Augmenting Endogenous Levels of Retinal Annexin A1 Suppresses Uveitis in Mice

PURPOSE: The purpose of this study was to examine the expression of the anti-inflammatory protein Annexin A1 (AnxA1) in mice and human retinae during uveitis and to determine whether local administration of human recombinant AnxA1 (hrAnxA1) can suppress uveitis in mice. METHODS: Retinal sections from mice (healthy normal and uveitis) and postmortem human (no history of eye disease (n = 5) and uveitis (n = 7)) were stained for AnxA1 expression and imaged by immunofluorescence microscopy. AnxA1 cellular expression was determined by colabeling with CD45, glial fibrillary acidic protein (GFAP), and Iba-1 cells, with additional staining of AnxA1 receptors formyl peptide receptor 1 (FPR1) and FPRL1/FPR2. Mice with acute endotoxin-induced uveitis and chronic experimental autoimmune uveitis were treated locally by intravitreal injection with hrAnxA1, and disease was assessed by clinical scoring and quantification of leukocyte infiltrate via flow cytometry. RESULTS: Constitutive expression of AnxA1 was observed in both healthy mouse and human retinae, and its expression increased during uveitis compared to healthy controls. AnxA1 colocalizes predominantly with CD45(+) cells, GFAP(+) macroglia, and to a lesser extent, Iba-1(+) myeloid cells. We also demonstrate that local treatment with hrAnxA1 attenuates the severity of uveitis in mice. CONCLUSIONS: These data indicate that locally expressed AnxA1 is elevated in the retina during intraocular inflammation. We demonstrate that local administration of hrAnxA1 to augment levels results in suppression of uveitis in mice. TRANSLATIONAL RELEVANCE: Our data suggest that elevated expression of retinal AnxA1 in human uveitis may be immunoregulatory and that local supplementation with hrAnxA1 may provide a potential novel treatment for inflammatory eye diseases such as noninfectious uveitis

Gene therapy restores vision in rd1 mice after removal of a confounding mutation in Gpr179

The rd1 mouse with a mutation in the Pde6b gene was the first strain of mice identified with a retinal degeneration. However, AAV-mediated gene supplementation of rd1 mice only results in structural preservation of photoreceptors, and restoration of the photoreceptor-mediated a-wave, but not in restoration of the bipolar cell-mediated b-wave. Here we show that a mutation in Gpr179 prevents the full restoration of vision in rd1 mice. Backcrossing rd1 with C57BL6 mice reveals the complete lack of b-wave in a subset of mice, consistent with an autosomal recessive Mendelian inheritance pattern. We identify a mutation in the Gpr179 gene, which encodes for a G-protein coupled receptor localized to the dendrites of ON-bipolar cells. Gene replacement in rd1 mice that are devoid of the mutation in Gpr179 successfully restores the function of both photoreceptors and bipolar cells, which is maintained for up to 13 months. Our discovery may explain the failure of previous gene therapy attempts in rd1 mice, and we propose that Grp179 mutation status should be taken into account in future studies involving rd1 mice

The use of histological analysis for the detection of somatic embryos in sugarcane

The aim of this study was to establish an in vitro system for the induction, maturation and regeneration of somatic embryo in sugarcane from buds of cultivar RB 867515. Embryogenic calluses were obtained on semi-solid MS medium supplemented with 4.42 mg L-1 2,4-D. After four weeks of culture of explants on the callus induction medium, globular structures were obtained. At the end of 20 days in maturation medium, somatic embryos were observed. Histological analysis showed somatic embryos with caulinarand root apex, protodermal tissue, and the vascular system, which  apparently has no connection with the vascular tissue explant that gave rise to it confirming the presence of the somatic embryo. The embryos were transferred to regeneration medium containing 1 mg L-1 GA3 and BAP, and after 1 to 2 weeks of culture, green points were observed, indicating the beginning of the formation of shoots. Key words: Saccharum spp, bud culture, 2.4-D, morphogenetic pathway, embryogenesis, plant regeneration

Exploitation promotes earlier sex change in a protandrous patellid limpet, Patella aspera Röding, 1798

