Objective:To investigate the inducing effects of sunitinib malate on expression of NKG2D ligands in nasopharyngeal carcinoma cell ABCG2high CNE2/DDP.Methods:ABCG2highCNE2/DDP cells and Allo-NK cells were isolated by magnetic activated cell sorting(MACS).Flow cytometry was used to evaluate the purity of isolated cells and the expression of NKG2D-ligands on target cells before and after incubation with sunitinib malate.Then the cytotoxic sensitivity of treated and un-treated ABCG2high CNE2/DDP cells to Allo-NK cells were measured by LDH releasing assay.Results:The positive rate of ABCG2 in ABCG2highCNE2/DDP cells was(91.40?2.32)%.More than 90% of isolated Allo-NK cells were proven to be CD3-CD16+CD56+ cells.The expression of MICA,MICB,ULBP1,ULBP2 and ULBP3 on ABCG2high CNE2/DDP cells incubated with sunitinib malate increased from(2.92?0.33)%,(4.27?0.33)%,(5.80?0.62)%,(11.10?3.15)%,and(7.75?1.14)% to(89.12?4.56)%,(66.10?2.22)%,(67.56?4.19)%,(69.37?8.83)%,and(63.28?3.31)%,respectively.At the E ∶T ratios of 10 ∶1 and 20 ∶1,the cytotoxic sensitivities of ABCG2high CNE2/DDP cells to Allo-NK cells increased from(15.32?13.86)% and(27.26?6.81)% to(41.12?4.12)% and(57.25?2.37)%,respectively,after treatment with sunitinib malate,with significantly difference found in the cytotoxic sensitivities of target cells in each group before and after sunitinib malate treatment(F=15.58,P=0.000).Conclusion:Sunitinib malate can up-regulate expression of NKG2D-ligands(MICA/B,ULBP1-3)in ABCG2high nasopharyngeal carcinoma cells,which results in higher cytotoxic sensitivity to Allo-NK cells.

Objective To study the effects of early mechanical strain magnitude on formation and differentiation of osteoclasts. Methods RAW 264.7 cells induced by macrophage colony-stimulating factors and osteoclast differentiation factors were subjected to 0, 1 000, 1 500, 2 000, 2 500 and 5 000 με mechanical straining for three days. The morphological changes, number of osteoclasts and proliferation of precursor cells were determined at day 7. The activity of the tartrate-reaistant acid phosphatase (TRAP) in the culture medium was detected at days 4 and 7. Results The number of osteoclasts was decreased in 2 500 με group, while it was increased in 5 000 με group. The proliferation of precursor cells was increased in 2 000 and 2 500 με group, while it was decreased significantly in 5 000 με group. There was no significant difference in the number of osteoclasts and proliferation of precursor cells among 1 000 με group, 1 500 με group and 0 με group. The activity of TRAP was decreased in 1 000, 1 500, 2 000, 2500 and 5 000 με groups at days 4 and 7 when compared with με group. Conclusions Early mechanical straining plays a direct role in formation and differentiation of osteoclasts. The high strain magnitude within physiological load inhibits osteoclast formation, while high strain magnitude beyond physiological load stimulates osteoclast formation. Low strain magnitude has nearly no impact on formation of osteoclasts. Early mechanical straining may inhibit differentiation of osteoclasts.

Dengue virus (DENV) envelope [E] protein is the major surface protein of the virions that indued neutralizing antibodies. The domain III of envelope protein (EDIII) is an immunogenic region that holds potential for the development of vaccines; however, the epitopes of DENV EDIII, especially neutralizing B-cell linear epitopes, have not been comprehensively mapped. We mapped neutralizing B-cell linear epitopes on DENV-1 EDIII using 27 monoclonal antibodies against DENV-1 EDIII proteins from mice immunized with the DENV-1 EDIII. Epitope recognition analysis was performed using two set of sequential overlapping peptides (16m and 12m) that spanned the entire EDIII protein from DENV-1, respectively. This strategy identified a DENV-1 type- specific and a group-specific neutralizing epitope, which were highly conserved among isolates of DENV-1 and the four DENV serotypes and located at two regions from DENV-1 E, namely amino acid residues 309-320 and 381-392(aa 309-320 and 381-392), respectively. aa310 -319(310KEVAETQHGT319)was similar among the four DENV serotypes and contact residues on aa 309 -320 from E protein were defined and found that substitution of residues E309 , V312, A313 and V320 in DENV-2, -3, -4 isolates were antigenically silent. We also identified a DENV-1 type-specific strain-restricted neutralizing epitope, which was located at the region from DENV-1 E, namely amino acid residues 329-348 . These novel type- and group-specific B-cell epitopes of DENV EDIII may aid help us elucidate the dengue pathogenesis and accelerate vaccine design.
Amino Acid Sequence ; Animals ; Antibodies, Neutralizing ; immunology ; Dengue ; virology ; Dengue Virus ; chemistry ; genetics ; immunology ; Epitope Mapping ; Epitopes, B-Lymphocyte ; chemistry ; genetics ; immunology ; Humans ; Mice ; Molecular Sequence Data ; Viral Envelope Proteins ; chemistry ; genetics ; immunology

