Anopheles gambiae pilot gene discovery project: identification of mosquito innate immunity genes from expressed sequence tags generated from immune-competent cell lines

Together with AIDS and tuberculosis, malaria is at the top of the list of devastating infectious diseases. However, molecular genetic studies of its major vector, Anopheles gambiae, are still quite limited. We have conducted a pilot gene discovery project to accelerate progress in the molecular analysis of vector biology, with emphasis on the mosquito's antimalarial immune defense. A total of 5,925 expressed sequence tags were determined from normalized cDNA libraries derived from immune-responsive hemocyte-like cell lines. The 3,242 expressed sequence tag-containing cDNA clones were grouped into 2,380 clone clusters, potentially representing unique genes. Of these, 1,118 showed similarities to known genes from other organisms, but only 27 were identical to previously known mosquito genes. We identified 38 candidate genes, based on sequence similarity, that may be implicated in immune reactions including antimalarial defense; 19 of these were shown experimentally to be inducible by bacterial challenge, lending support to their proposed involvement in mosquito immunity.     

Effects of ethanol on the properties of platelets and endothelial cells in model experiments

AIM: To investigate effects of ethanol on activity markers of atherosclerosis in an in vitro endothelial cell model. METHODS: After 24 h incubation with ethanol (0.0095%), human umbilical vein endothelial cells were stimulated for 1 h with lipopolysaccharide, and were then incubated in direct contact with activated platelets. Following this incubation, the expression of CD40L and CD62P on platelets, and the expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), urokinase plasminogen activator receptor (uPAR), and membrane-type 1 matrix metalloproteinase (MT1-MMP) on endothelial cells were measured by flow cytometry. RESULTS: The increased expression of VCAM-1 and uPAR on endothelial cells by proinflammatory stimulation with activated platelets was significantly reduced through pre-incubation with ethanol (P<0.05). Furthermore, platelets in direct contact with ethanol and with endothelial cells pre-incubated in ethanol showed a significant reduction in their CD40L expression (P<0.05). Ethanol had no significant effect on ICAM-1 and MT1-MMP expression on endothelial cells. CONCLUSION: Ethanol directly attenuates platelet activation and has significant endothelial cell-mediated effects on selected markers of atherosclerosis in vitro . These findings underline possible protective effects of ethanol on atherosclerosis.     

Impact of postoperative complications on readmission and long-term survival in patients following surgery for colorectal cancer

Purpose

It is well known that specific postoperative complications such as stroke influence readmissions and overall survival (OS) after surgery for colorectal cancer (CRC). Whether overall hospital morbidity is associated with increased risk of readmission and poorer long-term survival is unknown. New tools are available to accurately quantify overall morbidity, such as the comprehensive complication index (CCI). The aim is to evaluate the impact of complications on readmission and overall survival (OS) in patients operated for colorectal cancer.

Methods

Postoperative complications of patients undergoing surgery for CRC were assessed over a 5-year period using the Clavien-Dindo classification, and overall morbidity was assessed by using the CCI. Individual scores were analyzed regarding their association with readmission and OS by using the multivariate logistic and Cox proportional-hazards regression analysis, respectively.

Results

Two hundred eighty-four patients were operated for CRC, of which 22 (8%) were readmitted. One hundred five patients (37%) developed at least one postoperative complication during the hospital stay. While single complications or the use of severe complication only (grade ≥IIIb) was not associated with readmission, overall morbidity (CCI) predicted readmission (OR 1.02 (95% CI 1.0–1.04), p = 0.044). Similarly, morbidity assessed by the CCI had a significant negative predictive value on OS, e.g., patients with a CCI of 20 were 22% more likely to die within a 5-year follow-up, when compared to patients with a CCI of 10 (p = 0.022).

Conclusions

Overall combined morbidity as assessed by the CCI leads to more frequent readmission, and is associated with poorer long-term survival after surgery for CRC.

     

Are two‐stage hepatectomies associated with more complications than one‐stage procedures?

BackgroundTwo-stage liver resections with portal vein occlusion have become standard in patients with low volume future liver remnants. Whether they are associated with more complications is unclear. The aim of this study was to compare complications of one- and two-stage resections in a retrospective study.MethodsPatients with two-stage right liver resections with a previous portal vein occlusion were compared with patients with one-stage right liver resections between 2002 and 2010. Primary endpoints were the incidence of complications by severity. Secondary endpoints were mortality, post-operative liver- and kidney function tests, length of hospitalization and transfusion events. Logistic and linear regression analyses were performed to adjust for confounders.ResultsThe groups were comparable except for right trisectionectomies, pre-operative chemotherapy and underlying liver disease. Overall complications occurred in 25 out of 35 patients with two-stage and 106 out of 163 in one-stage procedures. Severe complications were observed in 47 out of 163 patients versus 9 out of 35 patients, respectively. Two-stage procedures had no increased adjusted risk for complications [relative risk (RR) 0.9, P = 0.79]. Mortality (5.7% versus 3.7%) and post-operative liver failure rates (2.9% versus 3.1%) were low. Secondary endpoints showed no adjusted differences in risk.ConclusionThis study suggests that liver resections in two stages are not associated with more post-operative complications than one-stage resections. These results should support the adoption of two-stage liver resections in selected patients.     

