AIM:To construct a eukaryotic expression plasmid for enhanced green fluorescent protein(EGFP)and ZNF580 fusion protein,and study its subcellular localization in the transfected MGC803 cells.METHODS:The primers were designed according to the cDNA encoding sequence of ZNF580 full-length open reading frame(1-172aa),ZNF580 amino terminus(1-93aa)and ZNF580 carboxyl terminus(94-172aa).The three cDNA segments of PCR were cloned into pGEM-T vector.Then they were subcloned respectively into plasmid pEGFP-C1(enhanced green fluorescent protein).The subcellular localization of the fusion protein in MGC803 cells transfected with the vector was monitored by autofluorescence microscopy.RESULTS:Restricted enzymes analysis and DNA sequencing showed that the sequences of the pEGFP-ZNF580(1-172),pEGFP-ZNF580(1-93)and pEGFP-ZNF580(94-172)transgenic plasmid were correct.The fusion proteins of EGFP-ZNF580(1-172)and pEGFP-ZNF580(94-172)were localized in the nuclei.CONCLUSION:The recombinant eukaryotic expression vector pEGFP-ZNF580 has been successfully constructed.The nuclear localization signal is within amino acid residues 94 and 172 of ZNF580 carboxyl terminus(C2H2 zinc finger domain).

Objective: To investigate the expressions of programmed death ligand 1（PD-L1）in triple-negative breast cancer (TNBC) and its correlation with angiogenesis. Methods: 120 cases of TNBC patients who underwent surgery in the Fourth Hospital of Hebei Medical University from March 1, 2011 to June 1, 2012 were collected. The tumor tissues of patients were surgically resected and confirmed by pathology. PD-L1 protein expression in TNBC tissues of 120 patients was detected by tissue microarray combined with immunohistochemistry, and its relationship with various clinical indicators was analyzed. Blood vessels and lymphatic vessels were labeled withCD34andD2-40todetectmicrovesseldensity(MVD)andlymphaticvesseldensity(LVD)inTNBC.Results:Thepositiveexpression rate of PD-L1 in the tumor cells and interstitial infiltrating lymphocytes fromTNBC was 56.7% (68/120); No correlation was found between PD-L1 protein expression and the gender, age, histological grade, clinical stage, or tumor size of patients with TNBC (P>0.05), but related to the lymph node metastasis (P<0.05) and vascular thrombus (P<0.05). TNBC with high PD-L1 expression exhibited high incidence of lymph node metastasis and formation of vascular thrombus, and the expression of PD-L1 was positively correlated with MVD (r=0.500, P=0.02) as well as LVD (r=0.662, P=0.01). Log-Rank test showed that the survival time of TNBC patients with positive PD-L1 protein expression was significantly shorter than that of patients with negative expression (P<0.05). Cox multivariate analysis suggested that PD-L1 protein expression could be an independent prognostic factor for TNBC overall survival. Conclusion: PD-L1 plays an important role in TNBC angiogenesis and lymphangiogenesis, and is closely related to TNBC invasion and metastasis; blocking PD1/PD-L1 signal pathway is expected to be an effective new strategy for TNBC treatment.

Objective To determine if the carbon dioxide (CO 2) pressure used in laparoscopic cholecystectomy (LC) has any effect on the occurrence of postoperative shoulder pain. Methods One hundred and twenty consecutive patients undergoing LC were prospectively divided into three groups(40 patients in each group): Patients in group A underwent LC with 10 mm Hg of CO 2, those in group B with 12 mm Hg of CO 2, and in group C with 14 mm Hg of CO 2. PaO 2?PaCO 2?pH before and after LC were recorded. Shoulder pain was recorded on a visual analogue pain scale 1,3,6,12,24,48,72,96 h after operation. Results The intraoperative CO 2 consumption was significantly higher in group C (F=11.38,P

