The influence of water motion on the growth rate of the kelp Laminaria hyperborea

The kelp Laminaria hyperborea is a dominant component of the subtidal nearshore ecosystem and is subjected to a heterogeneous wave and current climate. Water motion is known to influence physiological processes in macroalgae such as photosynthesis and nutrient uptake attributed to mass-transfer limitation. The study attempts to establish the effect of water motion on the growth rates of blades and elongation rates of the stipes of L. hyperborea at adjacent wave-exposed and wave-sheltered locations over a 12month period from field observations. The observations were supported by detailed physical and chemical measurements (light, temperature, seawater nutrient concentrations and hydrodynamics) and of tissue carbon and nitrogen concentrations together with δ¹³carbon. Despite a 30% difference in the root mean square of the velocity (Vel_rms) between the two survey locations, there was no evidence to suggest that water motion had any direct influence on the growth rates of either the blades or elongation of stipes of L. hyperborea. No significant differences were observed between either environmental or plant physiological variables between the sheltered and exposed locations. Using an integral velocity parameter (Vel_rms) the present study also highlighted the importance of the tidally induced current component of water flow in the subtidal zone. 

The influence of water motion on the growth rate of the kelp Laminaria digitata

The shallow water kelp Laminaria digitata, abundant in coastal zones of the North Atlantic, is exposed to a range of hydrodynamic environments that makes it ideal for assessing the role of water motion on their growth rate. Here we quantify the growth of L. digitata, as a factor of blade and stipe elongation, at sites adjacent to Strangford Lough, Northern Ireland under different hydrodynamic conditions over a one year period. A modelling approach was used to numerically determine both the temporal and spatial variability of the hydrodynamic environment. Ambient seawater nutrient concentrations, temperature and irradiance were measured as well as the internal nutrient status of the L. digitata populations. Kelp populations growing in the greatest and lowest water motion showed the lowest growth rates. Differences observed in growth rate could not be attributed to seawater nutrient availability, temperature or light. The internal nutrient status also suggested no influence on the observed differences in growth rate. Therefore if there are minimal differences in light, temperature and nutrients between sites, then populations of L. digitata exposed to different water motions are likely to exhibit different growth rates. It is suggested that the growth rate differences observed were a function of water motion with the possibility that, in response to the hydrodynamic forces experienced by the algal cells, L. digitata kelps in the high energy environments were putting more energy into strengthening cell walls rather than blade elongation

SOCS3 regulates onset and maintenance of TH2- mediated allergic responses

Members of the suppressor of cytokine signaling (SOCS) family are involved in the pathogenesis of many inflammatory diseases. SOCS-3 is predominantly expressed in T-helper type 2 (TH2) cells, but its role in TH2-related allergic diseases remains to be investigated. In this study we provide a strong correlation between SOCS-3 expression and the pathology of asthma and atopic dermatitis, as well as serum IgE levels in allergic human patients. SOCS-3 transgenic mice showed increased TH2 responses and multiple pathological features characteristic of asthma in an airway hypersensitivity model system. In contrast, dominant-negative mutant SOCS-3 transgenic mice, as well as mice with a heterozygous deletion of Socs3, had decreased TH2 development. These data indicate that SOCS-3 has an important role in regulating the onset and maintenance of TH2-mediated allergic immune disease, and suggest that SOCS-3 may be a new therapeutic target for the development of antiallergic drugs.

SOCS2 Can Enhance Interleukin-2 (IL-2) and IL-3 Signalling by Accelerating SOCS3 Degradation

Cytokine responses can be regulated by a family of proteins termed suppressors of cytokine signaling (SOCS) which can inhibit the JAK/STAT pathway in a classical negative-feedback manner. While the SOCS are thought to target signaling intermediates for degradation, relatively little is known about how their turnover is regulated. Unlike other SOCS family members, we find that SOCS2 can enhance interleukin-2 (IL-2)- and IL-3-induced STAT phosphorylation following and potentiate proliferation in response to cytokine stimulation. As a clear mechanism for these effects, we demonstrate that expression of SOCS2 results in marked proteasome-dependent reduction of SOCS3 and SOCS1 protein expression. Furthermore, we provide evidence that this degradation is dependent on the presence of an intact SOCS box and that the loss of SOCS3 is enhanced by coexpression of elongin B/C. This suggests that SOCS2 can bind to SOCS3 and elongin B/C to form an E3 ligase complex resulting in the degradation of SOCS3. Therefore, SOCS2 can enhance cytokine responses by accelerating proteasome-dependent turnover of SOCS3, suggesting a mechanism for the gigantism observed in SOCS2 transgenic mice.

A strategy for designing inhibitors of alpha-synuclein aggregation and toxicity as a novel treatment for Parkinson's disease and related disorders.

