Interplay between thermal and immune ecology: effect of environmental temperature on insect immune response and energetic costs after an immune challenge

Although the study of thermoregulation in insects has shown that infected animals tend to prefer higher temperatures than healthy individuals, the immune response and energetic consequences of this preference remain unknown. We examined the effect of environmental temperature and the energetic costs associated to the activation of the immune response of Tenebrio molitor larvae following a lipopolysaccharide (LPS) challenge. We measured the effect of temperature on immune parameters including phenoloxidase (PO) activity and antibacterial responses. Further as proximal and distal costs of the immune response we determined the standard metabolic rate (SMR) and the loss of body mass (m_b), respectively. Immune response was stronger at 30 °C than was at 10 or 20 °C. While SMR at 10 and 20 °C did not differ between immune treatments, at 30 °C SMR of LPS-treated larvae was almost 25–60% higher than SMR of PBS-treated and naïve larvae. In addition, the loss in m_b was 1.9 and 4.2 times higher in LPS-treated larvae than in PBS-treated and naïve controls. The immune responses exhibited a positive correlation with temperature and both, SMR and m_b change, were sensitive to environmental temperature. These data suggest a significant effect of environmental temperature on the immune response and on the energetic costs of immunity.     

Lymphangiogenesis and its regulation in human neuroblastoma

For the first time, we could detect lymph vessels in neuroblastoma (NB) by immunohistochemistry with the antibody D2_40. Furthermore, we demonstrate expression of the lymphangiogenic factors VEGF-C and VEGF-D and their receptors VEGFR-2 and VEGFR-3 in NB in vitro and in vivo by RT-PCR. However, addition of recombinant human VEGF-C or -D results in the absence of autocrine growth stimulus in NB cells. Treatment of NB cells with retinoic acid did not lead to a change in VEGF-C or VEGF-D mRNA expression. Incubation of the NB cells Lan-5 with 5-Aza-2'-deoxycytidine led to the up-regulation of VEGF-C mRNA expression, suggesting that the promotor of VEGF-C is methylated. Finally, VEGF-C mRNA expression could be effectively down-regulated by transfection with a specific siRNA in the NB cells Kelly. We conclude that lymphangiogenesis is involved in NB biology and that siRNA directed against VEGF-C may have a future role in anti-lymphangiogenic strategies in NB.     

Feasibility of Image-Guided Radiotherapy for Elderly Patients with Locally Advanced Rectal Cancer

Purpose

The study aims to assess the tolerance of elderly patients (70 years or older) with locally advanced rectal cancers to image-guided radiotherapy (IGRT). A retrospective review of 13 elderly patients with locally advanced rectal cancer who underwent preoperative chemoradiation using IGRT was performed. Grade 3–4 acute toxicities, survival, and long-term complications were compared to 17 younger patients (<70 years) with the same disease stage.

Results

Grade 3–4 hematologic toxicities occurred in 7.6% and 0% (p = 0.4) and gastrointestinal toxicities, and, in 15.2% and 5% (p = 0.5), of elderly and younger patients, respectively. Surgery was aborted in three patients, two in the elderly group and one in the younger group. One patient in the elderly group died after surgery from cardiac arrhythmia. After a median follow-up of 34 months, five patients had died, two in the elderly and three in the younger group. The 3-year survival was 90.9% and 87.5% (p = 0.7) for the elderly and younger group respectively. Two patients in the younger group developed ischemic colitis and fecal incontinence. There was no statistically significant difference in acute and late toxicities as well as survival between the two groups.

Conclusions and Clinical Relevance

Elderly patients with locally advanced rectal cancers may tolerate preoperative chemoradiation with IGRT as well as younger patients. Further prospective studies should be performed to investigate the potential of IGRT for possible cure in elderly patients with locally advanced rectal cancer.     

Climatic responses of tree-ring width and δ13C signatures of sessile oak (Quercus petraea Liebl.) on soils with contrasting water supply

