[LncRNA HULC promots HCC growth by downregulating miR-29].

Objective: To explore the effects of lncRNA HULC on hepatocellular carcinoma (HCC) growth by down-regulating miR-29. Methods: The expression levels of HULC and miR-29 in HCC tissues and cells were detected by real-time quantitative PCR (RT-qPCR), and the correlation analysis was performed. After HCC cells were transfected with HULC overexpressed plasmid or siRNA, the expressions of miR-29 and its target gene SETDB1 were determinate by RT-qPCR. According to the bioinformatic prediction of the miR-29 binding site in the HULC sequence, the report gene plasmids were constructed. The HCC cells were co-transfected with miR-29 mimics or miR-29 inhibitor, and the HULC targeted regulation of miR-29 was verified by dual luciferase reporter assay. The effect of miR-29 on the HULC-mediated proliferation in HCC cells was detected by cell count kit 8 (CCK-8) experiment. Expression of tumor proliferation antigen Ki-67 was detected by RT-qPCR.The Hep3B cells were inoculated in mice and miR-29 mimics and miR-29 negative control (NC) further injected into the lesions. The tumor volume was observed, and the expressions of tumor proliferation antigen ki-67 in tumor tissues were detected by immunohistochemical staining. Results: The expression of HULC was significantly up-regulated while the expression of miR-29 was significantly down-regulated in HCC tissues and cells (P<0.01). The level of HULC was negatively correlated with miR-29 in tumor tissues (r=-0.754, P<0.01) and HCC cells (r=-0.865, P<0.05). The in vitro experiments showed that, compared with the blank control group, the expression of miR-29 in HULC overexpressed Huh7 cells was significantly reduced, while the mRNA level of miR-29 target gene SETDB1 was increased (P<0.01). The expression of miR-29 was significantly increased in HULC deleted Hep3B cells, while the mRNA expression of SETDB1 was decreased (P<0.01). Double luciferase reporter gene assay showed that miR-29 mimics significantly inhibited the luciferase activity of Hep3B cells transfected with HULC wide type (psi-HULC-WT) plasmid but had no effect on Hep3B cells transfected with mutant plasmid (psi-HULC-Mut). However, the miR-29 inhibitor antagonized the inhibitory effect of miR-29 mimics on luciferase activity of psi-HULC-WT (P<0.01). Cell proliferation experiments showed that, compared with the control group, the proliferation ability of miR-29 mimics overexpressed Huh7 cells was significantly reduced.After 24, 48 and 72 hours of treatment, the proliferation rates of Huh7 cells in the HULC overexpressed group were (43.87±3.82)%, (83.45±7.46)% and (123.34±8.67)%, respectively, significantly higher than (13.45±1.77)%, (23.54±1.37)% and (38.21±2.09)% of control group (P<0.05). After treatment for 48 and 72 hours, the proliferation rates of miR-29 mimics transfected Huh7 cells were (57.10±1.94)% and (73.76±3.46)%, respectively, significantly lower than (83.45±7.46)% and (123.34±8.67)% of control group (P<0.05). After treatment for 48 and 72 hours, the proliferation rates of Huh7 cells transfected with miR-29 mimics and miR-29 inhibitor group were (76.45±3.24)% and (89.37±4.37)%, respectively, significant higher than (57.10%±1.94)% and (73.76±3.46)% of the control group (P<0.05). After 48 h transfection, the expression of Ki-67 in Huh7 transfected with miR-29 mimics was significantly inhibited compared with the control group (P<0.01). However, the expression of Ki-67 mRNA was increased in Huh7 cells transfected with miR-29 inhibitor (P<0.01). The results of in vivo experiments showed that the tumor volumes of the control group, miR-29 mimics group and miR-29 mimics + miR-29 inhibitors group were (504.0±19.6) mm(3), (310.0±24.3) mm(3) and (483.7±21.2) mm(3), respectively. Injection of miR-29 mimics reduced while miR-29 inhibitor promoted tumorigenesis ability of Huh7 in nude mice (P<0.01). The immunohistochemical staining showed that the average optical density values of Ki-67 protein in tumor tissues of the control group, miR-29 mimics group and miR-29 analogue+ miR-29 inhibitor group were 0.65±0.08, 0.36±0.07 and 0.56±0.06, respectively. The expression level of Ki-67 protein in miR-29 mimics group was significantly reduced (P<0.01) while increased in the miR-29 mimics+ miR-29 inhibitor group (P<0.01). Conclusion: LncRNA HULC promotes HCC growth by down-regulating miR-29.

[Significance of p16/Ki-67 double immunocytochemical staining in cervical cytology ASCUS, LSIL, and ASC-H].

