Correction: Safety verification for polysorbate 20, pharmaceutical excipient for intramuscular administration, in Sprague-Dawley rats and New Zealand White rabbits.

[This corrects the article DOI: 10.1371/journal.pone.0256869.].

Safety verification for polysorbate 20, pharmaceutical excipient for intramuscular administration, in Sprague-Dawley rats and New Zealand White rabbits.

Human serum albumin (HSA) has been widely used as a pharmaceutical excipient in Botulinum toxin serotype A (BoNT/A) products that are indicated for use in therapeutics and cosmetics. However, HSA as a human-derived material has some concerns, such as the potential risk of transmission of infectious agents, an insufficient supply, and difficulty in maintaining a certain quality. For those reasons, newly developed BoNT/A products (CORETOX®, Medytox, Inc., Republic of Korea) contained polysorbate 20, a non-human-derived excipient, to replace the HSA. However, most safety studies of polysorbate 20 have been conducted with non-invasive routes of administration, and thus there are a few studies on the safety of polysorbate 20 when administered intramuscularly. To secure the in vivo safety profile of polysorbate 20, a four-week repeated intramuscular dose toxicity study (0.02, 0.1, and 0.4 mg/kg, one injection every two weeks for a total of three injections) was conducted in 66 Sprague-Dawley (SD) rats. An intradermal irritation study was further conducted with 18 New Zealand White (NZW) rabbits. The toxicological evaluation of HSA (0.06 and 0.12 mg/kg) was also carried out as a comparative substance. Systemic and local toxicities were not observed in any of the SD rats or NZW rabbits based on clinical signs, body weight, hematology, clinical biochemistry, macroscopic findings on necropsy, histopathology of the injection site, and allergic reactions. The current study suggested that intramuscular administration of polysorbate 20 was considered to be safe at a level similar to that of HSA, which has an in vivo safety profile accumulated over the years. This provided the basis for the in vivo safety profile of polysorbate 20 administered intramuscularly and the scientific reliability of the use of polysorbate 20 as an alternative to HSA, which is used as an excipient for various pharmaceuticals in terms of its safety.

Comparative Analyses of Inflammatory Response and Tissue Integration of 14 Hyaluronic Acid-Based Fillers in Mini Pigs.

PURPOSE: Hyaluronic acid (HA)-based dermal fillers have been approved for various clinical indications, both cosmetic and medical. Previous studies that have assessed the performance of HA dermal fillers have primarily focused on evaluating filler durability, and only a few have studied their distribution within the tissues. The present study aimed to compare tissue integration of various types of HA dermal fillers having different clinical indications and varying injection depths. METHODS: To examine the local inflammatory response and distribution pattern of 14 HA dermal fillers (six Neuramis [NEU], one Belotero [BEL], three Juvéderm [JUV], and four Restylane [RES]), each product was injected intradermally and subcutaneously at the backs of two male miniature pigs. Histopathological evaluation and visual examination of the tissue sections were conducted 1 and 4 weeks after injection. RESULTS: Mean inflammatory cell infiltration scores tended to be lower in response to fillers from the NEU and BEL series than to those from the JUV and RES series after intradermal and subcutaneous injection. Furthermore, the inflammatory response to fillers with higher physicochemical properties specifically designed for injection into deeper layers of the skin tended to be slightly higher than those designated for injection into more superficial layers. There was no significant difference in tissue integration according to clinical indication and injection depth, although fillers from the NEU and BEL series exhibited better tissue integration than those from the JUV and RES series. CONCLUSION: Our findings not only suggest that the local inflammatory response and tissue integration differ across HA dermal filler products, but also that these parameters could vary according to the recommended clinical indication and injection depth of the products.

The development and validation of a predictive model for recurrence in rectal cancer based on radiological and clinicopathological data.

