Outside-in, inside-out: Proteomic analysis of endothelial stress mediated by 7-ketocholesterol.

Oxysterols are cholesterol oxidation products formed through enzymatic or autoxidation mechanisms. 7-ketocholeterol (7KC) is one of most abundant oxysterols found in atherosclerotic lesions. Its role in atherosclerosis pathogenesis has been broadly studied in a variety of models. The arterial microenvironment is a multicellular dynamic compartment that, among other systemic factors, is continuously stimulated by 7KC. Endothelial cells have a key role on that environment, being in intimate contact with both the blood stream and the vessel wall, the site of disease origin. 7KC has been shown to promote endothelial cell death and/or dysfunction, depending on its concentration. However, its contribution to the cell microenvironment through cell stimulation has not received much attention. Here we applied mass spectrometry-based proteomics followed by bioinformatics workflow to analyze the effect of a non-toxic 7KC concentration on endothelial cell protein expression and secretion in vitro. Trypsin digests were prepared from the secretome of the endothelial cells and from the total cell pellet after 24h exposure to 7KC. All samples were analyzed by high resolution and accurate mass nano-LC MS/MS. After database search and statistical analysis, differentially expressed proteins were selected for further studies. Our workflow identified 1805 secreted proteins and 2203 intracellular proteins, and of these, 48 and 53, respectively, were regulated. Regulated proteins upon 7KC exposure are involved in unfolded protein response, vascular homeostasis, and reduced control of angiogenesis. Moreover, blood coagulation was another main pathway regulated through Tissue Factor Pathway Inhibitor (TFPI), an antithrombotic agent associated with coronary disease that we found to be more than 2 times downregulated. Taken together, these data show differential endothelial protein regulation and secretion upon 7KC exposure for short time periods under non-toxic conditions. Herewith, these data support the role of 7KC in atherosclerosis pathophysiology and thus reinforce the deleterious effect of endothelial cells stress in the arterial microenvironment.

Detection of Plasmodium falciparum and Plasmodium vivax subclinical infection in non-endemic region: implications for blood transfusion and malaria epidemiology.

BACKGROUND: In Brazil, malaria is endemic in the Amazon River basin and non-endemic in the extra-Amazon region, which includes areas of São Paulo state. In this state, a number of autochthonous cases of malaria occur annually, and the prevalence of subclinical infection is unknown. Asymptomatic infections may remain undetected, maintaining transmission of the pathogen, including by blood transfusion. In these report it has been described subclinical Plasmodium infection in blood donors from a blood transfusion centre in São Paulo, Brazil. METHODS: In this cross-sectional study, representative samples of blood were obtained from 1,108 healthy blood donors at the Fundação Pró-Sangue Hemocentro de São Paulo, the main blood transfusion centre in São Paulo. Malaria exposure was defined by the home region (exposed: forest region; non-exposed: non-forest region). Real-time PCR was used to detect Plasmodium falciparum and Plasmodium vivax. Subclinical malaria cases were geo-referenced. RESULTS: Eighty-four (7.41%) blood donors tested positive for Plasmodium; 57 of these were infected by P. falciparum, 25 by P. vivax, and 2 by both. The prevalence of P. falciparum and P. vivax was 5.14 and 2.26, respectively. The overall prevalence ratio (PR) was 3.23 (95% confidence interval (CI) 2.03, 5.13); P. falciparum PR was 16.11 (95% CI 5.87, 44.21) and P. vivax PR was 0.47 (95% CI 0.2, 1.12). Plasmodium falciparum subclinical malaria infection in the Atlantic Forest domain was present in the mountain regions while P. vivax infection was observed in cities from forest-surrounded areas. CONCLUSIONS: The presence of Plasmodium in healthy blood donors from a region known as non-endemic, which is important in the context of transfusion biosafety, was described. Infected recipients may become asymptomatic carriers and a reservoir for parasites, maintaining their transmission. Furthermore, P. falciparum PR was positively associated with the forest environment, and P. vivax was associated with forest fragmentation.

