Dopamine-induced ascorbate release from retinal neurons involves glutamate release, activation of AMPA/Kainate receptors and downstream signaling pathways

Ascorbate, the reduced form of Vitamin C, is one of the most abundant and important low-molecular weight antioxidants in living tissues. Most animals synthesize Vitamin C, but some primates, including humans, have lost this capacity due to disruption in L-gulono-gamma-lactone oxidase gene. Because of this incapacity, those animals must obtain Vitamin C from the diet. Ascorbate is highly concentrated in the central nervous system (CNS), including the retina, and plays essential roles in neuronal physiology. Ascorbate transport into cells is controlled by Sodium Vitamin C Co-Transporters (SVCTs). There are four SVCT isoforms and SVCT2 is the major isoform controlling ascorbate transport in the CNS. Regarding ascorbate release from retinal neurons, Glutamate, by activating its ionotropic receptors leads to ascorbate release via the reversion of SVCT2. Moreover, dopamine, via activation of D1 receptor/cyclic AMP/EPAC2 pathway, also induces ascorbate release via SVCT2 reversion. Because the dopaminergic and glutamatergic systems are interconnected in the CNS, we hypothesized that dopamine could regulate ascorbate release indirectly, via the glutamatergic system. Here we reveal that dopamine increases the release of D-Aspartate from retinal neurons in a way independent on calcium ions and dependent on excitatory amino acid transporters. In addition, dopamine-dependent SVCT2 reversion leading to ascorbate release occurs by activation of AMPA/Kainate receptors and downstream ERK/AKT pathways. Overall, our data reveal a dopamine-to-glutamate signaling that regulates the bioavailability of ascorbate in neuronal cells.This work was supported by grants from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ), and Pró-Reitoria de Pesquisa, Pós-Graduação e Inovação da Universidade Federal Fluminense (PROPPI/UFF). TGE, ID, and NAO were recipients of graduate student fellowships from CAPES. RPC is a research fellow from CNPq and FAPERJ. CCP and RS hold employment contracts financed by national funds through FCT – Fundação para a Ciência e a Tecnologia, I.P., in the context of the program-contract described in paragraphs 4, 5 and 6 of art. 23 of Law no. 57/2016, of August 29, as amended by Law no. 57/2017 of July 19

Cell-Cell Communication between Malaria-Infected Red Blood Cells via Exosome-like Vesicles

Cell-cell communication is an important mechanism for information exchange promoting cell survival for the control of features such as population density and differentiation. We determined that Plasmodium falciparum-infected red blood cells directly communicate between parasites within a population using exosome-like vesicles that are capable of delivering genes. Importantly, communication via exosome-like vesicles promotes differentiation to sexual forms at a rate that suggests that signaling is involved. Furthermore, we have identified a P. falciparum protein, PfPTP2, that plays a key role in efficient communication. This study reveals a previously unidentified pathway of P. falciparum biology critical for survival in the host and transmission to mosquitoes. This identifies a pathway for the development of agents to block parasite transmission from the human host to the mosquito.Neta Regev-Rudzki, Danny W. Wilson, Teresa G. Carvalho, Xavier Sisquella, Bradley M. Coleman, Melanie Rug, Dejan Bursac, Fiona Angrisano, Michelle Gee, Andrew F. Hill, Jake Baum, Alan F. Cowma

Reconstruction and flux-balance analysis of the Plasmodium falciparum metabolic network

In the paper we present a metabolic reconstruction and flux-balance analysis (FBA) of Plasmodium falciparum, the primary agent of malaria. The compartmentalized metabolic network of the parasite accounts for 1001 reactions and 616 metabolites. Enzyme–gene associations were established for 366 genes and 75% of all enzymatic reactions.The model was able to reproduce phenotypes of experimental gene knockout and drug inhibition assays with up to 90% accuracy. The model also can be used to efficiently integrate mRNA-expression data to improve the accuracy of metabolic predictions.Using FBA of the reconstructed metabolic network, we identified 40 enzymatic drug targets (i.e. in silico essential genes) with no or very low sequence identity to human proteins.We experimentally tested one of the identified drug targets, nicotinate mononucleotide adenylyltransferase, using a recently discovered small-molecule inhibitor

First GIS analysis of modern stone tools used by wild chimpanzees (Pan troglodytes verus) in Bossou, Guinea, West Africa

Stone tool use by wild chimpanzees of West Africa offers a unique opportunity to explore the evolutionary roots of technology during human evolution. However, detailed analyses of chimpanzee stone artifacts are still lacking, thus precluding a comparison with the earliest archaeological record. This paper presents the first systematic study of stone tools used by wild chimpanzees to crack open nuts in Bossou (Guinea-Conakry), and applies pioneering analytical techniques to such artifacts. Automatic morphometric GIS classification enabled to create maps of use wear over the stone tools (anvils, hammers, and hammers/anvils), which were blind tested with GIS spatial analysis of damage patterns identified visually. Our analysis shows that chimpanzee stone tool use wear can be systematized and specific damage patterns discerned, allowing to discriminate between active and passive pounders in lithic assemblages. In summary, our results demonstrate the heuristic potential of combined suites of GIS techniques for the analysis of battered artifacts, and have enabled creating a referential framework of analysis in which wild chimpanzee battered tools can for the first time be directly compared to the early archaeological record.Leverhulme Trust [IN-052]; MEXT [20002001, 24000001]; JSPS-U04-PWS; FCT-Portugal [SFRH/BD/36169/2007]; Wenner-Gren Foundation for Anthropological Researc

