[Intein-Mediated Protein trans-Splicing of the Recombinant Streptavidin on Magnetosomes].

When expressing streptavidin recombinant polypeptide on magnetosomes (called bacterial magnetic nanoparticles, or BMPs), the presence of endogenous bacterial biotin might be detrimental. In the study, the streptavidin monomer fragment (S1-116) was fused with the intein N-terminal (termed precursor S1-116-IN), and S1-116-IN was expressed in E. coli (BL21). Meanwhile, the SA117-160 fragment was fused with the C-terminal intein, and then this chimeric polypeptide was expressed on magnetosomes by fusion with magnetosome membrance protein MamF. In the in vitro protein splicing system, the purified engineered magnetosomes (BMP-SA117-160-IC) and the S1-116-IN precursor were mixed. Intein-mediated trans-splicing reaction was induced to produce the functional magnetic beads BMP-SA. Our results indicate that intein-mediated protein trans-splicing may lead to efficient synthesis of the recombinant streptavidin on the magnetosomes, showing its promising potential to produce other functional magnetic nanoparticles.

[Knee stability after anterior cruciate ligament reconstruction with weight-bearing magnetic resonance imaging].

Objective: To investigate the application value of the weight-bearing magnetic resonance imaging (MRI) in evaluating the stability of knee joint after anterior cruciate ligament reconstruction. Methods: From July 2011 to August 2013, a total of 25 patients with anterior cruciate ligament rupture and reconstruction surgery in the Second Affiliated Hospital of Soochow University were enrolled in this study, including 18 males and 7 females, with an average age of (32±5) years. All the patients underwent the weight-bearing MRI, knee joint passive relaxation test (Kneelax 3), and Lysholm score before the surgery and 3 and 6 months after the surgery. The three examinations before and after the operation were analyzed by repeated measures of general linear model, and paired t test was used to compare the results before and after the operation. The correlation between the three preoperative examinations was statistically analyzed. Results: The relaxation data measured by Kneelax 3 after the surgery was significantly lower than that before the operation [(1.1±0.9) mm vs (6.1±1.3) mm, t=16.9, P<0.01]. The post-operative lateral tibial plateau anterior shift score was less than the pre-operative score [(3.0±0.7) mm vs (4.8±1.2) mm, t=6.2, P<0.01]. The post-operative Lysholm score was significantly higher than that before the operation (89±6 vs 64±14, t=-8.3, P<0.05). There was a negative correlation between the anterior displacement of the lateral platform and Lysholm score (r=-0.902, P<0.01). There was no correlation between anterior displacement and passive relaxation of the lateral platform in preoperative weight-bearing MRI. Conclusions: Anterior cruciate ligament reconstruction can improve the knee stability. The stability of knee joint can be evaluated by weight-bearing MRI. The anterior tibial displacement measured by the weight-bearing MRI is correlated with the clinical score.

Mur (MNS 10) screening with a novel loop-mediated isothermal amplification assay in Zhongshan, China.

BACKGROUND: The distribution of the Mur blood group antigen is 5-7% in the south of China, and a much higher prevalence is observed in some areas of the region. Anti-Mur can cause hemolytic disease of the newborn and severe transfusion reactions. OBJECTIVES: Genetic testing is more ideal than conventional serological tests because antibodies for detection are usually not available. METHODS: In this study, a novel loop-mediated isothermal amplification (LAMP) assay for the detection of Mur blood group antigen was established. RESULTS: Fifteen of 275 (5·5%) samples were confirmed by LAMP as Mur antigen positive. All the Mur antigen-positive samples were GP.Mur subtype which was confirmed with sequencing. CONCLUSION: The LAMP method has identical results with conventional serology method but more suitable for large-scale screening.

Imputation strategies for low-coverage whole-genome sequencing data and their effects on genomic prediction and genome-wide association studies in pigs.

