ECRG4 Expression in Normal Rat Tissues: Expression Study and Literature Review

The Esophageal Cancer Related Gene 4 (ECRG4) is a highly conserved tumour suppressor gene encoding various peptides (augurin, CΔ16 augurin, ecilin, argilin, CΔ16 argilin) which can be processed and secreted. In the present work, we examined ECRG4 expression and location in a wide range of rat organs and reviewed the available literature. ECRG4 mRNA was identified in all examined tissues by quantitative PCR (qPCR). ECRG4 immunoreaction was mainly cytoplasmic, and was detected in heart and skeletal muscles, smooth muscle cells showing only weak reactions. In the digestive system, ECRG4 immunostaining was stronger in the esophageal epithelium, bases of gastric glands, hepatocytes and pancreatic acinar epithelium. In the lymphatic system, immunoreactive cells were detectable in the thymus cortex, lymph node medulla and splenic red pulp. In the central and peripheral nervous systems, different neuronal groups showed different reaction intensities. In the endocrine system, ECRG4 immunoreaction was detected in the hypothalamic paraventricular and supraoptic nuclei, hypophysis, thyroid and parathyroid glands, adrenal zona glomerularis and medulla and Leydig cells, as well as in follicular and luteal cells of the ovary. In the literature, ECRG4 has been reported to inhibit cell proliferation and increase apoptosis in various cell types. It is down-regulated, frequently due to hypermethylation, in esophageal, prostate, breast and colon cancers, together with glioma (oncosuppressor function), although it is up-regulated in papillary thyroid cancer (oncogenic role). ECRG4 expression is also higher in non-proliferating cells of the lymphatic system. In conclusion, our identification of ECRG4 in many structures suggests the involvement of ECRG4 in the tumorigenesis of other organs and also the need for further research. In addition, on the basis of the location of ECRG4 in neurons and endocrine cells and the fact that it can be secreted, its role as a neurotransmitter/neuromodulator and endocrine factor must be examined in depth in the future.Key words: Augurin, gene expression, immunohistochemistry, rat, ECRG4     

The Local Complement Activation on Vascular Bed of Patients with Systemic Sclerosis: A Hypothesis-Generating Study

ObjectiveThe role of complement system in the pathogenesis of systemic sclerosis (SSc) has been debated during the last decade but an evident implication in this disease has never been found. We carried out an explorative study on SSc patients to evaluate the expression of soluble and local C5b-9 complement complex and its relation with a complement regulator, the Membrane Cofactor Protein (MCP, CD46) on skin vascular bed as target distinctive of SSc disease. We also analyzed two polymorphic variants in the complement activation gene cluster involving the MCP region.MethodsC5b-9 plasma levels of SSc patients and healthy subjects were analyzed by ELISA assay. Archival skin biopsies of SSc patients and controls were subjected to immunofluorescence analysis to detect C5b-9 and MCP on vascular endothelial cells. The expression of MCP was validated by immunoblot analysis with specific antibody. Polymorphic variants in the MCP gene promoter were tested by a quantitative PCR technique-based allelic discrimination method.ResultsEven though circulating levels of C5b-9 did not differ between SSc and controls, C5b-9 deposition was detected in skin biopsies of SSc patients but not in healthy subjects. MCP was significantly lower in skin vessels of SSc patients than in healthy controls and was associated with the over-expression of two polymorphic variants in the MCP gene promoter, which has been related to more aggressive phenotypes in other immune-mediated diseases.ConclusionsOur results firsty document the local complement activation with an abnormal expression of MCP in skin vessels of SSc patients, suggesting that a subset of SSc patients might be exposed to more severe organ complications and clinical evolution due to abnormal local complement activation.     

Palmitoylethanolamide Treatment Reduces Blood Pressure in Spontaneously Hypertensive Rats: Involvement of Cytochrome P450-Derived Eicosanoids and Renin Angiotensin System