Convergent biochemical and genetic evidence suggests that the formation of alpha-synuclein (alpha-syn) protein deposits is an important and, probably, seminal step in the development of Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). It has been reported that transgenic animals overexpressing human alpha-syn develop lesions similar to those found in the brain in PD, together with a progressive loss of dopaminergic cells and associated abnormalities of motor function. Inhibiting and/or reversing alpha-syn self-aggregation could, therefore, provide a novel approach to treating the underlying cause of these diseases. We synthesized a library of overlapping 7-mer peptides spanning the entire alpha-syn sequence, and identified amino acid residues 64-100 of alpha-syn as the binding region responsible for its self-association. Modified short peptides containing alpha-syn amino acid sequences from part of this binding region (residues 69-72), named alpha-syn inhibitors (ASI), were found to interact with full-length alpha-syn and block its assembly into both early oligomers and mature amyloid-like fibrils. We also developed a cell-permeable inhibitor of alpha-syn aggregation (ASID), using the polyarginine peptide delivery system. This ASID peptide was able to inhibit the DNA damage induced by Fe(II) in neuronal cells transfected with alpha-syn(A53T), a familial PD-associated mutation. ASI peptides without this delivery system did not reverse levels of Fe(II)-induced DNA damage. Furthermore, the ASID peptide increased (P

Identification of a novel population of Langerin+ dendritic cells.

Langerhans cells (LCs) are antigen-presenting cells that reside in the epidermis of the skin and traffic to lymph nodes (LNs). The general role of these cells in skin immune responses is not clear because distinct models of LC depletion resulted in opposite conclusions about their role in contact hypersensitivity (CHS) responses. While comparing these models, we discovered a novel population of LCs that resides in the dermis and does not represent migrating epidermal LCs, as previously thought. Unlike epidermal LCs, dermal Langerin(+) dendritic cells (DCs) were radiosensitive and displayed a distinct cell surface phenotype. Dermal Langerin(+) DCs migrate from the skin to the LNs after inflammation and in the steady state, and represent the majority of Langerin(+) DCs in skin draining LNs. Both epidermal and dermal Langerin(+) DCs were depleted by treatment with diphtheria toxin in Lang-DTREGFP knock-in mice. In contrast, transgenic hLang-DTA mice lack epidermal LCs, but have normal numbers of dermal Langerin(+) DCs. CHS responses were abrogated upon depletion of both epidermal and dermal LCs, but were unaffected in the absence of only epidermal LCs. This suggests that dermal LCs can mediate CHS and provides an explanation for previous differences observed in the two-model systems.

Getting to the root of neuronal signalling in plant-parasitic nematodes using RNA interference

A variety of genes expressed in preparasitic second-stage juveniles (J2) of plant-parasitic nematodes appear to be vulnerable to RNA interference (RNAi) in vitro by coupling double-stranded (ds)RNA soaking with the artificial stimulation of pharyngeal pumping. Also, there is mounting evidence that the in planta generation of nematode-specific double-stranded RNAs (dsRNAs) has real utility in the control of these pests. Although neuronally-expressed genes in Caenorhabditis elegans are commonly refractory to RNAi, we have discovered that neuronally-expressed genes in plant-parasitic nematodes are highly susceptible to RNAi and that silencing can be induced by simple soaking procedures without the need for pharyngeal stimulation. Since most front-line anthelmintics that are used for the control of nematode parasites of animals and humans act to disrupt neuromuscular coordination, we argue that intercellular signalling processes associated with neurons have much appeal as targets for transgenic plant-based control strategies for plant-parasitic nematodes. FMRFamide-like peptides (FLPs) are a large family of neuropeptides which are intimately associated with neuromuscular regulation, and our studies on flp gene function in plant-parasitic nematodes have revealed that their expression is central to coordinated locomotory activities. We propose that the high level of conservation in nervous systems across nematodes coupled with the RNAi-susceptibility of neuronally-expressed genes in plant-parasitic nematodes provides a valuable research tool which could be used to interrogate neuronal signalling processes in nematodes.

