Age-Dependent Changes in the Sphingolipid Composition of Mouse CD4+ T Cell Membranes and Immune Synapses Implicate Glucosylceramides in Age-Related T Cell Dysfunction

To determine whether changes in sphingolipid composition are associated with age-related immune dysfunction, we analyzed the core sphingolipidome (i.e., all of the metabolites through the first headgroup additions) of young and aged CD4⁺ T cells. Since sphingolipids influence the biophysical properties of membranes, we evaluated the compositions of immune synapse (IS) and non-IS fractions prepared by magnetic immuno-isolation. Broadly, increased amounts of sphingomyelins, dihydrosphingomyelins and ceramides were found in aged CD4⁺ T cells. After normalizing for total sphingolipid content, a statistically significant decrease in the molar fraction of glucosylceramides was evident in both the non-IS and IS fractions of aged T cells. This change was balanced by less dramatic increases in the molar fractions of sphingomyelins and dihydrosphingomyelins in aged CD4⁺ T cells. In vitro, the direct or enzymatic enhancement of ceramide levels decreased CD4⁺ T cell proliferation without regard for the age of the responding T cells. In contrast, the in vitro inhibition of glucosylceramidase preferentially increased the proliferation of aged CD4⁺ T cells. These results suggest that reductions in glucosylceramide abundance contribute to age-related impairments in CD4⁺ T cell function.     

An Overview of the Genetic Structure within the Italian Population from Genome-Wide Data

In spite of the common belief of Europe as reasonably homogeneous at genetic level, advances in high-throughput genotyping technology have resolved several gradients which define different geographical areas with good precision. When Northern and Southern European groups were considered separately, there were clear genetic distinctions. Intra-country genetic differences were also evident, especially in Finland and, to a lesser extent, within other European populations. Here, we present the first analysis using the 125,799 genome-wide Single Nucleotide Polymorphisms (SNPs) data of 1,014 Italians with wide geographical coverage. We showed by using Principal Component analysis and model-based individual ancestry analysis, that the current population of Sardinia can be clearly differentiated genetically from mainland Italy and Sicily, and that a certain degree of genetic differentiation is detectable within the current Italian peninsula population. Pair-wise F_ST statistics Northern and Southern Italy amounts approximately to 0.001 between, and around 0.002 between Northern Italy and Utah residents with Northern and Western European ancestry (CEU). The Italian population also revealed a fine genetic substructure underscoring by the genomic inflation (Sardinia vs. Northern Italy = 3.040 and Northern Italy vs. CEU = 1.427), warning against confounding effects of hidden relatedness and population substructure in association studies.     

Pristine Early Eocene Wood Buried Deeply in Kimberlite from Northern Canada

We report exceptional preservation of fossil wood buried deeply in a kimberlite pipe that intruded northwestern Canada’s Slave Province 53.3±0.6 million years ago (Ma), revealed during excavation of diamond source rock. The wood originated from forest surrounding the eruption zone and collapsed into the diatreme before resettling in volcaniclastic kimberlite to depths >300 m, where it was mummified in a sterile environment. Anatomy of the unpermineralized wood permits conclusive identification to the genus Metasequoia (Cupressaceae). The wood yields genuine cellulose and occluded amber, both of which have been characterized spectroscopically and isotopically. From cellulose δ¹⁸O and δ²H measurements, we infer that Early Eocene paleoclimates in the western Canadian subarctic were 12–17°C warmer and four times wetter than present. Canadian kimberlites offer Lagerstätte-quality preservation of wood from a region with limited alternate sources of paleobotanical information.     

Prognostic Impact of del(17p) and del(22q) as Assessed by Interphase FISH in Sporadic Colorectal Carcinomas

Background

Most sporadic colorectal cancer (sCRC) deaths are caused by metastatic dissemination of the primary tumor. New advances in genetic profiling of sCRC suggest that the primary tumor may contain a cell population with metastatic potential. Here we compare the cytogenetic profile of primary tumors from liver metastatic versus non-metastatic sCRC.

Methodology/Principal Findings

We prospectively analyzed the frequency of numerical/structural abnormalities of chromosomes 1, 7, 8, 13, 14, 17, 18, 20, and 22 by iFISH in 58 sCRC patients: thirty-one non-metastatic (54%) vs. 27 metastatic (46%) disease. From a total of 18 probes, significant differences emerged only for the 17p11.2 and 22q11.2 chromosomal regions. Patients with liver metastatic sCRC showed an increased frequency of del(17p11.2) (10% vs. 67%;p<.001) and del(22q11.2) (0% vs. 22%;p = .02) versusnon-metastatic cases. Multivariate analysis of prognostic factors for overall survival (OS) showed that the only clinical and cytogenetic parameters that had an independent adverse impact on patient outcome were the presence of del(17p) with a 17p11.2 breakpoint and del(22q11.2). Based on these two cytogenetic variables, patients were classified into three groups: low- (no adverse features), intermediate- (one adverse feature) and high-risk (two adverse features)- with significantly different OS rates at 5-years (p<.001): 92%, 53% and 0%, respectively.

