Proteomic basis of the antibody response to monkeypox virus infection examined in cynomolgus macaques and a comparison to human smallpox vaccination

Monkeypox is a zoonotic viral disease that occurs primarily in Central and West Africa. A recent outbreak in the United States heightened public health concerns for susceptible human populations. Vaccinating with vaccinia virus to prevent smallpox is also effective for monkeypox due to a high degree of sequence conservation. Yet, the identity of antigens within the monkeypox virus proteome contributing to immune responses has not been described in detail. We compared antibody responses to monkeypox virus infection and human smallpox vaccination by using a protein microarray covering 92-95% (166-192 proteins) of representative proteomes from monkeypox viral clades of Central and West Africa, including 92% coverage (250 proteins) of the vaccinia virus proteome as a reference orthopox vaccine. All viral gene clones were verified by sequencing and purified recombinant proteins were used to construct the microarray. Serum IgG of cynomolgus macaques that recovered from monkeypox recognized at least 23 separate proteins within the orthopox proteome, while only 14 of these proteins were recognized by IgG from vaccinated humans. There were 12 of 14 antigens detected by sera of human vaccinees that were also recognized by IgG from convalescent macaques. The greatest level of IgG binding for macaques occurred with the structural proteins F13L and A33R, and the membrane scaffold protein D13L. Significant IgM responses directed towards A44R, F13L and A33R of monkeypox virus were detected before onset of clinical symptoms in macaques. Thus, antibodies from vaccination recognized a small number of proteins shared with pathogenic virus strains, while recovery from infection also involved humoral responses to antigens uniquely recognized within the monkeypox virus proteome.     

Percutaneous septal ablation for left mid-ventricular obstructive hypertrophic cardiomyopathy: a case report

Background

Mid-ventricular obstructive hypertrophic cardiomyopathy (MVOHC) is a rare type of cardiomyopathy. The diagnosis is based on the hourglass appearance on the left ventriculogram and the presence of pressure gradient between apical and basal chamber of the ventriculum on the hemodynamic assessment.

Case presentation

The present case represents successful percutaneous treatment with septal ablation to patient with MVOHC associated with systolic anterior motion of the mitral valve and obstruction at both the mid-ventricular and outflow levels.

Conclusion

Alcohol septal ablation has been proposed as less invasive alternatives to surgery in patients with MVOHC.     

Combined effect of epirubicin and lymphokine-activated killer cells on the resistant human breast cancer cells

Accumulating evidence suggests the concept that epirubicin and lymphokine-activated killer (LAK) cells cytotoxicity may be mediated by free radicals generation and P-glycoprotein-positive (Pg-p⁺) cancer cells are more sensitive for LAK cells than their drug-sensitive parental lines. We tested this hypothesis further by exposing drug-sensitive (WT) and epirubicin-resistant MCF-7 human breast tumor cells to epirubicin and LAK cells. Subsequently, we monitored cell proliferation as a measure of cytotoxicity. The cytotoxicity of epirubicin, LAK, and LAK + epirubicin (1/10 of IC₅₀) was evaluated in 400-fold epirubicin resistant MCF-7 EPI^R (P-glycoprotein overexpressing) and drug-sensitive MCF-7 WT cells. IC₅₀ values were measured using the MTT cytotoxicity test. The MCF-7 EPI^R cells exhibited an increased susceptibility to LAK cells than did the MCF-7 WT cells. P-gp⁺ MCF-7 EPI^R cells were lysed by human LAK cells to a greater extend than were their drug-sensitive counterparts. LAK + epirubicin combined treatment increased susceptibility of MCF-7 WT and MCF-7 EPI^R cells to LAK cells cytotoxicity. For both cell lines, cytotoxicity was dependent upon the concentration of the epirubicin and effector cell/target cell (E/T) ratio. The resistance of MCF-7 EPI^R cells to epirubicin appears to be associated with a developed tolerance to superoxide, most likely because of a tree-fold increase in superoxide dismutase (SOD) activity and 13-fold augmented selenium dependent glutathione peroxidase (GSH-Px) activity. Acting in concert, these two enzymes would decrease the formation of hydroxyl radical from reduced molecular oxygen intermediates. The addition of SOD decreased cytotoxicity of epirubicin and LAK cells. Taken together, these observations support the role of oxygen radicals in the cytotoxicity mechanism of epirubicin and suggest further that the development of resistance to this drug by the MCF-7 EPI^R tumor cells may have a component linked to oxygen free radicals. It is proposed that production of reactive oxygen species by the treatment of epirubicin and LAK cells can cause cytotoxicity of MCF-7 WT and MCF-7 EPI^R cells. SOD, catalase, GSH-Px, GST (glutathione S-transferase), and GSH (reduced glutathione) must be considered as part of the intracellular antioxidant defense mechanism of MCF-7 WT and MCF-7 EPI^R cells against reactive oxygen species.     