Objective: To investigate the application value of p16/cell proliferation associated nuclear antigen (Ki-67) double-staining and human papillomavirus mRNA in the cytological screening. Methods: Two hundred and fifty-one cases who suffered from atypical squamous cell of undetermined significance (ASCUS), low-grade squamous intraepithelial lesion (LSIL), atypical squamous cell-cannot exclude high-grade squamous intraepithelial lesion (ASC-H) in ThinPrep cytologic test (TCT) were collected in Peking University First Hospital between October 2015 and March 2016. And p16/Ki-67 double-staining and hybrid capture Ⅱ (HC-Ⅱ) detection were performed on the cervical cells. The result was compared with the pathological result of colposcope guided biopsy. All statistical analysis was completed by Stata 12.0 statistical software analysis. The results of diagnostic tests were described by using the sensitivity, specificity, positive predictive value,negative predictive value, and the area under the receiver operating characteristic (ROC) curve. Results: (1) One hundred and eight cases of liquid based cytology diagnosis of ASCUS patients, the positive rate of p16/Ki-67 was 13.9% (15/108), 102 cases of liquid based cytology diagnosis of LSIL patients, the positive rate of p16/Ki-67 was 21.6% (22/102), 41 cases of liquid based cytology diagnosis of ASC-H patients, the positive rate of p16/Ki-67 was 39.0% (16/41), compared amongthree groups, the difference was statistically significant (χ(2)=78.516, P<0.05); cervical exfoliated cells p16/Ki-67 expression rate was 13.0%(28/215) in cervical low-grade lesions [cervical intraepithelial neoplasia (CIN) Ⅰ], which was 69.4%(25/36) in high level lesions (CIN Ⅱ-Ⅲ), the difference was statistically significant (χ(2)=7.932, P<0.05). (2) The specificity of p16/Ki-67 detection and diagnosis were higher than those of HC-Ⅱ in ASCUS, LSIL, and ASC-H (89.8% vs 71.4%, 83.3% vs 15.6%, 88.9% vs 40.7%; all P<0.05), meanwhile, the positive predictive value of p16/Ki-67 detection and diagnosis exceed those of HC-Ⅱ in ASCUS, LSIL, and ASC-H (33.3% vs 26.3%, 31.8% vs 12.6%, 81.3% vs 38.5%; all P<0.05). Moreover, the ROC curve of p16/Ki-67 were bigger than those of HC-Ⅱ in ASCUS, LSIL, and ASC-H (0.799 vs 0.696, 0.708 vs 0.531, 0.909 vs 0.561; all P<0.05). Conclusion: For patients with cytological diagnosis of ASCUS, LSIL, and ASC-H, p16/Ki-67 double staining method could be used as an effective method to assist in the diagnosis of high-grade cervical lesions, and the screening efficiency is superior to that of high-rist HPV.

[Comparison of clinical pathological characteristics in ovarian preserving patients with stage IB1 cervical adenocarcinoma and squamous cell carcinoma].

OBJECTIVE: To analyze the risk and prognostic of patients with stage IB1 cervical adenocarcinoma. METHODS: The clinical data of 139 patients with stage IB1 cervical adenocarcinoma treated at Department of Gynecology and Obstetrics in Peking University First Hospital from August 1994 to April 2015 were retrospectively reviewed, which included 38 cases of cervical adenocarcinoma and 101 cases of cervical squamous cell carcinoma. A comparison was made between ovarian preserving group and bilateral oophorectomy group, in order to justify the risk and prognosis of ovarian preserving patients. RESULTS: The 5-year cumulative survival rate of stage IB1 cervical adenocarcinoma and squamous cell carcinoma were 89.1% and 92.9% respectively with significant difference (P=0.034). One ovarian metastasis case was observed among the 32 cervical adenocarcinoma patients of bilateral oophorectomy, while another ovarian metastasis case was observed among 54 cervical squamous cell carcinoma patients of bilateral oophorectomy. The ovarian metastasis rate was 3.1% (1/32) and 1.8 % (1/54) respectively with no statistical difference (P=0.574). The cumulative 5-year survival of 6 ovarian preserving patients with cervical adenocarcinoma was 80.1%, while that of 47 ovarian preserving patients with cervical squamous cell carcinoma was 94.6% (P=0.127). There was no statistical difference between the survival curve of the two groups. CONCLUSION: The prognosis of stage IB1 cervical adenocarcinomas was somewhat poorer than that of cervical squamous cell carcinoma. However it was still reasonable to perform ovarian preservation among young patients of stage IB1 cervical adenocarcinoma with no high risk factors.

Clinical significance of plasma fibrinogen and D-dimer in predicting the chemotherapy efficacy and prognosis for small cell lung cancer patients.