Conclusions/Significance

Our results unravel the potential implication of del(17p11.2) in sCRC patients with liver metastasis as this cytogenetic alteration appears to be intrinsically related to an increased metastatic potential and a poor outcome, providing additional prognostic information to that associated with other cytogenetic alterations such as del(22q11.2). Additional prospective studies in larger series of patients would be required to confirm the clinical utility of the new prognostic markers identified.     

Association of Left Ventricular Mass with All-Cause Mortality,Myocardial Infarction and Stroke

Background

Our aim was to assess the association of left ventricular mass with mortality and nonfatal cardiovascular events.

Methodology/Principal Findings

Left ventricular mass was measured by echocardiography in 40138 adult patients (mean age 61.1±16.4 years, 52.5% male). The primary endpoint was all-cause mortality. Secondary endpoints included nonfatal myocardial infarction and nonfatal stroke. During a mean follow-up period of 5.6±3.9 years, 9181 patients died, 901 patients had a nonfatal myocardial infarction, and 2139 patients had a nonfatal stroke. Cumulative 10-year mortality was 26.8%, 31.9%, 37.4% and 46.4% in patients with normal, mildly, moderately and severely increased left ventricular mass, respectively (p<0.001). Ten-year rates of nonfatal myocardial infarction and stroke ranged from 3.2% and 6.7% in patients with normal left ventricular mass to 5.3% and 12.7% in those with severe increase in left ventricular mass, respectively. After multivariate adjustment, left ventricular mass remained an independent predictor of all-cause mortality (hazard ratio [HR] per 100 g increase 1.21, 95% confidence interval [CI] 1.14–1–27, p<0.001 in women, and HR 1.09, 95% CI 1.04–1–13, p<0.001 in men), myocardial infarction (HR 1.60, 95% CI 1.31–1.94, p<0.001 in women and HR 1.15, 95% CI 1.02–1.29, p = 0.019 in men) and stroke (HR 1.26, 95% CI 1.13–1.40, p<0.001 in women and HR 1.19, 95% CI 1.09–1.30, p<0.001 in men).

Conclusions/Significance

Left ventricular mass has a graded and independent association with all-cause mortality, myocardial infarction and stroke.     

Comprehensive Re-Sequencing of Adrenal Aldosterone Producing Lesions Reveal Three Somatic Mutations near the KCNJ5 Potassium Channel Selectivity Filter

Background

Aldosterone producing lesions are a common cause of hypertension, but genetic alterations for tumorigenesis have been unclear. Recently, either of two recurrent somatic missense mutations (G151R or L168R) was found in the potassium channel KCNJ5 gene in aldosterone producing adenomas. These mutations alter the channel selectivity filter and result in Na⁺ conductance and cell depolarization, stimulating aldosterone production and cell proliferation. Because a similar mutation occurs in a Mendelian form of primary aldosteronism, these mutations appear to be sufficient for cell proliferation and aldosterone production. The prevalence and spectrum of KCNJ5 mutations in different entities of adrenocortical lesions remain to be defined.

Materials and Methods

The coding region and flanking intronic segments of KCNJ5 were subjected to Sanger DNA sequencing in 351 aldosterone producing lesions, from patients with primary aldosteronism and 130 other adrenocortical lesions. The specimens had been collected from 10 different worldwide referral centers.

Results

G151R or L168R somatic mutations were identified in 47% of aldosterone producing adenomas, each with similar frequency. A previously unreported somatic mutation near the selectivity filter, E145Q, was observed twice. Somatic G151R or L168R mutations were also found in 40% of aldosterone producing adenomas associated with marked hyperplasia, but not in specimens with merely unilateral hyperplasia. Mutations were absent in 130 non-aldosterone secreting lesions. KCNJ5 mutations were overrepresented in aldosterone producing adenomas from female compared to male patients (63 vs. 24%). Males with KCNJ5 mutations were significantly younger than those without (45 vs. 54, respectively; p<0.005) and their APAs with KCNJ5 mutations were larger than those without (27.1 mm vs. 17.1 mm; p<0.005).

Discussion

Either of two somatic KCNJ5 mutations are highly prevalent and specific for aldosterone producing lesions. These findings provide new insight into the pathogenesis of primary aldosteronism.     