Effect of a taurine treatment on the regression of existing atherosclerotic lesions in rabbits fed on a high-cholesterol diet

We studied whether taurine has any regressive effect on existing atherosclerotic lesions and lipid peroxidation in rabbits fed on a high-cholesterol (HC) diet. The cholesterol, triglyceride, malondialdehyde (MDA) and diene conjugate (DC) levels, as well as the aortic histopathological findings were examined in rabbits that had been fed on a cholesterol-containing diet for 8 months [0.5% cholesterol (w/w) for 3 months and subsequently 0.25% cholesterol (w/w) for 5 months], and then for a further 4 months on a normal diet with or without taurine treatment [1% (w/v) in the drinking water]. High levels of lipid and lipid peroxide induced by the HC diet were observed to decline in the plasma, liver and aorta of atherosclerotic rabbits, as well as a slight retardation in aortic atherosclerotic lesions during the regression period. Although no significant differences in the lipid and lipid peroxide levels in the plasma and aorta were found between the regressed groups with or without the taurine treatment, the extent of atherosclerotic lesions in the aorta was less in the taurine-treated regressed group than in the non-treated regressed group. However, the liver MDA and DC levels were lower in the regressed rabbits with the taurine treatment in the non-treated group. These results indicate that the taurine treatment may accelerate the regression of cholesterol-induced atherosclerotic lesions in rabbits without having any effect on the plasma and aorta lipid and lipid peroxide levels.     

Toxic concentrations of selenite shortens repolarization phase of action potential in rat papillary muscle

Selenium (Se) has long been recognized as both an essential mammalian nutrient and a hazardous element. Sodium selenite is commonly used as a dietary supplement for the treatment of Se deficiency. On the other hand, chronic Se toxicity has been demonstrated to affect the major organs, including the heart, in experimental animals. This study examines the effects of high concentrations of extracellular selenite (in the range of 0.001–1 mM) application into the recording bath on the electrical properties of rat papillary muscles. Conventional glass semifloating microelectrodes were used to record intracellular action potentials (APs) in isolated rat papillary muscles. The amplitude of APs and the resting membrane potential of papillary muscles were not changed following a 20-min selenite (1 mM) application compared to the first minute of its application. Freshly isolated ventricular myocytes by an enzymatic method were used to determine the selenite-induced alterations in Na⁺ currents. Na⁺ currents, measured at 22°C, by the whole-cell patch-clamp technique, decreased by 38±8% in the presence of 1 mM selenite for 5 min. These selenite-induced effects were not reversed, but are restored by dithiothreitol (1 mM). These results demonstrated that toxic concentrations of selenite induced a significant shortening in AP duration as a result of an increase in the rate of repolarization. Our findings also suggest that a decrease in Na⁺ currents, in addition to Ca²⁺ currents, may play a role in the shortening of AP duration in rat papillary muscles.     

