Inhibition of HDAC Enzymes Contributes to Differential Expression of Pro-Inflammatory Proteins in the TLR-4 Signaling Cascade.

Class I and II histone deacetylases (HDAC) are considered important regulators of immunity and inflammation. Modulation of HDAC expression and activity is associated with altered inflammatory responses but reports are controversial and the specific impact of single HDACs is not clear. We examined class I and II HDACs in TLR-4 signaling pathways in murine macrophages with a focus on IκB kinase epsilon (IKKε) which has not been investigated in this context before. Therefore, we applied the pan-HDAC inhibitors (HDACi) trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA) as well as HDAC-specific siRNA. Administration of HDACi reduced HDAC activity and decreased expression of IKKε although its acetylation was increased. Other pro-inflammatory genes (IL-1ß, iNOS, TNFα) also decreased while COX-2 expression increased. HDAC 2, 3 and 4, respectively, might be involved in IKKε and iNOS downregulation with potential participation of NF-κB transcription factor inhibition. Suppression of HDAC 1-3, activation of NF-κB and RNA stabilization mechanisms might contribute to increased COX-2 expression. In conclusion, our results indicate that TSA and SAHA exert a number of histone- and HDAC-independent functions. Furthermore, the data show that different HDAC enzymes fulfill different functions in macrophages and might lead to both pro- and anti-inflammatory effects which have to be considered in therapeutic approaches.

The Specific IKKε/TBK1 Inhibitor Amlexanox Suppresses Human Melanoma by the Inhibition of Autophagy, NF-κB and MAP Kinase Pathways.

Inhibitor-kappaB kinase epsilon (IKKε) and TANK-binding kinase 1 (TBK1) are non-canonical IκB kinases, both described as contributors to tumor growth and metastasis in different cancer types. Several hints indicate that they are also involved in the pathogenesis of melanoma; however, the impact of their inhibition as a potential therapeutic measure in this "difficult-to-treat" cancer type has not been investigated so far. We assessed IKKε and TBK1 expression in human malignant melanoma cells, primary tumors and the metastasis of melanoma patients. Both kinases were expressed in the primary tumor and in metastasis and showed a significant overexpression in tumor cells in comparison to melanocytes. The pharmacological inhibition of IKKε/TBK1 by the approved drug amlexanox reduced cell proliferation, migration and invasion. Amlexanox did not affect the cell cycle progression nor apoptosis induction but significantly suppressed autophagy in melanoma cells. The analysis of potential functional downstream targets revealed that NF-кB and ERK pathways might be involved in kinase-mediated effects. In an in vivo xenograft model in nude mice, amlexanox treatment significantly reduced tumor growth. In conclusion, amlexanox was able to suppress tumor progression potentially by the inhibition of autophagy as well as NF-кB and MAP kinase pathways and might therefore constitute a promising candidate for melanoma therapy.

5-Amino-1-ß-D-Ribofuranosyl-Imidazole-4-Carboxamide (AICAR) Reduces Peripheral Inflammation by Macrophage Phenotype Shift.

The stimulation of the AMP-activated kinase (AMPK) by 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR) has been associated with antihyperalgesia and the inhibition of nociceptive signaling in the spinal cord in models of paw inflammation. The attenuated nociception comes along with a strongly reduced paw edema, indicating that peripheral antiinflammatory mechanisms contribute to antinociception. In this study, we investigated the impact of AICAR on the immune cell composition in inflamed paws, as well as the regulation of inflammatory and resolving markers in macrophages. By using fluorescence-activated cell sorting (FACS) analysis and immunofluorescence, we found a significantly increased fraction of proresolving M2 macrophages and anti-inflammatory interleukin (IL)-10 in inflamed tissue, while M1 macrophages and proinflammatory cytokines such as IL-1 were decreased by AICAR in wild type mice. In AMPKα2 knock-out mice, the M2 polarization of macrophages in the paw was missing. The results were supported by experiments in primary macrophage cultures which also showed a shift to a proresolving phenotype with decreased levels of proinflammatory mediators and increased levels of antiinflammatory mediators. However, in the cell cultures, we did not observe differences between the AMPKα2+/+ and -/- cells, thus indicating that the AICAR-induced effects are at least partially AMPK-independent. In summary, our results indicate that AICAR has potent antiinflammatory and proresolving properties in inflammation which are contributing to a reduction of inflammatory edema and antinociception.