We investigated climate–growth relationships (in terms of tree-ring width, basal area increment (BAI), and tree-ring δ¹³C signatures) of Quercus petraea in Central Europe (Luxembourg). Tree responses were assessed for 160 years and compared for sites with contrasting water supply (i.e. Cambisols vs. Regosols with 175 and 42 mm available water capacity, respectively). Oak trees displayed very low climate sensitivity, and climatic variables explained only 24 and 21 % of variance in tree-ring width (TRW) (Cambisol and Regosol sites, respectively). Contrary to our expectations, site-related differences in growth responses (i.e. BAI, δ¹³C signatures) to climate shifts were not significant. This finding suggests a high plasticity of oak trees in the study area. Despite a distinct growth depression found for all trees in the decade 1988–1997 (attributable to increasing annual mean temperatures by 1.1 °C), oak trees completely recovered in subsequent years. This indicates a high resilience of sessile oak to climate change. Shifts in δ¹³C_corr signatures were mainly affected by temperature, and peaks in δ¹³C_corr values (corrected for the anthropogenic increase in atmospheric CO₂) coincided with decadal maximum temperatures. Correlations between δ¹³C signatures and TRW (mainly affected by precipitation) were not significant. This finding suggests that wood growth often was disconnected from carbon assimilation (e.g. due to carbon storage in the trunk or allocation to seeds). Since the selection of drought-resistant tree species gains importance within the context of adaptive forest management strategies, Q. petraea proves to be an adaptive tree species in Central Europe’s forests under shifting climatic conditions.     

Natural selection in the tropical treehopper <Emphasis Type="Italic">Alchisme grossa</Emphasis> (Hemiptera: Membracidae) on two sympatric host-plants

The pronotum is the most distinctive and representative structure in treehoppers. Although several functions have been proposed for this structure, its involvement in fitness has not been formally evaluated. Given the high degree of maternal investment exhibited by the subsocial membracid Alchisme grossa (Hoplophorionini), the pronotum has been suggested as a shield for offspring protection. We performed selection gradient analyses on A. grossa in order to evaluate how natural selection is acting upon two traits associated with this structure considering three different fitness proxies: (1) number of eggs, (2) the ratio between the number of first-stage nymphs and the number of eggs (early survival), and (3) the number of third-stage nymphs (late survival). Since A. grossa feeds, oviposits and mates on two alternative host-plants (Brugmansia suaveolens and Solanum ursinum, both Solanaceae), we also evaluated and compared selection gradients between these host-plants. We found positive linear selection acting upon pronotum length considering the number of eggs and the late survival fitness proxies and positive linear selection acting on the distance between the suprahumeral horns considering the early survival proxy on females ovipositing on both host-plants. These results highlight the importance of the pronotum in treehoppers females’ fitness and suggest that maternal care appears to have a greater importance in the way that natural selection is operating than the host-plant where oviposition occurs.     

Combination unilateral amygdaloid and ventromedial hypothalamic lesions: evidence for a feeding pathway

Previous studies have reported hyperphagia and obesity in female rats with bilateral lesions of the most posterodorsal part of the amygdala. In rats with unilateral posterodorsal amygdaloid lesions, a dense pattern of anterograde degeneration appears in the ipsilateral ventromedial hypothalamus, but not the contralateral nucleus. In the present study, female rats with unilateral ventromedial hypothalamic lesions or sham lesions were given either sham lesions or unilateral lesions of the posterodorsal amygdala (PDA) 20 days later. Unilateral lesions of the ventromedial hypothalamus resulted in hyperphagia and excessive weight gain. Subsequent amygdaloid lesions that were contralateral to the initial hypothalamic lesions resulted in hyperphagia and additional excessive weight gains, but amygdaloid lesions ipsilateral to the initial hypothalamic lesions did not. It is concluded that the effects of the two lesions on body weight are not additive and that the PDA and ventromedial hypothalamus are part of the same ipsilateral pathway regulating feeding behavior and body weight regulation.     

Distal cis-acting elements restrict expression of the CyIIIb actin gene in the aboral ectoderm of the sea urchin embryo

The distal region of the S. purpuratus actin CyIIIb gene, between −400 and −1400 nucleotides, contains at least three distinct cis-acting elements (C1R, C1L and E1) which are necessary for correct expression of fusion reporter genes in transgenic sea urchin embryos. The contribution of these elements in the temporal and spatial regulation of the gene was analyzed by single and double site-directed mutagenesis in fusion constructs which carry the bacterial chloramphenicol acetyl transferase (CAT) gene as a reporter. Following microinjection of the transgenes in sea urchin embryos, the activity of the mutants was compared to the wild type in time and space by measuring CAT activity at the blastula and pluteus embryonic stages and by in situ hybridization to the CAT mRNA at pluteus stage. Our results indicate that E1 involved in the temporal regulation of CyIIIb and that all three elements are necessary and sufficient to confer aboral (dorsal) ectoderm specificity to the proximal promoter. This is achieved by suppressing the promoter's activity in all other tissues by the cooperative interaction of the cis-acting elements. The C1R element, binding site of the nuclear receptors SpCOUP-TF and SpSHR2, is by itself sufficient to restrict expression in the ectoderm, whereas the aboral ectoderm restricted expression requires in addition the presence of both C1L adn E1. It is therefore evident, that the actin CyIIIb gene is exclusively expressed in the aboral ectoderm by a combinatorial repression in all other cell lineages of the developing embryo.     

