Viruses (e.g. Human immunodeficiency virus, Human simplex virus and Prototype foamy virus) are obligate intracellular parasites and therefore depend on the cellular machinery for cellular trafficking. Bovine foamy virus (BFV) is a member of the Spumaretrovirinae subfamily of Retroviruses, however, details of its cellular trafficking remain unknown. In this study, we cloned the BFV gag gene into prokaryotic expression vector pET28a and purified the denaturalized Gag protein. The protein was used to immunize BALB/c mouse to produce antiserum, which could specifically recognize the BFV Gag protein in BFV-infected cells through western blot assay. Additionally, these results demonstrated that both the optimal and suboptimal cleavage of Gag protein occur in BFV-infected cells. Subsequently, the Gag antiserum was used to investigate subcellular localization of BFV. In immunofluorescence microscopy assays, colocalization microtubules (MTs) and assembling viral particles were clearly observed, which implied that BFV may transport along cellular MTs in host cells. Furthermore, MTs-depolymerizing assay indicated MTs were required for the efficient replication of BFV. In conclusion, our study suggests that BFV has evolved the mechanism to hijack the cellular cytoskeleton for its replication.

Objective To investigate the effect of Pingfei Oral Liquid (POL) on the distribution of mast cells (MCs) and the expression of interleukin 6 (IL-6) in the lung tissue of radiation pneumonia rats. MethodsForty-five SD rats were randomized into 3 groups : normal control,model group and POL group. The rat model of radiation lung fibro-sis was set up by a single X-ray dose of 20Gy irradiation over the whole chest of the rats. POL (20 g·kg-1·d-1,once a day, five times a week) was given orally one week before irradiation and the treatment lasted 5 weeks. MCs in the lung tissue were stained with toluidine blue firstly and then were counted 2, 4 and 8 weeks after irradiation. IL-6 protein expression of lung tissue was measured by immunohistochemical assay 8 weeks after irradiation, and mRNA ex-pression was determined with RT-PCR 4 weeks after irradiation. ResultsIt's showed the aggregation of large amount of pulmonary mast cells and increase of IL-6 protein expression 8 weeks after irradiation (P < 0.01).IL-6 mRNA expression in the irradiated lung of rats increased 4 weeks after irradiation (P < 0. 01). POL could reduce the aggrega- tion of MCs (P < 0. 01) and the expression of IL-6 protein (P < 0. 01) and mRNA (P < 0. 05) in the lung tissue. ConclusionPOL can prevent radiation pneumonia in rats by reducing the aggregation of mast cells and inhibiting IL-6 expression in the lung tissue.

Objective:To evaluate the effects of periodontitis on the expression of apoptosis-related proteins in pancreas of rats with Type 2 Diabetes Mellitus.Methods:Spontaneously type 2 diabetic OLETF rats were randomly divided into 2 groups:diabetes with or without periodontitis(diabetes group and combination group).LETO rats with the same germline and the same age but having normal glucose tolerance were randomly divided into control group and periodontitis group.20 weeks after periodontitis were established,all the rats were sacrificed and the pancreas were pathologically examined by HE staining.The expression of Bax,Bcl-2 and Caspase-3 in the pancreas islet were detected by immunohistochemistry staining and semi-quantitative analysis.Results:The expression of Bax, Bcl-2 and Caspase-3 in the pancreas islet was no significant difference between control and periodontitis groups(P=0.324,P=0.091,P=0.852).Compared with diabetes group,the expression of Bax and Caspase-3 in combination group showed a significant increase(P=0.000,P=0.000),and the expression of Bcl-2 was significantly decreased(P=0.022).Conclusion:Under healthy conditions,periodontitis has no effect on the expression of apoptosis-related proteins in rat pancreas islet.However,in rats with diabe-tes,periodontitis may affect the expression of apoptosis-related proteins in pancreas islet.

OBJECTIVETo assess the role of probiotics in the prevention of neonatal necrotizing enterocolitis (NEC) and to investigate the risk factors for NEC.

METHODSA total of 2528 hospitalized neonates between January 2002 and May 2005 were assigned into either receiving prophylactic use of probiotics bifoco (Prevention group, n=1182) or without probiotics supplementation (Control group, n = 1346). The incidence of NEC was compared between the two groups. The risk factors for NEC were investigated by conditional logistic regression multifactorial analysis.

RESULTSThere were 19 cases of NEC in the Control group (1.41%), but only 6 cases in the Prevention group (0.51%) (P < 0.05). Gestational age (OR = 5.521), hypoxicdouble ended arrowischemic encephalopathy (OR = 3.887), specticemia (OR = 4.854) and critical illness scores (OR = 5.989) were the risk factors for NEC, while the prophylactic use of probiotics was an independent protective factor for NEC (OR = 0.255).