OBJECTIVE: To develop a prediction model for recurrence by incorporating radiological and clinicopathological prognostic factors in rectal cancer patients. METHODS: All radiologic and clinicopathologic data of 489 patients with rectal cancer, retrospectively collected from a single institution between 2009 and 2013, were used to develop a predictive model for recurrence using the Cox regression. The model performance was validated on an independent cohort between 2015 and 2017 (N = 168). RESULTS: Out of 489 derivative patients, 103 showed recurrence after surgery. The prediction model was constructed with the following four significant predictors: distance from anal verge, MR-based extramural venous invasion, pathologic nodal stage, and perineural invasion (HR: 1.69, 2.09, 2.59, 2.29, respectively). Each factor was assigned a risk score corresponding to HR. The derivation and validation cohort were classified by sum of risk scores into 3 groups: low, intermediate, and high risk. Each of these groups showed significantly different recurrence rates (derivation cohort: 13.4%, 35.3%, 61.5 %; validation cohort: 6.2%, 23.7%, 64.7%). Our new model showed better performance in risk stratification, compared to recurrence rates of tumor node metastasis (TNM) staging in the validation cohort (stage I: 3.6%, II: 12%, III: 30.2%). The area under the receiver operating characteristic curve of the new prediction model was higher than TNM staging at 3-year recurrence in the validation cohort (0.853 vs. 0.731; p = .009). CONCLUSIONS: The new risk prediction model was strongly correlated with a recurrence rate after rectal cancer surgery and excellent for selection of high-risk group, who needs more active surveillance. KEY POINTS: • Multivariate analysis revealed four significant risk factors to be MR-based extramural venous invasion, perineural invasion, nodal metastasis, and the short distance from anal verge among the radiologic and clinicopathologic data. • Our new recurrence prediction model including radiologic data as well as clinicopathologic data showed high predictive performance of disease recurrence. • This model can be used as a comprehensive approach to evaluate individual prognosis and helpful for the selection of highly recurrent group who needs more active surveillance.

Critical role of neutralizing antibody for SARS-CoV-2 reinfection and transmission.

Cases of laboratory-confirmed SARS-CoV-2 reinfection have been reported in a number of countries. Further, the level of natural immunity induced by SARS-CoV-2 infection is not fully clear, nor is it clear if a primary infection is protective against reinfection. To investigate the potential association between serum antibody titres and reinfection of SARS-CoV-2, ferrets with different levels of NAb titres after primary SARS-CoV-2 infection were subjected to reinfection with a heterologous SARS-CoV-2 strain. All heterologous SARS-CoV-2 reinfected ferrets showed active virus replication in the upper respiratory and gastro-intestinal tracts. However, the high NAb titre group showed attenuated viral replication and rapid viral clearance. In addition, direct-contact transmission was observed only from reinfected ferrets with low NAb titres (<20), and not from other groups. Further, lung histopathology demonstrated the presence of limited inflammatory regions in the high NAb titre groups compared with control and low NAb groups. This study demonstrates a close correlation between a low NAb titre and SARS-CoV-2 reinfection in a recovered ferret reinfection model.

TGF-ß induced EMT and stemness characteristics are associated with epigenetic regulation in lung cancer.

Transforming growth factor-ß (TGF-ß) promotes tumor invasion and metastasis by inducing epithelial-mesenchymal transition (EMT). EMT is often related with acquisition of stemness characteristics. The objective of this study was to determine whether EMT and stemness characteristics induced by TGF-ß might be associated with epigenetic regulation in lung cancer. A human normal lung epithelial cell line and four lung cancer cell lines were treated with TGF-ß. Transcriptome analysis of BEAS-2B and A549 cells incubated with TGF-ß were analyzed through next-generation sequencing (NGS). Western blotting was carried out to investigate expression levels of epithelial and mesenchymal markers. Wound healing and Matrigel invasion assay, sphere formation assay, and in vivo mice tumor model were performed to evaluate functional characteristics of EMT and stemness acquisition. To investigate whether activation of EMT and stem cell markers might be involved in epigenetic regulation of lung cancer, experiment using a DNA methyltransferase inhibitor (5-azacytidine, AZA), methylation-specific PCR (MSP) and bisulfite sequencing were performed. NGS revealed changes in expression levels of EMT markers (E-cadherin, N-cadherin, fibronectin, vimentin, slug and snail) and stem cell markers (CD44 and CD87) in both BEAS-2B and A549 cells. Functional analysis revealed increased migration, invasion, sphere formation, and tumor development in mice after TGF-ß treatment. Expression of slug and CD87 genes was activated following treatment with AZA and TGF-ß. MSP and bisulfite sequencing indicated DNA demethylation of slug and CD87 genes. These results suggest that TGF-ß induced EMT and cancer stemness acquisition could be associated with activation of slug and CD87 gene by their promoter demethylation.

Risk of active tuberculosis development in contacts exposed to infectious tuberculosis in congregate settings in Korea.