N400-like magnetoencephalography responses modulated by semantic context,word frequency,and lexical class in sentences

Words have been found to elicit a negative potential at the scalp peaking at approximately 400 ms that is strongly modulated by semantic context. The current study used whole-head magnetoencephalography (MEG) as male subjects read sentences ending with semantically congruous or incongruous words. Compared with congruous words, sentence-terminal incongruous words consistently evoked a large magnetic field over the left hemisphere, peaking at approximately 450 ms. Source modeling at this latency with conventional equivalent current dipoles (ECDs) placed the N400 m generator in or near the left superior temporal sulcus. A distributed solution constrained to the cortical surface suggested a sequence of differential activation, beginning in Wernicke's area at approximately 250 ms, spreading to anterior temporal sites at approximately 270 ms, to Broca's area by approximately 300 ms, to dorsolateral prefrontal cortices by approximately 320 ms, and to anterior orbital and frontopolar cortices by approximately 370 ms. Differential activity was exclusively left-sided until >370 ms, and then involved right anterior temporal and orbital cortices. At the peak of the N400 m, activation in the left hemisphere was estimated to be widespread in the anterior temporal, perisylvian, orbital, frontopolar, and dorsolateral prefrontal cortices. In the right hemisphere, the orbital, as well as, weakly, the right anterior temporal cortices were activated. Similar but weaker field patterns were evoked by intermediate words in the sentences, especially to low-frequency words occurring in early sentence positions where there is little preceding context. The locations of the N400 m sources identified with the distributed solution correspond well with those previously demonstrated with direct intracranial recordings, and suggested by functional magnetic resonance imaging (fMRI). These results help identify a distributed cortical network that supports online semantic processing.     

Gender differences in cutaneous vascular and autonomic nervous response to local cooling

To investigate gender differences in cutaneous vascular reactivity to local cooling we performed cold exposure of one hand and measured laser Doppler (LD) flux on fingers ipsilaterally and contralaterally in a group of 10 healthy females and a group of 10 healthy males. The females were tested twice: in the early follicular and in the mid-luteal phase of the menstrual cycle. We related the characteristics of cutaneous vascular responses to indices of autonomic nervous system activity (heart rate, blood pressure, heart rate variability) at rest and during the cold challenge. In our experimental settings females exhibited greater cutaneous vascular response to local cooling at the ipsilateral site (LD flux decreased to 48.6 ± 6.2 % of the resting value), as compared to males (LD flux decreased to 71.9 ± 6.2 % of the resting value) as well as at the contralateral site (LD flux decreased to 68.2 ± 6.1 % in females and to 85.6 ± 3.8 % in males) (p < 0.05, Dunnett's test). The more pronounced cutaneous vascular response in females seems consistent with the finding of their lower sympathetic activity at rest as well as with their heart rate variability indices of greater sympathetic system reactivity to local cold exposure. Correspondingly, males showed a higher level of sympathetic nervous system activity at rest and a predominant reactivity of the parasympathetic system during local cooling at 15 °C. The females in our study, all of whom were premenopausal, exhibited intra-menstrual cycle variability only in cutaneous vascular response at the site of local cooling (LD flux decreased during the early follicular phase to 48.6 ± 6.2 % and during the mid-luteal phase to 29.2 ± 3.2 % of the resting value, p < 0.05, paired t-test), but not contralaterally. In addition, we found no intra-menstrual cycle differences in the indices of autonomic nervous system reactivity. Received: 18 October 2002, Accepted: 25 March 2003 Correspondence to Ksenija Cankar, DMD, PhD     

Visualization of the herpes simplex virus portal in situ by cryo-electron tomography

Herpes simplex virus type 1 (HSV-1), the prototypical herpesvirus, has an icosahedral nucleocapsid surrounded by a proteinaceous tegument and a lipoprotein envelope. As in tailed bacteriophages, the icosahedral symmetry of the capsid is broken at one of the 12 vertices, which is occupied by a dodecameric ring of portal protein, UL6, instead of a pentamer of the capsid protein, UL19. The portal ring serves as a conduit for DNA entering and exiting the capsid. From a cryo-EM reconstruction of capsids immuno-gold-labeled with anti-UL6 antibodies, we confirmed that UL6 resides at a vertex. To visualize the portal in the context of the assembled capsid, we used cryo-electron tomography to determine the three-dimensional structures of individual A-capsids (empty, mature capsids). The similarity in size and overall shape of the portal and a UL19 pentamer--both are cylinders of approximately 800 kDa--combined with residual noise in the tomograms, prevented us from identifying the portal vertices directly; however, this was accomplished by a computational classification procedure. Averaging the portal-containing subtomograms produced a structure that tallies with the isolated portal, as previously reconstructed by cryo-EM. The portal is mounted on the outer surface of the capsid floor layer, with its narrow end pointing outwards. This disposition differs from that of known phage portals in that the bulk of its mass lies outside, not inside, the floor. This distinction may be indicative of divergence at the level of portal-related functions other than its role as a DNA channel.