Bone and joint neuropathy in rats with type-2 diabetes

We have previously demonstrated that Goto-Kakizaki (GK) rats with spontaneous type-2 diabetes and peripheral neuropathy exhibit regional osteopathic changes. In the present study on 18 GK rats and 21 control Wistar rats, the occurrence of the sensory neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP), and the autonomic neuropeptide Y (NPY) was analysed in bone and joints, dorsal root ganglia and lumbar spinal cord by immunohistochemistry and radioimmunoassay (RIA). Immunohistochemistry disclosed a predominance of immunoreactivities in vessel-related nerve fibers, although some were also seen in free terminals. While SP, CGRP and NPY in periosteum, cortical bone and synovium was confined to neuronal tissue, the bone marrow in addition exhibited an abundance of NPY-positive megakaryocytes. Apart from this cellular source of NPY, the observations suggest that the three neuropeptides analysed in bone and joints are of neuronal origin. Quantification by RIA showed a significant decrease of NPY in cortical bone (-36%), bone marrow (-66%) and ankle (-29%) of GK rats. CGRP was decreased in the spinal cord (-19%) and dorsal root ganglia (-26%) but was unchanged in bone and joints, as with SP. Given the suggested anabolic role of NPY and CGRP on bone, neuropeptidergic deficit in diabetes may prove to be an important factor underlying the development of regional osteopenia.     

Ensembles of human MTL neurons "jump back in time" in response to a repeated stimulus

Episodic memory, which depends critically on the integrity of the medial temporal lobe (MTL), has been described as "mental time travel" in which the rememberer "jumps back in time." The neural mechanism underlying this ability remains elusive. Mathematical and computational models of performance in episodic memory tasks provide a specific hypothesis regarding the computation that supports such a jump back in time. The models suggest that a representation of temporal context, a representation that changes gradually over macroscopic periods of time, is the cue for episodic recall. According to these models, a jump back in time corresponds to a stimulus recovering a prior state of temporal context. In vivo single-neuron recordings were taken from the human MTL while epilepsy patients distinguished novel from repeated images in a continuous recognition memory task. The firing pattern of the ensemble of MTL neurons showed robust temporal autocorrelation over macroscopic periods of time during performance of the memory task. The gradually-changing part of the ensemble state was causally affected by the visual stimulus being presented. Critically, repetition of a stimulus caused the ensemble to elicit a pattern of activity that resembled the pattern of activity present before the initial presentation of the stimulus. These findings confirm a direct prediction of this class of temporal context models and may be a signature of the mechanism that underlies the experience of episodic memory as mental time travel. ? 2012 Wiley Periodicals, Inc.     

Gene Polymorphisms of Micrornas in Helicobacter pylori-Induced High Risk Atrophic Gastritis and Gastric Cancer

Background and aims

MicroRNAs (miRNAs) are known for their function as translational regulators of tumor suppressor or oncogenes. Single nucleotide polymorphisms (SNPs) in miRNAs related genes have been shown to affect the regulatory capacity of miRNAs and were linked with gastric cancer (GC) and premalignant gastric conditions. The purpose of this study was to evaluate potential associations between miRNA-related gene polymorphisms (miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608) and the presence of GC or high risk atrophic gastritis (HRAG) in European population.

Methods

Gene polymorphisms were analyzed in 995 subjects (controls: n = 351; GC: n = 363; HRAG: n = 281) of European descent. MiR-27a T>C (rs895819), miR-146a G>C (rs2910164), miR-196a-2 C>T (rs11614913), miR-492 G>C (rs2289030) and miR-608 C>G (rs4919510) SNPs were genotyped by RT-PCR.

Results

Overall, SNPs of miRNAs were not associated with the presence of GC or HRAG. We observed a tendency for miR-196a-2 CT genotype to be associated with higher risk of GC when compared to CC genotype, however, the difference did not reach the adjusted P-value (odds ratio (OR) - 1.46, 95% confidence interval (CI) 1.03-2.07, P = 0.032). MiR-608 GG genotype was more frequent in GC when compared to controls (OR −2.34, 95% CI 1.08–5.04), but significance remained marginal (P = 0.029). A similar tendency was observed in a recessive model for miR-608, where CC + CG vs GG genotype comparison showed a tendency for increased risk of GC with OR of 2.44 (95% CI 1.14–5.22, P = 0.021). The genotypes and alleles of miR-27a, miR-146a, miR-196a-2, miR-492 and miR-608 SNPs had similar distribution between histological subtypes of GC and were not linked with the presence of diffuse or intestinal-type GC.

Conclusions

Gene polymorphisms of miR-27a, miR-146a, miR-196a-2, miR-492, miR-492a and miR-608 were not associated with the presence of HRAG, GC or different histological subtypes of GC in European subjects.     