CONCLUSIONSThe prophylactic use of probiotics may reduce the incidence of NEC in neonates.

Case-Control Studies ; Enterocolitis, Necrotizing ; prevention & control ; Female ; Humans ; Infant, Newborn ; Male ; Probiotics ; therapeutic use

Objective To study the effect of glutamate on metabolism, shifts in glycolysis and lactate release in rat astrocytes.
Methods After 10 days, secondary cultured astrocytes were treated with 1 mmol/L glutamate for 1 h, and the oxygen consumption rates (OCR) and extra cellular acidification rate (ECAR) was analyzed using a Seahorse XF 24 Extracellular Flux Analyzer. Cell viability was then evaluated by MTT assay. Moreover, changes in extracellular lactate concentration induced by glutamate were tested with a lactate detection kit.
Results Compared with the control group, treatment with 1 mmol/L glutamate decreased the astrocytes’ maximal respiration and spare respiratory capacity but increased their glycolytic capacity and glycolytic reserve. Further analysis found that 1-h treatment with different concentrations of glutamate (0.1-1 mmol/L) increased lactate release from astrocytes, however the cell viability was not affected by the glutamate treatment.
Conclusion The current study provided direct evidence that exogenous glutamate treatment impaired the mitochondrial respiration capacity of astrocytes and enhanced aerobic glycolysis, which could be involved in glutamate injury or protection mechanisms in response to neurological disorders.

Objective To explore the clinical significance of some risk factor,such as blood calcium serum alkaline phosphatase (ALP) and hemoglobin (Hb) for predicting or diagnosing bone metastases from urinary bladder neoplasms.Methods A total of 35 patients diagnosed as urinary bladder neoplasms and bone metastasis based on the histopathologic analysis (bone metastasis group,BM),and 105 cases of age-,sex-and follow-up time matched urinary bladder neoplasms patients without bone metastasis (non bone metastasis group,NBM) were included in present study.The correlations between clinical-pathological parameters (blood calcium,ALP,Hb) and bone metastases from urinary bladder neoplasms were analyzed and the risk factors for predicting or diagnosing bone metastases were identified.Results The levels of blood calcium,ALP and Hb were significantly higher at first diagnosis and follow-up period in BM group than in NBM group (P＜0.05).Multivariate logistic regression analysis indicated that the levels of calcium,ALP and Hb at first diagnosis were the independent risk factors for predicting the later incidence of bone metastases (P＜0.05).The levels of calcium,ALP and Hb in follow-up period were the independent risk factors for diagnosing bone metastases (P＜0.05).Receiver operating characteristic (ROC) curves analysis showed that the levels of calcium,ALP,Hb may give a more accurate prediction on bone metastases (AUC:0.615,0.704,0.692;cut-off:2.55mmol/L,97.0U/L,101.5g/L,respectively) or diagnosing bone metastases (AUC:0.706,0.776,0.829;cut-off:2.46mmol/L,112.5U/L,103.5g/L,respectively).Conclusion The levels of blood calcium,ALP and Hb may be used to predict the risk of bone metastases for patients firstly diagnosed as urinary bladder neoplasms,and predict whether the patients will be complicated with bone metastases during follow-up period.

Astrocytoma ; genetics ; metabolism ; Brain Neoplasms ; genetics ; metabolism ; Frizzled Receptors ; genetics ; metabolism ; Glioblastoma ; genetics ; metabolism ; Glioma ; genetics ; metabolism ; Humans ; Paraffin Embedding ; RNA, Messenger ; metabolism ; Receptors, G-Protein-Coupled ; genetics ; metabolism ; Signal Transduction ; Wnt2 Protein ; genetics ; metabolism ; beta Catenin ; genetics ; metabolism

OBJECTIVETo isolate single chain antibody fragments (scFv) against cell surface molecules by pathfinder selection from an anti-KG1a cell scFv phage library.

METHODSThe anti-KG1a scFv library was enriched by KGla cell panning for three rounds, or unenriched, then processed for pathfinder selection respectively using anti-CD34 monoclonal antibody as pathfinder molecule. ScFv phage clones were randomly picked and identified by binding KG1a cells using immunofluorescein and flow cytometry. The KG1a+ clones were further identified by KG1a, HL60, U937, and CEM cell lines and ELISA. Their antigenic molecules on cell surface were digested by chymopapain and analyzed by flow cytometry. DNAs from ten positive clones were sequenced. The scFv clones with different primary structure were used to analyze the molecular weight of their antigens by Western blot.

