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Background: Randomized controlled trials (RCT) on the treatment of severe space-occupying infarction of the middle cerebral artery (malignant MCA infarction) showed that early decompressive hemicraniectomy (DHC) is life saving and improves outcome without promoting most severe disablity in patients aged 18-60 years. It is, however, unknown whether the results obtained in the randomized trials are reproducible in a broader population in and apart from an academical setting and whether hemicraniectomy has been implemented in clinical practice as recommended by national and international guidelines. In addition, they were not powered to answer further relevant questions, e. g. concerning the selection of patients eligible for and the timing of hemicraniectomy. Other important issues such as the acceptance of disability following hemicraniectomy, the existence of specific prognostic factors, the value of conservative therapeutic measures, and the overall complication rate related to hemicraniectomy have not been sufficiently studied yet. Methods/Design: DESTINY-R is a prospective, multicenter, open, controlled registry including a 12 months follow-up. The only inclusion criteria is unilateral ischemic MCA stroke affecting more than 50% of the MCA-territory. The primary study hypothesis is to confirm the results of the RCT (76% mRS <= 4 after 12 months) in the subgroup of patients additionally fulfilling the inclusion cirteria of the RCT in daily routine. Assuming a calculated proportion of 0.76 for successes and a sample size of 300 for this subgroup, the width of the 95% CI, calculated using Wilson's method, will be 0.096 with the lower bound 0.709 and the upper bound 0.805. Discussion: The results of this study will provide information about the effectiveness of DHC in malignant MCA infarction in a broad population and a real-life situation in addition to and beyond RCT. Further prospectively obtained data will give crucial information on open questions and will be helpful in the plannig of upcomming treatment studies.
Evaluation of a pathophysiological role of the interleukin-6-type cytokine oncostatin M (OSM) for human diseases has been complicated by the fact that mouse models of diseases targeting either OSM or the OSM receptor (OSMR) complex cannot fully reflect the human situation. This is due to earlier findings that human OSM utilizes two receptor complexes, glycoprotein 130 (gp130)/leukemia inhibitory factor receptor (LIFR) (type I) and gp130/OSMR (type II), both with wide expression profiles. Murine OSM on the other hand only binds to the gp130/OSMR (type II) receptor complex with high affinity. Here, we characterize the receptor usage for rat OSM. Using different experimental approaches (knock-down of the OSMR expression by RNA interference, blocking of the LIFR by LIF-05, an antagonistic LIF variant and stably transfected Ba/F3 cells) we can clearly show that rat OSM surprisingly utilizes both, the type I and type II receptor complex, therefore mimicking the human situation. Furthermore, it displays cross-species activities and stimulates cells of human as well as murine origin. Its signaling capacities closely mimic those of human OSM in cell types of different origin in the way that strong activation of the Jak/STAT, the MAP kinase as well as the PI3K/Akt pathways can be observed. Therefore, rat disease models would allow evaluation of the relevance of OSM for human biology.
Prevention of tissue damages at the site of Leishmania major inoculation can be achieved if the BALB/c mice are systemically given L. major antigen (LmAg)-loaded bone marrow-derived dendritic cells (DC) that had been exposed to CpG-containing oligodeoxynucleotides (CpG ODN). As previous studies allowed establishing that interleukin-4 (IL-4) is involved in the redirection of the immune response towards a type 1 profile, we were interested in further exploring the role of IL-4. Thus, wild-type (wt) BALB/c mice or DC-specific IL-4 receptor \(\alpha\) (IL-4R \(\alpha\))-deficient (CD11c\(^{cre}\)IL-4R \(\alpha^{-/lox}\) BALB/c mice were given either wt or IL-4R \(\alpha\)-deficient LmAg-loaded bone marrow-derived DC exposed or not to CpG ODN prior to inoculation of 2x10\(^5\) stationary-phase L. major promastigotes into the BALB/c footpad. The results provide evidence that IL4/IL-4R alpha-mediated signaling in the vaccinating DC is required to prevent tissue damage at the site of L. major inoculation, as properly conditioned wt DC but not IL-4R alpha-deficient DC were able to confer resistance. Furthermore, uncontrolled L. major population size expansion was observed in the footpad and the footpad draining lymph nodes of CD11c\(^{cre}\)IL-4R \(\alpha^{-/lox}\) mice immunized with CpG ODN-exposed LmAg-loaded IL-4R \(\alpha\)-deficient DC, indicating the influence of IL-4R \(\alpha\)-mediated signaling in host DC to control parasite replication. In addition, no footpad damage occurred in BALB/c mice that were systemically immunized with LmAg-loaded wt DC doubly exposed to CpG ODN and recombinant IL-4. We discuss these findings and suggest that the IL4/IL4R \(\alpha\) signaling pathway could be a key pathway to trigger when designing vaccines aimed to prevent damaging processes in tissues hosting intracellular microorganisms.
Brain serotonin (5-HT) is implicated in a wide range of functions from basic physiological mechanisms to complex behaviors, including neuropsychiatric conditions, as well as in developmental processes. Increasing evidence links 5-HT signaling alterations during development to emotional dysregulation and psychopathology in adult age. To further analyze the importance of brain 5-HT in somatic and brain development and function, and more specifically differentiation and specification of the serotonergic system itself, we generated a mouse model with brain-specific 5-HT deficiency resulting from a genetically driven constitutive inactivation of neuronal tryptophan hydroxylase-2 (Tph2). Tph2 inactivation (Tph2-/-) resulted in brain 5-HT deficiency leading to growth retardation and persistent leanness, whereas a sex- and age-dependent increase in body weight was observed in Tph2+/- mice. The conserved expression pattern of the 5-HT neuron-specific markers (except Tph2 and 5-HT) demonstrates that brain 5-HT synthesis is not a prerequisite for the proliferation, differentiation and survival of raphe neurons subjected to the developmental program of serotonergic specification. Furthermore, although these neurons are unable to synthesize 5-HT from the precursor tryptophan, they still display electrophysiological properties characteristic of 5-HT neurons. Moreover, 5-HT deficiency induces an up-regulation of 5-HT\(_{1A}\) and 5-HT\(_{1B}\) receptors across brain regions as well as a reduction of norepinephrine concentrations accompanied by a reduced number of noradrenergic neurons. Together, our results characterize developmental, neurochemical, neurobiological and electrophysiological consequences of brain-specific 5-HT deficiency, reveal a dual dose-dependent role of 5-HT in body weight regulation and show that differentiation of serotonergic neuron phenotype is independent from endogenous 5-HT synthesis.
High background noise is an impediment to signal detection and perception. We report the use of multiple solutions to improve signal perception in the acoustic and visual modality by the Bornean rock frog, Staurois parvus. We discovered that vocal communication was not impaired by continuous abiotic background noise characterised by fast-flowing water. Males modified amplitude, pitch, repetition rate and duration of notes within their advertisement call. The difference in sound pressure between advertisement calls and background noise at the call dominant frequency of 5578 Hz was 8 dB, a difference sufficient for receiver detection. In addition, males used several visual signals to communicate with conspecifics with foot flagging and foot flashing being the most common and conspicuous visual displays, followed by arm waving, upright posture, crouching, and an open-mouth display. We used acoustic playback experiments to test the efficacy-based alerting signal hypothesis of multimodal communication. In support of the alerting hypothesis, we found that acoustic signals and foot flagging are functionally linked with advertisement calling preceding foot flagging. We conclude that S. parvus has solved the problem of continuous broadband low-frequency noise by both modifying its advertisement call in multiple ways and by using numerous visual signals. This is the first example of a frog using multiple acoustic and visual solutions to communicate in an environment characterised by continuous noise.
