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Objective:To observe effect of Jingulian capsule on the proliferation of human breast cancer MDA-MB-231 cells and investigate its action mechanism against triple negative breast cancer （TNBC）. Method:The ingredients of Jingulian capsule were identified by ultra-performance liquid chromatography-tandem mass spectrometry（UPLC-MS/MS）. The inhibitory effect of Jingulian capsule at different doses （0.125，0.25，0.5，1，and 2 g·L<sup>-1</sup>） against the proliferation of MDA-MB-231 cells were detected by methyl thiazolyl tetrazolium （MTT） assay. After treatment for 24 h， the morphological changes in nuclear apoptosis of MDA-MB-231 cells were detected by Hoechst 33258 staining. The effect of different concentrations of Jingulian capsule on the apoptosis and cycle of MDA-MB-231 cells after different treatment time were determined by flow cytometry. The protein expression levels of Poly-ADP-ribose polymeras （PARP）， proto-oncogene c-Myc， cyclin B<sub>1</sub>， and phosphorylated extracellular signal-regulated kinase （p-ERK） in each group were assayed by Western blot. Result:A total of 113 compounds in Jingulian capsule were identified by UPLC-MS/MS. As revealed by MTT assay，compared with blank group，Jingulian capsule （0.125，0.25，0.5，1，2 g·L<sup>-1</sup>） significantly inhibited viability of MDA-MB-231 cells （<italic>P</italic><0.01）， with the half maximal inhibitory concentration （ IC<sub>50</sub>） of（0.13±0.02）g·L<sup>-1</sup>. According to flow cytometry，compared with the blank group，Jingulian capsule at 1 g·L<sup>-1</sup> significantly induced the apoptosis of MDA-MB-231 cells （<italic>P</italic><0.05）and Jingulian capsule at 0.5， 1 g·L<sup>-1</sup> obviously increased the number of MDA-MB-231 cells in S phase （<italic>P</italic><0.05，<italic>P</italic><0.01）. The results of Western blotting demonstrated that the protein expression levels of PARP，c-Myc，and cyclin B<sub>1</sub> in 0.5， 1 g·L<sup>-1 </sup>Jingulian capsule groups were remarkably down-regulated as compared with those in the blank group（<italic>P</italic><0.01）， and the protein expression level of p-ERK in 1 g·L<sup>-1 </sup>Jingulian capsule group was also down-regulated （<italic>P</italic><0.01）. Conclusion:Jingulian capsule is able to inhibit the proliferation of MDA-MB-231 cells，induce S phase cell cycle arrest， and promote their apoptosis， which may be related to the inactivation of the MAPK signaling pathway.
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Objective:To illustrate the effect and mechanism of ibrutinib,a Bruton's tyrosine kinase(BTK)inhibitor that inhibits diffuse large B-cell lymphoma(DLBCL)cell survival.Methods:DLBCL cell lines SUDHL-10 and HBL-1 were treated with ibrutinib at different concentrations.A MTT assay was used to detect the inhibition of cell proliferation.Cell apoptosis was analyzed by Annexin V-binding assay,as well as flow cytometry and DAPI staining.The expression of phosphorylated BTK,AKT and ERK was detected by Western blot. DLBCL cells were co-cultured with MSC.The inhibitory effect of ibrutinib on DLBCL cells in tumor microenvironment was assessed in clonogenicity in vitro and in a tumor-bearing non-obese diabetic/severe combined immunodeficient mice in vivo.Results:Up to 2.5 μmol/L and high concentrations of ibrutinib significantly inhibited the proliferation of DLBCL cells in a dose-dependent manner.Approx-imately 1 and 2.5 μmol/L ibrutinib was added on SUDHL-10 cells for 24 h,and the cell apoptotic rates were(21.73±3.64)% and(34.71± 2.36)%,respectively.Both were superior to that of the control group(3.55±1.89)%(P<0.05).Both two DLBCL cell lines pretreated with 5 and 10 μmol/L ibrutinib for 24 h and exhibited nuclear shrinkage at 5 μmol/L and nuclear fragmentation at 10 μmol/L.The expres-sion of phosphorylated BTK,AKT,and ERK decreased significantly after ibrutinib treatment.Ibrutinib inhibited clonogenicity in vitro(P<0.01)and cell proliferation and growth in vivo of DLBCL cells in co-culture system.The differences were statistically significant.Conclu-sion:Ibrutinib can inhibit the proliferation and induce apoptosis of SUDHL-10 and HBL-1 cell lines through a mechanism of blocking the AKT and ERK signaling pathways,as well as the proliferation of DLBCL cells in tumor microenvironment.This finding can significant-ly benefit DLBCL treatment.