The Role of AlphαSynuclein in Mouse Models of Acute, Inflammatory and Neuropathic Pain.

(1) AlphαSynuclein (αSyn) is a synaptic protein which is expressed in the nervous system and has been linked to neurodegenerative diseases, in particular Parkinson's disease (PD). Symptoms of PD are mainly due to overexpression and aggregation of αSyn and include pain. However, the interconnection of αSyn and pain has not been clarified so far. (2) We investigated the potential effects of a αSyn knock-out on the nociceptive behaviour in mouse models of acute, inflammatory and neuropathic pain. Furthermore, we assessed the impact of αSyn deletion on pain-related cellular and molecular mechanisms in the spinal cord in these models. (3) Our results showed a reduction of acute cold nociception in αSyn knock-out mice while responses to acute heat and mechanical noxious stimulation were similar in wild type and knock-out mice. Inflammatory nociception was not affected by αSyn knock-out which is also mirrored by unaltered inflammatory gene expression. In contrast, in the SNI model of neuropathic pain, αSyn knock-out mice showed decreased mechanical allodynia as compared to wild type mice. This effect was associated with reduced proinflammatory mechanisms and suppressed activation of MAP kinase signalling in the spinal cord while endogenous antinociceptive mechanisms are not inhibited. (4) Our data indicate that αSyn plays a role in neuropathy and its inhibition might be useful to ameliorate pain symptoms after nerve injury.

Depression and suicidality among psychiatric residents - results from a multi-country study.

BACKGROUND: Previous studies have highlighted risks for depression and suicide in medical cohorts, but evidence regarding psychiatric residents is missing. This study aimed to determine rates of depression, suicide ideation and suicide attempt among psychiatric residents and to identify associated individual, educational and work-related risk factors. METHODS: A total of 1980 residents from 22 countries completed the online survey which collected data on depression (PHQ-9), suicidality (SIBQ), socio-demographic profiles, training, and education. Generalized linear modeling and logistic regression analysis were used to predict depression and suicide ideation, respectively. RESULTS: The vast majority of residents did not report depression, suicide ideation or attempting suicide during psychiatric training. Approximately 15% (n = 280) of residents met criteria for depression, 12.3% (n = 225) reported active suicide ideation, and 0.7% (n = 12) attempted suicide during the training. Long working hours and no clinical supervision were associated with depression, while more completed years of training and lack of other postgraduate education (e.g. PhD or psychotherapy training) were associated with increased risk for suicide ideation during psychiatric training. Being single and female was associated with worse mental health during training. LIMITATIONS: Due to the cross-sectional nature of the study, results should be confirmed by longitudinal studies. Response rate was variable but the outcome variables did not statistically significantly differ between countries with response rates of more or less than 50%. CONCLUSION: Depression rates among psychiatric residents in this study were lower than previously reported data, while suicide ideation rates were similar to previous reports. Poor working and training conditions were associated with worse outcomes. Training programmes should include effective help for residents experiencing mental health problems so that they could progress through their career to the benefit of their patients and wider society.

A prospective, randomized study of computer-assisted and conventional total knee arthroplasty. Three-dimensional evaluation of implant alignment and rotation.

