Ultrasonic Elastography Research Based on a Multicenter Study: Adding Strain Ratio after 5-Point Scoring Evaluation or Not

Background

This study aimed to confirm whether strain ratio should be added after evaluation of lesions with 5-point elasticity scoring for differentiating benign and malignant breast lesions on ultrasonographic elastography(UE).

Materials and Methods

From June 2010 to March 2012, 1080 consecutive female patients with breast lesions were recruited into a multicenter retrospective study, which involved 8 centers across China. Each institutional ethic review board approved the study, and all the patients gave written informed consent. All the patients underwent the UE procedure and the strain ratios were calculated and the final diagnosis was made by histological findings. The sensitivity, specificity, accuracy, PPV and NPV were calculated for each of the two evaluation systems and the areas under the ROC curve were compared.

Results

The strain ratios of benign lesions (mean, 2.6±2.0) and malignant lesions (mean,7.9±5.8) were significantly different (p <0.01). When the cutoff point was 3.01, strain ratio method had 79.8% sensitivity, 82.8% specificity, and 81.3% accuracy, while the 5-point scoring method had 93.1% sensitivity, 73.0% specificity, and 76.8% accuracy. The areas under the ROC curve with the strain ratio method and 5-point scoring method were 0.863 and 0.865, respectively(p>0.05). The strain ratio method shows better a diagnosis performance of the lesions with elasticity score 3 and 4.

Conclusions

Although the two UE methods have similar diagnostic performance, separate calculation of the strain ratios seems compulsory, especially for the large solid breast lesions and the lesions with elasticity score 3 and 4.     

Integrin-binding Protein Nischarin Interacts with Tumor Suppressor Liver Kinase B1 (LKB1) to Regulate Cell Migration of Breast Epithelial Cells

Biallelic inactivation of LKB1, a serine/threonine kinase, has been detected in 30% of lung adenocarcinomas, and inhibition of breast tumor growth has been demonstrated. We have identified the tumor suppressor, Nischarin, as a novel binding partner of LKB1. Our mapping analysis shows that the N terminus of Nischarin interacts with amino acids 44–436 of LKB1. Time lapse microscopy and Transwell migration data show that the absence of both Nischarin and LKB1 from an invasive breast cancer cell line (MDA-MB-231) enhances migration as measured by increased distance and speed of migrating cells. Our data suggest that this is a result of elevated PAK1 and LIMK1 phosphorylation. Moreover, the absence of Nischarin and LKB1 increased tumor growth in vivo. Consistent with this, the percentage of S phase cells was increased, as demonstrated by flow cytometry and enhanced cyclin D1. The absence of Nischarin and LKB1 also led to a dramatic increase in the formation of lung metastases. Our studies, for the first time, demonstrate functional interaction between LKB1 and Nischarin to inhibit cell migration and breast tumor progression. Mechanistically, we show that these two proteins together regulate PAK-LIMK-Cofilin and cyclin D1/CDK4 pathways.     

<Emphasis Type="Italic">Strboh A</Emphasis> homologue of NADPH oxidase regulates wound-induced oxidative burst and facilitates wound-healing in potato tubers

During 30-months of storage at 4°C, potato (Solanum tuberosum L.) tubers progressively lose the ability to produce superoxide in response to wounding, resist microbial infection, and develop a suberized wound periderm. Using differentially aged tubers, we demonstrate that Strboh A is responsible for the wound-induced oxidative burst in potato and aging attenuates its expression. In vivo superoxide production and NADPH oxidase (NOX) activity from 1-month-old tubers increased to a maximum 18–24 h after wounding and then decreased to barely detectable levels by 72 h. Wounding also induced a 68% increase in microsomal protein within 18 h. These wound-induced responses were lost over a 25- to 30-month storage period. Superoxide production and NOX activity were inhibited by diphenylene iodonium chloride, a specific inhibitor of NOX, which in turn effectively inhibited wound-healing and increased susceptibility to microbial infection and decay in 1-month-old tubers. Wound-induced superoxide production was also inhibited by EGTA-mediated destabilization of membranes. The ability to restore superoxide production to EGTA-treated tissue with Ca⁺² declined with advancing tuber age, likely a consequence of age-related changes in membrane architecture. Of the five homologues of NOX (Strboh A-D and F), wounding induced the expression of Strboh A in 6-month-old tubers but this response was absent in tubers stored for 25–30 months. Strboh A thus mediates the initial burst of superoxide in response to wounding of potato tubers; loss of its expression increases the susceptibility to microbial infection and contributes to the age-induced loss of wound-healing ability.     