The uncertainty resulting from missing genotypes in low-coverage whole-genome sequencing (LCWGS) data complicates genotype imputation. The aim of this study is to find out an optimal strategy for accurately imputing LCWGS data and assess its effectiveness for genomic prediction (GP) and genome-wide association study (GWAS) on economically important traits of Large White pigs. The LCWGS data of 1 423 Large White pigs were imputed using three different strategies: (1) using the high-coverage whole-genome sequencing (HCWGS) of 30 key progenitors as the reference panel (Ref_LG); (2) mixing HCWGS of key progenitors with LCWGS (Mix_HLG) and (3) self-imputation in LCWGS (Within_LG). Additionally, to compare the imputation effects of LCWGS, we also imputed SNP chip data of 1 423 Large White pigs to the whole-genome sequencing level using the reference panel consisting of key progenitors (Ref_SNP). To evaluate effects of the imputed sequencing data, we compared the accuracies of GP and statistical power of GWAS for four reproductive traits based on the chip data, sequencing data imputed from chip data and LCWGS data using an optimal strategy. The average imputation accuracies of the Within_LG, Ref_LG and Mix_HLG were 0.9893, 0.9899 and 0.9875, respectively, which were higher than that of the Ref_SNP (0.8522). Using the imputed sequencing data from LCWGS with the Ref_LG imputation strategy, the accuracies of GP for four traits improved by approximately 0.31-1.04% compared to the chip data, and by 0.7-1.05% compared to the imputed sequencing data from chip data. Furthermore, by using the sequence data imputed from LCWGS with the Ref_LG, 18 candidate genes were identified to be associated with the four reproductive traits of interest in Large White pigs: total number of piglets born - EPC2, MBD5, ORC4 and ACVR2A; number of piglets born healthy - IKBKE; total litter weight of piglets born alive - HSPA13 and CPA1; gestation length - GTF2H5, ITGAV, NFE2L2, CALCRL, ITGA4, STAT1, HOXD10, MSTN, COL5A2 and STAT4. With the exception of EPC2, ORC4, ACVR2A and MSTN, others represent novel candidates. Our findings can provide a reference for the application of LCWGS data in livestock and poultry.

[The oncologic and functional outcomes of supracricoid partial laryngectomy for the treatment of laryngeal cancer].

Objective: To evaluate the oncologic and functional outcomes of laryngeal squamous cell carcinomas treated by supracricoid laryngectomy. Methods: The clinical data of 134 patients with laryngeal cancer who underwent supracricoid laryngectomy with cricohyoidoepiglottopexy (CHEP) or cricohyoidopexy (CHP) between July 2005 and April 2014 at Shandong Provincial ENT Hospital were retrospectively reviewed. Ninety-one patients including 31 cases of stage Ⅰ, 36 of stage Ⅱ, 18 of stage Ⅲ and 6 of stage Ⅳ underwent CHEP and 43 patients underwent CHP. Two patients received CHEP due to recurrence after open surgery and laser surgery. Three patients received CHP due to the recurrence of disease after open surgery and postradiotherapy persistence of disease. The Kaplan-Meier method was used to calculate the 3-year and 5-year survival rates. The Chi-square test was used to compare the survival rates between different surgical procedures. Results: All 91 patients who underwent CHEP had successful removals of PEG tubes, and 88 (96.7%) of them had tracheostomy tube decannulation. Among 43 patients with CHP, 42(97.6%) cases removal of PEG tubes(97.6%), including and 40(93.0%) cases with tracheostomy tube decannulation. There was one patient with local recurrence in all cases. In CHEP group, 3-year local control rate was 98.2%; 3-year and 5-year overall survival rate were 94.5% and 93.9%, respectively. In CHP group, 3-year local control rate was 97.6%; 3-year and 5-year overall survival rates were 86.0% and 83.3%, respectively. Pharyngeal fistula appeared in 2 cases of CHEP group and 4 cases of CHP group, and all of them were cured by conservative treatment. Conclusion: Supracricoid laryngectomy shows excellent oncologic and functional results for treatment of laryngeal cancer while maintaining laryngeal functions, especially in terms of local control rate and tracheostomy tube decannulation.

Involvement of circular RNA SMARCA5/microRNA-620 axis in the regulation of cervical cancer cell proliferation, invasion and migration.