The Epstein-Barr Virus (EBV) -encoded EBNA2 protein, which is essential for the in vitro transformation of B-lymphocytes, interferes with cellular processes by binding to proteins via conserved sequence motifs. Its Arginine-Glycine (RG) repeat element contains either symmetrically or asymmetrically di-methylated arginine residues (SDMA and ADMA, respectively). EBNA2 binds via its SDMA-modified RG-repeat to the survival motor neurons protein (SMN) and via the ADMA-RG-repeat to the NP9 protein of the human endogenous retrovirus K (HERV-K (HML-2) Type 1). The hypothesis of this work was that the methylated RG-repeat mimics an epitope shared with cellular proteins that is used for interaction with target structures. With monoclonal antibodies against the modified RG-repeat, we indeed identified cellular homologues that apparently have the same surface structure as methylated EBNA2. With the SDMA-specific antibodies, we precipitated the Sm protein D3 (SmD3) which, like EBNA2, binds via its SDMA-modified RG-repeat to SMN. With the ADMA-specific antibodies, we precipitated the heterogeneous ribonucleoprotein K (hnRNP K). Specific binding of the ADMA-antibody to hnRNP K was demonstrated using E. coli expressed/ADMA-methylated hnRNP K. In addition, we show that EBNA2 and hnRNP K form a complex in EBV-infected B-cells. Finally, hnRNP K, when co-expressed with EBNA2, strongly enhances viral latent membrane protein 2A (LMP2A) expression by an unknown mechanism as we did not detect a direct association of hnRNP K with DNA-bound EBNA2 in gel shift experiments. Our data support the notion that the methylated surface of EBNA2 mimics the surface structure of cellular proteins to interfere with or co-opt their functional properties.
Dyslexia affects 5-10% of school-aged children and is therefore one of the most common learning disorders. Research on auditory event related potentials (AERP), particularly the mismatch negativity (MMN) component, has revealed anomalies in individuals with dyslexia to speech stimuli. Furthermore, candidate genes for this disorder were found through molecular genetic studies. A current challenge for dyslexia research is to understand the interaction between molecular genetics and brain function, and to promote the identification of relevant endophenotypes for dyslexia. The present study examines MMN, a neurophysiological correlate of speech perception, and its potential as an endophenotype for dyslexia in three groups of children. The first group of children was clinically diagnosed with dyslexia, whereas the second group of children was comprised of their siblings who had average reading and spelling skills and were therefore "unaffected'' despite having a genetic risk for dyslexia. The third group consisted of control children who were not related to the other groups and were also unaffected. In total, 225 children were included in the study. All children showed clear MMN activity to/da/-/ba/ contrasts that could be separated into three distinct MMN components. Whilst the first two MMN components did not differentiate the groups, the late MMN component (300-700 ms) revealed significant group differences. The mean area of the late MMN was attenuated in both the dyslexic children and their unaffected siblings in comparison to the control children. This finding is indicative of analogous alterations of neurophysiological processes in children with dyslexia and those with a genetic risk for dyslexia, without a manifestation of the disorder. The present results therefore further suggest that the late MMN might be a potential endophenotype for dyslexia.
Background: Regular exercise is beneficial for cardiovascular health but a recent meta-analysis indicated a relationship between extensive endurance sport and a higher risk of atrial fibrillation, an independent risk factor for stroke. However, data on the frequency of cardiac arrhythmias or (clinically silent) brain lesions during and after marathon running are missing.
Methods/Design: In the prospective observational "Berlin Beat of Running" study experienced endurance athletes underwent clinical examination (CE), 3 Tesla brain magnetic resonance imaging (MRI), carotid ultrasound imaging (CUI) and serial blood sampling (BS) within 2-3 days prior (CE, MRI, CUI, BS), directly after (CE, BS) and within 2 days after (CE, MRI, BS) the 38\(^{th}\) BMW BERLIN-MARATHON 2011. All participants wore a portable electrocardiogram (ECG)-recorder throughout the 4 to 5 days baseline study period. Participants with pathological MRI findings after the marathon, troponin elevations or detected cardiac arrhythmias will be asked to undergo cardiac MRI to rule out structural abnormalities. A follow-up is scheduled after one year.
Results: Here we report the baseline data of the enrolled 110 athletes aged 36-61 years. Their mean age was 48.8 \(\pm\) 6.0 years, 24.5% were female, 8.2% had hypertension and 2.7% had hyperlipidaemia. Participants have attended a mean of 7.5 \(\pm\) 6.6 marathon races within the last 5 years and a mean of 16 \(\pm\) 36 marathon races in total. Their weekly running distance prior to the 38\(^{th}\) BMW BERLIN-MARATHON was 65 \(\pm\) 17 km. Finally, 108 (98.2%) Berlin Beat-Study participants successfully completed the 38\(^{th}\) BMW BERLIN-MARATHON 2011.
Discussion: Findings from the "Berlin Beats of Running" study will help to balance the benefits and risks of extensive endurance sport. ECG-recording during the marathon might contribute to identify athletes at risk for cardiovascular events. MRI results will give new insights into the link between physical stress and brain damage.
Background: The diagnostic and pathophysiological relevance of antibodies to aquaporin-4 (AQP4-Ab) in patients with neuromyelitis optica spectrum disorders (NMOSD) has been intensively studied. However, little is known so far about the clinical impact of AQP4-Ab seropositivity.
Objective: To analyse systematically the clinical and paraclinical features associated with NMO spectrum disorders in Caucasians in a stratified fashion according to the patients' AQP4-Ab serostatus.
Methods: Retrospective study of 175 Caucasian patients (AQP4-Ab positive in 78.3%).
Results: Seropositive patients were found to be predominantly female (p < 0.0003), to more often have signs of co-existing autoimmunity (p < 0.00001), and to experience more severe clinical attacks. A visual acuity of <= 0.1 during acute optic neuritis (ON) attacks was more frequent among seropositives (p < 0.002). Similarly, motor symptoms were more common in seropositive patients, the median Medical Research Council scale (MRC) grade worse, and MRC grades <= 2 more frequent, in particular if patients met the 2006 revised criteria (p < 0.005, p < 0.006 and p < 0.01, respectively), the total spinal cord lesion load was higher (p < 0.006), and lesions >= 6 vertebral segments as well as entire spinal cord involvement more frequent (p < 0.003 and p < 0.043). By contrast, bilateral ON at onset was more common in seronegatives (p < 0.007), as was simultaneous ON and myelitis (p < 0.001); accordingly, the time to diagnosis of NMO was shorter in the seronegative group (p < 0.029). The course of disease was more often monophasic in seronegatives (p < 0.008). Seropositives and seronegatives did not differ significantly with regard to age at onset, time to relapse, annualized relapse rates, outcome from relapse (complete, partial, no recovery), annualized EDSS increase, mortality rate, supratentorial brain lesions, brainstem lesions, history of carcinoma, frequency of preceding infections, oligoclonal bands, or CSF pleocytosis. Both the time to relapse and the time to diagnosis was longer if the disease started with ON (p < 0.002 and p < 0.013). Motor symptoms or tetraparesis at first myelitis and > 1 myelitis attacks in the first year were identified as possible predictors of a worse outcome.
In this study, we have analysed the apoptotic effects of the ubiquitous environmental toxin benzo[ a] pyrene (BP) in HaCaT cells and human keratinocytes. Although prolonged exposure to BP was not cytotoxic on its own, a strong enhancement of CD95 (Fas)-mediated apoptosis was observed with BP at concentrations activating the aryl hydrocarbon receptor (AhR). Importantly, the ultimately mutagenic BP-metabolite, that is, (+)-anti-BP-7,8-diol-9,10-epoxide (BPDE), failed to enhance CD95-mediated cell death, suggesting that the observed pro-apoptotic effect of BP is neither associated with DNA adducts nor DNA-damage related signalling. CD95-induced apoptosis was also enhanced by beta-naphtoflavone, a well-known agonist of the AhR that does not induce DNA damage, thus suggesting a crucial role for AhR activation. Consistently, BP failed to sensitise for CD95L-induced apoptosis in AhR knockdown HaCaT cells. Furthermore, inhibition of CYP1A1 and/or 1B1 expression did not affect the pro-apoptotic crosstalk. Exposure to BP did not increase expression of CD95, but led to augmented activation of caspase-8. Enhancement of apoptosis was also observed with the TRAIL death receptors that activate caspase-8 and apoptosis by similar mechanisms as CD95. Together, these observations indicate an interference of AhR signalling with the activity of receptor-associated signalling intermediates that are shared by CD95 and TRAIL receptors. Our data thus suggest that AhR agonists can enhance cytokine-mediated adversity upon dermal exposure.
