9.81 kPa (TypeⅠ);12 cases,ALPP 6.38-9.81 kPa (TypeⅡ);22 cases,ALPP

Objective To identify the effects of virus specific amino acids in the fusion active domains of paramyxovirus fusion proteins on the specific membrane fusion. Methods Site-directed mutagenesis was used to obtain mutants in the identified fusion active domains of Newcastle disease virus (NDV) fusion protein (F) and human parainfluenza virus (hPIV) fusion protein (F). All the mutant F genes were co-expressed with their homol-ogous or heterogenous hemagglutinin-neuraminidase (HN) genes in eukaryocytes. The fusion functions of mutants were assayed by Giemsa staining and reporter gene method. The expression efficiencies of mutants were assayed by fluorescence-activated cell sorter (FACS). Results In the NDV F mutants, N150D-L152D had 46.31% fusion activity of wide type. The fusion activities of N257D-N258D-Q259E, G271D-N272D and Q279E-Q281E almost disappeared, and they had only 1.25%, 3.14% and 2.23% of fusion activities, respectively, compared with wide type. N296D-N297D had 97.68% fusion activity of wide type. In the hPIV F mutants, D143A-E145A had 32.63% fusion activity of wide type. The fusion activity of E223Q-K224A almost disappeared, and it had only 1.91% fusion activity of wide type. K263A-R265A, D268A-D270A and R475A-R476A had 14.63%, 19.52% and 28.95% of fusion activities respectively compared with wild type. The analysis of FACS indicated that proteins of NDV F N257D-N258D-Q259E, G271D-N272D, Q279E-Q281E and hPIV F E223Q-K224A were not expressed on the cell surface, while proteins of the rest mutants were expressed nearly as the same as the wide types. Con-clusion As to NDV F, the amino acids of N257, N258, Q259, G271, N272, Q279 and Q281 were significant to the specific membrane fusion, and N150 and L152 were also important, but N296 and N297 were not. For hPIV F, the amino acids of E223 and K224 were significant to the specific membrane fusion, and D143, E145, K263, 11265, D268, D270, R475 and R476 were also important.

Objective To explore the impact of puerarin treatment on autophagy in rats with traumatic brain injury (TBⅠ) and the underlying mechanism.Methods Seventy five Sprague-Dawley (SD) rats were randomized into 5 groups:sham group (S group,n =15),traumatic brain injury group (TBⅠ group,n =15),TBⅠ + puerarin treatment group (TBⅠ + Pue group,n =15),TBⅠ + JNK inhibitor group (TBⅠ + SP group,n =15),and TBⅠ + JNK activator + Pue (TBⅠ + An + Pue group,n =15).Feeney method was applied to make rats with TBⅠ model.Mter that,head water content and neurological deficit score (NDS) were measured and recorded at day 1,3 and 7 in each group.Western blot was used to measure the JNK activity and autophagic marker proteins,including LC3B and Beclin1.Results Compared to S group,the head water content and NDS were decreased significantly among the others (P ＜ 0.05).The head water content and NDS in TBⅠ + Pue and TBⅠ + SP groups was decreased remarkably compared with TBⅠ group.Combined with puerarin and animycin treatments failed to reduce head water content and NDS compared to the TBⅠ + Pue group.Activated autophagy could be observed in TBⅠ group compared to S group.Compared to group S,LC3Ⅱ,Beclin1 and P-JNK1 were increased significantly.Pue and SP could reduce their expressions,respectively.Combined with puerarin and animycin treatments failed to reduce LC3Ⅱ,Beclin1 and P-JNK1 compared to TBⅠ + Pue group.Conclusions Puerarin could protect rats with TBⅠ via inhibiting autophagy,JNK signal pathway could involve the process of puerarin regulating autophagy.

ObjectiveTo construct the eukaryotic expression vector of human HYAL1 gene and obtain MCF-7 and ZR-75-30 cell clones expressing HYAL1 gene stably.MethodsThe cDNA encoding HYAL1 gene of human breast cancer was amplified by RT-PCR from the total RNA isolated from human MDA-MB-435S cells and inserted into pcDNA3.1/V5-His-TOPO vector.The recombinant plasmid was transferred into MCF-7 and ZR-75-30 cells.ResultsA 1332-bp DNA fragment was successfully amplified from human MDA-MB-435S cell.Restriction enzyme digestion analysis and DNA sequencing showed that HYAL1 gene was inserted into recombinant vector.RT-PCR analysis revealed that HYAL1 gene could be expressed stably in the transfected MCF-7 and ZR-75-30 and it had strong invasive potential.ConclusionThe eukaryotic expression vector of human HYAL1 gene was successfully constructed.MCF-7 and ZR-75-30 cell clones that can express HYAL1 gene were obtained and can promote the invasion.

Objective To study the effects of HYAL1 gene overexpression on invasive, angiogenic and proliferative ability of breast cancer cell lines MCF-7 and ZR-75-30. Methods Double-chamber co-culture technique was applied to construct the invasive model and angiogenic model in vitro, which was used to detect the invasive and angiogenic potential of breast cancer cell; MTT and flow cytometry were used to detect the proliferation of breast cancer cells. Results Breast cancer cells overexpressing HYAL1 gene showed stronger invasive potential and angiogenic potential than control cells, but had no significant difference on proliferative potential. Conclusion Overexpression of HYAL1 gene can promote the invasion and angiogenesis of breast cancer cells in vitro, but not affect the proliferation.