OBJECTIVE: Circular RNAs (circRNAs) have emerged as crucial regulators for tumor progression. However, the effects of circRNAs are not entirely clear in cervical cancer (CC). The objective of this study was to investigate the function and regulation mechanism of circular RNA SMARCA5 (circSMARCA5, also named hsa_circ_0001445) in cervical cancer. PATIENTS AND METHODS: circSMARCA5 and miR-620 expressions were analyzed by qRT-PCR assay. The cell proliferation, cell cycle, migration, invasion abilities were measured by CCK-8 and colony formation, Flow cytometry, Wound-healing and transwell assays. In addition, the interaction between circSMARCA5 and miR-620 was accessed by luciferase reporter assay. RESULTS: The results indicated that circSMARCA5 was downregulated in CC; overexpression of circSMARCA5 inhibited CC cell proliferation, migration and invasion, and induced cell cycle arrest. In addition, we found that circSMARCA5 could bind to miR-620 and significantly downregulated its expression. Furthermore, the results revealed that circSMARCA5 suppressed proliferation and invasion by miR-620 in CC. CONCLUSIONS: We suggested that circSMARCA5/miR-620 regulatory axis is involved in the development of CC and may serve as a potential therapy target.

[Significance of retropharyngeal node dissection in treatment of hypopharyngeal carcinoma].

Objective: investigate the incidence of retropharyngeal lymph node (RPLN) metastasis and the risk factors for RPLN metastasis in hypopharyngeal cancer, and the relationship of planned dissection of the RPLN with the survival and tumor control rates in patients with hypopharyngeal cancer. Methods: A total of 203 patients with hypopharyngeal squamous cell carcinoma who underwent radical surgery as initial treatment from February 2011 to July 2015 were analyzed retrospectively. There were 167 cases of pyriform sinus carcinoma, 23 cases of posterior pharyngeal wall carcinoma, and 13 cases of postcricoid carcinoma. Results: The incidence of RPLN metastasis in HPC was 17.7%, with a highest rate of 43.5% in pharyngeal wall carcinoma. The incidence of RPLN metastasis in T3-4 pyriform sinus carcinoma was 18.3%, which significantly higher than 2.8% in T1-2 cases(χ(2)=5.360, P=0.020). The rate of RPLN metastasis was 23.8% in N2b-3 and 8.6% in N0-2a, with a statistically significant difference(χ(2)=7.637, P=0.006). There was no statistically significant difference in overall survival rates between patients with and without RPLN metastasis(P>0.05). Data were analyzed by SPSS 13.0 software. Conclusions: RPLN metastasis is not rare in hypopharyngeal carcinoma. Planned dissection of the RPLN should be performed with the initial surgery in patients with advanced hypopharyngeal cancer, especially posterior pharyngeal wall carcinoma, T3-4 pyriform sinus carcinoma and staged N2b-3 disease, which can reduce the regional recurrence rate and provided with a better prognosis.

[Endoscopic screening for upper gastrointestinal second primary malignancies in patients with hypopharyngeal squamous cell carcinoma].

Objective: To evaluate the usefullness of flexible esophagoscopy and chromoendoscopy with Lugol's solution in the detection of synchronous esophageal neoplasm in patients with hypopharyngeal squamous cell carcinoma (HSCC). Methods: A retrospective review of 96 cases with HSCC that received surgical treatment from March 2016 to March 2017 was accomplished. In these patients, the site of origin were pyriform sinus (n=75), posterior pharyngeal wall (n=11) and postcricoid (n=10). Esophagoscopy was prospectively performed on all patients before treatment for HSCC. All patients underwent conventional white-light endoscopic examination with Lugol chromoendoscopy and narrow band image. Suspicious areas of narrow band image or Lugol-voiding lesions were observed and biopsied. The treatment strategy of primary HSCC was modified according to the presence of synchronous esophageal squamous cell neoplasms by a multidisciplinary approach. Results: Ninety-six patients were enrolled (age ranging from 37-80 years). All patients did not have previous treatment.Histopathological analysis revealed middle to high-grade dysplasia in 5 cases, Tis cancer in 5 cases, cancer in 16 cases and inflammation or normal findings in the others. Four cases were treated with endoscopic submucosal dissection before hypopharygeal surgery, 3 cases with lower esophageal cancers were treated with gastric pull-up combined with free jejunal flap after total circumferential pharyngolaryngectomy (TCPL) and certical esophagectomy, and 14 cases were treated with TCPL, total esophagectomy and gastric pull-up. Conclusions: Esophagoscopy is a feasible and justified procedure in HSCC cases as it enhances the detection of premalignant lesion or second primary cancer. Routine esophagoscopy for detecting synchronous second primary tumor should be recommended for patients with HSCC. The treatment strategy for primary HSCC is modified according to the presence of synchronous second primary tumor.