Physiological Notch Signaling Maintains Bone Homeostasis via RBPjk and Hey Upstream of NFATc1
(2012)
Notch signaling between neighboring cells controls many cell fate decisions in metazoans both during embryogenesis and in postnatal life. Previously, we uncovered a critical role for physiological Notch signaling in suppressing osteoblast differentiation in vivo. However, the contribution of individual Notch receptors and the downstream signaling mechanism have not been elucidated. Here we report that removal of Notch2, but not Notch1, from the embryonic limb mesenchyme markedly increased trabecular bone mass in adolescent mice. Deletion of the transcription factor RBPjk, a mediator of all canonical Notch signaling, in the mesenchymal progenitors but not the more mature osteoblast-lineage cells, caused a dramatic high-bone-mass phenotype characterized by increased osteoblast numbers, diminished bone marrow mesenchymal progenitor pool, and rapid age-dependent bone loss. Moreover, mice deficient in Hey1 and HeyL, two target genes of Notch-RBPjk signaling, exhibited high bone mass. Interestingly, Hey1 bound to and suppressed the NFATc1 promoter, and RBPjk deletion increased NFATc1 expression in bone. Finally, pharmacological inhibition of NFAT alleviated the high-bone-mass phenotype caused by RBPjk deletion. Thus, Notch-RBPjk signaling functions in part through Hey1-mediated inhibition of NFATc1 to suppress osteoblastogenesis, contributing to bone homeostasis in vivo.
Primary osteoporosis is an age-related disease characterized by an imbalance in bone homeostasis. While the resorptive aspect of the disease has been studied intensely, less is known about the anabolic part of the syndrome or presumptive deficiencies in bone regeneration. Multipotent mesenchymal stem cells (MSC) are the primary source of osteogenic regeneration. In the present study we aimed to unravel whether MSC biology is directly involved in the pathophysiology of the disease and therefore performed microarray analyses of hMSC of elderly patients (79-94 years old) suffering from osteoporosis (hMSC-OP). In comparison to age-matched controls we detected profound changes in the transcriptome in hMSC-OP, e.g. enhanced mRNA expression of known osteoporosis-associated genes (LRP5, RUNX2, COL1A1) and of genes involved in osteoclastogenesis (CSF1, PTH1R), but most notably of genes coding for inhibitors of WNT and BMP signaling, such as Sclerostin and MAB21L2. These candidate genes indicate intrinsic deficiencies in self-renewal and differentiation potential in osteoporotic stem cells. We also compared both hMSC-OP and non-osteoporotic hMSC-old of elderly donors to hMSC of similar to 30 years younger donors and found that the transcriptional changes acquired between the sixth and the ninth decade of life differed widely between osteoporotic and non-osteoporotic stem cells. In addition, we compared the osteoporotic transcriptome to long term-cultivated, senescent hMSC and detected some signs for pre-senescence in hMSC-OP. Our results suggest that in primary osteoporosis the transcriptomes of hMSC populations show distinct signatures and little overlap with non-osteoporotic aging, although we detected some hints for senescence-associated changes. While there are remarkable inter-individual variations as expected for polygenetic diseases, we could identify many susceptibility genes for osteoporosis known from genetic studies. We also found new candidates, e.g. MAB21L2, a novel repressor of BMP-induced transcription. Such transcriptional changes may reflect epigenetic changes, which are part of a specific osteoporosis-associated aging process.
GABA(A) receptors are clustered at synaptic sites to achieve a high density of postsynaptic receptors opposite the input axonal terminals. This allows for an efficient propagation of GABA mediated signals, which mostly result in neuronal inhibition. A key organizer for inhibitory synaptic receptors is the 93 kDa protein gephyrin that forms oligomeric superstructures beneath the synaptic area. Gephyrin has long been known to be directly associated with glycine receptor beta subunits that mediate synaptic inhibition in the spinal cord. Recently, synaptic GABA(A) receptors have also been shown to directly interact with gephyrin and interaction sites have been identified and mapped within the intracellular loops of the GABA(A) receptor alpha 1, alpha 2, and alpha 3 subunits. Gephyrin-binding to GABA(A) receptors seems to be at least one order of magnitude weaker than to glycine receptors (GlyRs) and most probably is regulated by phosphorylation. Gephyrin not only has a structural function at synaptic sites, but also plays a crucial role in synaptic dynamics and is a platform for multiple protein-protein interactions, bringing receptors, cytoskeletal proteins and downstream signaling proteins into close spatial proximity.
A recent genome-wide association study in patients with panic disorder (PD) identified a risk haplotype consisting of two single-nucleotide polymorphisms (SNPs) (rs7309727 and rs11060369) located in intron 3 of TMEM132D to be associated with PD in three independent samples. Now we report a subsequent confirmation study using five additional PD case-control samples (n = 1670 cases and n 2266 controls) assembled as part of the Panic Disorder International Consortium (PanIC) study for a total of 2678 cases and 3262 controls in the analysis. In the new independent samples of European ancestry (EA), the association of rs7309727 and the risk haplotype rs7309727-rs11060369 was, indeed, replicated, with the strongest signal coming from patients with primary PD, that is, patients without major psychiatric comorbidities (n 1038 cases and n 2411 controls). This finding was paralleled by the results of the meta-analysis across all samples, in which the risk haplotype and rs7309727 reached P-levels of P = 1.4e-8 and P = 1.1e-8, respectively, when restricting the samples to individuals of EA with primary PD. In the Japanese sample no associations with PD could be found. The present results support the initial finding that TMEM132D gene contributes to genetic susceptibility for PD in individuals of EA. Our results also indicate that patient ascertainment and genetic background could be important sources of heterogeneity modifying this association signal in different populations.
The photoluminescence of annealed Cd\(_x\)Zn\(_{1-x}\)Se quantum dots (QDs) under the influence of an external magnetic field has been studied in this paper. Post-growth annealing was performed for different annealing times. Above a critical annealing time, the QD luminescence shows a pronounced red-shift of the Zeeman split magnetic subcomponents. This observation is in contrast to the blue-shift caused by the diamagnetic behavior that is usually observed in non-magnetic QDs. We attribute our finding to the paramagnetism caused by the mixing of heavy and light hole states. Hence, post-growth thermal annealing treatment might be employed to render undoped epitaxial QDs intrinsically magnetic in a controlled manner. Two theoretical models were developed: a few-particle model to account for excitonic complex effects and a multiband calculation that describes the valence band hybridization. Contrasting the two models allowed us to unambiguously elucidate the nature of such an effect.
The transcription factor Miz1 forms repressive DNA-binding complexes with the Myc, Gfi-1 and Bcl-6 oncoproteins. Known target genes of these complexes encode the cyclin-dependent kinase inhibitors (CKIs) cdkn2b (p15\(^{Ink4}\)), cdkn1a (p21\(^{Cip1}\)), and cdkn1c (p57\(^{Kip2}\)). Whether Miz1-mediated repression is important for control of cell proliferation in vivo and for tumor formation is unknown. Here we show that deletion of the Miz1 POZ domain, which is critical for Miz1 function, restrains the development of skin tumors in a model of chemically-induced, Ras-dependent tumorigenesis. While the stem cell compartment appears unaffected, interfollicular keratinocytes lacking functional Miz1 exhibit a reduced proliferation and an accelerated differentiation of the epidermis in response to the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Tumorigenesis, proliferation and normal differentiation are restored in animals lacking cdkn1a, but not in those lacking cdkn2b. Our data demonstrate that Miz1-mediated attenuation of cell cycle arrest pathways via repression of cdkn1a has a critical role during tumorigenesis in the skin.
Background: There are few studies of the effects of nasal snuff and environmental factors on the risk of nasal cancer. This study aimed to investigate the impact of using nasal snuff and of other risk factors on the risk of nasal cancer in German men.
Methods: A population-based case-control study was conducted in the German Federal States of Bavaria and Baden-Wurttemberg. Tumor registries and ear, nose and throat departments provided access to patients born in 1926 or later.
