The role of autophagy in survival response induced by 27-hydroxycholesterol in human promonocytic cells.

Autophagy has been shown to be stimulated in advanced atherosclerotic plaques by metabolic stress, inflammation and oxidized lipids. The lack of published studies addressing the potential stimulation of pro-survival autophagy by oxysterols, a family of cholesterol oxidation products, has prompted our study. Thus, the goal of the current study is to elucidate the molecular mechanism of the autophagy induced by 27-hydroxycholesterol (27-OH), that is one of the most abundant oxysterols in advanced atherosclerotic lesions, and to assess whether the pro-oxidant effect of the oxysterol is involved in the given response. Here we showed that 27-OH, in a low micromolar range, activates a pro-survival autophagic response in terms of increased LC3 II/LC3 I ratio and Beclin 1, that depends on the up-regulation of extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K)/Akt pathways as a potential result of an intracellular reactive oxygen species increase provoked by the oxysterol in human promonocytic U937 cells. Moreover, 27-OH induced autophagy is dependent on the relation between nuclear factor erythroid 2 p45-related factor 2 (Nrf2)-dependent antioxidant response and p62. The data obtained highlight the involvement of cholesterol oxidation products in the pathogenesis of oxidative stress related chronic diseases like atherosclerosis. Therefore, deeply understanding the complex mechanism and generating synthetic or natural molecules targeting this survival mechanism might be very promising tools in the prevention of such diseases. Keywords: Oxysterols, 27-hydroxycholesterol, Autophagy, ROS, Survival signalin

Molecular detection and prevalence of feline hemotropic mycoplasmas in Istanbul, Turkey

The aim of this study was to investigate Mycoplasma spp. species in blood samples of the domestic cats from the province of Istanbul, Turkey. Three hundred eighty four blood samples of client-owned cats were used for the identification of Mycoplasma haemofelis (Mhf), Candidatus Mycoplasma haemominutum (CMhm) and Candidatus Mycoplasma turicensis (CMt) by Polymerase Chain Reaction (PCR) and Restriction Fragment Length Polymorphism (RFLP) assays. Out of 384 blood samples, 74 (19.3%) were positive for one of Mycoplasma species. The total prevalence of Mhf, CMhm and CMt infections was 9.9%, 17.7% and 0.8% respectively. The most common species was CMhm. Co-infections were mostly with Mhf/CMhm and the frequency was 8.1%. Two cats were infected with three species. The current study was the first molecular prevalence study of hemotropic mycoplasmas in Istanbul, reporting the presence of CMt for the first time in Turkey. Prevalence of feline mycoplasma was notably high in Istanbul and PCR assay could be preferred rather than the microscopic examination for the diagnosis

Molecular Detection And Typing Of Anaplasma Species In Small Ruminants In Thrace Region Of Turkey

This study was conducted to determine the presence and distribution of Anaplasma ovis and Anaplasma phagocytophilum in small ruminants in Istanbul, Tekirdag, Edirne and Kirklareli provinces in Thrace region of northwestern Turkey during May-September in 2014. A total of 423 blood samples (216 sheep and 207 goats) were collected randomly from small ruminants regardless of the clinical symptoms. Species-specific polymerase chain reaction (PCR) assays, targeting the major surface protein 4 (msp4), were employed for identification of A. ovis and A. phagocytophilum and selected products were confirmed via sequencing. A total of 230 small ruminants (54.37%) were found to be infected with A. ovis and/or A. phagocytophilum. The rates of infected animals for A. ovis and A. phagocytophilum were 50.83% (215/423) and 8.51% (36/423) respectively. Coinfection rate in small ruminants was determined as 4.96% (21/423). Sequence diversity rates of 0-0.94% for A. ovis and 0.41-2.49% for A. phagocytophilum have been observed. This is the first detection of A. ovis and A. phagocytophilum in sheep and goats in Thrace region of northwestern Turkey via polymerase chain reaction and sequence characterization. Further researches are needed to determine the vectors, vector-host interactions and genotypic variants that may affect the presence and distribution of Anaplasma species in the region.WoSScopu

Serum protein electrophoresis in dogs with intestinal parasites

The serum of 66 dogs with intestinal parasites (showing gastrointestinal problems caused by taeniosis, coccidiosis, ancylostomosis, trichuriosis and ascarididosis) was examined by electrophoresis. There were 6 dogs with coccidiosis, 6 dogs with ancylostomosis, 6 dogs with trichuriosis. 24 dogs with taeniosis and 24 dogs with ascarididosis. After agar gel protein electorphoresis of the serum samples, alpha 1 globulin levels were significantly lower in the coccidiosis group than in the other groups (p<0.05). While alpha 2 globulin levels increased in the ancylostomosis group (p<0.05), these levels decreased significantly in the dogs with ascarididosis (p<0.05). There was no change in beta and gamma globulin levels in any of the groups. While the protein level increased in the dogs with taeniosis (p<0.05), these levels were lower in the coccididosis group than in the other groups (p<0.05)