Objective:To express the glycoprotein D of herpes simplex virus type 2 (gD2) in the insect cells,and to determine its immunogenicity.Methods:HSV-2 genome was used as the template for amplification of gD2 extracellular domain fragment gene by PCR.The PCR product was inserted into the vector Bacmind,and the constructed recombinant plasmid gD2-Bacmind was transfected into the sf9 cells to package the recombinant baculovirus.The Sf9 cells were infected by recombinant baculovirus seed derived from the forth passage(P4),the titer of P4 recombinant baculovirus was detected by a plaque assay and the expression of recombinant protein gD2 was determined by Western blotting method.The supernatant of infected cells was collected and purified by Ni-NTA affinity chromatography to obtain the target protein gD2,the purified gD2 protein was used to immunize the BALB/c mice in 0, 2, 4 weeks (gD2 group),and PBS was used as negative control(PBS group);the titers of gD2 specific IgG in serum were detected by ELISA assay.Results: The PCR analysis and sequencing results proved that gD2-Bacmind was constructed correctly.The titer of recombinant baculovirus was 2.0×109 pfu·mL-1,the purified gD2 was about 37 000 with expectation,the percentage of gD2 in total protein was 90%.The average value of Log10 of titer of gD2 specific IgG in serum detected by ELISA assay in gD2 group at the sixth week was 4.34,and there was significant difference compared with PBS group(P<0.01).Conclusion: The gD2 expressed by insect-baculovirus expression vector system has the immunogenicity and can be selected as candidate protein for HSV-2 vaccine.

In the practice of building its humanistic environment of a research hospital,Nanjing Drum Tower hospital adheres to such humanistic characteristics of the hospital as humanistic concept,planning, environment,management,service,and quality.Furthermore,the hospital upholds such keys as learning, innovation, cooperation, undertaking, competition and development. High focus, high starting point planning,persistence,and down-to-earth efforts,effectively promoting hospital development.

Objective: To investigate the effect of extensive retraction clefts (RC, >20% of tumor volume) on prognosis in invasive breast carcinoma of no specific type (IBC-NST). Methods: A total of 2 184 cases of IBC-NST diagnosed at the Fourth Hospital of Hebei Medical University from January 2006 to December 2008 were collected. All the cases were diagnosed according to the latest guideline and standard. After excluding cases of shrinkage due to tissue fixation, 483 cases with RC were identified, and the clinical and pathological features were retrospectively analyzed. Results: Among the 483 cases, the mean tumor size was 2.0 cm (range 0.8 to 4.8 cm). Two hundred and thirty-two cases were moderately differentiated (48.0%), 97 were well differentiated (20.1%), 154 were poorly differentiated (31.9%); 382 (79.1%) cases were of stages Ⅰ and Ⅱ. A total of 177 cases (36.7%) had lymphatic invasion; nodal metastasis were found in 202 cases (41.8%). Extensive RC was found in 237 of 483 cases (49.1%). Follow-up information was available in 407 patients, and 46 died of breast cancer with survival time from 37 to 103 months. Multivariate analysis of extensive RC showed that tumor size, histological grade and nodal metastasis were risk factors of patients with IBC-NST (P<0.05). Lymphatic invasion and nodal metastasis were risk factors for extensive RCs in patients with IBC-NST (P<0.05). There was a high probability of lymph node metastasis in patients of extensive RC without lymphatic invasion, and the difference was statistically significant (P<0.05). Conclusions Both lymphatic invasion and nodal metastasis are risk factors of extensive RC. The presence of extensive RC in IBC-NST patients is correlated with poor outcome. Tumors with lymphatic invasion are more likely to show extensive RC.

Objective: To investigate the grading system for lymph vessel tumor emboli and its prognostic value in patients with invasive carcinomas of no special type (ICNST) of the breast. Methods: Clinical data of 466patients with ICNST were collected from January 2006 to December 2008 in the Fourth Hospital of Hebei Medical University. The expression levels of D2-40, estrogen receptor(ER), progesterone receptor(PR) and human epidermal growth factor receptor 2 (HER-2) were analyzed using immunohistochemical staining. Grades for lymph vessel tumoremboli were classified based on the number of mitotic and apoptotic figures in tumor cells under a high-power field. Correlation analysis was performed using Spearman rank correlation test. Kaplan-meier curves and Log-rank tests were used to analyze the survival rate. Multivariate Cox proportional hazard model was used to analyze the prognostic factors. Results: Among the 466 patients, grades for lymph vessel tumor emboli were categorized as follows: 280 cases were grade 0 (60.1%); 112 cases were grade 1 (24.0%); 58 cases were grade 2 (12.5%); 16 cases were grade 3 (3.4%). Correlation analyses showed that lymph vessel tumor emboli grading system was positively correlated with lymph node metastasis (r=0.365, P<0.001). Kaplan-Meier univariant analysis showed that histological grading, lymph vessel tumor emboli grading system, lymph node metastasis, the expression levels of ER, PR and HER-2 and molecular typing were associated with prognosis of patients (P<0.05 for all). Multivariate analysis of Cox proportional hazard model showed that lymph vessel tumor emboli grading system and lymph node metastasis were independent prognostic factors in patients with ICNST(P<0.05 for all). Conclusion Grading system for lymph vessel tumor emboli canpredict the clinical outcome of patients with ICNST.