Objective:To analyze the factors related to metastasis of contralateral central lymph node (CLN) in cN0 papillary thyroid car-cinoma (PTC) and discuss the indications for CLN dissection. Methods:We enrolled 149 unilateral PTC patients who underwent total thyroidectomy and prophylactic bilateral (CLN) dissection. This work analyzed the relationship of gender, age, extrathyroidal extension, multifocality, thyroiditis, ipsilateral central lymph nodes, and prelaryngeal lymph node with CLNs. Results:The rates of metastasis to ip-silateral and contralateral central compartments were 73.2%and 23.5%, respectively. In univariate analysis, gender, age, tumor size, multifocality, and thyroiditis were not important in predicting contralateral central compartment lymph node metastasis (P=0.792, 0.097, 0.531, 0.269, and 1.000, respectively);by contrast, extrathyroidal extension (P=0.017), prelaryngeal lymph nodes (P=0.006), and ipsilateral CLNs (P<0.001) are related to CLN metastasis. However, multivariate analysis showed that ipsilateral central metastasis was an independent risk factor for lymph node metastasis in the contralateral central region when the number of ipsilateral central metas-tases is≥3 (P=0.010). Conclusion:Extracapsular invasion, prelaryngeal lymph nodes, and ipsilateral CLN influence the metastases of CLN. Bilateral CLN dissection should be performed when the number of ipsilateral central metastases is≥3 and there is merger of ex-tra-laryngeal lymph nodes or capsule invasion.

Objective To investigate the relationship between subgroups of central lymph node metastasis (sCLNM) and lateral lymph node metastasis (LNM) of unilatal papillary thyroid carcinoma (uPTC) with cervical lymph node negative(cN0).Methods The clinical and pathological data of 161 patients with cN0-uPTC who underwent total thyroidectomy+central lymph node dissection+lateral lymph node dissection from Jan.2016 to Dec.2016 were retrospectively analyzed.The relationship between the lymph node metastasis of each subarea in the central area of the affected side and the lymph node metastasis of the affected side was investigated.Results Binary logistic regression analysis of cN0-uPTC subregions in the affected central region showed:pre-laryngeal lymph node metastasis,pre-tracheal lymph node metastasis and paratracheal lymph node metastasis were independent risk factors for lymph node metastasis in the affected lateral region(P=0.008,0.016,0.035,respectively).Prelaryngeal lymph node metastasis was an independent risk factor for lymph node metastasis in the affected area Ⅱ (P=0.015).Pre-tracheal lymph node metastasis was an independent risk factor for lymph node metastasis in affected area Ⅲ (P=0.004).Pre-tracheal and para-tracheal lymph node metastasis were independent risk factors for lymph node metastasis in the affected Ⅳ area (P=0.035,0.011,respectively).Conclusions The lymph node metastasis pathway of thyroid cancer had certain regularity.The pre-laryngeal lymph node metastasis has the prediction value for the lymph node metastasis of the affected area Ⅱ.The pre-tracheal lymph node metastasis has the prediction value for the lymph node metastasis of the affected area Ⅲ.The pre-tracheal and paratracheal lymph node metastasis have the prediction value for lymph node metastasis of the affected area Ⅳ.Lymph node dissection in affected areas Ⅲ and Ⅳ needs to be considered in patients with pre-tracheal or paratracheal lymph node metastases.On this basis,lymph node dissection on the affected areas Ⅱ,Ⅲ,and Ⅳ might be considered if there is pre-laryngeal lymph node metastasis at the same time.

Objective:To retrospectively analyze the regularity and risk factors of skip metastasis (central lymph node negative and lat-eral lymph node positive) in papillary thyroid carcinoma (PTC). Methods:A total of 521 PTC patients underwent total thyroidectomy and central plus lateral lymph node dissection at The First Affiliated Hospital of Chongqing Medical University from January 2013 to De-cember 2016. Clinicopathological characteristics of the patients were collected and analyzed. Results:Skip metastasis rate of PTC was 8.3％(43/521). Tumors in the upper lobe (OR=3.401, 95％CI:1.770-6.536;P=0.001) and in the lateral part (OR=3.424, 95％CI:1.182-9.920;P=0.023) of the thyroid, as well as age above 45 (OR=2.856, 95％CI:1.488-5.482;P=0.002), were independent risk factors for skip metastases for this disease. Clinically node-negative (cN0) PTC patients with tumors in the upper lobe had higher possibility of skip metastases than those with clinically involved lateral neck nodes(cN1b) (P=0.022). Conclusion:Skip metastasis of PTC is not un-common. Thus, preoperative clinical assessment and imaging examination for lateral lymph node is necessary, especially for PTC pa-tients who are above 45 years old and with tumors in the upper lobe and/or unilateral area of thyroid. The lateral lymph node dissec-tion should be performed when necessary.