Características biológicas das células-tronco mesenquimais: [revisão] / Biological characteristics of mesenchymal stem cells: [review]

Células-tronco são células indiferenciadas. Como tal, apresentam uma série de características que as tornam candidatas à utilização terapêutica. As principais características das células-tronco são a capacidade de autorrenovação e de se diferenciarem em diversos tipos celulares. Desta forma, acredita-se que células-tronco presentes nos diferentes tecidos tenham papel regenerativo quando estes sofrem uma lesão ou injúria. Entre os tecidos conhecidos por apresentarem células-tronco após a vida pós-natal, a medula óssea foi a mais estudada, por muitos anos, como fonte tanto de células-tronco hematopoéticas quanto de células-tronco mesenquimais, também denominadas de células mesenquimais estromais da medula óssea ou células estromais mesenquimais multipotentes. Estas células são um grupo de células clonogênicas, presentes no estroma da medula óssea, que, quando submetidas a diferentes estímulos apropriados, são capazes de se diferenciarem em várias linhagens de células, como a osteogênica, a condrogênica e a adipogênica e, possivelmente, em outros tipos celulares não mesodérmicos, como células neurais ou hepatócitos. Nesta revisão, as principais características das células-tronco mesenquimais serão abordadas, incluindo os marcadores moleculares e de membrana, as características de divisão e de diferenciação, a heterogeneidade e as aplicações clínicas potenciais.

Stem cells are undifferentiated cells. They show various characteristics that make them suitable for clinical applications. The main stem cell characteristics are their capacity of autorenewal and of differentiation into different cell lines so it is quite possible that stem cells in different tissues exhibit a regenerative role when these tissues are injured. Bone marrow is the best studied tissue as a source of hematopoietic stem cells as well as mesenchymal stem cells (also known as mesenchymal stromal cells or mesenchymal stromal multipotent cells); clonogenic cells in the bone marrow stroma. They are able to differentiate under specific stimuli in several cell lines including osteogenic, chondrogenic and adipogenic cells, and probably in other nonmesodermic cell lines such as neural cells or hepatocytes. Here the main characteristics of mesenchymal stem cells will be discussed, including the molecular and membrane markers, the division and differentiation properties, the heterogeneity, and the potential clinical applications.

Células-tronco do líquido amniótico / Amniotic fluid stem cells

Desde o primeiro isolamento e cultivo de células-tronco embrionárias humanas, há mais de 10 anos, seu uso na pesquisa e terapia foi inibida por considerações éticas complexas e pelo risco de transformação maligna destas células indiferenciadas após transplante no paciente. As células-tronco adultas são eticamente aceitas e o risco de transformação maligna é muito baixo. Entretanto, seu potencial de diferenciação e sua capacidade proliferativa são limitados. Cerca de 6 anos atrás, a descoberta de célulastronco no líquido amniótico que expressavam Oct-4, um marcador específico de pluripotencialidade, com alta capacidade de proliferação e diferenciação, iniciou um novo campo promissor na área das células-tronco. Estas células têm potencial de se diferenciar em células dos três folhetos germinativos. Não formam tumores in vivo e não levantam os questionamentos éticos associados com as células-tronco embrionárias humanas. Futuras investigações revelarão se as células-tronco do líquido amniótico realmente irão representar um tipo intermediário com vantagens em relação tanto às células-tronco embrionárias quanto às adultas. Este artigo faz uma revisão acerca destes tópicos e das características biológicas das células-tronco do líquido amniótico.

Since the first successful isolation and cultivation of human embryonic stem cells about 10 years ago, their use for research and therapy has been constrained by complex ethical considerations as well as by the risk of development of malignancies of undifferentiated embryonic stem cells after transplantation into the patient. Adult stem cells are ethically acceptable and the risk of tumor development is low. However, their differentiation potential and proliferative capacity are limited. About 6 years ago, the discovery of Oct-4 expressing amniotic fluid stem cells, a specific marker of pluripotency, with a high proliferative capacity, and multilineage differentiation potential, initiated a promising field of research. These cells, indeed, have the potential to differentiate into cells of all three embryonic germ layers. They do not form tumors in vivo and do not raise ethical concerns. Further investigation will reveal whether these cells really are an intermediate cell type with advantages over both embryonic and adult stem cells. This article reviews the biological characteristics of amniotic fluid stem cells.