Objective:To investigate the effect of phosphorylated HSP27 on the proliferation and metastasis of nasopharyngeal carcinoma and its molecular mechanism. Methods:①Western blot assay was used to detect the expression levels of HSP27 and p-HSP27 in CNE1 and CNE2 cells of nasopharyngeal carcinoma. Inhibited the phosphorylation of HSP27, Transwell assay detected the metastasis ability of nasopharyngeal carcinoma cells. ②Nasopharyngeal carcinoma cell lines（CNE2-OE1 and CNE2-OE2） overexpressing phosphorylated and dephosphorylated mutants of HSP27 were synthesized, empty vector transfected CNE2 cells（CNE2-NC） were used as controls. The proliferation ability of the three groups of cells was detected by CCK8, and the expression levels of CyclinD1, Bax and Bcl-2 were detected by Western blot. Transwell was used to detect the migration and invasion ability of cells, and Western blot was used to detect the expression levels of E-cadherin, Vimentin, MMP2 and MMP9. Results:The expression level of HSP27 in CNE2 was higher than that of CNE1 cells, while the expression level of p-HSP27 was opposite. After inhibition of HSP27 phosphorylation, the invasion and migration ability of CNE1 cells decreased significantly, with no significant change in CNE2 cells. Compared with CNE2-NC, the growth rate of CNE2-OE1 decreased, and the growth rate of CNE2-OE2 increased. The expression level of CyclinD1 was down-regulated in CNE2-OE1 and higher in CNE2-OE2. The expression level of Bax in CNE2-OE1 was increased, while the expression of Bcl-2 was decreased. There was no significant change in the expression of Bax and Bcl-2 in CNE2-OE2. Compared with CNE2-NC, the migration ability of CNE2-OE1 was enhanced and the invasion ability was weakened, while the migration ability of CNE2-OE2 was weakened and the invasion ability was enhanced. There was no significant difference in the expression levels of E-cadherin was decreased in CNE2-OE1 and increased in CNE2-OE2. There was no significant difference in the expression levels of Vimentin among the three groups. The expression levels of MMP2 and MMP9 were up-regulated in CNE2-OE2, and slightly down-regulated in CNE2-OE1. Conclusion:HSP27 and p-HSP27 were differentially expressed in different nasopharyngeal carcinoma cells, and the metastasis ability of nasopharyngeal carcinoma was not only related to the expression level of HSP27, but also related to the level of p-HSP27. The p-HSP27 inhibited CNE 2 cell proliferation and promoted their apoptosis. As p-HSP27 plays different roles in different stages of nasopharyngeal carcinoma metastasis, it can increase the migration ability of CNE2 cells and reduce its invasion ability. p-HSP27 may induce EMT changes in CNE2 by down-regulating E-cadherin, thus promoting CNE2 migration, and may inhibit CNE2 invasion by down-regulating MMPs expression.
Humans ; Cell Proliferation ; Cell Line, Tumor ; Nasopharyngeal Carcinoma/pathology* ; Nasopharyngeal Neoplasms/pathology* ; Cell Movement ; HSP27 Heat-Shock Proteins/metabolism* ; Phosphorylation ; Neoplasm Metastasis ; Matrix Metalloproteinase 9/metabolism* ; Neoplasm Invasiveness ; Matrix Metalloproteinase 2/metabolism* ; Heat-Shock Proteins/metabolism* ; Cyclin D1/metabolism* ; Molecular Chaperones/metabolism* ; Cadherins/metabolism*