Ca2+-selective transient receptor potential V channel architecture and function require a specific ankyrin repeat

Transient receptor potential (TRP) proteins form cation-conducting ion channels with currently 28 known genes encoding TRP channel monomers in mammals. These monomers are thought to coassemble to form homo- or heterotetrameric channels, but the signals governing their assembly are unknown. Within the TRPV subgroup, TRPV5 and TRPV6 show exclusive calcium selectivity and play an important role in calcium uptake. To identify signals that mediate assembly of functional TRPV6, we screened domains for self-association using co-immunoprecipitation, sucrose gradient centrifugation, bacterial two-hybrid assays, and patch clamp analysis. Of the two identified interaction domains within the N-terminal region, we showed that the first domain encompassing the third ankyrin repeat is the stringent requirement for physical assembly of TRPV6 subunits and when transferred to an unrelated protein enables its interaction with TRPV6. Deletion of this repeat or mutation of critical residues within this repeat rendered nonfunctional channels that do not co-immunoprecipitate or form tetramers. Suppression of dominant-negative inhibitors of TRPV6-specific currents was achieved by deletion of ankyrin (ANK) 3. We propose that the third ANK repeat initiates a molecular zippering process that proceeds past the fifth ANK repeat and creates an intracellular anchor that is necessary for functional subunit assembly.     

TorsinA folding and N-linked glycosylation are sensitive to redox homeostasis

The Endoplasmic Reticulum (ER) is responsible for the folding and post-translational modification of secretory proteins, as well as for triaging misfolded proteins. During folding, there is a complex yet only partially understood interplay between disulfide bond formation, which is an enzyme catalyzed event in the oxidizing environment of the ER, along with other post-translational modifications (PTMs) and chaperone-supported protein folding. Here, we used the glycoprotein torsinA as a model substrate to explore the impact of ER redox homeostasis on PTMs and protein biogenesis. TorsinA is a AAA+ ATPase with unusual oligomeric properties and controversial functions. The deletion of a C-terminal glutamic acid residue (?E) is associated with the development of Early-Onset Torsion Dystonia, a severe movement disorder. TorsinA differs from other AAA+ ATPases since it is an ER resident, and as a result of its entry into the ER torsinA contains two N-linked glycans and at least one disulfide bond. The role of these PTMs on torsinA biogenesis and function and the identity of the enzymes that catalyze them are poorly defined. Using a yeast torsinA expression system, we demonstrate that a specific protein disulfide isomerase, Pdi1, affects the folding and N-linked glycosylation of torsinA and torsinA?E in a redox-dependent manner, suggesting that the acquisition of early torsinA folding intermediates is sensitive to perturbed interactions between Cys residues and the quality control machinery. We also highlight the role of specific Cys residues during torsinA biogenesis and demonstrate that torsinA?E is more sensitive than torsinA when these Cys residues are mutated.     

Variable Expression of PIK3R3 and PTEN in Ewing Sarcoma Impacts Oncogenic Phenotypes

Ewing Sarcoma is an aggressive malignancy of bone and soft tissue affecting children and young adults. Ewing Sarcoma is driven by EWS/Ets fusion oncoproteins, which cause widespread alterations in gene expression in the cell. Dysregulation of receptor tyrosine kinase signaling, particularly involving IGF-1R, also plays an important role in Ewing Sarcoma pathogenesis. However, the basis of this dysregulation, including the relative contribution of EWS/Ets-dependent and independent mechanisms, is not well understood. In the present study, we identify variable expression of two modifiers of PI3K signaling activity, PIK3R3 and PTEN, in Ewing Sarcoma, and examine the consequences of this on PI3K pathway regulation and oncogenic phenotypes. Our findings indicate that PIK3R3 plays a growth-promotional role in Ewing Sarcoma, but suggest that this role is not strictly dependent on regulation of PI3K pathway activity. We further show that expression of PTEN, a well-established, potent tumor suppressor, is lost in a subset of Ewing Sarcomas, and that this loss strongly correlates with high baseline PI3K pathway activity in cell lines. In support of functional importance of PTEN loss in Ewing Sarcoma, we show that re-introduction of PTEN into two different PTEN-negative Ewing Sarcoma cell lines results in downregulation of PI3K pathway activity, and sensitization to the IGF-1R small molecule inhibitor OSI-906. Our findings also suggest that PTEN levels may contribute to sensitivity of Ewing Sarcoma cells to the microtubule inhibitor vincristine, a relevant chemotherapeutic agent in this cancer. Our studies thus identify PIK3R3 and PTEN as modifiers of oncogenic phenotypes in Ewing Sarcoma, with potential clinical implications.