Development of a chemiluminescence immunoassay using recombinant non-structural epitope-based proteins to accurately differentiate foot-and-mouth disease virus-infected and vaccinated bovines.

The contamination of inactivated vaccine with non-structural proteins (NSPs) leads to a high false-positive rate, which is a substantial barrier to accurately differentiate foot-and-mouth disease virus (FMDV)-infected animals from vaccinated animals. To address this problem, a new chemiluminescence immunoassay (CLIA) method was developed to detect antibodies targeting the two recombinant epitope-based proteins located in 3A and 3B. The 3Aepitp-3Bepitp CLIA exhibited a diagnostic sensitivity of 94.0% and a diagnostic specificity of 97.5% for the detection of serum samples (naïve bovines, n = 52, vaccinated bovines, n = 422, infected bovines, n = 116) from animals with known status. The CLIA method also had a concordance rate of 88.1% with the PrioCHECK FMDV NSP ELISA based on the detection of 270 serum samples from the field. Importantly, the 3Aepitp-3Bepitp CLIA produced no false-positives when used to detect FMDV in samples from bovines that had been vaccinated up to five times, and it was demonstrated a low false-positive rate when the bovines had been vaccinated up to ten (2.15%) and fifteen times (5.93%). Therefore, the 3Aepitp-3Bepitp CLIA detects FMDV in samples from frequently vaccinated bovines with high accuracy and represents an alternative method to differentiate FMDV-infected and vaccinated bovines.

Propofol suppresses invasion and induces apoptosis of osteosarcoma cell in vitro via downregulation of TGF-ß1 expression.

OBJECTIVE: Osteosarcoma (OS) is the most common malignant tumor of the bone, with a high mortality rate and poor prognosis. Propofol has been proposed to play a role of antitumor in various cancers. However, the functions and mechanisms of propofol in OS is still not clear. MATERIALS AND METHODS: The different concentrations of propofol were co-incubated with osteosarcoma MG-63 lines for 72 hrs. Cell proliferation, apoptosis, and invasion were detected by MTT assay, Flow cytometry analysis, and Matrigel invasion assay. Western blot was used to detect the TGF-ß1 protein levels. MG-63 cells were treated with human recombinant TGF-ß1 (rh TGF-ß1) to assess the role of TGF-ß1 in propofol-induced anti-tumor activity. RESULTS: Propofol significantly inhibited cell proliferation and invasion and promoted apoptosis of MG-63 lines cells. Propofol also efficiently reduced TGF-ß1 expression. Moreover, restoration of TGF-ß1 by rhTGF-ß1 treatment reversed the effects of propofol on the biological behavior of OS cells. CONCLUSIONS: Propofol can effectively inhibit proliferation and invasion and induce apoptosis of OS cells through, at least partly, downregulation of TGF-ß1 expression.

Effect of Xin Mai Jia on atherosclerosis in rats.

We investigated the therapeutic effect of Xin Mai Jia (XMJ) on atherosclerosis (AS) in rats. Rat models of AS were established by peritoneally injecting vitamin D, feeding a high-fat diet, and inducing balloon injuries in rats. The stomachs of the rats were irrigated continuously for 10 weeks with XMJ. Blood lipid- and hemorheology-related indices of blood samples were detected. Pathological changes in the right common carotid arterial tissues were also determined. The protein expression levels of endothelial nitric oxide synthase, angio-tensin-1, and endothelin-1 were determined by western blotting. XMJ reduced cholesterol, trigylecride, and low-density lipoprotein levels as well as blood viscosity, sedimentation, and hematocrit. Furthermore, XMJ alleviated vascular endothelial injury and reduced/eliminated atherosclerotic plaques. In contrast, XMJ significantly increased the endothelium-dependent relaxing response of the AS rat models. The western blotting results showed that XMJ upregulated endothelial nitric oxide synthase but downregulated angiotensin-1 and endothelin-1. XMJ prevented the development of AS by regulating blood lipid levels, hemorheology, and vascular function.

Knockdown of PRAME enhances adriamycin-induced apoptosis in chronic myeloid leukemia cells.

OBJECTIVE: Leukemia is resistant to currently available chemotherapy, and new strategies have been proposed to improve its efficacy. Such an approach requires know of the mechanisms involved in the resistance and survival of leukemia cells. Previous studied has found that Preferentially Expressed Antigen of Melanoma (PRAME) is overexpressed in the leukemia cells, and knockdown of PRAME promoted apoptosis in leukemia K562 cells. In the present study, we investigated whether inhibition of PRAME could sensitize K562 cells to chemotherapy. MATERIALS AND METHODS: K562 cells were treated with PRAME siRNA, and/or adriamycin (ADR), and cell viability and apoptosis, mRNA and protein expression levels were, then, evaluated. Furthermore, the efficacy of PRAME siRNA combined with ADR was further examined in established xenograft models. RESULTS: PRAME suppression was sufficient to induce spontaneous apoptosis of K562 cells. PRAME knockdown showed antiproliferative effects and induced tumor regression in established K562 xenograft models. ADR showed antitumor activity against K562 cells, co-treatment with PRAME siRNA induced an increased apoptosis rate than the sum of the single-treatment rates and promoted tumor regression without enhanced body weight loss in the K562 xenograft models. CONCLUSIONS: PRAME is responsible for the inherent low levels of spontaneous apoptosis in K562 cells. The combination of PRAME siRNA with ADR induced more intense apoptosis compared with each single treatment. PRAME siRNA in combination with ADR is well tolerated and shows greater efficacy than either agent alone in mouse xenograft models.

Comparative characterization of microRNA profiles of different genotypes of Toxoplasma gondii.

