Effects of P-Glycoprotein and Its Inhibitors on Apoptosis in K562 Cells

P-glycoprotein (P-gp) is a major factor in multidrug resistance (MDR) which is a serious obstacle in chemotherapy. P-gp has also been implicated in causing apoptosis of tumor cells, which was shown to be another important mechanism of MDR recently. To study the influence of P-gp in tumor cell apoptosis, K562/A cells (P-gp+) and K562/S cells (P-gp−) were subjected to doxorubicin (Dox), serum withdrawal, or independent co-incubation with multiple P-gp inhibitors, including valspodar (PSC833), verapamil (Ver) and H108 to induce apoptosis. Apoptosis was simultaneously detected by apoptotic rate, cell cycle by flow cytometry and cysteine aspartic acid-specific protease 3 (caspase 3) activity by immunoassay. Cytotoxicity and apoptosis induced by PSC833 were evaluated through an MTT method and apoptosis rate, and cell cycle combined with caspase 3 activity, respectively. The results show that K562/A cells are more resistant to apoptosis and cell cycle arrest than K562/S cells after treatment with Dox or serum deprivation. The apoptosis of K562/A cells increased after co-incubation with each of the inhibitors of P-gp. P-gp inhibitors also enhanced cell cycle arrest in K562/A cell. PSC833 most strikingly decreased viability and led to apoptosis and S phase arrest of cell cycle in K562/A cells. Our study demonstrates that P-gp inhibits the apoptosis of tumor cells in addition to participating in the efflux of intracellular chemotherapy drugs. The results of the caspase 3 activity assay also suggest that the role of P-gp in apoptosis avoidance is caspase-related

In Vivo Quantitation of Collagen/Gelatin Fragments Using HPLC-MS/MS Detection of Peptide Markers

Collagen/gelatin fragments are often polydisperse and heterogeneous, making them difficult for quantitation, especially during in vivo circulation. In this study, a new quantitation method was established based on the detection of collagen-specific peptide markers using HPLC-MS/MS. Blood samples were collected and digested for collagen-specific peptides identification and content determination after the infusion of bovine gelatin fragments into rabbits. Thirteen collagen-specific peptides, of which five were present in bovine collagen but absent in human collagen, were identified by MS detection and could be used as peptide markers for the quantitation of gelatin fragments in blood. One of the bovine collagen specific-peptides was then used as a peptide marker, and its detection linearity range and limit of quantification were confirmed. The pharmacokinetic parameters, such as half-life and elimination rate, and also the pharmacokinetic model of the infused gelatin fragments were determined from the time-concentration curve. This peptide marker based method offers better selectivity towards collagen/gelatin fragments quantitation and could find wider applications in the field of collagen/gelatin based products such as drug delivery materials, and nanoparticles.</p

Characteristics and sources of black carbon aerosol in a mega-city in the western Yangtze River Delta, China

International audienceA single particle soot photometer (SP2) was deployed in urban Nanjing, located in the Yangtze River Delta (China), to investigate the mixing state and sources of ambient refractory black carbon (rBC) from 26 January to 25 February 2014, along with an in-situ measurement of submicron aerosol chemical species by an aerodyne aerosol chemical speciation monitor (ACSM). The results showed that anthropogenic activities associated with firework emissions can be a significant source for rBC-containing particles during the period of the Chinese New Year, resulting from the evident peaks of rBC at midnight. During the residual periods, namely regular day (RD), the diurnal cycles of rBC presented two typical peaks that can be attributed to a synergistic influence of local traffic emissions and boundary layer changes throughout a day. Three coating factors, including organics, sulfate, and nitrate (-rich), were resolved using a positive matrix factorization (PMF) approach to explain the potential contribution of non-rBC coatings (i.e., organics, sulfate, and nitrate) to the coating thickness of rBC-containing particles. As the results show, organic aerosols (OAs) might be a major contributor to the coating thickness of rBC-coating particles during the whole period. The relative coating thickness (a ratio between coated particle size to BC core) exhibited a positive relationship with sulfate, indicative of the favorable coating factor during the episode caused by firework emissions. Source apportionment of rBC was performed via a multiple linear regression between the total rBC mass and each ACSM-PMF factor (rBC-ACSM-PMF). On average, biomass burning emissions accounted for 43%, being the largest contributor during the RD period, whereas local traffic emissions played a major role during the new year time

Seasonal size distribution and mixing state of black carbon aerosols in a polluted urban environment of the Yangtze River Delta region, China

The optical properties of black carbon aerosols (BC) are determined by the particles size and the associated non-BC materials, which may be source-related or modified during secondary processing. The one-year long monitoring of BC was first conducted using a Single Particle Soot Photometer (SP2) from December 2013 to November 2014 in Nanjing, a megacity in the Yangtze River Delta region of China. The seasonal variation in the BC size distribution and mixing state were investigated. There was no apparent systematic variation in the mean BC core mass median diameter between seasons, as these values were 226 +/- 12 nm, 217 +/- 13 nm, 211 +/- 15 nm and 221 +/- 12 nm for winter, spring, summer and autumn respectively. The mixing state of BC was quantified as the bulk relative coating thickness (defined as particle size D(p )over core size D-c, D-p/D-c), which ranged from 1.05 to 2.65. The BC was found to be significantly more coated in the winter (D-p/D-c = 1.50 +/- 0.30) than in other seasons (D-p/D-c = 1.27 +/- 0.09, 1.28 +/- 0.10, 1.27 +/- 0.11 in spring, summer and autumn respectively). Higher levels of coating during the winter may due to the contributions of the primary source (with the highest BC mass loadings between seasons) or secondary processes such as low temperature that facilitated the condensation. It was found that the photochemical process may enhance the coatings on BC in summer. At nighttime, the reduced and stabilized planetary boundary layer and the nighttime secondary formation may also lead to BC becoming well mixed with other components. Moreover. BC was shown to be less coated when the NOx concentration was high. However, during all seasons, the BC coating was strongly correlated with other non-BC particulate mass, which suggests that at higher pollution levels BC was more significantly coated with other existing materials through coagulation or condensation by other secondary species

Suppression of DLBCL Progression by the E3 Ligase Trim35 Is Mediated by CLOCK Degradation and NK Cell Infiltration

The majority of diffuse large B-cell lymphoma (DLBCL) patients develop relapsed or refractory disease after standard ruxolitinib, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) chemotherapy, which is partly related to a dysregulated tumor immune microenvironment. However, how the infiltration of immune cells is appropriately regulated is poorly understood. Herein, we show that the E3 ubiquitin ligase Trim35 is expressed at low levels in human DLBCL tissues. We also show that overexpression of Trim35 suppresses DLBCL cell proliferation and correlates with inferior survival in DLBCL patients. Our mechanistic study shows that Trim35 functions as an E3 ligase to mediate the ubiquitination and degradation of CLOCK, a key regulator of circadian rhythmicity. High expression of Trim35 correlates with NK cell infiltration in DLBCL, partly due to the degradation of CLOCK. Consistently, patients with high expression of CLOCK show poor overall survival. Overall, these findings suggest that Trim35 suppresses the progression of DLBCL by modulating the tumor immune microenvironment, indicating that it may be a promising diagnostic and prognostic biomarker in DLBCL