[Treatment and prognosis of 264 patients with hypopharyngeal carcinoma].

Objective: To investigate the key factors influencing the prognosis of hypopharyngeal carcinoma and the therapeutic methods improving the efficacy of treatments for hypopharyngeal carcinoma. Methods: Two hundred and sixty-four cases of hypopharyngeal squamous cell carcinoma treated from May 2010 to May 2015 were analyzed retrospectively. There were 211 cases of pyriform sinus carcinoma, 37 cases of posterior pharyngeal wall carcinoma, and 16 cases of postcricoid carcinoma. According to UICC 2002 criteria, 2 cases were for stage Ⅰ, 14 for stage Ⅱ, 32 for stage Ⅲ and 216 for stage Ⅳ. Postoperative circumferential defects existed in 112 (42.4%) cases, and 86 of them were reconstructed with free jejunum transplantation. Among all cases, 54 patients (20.5%) had the preservation of laryngeal functions after surgery and 210 patients (79.5%) with total laryngectomy; 238 cases (90.2%) underwent bilateral cervical lymph node dissection and 203 patients received posterior pharyngeal lymph node exploration and dissection, with positive metastases for posterior pharyngeal lymph nodes in 36 cases (17.7%). Eight cases with cervical lymph node metastasis extensively involving the soft tissue, prevertebral fascia or encases carotid artery received preoperative radiotherapy of 50 Gy. After surgery 13 patients received concurrent radiotherapy and chemotherapy, 337 underwent adjuvant radiotherapy with a dose of 50-60 Gy each, and 14 patients did not receive radiotherapy or did not completed their radiotherapy programs. SPSS 13.0 saftware was used to analyze the data. Results: All patients were followed up for more than 2 years. With Kaplan-Meier method, the 2-, 3- and 5-years survival rates were 69.6%, 62.8% and 51.3%, respectively. There were significant differences in 3-year survival rates between T1-2 group (75.5%) and T3-4 group (59.2%) (χ(2)=4.282 P=0.039), N0 group (81.6%) and N+ group (58.2%) (χ(2)=6.802 P=0.009), laryngeal functions preserved (81.8%) and unpreserved group (58.9%) (χ(2)=5.314 P=0.021). Multivariate Logistic regression analysis showed that cervical lymph node metastasis was an independent prognostic factor (P=0.027). The success rate of free jejunum transplantation was 98.8%. Local recurrence, cervical lymph node recurrence, second primary cancer, and distant metastasis accounted respectively for 11.2%, 18.8%, 12.5% and 45.0% of death cases. Conclusions: The prognosis-associated factors for hypopharyngeal carcinoma should be taken into account, including the evaluation of the carcinogenesis of the mucosal area, early screening of premalignant lesion or second primary cancer in the esophagus and dissection of the posterior pharyngeal lymph nodes, which will help to improve the local control rate and recent survival rate in patients with hypopharyngeal cancer.

Arterial spin-labeling perfusion MRI stratifies progression-free survival and correlates with epidermal growth factor receptor status in glioblastoma.