BACKGROUND: Despite the use of modern instruments in total knee arthroplasty, component malalignment remains a problem. Whether a computer-assisted implantation technique can improve the accuracy of the spatial positioning of an implant is a matter of debate. The objective of this study was to determine whether computer-assisted total knee arthroplasty is superior to the conventional surgical method with regard to the precision of implant positioning. METHODS: The spatial positioning of the implant in sixty total knee arthroplasties (thirty-two imageless computer-assisted and twenty-eight conventional implantations) was determined three-dimensionally with use of computed tomographic measurement, which allowed derotation and full extension of the knee in order to avoid projection-related imaging errors. RESULTS: The overall mechanical axis showed a range of between 4.8 degrees of valgus and 6.6 degrees of varus alignment in the frontal plane for conventionally implanted arthroplasty components compared with a significantly smaller range of between 2.9 degrees of valgus and 3.1 degrees of varus alignment for computer-assisted implantations (p = 0.004). In relation to the tibial implant, the mean deviation (and standard deviation) from the mechanical axis was 2.0 degrees +/- 1.7 degrees for the conventional surgical method and 1.4 degrees +/- 0.9 degrees for the navigated implantation. The rotational deviation from the referenced axis of the femoral component was between 3.3 degrees of internal rotation and 5.0 degrees of external rotation for the conventional implantation method, with a mean deviation of 0.1 degrees +/- 2.2 degrees. Femoral components implanted with computer assistance showed a deviation of between 4.7 degrees of internal rotation and 2.2 degrees of external rotation, with a mean deviation of 0.3 degrees +/- 1.4 degrees. CONCLUSIONS: In this study, with our technique of filtering out projection-related imaging errors, computer-assisted implantation of total knee replacements improved the frontal and sagittal alignment of the femoral component but not of the tibial component. We found that the rotational alignment of the component was not improved through navigation by solely referencing to the epicondylar axis for the femur and the tuberosity for the tibia.

Influence of age and gender on risperidone plasma concentrations.

There is limited information on gender- and age-specific effects on plasma concentrations of risperidone and its active metabolite, 9-hydroxyrisperidone. The present study investigated dose- and weight-adjusted plasma concentrations of risperidone and its metabolite in three age groups (45 years, 45-60 years, over 60 years). Gender-specific differences were examined in the whole sample and for the premenopausal subgroup. One hundred and twenty-nine patients (18-93 years) were included in the study, 52 (40%) male and 77 (60%) female. Concentrations of risperidone and 9-hydroxyrisperidone were measured at steady-state by high-performance liquid chromatography with electrochemical detection (HPLC-ED). When total plasma concentrations (risperidone plus 9-hydroxyrisperidone) were adjusted for daily maintenance dose (ng/mL/mg C/D ratio), significant differences between all age groups were found. We found a mean increase of the C/D ratio by 34.8% per decade in patients older than 42 years. No significant sex-related differences in the average plasma concentrations were observed for the whole sample and for the premenopausal subgroup. This study shows clear evidence of higher risperidone total plasma concentrations for patients over 40 years of age. This linear increase (over 30% per decade) may then lead to an increased incidence of adverse effects in elderly patients.

Cognitive impairment: assessment with brain magnetic resonance imaging and proton magnetic resonance spectroscopy.

BACKGROUND: In vivo proton magnetic resonance spectroscopy is a safe and noninvasive tool that can be used to study aspects of brain chemistry and metabolism. This study was designed to evaluate its role in routine application to reveal the diagnostic reasons for cognitive impairment. METHOD: 37 Alzheimer's disease patients (NINCDS-ADRDA criteria), 31 patients with subcortical ischemic vascular dementia (Chui et al. criteria), and 13 subjects with subjective cognitive impairment (DSM-IV criteria) were included in this retrospective study. Magnetic resonance images were used for atrophy rating; additionally, proton magnetic resonance spectroscopy was performed. RESULTS: Significantly reduced N-acetylaspartate levels (p <.05) were found in both patients with Alzheimer's disease and patients with subcortical ischemic vascular dementia compared to the group with subjective memory complaints. The ratios of N-acetylaspartate/creatine and N-acetylaspartate/myo-inositol were significantly lower in Alzheimer's disease patients compared to patients with vascular dementia (p =.012) or patients with subjective memory impairment (p =.002). N-acetylaspartate/creatine and N-acetylaspartate/myo-inositol ratios were positively correlated to the degree of cerebral atrophy. Disoriented patients displayed a low N-acetylaspartate/creatine ratio. In contrast, we were not able to relate concurrent psychotic or behavioral symptoms to any spectroscopic parameter. CONCLUSION: This study indicates that proton magnetic resonance spectroscopy parameters could provide additional information in differentiating between Alzheimer's disease, subcortical ischemic vascular dementia, and subjective cognitive impairment. Therefore, this method can contribute to the routine diagnosis of dementia. Psychiatric and behavioral symptoms associated with dementia or due to a major psychiatric disorder cannot be related to changes in the measured proton magnetic resonance spectroscopy parameters.

