Combinations of Genetic Data Present in Bipolar Patients,but Absent in Control Persons

The main objective of the study was to find combinations of genetic variants significantly associated with bipolar disorder. In a previous study of bipolar disorder, combinations of three single nucleotide polymorphism (SNP) genotypes taken from 803 SNPs were analyzed, and four clusters of combinations were found to be significantly associated with bipolar disorder. In the present study, combinations of four SNP genotypes taken from the same 803 SNPs were analyzed, and one cluster of combinations was found to be significantly associated with bipolar disorder. Combinations from the new cluster and from the four previous clusters were identified in the genomes of 209 of the 607 patients in the study whereas none of the 1355 control participants had any of these combinations in their genome.     

Beneficial Effect of Verminephrobacter Nephridial Symbionts on the Fitness of the Earthworm Aporrectodea tuberculata

Almost all lumbricid earthworms (Oligochaeta: Lumbricidae) harbor species-specific Verminephrobacter (Betaproteobacteria) symbionts in their nephridia (excretory organs). The function of the symbiosis, and whether the symbionts have a beneficial effect on their earthworm host, is unknown; however, the symbionts have been hypothesized to enhance nitrogen retention in earthworms. The effect of Verminephrobacter on the life history traits of the earthworm Aporrectodea tuberculata (Eisen) was investigated by comparing the growth, development, and fecundity of worms with and without symbionts given high (cow dung)- and low (straw)-nutrient diets. There were no differences in worm growth or the number of cocoons produced by symbiotic and aposymbiotic worms. Worms with Verminephrobacter symbionts reached sexual maturity earlier and had higher cocoon hatching success than worms cured of their symbionts when grown on the low-nutrient diet. Thus, Verminephrobacter nephridial symbionts do have a beneficial effect on their earthworm host. Cocoons with and without symbionts did not significantly differ in total organic carbon, total nitrogen, or total hydrolyzable amino acid content, which strongly questions the hypothesized role of the symbionts in nitrogen recycling for the host.Symbiosis has long been recognized as a source of evolutionary innovation (24), and the acquisition of symbionts can enable animal hosts to exploit previously inaccessible niches (3). The phylum Annelida is no exception to this; chemosynthetic symbionts in marine annelids (e.g., the giant tubeworm Riftia sp. and other gutless marine oligochaetes [9]) gain energy from the oxidation of reduced sulfur compounds and fix CO₂ and supply their animal host with fixed carbon. A more obscure partnership is known from the bone-eating annelid Osedax sp., where endosymbionts help degrade the bones of whale carcasses, the only known habitat of the worms (33). The medicinal leech Hirudo sp., like other blood-feeding animals (3, 7), has symbionts that are thought to produce essential vitamins missing from a blood meal (13). In addition, leeches have a number of symbionts of unknown function in their nephridia (excretory organs) (18). Earthworms (Oligochaeta: Lumbricidae) have also long been known to harbor symbiotic bacteria in their nephridia (19, 36). The function of this symbiosis, however, is still not known, but the stability of the symbiosis over evolutionary time (23) suggests that the symbionts benefit the host.The earthworm symbionts reside in the nephridia and have therefore been proposed to be involved in internal recycling of nitrogen in the host (29). The earthworm nephridia play an important role in both nitrogenous waste excretion and osmoregulation (20). The nephridia are found in pairs in each segment of the worm and consist of a coiled tube leading from the coelom to the exterior (Fig. ​(Fig.1).1). The tube forms three loops, and the symbiotic bacteria are situated in the ampulla in the second loop, where they form a dense population lining the lumen wall (19, 36).Open in a separate windowFIG. 1.The nephridia are found as paired organs in each segment of the worm. The nephridostome (the inlet to the nephridia) protrudes into the previous segment. The nephridial tube forms three loops and finally empties out through the body wall via the nephridopore. The symbionts (black) reside in the ampulla in the second loop. (Modified from reference 23 with permission.)The symbionts form the monophyletic genus Verminephrobacter (Betaproteobacteria) (30, 36); they are species specific and present in almost all lumbricid earthworms (23). The Verminephrobacter symbionts are transmitted vertically via the cocoon, where they are deposited along with eggs and sperm (5). During embryogenesis, the symbionts migrate into the developing nephridia, and after the worm hatches, the symbionts can no longer infect (5, 6). By taking advantage of the vertical transmission mode, it has been possible to establish symbiont-free earthworm cultures in the laboratory through controlled antibiotic treatment of newly deposited cocoons (5; this study). Separation of the symbiotic partners allows studies of the effect of the symbionts on their earthworm hosts.Pandazis (29) hypothesized that the symbionts enhance earthworm nitrogen retention by excreting proteolytic enzymes that will degrade peptides and proteins lost in urine; this would allow the earthworm to reabsorb the resulting amino acids. As a consequence, earthworms cured of their symbionts should have a lower fitness level than control worms when grown under nitrogen-limiting conditions. To test this hypothesis, growth, development, fecundity, and cocoon hatching success were compared for symbiotic and aposymbiotic earthworms of the species Aporrectodea tuberculata (Eisen) under high and low nutrient availability conditions.     