Contact investigation is an important and effective active case-finding strategy, but there is a lack of research on congregate settings in countries with an intermediate incidence. This study determined the incidence of and risk factors for tuberculosis (TB) development after exposure in congregate settings. This retrospective cohort study included 116,742 contacts identified during the investigation of 2,609 TB cases diagnosed from January to December 2015. We searched the Korean National Tuberculosis Surveillance System TB registry to identify contacts that developed active TB during follow-up until May 2018. During the mean observation period of 2.9 years, 499 of 116,742 contacts (0.4%) developed new active TB. From these contacts, 404 (81.0%) developed TB within 2 years after exposure. The 2-year Kaplan-Meier cumulative risk for TB was the highest in contacts aged ≥65 years [1%; 95% confidence interval (CI), 0.8-1.3]. Contacts with LTBI who completed chemoprophylaxis exhibited a lower risk of active TB development than those without chemoprophylaxis (adjusted hazard ratio, 0.16; 95% CI, 0.08-0.29). Aggressive contact investigation is effective for the early detection and prevention of TB in congregate settings. The risk of progression to active TB among contacts with LTBI can be minimised by the completion of chemoprophylaxis.

Spoligotype Variation of Mycobacterium tuberculosis Strains Prevailing in Korea.

Tuberculosis (TB) is an ongoing global health problem, including in South Korea. To manage TB efficiently, it is necessary to understand the epidemiology, transmission route, and characteristics of prevailing Mycobacterium tuberculosis strains. In this study, we investigated microevolutions over time in the spoligotype patterns of M. tuberculosis isolated from TB patients in Korea. We collected 1,055 clinical M. tuberculosis isolates from 16 provinces in Korea from 1994 to 2006 and analyzed them by spoligotyping. We observed 26 subfamilies, including two large predominant families: a Beijing family (72.7%) and the T family (19.1%). Specifically, the abundance of spoligotype SIT269 from the Beijing-like subfamily significantly increased in the 2000s relative to the 1990s in Korea. This study provides an overview of the M. tuberculosis genotype trends over time in Korea. These data also indicate that we should consider the influence of the newly growing SIT269 subtype identified in the Beijing family.

Effects of birth months on rheumatic diseases in South Korea: a nationwide case-control study.

OBJECTIVES: Birth month/season impacts the development of certain diseases. However, the effect of birth month/season on the development of rheumatic diseases has not been thoroughly investigated. Thus, the objective of this study was to determine whether birth month/season might affect the development of rheumatic diseases. METHODS: Birth month patterns of patients with various rheumatic diseases were compared with those of the general population. The dataset included 17,247,458 individuals from the health insurance review and assessment service database of Korea. RESULTS: Among 24 rheumatic diseases, the development of Crohn's disease, ulcerative colitis (UC), rheumatoid arthritis, Sjögren's syndrome, polymyalgia rheumatica, ankylosing spondylitis (AS), gout, and fibromyalgia (FM) was significantly associated with birth month/season. UC and AS were more prevalent in individuals born in February/winter. On the contrary, those who were born in June or July/summer were at a higher risk of gout and FM. CONCLUSIONS: Seasonal variations in infectious agents, sun exposure, and food ingestion during gestation or early infancy seem to explain the association between birth month/season and development of rheumatic diseases.

The risk of active tuberculosis among individuals living in tuberculosis-affected households in the Republic of Korea, 2015.

BACKGROUND: In the Republic of Korea (ROK), compared to other high-income countries, tuberculosis (TB) prevalence is relatively high. Active TB and latent TB infection (LTBI) surveillance of individuals living in TB-affected households has been conducted for several years. Although active case finding is an important strategy in low-prevalence, high-income countries, its effectiveness in a high prevalence setting is unclear. This study evaluated the risk of TB in household contact by calculating the incidence of TB among household contacts and comparing it with the general population of the ROK. METHODS: A retrospective cohort study, including 36,133 household-contacts of 17,958 TB patients reported in 2015, was conducted. The data was extracted from the Korean National TB Surveillance System (web-based TB cases notification system, KNTSS). The Cox proportional hazard regression model was used to evaluate risk factors for incidence of TB. A P-value < .05 was considered statistically significant. RESULTS: In this study, 319 (0.9%) of 36,133 household-contacts were reported as having TB within 1 year, which is a higher rate than the rate for the general population in the ROK. The rate of TB reported for contacts that had completed LTBI treatment (0.6%) was lower than for the LTBI group without treatment (4.6%). In multivariate analysis, age older than 65 (p < .001), being a spouse of a TB patient (p = .007), and LTBI without treatment (p = .013) were each a risk factor for TB incidence among contacts. Younger age (p < .001), presence of a cough (p < .001), testing positive for acid-fast bacilli (AFB; p < .001), and cavity on radiograph (p < .001) of the index patient were also statistically significant risk factors. CONCLUSIONS: Individuals living in TB-affected households are at high risk of developing TB in the ROK and active case finding among them is a strategy effective in the early detection and prevention of TB.