An acenocoumarol dosing algorithm using clinical and pharmacogenetic data in spanish patients with thromboembolic disease

Appropriate dosing of coumarins is difficult to establish, due to significant inter-individual variability in the dose required to obtain stable anticoagulation. Several genetic and other clinical factors have been associated with the coumarins dose, and some pharmacogenetic-guided dosing algorithms for warfarin and acenocoumarol have been developed for mixed populations. We recruited 147 patients with thromboembolic disease who were on stable doses and with an international normalized ratio (INR) between 2 and 3. We ascertained the influence of clinical and genetic variables on the stable acenocoumarol dose by multiple linear regression analysis in a derivation cohort (DC; n = 117) and developed an algorithm for dosing that included clinical factors (age, body mass index and concomitant drugs) and genetic variations of VKORC1, CYP2C9, CYP4F2 and APOE. For purposes of comparison, a model including only clinical data was created. The clinical factors explained 22% of the dose variability, which increased to 60.6% when pharmacogenetic information was included (p<0.001); CYP4F2 and APOE variants explained 4.9% of this variability. The mean absolute error of the predicted acenocoumarol dose (mg/week) obtained with the pharmacogenetic algorithm was 3.63 vs. 5.08 mg/week with the clinical algorithm (95% CI: 0.88 to 2.04). In the testing cohort (n = 30), clinical factors explained a mere 7% of the dose variability, compared to 39% explained by the pharmacogenetic algorithm. Considering a more clinically relevant parameter, the pharmacogenetic algorithm correctly predicted the real stable dose in 59.8% of the cases (DC) vs. only 37.6% predicted by the clinical algorithm (95% CI: 10 to 35). Therefore the number of patients needed to genotype to avoid one over- or under-dosing was estimated to be 5.     

The neural bases of social intention understanding: the role of interaction goals

Decoding others' intentions is a crucial aspect of social cognition. Neuroimaging studies suggest that inferring immediate goals engages the neural system for action understanding (i.e. mirror system), while the decoding of long-term intentions requires the system subserving the attribution of mental states (i.e. mentalizing). A controversial issue, stimulated by recent inconsistent results, concerns whether the two systems are concurrently vs. exclusively involved in intention understanding. This issue is particularly relevant in the case of social interactions, whose processing has been mostly, but not uncontroversially, associated with the mentalizing system. We tested the alternative hypothesis that the relative contribution of the two systems in intention understanding may also depend on the shared goal of interacting agents. To this purpose, 27 participants observed social interactions differing in their cooperative vs. affective shared goal during functional-Magnetic-Resonance-Imaging. The processing of both types of interactions activated the right temporo-parietal junction involved in mentalizing on action goals. Additionally, whole-brain and regions-of-interest analyses showed that the action understanding system (inferior prefrontal-parietal cortex) was more strongly activated by cooperative interactions, while the mentalizing-proper system (medial prefrontal cortex) was more strongly engaged by affective interactions. These differences were modulated by individual differences in empathizing. Both systems can thus be involved in understanding social intentions, with a relative weighting depending on the specific shared goal of the interaction.     

Role of Toll Like Receptors in Irritable Bowel Syndrome: Differential Mucosal Immune Activation According to the Disease Subtype

Background

The irritable bowel syndrome (IBS) is a functional gastrointestinal disorder whose pathogenesis is not completely understood. Its high prevalence and the considerable effects on quality of life make IBS a disease with high social cost. Recent studies suggest that low grade mucosal immune activation, increased intestinal permeability and the altered host-microbiota interactions that modulate innate immune response, contribute to the pathophysiology of IBS. However, the understanding of the precise molecular pathophysiology remains largely unknown.

Methodology and Findings

In this study our objective was to evaluate the TLR expression as a key player in the innate immune response, in the colonic mucosa of IBS patients classified into the three main subtypes (with constipation, with diarrhea or mixed). TLR2 and TLR4 mRNA expression was assessed by real time RT-PCR while TLRs protein expression in intestinal epithelial cells was specifically assessed by flow cytometry and immunofluorescence. Mucosal inflammatory cytokine production was investigated by the multiplex technology. Here we report that the IBS-Mixed subgroup displayed a significant up-regulation of TLR2 and TLR4 in the colonic mucosa. Furthermore, these expressions were localized in the epithelial cells, opening new perspectives for a potential role of epithelial cells in host-immune interactions in IBS. In addition, the increased TLR expression in IBS-M patients elicited intracellular signaling pathways resulting in increased expression of the mucosal proinflammatory cytokines IL-8 and IL1β.

Conclusions

Our results provide the first evidence of differential expression of TLR in IBS patients according to the disease subtype. These results offer further support that microflora plays a central role in the complex pathophysiology of IBS providing novel pharmacological targets for this chronic gastrointestinal disorder according to bowel habits.