Epinephrine infusion increases adipose interleukin-6 gene expression and systemic levels in humans.

Exercise increases IL-6 mRNA in subcutaneous adipose tissue; however, the immediate signal for the IL-6 induction is unknown. We, therefore, explored the possible role of epinephrine in the induction of IL-6 in adipose tissue. Subcutaneous adipose tissue biopsies and blood samples were obtained from eight healthy men (mean age 27 yr, mean height 184 cm, mean weight 83 kg) in response to epinephrine infusion or in response to saline infusion. The rate of epinephrine infusion was such that circulating epinephrine concentrations mimicked that typically seen during exercise. The level of IL-6 mRNA in subcutaneous adipose tissue increased 26-fold (95% confidence interval, 9- to 166-fold) at 3 h of epinephrine infusion compared with controls (P=0.028). In addition, plasma levels of IL-6 increased in response to epinephrine infusion (P <0.001). However, epinephrine did not affect the IL-6 receptor mRNA. In conclusion, epinephrine acutely increases IL-6 mRNA levels in subcutaneous adipose tissue as well as circulating IL-6 levels in healthy men.     

Alignment-independent comparisons of human gastrointestinal tract microbial communities in a multidimensional 16S rRNA gene evolutionary space

We present a novel approach for comparing 16S rRNA gene clone libraries that is independent of both DNA sequence alignment and definition of bacterial phylogroups. These steps are the major bottlenecks in current microbial comparative analyses. We used direct comparisons of taxon density distributions in an absolute evolutionary coordinate space. The coordinate space was generated by using alignment-independent bilinear multivariate modeling. Statistical analyses for clone library comparisons were based on multivariate analysis of variance, partial least-squares regression, and permutations. Clone libraries from both adult and infant gastrointestinal tract microbial communities were used as biological models. We reanalyzed a library consisting of 11,831 clones covering complete colons from three healthy adults in addition to a smaller 390-clone library from infant feces. We show that it is possible to extract detailed information about microbial community structures using our alignment-independent method. Our density distribution analysis is also very efficient with respect to computer operation time, meeting the future requirements of large-scale screenings to understand the diversity and dynamics of microbial communities.     

Changes in brain levels of N-acylethanolamines and 2-arachidonoylglycerol in focal cerebral ischemia in mice

The N -acylethanolamines (NAEs) and 2-arachidonoylglycerol (2-AG) are bioactive lipids that can modulate inflammatory responses and protect neurons against glutamatergic excitotoxicity. We have used a model of focal cerebral ischemia in young adult mice to investigate the relationship between focal cerebral ischemia and endogenous NAEs. Over the first 24 h after induction of permanent middle cerebral artery occlusion, we observed a time-dependent increase in all the investigated NAEs, except for anandamide. Moreover, we found an accumulation of 2-AG at 4 h that returned to basal level 12 h after induction of ischemia. Accumulation of NAEs did not depend on regulation of N -acylphosphatidylethanolamine-hydrolyzing phospholipase D or fatty acid amide hydrolase. Treatment with the fatty acid amide hydrolase inhibitor URB597 (cyclohexyl carbamic acid 3'-carbamoyl-biphenyl-3-yl ester; 1 mg/kg; i.p.) 1.5 h before arterial occlusion decreased the infarct volume in our model system. Our results suggest that NAEs and 2-AG may be involved in regulation of neuroprotection during focal cerebral ischemia in mice.     

Distribution of neuropeptides in the primary olfactory center of the heliothine moth <Emphasis Type="Italic">Heliothis virescens</Emphasis>

