. Outside of this window, in early stages of cirrhosis and in very advanced cirrhosis, NSBB may be ineffective and even potentially harmful. In this paper we review the beneficial effects and potential harms of beta-blocker therapy in cirrhosis and underline the most recent recommendations for their use in very advanced cases of liver disease.

PLATELET INDICES AND LIVER FIBROSIS EVALUATION IN CHRONIC HEPATITIS C.

UNLABELLED: Recently, several studies have reported that the mean platelet volume and platelet distribution width may give information about liver fibrosis severity in chronic hepatitis C. The aim of the present study was to evaluate whether platelet indices correlate with hepatic fibrosis measured by transient elastography in patients with chronic hepatitis C. MATERIALS AND METHODS: Patients with chronic hepatitis C were prospectively enrolled. Samples for complete blood count and routine biochemical parameters were collected and analyzed in the same day with liver fibrosis assessment by transient elastography. Mean platelet volume, platelet large cell ratio and platelet distribution width were compared with stages of liver fibrosis. Statistical analysis was carried out using SPSS 17.0 software. A P-value of less than 0.05 was considered statistically significant. RESULTS: There were 139 patients with chronic hepatitis C (70.5% males, mean age 54.8 +/- 16.7 years). Compared with mild/moderate liver fibrosis patients, those with advanced liver fibrosis had an increased mean platelet volume (10.4 +/- 0.7 vs. 10.9 +/- 0.9, p < 0.002), platelet large cell ratio (28.5 +/- 5.3 vs. 32.5 +/- 7.2, P < 0.0001), and platelet distribution width (12.8 +/- 1.5 vs. 14.1 +/- 2.7, P = 0.003). CONCLUSIONS: Increased platelet indices were associated with advanced liver fibrosis stages evaluated by transient elastography in patients with chronic hepatitis C.

L-carnitine supplementation does not promote weight loss in ovariectomized rats despite endurance exercise.

In this five-week study, we tested the hypotheses that free access to a maintenance diet supplemented with L-carnitine (L-C) would reduce body fat in adult, sedentary, ovariectomized (OVX) rats, and that there would be an additive effect of L-C on weight reduction in swim-trained animals. As expected, serum carnitine was higher in rats fed the L-C diet, and the OVX-induced weight gain and abdominal fat were counteracted by swimming. L-C supplementation did not reduce the weight gain or abdominal fat in these adult female rats, Moreover, though not reaching statistical significance, rats that were fed L-C demonstrated a tendency for greater weight gain than their basal-fed counterparts despite no difference in energy intake. If the results of this study on ovariectomized rats can be translated to postmenopausal women, moderate intensity exercise may be recommended, but L-C supplementation, with no energy restriction, may be contraindicated as a weight loss method in this cohort.

Randomised clinical trial: alisporivir combined with peginterferon and ribavirin in treatment-naïve patients with chronic HCV genotype 1 infection (ESSENTIAL II).

BACKGROUND: Alisporivir (ALV) is an oral, host-targeting agent with pangenotypic anti-hepatitis C virus (HCV) activity and a high barrier to resistance. AIM: To evaluate efficacy and safety of ALV plus peginterferon-α2a and ribavirin (PR) in treatment-naïve patients with chronic HCV genotype 1 infection. METHODS: Double-blind, randomised, placebo-controlled, Phase 3 study evaluating ALV 600 mg once daily [response-guided therapy (RGT) for 24 or 48 weeks or 48 weeks fixed duration] or ALV 400 mg twice daily RGT with PR, compared to PR alone. Following a Food and Drug Administration partial clinical hold, ALV/placebo was discontinued and patients completed treatment with PR only. At that time, 87% of patients had received ≥12 weeks and 20% had received ≥24 weeks of ALV/PR triple therapy. RESULTS: A total of 1081 patients were randomised (12% cirrhosis, 55% CT/TT IL28B). Addition of ALV to PR improved virological response in a dose-dependent fashion. Overall, sustained virological response (SVR12; primary endpoint) was 69% in all ALV groups vs. 53% in PR control. Highest SVR12 (90%) was achieved in patients treated with ALV 400 mg twice daily and PR for >24 weeks. Seven cases of pancreatitis were reported, with similar frequency between ALV/PR and PR control groups (0.6% vs. 0.8% respectively). Adverse events seen more frequently with ALV/PR than with PR alone were anaemia, thrombocytopenia, hyperbilirubinaemia and hypertension. CONCLUSIONS: Alisporivir, especially the 400 mg twice daily regimen, increased efficacy of PR therapy in treatment-naïve patients with HCV genotype 1 infection. The mechanism of action and pangenotypic activity suggest that alisporivir could be useful in interferon-free combination regimens.

Diurnal variation of Cobb angle measurement in adolescent idiopathic scoliosis.

To investigate the possible influence of gravity on daily variations of curve magnitude in adolescent idiopathic scoliosis, standardized anteroposterior standing radiographs of the spine were taken at 8:00 AM and repeated in the same conditions at 8:00 PM on the same day, 19 girls with idiopathic scoliosis, aged 10-16 years were included. The Cobb angle of the primary thoracic curve was measured blindly by four skilled orthopaedic surgeons. The mean interobserver variation in measuring Cobb angles on the 38 radiographs was 1.6 degrees. The average Cobb angle on morning radiographs was 60 degrees (range, 42-91 degrees) as compared to 65 degrees (range, 47-89 degrees) in the afternoon, a significant (P < 0.001) difference using a paired two-sided Student t test. It was concluded that there is a statistically and clinically significant daily increase of curve severity in moderate to severe idiopathic scoliosis.

Endoscopic intravascular esophageal pressure measurements in cirrhotic patients: response to metoclopramide.

This study reports the effects of metoclopramide (10 mg i.v.) on intravascular esophageal variceal pressure in 12 patients with alcoholic cirrhosis. The esophageal variceal pressure was measured by the direct variceal puncture technique. Metoclopramide caused a reduction in the variceal pressure in 10 out of 12 patients; overall, there was a decrease from 21.5 +/- 5.0 mmHg to 14.0 +/- 3.4 mmHg (p < 0.001). In conclusion, intravenous metoclopramide, which increases lower esophageal sphincter pressure, significantly decreases intravascular variceal pressure in cirrhotic patients.

Small intestine biopotentials in rats after hypokinesia.

To study the effect of hypokinesia on rats small intestine (jejunum and ileum) biopotentials it was first necessary to characterize it. Biopotentials were recorded by intracellular placed microelectrodes from oral and caudal segments of the small intestine. The character of rats small intestine biopotentials differs from that of other species (man, cat, rabbit, dog, e.a.), the slow waves (SW) being smaller and the frequency of basal electrical rhythm higher (31.23 c/min orally and 24.50 caudally). Spike potentials are inscribed on the descending slope of SW but frequently delayed in each successive wave with a regular interval. Hypokinesia obtained by keeping rats in small cages for two weeks create only little changes in intestine biopotentials. The only clear difference was the increase of the slow waves amplitude. The other parameters were not specifically changed.