Proteomic analysis of chicken peripheral blood mononuclear cells after infection by Newcastle disease virus

Characteristic clinical manifestations of Newcastle disease include leukopenia and immunosuppression. Peripheral blood mononuclear cells (PBMCs) are the main targets of Newcastle disease virus (NDV) infection. To survey changes in proteomic expression in chicken PBMCs following NDV infection, PBMC proteins from 30 chickens were separated using two-dimensional electrophoresis (2-DE) and subjected to mass spectrometry analysis. Quantitative intensity analysis showed that the expression of 78 proteins increased more than two-fold. Thirty-five proteins exhibited consistent changes in expression and 13 were identified as unique proteins by matrix assisted laser desorption ionization-time of flight mass spectrometer/mass spectrometer including three that were down-regulated and 10 that were up-regulated. These proteins were sorted into five groups based on function: macromolecular biosynthesis, cytoskeleton organization, metabolism, stress responses, and signal transduction. Furthermore, Western blot analysis confirmed the down-regulation of integrin-linked kinase expression and up-regulation of lamin A production. These data provide insight into the in vivo response of target cells to NDV infection at the molecular level. Additionally, results from this study have helped elucidate the molecular pathogenesis of NDV and may facilitate the development of new antiviral therapies as well as innovative diagnostic methods.

Expresión disminuida de caspasa 3 asociada al polimorfismo del gen del antígeno-4 asociado a linfocito T-citotóxico (CTLA4) en pacientes chilenos con diabetes tipo 1 / Decreased caspase 3 expression and cytotoxic T lymphocyte antigen-4 polymorphism in Chilean patients with type 1 diabetes

Background: Several polymorphisms of the CTLA4 gene have been associated with autoimmune diseases. The activation of induced cell death is the major event and caspase 3 represents the main protein for the apoptotic machinery, especially in lymphocytes. Aim: To correlate CTLA4 polymorphisms with caspase 3 expression in peripheral blood mononuclear cells (PBMC) simulating in vitro the glucose effect. Material and Methods: CTLA4 polymorphisms were determined by restriction fragment length polymorphisms (RFLPs). PBMC from 21 patients with type 1 diabetes aged 8.5 ± 4.3 years and 21 healthy subjects aged 18.3 ± 1.8 years, were stimulated under normal (5 mM) and toxic (14 mM) glucose conditions to assess its effect on the expression and activity of caspase 3. Relative abundance of caspase 3 mRNA was measured by semi quantitative RT-PCR and its activity, by a colorimetric assay. Results: When stimulated with 14mM glucose, PBMC of G allele carriers with type 1 diabetes had significantly lower relative mRNA abundance of caspase 3 (median value = 0.12, range 0.01-0.70 AU) compared with non-carriers (median value = 0.81, range 0.06-1.09 AU). When the incubation was carried out with the lower glucose concentration, a similar profile of caspase 3 activity was observed in diabetic patients carrying G allele (median value = 0.57, range 0.13-1.20 AU) as compared with non-carriers (median value = 0.89, range 0.14-5.50 AU). No significant changes after stimulating with glucose, were observed in PBMCs of the control group. Conclusions: PBMC of recently diagnosed patients with T1D, carrying the G allele in + 49A/G polymorphisms of CTLA4, have a decreased expression and activity of caspase 3.

Telomerase Activity and the Risk of Lung Cancer

Telomerase play a key role in the maintenance of telomere length and chromosome integrity. We have evaluated the association between telomerase activity and the risk of lung cancer in peripheral blood. Telomerase activity in peripheral blood mononuclear cells was measured by a PCR-designed telomeric repeat amplification protocol in 63 lung cancer patients and 190 healthy controls that were matched for age, gender, and smoking status. Telomerase activity was significantly lower in the lung cancer patients than in controls (mean +/- standard deviation; 1.32 +/- 1.65 vs 2.60 +/- 3.09, P < 1 x 10(-4)). When telomerase activity was categorized into quartiles based on telomerase activity in the controls, the risk of lung cancer increased as telomerase activity reduced (Ptrend = 1 x 10(-4)). Moreover, when the subjects were categorized based on the median value of telomerase activity, subjects with low telomerase activity were at a significantly increased risk of lung cancer compared to subjects with high telomerase activity (adjusted odds ratio = 3.05, 95% confidence interval = 1.60-5.82, P = 7 x 10-4). These findings suggest that telomerase activity may affect telomere maintenance, thereby contributing to susceptibility to lung cancer.

Lentinula edodes (Shiitake) mushroom extract protects against hydrogen peroxide induced cytotoxicty in peripheral blood mononuclear cells.