Exploitation of organisms can prompt the reduction in the number and size of target populations consequently affecting reproductive output and replenishment. Here, we investigated the effects of exploitation on the population structure of a protandrous patellid limpet, Patella aspera, an overexploited Macaronesian endemic. Timed dives were used to collect animals across eleven islands of Macaronesia. Individuals were inspected for sex, size, and gonad stage. Using catch effort (time per person) per island coastal perimeter as a surrogate for exploitation intensity, we found that limpet abundance (CPUE) and mean size tended to decrease with exploitation intensity. When considering the sex of animals separately, the size of the largest male, but not females, decreased with exploitation. In contrast, the size of the smallest male remained relatively consistent, whereas the size of the smallest female decreased significantly with exploitation. As exploitation is mostly targeting larger individuals, results suggest that males are compensating the removal of larger females, by undergoing sex change at smaller and presumably earlier sizes. These results have wider implications for the conservation of P. aspera, as a reduction in female size will likely affect the numbers of oocytes produced, hence fecundity. Regulations promoting the protection of the larger-sized animals should be enforced to safeguard the replenishment of the population

Flow cytometric analysis of inflammatory and resident myeloid populations in mouse ocular inflammatory models

Myeloid cells make a pivotal contribution to tissue homeostasis during inflammation. Both tissue-specific resident populations and infiltrating myeloid cells can cause tissue injury through aberrant activation and/or dysregulated activity. Reliable identification and quantification of myeloid cells within diseased tissues is important to understand pathological inflammatory processes. Flow cytometry is a valuable technique for leukocyte analysis, but a standardized flow cytometric method for myeloid cell populations in the eye is lacking. Here, we validate a reproducible flow cytometry gating approach to characterize myeloid cells in several commonly used models of ocular inflammation. We profile and quantify myeloid subsets across these models, and highlight the value of this strategy in identifying phenotypic differences using Ccr2-deficient mice. This method will aid standardization in the field and facilitate future investigations into the roles of myeloid cells during ocular inflammation

Zootherapeutics utilized by residents of the community Poço Dantas, Crato-CE, Brazil

<p>Abstract</p> <p>Background</p> <p>Animals have been used as a source of medicine in Brazil since ancient times, and have played a significant role in healing practices. Specifically in Northeast Brazil, zootherapy is a very common practice, and together with medicinal plants, it plays an important role as a therapeutic alternative. In the state of Ceara, no works have been carried out on rural communities with regard to use of zootherapeutics, even though the practice of zootherapy is common in this region. Therefore, the aim of this study was to analyze the use of medicinal animals in a rural community (Poco Dantas) in the municipality of Crato, Ceara, Brazil.</p> <p>Methods</p> <p>The field survey was carried out from October 2008 to January 2009 by conducting interviews using structured questionnaires with 72 people (33 men and 39 women), who provided information on animal species used as remedies, body parts used to prepare the remedies, and ailments for which the remedies were prescribed. We calculated the informant consensus factor (ICF) to determine the consensus over which species are effective for particular ailments, as well as the species use value (UV) to determine the extent of utilization of each species.</p> <p>Results</p> <p>A total of 29 species, distributed in 17 families were categorized as having some medicinal property. The taxa most represented were: mammals (9), insects (7), reptiles and birds (4). <it>Progne chalybea</it>, a species not previously recorded as being of medicinal use, was cited in the present work, where it is utilized in the treatment of alcoholism. The animals are used in the treatment of 34 diseases or symptoms, where sore throat, inflammations and cough are the ailments with the greatest number of citations.</p> <p>Conclusion</p> <p>The data show that zootherapy represents an important therapeutic alternative for the inhabitants of the community. New studies on medicinal fauna should be conducted with the aim of determining the exploitation level of the species utilized, promoting sustainable development of medicinal species that are eventually threatened, and preserving and disseminating the knowledge developed by traditional individuals of the community.</p

Evaluation of microvessel density and p53 expression in pancreatic adenocarcinoma

OBJECTIVE: To evaluate the prognostic significance of microvessel density and p53 expression in pancreatic cancer. METHODS: Between 2008 and 2012, 49 patients with pancreatic adenocarcinoma underwent resection with curative intention. The resected specimens were immunohistochemically stained with anti-p53 and anti-CD34 antibodies. Microvessel density was assessed by counting vessels within ten areas of each tumoral section a highpower microscope. RESULTS: The microvessel density ranged from 21.2 to 54.2 vessels/mm2. Positive nuclear staining for p53 was found in 20 patients (40.6%). The overall median survival rate after resection was 24.1 months and there were no differences in survival rates related to microvessel density or p53 positivity. Microvessel density was associated with tumor diameter greater than 3.0 cm and with R0 resection failure. CONCLUSIONS: Microvessel density was associated with R1 resection and with larger tumors. p53 expression was not correlated with intratumoral microvessel density in pancreatic adenocarcinoma