Objective: To investigate the mechanisms by which D-methionine (D-Met) eradicates Porphyromonas gingivalis biofilms by suppressing cyclic dimeric GMP (c-di-GMP) levels. Methods : Cell viability, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were measured to determine the effective concentrations of D-Met, which were subsequently used in the following experiments. During the P. gingivalis biofilm formation inhibition experiment and the mature biofilm disassembly experiment, biofilm biomass, exopolysaccharide (EPS), biofilm morphology, integrity of the cell membrane, and the level of c-di-GMP were determined. Results : D-Met < 40 mmol/L was biocompatible. During the biofilm formation inhibition and mature biofilm disassembly experiments, D-Met ≥ 20 mmol/L decreased the biofilm biomass and the production of EPS. SEM analysis showed that the extracellular matrix and bacterial density were drastically reduced by D-Met ≥ 20 mmol/L. TEM detection showed that 35 mmol/L D-Met ruptured the cell membrane during biofilm formation and increased the permeability of the cell membrane in the disassembly phase of mature biofilms. C-di-GMP levels decreased with increasing concentrations of D-Met in a concentration-dependent manner. Conclusion D-Met ≥ 20 mmol/L could eradicate P. gingivalis biofilms by suppressing c-di-GMP levels.

Occlusal disorder is an abnormal condition in which the static and dynamic relations between the upper and lower teeth are not well aligned . The most common occlusal disorder in clinical practice is the inability to reach the intercuspal position due to early contact of individual points or occlusal interference due to occlusal high points, which can lead to periodontal tissue damage, decreased masticatory function, temporomandibular joint and muscle discomfort; these results can occur through the overactivation of the locus coeruleus-sympathetic-adrenal medullary system and hypothalamic-pituitary-adrenal axis induce elevated serum corticosteroid levels, which leads to chronic stress in the body. This article reviews the effects of chronic stress caused by occlusal disorder on bone tissue, stomatognathic system, emotional health and cognitive function. It has been found that occlusal disorders not only result in the loss of bone in the oral cavity, the reduction of bone mass in the whole body and damage to the local function of the stomatognathic system but also negatively affect the body’s anxiety, sleep, cognitive function and spatial memory ability as a result of the neuroendocrine changes . In recent years, concern about occlusal disorders has been on the rise. Early detection and timely adjustment of uncoordinated occlusion has become an issue that cannot be ignored in the clinic.

Background: There is still controversy whether the existence of esophageal heterotopic gastric mucosa (EHGM) and its histological type are related to the laryngopharyngeal symptoms. Aims: To analyze the clinical and histological characteristics of EHGM and its correlation with gastroesophageal reflux. Methods: A retrospective study was conducted in consecutive gastroscopy-proved EHGM cases from September 2018 to January 2020 at the Second Affiliated Hospital of Baotou Medical College. Besides clinical data review and questionnaire survey on reflux symptoms, histological typing of EHGM and immunohistochemistry were also performed in some cases. Results: A total of 1 229 cases of EHGM were recruited. The male-to-female ratio was 1.67:1, and middle-aged people were predominant. Most of the heterotopic mucosa were located 15-18 cm away from the incisors, and were mainly single. Two hundred and ninety-four cases (23.9%) were complicated with reflux esophagitis (RE), of which Los Angeles grade A and B accounted for 96.6%. Regurgitation/acid reflux (15.5 %) and heartburn (12.3%) were the most common esophageal symptoms, while extraesophageal symptoms were rare. Histological typing was obtained in 57 cases, of which, 37 (64.9%) were cardia-type, 18 (31.6%) were fundic-type, and 2 (3.5%) were mixed type. There were no significant differences in gender, age, location and number of EHGM, expression levels of H