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Objectives To investigate the clinical, pathological and endoscopic characteristics of colorectal polyps treated by endoscopic mucosal resection (EMR), then evaluate its therapeutic effect and security, and analyze its significance in diagnosis and treatment. Methods Clinical and pathological data of 437 patients (687 lesions), who underwent EMR from August, 2014 to August, 2015 were collected. A retrospective analysis was made, and lesions were statistically analyzed by variables such as age, sex, endoscopic features and clinical and pathological characteristics, and so on, according to the group procedure of adenoma cancerization and 'high risk adenomas - low risk adenomas - non-adenoma polyps' to observe the curative effect and safety on EMR. Results 48.28% of the patients who received colonoscopy examines were detected with polyps, and 34.90% of them were operated EMR, including advanced adenomas, which were 17.08% of the total quantity. From non-adenoma to advanced adenoma, with a increasing possibility of carcinogenesis, the location of polyps are transferred from proximal colon to distal colon and lesions are readily to be pedunculated ones with a higher level of mucosal lobulation and other kinds of mucosal changing. The degeneration of the colorectum is associated with pedunculated polyps, distal colorectal polyps, the size (>1.0 cm), adenomas containing villous structures and mucosal lobulation. As for EMR treatment, the en bloc resection rate can reach as high as 99.70%, with a complication rate of 1.14%. Conclusions Some special characteristics can be found in the site and the endoscopic feature of advanced adenomas and degenerated adenomas. To polyps sizing in 0.5 ~ 3.0 cm, EMR with endoclips has a remarkable effect and a low complication incidence. It can reduce the process of colorectal adenoma degenerates to multiple colorectal adenocarcinoma, which can be regarded as an effective method to prevent and cure the colorectal carcinoma.
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Objective:To illustrate the effect and mechanism of ibrutinib,a Bruton's tyrosine kinase(BTK)inhibitor that inhibits diffuse large B-cell lymphoma(DLBCL)cell survival.Methods:DLBCL cell lines SUDHL-10 and HBL-1 were treated with ibrutinib at different concentrations.A MTT assay was used to detect the inhibition of cell proliferation.Cell apoptosis was analyzed by Annexin V-binding assay,as well as flow cytometry and DAPI staining.The expression of phosphorylated BTK,AKT and ERK was detected by Western blot. DLBCL cells were co-cultured with MSC.The inhibitory effect of ibrutinib on DLBCL cells in tumor microenvironment was assessed in clonogenicity in vitro and in a tumor-bearing non-obese diabetic/severe combined immunodeficient mice in vivo.Results:Up to 2.5 μmol/L and high concentrations of ibrutinib significantly inhibited the proliferation of DLBCL cells in a dose-dependent manner.Approx-imately 1 and 2.5 μmol/L ibrutinib was added on SUDHL-10 cells for 24 h,and the cell apoptotic rates were(21.73±3.64)% and(34.71± 2.36)%,respectively.Both were superior to that of the control group(3.55±1.89)%(P<0.05).Both two DLBCL cell lines pretreated with 5 and 10 μmol/L ibrutinib for 24 h and exhibited nuclear shrinkage at 5 μmol/L and nuclear fragmentation at 10 μmol/L.The expres-sion of phosphorylated BTK,AKT,and ERK decreased significantly after ibrutinib treatment.Ibrutinib inhibited clonogenicity in vitro(P<0.01)and cell proliferation and growth in vivo of DLBCL cells in co-culture system.The differences were statistically significant.Conclu-sion:Ibrutinib can inhibit the proliferation and induce apoptosis of SUDHL-10 and HBL-1 cell lines through a mechanism of blocking the AKT and ERK signaling pathways,as well as the proliferation of DLBCL cells in tumor microenvironment.This finding can significant-ly benefit DLBCL treatment.
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Objective: To explore the diagnostic value of blood N-terminal pro-brain natriuretic peptide (NT-proBNP) and interleukin-17(IL- 17) for incomplete Kawasaki disease. Methods: Patients with Kawasaki disease, Incomplete Kawasaki disease and unclear infectious fever were included in this retrospective study. Their clinical features, and laboratory test results of blood NT-proBNP and IL-17 were collected and compared. Results: 766 patients with complete clinical information were recruited, consisting of 291 cases of Kawasaki disease, 74 cases of incomplete Kawasaki disease, and 401 cases of unclear infectious diseases. When the consistency with indicator 2 and 3 in Kawasaki disease diagnosis criteria was assessed with blood IL-17 ≥11.55 pg/mL and blood NT-proBNP ≥ 225.5 pg/dL as the criteria, the sensitivity and specificity for distinguishing incomplete Kawasaki disease and infectious diseases reached 86.5% and 94.8%, respectively. When we chose the consistency with indicator 1 and 2 in Kawasaki disease diagnosis criteria, the appearance of decrustation and/or the BCG erythema, blood IL-17 ≥11.55 pg/mL and blood NT-Pro BNP ≥225.5 pg/dL as the criteria, the sensitivity and specificity for distinguishing incomplete Kawasaki disease and infectious diseases was 43.2% and 100%, respectively. Conclusion: Blood NT-proBNP and IL-17 are useful laboratory indicators for distinguishing incomplete Kawasaki disease and infectious diseases at the early stage.