Palmitoylethanolamide (PEA), a peroxisome proliferator-activated receptor-α agonist, has been demonstrated to reduce blood pressure and kidney damage secondary to hypertension in spontaneously hypertensive rat (SHR). Currently, no information is available concerning the putative effect of PEA on modulating vascular tone. Here, we investigate the mechanisms underpinning PEA blood pressure lowering effect, exploring the contribution of epoxyeicosatrienoic acids, CYP-dependent arachidonic acid metabolites, as endothelium-derived hyperpolarizing factors (EDHF), and renin angiotensin system (RAS) modulation. To achieve this aim SHR and Wistar-Kyoto rats were treated with PEA (30 mg/kg/day) for five weeks. Functional evaluations on mesenteric bed were performed to analyze EDHF-mediated vasodilation. Moreover, mesenteric bed and carotid were harvested to measure CYP2C23 and CYP2J2, the key isoenzymes in the formation of epoxyeicosatrienoic acids, and the soluble epoxide hydrolase, which is responsible for their degradation in the corresponding diols. Effect of PEA on RAS modulation was investigated by analyzing angiotensin converting enzyme and angiotensin receptor 1 expression. Here, we showed that EDHF-mediated dilation in response to acetylcholine was increased in mesenteric beds of PEA-treated SHR. Western blot analysis revealed that the increase in CYP2C23 and CYP2J2 observed in SHR was significantly attenuated in mesenteric beds of PEA-treated SHR, but unchanged in the carotids. Interestingly, in both vascular tissues, PEA significantly decreased the soluble epoxide hydrolase protein level, accompanied by a reduced serum concentration of its metabolite 14-15 dihydroxyeicosatrienoic acid, implying a reduction in epoxyeicosatrienoic acid hydrolisis. Moreover, PEA treatment down-regulated angiotensin receptor 1 and angiotensin converting enzyme expression, indicating a reduction in angiotensin II-mediated effects. Consistently, a damping of the activation of angiotensin receptor 1 underlying pathways in mesenteric beds was shown in basal conditions in PEA-treated SHR. In conclusion, our data demonstrate the involvement of epoxyeicosatrienoic acids and renin angiotensin system in the blood pressure lowering effect of PEA.     

In vivo moment generation and architecture of the human plantar flexors after different shortening–stretch cycles velocities

The purpose of this study was to examine the moment generation of the human plantar flexors and the architecture of the gastrocnemius medialis muscle during and after shortening–stretch cycles in vivo. Fourteen male subjects (30 ± 7 years, 177 ± 7 cm, 80 ± 9 kg) performed a series of electro-stimulated shortening–stretch plantar flexion contractions. The shortening–stretch cycles were performed at three constant angular velocities (25°/s, 50°/s, 100°/s), two amplitudes (15° and 25° ankle angle changes) and at two different stimulation frequencies (30 Hz and 85 Hz). The resultant ankle joint moments were calculated through inverse dynamics. Pennation angle and fascicle length of the m. gastrocnemius medialis at rest and during contractions were measured using ultrasonography. The corresponding ankle moments, kinematics and changes in muscle architecture were analysed at seven time intervals. A three-way analysis of variance (amplitude × velocity × stimulation frequency) and post-hoc test with Bonferroni correction were used to check the amplitude, velocity and stimulation level related effects on moment enhancement (α = 0.05). The results show an ankle joint moment enhancement after shortening–stretch cycles influenced by muscle architectural changes. We found 2–3% isometric ankle joint moment enhancement at steady state, 1.5–2.0 s after the shortening–stretch cycle. However, the observed alteration in muscle architecture after the imposed perturbation, could lead to an underestimation (1–3%) of joint moment enhancement due to the force–length relationship of the triceps surae. Furthermore, the enhancement observed was independent of the shortening–stretch amplitude, velocity and stimulation frequency.     

Structural and functional link between the mitochondrial network and the endoplasmic reticulum

Mitochondrial and endoplasmic reticulum (ER) networks are fundamental for the maintenance of cellular homeostasis and for determination of cell fate under stress conditions. Recent structural and functional studies revealed the interaction of these networks. These zones of close contact between ER and mitochondria called MAM (mitochondria associated membranes) support communication between the two organelles including bioenergetics and cell survival. The existence of macromolecular complexes in these contact sites has also been revealed. In this contribution, we will review: (i) the ER and mitochondria structure and their dynamics, (ii) the basic principles of ER mitochondrial Ca²⁺ transport, (iii) the physiological/pathological role of this cross-talk.     