RESULTSOne hundred and two KG1a+ scFv phage clones were isolated from 144 enriched and 96 unenriched scFv phage library respectively, among which 47 bound KG1a, HL60, U937, and CEM cells, 55 bound KG1a cells exclusively. None of 28 KG1a+, HL60-, U937-, and CEM- scFv clones bound to the CD34 antigen, as confirmed by ELISA, although most of their antigens were sensitive to chymopapain digestion. DNA sequences from ten positive clones showed that they were from four different clones. They bound antigens with different molecular weight.

CONCLUSIONSOne hundred and two scFv phage clones specific for hematopoietic stem and progenitor cells have been isolated from an anti-KG1a cell scFv phage library. The pathfinder selection has showed advantages to improve the screening efficacy of scFv phage clones against antigens, which present at very low densities on the cell surface.

Antibodies, Anti-Idiotypic ; immunology ; Antibodies, Monoclonal ; genetics ; Antibody Specificity ; Bacteriophages ; genetics ; Cloning, Molecular ; Hematopoietic Stem Cells ; immunology ; Humans ; Immunoglobulin Fc Fragments ; biosynthesis ; genetics ; immunology ; Immunoglobulin Fragments ; genetics ; immunology ; Immunoglobulin Variable Region ; genetics ; immunology ; Peptide Library ; Single-Chain Antibodies

OBJECTIVETo assess the anti-nociception and anti-inflammation pharmacodynamics of Asarum heterotropoides var. mandshuricum and A. sieboldii.

METHODBoth the writhing test and hot plate test were conducted to assess the anti-nociceptive effect of Asarum and Xylene-induced mouse ear edema was conducted to assess the anti-inflammatory effect of Asarum.

RESULTTwelve samples of A. heterotropoides var. mandshuricum and A. sieboldii from different producing areas showed anti-nociceptive and anti-inflammatory effects. Specifically, 27% to 61% of the seven samples of A. heterotropoides var. mandshuricum showed anti-nociceptive effect and while 34% to 48% of A. sieboldi showed anti-nociceptive effect. The inflammatory inhibition rate of A. heterotropoides var. mandshuricum produced in six producing areas (38%-57%) is higher than that of A. heterotropoides var. mandshuricum produced in five producing areas (34%-48%). The same kind of Asarum produced in different areas showed significant differences. A. heterotropoides var. mandshuricum produced in Jilin province (38%-57%) showed better anti-nociceptive effect than sample produced in Heilongjiang province (34%) in writhing test. A. heterotropoides var. mandshuricum produced in Heilongjiang (43%) province showed a better anti-nociceptive effect than samples produced in Liaoning province (29%-36%) in hot plate test. A. sieboldii produced in Shaanxi province (47%-49%) showed a better anti-nociceptive effect than samples produced in Hubei province (40%) in writhing test. A. sieboldii produced in Shaanxi province (45%-59%) showed better anti-nociceptive effect than samples produced in Chongqing (40%) in hot plate test. A. heterotropoides var. mandshuricum produced in Jilin province (51%-63%) showed better anti-inflammatory effect than samples produced in Heilongjiang province (50%). In totality, the results from analysis of geoherbalism showed that famous-region A. heterotropoides var. mandshuricum and A. sieboldii had a better anti-nociception effect than Asarum produced in other producing areas, famous-region A. heterotropoides var. mandshuricum had a better effect than those produced in other producing areas in anti-inflammation. But famous-region A. sieboldii showed no obvious difference from those produced in other producing areas in anti-inflammation.

CONCLUSIONAll samples of Asarum showed anti-nociceptive and anti-inflammatory effects, but with significant differences among Asarum produced in different areas, indicating the eoherbalism to some extent.

Analgesics ; pharmacology ; Animals ; Anti-Inflammatory Agents ; pharmacology ; Asarum ; Female ; Male ; Mice ; Mice, Inbred ICR ; Plant Extracts ; pharmacology

Objective To investigate the changes of metabolic pathways in the process of immunesuppression in mice caused by cyclophosphamide by metabolomic analysis. Methods Flow cytometry was used to detect the changes of immune cells, and the results were analyzed using partial least squares model. The potential biomarkers were screened by variable importance in projection ( VIP) parameters in the partial least squares model, and the differential metabolites were determined by statistical analysis. The differential metabolites were identified using the metabolomics rapid identification and analysis software. Nine kinds of metabolites were identified by searching and calculation, and result of pathway enrich-ment showed three differential metabolic pathways. Results Cyclophosphamide had a significant effect on the biosynthesis of unsaturated fatty acids, metabolism of mitochondrial fatty acids, metabolism of glycophospholipid, biosynthesis of steroid hormones, metabolism of arachidonic acid, metabolism of fatty acids, and the biosynthesis of pyrimidine. Conclusions The most important metabolic pathways affected by cyclophosphamide in the normal body are the metabolism of arachidonic acid, glycophospholipid and pyrimidine.