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In order to explore the potential influences of the disulfide bridge on the physical and chemical properties of PrP protein, the expressed recombinant human wild-type PrP protein was purified for using in an established redox process for the reduction and oxidation of the ethanethiol group within PrP. Sedimentation tests illustrated that redox process remarkably promoted the aggregation of recombinant PrP. Thioflavin T binding assay revealed an enhanced fibrillization of the recombinant human PrP after redox process. Far-UV circular dichroism demonstrated that the PrP treated with redox process showed a significant p-sheet rich structure. Furthermore, PrP-specific Western blot identified that the recombinant PrP after redox possessed stronger proteinase K-resistance. Those data indicates that the formation of the disulfide bridge induces the alteration of the secondary structure and enhances the progresses of aggregation and fibrillization of PrP protein.
Subject(s)
Humans , Amyloid , Chemistry , Endopeptidase K , Metabolism , Oxidation-Reduction , Prions , Chemistry , Metabolism , Protein Multimerization , Protein Structure, Secondary , Proteolysis , Sulfhydryl Compounds , ChemistryABSTRACT
<p><b>OBJECTIVE</b>To create transgenic mice expressing hamster- and human-PRNP as a model for understanding the physiological function and pathology of prion protein (PrP), as well as the mechanism of cross-species transmission of transmissible spongiform encephalopathies (TSEs).</p><p><b>METHODS</b>Hamster and human-PRNP transgenic mice were established by conventional methods. The copy number of integrated PRNP in various mouse lines was mapped by real-time PCR. PRNP mRNA and protein levels were determined by semi-quantitative RT-PCR, real-time RT-PCR, and western blot analysis. Histological analyses of transgenic mice were performed by hematoxylin and eosin (H & E) staining and immunohistochemical (IHC) methods.</p><p><b>RESULTS</b>Integrated PRNP copy number in various mouse lines was 53 (Tg-haPrP1), 18 (Tg-huPrP1), 3 (Tg-huPrP2), and 16 (Tg-huPrP5), respectively. Exogenous PrPs were expressed at both the transcriptional and translational level. Histological assays did not detect any abnormalities in brain or other organs.</p><p><b>CONCLUSION</b>We have established one hamster-PRNP transgenic mouse line and three human-PRNP transgenic mouse lines. These four transgenic mouse lines provide ideal models for additional research.</p>
Subject(s)
Animals , Cricetinae , Humans , Mice , Blotting, Western , DNA , Genetics , Disease Models, Animal , Immunohistochemistry , Mice, Inbred C57BL , Mice, Transgenic , Organ Specificity , Plasmids , Prion Diseases , Genetics , Prion Proteins , Prions , Genetics , Real-Time Polymerase Chain Reaction , Transcription, GeneticABSTRACT
The solid-phase method to synthesis HCV polypetides was used.The connecting rate and the polypeptides' productivity and purity were compared when Wang resin and 2-chlorotrity choloride resin acting as vectors.By the identification of mass spectrum,the products' molecular weight was analysed.By ninhydrin method we compared to the connecting rate between two resins and amino acid.By RP-HPLC we carried out the analysis and purity of hetero peptides.The result showed the first amino acid's connecting rate along with the purity and productivity which we need when 2-chlorotrityl choloride resin acted as vector was notably higher than those when Wang resin acted as vector.So 2-chlorotrityl choloride as vector is used to synthesize HCV polypeptides,which adapts to the demand of big scale's industrialization production.
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<p><b>OBJECTIVE</b>To explore the effect of serum from partial hepatectomy (PH) rat and hepatocyte growth factor (HGF) on expression of albumin and AFP of bone marrow cells.</p><p><b>METHODS</b>The bone marrow mono-nucleated cells were separated from SD rats and cultured in three groups: (1) The medium only group as control was added normal fetal bovine serum; (2) Rat hepatic injury serum group (was added 15% rat serum from 2-AAF+PH model); (3) HGF group (HGF 20 ng/ml). The role of these factors was determined by RT-PCR, immunohistochemistry (IHC) and Western blot, using AFP and albumin as special hepatocytic markers.</p><p><b>RESULTS</b>By immunohistochemical staining and Western blot, the fresh bone marrow cells were AFP-negative, same as the cells cultured with medium only group. While bone marrow cells, co-cultured with rat hepatic injury serum or HGF at day 10 and 20, expressed AFP protein. AFP mRNA expression could be found in bone marrow cells after 10 and 20 days cultured with rat hepatic injury serum or HGF, but not in fresh bone marrow cells and bone marrow cells cultured with medium only. Albumin mRNA expression was weak in fresh bone marrow cell and increased in groups 2 and 3.</p><p><b>CONCLUSION</b>The rat hepatic injury serum or HGF could stimulate the expression of AFP protein and it's mRNA of bone marrow cells. Also they can stimulate albumin mRNA expression. It seems that, in bone marrow, there is a kind of cells so called bone marrow derived liver stem cell which can express albumin mRNA in a weak style.</p>