Stresses applied for the re-programming of plant microspores towards in vitro embryogenesis

Microspore embryogenesis is the most commonly used method to produce doubled haploids. It is based on the ability of a single haploid cell, the microspore, to de-differentiate and regenerate into a whole plant after being exposed to stresses, such as low or high temperatures, carbon starvation and colchicine. Some stresses such as temperature treatments and carbon starvation have been used with success in many plant species, whereas others such as colchicine had limited application in a few species. Reports on the application of whole plant treatments with feminizing agents on inflorescences and buds are scarce. Furthermore, the technical means to apply some stresses such as γ-irradiation are not readily available. Recently, novel stresses such as pH, inducer chemicals, carrageenan oligosaccharides and heavy metals were reported to induce microspore embryogenesis. It remains to be seen, however, whether these stresses are effective in a wider range of species. Finally, pretreatment of cultured cells with high concentrations of 2,4-D efficiently induces somatic embryogenesis in several species (carrot, alfalfa). However, reports on the use of this particular chemical stress are not available in microspore embryogenesis. The paper presented here gives an overview of various stresses and mechanisms of action of these stresses in inducing microspore embryogenesis.     

Physiological and Biochemical Mechanisms of Salinity Tolerance in <i>Carex morrowii</i> Boott

Carex species are widely used in many parts of the world and contain a large number of ecologically diverse species. Among the Carex species, some of them are known to be glycophytes, while others are halophytes. Carex morrowii Boott (Cyperaceae) is resistant to trample through their root structure and has an essential ornamental value in the landscape with their leaves. However, no information was found about the level of salinity tolerance/ sensitivity of the Carex morrowii among these species. In the present study, changes in trace element contents (Na, K, Ca, Cu, Mn, Mg, Ni, Fe, P, Zn, and N) and their transport from roots to leaves, osmotic regulation, alterations in chlorophyll and carotenoid contents, nitrogen assimilation (nitrate reductase activity; NRA) and total soluble protein content in both roots and leaves of Carex morrowii under different salinity concentrations (50 mM, 100 mM, 200 mM and 300 mM NaCl) were examined in detail. Our study provides the first detailed data concerning the responses of leaves and roots and the determination of the level of salinity tolerance/sensitivity of the Carex morrowii. The K+ /Na+ ratio was preserved up to 200 mM NaCl, and accordingly, the element uptake and transport ratios showed that they could control moderate NaCl levels. Ca homeostasis that is maintained even in 200 mM NaCl concentration can be effective in maintaining the structural integrity and selective permeability of the cell membranes, while 300 mM NaCl concentration caused decreased photosynthetic pigments, and deterioration in element content and compartmentation. Moreover, these data suggest that plant parts of Carex morrowii respond differently against varied levels of salinity stress. Although the decrease in NR activity at 200 mM and 300 mM NaCl concentrations in the leaves, NR activity was maintained in the roots. Consequently, Carex morrowii is moderately tolerant to salinity and the carotenoid content and osmotic regulation of Carex morrowii appears to be instrumental in its survival at different salinity levels. Especially the roots of Carex morrowii have a remarkable role in salinity tolerance.     

Vascular adhesion and transendothelial migration of eosinophil leukocytes

Tissues respond to injury with inflammation in an effort to protect and repair the damaged site. During inflammation, leukocytes typically accumulate in response to certain chemicals produced within the tissue itself. The passage of leukocytes through the vascular lumen into tissues occurs in several phases, including rolling, activation, firm adhesion, transendothelial migration, and subendothelial migration. Although infiltration of eosinophil leukocytes is one of the most important aspects of allergic inflammatory reactions, eosinophils also participate in nonallergic inflammation. Eosinophil accumulation is regulated not only by endothelial adhesion molecules, but also by interactions between eosinophil adhesion molecules and extracellular matrix elements. This review summarizes the regulation of eosinophil leukocyte adhesion and migration. A better understanding of eosinophil recruitment responses may lead to the development of novel therapeutics for chronic allergic diseases.     