Results: Telephone interviews were conducted with 427 cases (mean age 62.1 years) and 2.401 population-based controls (mean age 60.8 years). Ever-use of nasal snuff was associated with an odds ratio (OR) for nasal cancer of 1.45 (95% confidence interval [CI] 0.88-2.38) in the total study population, whereas OR in smokers was 2.01 (95% CI 1.00-4.02) and in never smokers was 1.10 (95% CI 0.43-2.80). The OR in ever-smokers vs. never-smokers was 1.60 (95% CI 1.24-2.07), with an OR of 1.06 (95% CI 1.05-1.07) per pack-year smoked, and the risk was significantly decreased after quitting smoking. Exposure to hardwood dust for at least 1 year resulted in an OR of 2.33 (95% CI 1.40-3.91) in the total population, which was further increased in never-smokers (OR 4.89, 95% CI 1.92-12.49) in analyses stratified by smoking status. The OR for nasal cancer after exposure to organic solvents for at least 1 year was 1.53 (1.17-2.01). Ever-use of nasal sprays/nasal lavage for at least 1 month rendered an OR of 1.59 (1.04-2.44). The OR after use of insecticides in homes was 1.48 (95% CI 1.04-2.11).
Conclusions: Smoking and exposure to hardwood dust were confirmed as risk factors for nasal carcinoma. There is evidence that exposure to organic solvents, and in-house use of insecticides could represent novel risk factors. Exposure to asbestos and use of nasal snuff were risk factors in smokers only.
To highlight human impact on biodiversity in the Lamto region, termites were studied with regard to their use as bio-indicators of habitat change in the tropics. Using a standardized method, termites were sampled in the three most common habitat types, i.e., in semi-deciduous forest, savanna woodland, and annually burned savanna, all inside Lamto Reserve and its surrounding rural domain. Termite species richness fell from 25 species in the Lamto forest to 13 species in the rural area, involving strong modification in the species composition (species turnover = 59 %). In contrast, no significant change in diversity was found between the Lamto savannas and the rural ones. In addition, the relative abundance of termites showed a significantly greater decline in the rural domain, even in the species Ancistrotermes cavithorax (Sjostedt) (Isoptera: Termitidae), which is known to be ecologically especially versatile. Overall, the findings of this study suggest further investigation around Lamto Reserve on the impact of human activities on biodiversity, focusing on forest conversion to land uses (e.g. agricultural and silvicultural systems).
Background: During early stages of brain development, secreted molecules, components of intracellular signaling pathways and transcriptional regulators act in positive and negative feed-back or feed-forward loops at the mid-hindbrain boundary. These genetic interactions are of central importance for the specification and subsequent development of the adjacent mid-and hindbrain. Much less, however, is known about the regulatory relationship and functional interaction of molecules that are expressed in the tectal anlage after tectal fate specification has taken place and tectal development has commenced.
Results: Here, we provide experimental evidence for reciprocal regulation and subsequent cooperation of the paired-type transcription factors Pax3, Pax7 and the TALE-homeodomain protein Meis2 in the tectal anlage. Using in ovo electroporation of the mesencephalic vesicle of chick embryos we show that (i) Pax3 and Pax7 mutually regulate each other's expression in the mesencephalic vesicle, (ii) Meis2 acts downstream of Pax3/7 and requires balanced expression levels of both proteins, and (iii) Meis2 physically interacts with Pax3 and Pax7. These results extend our previous observation that Meis2 cooperates with Otx2 in tectal development to include Pax3 and Pax7 as Meis2 interacting proteins in the tectal anlage.
Conclusion: The results described here suggest a model in which interdependent regulatory loops involving Pax3 and Pax7 in the dorsal mesencephalic vesicle modulate Meis2 expression. Physical interaction with Meis2 may then confer tectal specificity to a wide range of otherwise broadly expressed transcriptional regulators, including Otx2, Pax3 and Pax7.
The etiology of emotion-related disorders such as anxiety or affective disorders is considered to be complex with an interaction of biological and environmental factors. Particular evidence has accumulated for alterations in the dopaminergic and noradrenergic system - partly conferred by catechol-O-methyltransferase (COMT) gene variation - for the adenosinergic system as well as for early life trauma to constitute risk factors for those conditions. Applying a multi-level approach, in a sample of 95 healthy adults, we investigated effects of the functional COMT Val158Met polymorphism, caffeine as an adenosine A2A receptor antagonist (300 mg in a placebo-controlled intervention design) and childhood maltreatment (CTQ) as well as their interaction on the affect-modulated startle response as a neurobiologically founded defensive reflex potentially related to fear- and distress-related disorders. COMT val/val genotype significantly increased startle magnitude in response to unpleasant stimuli, while met/met homozygotes showed a blunted startle response to aversive pictures. Furthermore, significant gene-environment interaction of COMT Val158Met genotype with CTQ was discerned with more maltreatment being associated with higher startle potentiation in val/val subjects but not in met carriers. No main effect of or interaction effects with caffeine were observed. Results indicate a main as well as a GxE effect of the COMT Val158Met variant and childhood maltreatment on the affect-modulated startle reflex, supporting a complex pathogenetic model of the affect-modulated startle reflex as a basic neurobiological defensive reflex potentially related to anxiety and affective disorders.
Inflammation following ischaemic stroke attracts high priority in current research, particularly using human-like models and long-term observation periods considering translational aspects. The present study aimed on the spatio-temporal course of macrophage-like cell accumulation after experimental thromboembolic stroke and addressed microglial and astroglial reactions in the ischaemic border zone. Further, effects of tissue plasminogen activator (tPA) as currently best treatment for stroke and the potentially neuroprotective co-administration of hyperbaric oxygen (HBO) were investigated. Rats underwent middle cerebral artery occlusion and were assigned to control, tPA or tPA+HBO. Twenty-four hours, 7, 14 and 28 days were determined as observation time points. The accumulation of macrophage-like cells was semiquantitatively assessed by CD68 staining in the ischaemic area and ischaemic border zone, and linked to the clinical course. CD11b, ionized calcium binding adaptor molecule 1 (Iba), glial fibrillary acidic protein (GFAP) and Neuronal Nuclei (NeuN) were applied to reveal delayed glial and neuronal alterations. In all groups, the accumulation of macrophage-like cells increased distinctly from 24 hours to 7 days post ischaemia. tPA+HBO tended to decrease macrophage-like cell accumulation at day 14 and 28. Overall, a trend towards an association of increased accumulation and pronounced reduction of the neurological deficit was found. Concerning delayed inflammatory reactions, an activation of microglia and astrocytes with co-occurring neuronal loss was observed on day 28. Thereby, astrogliosis was found circularly in contrast to microglial activation directly in the ischaemic area. This study supports previous data on long-lasting inflammatory processes following experimental stroke, and additionally provides region-specific details on glial reactions. The tendency towards a decreasing macrophage-like cell accumulation after tPA+HBO needs to be discussed critically since neuroprotective properties were recently ascribed to long-term inflammatory processes.
Can Joint Carbon and Biodiversity Management in Tropical Agroforestry Landscapes Be Optimized?
(2012)
Managing ecosystems for carbon storage may also benefit biodiversity conservation, but such a potential 'win-win' scenario has not yet been assessed for tropical agroforestry landscapes. We measured above-and below-ground carbon stocks as well as the species richness of four groups of plants and eight of animals on 14 representative plots in Sulawesi, Indonesia, ranging from natural rainforest to cacao agroforests that have replaced former natural forest. The conversion of natural forests with carbon stocks of 227-362 Mg C ha\(^{-1}\) to agroforests with 82-211 Mg C ha\(^{-1}\) showed no relationships to overall biodiversity but led to a significant loss of forest-related species richness. We conclude that the conservation of the forest-related biodiversity, and to a lesser degree of carbon stocks, mainly depends on the preservation of natural forest habitats. In the three most carbon-rich agroforestry systems, carbon stocks were about 60% of those of natural forest, suggesting that 1.6 ha of optimally managed agroforest can contribute to the conservation of carbon stocks as much as 1 ha of natural forest. However, agroforestry systems had comparatively low biodiversity, and we found no evidence for a tight link between carbon storage and biodiversity. Yet, potential win-win agroforestry management solutions include combining high shade-tree quality which favours biodiversity with cacao-yield adapted shade levels.
Background: The weight that gene copy number plays in transcription remains controversial; although in specific cases gene expression correlates with copy number, the relationship cannot be inferred at the global level. We hypothesized that genes steadily expressed by 15 melanoma cell lines (CMs) and their parental tissues (TMs) should be critical for oncogenesis and their expression most frequently influenced by their respective copy number.
