Objective:To apply lectin affinity chromatography and glycoproteomics-based LC-MS/MS to preliminarily investigate the possible potential plasma biomarkers of Opisthorchis viverrini(OV)-associated CCA in OV/dimethylnit...Objective:To apply lectin affinity chromatography and glycoproteomics-based LC-MS/MS to preliminarily investigate the possible potential plasma biomarkers of Opisthorchis viverrini(OV)-associated CCA in OV/dimethylnitrosamine(DMN)-induced CCA hamster model.Methods:Nine Syrian hamsters were divided into 3 groups as follows(n=3 each):normal(healthy control group);OV group;and OV/DMN group(CCA group).Pooled plasma samples collected from animals in each group at the 6th month post-infection with OV metacercarae were subjected to glycoproteomics analysis.Glycoproteins in the pooled sample from each group were initially isolated by concanavilin A(Con A)-based affinity chromatography.The expression of glycoproteins isolated by both enrichment methods were determined using LCMS/MS.Results:Among the 24 Con A-binding glycoproteins isolated,two proteins,N-myc downstream regulated gene 1(NDRG1)and fetuin-B(FETUB)were found up-regulated only in the samples from the OV and control groups,but not in the OV/DMN(CCA)groups.On the other hand,one protein,i.e.,NSFL1 cofactor p47 isoform x3(NSFL1C)was found only in the samples from OV/DMN(CCA)and control groups,but not in the OV group.The remaining 21 proteins were upregulated in the samples from all groups.Conclusions:NDRG1,FETUB and NSFL1 C glycoproteins isolated by Con A-based affinity chromatography could be potential biomarkers for CCA.Plasma samples with negative for NDRG1 and FETUB proteins but positive for NSFL1 C are likely to be OV-associated CCA.Nevertheless,this conclusion remains to be confirmed whether this battery test can discriminate OV-associated CCA from other risk factors.展开更多
Objective:To generate insights into the mechanism of NVP induced hepatotoxicity.Methods:Liver(HepG2)cells were cultured with various concentrations of NVP.This cell line was chosen because it has low expression of cyt...Objective:To generate insights into the mechanism of NVP induced hepatotoxicity.Methods:Liver(HepG2)cells were cultured with various concentrations of NVP.This cell line was chosen because it has low expression of cytochrome P450,allowing evaluation of the effects of NVP rather than specific metabolites.Cytotoxicity was determined using a proliferation assay and cell numbers were monitored using trypan blue exclusion assay for long term culture experiments and apoptosis induction was determined by morphological and biochemical investigation.Results:HepG2 cells treated with the highest concentration of NVP tested(819μM)initially showed a rounded morphology and all cells had died by week three of exposure.Nuclear condensation and fragmentation,increased Annexin V/propidium iodide staining and caspase 9 activation all supported the induction of apoptosis in HepG2 cells in response to NVP treatment.Conclusions:There is a clear induction of apoptosis in response to NVP which suggests that NVP has significant cytotoxicity,over and above any cytotoxicity of metabolites and may contribute directly to patient hepatotoxicity.展开更多
基金supported by the National Research University Project(NRU)of Thailand Office of Higher Education Commission,Ministry of Education of Thailand and Thammasat University(Excellence Center in Pharmacology and Molecular Biology of Malaria and Cholangiocarcinoma)
文摘Objective:To apply lectin affinity chromatography and glycoproteomics-based LC-MS/MS to preliminarily investigate the possible potential plasma biomarkers of Opisthorchis viverrini(OV)-associated CCA in OV/dimethylnitrosamine(DMN)-induced CCA hamster model.Methods:Nine Syrian hamsters were divided into 3 groups as follows(n=3 each):normal(healthy control group);OV group;and OV/DMN group(CCA group).Pooled plasma samples collected from animals in each group at the 6th month post-infection with OV metacercarae were subjected to glycoproteomics analysis.Glycoproteins in the pooled sample from each group were initially isolated by concanavilin A(Con A)-based affinity chromatography.The expression of glycoproteins isolated by both enrichment methods were determined using LCMS/MS.Results:Among the 24 Con A-binding glycoproteins isolated,two proteins,N-myc downstream regulated gene 1(NDRG1)and fetuin-B(FETUB)were found up-regulated only in the samples from the OV and control groups,but not in the OV/DMN(CCA)groups.On the other hand,one protein,i.e.,NSFL1 cofactor p47 isoform x3(NSFL1C)was found only in the samples from OV/DMN(CCA)and control groups,but not in the OV group.The remaining 21 proteins were upregulated in the samples from all groups.Conclusions:NDRG1,FETUB and NSFL1 C glycoproteins isolated by Con A-based affinity chromatography could be potential biomarkers for CCA.Plasma samples with negative for NDRG1 and FETUB proteins but positive for NSFL1 C are likely to be OV-associated CCA.Nevertheless,this conclusion remains to be confirmed whether this battery test can discriminate OV-associated CCA from other risk factors.
基金supported by The National Research Council of Thailand(NRCT)Office of the Higher Education Commission and Chiang Mai University under the National Research Universities Initiative
文摘Objective:To generate insights into the mechanism of NVP induced hepatotoxicity.Methods:Liver(HepG2)cells were cultured with various concentrations of NVP.This cell line was chosen because it has low expression of cytochrome P450,allowing evaluation of the effects of NVP rather than specific metabolites.Cytotoxicity was determined using a proliferation assay and cell numbers were monitored using trypan blue exclusion assay for long term culture experiments and apoptosis induction was determined by morphological and biochemical investigation.Results:HepG2 cells treated with the highest concentration of NVP tested(819μM)initially showed a rounded morphology and all cells had died by week three of exposure.Nuclear condensation and fragmentation,increased Annexin V/propidium iodide staining and caspase 9 activation all supported the induction of apoptosis in HepG2 cells in response to NVP treatment.Conclusions:There is a clear induction of apoptosis in response to NVP which suggests that NVP has significant cytotoxicity,over and above any cytotoxicity of metabolites and may contribute directly to patient hepatotoxicity.