Lentinula edodes (Berk) Pegler, commonly known as Shiitake mushroom has been used as medicinal food in Asian countries, especially in China and Japan and is believed to possess strong immunomodulatory property. In the present study, the methanolic extract of the fruit bodies of L. edodes was investigated for cytoprotective effect against H2O2-induced cytotoxicity in human peripheral blood mononuclear cells (PBMCs) by measuring the activities of xanthine oxidase (XO) and glutathione peroxidase (GPx) . H2O2 at a concentration of 5 μM caused 50% inhibition of PBMCs viability. The extract improved the PBMC viability and exerted a dose-dependent protection against H2O2-induced cytotoxicity. At 100 μg/ml of extract concentration, the cell viability increased by 60% compared with the PBMCs incubated with H2O2 alone. The extract also inhibited XO activity in PBMC, while showing moderate stimulatory effect on GPx. However, in the presence of H2O2 alone, both the enzyme activities were increased significantly. The GPx activity increased, possibly in response to the increased availability of H2O2 in the cell. When the cells were pretreated with the extract and washed (to remove the extract) prior to the addition of H2O2, the GPx and XO activities as well as the cell viability were comparable to those when incubated with the extract alone. Thus, it is suggested that one of the possible mechanisms via which L. edodes methanolic extract confers protection against H2O2-induced oxidative stress in PBMC is by inhibiting the superoxide-producing XO and increasing GPx activity which could rapidly inactivate H2O2.

14kDa protein molecule isolated from garlic suppresses indoleamine 2, 3-dioxygenase metabolites in mononuclear cells in vitro

A wide range of biological activities of garlic in vitro and in vivo have been verified including its antioxidant, antitumor and anti-inflammatory effects. Indoleamine 2,3-dioxygenase [IDO] is an enzyme widely distributed in mammals and is inducible preferentially by IFN-?. IDO degrades the essential amino acid tryptophan to form N-formyl kynurenine. In the present in vitro study, the modulatory effect of 14kDa molecule isolated from garlic on IDO induction was tested. Cultures of mononuclear cells were exposed to 14kDa garlic fraction. Then, their proliferation responses and IDO metabolites were measured. A significant down-regulatory effect of garlic on IDO activity was found and also the proliferation responses of mononuclear cells increased. If these results are verified in vivo, an explanation will be provided on how garlic may interfere in IDO induction, which paves the way for elucidating its specific therapeutic effect in preventing tumor progress

Expression of heme oxygenase-1 in the peripheral blood mononuclear cells from asthmatic patients / 华中科技大学学报（医学）（英德文版）

To explore the expression of heme oxygenase-1 (HO-1) in the peripheral blood mononuclear cells (PBMCs) and its relationship with pulmonary ventilation function in asthmatic patients, 18 asthmatic patients and 18 healthy subjects were selected. HO-1 protein and mRNA levels in PBMCs were measured by immunohistochemical staining and reverse transcription-polymerase chain reaction (RT-PCR), respectively. Blood carbon monoxide Hb (COHb), serum total IgE and pulmonary ventilatory function were observed. Our results showed that the percentage of cells positive for immunohistochemical staining of HO-1 were significantly higher in asthmatic patients (41.72 +/- 7.44) % than that in with healthy subjects (10.45 +/- 4.36) % (P < 0.001) and the optical density of PBMC HO-1 mRNA was higher in asthmatic patients (26.05 +/- 4.14) than that in healthy subjects (10. 82 +/- 4.26) (P < 0.001). The relation analysis showed that PBMC HO-1 protein and mRNA levels had significantly negative relation with FEV1%, PEFR, MEFR50%, respectively (r = -0.51-0.89, P < 0.05-0.001, respectively) and a positive relation with COHb and serum total IgE (r = 0.48-0. 85, 0.05-0.001, respectively). It is concluded that the expression of PBMC HO-1 protein and mRNA increased significantly in asthmatic patients, and HO-1 may play a significant role in the pathogenesis of asthma. The expression of HO-1 may bear a relation with severity of asthma.

In vitro HIV infection of primate lymphocytes

El propósito de este estudio fue el de evaluar la capacidad de los virus del SIDA (VIH-1 y VIH-2) para multiplicarse en las células mononuclearres de la sangre periférica (CMSP) de cuatro especies de primates. CMSP de Cebus apella, patas (Erythrocebus patas), monos verdes (cercopithecus aethiops sabeus) y rhesus (Macaca mulatta) fueron infectados "in vitro" con VIH-1 y con VIH-2. La multiplicación de estos virus se determinó midiendo la actividad de la enzima retrotranscriptasa en los cultivos infectados. Ambos virus produjeron efectos citipáticos en dichos cultivos. Se observó un bajo nivel de multiplicación de los virus VIH-1 y VIH-2 en las células provenientes de monos Cebus. Sin embargo, el virus VIH-2 se multiplicó eficientemente en CMSP de monos rhesus. La capacidad que posee el virus de la inmunodeficiencia humana tipo 2, (VIH-2) de multiplicarse en estas células, podría ser utilizada para en la evaluación "in vivo" de productos antivirales y de vacunas