ObjectiveConstruction of a negative control line for the Drosophila transgenic system based on ΦC31 integrase and vector plasmid pUASTattB to provide a more scientific negative control for transgenic Drosophila research experiments. MethodsThe vector plasmid pUASTattB was microinjected into four different genetic backgrounds Drosophila lines attP-25C6, attP-68A4, attP-75B1 and attP-86F8 embryos carrying ΦC31 integrase. All of the injected embryos were incubuated to get G0 adults, and each of them was crossed with balancer stock ywR13S separately in a single vial (1 adult of the G0 generation and 3 of the ywR13S in each vial). The probability of successful insertion was calculated by observing the colour of the compound eyes of the G1 generation of Drosophila to determine whether there was a mini-White insertion. The G1 generation Drosophila adults successfully inserted into mini-White were then selected to make single-vial crosses (one G1 generation male Drosophila crossed with three virgins of balancer Drosophila line) with each of the three balancer Drosophila strains DB, ywR13S and yw122, respectively, for balanced seed preservation. The genomic DNA of the conserved Drosophila lines was extracted and the vector plasmid pUASTattB was identified for transfer by PCR. Results12 Drosophila strains were obtained, all of which were red-eyedDrosophila melanogaster carrying the mini-White marker, and were identified by PCR as having the pUASTattB sequence insertion. ConclusionThe 12 transgenic Drosophila strains can meet the negative control requirements for the transgenic fly research experiments that constructed with pUASTattB as the vector basically, enriching the Drosophila resources in the National Drosophila Resource Center of China.

Immunotherapy has achieved objective response rates of 20%-30% in patients with recurrent or metastatic nasopharyngeal carcinoma, but fewer people are benefiting. Studies have shown that patients with nasopharyngeal carcinoma carrying high expression of programmed death-1/programmed death-ligand 1 and/or high tumor mutation burden have a significant response to immunotherapy. Biomarkers of the tumor microenvironment, especially tumor infiltrating lymphocyte, are abundant in nasopharyngeal carcinoma, varying from different Epstein-Barr virus states, which can also play a predictive role of immunotherapy efficacy. Other biomarkers, such as mismatch repair-deficient, have a low incidence in nasopharyngeal carcinoma and limited predictive power. Combined detection of different types of immunotherapeutic biomarkers is more helpful to identify suitable populations for immunotherapy.

Objective To study the protective effects of curcumin on oxidative stress injury of H9C2 cardiac myocytes induced by H2O2.Methods The oxidative stress injury model in H9C2 cardiac myocytes was established in vitro, and cells were divided into the control group, H2O2 group (200μmol/L), low- (10μmol/L), medium- (20μmol/L), high- (40μmol/L) dose curcumin, and each group set six holes. After H2O2 stimulation for 24 h, the morphology changes were observed by microscope, and the survival rates were measured using CCK8 method. The activity of LDH, CK, and AST, and the content of MDA in culture medium were detected, and the activities of SOD, CAT, and GSH-Px in cardiac myocytes were also determined.Results Compared with the H2O2 group, the activity of LDH (247.36 ± 28.04 U/L, 195.14 ± 21.37 U/L, 132.27 ± 16.33 U/Lvs. 247.58± 28.34 U/L), AST (67.27 ± 10.58 U/ml, 54.81 ± 8.60 U/ml, 42.14 ± 5.95 U/mlvs.84.34 ± 12.36 U/ml), CK (1.34 ± 0.66 U/ml, 0.95 ± 0.42 U/ml, 0.71 ± 0.39 U/ml vs.1.56 ± 0.74 U/ml) and the content of MDA (227.39 ± 32.05 nmol/ml, 185.11 ± 24.37 nmol/ml, 143.50 ± 16.37 nmol/mlvs. 274.19 ± 36.51 nmol/ml) in culture medium of curcumin were significantly decreased (P<0.05). The morphology of H9C2 cardiac myocytes in curcumin group was improved, the survival rate was significantly increased (P<0.05). The activity of SOD (17.67± 1.36 U/mg, 21.54 ± 1.72 U/mg, 28.37 ± 2.36 U/mgvs. 14.37 ± 1.22 U/mg), CAT (19.58 ± 3.87 U/mg, 25.34 ± 4.06 U/mg, 30.21 ± 4.83 U/mgvs. 14.77 ± 3.25 U/mg) and GSH-Px (1.99 ± 1.17 U/mg, 2.56 ± 1.29 U/mg, 3.04 ± 0.45 U/mgvs. 1.67 ± 0.87 U/mg) in cardiac myocytes were significantly increased (P<0.05). Conclusions Curcumin can effectively improve oxidative stress injury of H9C2 cardiac myocytes induced by H2O2, and curcumin has dose-dependent protective effects against the oxidative stress of H9C2 cardiac myocytes induced by H2O2.