Unveiling introgression in bumblebee (Bombus terrestris) populations through mitogenome-based markers

The bumblebee, Bombus terrestris, is an important pollinator commercially used on a global scale. The exported subspecies B. t. terrestris has colonised diverse environments, in some cases displacing wild pollinators to the verge of local extinction. In this sense, the native Iberian subspecies B. t. lusitanicus may be threatened by the subspecies B. t. terrestris, naturally distributed from the Pyrenees to Central Europe but also observed in southern Spain due to escapes from commercial nests. Mitochondrial genomes have a low recombination rate and a small effective population size owing to their maternal inheritance, thus providing an accurate approach to study hybridisation events between populations. Therefore, we present the sequences of the mitogenomes of both subspecies as a molecular framework to select suitable markers to detect possible introgression events between them. We used metagenomics to obtain approximately 17 kbp of the mitogenome from both subspecies. Their mitogenomes differed in 358 bp (excluding the AT-rich region). Four mitogenomic fragments were selected to be tested as subspecific diagnostic markers. A RFLP detected in the gene nad2 (NADH dehydrogenase subunit 2) has proven to be an efficient, quick and cost-effective tool to assess the dispersion of the non-endemic subspecies into Iberian native populations. Subspecific haplotypes were observed in both morphological subspecies, suggesting introgression events in the northern natural contact area and in the new human-mediated contact area in the south of the Iberian Peninsula.     

Vesicular glutamate transporter modulates sex differences in dopamine neuron vulnerability to age‐related neurodegeneration

Age is the greatest risk factor for Parkinson''s disease (PD) which causes progressive loss of dopamine (DA) neurons, with males at greater risk than females. Intriguingly, some DA neurons are more resilient to degeneration than others. Increasing evidence suggests that vesicular glutamate transporter (VGLUT) expression in DA neurons plays a role in this selective vulnerability. We investigated the role of DA neuron VGLUT in sex‐ and age‐related differences in DA neuron vulnerability using the genetically tractable Drosophila model. We found sex differences in age‐related DA neurodegeneration and its associated locomotor behavior, where males exhibit significantly greater decreases in both DA neuron number and locomotion during aging compared with females. We discovered that dynamic changes in DA neuron VGLUT expression mediate these age‐ and sex‐related differences, as a potential compensatory mechanism for diminished DA neurotransmission during aging. Importantly, female Drosophila possess higher levels of VGLUT expression in DA neurons compared with males, and this finding is conserved across flies, rodents, and humans. Moreover, we showed that diminishing VGLUT expression in DA neurons eliminates females'' greater resilience to DA neuron loss across aging. This offers a new mechanism for sex differences in selective DA neuron vulnerability to age‐related DA neurodegeneration. Finally, in mice, we showed that the ability of DA neurons to achieve optimal control over VGLUT expression is essential for DA neuron survival. These findings lay the groundwork for the manipulation of DA neuron VGLUT expression as a novel therapeutic strategy to boost DA neuron resilience to age‐ and PD‐related neurodegeneration.     

Piericidin A Aggravates Tau Pathology in P301S Transgenic Mice

ObjectiveThe P301S mutation in exon 10 of the tau gene causes a hereditary tauopathy. While mitochondrial complex I inhibition has been linked to sporadic tauopathies. Piericidin A is a prototypical member of the group of the piericidins, a class of biologically active natural complex I inhibitors, isolated from streptomyces spp. with global distribution in marine and agricultural habitats. The aim of this study was to determine whether there is a pathogenic interaction of the environmental toxin piericidin A and the P301S mutation.MethodsTransgenic mice expressing human tau with the P301S-mutation (P301S^+/+) and wild-type mice at 12 weeks of age were treated subcutaneously with vehicle (N = 10 P301S^+/+, N = 7 wild-type) or piericidin A (N = 9 P301S^+/+, N = 9 wild-type mice) at a dose of 0.5 mg/kg/d for a period of 28 days via osmotic minipumps. Tau pathology was measured by stereological counts of cells immunoreative with antibodies against phosphorylated tau (AD2, AT8, AT180, and AT100) and corresponding Western blot analysis.ResultsPiericidin A significantly increased the number of phospho-tau immunoreactive cells in the cerebral cortex in P301S^+/+ mice, but only to a variable and mild extent in wild-type mice. Furthermore, piericidin A led to increased levels of pathologically phosphorylated tau only in P301S^+/+ mice. While we observed no apparent cell loss in the frontal cortex, the synaptic density was reduced by piericidin A treatment in P301S^+/+ mice.DiscussionThis study shows that exposure to piericidin A aggravates the course of genetically determined tau pathology, providing experimental support for the concept of gene-environment interaction in the etiology of tauopathies.