Apoptosis of non-parasitized red blood cells in malaria: a putative mechanism involved in the pathogenesis of anaemia

<p>Abstract</p> <p>Background</p> <p>Severe anaemia is a common complication of <it>Plasmodium falciparum </it>malaria in hyperendemic regions. Premature elimination of non-parasitized red blood cells (nRBC) has been considered as one mechanism involved in the genesis of severe malaria anaemia. It has been reported that apoptosis can occur in RBC and, consequently, this cell death process could contribute to anaemia. This study was performed to evaluate the susceptibility of nRBC to apoptosis in a malaria anaemia murine model.</p> <p>Methods</p> <p>Balb/c mice were intraperitonially inoculated with 1 × 10⁶<it>P. yoelii </it>17XL parasitized RBC (pRBC) and, then, parasitaemia and anaemia were monitored. Apoptosis in both pRBC and nRBC was assessed during early and late phases of infection by flow cytometry using Syto 16 and annexin V-PE double staining and forward scatter measurement.</p> <p>Results</p> <p>As expected, experimental infection of Balb/c mice with <it>Plasmodium yoelii </it>17XL parasites was characterized by progressive increase of parasitaemia and acute anaemia, leading to death. Flow cytometry analysis showed that a number of pRBC was in the apoptotic process. It was noteworthy that the increase of nRBC apoptosis levels occurred in the late phase of infection, when anaemia degree was notably accentuated, while no significant alteration was observed in the early phase.</p> <p>Conclusion</p> <p>The increased levels of nRBC apoptosis herein firstly reported, in malaria infection could represent a putative mechanism worsening the severity of malarial anaemia.</p

Essential Hypertension Is Associated With Changes in Gut Microbial Metabolic Pathways A Multisite Analysis of Ambulatory Blood Pressure

Recent evidence supports a role for the gut microbiota in hypertension, but whether ambulatory blood pressure is associated with gut microbiota and their metabolites remains unclear. We characterized the function of the gut microbiota, their metabolites and receptors in untreated human hypertensive participants in Australian metropolitan and regional areas. Ambulatory blood pressure, fecal microbiome predicted from 16S rRNA gene sequencing, plasma and fecal metabolites called short-chain fatty acid, and expression of their receptors were analyzed in 70 untreated and otherwise healthy participants from metropolitan and regional communities. Most normotensives were female (66%) compared with hypertensives (35%, P<0.01), but there was no difference in age between the groups (59.2±7.7 versus 60.3±6.6 years old). Based on machine learning multivariate covariance analyses of de-noised amplicon sequence variant prevalence data, we determined that there were no significant differences in predicted gut microbiome α- and β-diversity metrics between normotensives versus essential or masked hypertensives. However, select taxa were specific to these groups, notably Acidaminococcus spp., Eubacterium fissicatena, and Muribaculaceae were higher, while Ruminococcus and Eubacterium eligens were lower in hypertensives. Importantly, normotensive and essential hypertensive cohorts could be differentiated based on gut microbiome gene pathways and metabolites. Specifically, hypertensive participants exhibited higher plasma acetate and butyrate, but their immune cells expressed reduced levels of short-chain fatty acid-activated GPR43 (G-protein coupled receptor 43). In conclusion, gut microbial diversity did not change in essential hypertension, but we observed a significant shift in microbial gene pathways. Hypertensive subjects had lower levels of GPR43, putatively blunting their response to blood pressure-lowering metabolites

Harmonic Analysis of Boolean Networks: Determinative Power and Perturbations

Consider a large Boolean network with a feed forward structure. Given a probability distribution on the inputs, can one find, possibly small, collections of input nodes that determine the states of most other nodes in the network? To answer this question, a notion that quantifies the determinative power of an input over the states of the nodes in the network is needed. We argue that the mutual information (MI) between a given subset of the inputs X = {X_1, ..., X_n} of some node i and its associated function f_i(X) quantifies the determinative power of this set of inputs over node i. We compare the determinative power of a set of inputs to the sensitivity to perturbations to these inputs, and find that, maybe surprisingly, an input that has large sensitivity to perturbations does not necessarily have large determinative power. However, for unate functions, which play an important role in genetic regulatory networks, we find a direct relation between MI and sensitivity to perturbations. As an application of our results, we analyze the large-scale regulatory network of Escherichia coli. We identify the most determinative nodes and show that a small subset of those reduces the overall uncertainty of the network state significantly. Furthermore, the network is found to be tolerant to perturbations of its inputs