Objective To investigate the clinical pathological characteristics, diagnosis and treatment of breast rhabdomyosarcoma, and to enhance the awareness of malignancy infiltration to bone marrow (BM). Methods The data of one case of Rhabdomyosarcoma of breast were analyzed retrospectively. BM aspirate and biopsy, morphology, immunology, cytogenetics, molecular biology (MICM) in different parts of BM, peripheral blood smear, fine puncture of breast mass, final biopsy of breast mass by Mammotome System and whole body PET-CT were performed. The immunochemistry stain of specimen of breast mass was used. Results The peripheral blood smear of this patient showed immature erythrocytes, leucocytes and classification of unknown cells which were consistent with BM morphology. The results of BM aspirate and biopsy depicted a hypercellular specimen with disseminated unknown cells infiltration. Unknown cells were positive for CD56 and negative for any hematopoietic markers by flow cytometry. The whole body PET-CT showed that uptake of 18F-FDG of bilateral breast and whole BM was increased, whereas the mass of breast was not presented by CT. PET-CT suggested a probable malignant hematologic disease. The enough specimen of breast mass got from Mammotome System showed embryonal rhabdomyosarcoma, and the tumor cells were positive for MyoD1, Vimentin and Desmin. Conclusions It is a challenge for early diagnosis of solid sarcoma with unknown origin which diffusely infiltrating into BM. Negative expression of hematopoietic markers by flow cytometry plays a role on differential diagnosis in this setting, whereas PET-CT only provides a valuable reference. Enough specimen and immunohistochemical staining could provide solid evidences of diagnosis.

Objective To explore the clinical features and causes of misdiagnosis of the patients with acquired deficiency of vitamin K-dependent coagulation factors (ADVKDCF). Methods Retrospective analysis was performed with the data from 62 patients with ADVKDCF for etiological factors, clinical manifestations,laboratory examinations, diagnosis and treatments. Results Among the 62 patients, 51 patients were with unknown causes( subgroup A) and 11 were with clear histories of anticoagulant rodenticide poisoning( subgroup B). The presentations of hemorrhage of the patients varied with hematuria as the most common first symptom,followed by skin, mucosa, muscle, internal organs bleeding (28/62). The most common hemorrhage symptom is hematuria. 35 of the 62 patients had hemoglobin(Hb) levels less than 100 g/L due to blood loss( the lowest level was 32 g/L). Thirty-eight patients were misdiagnosed at the first visit and the median time from hemorrhage manifestation to definite diagnosis was 8 days (range,2 to 192 days). ADVKDCF was mostly misdiagnosed as the urinary system diseases (23/38), followed by hemophilia (8/38). Laboratory examinations showed normal platelet count , throm bin time (TT) and normal fibrinogen(Fg) concentration, but prolonged plasma prothrombin time (PT), activated partial prothrombin time (APTT) and international normalized ration (INR). All of patients received high dose vitamin K ( intravenous vitamin K1 with a initial dose of 20 to 240 mg/d and then oral vitamin K4 maintenance) . The bleeding symptoms disappeared 1 day after treatment and the Hb levels increased dramatically. There were significant differences in PT, APTT and INR of the patients before and after treatment( P ＜0. 01 ). Followed by a median follow - up of 8 months , no patient had severe adverse effects or recurrence. Conclusion The hemorrhage presentations of the patients with ADVKDCF are various. The most common hemorrhage symptom is hematuria. The misdiagnosis rate of ADVKDCF is high with urinary systems disorders as the most common misdiagnosis. Sequential treatment with vitamin K is an effective and safe method to prevent recurrence. Early detection of coagulation function is helpful to reduce misdiagnosis possibility.