Physical,Behavioural and Genetic Predictors of Adult Hypertension: The Findings of the Kaunas Cardiovascular Risk Cohort Study

Background

The roots of adult hypertension go back to childhood. This study aimed to examine the independent effects of physical, behavioural and genetic factors identified in childhood and mid-adulthood for prediction of adult hypertension.

Methods

The study subjects were participants of the Kaunas Cardiovascular Risk Cohort study started in 1977 (n = 1082, age 12–13 years). In 2012, a total of 507 individuals (63.9% of eligible sample) participated in the 35-year follow-up survey. Health examination involved measurements of blood pressure (BP), anthropometric parameters, and interview about health behaviours. Subjects were genotyped for AGT (M235T), ACE (I/D, rs4340), ADM (rs7129220), and CACNB2 (rs12258967) genes polymorphisms. A genetic risk score was calculated as the sum of the number of risk alleles at each of four single nucleotide polymorphisms.

Results

AGT TT genotype male carriers had the highest mean values of systolic BP in childhood. In females, ADM genotype AA was associated with the highest values of systolic and diastolic BP, while CACNB2 genotype CC carriers had the highest values of diastolic BP in childhood. Systolic and diastolic BP in childhood, gain in BMI from childhood to adulthood, and risky alcohol consumption predicted hypertension in middle-aged men. In women, genetic risk score together with diastolic BP in childhood and gain in BMI were significant predictors of adult hypertension. The comparison of four nested logistic regression models showed that the prediction of hypertension improved significantly after the addition of BMI gain. Genetic risk score had a relatively weak effect on the improvement of the model performance in women.

Conclusions

BP in childhood and the gain in BMI from childhood to adulthood were significant predictors of adult hypertension in both genders. Genetic risk score in women and risky alcohol consumption in men were independently related with the risk of adult hypertension.     

Fibromyalgia Patients Had Normal Distraction Related Pain Inhibition but Cognitive Impairment Reflected in Caudate Nucleus and Hippocampus during the Stroop Color Word Test

The mechanisms causing cognitive problems in chronic pain patients are not well understood. We used the Stroop color word task (SCWT) to investigate distraction-induced analgesia, cognitive performance, and cerebral activation patterns in 29 fibromyalgia (FM) patients (mean age 49.8 years, range 25–64 years) and 31 healthy controls (HC) (mean age 46.3 years, range 20–63 years). In the first study, SCWT was used to investigate distraction-induced analgesia in FM patients. Two versions of the task were applied, one with only congruent color-word images and one with incongruent images. Pressure pain thresholds were assessed using a pressure algometer before, during, and following SCWT. In the second study, reaction times (RTs) were assessed and functional magnetic resonance imaging (fMRI) was used to investigate cerebral activation patterns in FM patients and HC during the SCWT. An event-related task mixing incongruent and congruent images was used. In study one, we found reduced pressure pain sensitivity during SCWT in both groups alike and no statistically significant differences were seen between the incongruent and congruent conditions. The study two revealed longer RTs during the incongruent compared to the congruent condition in both groups. FM patients had longer RTs than HC in both conditions. Furthermore, we found a significant interaction between group and congruency; that is, the group differences in RTs were more pronounced during the incongruent condition. This was reflected in a reduced activation of the caudate nucleus, lingual gyrus, temporal areas, and the hippocampus in FM patients compared to HC. In conclusion, we found normal pain inhibition during SWTC in FM patients. The cognitive difficulties seen in FM patients, reflected in longer RTs, were related to reduced activation of the caudate nucleus and hippocampus during incongruent SCWT, which most likely affected the mechanisms of cognitive learning in FM patients.     

Identification of a Novel Bacterial Outer Membrane Interleukin-1Β-Binding Protein from Aggregatibacter actinomycetemcomitans

Aggregatibacter actinomycetemcomitans is a gram-negative opportunistic oral pathogen. It is frequently associated with subgingival biofilms of both chronic and aggressive periodontitis, and the diseased sites of the periodontium exhibit increased levels of the proinflammatory mediator interleukin (IL)-1β. Some bacterial species can alter their physiological properties as a result of sensing IL-1β. We have recently shown that this cytokine localizes to the cytoplasm of A. actinomycetemcomitans in co-cultures with organotypic gingival mucosa. However, current knowledge about the mechanism underlying bacterial IL-1β sensing is still limited. In this study, we characterized the interaction of A. actinomycetemcomitans total membrane protein with IL-1β through electrophoretic mobility shift assays. The interacting protein, which we have designated bacterial interleukin receptor I (BilRI), was identified through mass spectrometry and was found to be Pasteurellaceae specific. Based on the results obtained using protein function prediction tools, this protein localizes to the outer membrane and contains a typical lipoprotein signal sequence. All six tested biofilm cultures of clinical A. actinomycetemcomitans strains expressed the protein according to phage display-derived antibody detection. Moreover, proteinase K treatment of whole A. actinomycetemcomitans cells eliminated BilRI forms that were outer membrane specific, as determined through immunoblotting. The protein was overexpressed in Escherichia coli in both the outer membrane-associated form and a soluble cytoplasmic form. When assessed using flow cytometry, the BilRI-overexpressing E. coli cells were observed to bind 2.5 times more biotinylated-IL-1β than the control cells, as detected with avidin-FITC. Overexpression of BilRI did not cause binding of a biotinylated negative control protein. In a microplate assay, soluble BilRI bound to IL-1β, but this binding was not specific, as a control protein for IL-1β also interacted with BilRI. Our findings suggest that A. actinomycetemcomitans expresses an IL-1β-binding surface-exposed lipoprotein that may be part of the bacterial IL-1β-sensing system.     