ATP synthase complex of Plasmodium falciparum: dimeric assembly in mitochondrial membranes and resistance to genetic disruption

The rotary nanomotor ATP synthase is a central player in the bioenergetics of most organisms. Yet the role of ATP synthase in malaria parasites has remained unclear, as blood stages of Plasmodium falciparum appear to derive ATP largely through glycolysis. Also, genes for essential subunits of the F(O) sector of the complex could not be detected in the parasite genomes. Here, we have used molecular genetic and immunological tools to investigate the localization, complex formation, and functional significance of predicted ATP synthase subunits in P. falciparum. We generated transgenic P. falciparum lines expressing seven epitope-tagged canonical ATP synthase subunits, revealing localization of all but one of the subunits to the mitochondrion. Blue native gel electrophoresis of P. falciparum mitochondrial membranes suggested the molecular mass of the ATP synthase complex to be greater than 1 million daltons. This size is consistent with the complex being assembled as a dimer in a manner similar to the complexes observed in other eukaryotic organisms. This observation also suggests the presence of previously unknown subunits in addition to the canonical subunits in P. falciparum ATP synthase complex. Our attempts to disrupt genes encoding β and γ subunits were unsuccessful, suggesting an essential role played by the ATP synthase complex in blood stages of P. falciparum. These studies suggest that, despite some unconventional features and its minimal contribution to ATP synthesis, P. falciparum ATP synthase is localized to the parasite mitochondrion, assembled as a large dimeric complex, and is likely essential for parasite survival.     

Assessment of association between BRAF-V600E mutation status in melanomas and clinical response to ipilimumab

Ipilimumab, a fully human monoclonal antibody against cytotoxic T lymphocyte antigen-4, has demonstrated significant improvement in overall survival in previously treated advanced melanoma patients. The BRAF inhibitor, vemurafenib, has shown up to 78% objective response rates in melanoma patients harboring the BRAF-V600E mutation but not in patients lacking the mutation. As an immune potentiator, the mechanism of action of ipilimumab may not be dependent of the activity of the BRAF pathway. To test this, we investigated whether the clinical activity of ipilimumab would be affected by the BRAF-V600E mutation status of the tumors. Thus, this retrospective analysis was carried using a set of tumor biopsies from a completed phase II clinical trial. CA184004 was a randomized, double-blind, multicenter trial of 82 previously treated or untreated patients with unresectable stage III/IV melanoma. Patients received ipilimumab 3 or 10 mg/kg every 3 weeks for four doses followed by maintenance dosing in eligible patients. The BRAF-V600E mutation status for 80 patients was determined in tumor biopsies by PCR-based assays. Data on disease control were available for 69 patients with evaluated BRAF-V600E mutation status. Rates of objective responses and stable disease in patients with BRAF-V600E mutation positive tumors (30%) were comparable to those in patients with the wild-type gene (~33%). Eleven patients displayed Durable Disease Control (DDC) of which 55% had BRAF-V600E mutation positive tumors and 45% did not. In the 48 patients showing no DDC, the mutation frequency was 50%. In this study, no association between BRAF-V600E mutation status of melanoma tumors and DDC after treatment with ipilimumab was detected.     

Advances in selectable marker genes for plant transformation

Plant transformation systems for creating transgenics require separate process for introducing cloned DNA into living plant cells. Identification or selection of those cells that have integrated DNA into appropriate plant genome is a vital step to regenerate fully developed plants from the transformed cells. Selectable marker genes are pivotal for the development of plant transformation technologies because marker genes allow researchers to identify or isolate the cells that are expressing the cloned DNA, to monitor and select the transformed progeny. As only a very small portion of cells are transformed in most experiments, the chances of recovering transgenic lines without selection are usually low. Since the selectable marker gene is expected to function in a range of cell types it is usually constructed as a chimeric gene using regulatory sequences that ensure constitutive expression throughout the plant. Advent of recombinant DNA technology and progress in plant molecular biology had led to a desire to introduce several genes into single transgenic plant line, necessitating the development of various types of selectable markers. This review article describes the developments made in the recent past on plant transformation systems using different selection methods adding a note on their importance as marker genes in transgenic crop plants.