Results: Functional interpretation of 3,030 transcripts concordantly expressed (Pearson's correlation coefficient p-value < 0.05) by CMs and TMs confirmed an enrichment of functions crucial to oncogenesis. Among them, 968 were expressed according to the transcriptional efficiency predicted by copy number analysis (Pearson's correlation coefficient p-value < 0.05). We named these genes, "genomic delegates" as they represent at the transcriptional level the genetic footprint of individual cancers. We then tested whether the genes could categorize 112 melanoma metastases. Two divergent phenotypes were observed: one with prevalent expression of cancer testis antigens, enhanced cyclin activity, WNT signaling, and a Th17 immune phenotype (Class A). This phenotype expressed, therefore, transcripts previously associated to more aggressive cancer. The second class (B) prevalently expressed genes associated with melanoma signaling including MITF, melanoma differentiation antigens, and displayed a Th1 immune phenotype associated with better prognosis and likelihood to respond to immunotherapy. An intermediate third class (C) was further identified. The three phenotypes were confirmed by unsupervised principal component analysis.
Conclusions: This study suggests that clinically relevant phenotypes of melanoma can be retraced to stable oncogenic properties of cancer cells linked to their genetic back bone, and offers a roadmap for uncovering novel targets for tailored anti-cancer therapy.
Basing on the assumption that frontotemporal lobar degeneration (FTLD), schizophrenia and bipolar disorder (BPD) might share common aetiological mechanisms, we analyzed genetic variation in the FTLD risk gene progranulin (GRN) in a German population of patients with schizophrenia (n=271) or BPD (n=237) as compared with 574 age-, gender-and ethnicity-matched controls. Furthermore, we measured plasma progranulin levels in 26 German BPD patients as well as in 61 Italian BPD patients and 29 matched controls. A significantly decreased allelic frequency of the minor versus the wild-type allele was observed for rs2879096 (23.2 versus 34.2%, P<0.001, OR: 0.63, 95% CI: 0.49-0.80), rs4792938 (30.7 versus 39.7%, P=0.005, OR: 0.70, 95% CI: 0.55-0.89) and rs5848 (30.3 versus 36.8, P=0.007, OR: 0.71, 95% CI: 0.56-0.91). Mean +/- SEM progranulin plasma levels were significantly decreased in BPD patients, either Germans or Italians, as compared with controls (89.69 +/- 3.97 and 116.14 +/- 5.80 ng/ml, respectively, versus 180.81 +/- 18.39 ng/ml P<0.001) and were not correlated with age. In conclusion, GRN variability decreases the risk to develop BPD and schizophrenia, and progranulin plasma levels are significantly lower in BPD patients than in controls. Nevertheless, a larger replication analysis would be needed to confirm these preliminary results.
The 2007 flood in the Sahel: causes, characteristics and its presentation in the media and FEWS NET
(2012)
During the rainy season in 2007, reports about exceptional rains and floodings in the Sahel were published in the media, especially in August and September. Institutions and organizations like the World Food Programme (WFP) and FEWS NET put the events on the agenda and released alerts and requested help. The partly controversial picture was that most of the Sahel faced a crisis caused by widespread floodings. Our study shows that the rainy season in 2007 was exceptional with regard to rainfall amount and return periods. In many areas the event had a return period between 1 and 50 yr with high spatial heterogeneity, with the exception of the Upper Volta basin, which yielded return periods of up to 1200 yr. Despite the strong rainfall, the interpretation of satellite images show that the floods were mainly confined to lakes and river beds. However, the study also proves the difficulties in assessing the meteorological processes and the demarcation of flooded areas in satellite images without ground truthing. These facts and the somewhat vague and controversial reports in the media and FEWS NET demonstrate that it is crucial to thoroughly analyze such events at a regional and local scale involving the local population.
Background: Genes that, when mutated, cause Fanconi anemia or greatly increase breast cancer risk encode for proteins that converge on a homology-directed DNA damage repair process. Mutations in the SLX4 gene, which encodes for a scaffold protein involved in the repair of interstrand cross-links, have recently been identified in unclassified Fanconi anemia patients. A mutation analysis of SLX4 in German or Byelorussian familial cases of breast cancer without detected mutations in BRCA1 or BRCA2 has been completed, with globally negative results.
Methods: The genomic region of SLX4, comprising all exons and exon-intron boundaries, was sequenced in 94 Spanish familial breast cancer cases that match a criterion indicating the potential presence of a highly-penetrant germline mutation, following exclusion of BRCA1 or BRCA2 mutations.
Results: This mutational analysis revealed extensive genetic variation of SLX4, with 21 novel single nucleotide variants; however, none could be linked to a clear alteration of the protein function. Nonetheless, genotyping 10 variants (nine novel, all missense amino acid changes) in a set of controls (138 women and 146 men) did not detect seven of them. Conclusions: Overall, while the results of this study do not identify clearly pathogenic mutations of SLX4 contributing to breast cancer risk, further genetic analysis, combined with functional assays of the identified rare variants, may be warranted to conclusively assess the potential link with the disease.
The development of insecticides requires valid risk assessment procedures to avoid causing harm to beneficial insects and especially to pollinators such as the honeybee Apis mellifera. In addition to testing according to current guidelines designed to detect bee mortality, tests are needed to determine possible sublethal effects interfering with the animal's vitality and behavioral performance. Several methods have been used to detect sublethal effects of different insecticides under laboratory conditions using olfactory conditioning. Furthermore, studies have been conducted on the influence insecticides have on foraging activity and homing ability which require time-consuming visual observation. We tested an experimental design using the radiofrequency identification (RFID) method to monitor the influence of sublethal doses of insecticides on individual honeybee foragers on an automated basis. With electronic readers positioned at the hive entrance and at an artificial food source, we obtained quantifiable data on honeybee foraging behavior. This enabled us to efficiently retrieve detailed information on flight parameters. We compared several groups of bees, fed simultaneously with different dosages of a tested substance. With this experimental approach we monitored the acute effects of sublethal doses of the neonicotinoids imidacloprid (0.15-6 ng/bee) and clothianidin (0.05-2 ng/bee) under field-like circumstances. At field-relevant doses for nectar and pollen no adverse effects were observed for either substance. Both substances led to a significant reduction of foraging activity and to longer foraging flights at doses of >= 0.5 ng/bee (clothianidin) and >= 1.5 ng/bee (imidacloprid) during the first three hours after treatment. This study demonstrates that the RFID-method is an effective way to record short-term alterations in foraging activity after insecticides have been administered once, orally, to individual bees. We contribute further information on the understanding of how honeybees are affected by sublethal doses of insecticides.
Background: CEACAM3 is a granulocyte receptor mediating the opsonin-independent recognition and phagocytosis of human-restricted CEACAM-binding bacteria. CEACAM3 function depends on an intracellular immunoreceptor tyrosine-based activation motif (ITAM)-like sequence that is tyrosine phosphorylated by Src family kinases upon receptor engagement. The phosphorylated ITAM-like sequence triggers GTP-loading of Rac by directly associating with the guanine nucleotide exchange factor (GEF) Vav. Rac stimulation in turn is critical for actin cytoskeleton rearrangements that generate lamellipodial protrusions and lead to bacterial uptake.
Principal Findings: In our present study we provide biochemical and microscopic evidence that the adaptor proteins Nck1 and Nck2, but not CrkL, Grb2 or SLP-76, bind to tyrosine phosphorylated CEACAM3. The association is phosphorylation-dependent and requires the Nck SH2 domain. Overexpression of the isolated Nck1 SH2 domain, RNAi-mediated knock-down of Nck1, or genetic deletion of Nck1 and Nck2 interfere with CEACAM3-mediated bacterial internalization and with the formation of lamellipodial protrusions. Nck is constitutively associated with WAVE2 and directs the actin nucleation promoting WAVE complex to tyrosine phosphorylated CEACAM3. In turn, dominant-negative WAVE2 as well as shRNA-mediated knock-down of WAVE2 or the WAVE-complex component Nap1 reduce internalization of bacteria.
Conclusions: Our results provide novel mechanistic insight into CEACAM3-initiated phagocytosis. We suggest that the CEACAM3 ITAM-like sequence is optimized to co-ordinate a minimal set of cellular factors needed to efficiently trigger actin-based lamellipodial protrusions and rapid pathogen engulfment.