Objective To investigate the mechanisms and effects of Sorafenib on cytotoxic sensitivity of allo-reactive natural killer(Allo-NK) cells against human multi-drug resistant nasopharyngeal carcinoma CNE2/DDP cells which expressing highly ATP-binding cassette superfamily G member 2(ABCG2)(abbr.as ABCG2HighCNE2/DDP cells).Methods ABCG2HighCNE2/DDP and Allo-NK cells were isolated by magnetic bead technique.The target cells were divided into 3 groups: a) treated group(ABCG2HighCNE2/DDP cells incubated with 10 ng/ml sorafenib for 4h);b) untreated group(conventionally cultured ABCG2HighCNE2/DDP cells);and c) control group(conventionally cultured K562 cells).Expression rates of ABCG2 in treated and untreated groups,and of five NKG2D-ligands(MICA,MICB,ULBP1,ULBP2,ULBP3) were evaluated by flow cytometry.The cytotoxic effects of NK cells against different groups of target cells were detected with LDH releasing assay.Results Expression rate of ABCG2 in isolated CNE2/DDP cells was 91.40%?2.32%.The purity of sorted CD3-CD16+CD56+ Allo-NK cells was 90% and higher.The expression rates of NKG2D-ligands(MICA,MICB,ULBP1,ULBP2 and ULBP3) in untreated group were 2.92%?0.33%,4.27%?0.33%,5.80%?0.62%,11.10%?3.15% and 7.75%?1.14%,respectively,which were remarkablely higher than that in treated group(10.38%?1.23%,10.68%?1.26%,11.62%?1.22%,43.24%?4.42% and 11.91%?0.88%,respectively,P

Objective:To explore the effects of G protein-coupled receptor 55 (GPR55) antagonist CID16020046 on renal fibrosis in mice, and provide a new method and idea for the treatment of renal fibrosis.Methods:(1) GPR55 overexpression and GPR55 antagonist CID16020046 were used in renal fibroblasts (NRK-49F) of rats, respectively. Meanwhile,transforming growth factor-β1 (TGF-β1) was applied in the NRK-49F cells to observe the expression of fibrosis-related factors and inflammatory factors. (2) A mouse model of renal fibrosis with unilateral ureteral obstruction (UUO) was established in vivo. Eight-week-old male C57BL/6J mice (20-25 g) were randomly divided into three groups according to the random number table method: sham group ( n=6), model group (UUO group, n=7), model + CID16020046 drug (UUO+CID group, n=8). The drug CID16020046 (10 mg/kg) was intraperitoneally injected 1 day before modeling, on the day of modeling and every day after surgery in UUO+CID group, and the corresponding dose of 0.9% normal saline was injected intraperitoneally in sham and UUO groups.The mice were sacrificed for sampling 7 days after UUO surgery, and their renal function indicators, liver transaminase, and cardiac markers were examined. Western blotting and quantitative real-time PCR were used to detect the expression of renal fibrosis-related factors and inflammatory factors. Immunohistochemistry staining, Sirius red staining and Masson trichrome staining were used to detect the pathological changes of renal tissues. Results:(1) After NRK-49F cells were stimulated by TGF-β1, the mRNA and protein expression levels of GPR55 were significantly increased (both P<0.05). There was no statistically significant difference in the mRNA expression of fibrosis-related factors fibronectin and collagen Ⅰ, and inflammatory factors interleukin-1β and tumor necrosis factor-α between TGF-β1 group and TGF-β1 + GPR55 overexpression group (all P>0.05). Compared with the TGF-β1 group, the protein expression levels of fibrosis-related factors alpha-smooth muscle actin (α-SMA) and vimentin, and the mRNA expression levels of collagen Ⅰ and α-SMA were lower in the TGF-β1 + CID group (all P<0.05). (2) Compared with sham group, the mRNA and protein expression levels of GPR55 in UUO group were higher (both P<0.05). The serum creatinine in the UUO+CID group was lower compared to the UUO group ( P<0.05). There was no statistically significant difference in blood urea nitrogen, alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase and creatine kinase isoenzyme between UUO group and UUO+CID group (all P>0.05). Compared with the UUO group, the protein expression levels of renal fibrosis-related factors fibronectin, collagen Ⅰ and vimentin, and the mRNA expression levels of fibronectin, collagen Ⅰ, collagen Ⅲ and α-SMA were lower in the UUO+CID group (all P<0.05). The degree of renal tubular dilation and interstitial collagen fiber deposition in the UUO+CID group was significantly reduced compared to the UUO group (all P<0.05). Conclusions:CID16020046 can reduce serum creatinine in UUO mice, protect renal function, and simultaneously decrease the expression of fibrosis-related factors in renal fibroblasts and mouse kidney tissues, thereby alleviating renal fibrosis.