BACKGROUND AND PURPOSE: Glioblastoma is a common primary brain tumor with a poor but variable prognosis. Our aim was to investigate the feasibility of MR perfusion imaging by using arterial spin-labeling for determining the prognosis of patients with glioblastoma. MATERIALS AND METHODS: Pseudocontinuous arterial spin-labeling with 3D background-suppressed gradient and spin-echo was acquired before surgery on 53 patients subsequently diagnosed with glioblastoma. The calculated CBF color maps were visually evaluated by 3 independent readers blinded to patient history. Pathologic and survival data were correlated with CBF map findings. Arterial spin-labeling values in tumor tissue were also quantified by using manual fixed-size ROIs. RESULTS: Two perfusion patterns were characterized by visual evaluation of CBF maps on the basis of either the presence (pattern 1) or absence (pattern 2) of substantial hyperperfused tumor tissue. Evaluation of the perfusion patterns was highly concordant among the 3 readers (κ = 0.898, P < .001). Pattern 1 (versus pattern 2) was associated with significantly shorter progression-free survival by Kaplan-Meier analysis (median progression-free survival of 182 days versus 485 days, P < .01) and trended with shorter overall survival (P = .079). There was a significant association between pattern 1 and epidermal growth factor receptor variant III expression (P < .01). CONCLUSIONS: Qualitative evaluation of arterial spin-labeling CBF maps can be used to stratify survival and predict epidermal growth factor receptor variant III expression in patients with glioblastoma.

Effects of shrimp consumption on plasma lipoproteins.

Shrimp is very low in total fat, yet it has a high cholesterol content. Although shrimp is a popular food in the American diet, many people avoid it because of its high cholesterol content. The objective of this study was to test the effect of the addition of cholesterol from shrimp to a low-fat baseline diet as well as to compare the effect of an equal amount of dietary cholesterol derived from shrimp or egg on the plasma lipoprotein pattern of normolipidemic subjects. In a randomized crossover trial, a diet containing 300 g shrimp/d, which supplied 590 mg dietary cholesterol/d, significantly increased low-density-lipoprotein (LDL) cholesterol by 7.1% (P = 0.014) and high-density-lipoprotein (HDL) cholesterol by 12.1% (P = 0.0001) when compared with a baseline diet matched for fat content but containing only 107 mg cholesterol/d. However, because the percentage increase in LDL cholesterol was less than for HDL cholesterol, the shrimp diet did not worsen the ratio of total cholesterol to HDL cholesterol or the ratio of LDL to HDL cholesterol. Moreover, shrimp consumption decreased triacylglycerol (triglyceride) concentrations by 13% (P = 0.004). The diet containing two large eggs per day with 581 mg dietary cholesterol/d also raised LDL- and HDL-cholesterol concentrations compared with baseline, but the percentage increase in LDL cholesterol (10.2%, P = 0.0001) was more than for HDL cholesterol (7.6%, P = 0.004) and there was a trend toward worse lipoprotein ratios. In a comparison of the two high-cholesterol diets, the shrimp diet produced significantly lower ratios of total to HDL cholesterol and lower ratios of LDL to HDL cholesterol than the egg diet as well as lower triacylglycerol concentrations. We conclude that moderate shrimp consumption in normolipidemic subjects will not adversely affect the overall lipoprotein profile and can be included in "heart healthy" nutritional guidelines.

On prediction of future observation in growth curve model.

Rao proposed and compared several approaches for predicting future observations in a growth curve model. The assessment of associated prediction efficiency for different prediction methods were evaluated by Cross-Validation Assessment Error (CVAE). He used three data sets, each with a limited number of subjects (13-27) and also with a limited number of repeated measurements (4-7) per subject, to illustrate the prediction methods. In the present paper, we applied four of the prediction methods discussed by Rao, on a data set with a relatively large number of subjects (174) and also with a larger number of measurements (21) per subject, using the polynomial function and log-linear function. We propose to use the restricted cubic spline function as an alternative growth curve model and compare its performance with the polynomial function and log-linear function. It turns out that, at least for larger data sets such as that used in this paper, the prediction methods perform somewhat better when the growth is described by restricted cubic spline function than when the growth is described by polynomial function and log-linear function.