The larvae of the cabbage root fly induce serious damage to cultivated crops of the family Brassicaceae. We here report the biochemical characterisation of neuropeptides from the central nervous system and neurohemal organs, as well as regulatory peptides from enteroendocrine midgut cells of the cabbage maggot. By LC-MALDI-TOF/TOF and chemical labelling with 4-sulfophenyl isothiocyanate, 38 peptides could be identified, representing major insect peptide families: allatostatin A, allatostatin C, FMRFamide-like peptides, kinin, CAPA peptides, pyrokinins, sNPF, myosuppressin, corazonin, SIFamide, sulfakinins, tachykinins, NPLP1-peptides, adipokinetic hormone and CCHamide 1. We also report a new peptide (Yamide) which appears to be homolog to an amidated eclosion hormone-associated peptide in several Drosophila species. Immunocytochemical characterisation of the distribution of several classes of peptide-immunoreactive neurons and enteroendocrine cells shows a very similar but not identical peptide distribution to Drosophila. Since peptides regulate many vital physiological and behavioural processes such as moulting or feeding, our data may initiate the pharmacological testing and development of new specific peptide-based protection methods against the cabbage root fly and its larva.
Acute bacterial meningitis is a life-threatening disease in humans. Discussed as entry sites for pathogens into the brain are the blood-brain and the blood-cerebrospinal fluid barrier (BCSFB). Although human brain microvascular endothelial cells (HBMEC) constitute a well established human in vitro model for the blood-brain barrier, until now no reliable human system presenting the BCSFB has been developed. Here, we describe for the first time a functional human BCSFB model based on human choroid plexus papilloma cells (HIBCPP), which display typical hallmarks of a BCSFB as the expression of junctional proteins and formation of tight junctions, a high electrical resistance and minimal levels of macromolecular flux when grown on transwell filters. Importantly, when challenged with the zoonotic pathogen Streptococcus suis or the human pathogenic bacterium Neisseria meningitidis the HIBCPP show polar bacterial invasion only from the physiologically relevant basolateral side. Meningococcal invasion is attenuated by the presence of a capsule and translocated N. meningitidis form microcolonies on the apical side of HIBCPP opposite of sites of entry. As a functionally relevant human model of the BCSFB the HIBCPP offer a wide range of options for analysis of disease-related mechanisms at the choroid plexus epithelium, especially involving human pathogens.
DOT1A-dependent H3K76 methylation is required for replication regulation in Trypanosoma brucei
(2012)
Cell-cycle progression requires careful regulation to ensure accurate propagation of genetic material to the daughter cells. Although many cell-cycle regulators are evolutionarily conserved in the protozoan parasite Trypanosoma brucei, novel regulatory mechanisms seem to have evolved. Here, we analyse the function of the histone methyltransferase DOT1A during cell-cycle progression. Over-expression of DOT1A generates a population of cells with aneuploid nuclei as well as enucleated cells. Detailed analysis shows that DOT1A over-expression causes continuous replication of the nuclear DNA. In contrast, depletion of DOT1A by RNAi abolishes replication but does not prevent karyokinesis. As histone H3K76 methylation has never been associated with replication control in eukaryotes before, we have discovered a novel function of DOT1 enzymes, which might not be unique to trypanosomes.
Merkel cell carcinoma (MCC) is a highly aggressive skin cancer with an increasing incidence. The understanding of the molecular carcinogenesis of MCC is limited. Here, we scrutinized the PI3K/AKT pathway, one of the major pathways activated in human cancer, in MCC. Immunohistochemical analysis of 41 tumor tissues and 9 MCC cell lines revealed high levels of AKT phosphorylation at threonine 308 in 88% of samples. Notably, the AKT phosphorylation was not correlated with the presence or absence of the Merkel cell polyoma virus (MCV). Accordingly, knock-down of the large and small T antigen by shRNA in MCV positive MCC cells did not affect phosphorylation of AKT. We also analyzed 46 MCC samples for activating PIK3CA and AKT1 mutations. Oncogenic PIK3CA mutations were found in 2/46 (4%) MCCs whereas mutations in exon 4 of AKT1 were absent. MCC cell lines demonstrated a high sensitivity towards the PI3K inhibitor LY-294002. This finding together with our observation that the PI3K/AKT pathway is activated in the majority of human MCCs identifies PI3K/AKT as a potential new therapeutic target for MCC patients.
Intraocular pressure (IOP) is a highly heritable risk factor for primary open-angle glaucoma and is the only target for current glaucoma therapy. The genetic factors which determine IOP are largely unknown. We performed a genome-wide association study for IOP in 11,972 participants from 4 independent population-based studies in The Netherlands. We replicated our findings in 7,482 participants from 4 additional cohorts from the UK, Australia, Canada, and the Wellcome Trust Case-Control Consortium 2/Blue Mountains Eye Study. IOP was significantly associated with rs11656696, located in GAS7 at 17p13.1 (p = 1.4 x 10\(^{-8}\)), and with rs7555523, located in TMCO1 at 1q24.1 (p = 1.6 x 10\(^{-8}\)). In a meta-analysis of 4 case-control studies (total N = 1,432 glaucoma cases), both variants also showed evidence for association with glaucoma (p = 2.4 x 10\(^{-2}\) for rs11656696 and p = 9.1 x 10\(^{-4}\) for rs7555523). GAS7 and TMCO1 are highly expressed in the ciliary body and trabecular meshwork as well as in the lamina cribrosa, optic nerve, and retina. Both genes functionally interact with known glaucoma disease genes. These data suggest that we have identified two clinically relevant genes involved in IOP regulation.
Recently, several classifiers that combine primary tumor data, like gene expression data, and secondary data sources, such as protein-protein interaction networks, have been proposed for predicting outcome in breast cancer. In these approaches, new composite features are typically constructed by aggregating the expression levels of several genes. The secondary data sources are employed to guide this aggregation. Although many studies claim that these approaches improve classification performance over single genes classifiers, the gain in performance is difficult to assess. This stems mainly from the fact that different breast cancer data sets and validation procedures are employed to assess the performance. Here we address these issues by employing a large cohort of six breast cancer data sets as benchmark set and by performing an unbiased evaluation of the classification accuracies of the different approaches. Contrary to previous claims, we find that composite feature classifiers do not outperform simple single genes classifiers. We investigate the effect of (1) the number of selected features; (2) the specific gene set from which features are selected; (3) the size of the training set and (4) the heterogeneity of the data set on the performance of composite feature and single genes classifiers. Strikingly, we find that randomization of secondary data sources, which destroys all biological information in these sources, does not result in a deterioration in performance of composite feature classifiers. Finally, we show that when a proper correction for gene set size is performed, the stability of single genes sets is similar to the stability of composite feature sets. Based on these results there is currently no reason to prefer prognostic classifiers based on composite features over single genes classifiers for predicting outcome in breast cancer.
Late-stage age-related macular degeneration (AMD) is a common sight-threatening disease of the central retina affecting approximately 1 in 30 Caucasians. Besides age and smoking, genetic variants from several gene loci have reproducibly been associated with this condition and likely explain a large proportion of disease. Here, we developed a genetic risk score (GRS) for AMD based on 13 risk variants from eight gene loci. The model exhibited good discriminative accuracy, area-under-curve (AUC) of the receiver-operating characteristic of 0.820, which was confirmed in a cross-validation approach. Noteworthy, younger AMD patients aged below 75 had a significantly higher mean GRS (1.87, 95% CI: 1.69-2.05) than patients aged 75 and above (1.45, 95% CI: 1.36-1.54). Based on five equally sized GRS intervals, we present a risk classification with a relative AMD risk of 64.0 (95% CI: 14.11-1131.96) for individuals in the highest category (GRS 3.44-5.18, 0.5% of the general population) compared to subjects with the most common genetic background (GRS -0.05-1.70, 40.2% of general population). The highest GRS category identifies AMD patients with a sensitivity of 7.9% and a specificity of 99.9% when compared to the four lower categories. Modeling a general population around 85 years of age, 87.4% of individuals in the highest GRS category would be expected to develop AMD by that age. In contrast, only 2.2% of individuals in the two lowest GRS categories which represent almost 50% of the general population are expected to manifest AMD. Our findings underscore the large proportion of AMD cases explained by genetics particularly for younger AMD patients. The five-category risk classification could be useful for therapeutic stratification or for diagnostic testing purposes once preventive treatment is available.