Contralateral but not systemic administration of bupivacaine reduces acute inflammation in the rat hindpaw

The effects of contralateral treatment with local anesthetics following acute hindpaw inflammation were investigated in rats. Inflammation was induced by unilateral injection of either 50 or 100 &#119 l of 1% carrageenan into the right paw. Contralateral injection of either bupivacaine or saline was given immediately before the carrageenan. Hindpaw edema and withdrawal responses to thermal and mechanical stimulation were evaluated after 3, 6 and 24 h. The results showed that the proinflammatory effects of carrageenan were strongest at 6 h after the injection of 100 mul carrageenan with bilaterally decreased withdrawal latencies and ipsilateral edema formation. Contralateral treatment with bupivacaine (1.25, 2.5 or 5 mg/ml) dosedependently reduced nociceptive behavior for 3-24 h. The edema was also reduced at 6 h. No effects on pain-related behavior were observed following systemic administration of bupivacaine. Sciatic nerve ligation on the contralateral side or intrathecal administration of saline significantly reduced the effects of bupivacaine when respectively compared with shamoperation and subcutaneous saline injection. Contralateral treatment with bupivacaine into the knee joint induced the same anti-nociceptive effect as administered into the paw. Our findings indicate that contralateral administration of bupivacaine induces long-lasting anti-nociceptive effects and may serve as a new or complementary treatment approach in acute inflammatory pain conditions.     

Generation of menangle virus nucleocapsid-like particles in yeast Saccharomyces cerevisiae

Menangle virus (MenV), which was isolated in Australia in 1997 during an outbreak of severe reproductive disease in pigs, is a novel member of the genus Rubulavirus in the family Paramyxoviridae. Although successfully eradicated from the affected piggery, fruit bats are considered to be the natural reservoir of the virus and therefore an ongoing risk of re-introduction to the pig population exists. Accordingly, reagents to facilitate serological surveillance are required to enhance the diagnostic capability for MenV, which is a newly recognized cause of disease in pigs with the potential to severely affect production in naive breeding herds. To address this need, recombinant MenV nucleocapsid (N) protein was expressed in the yeast Saccharomyces cerevisiae. Using the expression vector pFGG3 under control of the GAL7 promoter, high yields of recombinant MenV N protein were obtained. Electron microscopy demonstrated that purified recombinant N protein self-assembled into nucleocapsid-like particles which were identical in density and morphology, although not in length, to authentic nucleocapsids from virus-infected cells. Electron microscopy analysis also showed that yeast-expressed N protein which lacked the C-terminal tail (amino acid residues 400–519) formed significantly longer and denser nucleocapsid-like particles. Nucleocapsid-like particles derived from the full-length recombinant protein were stable and readily purified by CsCl gradient ultracentrifugation. When used as coating antigen in an indirect ELISA, the recombinant N protein reacted with sera derived from pigs experimentally infected with MenV and a simple serological assay to detect MenV-specific antibodies in pigs, fruit bats and humans could be designed on this basis.     

TAGLN2 regulates T cell activation by stabilizing the actin cytoskeleton at the immunological synapse

The formation of an immunological synapse (IS) requires tight regulation of actin dynamics by many actin polymerizing/depolymerizing proteins. However, the significance of actin stabilization at the IS remains largely unknown. In this paper, we identify a novel function of TAGLN2—an actin-binding protein predominantly expressed in T cells—in stabilizing cortical F-actin, thereby maintaining F-actin contents at the IS and acquiring LFA-1 (leukocyte function-associated antigen-1) activation after T cell receptor stimulation. TAGLN2 blocks actin depolymerization and competes with cofilin both in vitro and in vivo. Knockout of TAGLN2 (TAGLN2^−/−) reduced F-actin content and destabilized F-actin ring formation, resulting in decreased cell adhesion and spreading. TAGLN2^−/− T cells displayed weakened cytokine production and cytotoxic effector function. These findings reveal a novel function of TAGLN2 in enhancing T cell responses by controlling actin stability at the IS.