The present study compared the miRNA expression profiles of five Toxoplasma gondii strains, namely RH (Type I, ToxoDB10), TgXD (Type I, ToxoDB10), PRU (Type II, ToxoDB1), QHO (Type II, ToxoDB1) and TgC7 (ToxoDB9), by Solexa deep sequencing, bioinformatics analysis and real-time quantitative PCR. A total of 7, 15, 10, 12 and 10 miRNAs were found from RH, TgXD, PRU, QHO and TgC7 strains, respectively. Thirteen miRNAs were shared by three genotypes, with only one miRNA shared by all of the 5 strains and others shared by 2 or more strains. A large number of targets ranging from 1 to 185 were identified for commonly shared miRNAs and strain-specific miRNAs with complete or nearly complete complementarity. Functional prediction showed that these targets were mostly focused on catalytic activity (191 targets) and binding activity (183 targets). Nonetheless, the majority of targets and most of the miRNAs are related to the virulence or invasion proteins of different strains of T. gondii, including ROP and MIC, as well as some other proteins, such as AMA1, GRA and RHO. The present study characterized comparatively the miRNA profiles of 3 different genotypes of T. gondii, identified genotype-shared miRNAs and strain-specific miRNAs.

Modulation of mouse macrophage proteome induced by Toxoplasma gondii tachyzoites in vivo.

Toxoplasma gondii is an obligate intracellular protozoan parasite, which can invade and multiply within the macrophages of humans and most warm-blooded animals. Macrophages are important effector cells for the control and killing of intracellular T. gondii, and they may also serve as long-term host cells for the replication and survival of the parasite. In the present study, we explored the proteomic profile of macrophages of the specific pathogen-free Kunming mice at 24 h after infection with tachyzoites of the virulent T. gondii RH strain using two-dimensional gel electrophoresis combined with matrix-assisted laser desorption ionization time-of-flight (TOF)/TOF tandem mass spectrometry. Totally, 60 differentially expressed protein spots were identified. Among them, 52 spots corresponded to 38 proteins matching to proteins of the mouse, including actin, enolase, calumenin, vimentin, plastin 2, annexin A1, cathepsin S, arginase-1, arachidonate 12-lipoxygenase, and aminoacylase-1. Functional prediction using Gene Ontology database showed that these proteins were mainly involved in metabolism, structure, protein fate, and immune responses. The findings provided an insight into the interactive relationship between T. gondii and the host macrophages, and will shed new lights on the understanding of molecular mechanisms of T. gondii pathogenesis.

Antagonistic effects of two herbs in Zuojin Wan, a traditional Chinese medicine formula, on catecholamine secretion in bovine adrenal medullary cells.

In order to research the target of superior efficacy and lesser side effects, combination of herbal materials has been applied to phytotherapy for thousands of years in China and some other countries. Zuojin Wan (ZJW), a famous traditional Chinese medicine formula, is used in treating gastric diseases in China. It is composed of two herbs, Rhizoma Coptidis (RC) and Fructus Evodiae (FE) in the ratio of 6: 1(w/w). In the present study, we examined the effects of ZJW, RC, FE and active components isolated from these herbs on catecholamine (CA) secretion and intracellular calcium ([Ca(2+)](i)) in cultured bovine adrenal medullary cells. Extracts of ZJW and RC and berberine, palmatine and jatrorrhizine, components of RC, all inhibited CA secretion and rise in [Ca(2+)](i) induced by acetylcholine (ACh), veratridine (Ver) and/or 56 mM K(+). On the other hand, extract of FE, evodiamine and rutaecarpine, components of FE, stimulated CA secretion and rise in [Ca(2+)](i) induced by ACh. Furthermore, different proportions of RC and FE caused different responses in CA secretion. The present findings suggest that two herbs in ZJW have opposite effects, i.e., inhibitory effect of RC and stimulatory effect of FE, on CA secretion induced by acetylcholine in cultured bovine adrenal medullary cells.

Pulmonary venous anastomosis in lung transplantation without donor left atrium. Experimental and clinical results.

Single lung transplantation now is a therapeutic option for some patients with end-stage lung disease. Cyclosporine immunosuppression and refinements in bronchial anastomosis have been responsible for recent successes. Since 1953, the usual pulmonary venous anastomosis, both in experimental animals and in humans, has been an atrium-to-atrium connection. This technique may limit the availability of usable donor lungs, since the donor heart, along with the atrium, is usually harvested for another recipient. Although techniques can be developed to allow both transplant teams to harvest atrial tissue, this study was undertaken to determine if, in fact, anastomosis with donor left atrium is necessary. Twenty-four dogs were anesthetized and a left thoracotomy performed. After heparinization (3 mg/kg), the pulmonary artery and left atrium were occluded. One of four different pulmonary venous anastomoses was performed at 3.5x magnification: superior pulmonary vein end to end (group I), inferior pulmonary vein end to end (group II), superior pulmonary vein implantation into left atrium (group III), and left atrium-to-left atrium anastomosis as control (group IV). Everting mattress sutures of 7-0 polypropylene were used in groups I, II, and III and 6-0 in group IV. Average crossclamp time for group I, group II, and group IV was 20 minutes. The average crossclamp time for group III was 10 minutes. All anastomoses were patent at the time of 1-week reevaluation. Gross and microscopic examination demonstrated establishment of an intimal lining; organized nonocclusive thrombus was present in only one anastomosis. We conclude that atrium-to-atrium anastomosis is not necessary for a successful single lung transplantation, and that transplantation of a single lobe is feasible. The best alternative is implantation of the pulmonary vein into the left atrium, which will easily allow use of the heart and both lungs from a single donor to different recipients. We have used this anastomosis in one patient without difficulty.