Background The levels and clinical relevance of Th17 cells and other interleukin-17-producing cells have not been analyzed in chronic lymphocytic leukemia. The objective of this study was to quantify blood and tissue levels of Th17 and other interleukin-17-producing cells in patients with this disease and correlate blood levels with clinical outcome.
Design and Methods: Intracellular interleukin-17A was assessed in blood and splenic mononuclear cells from patients with chronic lymphocytic leukemia and healthy subjects using flow cytometry. Interleukin-17A-producing cells were analyzed in formalin-fixed, paraffin-embedded spleen and lymph node sections using immunohistochemistry and immunofluorescence.
Results: The absolute numbers of Th17 cells in peripheral blood mononuclear cells and the percentages of Th17 cells in spleen cell suspensions were higher in patients with chronic lymphocytic leukemia than in healthy subjects; in six out of eight paired chronic lymphocytic leukemia blood and spleen sample comparisons, Th17 cells were enriched in spleen suspensions. Circulating Th17 levels correlated with better prognostic markers and longer overall survival of the patients. Two "non-Th17" interleukin-17-expressing cells were identified in chronic lymphocytic leukemia spleens: proliferating cells of the granulocytic lineage and mature mast cells. Granulocytes and mast cells in normal spleens did not express interleukin-17. Conversely, both chronic lymphocytic leukemia and healthy lymph nodes contained similar numbers of interleukin-17+ mast cells as well as Th17 cells.
Conclusions: Th17 cells are elevated in chronic lymphocytic leukemia patients with better prognostic markers and correlate with longer survival. Furthermore, non-Th17 interleukin-17A-expressing cells exist in chronic lymphocytic leukemia spleens as maturing granulocytes and mature mast cells, suggesting that the microenvironmental milieu in leukemic spleens promotes the recruitment and/or expansion of Th17 and other IL-17-expressing cells. The pathophysiology of Th17 and non-Th17-interleukin-producing cells in chronic lymphocytic leukemia and their distributions and roles in this disease merit further study.
Multiple myeloma (MM) is a lethal human cancer characterized by a clonal expansion of malignant plasma cells in bone marrow. Mouse models of human MM are technically challenging and do not always recapitulate human disease. Therefore, new mouse models for MM are needed. Mineral-oil induced plasmacytomas (MOPC) develop in the peritoneal cavity of oil-injected BALB/c mice. However, MOPC typically grow extramedullary and are considered poor models of human MM. Here we describe an in vivo-selected MOPC315 variant, called MOPC315.BM, which can be maintained in vitro. When injected i.v. into BALB/c mice, MOPC315.BM cells exhibit tropism for bone marrow. As few as 10\(^4\) MOPC315.BM cells injected i.v. induced paraplegia, a sign of spinal cord compression, in all mice within 3-4 weeks. MOPC315.BM cells were stably transfected with either firefly luciferase (MOPC315.BM.Luc) or DsRed (MOPC315.BM.DsRed) for studies using noninvasive imaging. MOPC315.BM.Luc cells were detected in the tibiofemoral region already 1 hour after i.v. injection. Bone foci developed progressively, and as of day 5, MM cells were detected in multiple sites in the axial skeleton. Additionally, the spleen (a hematopoietic organ in the mouse) was invariably affected. Luminescent signals correlated with serum myeloma protein concentration, allowing for easy tracking of tumor load with noninvasive imaging. Affected mice developed osteolytic lesions. The MOPC315.BM model employs a common strain of immunocompetent mice (BALB/c) and replicates many characteristics of human MM. The model should be suitable for studies of bone marrow tropism, development of osteolytic lesions, drug testing, and immunotherapy in MM.
A new flavanone-chromone biflavonoid, preussianone (1), has been isolated from the leaves of Garcinia preussii, along with four known biflavonoids. The absolute stereostructures were elucidated by chemical, spectroscopic, and chiroptical methods. The biological properties of the new biflavonoid against several bacterial strains were evaluated.
Background. Nitric oxide (NO)-signal transduction plays an important role in renal ischemia/reperfusion (I/R) injury. NO produced by endothelial NO-synthase (eNOS) has protective functions whereas NO from inducible NO-synthase (iNOS) induces impairment. Rosiglitazone (RGZ), a peroxisome proliferator-activated receptor (PPAR)-gamma agonist exerted beneficial effects after renal I/R injury, so we investigated whether this might be causally linked with NOS imbalance. Methods. RGZ (5 mg/kg) was administered i.p. to SD-rats (f) subjected to bilateral renal ischemia (60 min). Following 24 h of reperfusion, inulin-and PAH-clearance as well as PAH-net secretion were determined. Morphological alterations were graded by histopathological scoring. Plasma NOx-production was measured. eNOS and iNOS expression was analyzed by qPCR. Cleaved caspase 3 (CC3) was determined as an apoptosis indicator and ED1 as a marker of macrophage infiltration in renal tissue. Results. RGZ improves renal function after renal I/R injury (PAH-/inulin-clearance, PAH-net secretion) and reduces histomorphological injury. Additionally, RGZ reduces NOx plasma levels, ED-1 positive cell infiltration and CC3 expression. iNOS-mRNA is reduced whereas eNOS-mRNA is increased by RGZ. Conclusion. RGZ has protective properties after severe renal I/R injury. Alterations of the NO pathway regarding eNOS and iNOS could be an explanation of the underlying mechanism of RGZ protection in renal I/R injury.
Behavioural Analyses of Quinine Processing in Choice, Feeding and Learning of Larval Drosophila
(2012)
Gustatory stimuli can support both immediate reflexive behaviour, such as choice and feeding, and can drive internal reinforcement in associative learning. For larval Drosophila, we here provide a first systematic behavioural analysis of these functions with respect to quinine as a study case of a substance which humans report as "tasting bitter". We describe the dose-effect functions for these different kinds of behaviour and find that a half-maximal effect of quinine to suppress feeding needs substantially higher quinine concentrations (2.0 mM) than is the case for internal reinforcement (0.6 mM). Interestingly, in previous studies (Niewalda et al. 2008, Schipanski et al 2008) we had found the reverse for sodium chloride and fructose/sucrose, such that dose-effect functions for those tastants were shifted towards lower concentrations for feeding as compared to reinforcement, arguing that the differences in dose-effect function between these behaviours do not reflect artefacts of the types of assay used. The current results regarding quinine thus provide a starting point to investigate how the gustatory system is organized on the cellular and/or molecular level to result in different behavioural tuning curves towards a bitter tastant.
Background: Cytokines such as interleukin 6 (IL-6) have been implicated in dual functions in neuropsychiatric disorders. Little is known about the genetic predisposition to neurodegenerative and neuroproliferative properties of cytokine genes. In this study the potential dual role of several IL-6 polymorphisms in brain morphology is investigated.
Methodology: In a large sample of healthy individuals (N = 303), associations between genetic variants of IL-6 (rs1800795; rs1800796, rs2069833, rs2069840) and brain volume (gray matter volume) were analyzed using voxel-based morphometry (VBM). Selection of single nucleotide polymorphisms (SNPs) followed a tagging SNP approach (e. g., Stampa algorigthm), yielding a capture 97.08% of the variation in the IL-6 gene using four tagging SNPs. Principal findings/results In a whole-brain analysis, the polymorphism rs1800795 (-174 C/G) showed a strong main effect of genotype (43 CC vs. 150 CG vs. 100 GG; x = 24, y = -10, z = -15; F(2,286) = 8.54, p(uncorrected) = 0.0002; p(AlphaSim-corrected) = 0.002; cluster size k = 577) within the right hippocampus head. Homozygous carriers of the G-allele had significantly larger hippocampus gray matter volumes compared to heterozygous subjects. None of the other investigated SNPs showed a significant association with grey matter volume in whole-brain analyses.
Conclusions/significance: These findings suggest a possible neuroprotective role of the G-allele of the SNP rs1800795 on hippocampal volumes. Studies on the role of this SNP in psychiatric populations and especially in those with an affected hippocampus (e.g., by maltreatment, stress) are warranted.