Nitric oxide synthase gene therapy enhances the toxicity of cisplatin in cancer cells

BACKGROUND AIMS: Cell-based gene therapy is an alternative to viral and non-viral gene therapy. Emerging evidence suggests that mesenchymal stem cells (MSC) are able to migrate to sites of tissue injury and have immunosuppressive properties that may be useful in targeted gene therapy for sustained specific tissue engraftment. METHODS: In this study, we injected intravenously (i.v.) 1x10(6) MSC, isolated from green fluorescent protein (GFP) transgenic rats, into Rif-1 fibrosarcoma-bearing C3H/HeN mice. The MSC had been infected using a lentiviral vector to express stably the luciferase reporter gene (MSC-GFP-luci). An in vivo imaging system (IVIS 200) and Western blotting techniques were used to detect the distribution of MSC-GFP-luci in tumor-bearing animals. RESULTS: We observed that xenogenic MSC selectively migrated to the tumor site, proliferated and expressed the exogenous gene in subcutaneous fibrosarcoma transplants. No MSC distribution was detected in other organs, such as the liver, spleen, colon and kidney. We further showed that the FGF2/FGFR pathways may play a role in the directional movement of MSC to the Rif-1 fibrosarcoma. We performed in vitro co-culture and in vivo tumor growth analysis, showing that MSC did not affect the proliferation of Rif-1 cells and fibrosarcoma growth compared with an untreated control group. Finally, we demonstrated that the xenogenic MSC stably expressing inducible nitric oxide synthase (iNOS) protein transferred by a lentivirus-based system had a significant inhibitory effect on the growth of Rif-1 tumors compared with MSC alone and the non-treatment control group. CONCLUSIONS: iNOS delivered by genetically modified iNOS-MSC showed a significant anti-tumor effect both in vitro and in vivo. MSC may be used as a target gene delivery vehicle for the treatment of fibrosarcoma and other tumors

Inhibition of Advanced Glycation and Absence of Galectin-3 Prevent Blood-Retinal Barrier Dysfunction during Short-Term Diabetes

Breakdown of the inner blood-retinal barrier (iBRB) occurs early in diabetes and is central to the development of sight-threatening diabetic macular edema (DME) as retinopathy progresses. In the current study, we examined how advanced glycation end products (AGEs) forming early in diabetes could modulate vasopermeability factor expression in the diabetic retina and alter inter-endothelial cell tight junction (TJ) integrity leading to iBRB dysfunction. We also investigated the potential for an AGE inhibitor to prevent this acute pathology and examined a role of the AGE-binding protein galectin-3 (Gal-3) in AGE-mediated cell retinal pathophysiology. Diabetes was induced in C57/BL6 wild-type (WT) mice and in Gal-3(-/-) transgenic mice. Blood glucose was monitored and AGE levels were quantified by ELISA and immunohistochemistry. The diabetic groups were subdivided, and one group was treated with the AGE-inhibitor pyridoxamine (PM) while separate groups of WT and Gal-3(-/-) mice were maintained as nondiabetic controls. iBRB integrity was assessed by Evans blue assay alongside visualisation of TJ protein complexes via occludin-1 immunolocalization in retinal flat mounts. Retinal expression levels of the vasopermeability factor VEGF were quantified using real-time RT-PCR and ELISA. WT diabetic mice showed significant AGE -immunoreactivity in the retinal microvasculature and also showed significant iBRB breakdown (P < .005). These diabetics had higher VEGF mRNA and protein expression in comparison to controls (P < .01). PM-treated diabetics had normal iBRB function and significantly reduced diabetes-mediated VEGF expression. Diabetic retinal vessels showed disrupted TJ integrity when compared to controls, while PM-treated diabetics demonstrated near-normal configuration. Gal-3(-/-) mice showed significantly less diabetes-mediated iBRB dysfunction, junctional disruption, and VEGF expression changes than their WT counterparts. The data suggests an AGE-mediated disruption of iBRB via upregulation of VEGF in the diabetic retina, possibly modulating disruption of TJ integrity, even after acute diabetes. Prevention of AGE formation or genetic deletion of Gal-3 can effectively prevent these acute diabetic retinopathy changes.

Langerhans cells are critical in the development of atopic dermatitis-like inflammation and symptoms in mice

Genetic or vitamin D3-induced overexpression of thymic stromal lymphopoietin (TSLP) by keratinocytes results in an atopic dermatitis (AD)-like inflammatory phenotype in mice echoing the discovery of high TSLP expression in epidermis from AD patients. Although skin dendritic cells (DC) are suspected to be involved in AD, direct evidence of a pathogenetic role for skin DC in TSLP-induced skin inflammation has not yet been demonstrated. In a mouse model of AD, i.e. mice treated with the low-calcemic vitamin D3 analogue, MC903, we show that epidermal Langerhans cells (LC)-depleted mice treated with MC903 do neither develop AD-like inflammation nor increased serum IgE as compared to vitamin D3 analogue-treated control mice. Accordingly, we show that, in mice treated with MC903 or in K14-TSLP transgenic mice, expression of maturation markers by LC is increased whereas maturation of dermal DC is not altered. Moreover, only LC are responsible for the polarization of naive CD4+ T cells to a Th2 phenotype, i.e. decrease in interferon-gamma and increase in interleukin (IL)-13 production by CD4+ T cells. This effect of LC on T-lymphocytes does not require OX40-L/CD134 and is mediated by a concomitant down-regulation of IL-12 and CD70. Although it was previously stated that TSLP up-regulates the production of thymus and activation-regulated chemokine (TARC)/chemokine (C-C motif) ligand 17 (CCL17) and macrophage-derived chemokine (MDC)/CCL22 by human LC in vitro, our work shows that production of these Th2- cell attracting chemokines is increased only in keratinocytes in response to TSLP overexpression. These results demonstrate that LC are required for the development of AD in mouse models of AD involving epidermal TSLP overexpression.