Differential effects of antigens from L. braziliensis isolates from disseminated and cutaneous leishmaniasis on in vitro cytokine production

BACKGROUND: Disseminated leishmaniasis is an emerging infectious disease, mostly due to L. braziliensis, which has clinical and histopathological features distinct from cutaneous leishmaniasis. METHODS: In the current study we evaluated the in vitro production of the cytokines IFN-γ, TNF-α, IL-5 and IL-10 by peripheral blood mononuclear cells (PBMC) from 15 disseminated leishmaniasis and 24 cutaneous leishmaniasis patients upon stimulation with L. braziliensis antigens genotyped as disseminated leishmaniasis or cutaneous leishmaniasis isolates. RESULTS: Regardless of the source of L. braziliensis antigens, PBMC from cutaneous leishmaniasis patients produced significantly higher IFN-γ than PBMC from disseminated leishmaniasis patients. Levels of TNF-α by PBMC from cutaneous leishmaniasis patients were significantly higher than disseminated leishmaniasis patients only when stimulated by genotyped cutaneous leishmaniasis antigens. The levels of IL-5 and IL-10 production by PBMC were very low and similar in PBMCs from both disseminated leishmaniasis and cutaneous leishmaniasis patients. The immune response of each patient evaluated by the two L. braziliensis antigens was assessed in a paired analysis in which we showed that L. braziliensis genotyped as disseminated leishmaniasis isolate was more potent than L. braziliensis genotyped as cutaneous leishmaniasis isolate in triggering IFN-γ and TNF-α production in both diseases and IL-5 only in cutaneous leishmaniasis patients. CONCLUSION: This study provides evidence that antigens prepared from genotypically distinct strains of L. braziliensis induce different degrees of immune response. It also indicates that both parasite and host play a role in the outcome of L. braziliensis infection

Optimized Enzymatic Synthesis of Hesperidin Fatty Acid Esters in a Two-Phase System Containing Ionic Liquid

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Response surface methodology (RSM) based on a five-level, three-variable central composite design (CCD) was employed for modeling and optimizing the conversion yield of the enzymatic acylation of hesperidin with decanoic acid using immobilized Candida antarctica lipase B (CALB) in a two-phase system containing [bmim]BF(4). The three variables studied (molar ratio of hesperidin to decanoic acid, [bmim]BF(4)/acetone ratio and lipase concentration) significantly affected the conversion yield of acylated hesperidin derivative. Verification experiments confirmed the validity of the predicted model. The lipase showed higher conversion degree in a two-phase system using [bmim] BF4 and acetone compared to that in pure acetone. Under the optimal reaction conditions carried out in a single-step biocatalytic process when the water content was kept lower than 200 ppm, the maximum acylation yield was 53.6%.16871717182Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [09/09224-3

Ameloblastic neoplasia spectrum : a cross-sectional study of MMPS expression and proliferative activity

Objective. To compare the proliferation and the expression of matrix metalloproteinases (MMPs; MMP-2 and MMP-9) in solid and unicystic ameloblastomas with ameloblastic carcinomas. Study Design.Five cases of ameloblastic carcinoma (AC), 18 cases of solid ameloblastoma (SA), and seven of unicystic ameloblastoma (UA) were selected. The immunohistochemical expression of MMPs was assessed by the percentage of positive tumor cells and stained stroma. The mean argyrophilic nucleolar organizer region (AgNOR) and the percentage of cells with more than one AgNOR per nucleus were evaluated. Results. Statistically significant higher mean AgNOR was observed in AC than in SA and UA. MMP-2 was expressed similarly in tumor and stroma among groups. MMP-9 was higher in the stroma of SA than that of UA (P = .0484). Conclusions. The cell proliferation was related to the greater aggressiveness of AC. High expression of MMP-2 and MMP-9 in all lesions highlighted the importance of these enzymes in the biology of ameloblastic tumors

Removal of Heterologous Sequences from Plasmodium falciparum Mutants Using FLPe-Recombinase

Genetically-modified mutants are now indispensable Plasmodium gene-function reagents, which are also being pursued as genetically attenuated parasite vaccines. Currently, the generation of transgenic malaria-parasites requires the use of drug-resistance markers. Here we present the development of an FRT/FLP-recombinase system that enables the generation of transgenic parasites free of resistance genes. We demonstrate in the human malaria parasite, P. falciparum, the complete and efficient removal of the introduced resistance gene. We targeted two neighbouring genes, p52 and p36, using a construct that has a selectable marker cassette flanked by FRT-sequences. This permitted the subsequent removal of the selectable marker cassette by transient transfection of a plasmid that expressed a 37°C thermostable and enhanced FLP-recombinase. This method of removing heterologous DNA sequences from the genome opens up new possibilities in Plasmodium research to sequentially target multiple genes and for using genetically-modified parasites as live, attenuated malaria vaccines