Liver and kidney toxicity in chronic use of opioids: An experimental long term treatment model

In this study, histopathological and biochemical changes due to chronic usage of morphine or tramadol in liver and kidney were assessed in rats. Thirty male Wistar rats (180–220 g) were included and divided into three groups. Normal saline (1 ml) was given intraperitoneally as placebo in the control group (n = 10). Morphine group (n = 10) received morphine intraperitoneally at a dose of 4, 8, 10 mg/kg/day in the first, second and the third ten days of the study, respectively. Tramadol group (n = 10), received the drug intraperitoneally at doses of 20, 40 and 80 mg/kg/day in the first, second and the third ten days of the study, respectively. Aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), creatinin, blood urea nitrogen (BUN) and malondialdehyde (MDA) levels were measured in the serum. Liver and kidney specimens were evaluated by light microscopy. Serum ALT, AST, LDH, BUN and creatinin levels were significantly higher in morphine group compared to the control group. Serum LDH, BUN and creatinin levels were significantly increased in the morphine group compared to the tramadol group. The mean MDA level was significantly higher in morphine group compared to the tramadol and control groups (P<0.05). Light microscopy revealed severe centrolobular congestion and focal necrosis in the liver of morphine and tramadol groups, but perivenular necrosis was present only in the morphine group. The main histopathologic finding was vacuolization in tubular cells in morphine and tramadol groups. Our findings pointed out the risk of increased lipid peroxidation, hepatic and renal damage due to long term use of opioids, especially morphine. Although opioids are reported to be effective in pain management, their toxic effects should be kept in mind during chronic usage Presented at the 10th XX Annual ESRA Congress, 6–9 April 2002, Warsaw, Poland.     

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods

IntroductionThe clinical significance of Blastocystis sp. and Dientamoeba fragilis in patients with gastrointestinal symptoms is a controversial issue. Since the pathogenicity of these protists has not been fully elucidated, testing for these organisms is not routinely pursued by most laboratories and clinicians. Thus, the prevalence of these organisms and the subtypes of Blastocystis sp. in human patients in Turkey are not well characterized. This study aimed to determine the prevalence of Blastocystis sp. and D. fragilis in the diarrheic stool samples of immunodeficient and immunocompetent patients using conventional and molecular methods and to identify Blastocystis sp. subtypes using next generation sequencing.Material and methodsIndividual stool specimens were collected from 245 immunodeficient and 193 immunocompetent diarrheic patients between March 2017 and December 2019 at the Gazi University Training and Research Hospital in Ankara, Turkey. Samples were screened for Blastocystis sp. and D. fragilis by conventional and molecular methods. Molecular detection of both protists was achieved by separate qPCRs targeting a partial fragment of the SSU rRNA gene. Next generation sequencing was used to identify Blastocystis sp. subtypes.ResultsThe prevalence of Blastocystis sp. and D. fragilis was 16.7% and 11.9%, respectively as measured by qPCR. The prevalence of Blastocystis sp. and D. fragilis was lower in immunodeficient patients (12.7% and 10.6%, respectively) compared to immunocompetent patients (21.8% and 13.5%, respectively). Five Blastocystis sp. subtypes were identified and the following subtype distribution was observed: ST3 54.4% (n = 37), ST2 16.2% (n = 11), ST1 4.4% (n = 3), ST6 2.9% (n = 2), ST4 1.5% (n = 1), ST2/ST3 11.8% (n = 8) and ST1/ST3 8.8% (n = 6). There was no statistically significant difference in the distribution of Blastocystis sp. subtypes between immunocompetent and immunodeficient patients.Conclusion and recommendationOur findings demonstrated that Blastocystis sp. and D. fragilis are commonly present in immunocompetent and immunodeficient patients with diarrhea. This study is the first to use next generation sequencing to address the presence of Blastocystis sp. mixed subtypes and intra-subtype variability in clinical samples in Turkey.