Neuropeptides are a diverse widespread class of signaling substances in the nervous system. As a basis for the analysis of peptidergic neurotransmission in the insect olfactory system, we have studied the distribution of neuropeptides in the antennal lobe of the moth Heliothis virescens. Immunocytochemical experiments with antisera recognizing A-type allatostatins (AST-As), Manduca sexta allatotropin (Mas-AT), FMRFamide-related peptides (FaRPs), and tachykinin-related peptides (TKRPs) have shown that members of all four peptide families are present in local interneurons of the antennal lobe. Whereas antisera against AST-As, Mas-AT, and FaRPs give similar staining patterns characterized by dense meshworks of processes confined to the core of all antennal-lobe glomeruli, TKRPs are present only in neurons with blebby processes distributed throughout each glomerulus. In addition to local neurons, a pair of centrifugal neurons with cell bodies in the lateral subesophageal ganglion, arborizations in the antennal lobe, and projections in the inner antenno-cerebral tracts exhibits tachykinin immunostaining. Double-label immunofluorescence has detected the co-localization of AST-As, Mas-AT, and FaRPs in certain local interneurons, whereas TKRPs occurs in a distinct population. MALDI-TOF mass spectrometry has revealed nearly 50 mass peaks in the antennal lobe. Seven of these masses (four AST-As, two N-terminally extended FLRFamides, and Mas-AT) match known moth neuropeptides. The data thus show that local interneurons of the moth antennal lobe are highly differentiated with respect to their neuropeptide content. The antennal lobe therefore represents an ideal preparation for the future analysis of peptide signaling in insect brain.     

Ratiometric Imaging of Extracellular pH in Bacterial Biofilms with C-SNARF-4

pH in the extracellular matrix of bacterial biofilms is of central importance for microbial metabolism. Biofilms possess a complex three-dimensional architecture characterized by chemically different microenvironments in close proximity. For decades, pH measurements in biofilms have been limited to monitoring bulk pH with electrodes. Although pH microelectrodes with a better spatial resolution have been developed, they do not permit the monitoring of horizontal pH gradients in biofilms in real time. Quantitative fluorescence microscopy can overcome these problems, but none of the hitherto employed methods differentiated accurately between extracellular and intracellular microbial pH and visualized extracellular pH in all areas of the biofilms. Here, we developed a method to reliably monitor extracellular biofilm pH microscopically with the ratiometric pH-sensitive dye C-SNARF-4, choosing dental biofilms as an example. Fluorescent emissions of C-SNARF-4 can be used to calculate extracellular pH irrespective of the dye concentration. We showed that at pH values of <6, C-SNARF-4 stained 15 bacterial species frequently isolated from dental biofilm and visualized the entire bacterial biomass in in vivo-grown dental biofilms with unknown species composition. We then employed digital image analysis to remove the bacterial biomass from the microscopic images and adequately calculate extracellular pH values. As a proof of concept, we monitored the extracellular pH drop in in vivo-grown dental biofilms fermenting glucose. The combination of pH ratiometry with C-SNARF-4 and digital image analysis allows the accurate monitoring of extracellular pH in bacterial biofilms in three dimensions in real time and represents a significant improvement to previously employed methods of biofilm pH measurement.     

A pilot study testing the feasibility of skin temperature monitoring to reduce recurrent foot ulcers in patients with diabetes – a randomized controlled trial

Background

Although monitoring foot skin temperatures has been associated with diabetic foot ulcer recurrence, no studies have been carried out to test the feasibility among European Caucasians. Moreover, the educational and/or motivational models that promote cognitive or psychosocial processes in these studies are lacking. Thus, we conducted a pilot randomized controlled trial to test the feasibility of monitoring foot skin temperatures in combination with theory-based counselling to standard foot care to reduce diabetic foot ulcer recurrence.

Methods

In a single-blinded nurse-led 1-year controlled trial, conducted at a hospital setting in Norway, 41 patients with diabetic neuropathy and previous foot ulcer were randomized to the intervention (n?=?21) or control groups (n?=?20). All participants were instructed in foot care and recording observations daily. Additionally, the intervention group was taught how to monitor and record skin temperature at baseline, and received counselling every third month supporting them to use the new treatment. Subjects observing temperature differences >2.0 °C between corresponding sites on the left and right foot on two consecutive days were asked to contact the study nurse and reduce physical activity. Fisher exact test was used to evaluate the effect of the intervention on the proportion of subjects with a foot ulcer. Kaplan-Meier survival analysis was performed to compare the two groups in regard to the time to development of a foot ulcer.

Results

In the intervention group, 67 % (n?=?14/21) monitored and recorded skin temperatures ≥80 % of the time while 70 % (n?=?14/20) of the controls recorded foot inspections. Foot ulcer incidence was 39 % (7/21) vs. 50 % (10/20) in the intervention and control groups, respectively (ns).

Conclusions

This feasibility study showed that the addition of counselling to promote self-monitoring of skin temperature to standard care to prevent recurrence of foot ulcer is feasible in patients with diabetes in Norway. Home skin temperature monitoring was performed as frequently by the intervention group as usual foot observations in the controls despite the extra effort required. We did not detect a difference in foot ulcer recurrence between groups, but our study may inform future full scale studies.

Trial registration

Clinicaltrials.gov NCT01269502