Foxp3+ T cells induce Perforin-Dependent Dendritic Cell Death in Tumor-Draining Lymph Nodes.

Regulatory T (Treg) cells limit the onset of effective antitumor immunity, through yet-ill-defined mechanisms. We showed the rejection of established ovalbumin (OVA)-expressing MCA101 tumors required both the adoptive transfer of OVA-specific CD8(+) T cell receptor transgenic T cells (OTI) and the neutralization of Foxp3(+) T cells. In tumor-draining lymph nodes, Foxp3(+) T cell neutralization induced a marked arrest in the migration of OTI T cells, increased numbers of dendritic cells (DCs), and enhanced OTI T cell priming. Using an in vitro cytotoxic assay and two-photon live microscopy after adoptive transfer of DCs, we demonstrated that Foxp3(+) T cells induced the death of DCs in tumor-draining lymph nodes, but not in the absence of tumor. DC death correlated with Foxp3(+) T cell-DC contacts, and it was tumor-antigen and perforin dependent. We conclude that Foxp3(+) T cell-dependent DC death in tumor-draining lymph nodes limits the onset of CD8(+) T cell responses.

NADPH oxidase-4 mediates protection against chronic load-induced stress in mouse hearts by enhancing angiogenesis

Cardiac failure occurs when the heart fails to adapt to chronic stresses. Reactive oxygen species (ROS)-dependent signaling is implicated in cardiac stress responses but the role of different ROS sources remains unclear. Here, we report that NADPH oxidase-4 (Nox4) facilitates cardiac adaptation to chronic stress. Unlike other Nox proteins, Nox4 activity is regulated mainly by its expression level which increased in cardiomyocytes during stresses such as pressure overload or hypoxia. To investigate the functional role of Nox4 during the cardiac response to stress, we generated mice with a genetic deletion of Nox4 or a cardiomyocyte-targeted overexpression of Nox4. Basal cardiac function was normal in both models but Nox4-null animals developed exaggerated contractile dysfunction, hypertrophy and cardiac dilatation during exposure to chronic overload whereas Nox4-transgenic mice were protected. Investigation of mechanisms underlying this protective effect revealed a significant Nox4-dependent preservation of myocardial capillary density after pressure overload. Nox4 enhanced stress-induced activation of cardiomyocyte Hif1 and the release of VEGF, resulting in an increased paracrine angiogenic activity. These data indicate that cardiomyocyte Nox4 is a novel inducible regulator of myocardial angiogenesis, a key determinant of cardiac adaptation to overload stress. Our results also have wider relevance to the use of non-specific antioxidant approaches in cardiac disease and may provide an explanation for the failure of such strategies in many settings.

High incidence of proviral integrations in the Hoxa locus in a new model of E2a-PBX1-induced B-cell leukemia

Relevant mouse models of E2a-PBX1-induced pre-B cell leukemia are still elusive. We now report the generation of a pre-B leukemia model using E2a-PBX1 transgenic mice, which lack mature and precursor T-cells as a result of engineered loss of CD3epsilon expression (CD3epsilon(-/-)). Using insertional mutagenesis and inverse-PCR, we show that B-cell leukemia development in the E2a-PBX1 x CD3epsilon(-/-) compound transgenic animals is significantly accelerated when compared to control littermates, and document several known and novel integrations in these tumors. Of all common integration sites, a small region of 19 kb in the Hoxa gene locus, mostly between Hoxa6 and Hoxa10, represented 18% of all integrations in the E2a-PBX1 B-cell leukemia and was targeted in 86% of these leukemias compared to 17% in control tumors. Q-PCR assessment of expression levels for most Hoxa cluster genes in these tumors revealed an unprecedented impact of the proviral integrations on Hoxa gene expression, with tumors having one to seven different Hoxa genes overexpressed at levels up to 6600-fold above control values. Together our studies set the stage for modeling E2a-PBX1-induced B-cell leukemia and shed new light on the complexity pertaining to Hox gene regulation. In addition, our results show that the Hoxa gene cluster is preferentially targeted in E2a-PBX1-induced tumors, thus suggesting functional collaboration between these oncogenes in pre-B-cell tumors.