Survival signals induced by low concentrations of 27-hydroxycholesterol in human monocytic cells via ERK activation

Oxysterols are oxidized derivatives of cholesterol, most likely contributing to the development of atherosclerosis. On the basis of the clear findings by previous workers about oxysterol-mediated induction of survival pathways aside that of death pathways, we aimed to identify the main genes and related products involved in the transduction of survival signals and elucidate the relevant molecular mechanisms in human macrophagic cells challenged with 27-hydroxycholesterol in the low micromolar range

Loss of PRMT2 in myeloid cells in normoglycemic mice phenocopies impaired regression of atherosclerosis in diabetic mice

Abstract The regression, or resolution, of inflammation in atherosclerotic plaques is impaired in diabetes. However, the factors mediating this effect remain incomplete. We identified protein arginine methyltransferase 2 (PRMT2) as a protein whose expression in macrophages is reduced in hyperglycemia and diabetes. PRMT2 catalyzes arginine methylation to target proteins to modulate gene expression. Because PRMT2 expression is reduced in cells in hyperglycemia, we wanted to determine whether PRMT2 plays a causal role in the impairment of atherosclerosis regression in diabetes. We, therefore, examined the consequence of deleting PRMT2 in myeloid cells during the regression of atherosclerosis in normal and diabetic mice. Remarkably, we found significant impairment of atherosclerosis regression under normoglycemic conditions in mice lacking PRMT2 (Prmt2 −/−) in myeloid cells that mimic the decrease in regression of atherosclerosis in WT mice under diabetic conditions. This was associated with increased plaque macrophage retention, as well as increased apoptosis and necrosis. PRMT2-deficient plaque CD68+ cells under normoglycemic conditions showed increased expression of genes involved in cytokine signaling and inflammation compared to WT cells. Consistently, Prmt2 −/− bone marrow-derived macrophages (BMDMs) showed an increased response of proinflammatory genes to LPS and a decreased response of inflammation resolving genes to IL-4. This increased response to LPS in Prmt2 −/− BMDMs occurs via enhanced NF-kappa B activity. Thus, the loss of PRMT2 is causally linked to impaired atherosclerosis regression via a heightened inflammatory response in macrophages. That PRMT2 expression was lower in myeloid cells in plaques from human subjects with diabetes supports the relevance of our findings to human atherosclerosis

Nrf2 antioxidant defense is involved in survival signaling elicited by 27-hydroxycholesterol in human promonocytic cells

Cholesterol oxidation products such as oxysterols are considered critical factors in the atherosclerotic plaque formation since they induce oxidative stress, inflammation and apoptotic cell death. 27-hydroxycholesterol (27-OH) is one of the most represented oxysterols in atherosclerotic lesions. We recently showed that relatively low concentrations of 27-OH generated a strong survival signaling through an early and transient increase of cellular ROS level, that enhanced MEK-ERK/PI3K-Akt phosphorylation, in turn responsible of a sustained quenching of ROS production. It remains to identify the link between ERK/Akt up-regulation and the consequent quenching effect on ROS intracellular level that efficiently and markedly delay the pro-apoptotic effect of the oxysterol. Here we report on the potent activation of Nrf2 redox-sensitive transcription factor by low micromolar amount of 27-OH added to U937 promonocytic cells. The 27-OH-exerted induction of Nrf2 and subsequently of the target genes, HO-1 and NQO-1, was proved to be: (i) dependent upon the activation of ERK and Akt pathways, (ii) directly responsible for the quenching of intracellular oxidative stress and by this way (iii) ultimately responsible for the observed oxysterol-induced pro-survival response

Survival signaling elicited by 27-hydroxycholesterol through the combined modulation of cellular redox state and ERK/Akt phosphorylation

The oxysterol 27-hydroxycholesterol (27-OH) is increasingly considered to be involved in a variety of pathophysiological processes, having been shown to modulate cell proliferation and metabolism, and also to exert proinflammatory and proapoptotic effects. This study aimed to elucidate the molecular pathways whereby 27-OH may generate survival signals in cells of the macrophage lineage, and to clarify whether its known prooxidant effect is involved in that process. A net up-regulation of survival signaling, involving the extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K)/Akt phosphorylation pathways, was observed in U937 promonocytic cells cultivated over time in the presence of a low micromolar concentration of the oxysterol. Interestingly, the up-regulation of both kinases was shown to be closely dependent on an early 27-OH-induced intracellular increase of reactive oxygen species (ROS). In turn, stimulation of ERK and PI3K/Akt both significantly quenched ROS steady state and markedly phosphorylated Bad, thereby determining a marked delay of the oxysterol׳s proapoptotic action. The 27-OH-induced survival pathways thus appear to be redox modulated and, if they occur within or nearby inflammatory cells during progression of chronic diseases such as cancer and atherosclerosis, they could significantly impact the growth and evolution of such diseases