PURPOSE: Elevated plasma fibrinogen and D-dimer levels indicate activation of hemostasis and fibrinolysis, and this activation is required for tumor angiogenesis, metastasis, and invasion. Previous studies demonstrated that the plasma fibrinogen and D-dimer levels correlate with patient's prognosis in several solid tumors. The aim of this study is to examine the relationship between plasma fibrinogen and D-dimer levels before and during chemotherapy and treatment response and survival in patients with small cell lung cancer (SCLC). METHODS: Plasma fibrinogen and D-dimer levels before and during chemotherapy were prospectively measured in 74 SCLC patients who received first-line therapy. The results were analyzed for correlation between fibrinogen and D-dimer levels and treatment response, as well as progressive-free survival (PFS) and overall survival (OS). RESULTS: The levels of fibrinogen and D-dimer in SCLC patients before (C0) and after two cycles (C2) of chemotherapy were significantly higher than those in controls. Fibrinogen and D-dimer levels decreased during chemotherapy, and changes in fibrinogen and D-dimer levels between at C0 and at C2 were associated with treatment response. No matter which disease stage, patients with fibrinogen or D-dimer positivities at C0 and C2 time points had worse PFS and OS than those with fibrinogen or D-dimer negativities. Multivariate analyses revealed that fibrinogen and D-dimer positivities after two chemotherapy cycles were independently unfavorable factors for PFS and OS. CONCLUSION: Fibrinogen and D-dimer levels after two cycles of chemotherapy are predictors for response on chemotherapy and prognosis in SCLC patients.

Prognostic and predictive value of plasma D-dimer in advanced non-small cell lung cancer patients undergoing first-line chemotherapy.

PURPOSE: Previous studies have demonstrated that plasma D-dimer, a degradation product of cross-linked fibrin, correlated with tumor stage and prognosis in cancer patients. The aim of this study is to examine whether plasma D-dimer levels before and during chemotherapy predict tumor response and survival in advanced NSCLC patients undergoing first line chemotherapy. METHODS: Plasma D-dimer levels before (B0), and after one (B1) and two (B2) cycles of chemotherapy in 82 patients with advanced NSCLC were measured and correlated with treatment response, clinical features, progression-free survival (PFS) and overall survival. RESULTS: A significant correlation was identified between changes in D-dimer levels before and after two chemotherapy cycles and treatment response. In addition, there were significant correlations between D-dimer positivity at B0, B1 and B2 time points and tumor stage, number of metastatic sites and treatment response. Patients with positivity of D-dimer at B0, B1 and B2 had significantly shorter PFS compared with those with negativity. Notably, positivity of D-dimer at B1 and B2 time points was an independent predictive factor for unfavorable PFS. CONCLUSIONS: The positivity of D-dimer before and during chemotherapy is a predictor of treatment response and worse PFS in patients with advanced NSCLC. D-dimer levels provide prognostic information in addition to that of imaging studies.

Clinical significance of survivin and VEGF mRNA detection in the cell fraction of the peripheral blood in non-small cell lung cancer patients before and after surgery.

PURPOSE: The aim of this study was to evaluate the predictive and prognostic value of peripheral blood survivin and VEGF mRNA expression levels in non-small cell lung cancer (NSCLC) patients. PATIENTS AND METHODS: Fifty-eight patients with stage I-IIIA NSCLC who underwent surgical resection were enrolled in this study. Thirty-six patients with benign lung disease (BLD) entered this study as control group. Quantitative real-time PCR was used to detect survivin and VEGF mRNA levels in the cell fraction of peripheral blood in NSCLC patients before and after surgery and BLD patients. The relationship between blood survivin and VEGF mRNA levels and patients clinicopathologic parameters and prognostic factors were investigated. RESULTS: The levels of survivin and VEGF mRNA were decreased significantly after surgery in NSCLC patients (P = 0.024 and P = 0.012 respectively). Tumor recurrence was significantly more frequent in NSCLC patients with survivin and VEGF mRNA positivity postoperation than in patients without (P = 0.003 and P = 0.006, respectively). Patients with survivin or VEGF mRNA positivity postoperation had markedly shorter disease-free survival (DFS) and overall survival (OS) than patients without (P = 0.023 and P = 0.016 for survivin; P = 0.031 and P = 0.025 for VEGF, respectively). Multivariate analysis showed that survivin positivity preoperation (P = 0.026, P = 0.041, respectively) and postoperation (P = 0.003, P = 0.005, respectively) and VEGF mRNA positivity postoperation (P = 0.007, P = 0.009, respectively) were independently associated with DFS and OS. CONCLUSION: Although the levels of surviving and VEGF mRNA were decreased significantly after surgery, postoperative detections of survivin and VEGF mRNA by quantitative real-time PCR could be used as tools to monitor tumor recurrence and predict prognosis.

Serum proteomic features for detection of endometrial cancer.

To find new potential biomarkers for detection of endometrial cancer (EC), 70 serum samples including 40 from EC patients and 30 from normal healthy females were detected by surface-enhanced laser desorption-ionization time-of-flight mass spectrometry (SELDI-TOF-MS) using WCX2 (weak cation exchange) protein chip. Mass spectra were then assessed with three powerful data-mining tools: a tree classifier, Biomarker Wizard software, and Biomarker Patterns System. The diagnostic pattern combined with 13 potential biomarkers could differentiate EC patients from healthy persons, with a specificity of 100%, sensitivity of 92.5%, and total coincidence of 95.7%. The combination of surface-enhanced laser desorption-ionization with bioinformatics tools could help find new biomarkers and establish with high sensitivity and specificity for the detection of EC.