The individual sensitivity for ones internal bodily signals ("interoceptive awareness") has been shown to be of relevance for a broad range of cognitive and affective functions. Interoceptive awareness has been primarily assessed via measuring the sensitivity for ones cardiac signals ("cardiac awareness") which can be non-invasively measured by heartbeat perception tasks. It is an open question whether cardiac awareness is related to the sensitivity for other bodily, visceral functions. This study investigated the relationship between cardiac awareness and the sensitivity for gastric functions in healthy female persons by using non-invasive methods. Heartbeat perception as a measure for cardiac awareness was assessed by a heartbeat tracking task and gastric sensitivity was assessed by a water load test. Gastric myoelectrical activity was measured by electrogastrography (EGG) and subjective feelings of fullness, valence, arousal and nausea were assessed. The results show that cardiac awareness was inversely correlated with ingested water volume and with normogastric activity after water load. However, persons with good and poor cardiac awareness did not differ in their subjective ratings of fullness, nausea and affective feelings after drinking. This suggests that good heartbeat perceivers ingested less water because they subjectively felt more intense signals of fullness during this lower amount of water intake compared to poor heartbeat perceivers who ingested more water until feeling the same signs of fullness. These findings demonstrate that cardiac awareness is related to greater sensitivity for gastric functions, suggesting that there is a general sensitivity for interoceptive processes across the gastric and cardiac modality.
Background: Clearance of apoptotic neutrophils in the lung is an essential process to limit inflammation, since they could become a pro-inflammatory stimulus themselves. The clearance is partially mediated by alveolar macrophages, which phagocytose these apoptotic cells. The phagocytosis of apoptotic immune cells by monocytes in vitro has been shown to be augmented by several constituents of pulmonary surfactant, e. g. phospholipids and hydrophobic surfactant proteins. In this study, we assessed the influence of exogenous poractant alfa (Curosurf (R)) instillation on the in vivo phagocytosis of apoptotic neutrophils by alveolar macrophages.
Methods: Poractant alfa (200 mg/kg) was instilled intratracheally in the lungs of three months old adult male C57/Black 6 mice, followed by apoptotic neutrophil instillation. Bronchoalveloar lavage was performed and alveolar macrophages and neutrophils were counted. Phagocytosis of apoptotic neutrophils was quantified by determining the number of apoptotic neutrophils per alveolar macrophages.
Results: Exogenous surfactant increased the number of alveolar macrophages engulfing apoptotic neutrophils 2.6 fold. The phagocytosis of apoptotic neutrophils was increased in the presence of exogenous surfactant by a 4.7 fold increase in phagocytosed apoptotic neutrophils per alveolar macrophage.
Conclusions: We conclude that the anti-inflammatory properties of surfactant therapy may be mediated in part by increased numbers of alveolar macrophages and increased phagocytosis of apoptotic neutrophils by alveolar macrophages.
We present a quantitative 3D analysis of the motility of the blood parasite Trypanosoma brucei. Digital in-line holographic microscopy has been used to track single cells with high temporal and spatial accuracy to obtain quantitative data on their behavior. Comparing bloodstream form and insect form trypanosomes as well as mutant and wildtype cells under varying external conditions we were able to derive a general two-state-run-and-tumble-model for trypanosome motility. Differences in the motility of distinct strains indicate that adaption of the trypanosomes to their natural environments involves a change in their mode of swimming.
Background: Because most human stroke victims are elderly, studies of experimental stroke in the aged rather than the young rat model may be optimal for identifying clinically relevant cellular responses, as well for pinpointing beneficial interventions.
Methodology/Principal Findings: We employed the Affymetrix platform to analyze the whole-gene transcriptome following temporary ligation of the middle cerebral artery in aged and young rats. The correspondence, heat map, and dendrogram analyses independently suggest a differential, age-group-specific behaviour of major gene clusters after stroke. Overall, the pattern of gene expression strongly suggests that the response of the aged rat brain is qualitatively rather than quantitatively different from the young, i.e. the total number of regulated genes is comparable in the two age groups, but the aged rats had great difficulty in mounting a timely response to stroke. Our study indicates that four genes related to neuropathic syndrome, stress, anxiety disorders and depression (Acvr1c, Cort, Htr2b and Pnoc) may have impaired response to stroke in aged rats. New therapeutic options in aged rats may also include Calcrl, Cyp11b1, Prcp, Cebpa, Cfd, Gpnmb, Fcgr2b, Fcgr3a, Tnfrsf26, Adam 17 and Mmp14. An unexpected target is the enzyme 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 1 in aged rats, a key enzyme in the cholesterol synthesis pathway. Post-stroke axonal growth was compromised in both age groups.
Conclusion/Significance: We suggest that a multi-stage, multimodal treatment in aged animals may be more likely to produce positive results. Such a therapeutic approach should be focused on tissue restoration but should also address other aspects of patient post-stroke therapy such as neuropathic syndrome, stress, anxiety disorders, depression, neurotransmission and blood pressure.
RNase P processes the 5'-end of tRNAs. An essential catalytic RNA has been demonstrated in Bacteria, Archaea and the nuclei of most eukaryotes; an organism-specific number of proteins complement the holoenzyme. Nuclear RNase P from yeast and humans is well understood and contains an RNA, similar to the sister enzyme RNase MRP. In contrast, no protein subunits have yet been identified in the plant enzymes, and the presence of a nucleic acid in RNase P is still enigmatic. We have thus set out to identify and characterize the subunits of these enzymes in two plant model systems. Expression of the two known Arabidopsis MRP RNA genes in vivo was verified. The first wheat MRP RNA sequences are presented, leading to improved structure models for plant MRP RNAs. A novel mRNA encoding the central RNase P/MRP protein Pop1p was identified in Arabidopsis, suggesting the expression of distinct protein variants from this gene in vivo. Pop1p-specific antibodies precipitate RNase P activity and MRP RNAs from wheat extracts. Our results provide evidence that in plants, Pop1p is associated with MRP RNAs and with the catalytic subunit of RNase P, either separately or in a single large complex.
Animals acquire predictive values of sensory stimuli through reinforcement. In the brain of Drosophila melanogaster, activation of two types of dopamine neurons in the PAM and PPL1 clusters has been shown to induce aversive odor memory. Here, we identified the third cell type and characterized aversive memories induced by these dopamine neurons. These three dopamine pathways all project to the mushroom body but terminate in the spatially segregated subdomains. To understand the functional difference of these dopamine pathways in electric shock reinforcement, we blocked each one of them during memory acquisition. We found that all three pathways partially contribute to electric shock memory. Notably, the memories mediated by these neurons differed in temporal stability. Furthermore, combinatorial activation of two of these pathways revealed significant interaction of individual memory components rather than their simple summation. These results cast light on a cellular mechanism by which a noxious event induces different dopamine signals to a single brain structure to synthesize an aversive memory.
Background: Invasion of intestinal epithelial cells by Salmonella enterica serovar Typhimurium (S. Typhimurium) requires expression of the extracellular virulence gene expression programme (STEX), activation of which is dependent on the signalling molecule guanosine tetraphosphate (ppGpp). Recently, next-generation transcriptomics (RNA-seq) has revealed the unexpected complexity of bacterial transcriptomes and in this report we use differential RNA sequencing (dRNA-seq) to define the high-resolution transcriptomic architecture of wildtype S. Typhimurium and a ppGpp null strain under growth conditions which model STEX. In doing so we show that ppGpp plays a much wider role in regulating the S. Typhimurium STEX primary transcriptome than previously recognised.
Results: Here we report the precise mapping of transcriptional start sites (TSSs) for 78% of the S. Typhimurium open reading frames (ORFs). The TSS mapping enabled a genome-wide promoter analysis resulting in the prediction of 169 alternative sigma factor binding sites, and the prediction of the structure of 625 operons. We also report the discovery of 55 new candidate small RNAs (sRNAs) and 302 candidate antisense RNAs (asRNAs). We discovered 32 ppGpp-dependent alternative TSSs and determined the extent and level of ppGpp-dependent coding and non-coding transcription. We found that 34% and 20% of coding and non-coding RNA transcription respectively was ppGpp-dependent under these growth conditions, adding a further dimension to the role of this remarkable small regulatory molecule in enabling rapid adaptation to the infective environment.
Conclusions: The transcriptional architecture of S. Typhimurium and finer definition of the key role ppGpp plays in regulating Salmonella coding and non-coding transcription should promote the understanding of gene regulation in this important food borne pathogen and act as a resource for future research.