Treatment patterns in inpatients with bipolar disorder at a psychiatric university hospital over a 9-year period: focus on mood stabilizers.

The increasing number of pharmacological treatment options for bipolar disorder seems to be paralleled by the number of evidence-based guidelines published previously. The aim of this study was to systematically examine the adherence to published guidelines and any change in prescription habits over time in a psychiatric hospital setting. This is a retrospective study of 531 bipolar in patients who were consecutively admitted to the Department for Psychiatry and Psychotherapy in Innsbruck. Their complete medical histories were evaluated for psychotropic medications, with a special focus on mood stabilizers (MSs). To compare the use of individual MSs or combinations with other psychotropic medications in two preselected observation periods (1999-2003 and 2004-2007), we used Fisher's exact test. Overall, the proportion of patients receiving at least one MS increased significantly from 1999-2003 to 2004-2007 (74.1 vs. 83.1%, P=0.011). Among the individual MSs, valproate was used most frequently in both time periods, showing a significant increase (P<0.001). Prescriptions of quetiapine (P<0.001) and lamotrigine (P=0.033) increased significantly, carbamazepine showed a significant decrease (P<0.001). Prescriptions of lithium and olanzapine decreased without reaching significance. The significant increase in the prescription of MS reflects the increasing awareness and implementation of recent evidence-based medicine guidelines into clinical practice. Clinical decision making, usually made on the basis of individual clinical experience, should always be reevaluated using periodically updated evidence-based medicine guidelines.

Stability and catalytic activity of alpha-amylase from barley malt at different pressure-temperature conditions.

The impact of high hydrostatic pressure and temperature on the stability and catalytic activity of alpha-amylase from barley malt has been investigated. Inactivation experiments with alpha-amylase in the presence and absence of calcium ions have been carried out under combined pressure-temperature treatments in the range of 0.1-800 MPa and 30-75 degrees C. A stabilizing effect of Ca(2+) ions on the enzyme was found at all pressure-temperature combinations investigated. Kinetic analysis showed deviations of simple first-order reactions which were attributed to the presence of isoenzyme fractions. Polynomial models were used to describe the pressure-temperature dependence of the inactivation rate constants. Derived from that, pressure-temperature isokinetic diagrams were constructed, indicating synergistic and antagonistic effects of pressure and temperature on the inactivation of alpha-amylase. Pressure up to 200 MPa significantly stabilized the enzyme against temperature-induced inactivation. On the other hand, pressure also hampers the catalytic activity of alpha-amylase and a progressive deceleration of the conversion rate was detected at all temperatures investigated. However, for the overall reaction of blocked p-nitrophenyl maltoheptaoside cleavage and simultaneous occurring enzyme inactivation in ACES buffer (0.1 M, pH 5.6, 3.8 mM CaCl(2)), a maximum of substrate cleavage was identified at 152 MPa and 64 degrees C, yielding approximately 25% higher substrate conversion after 30 min, as compared to the maximum at ambient pressure and 59 degrees C.

Effects of age and sex on olanzapine plasma concentrations.

Age and sex may influence both efficacy and side effects of second-generation antipsychotics. Women and elderly patients tend to have a higher prevalence for several side effects. Higher plasma levels in these groups of patients may be one reason. We studied the hypothesis that steady-state olanzapine plasma concentrations depend on age and sex. Sixty-seven inpatients on stable olanzapine dose were referred to routine therapeutic drug monitoring of olanzapine. Plasma levels were determined by high-performance liquid chromatography with electrochemical detection. Obtained data were then analyzed by analysis of covariance. Olanzapine plasma levels showed a marked sex difference with significantly higher mean concentrations in female patients (adjusted mean concentrations, 18.5 ng/mL for men and 31.7 ng/mL for women; P = 0.003). On average, the weight-corrected concentration/dose ratios shown by women were 33.5% higher than those shown by men, irrespective of age. Regarding the effect of age, weight-corrected concentration/dose ratios increased by an average of 9.4% per decade of life. All results were adjusted for smoking. Comedication did not significantly influence these results. In conclusion, age and sex are important variables to consider when prescribing olanzapine for women and in the elderly.