Clinical profiles of early and tuberculosis-related mortality in South Korea between 2015 and 2017: a cross-sectional study.

BACKGROUND: Although the incidence of tuberculosis (TB) has decreased in South Korea, the mortality rate remains high. TB mortality is a key indicator for TB control interventions. The purpose of this study was to assess early and TB-related mortality during anti-TB treatment and describe the associated clinical characteristics. METHODS: A multicenter cross-sectional study was performed across South Korea. Patients with pulmonary TB who died during anti-TB treatment and whose records were submitted to the national TB surveillance system between 2015 and 2017 were enrolled. All TB deaths were categorized based on cause (TB-related or non-TB-related) and timing (early or late). We identified statistical associations using the frequency table, chi-square test, and binary logistic regression. RESULTS: Of 5595 notifiable mortality cases, 3735 patients with pulmonary TB were included in the analysis. There were 2541 (68.0%) male patients, and 2935 (78.6%) mortality cases were observed in patients older than 65 years. There were 944 (25.3%) cases of TB-related death and 2545 (68.1%) cases of early death. Of all cases, 187 (5.0%) patients were diagnosed post-mortem and 38 (1.0%) patients died on the first day of treatment. Low body mass index (adjusted odds ratio (aOR) = 1.26; 95% confidence interval (CI) = 1.08-1.48), no reported illness (aOR = 1.36; 95% CI = 1.10-1.68), bilateral disease on chest X-ray (aOR = 1.30; 95% CI = 1.11-1.52), and positive acid-fast bacilli smear result (aOR = 1.30; 95% CI = 1.11-1.52) were significantly associated with early death, as well as TB-related death. Acute respiratory failure was the most common mode of non-TB-related death. Malignancy was associated with both late (aOR = 0.71; 95% CI = 0.59-0.89) and non-TB-related (aOR = 0.35; 95% CI = 0.26-0.46) death. CONCLUSIONS: A high proportion of TB death was observed in elderly patients and attributed to non-TB-related causes. Many TB-related deaths occurred during the intensive phase, particularly within the first month. Further studies identifying risk factors for different causes of TB death at different phases of anti-TB treatment are warranted for early targeted intervention in order to reduce TB mortality.

Comparison of PANAMutyper and PNAClamp for Detecting KRAS Mutations from Patients With Malignant Pleural Effusion.

BACKGROUND/AIM: KRAS is one of the frequently mutated genes in human cancers and often relates with drug resistance and poor prognosis. PANAMutyper™ is a novel technology that integrates PNAClamp™ and PANA S-Melting™. In the present study, PANAMutyper™ and PNAClamp™ were compared for the detection of KRAS mutations using different samples of patients with malignant pleural effusion. PATIENTS AND METHODS: A total of 103 patients (including 56 lung adenocarcinoma, 10 lung squamous carcinoma, 17 small cell lung cancer, 3 large cell lung cancer, 3 stomach cancer, 2 ovarian cancer, and others) with malignant pleural effusion were investigated using matched tumor tissue, cell block, and pleural effusion samples. The diagnostic performance of these two methods was compared. RESULTS: KRAS mutations were detected in 18 (17.5%) of 103 patients using tissue, cell block, and pleural effusion samples. All 18 patients with KRAS mutations were detected by PANAMutyper™ using any sample type, however, only 7 cases were detected by PNAClamp™. Among the subtypes of KRAS mutations, substitution in codon 12, 35G>T was the most frequent, followed by substitution in codon 12, 35G>A and codon 12, 34G>A. In pleural effusion specimens, PANAMutyper™ showed a better diagnostic performance compared to PNAClamp™. CONCLUSION: PANAMutyper™ had a diagnostic superiority for the detection of KRAS mutations in patients with malignant pleural effusion compared to PNAClamp™, although there was a concordance between PANAMutyper™ and PNAClamp™ results. Therefore, PANAMutyper™ can be used for a more sensitive and accurate detection of KRAS mutations.

Comparison of PNA Clamping-assisted Fluorescence Melting Curve Analysis and PNA Clamping in Detecting EGFR Mutations in Matched Tumor Tissue, Cell Block, Pleural Effusion and Blood of Lung Cancer Patients With Malignant Pleural Effusion.

BACKGROUND/AIM: This study compared the efficacy of PANAMutyper™, a novel technology that integrates PNAClamp™ and PANA S-Melting™, and PNAClamp™ alone for the detection of EGFR mutations in lung cancer patients. MATERIALS AND METHODS: PANAMutyper™ and PNAClamp™ were used to assess the EGFR mutation status in tissue, cell block, pleural effusion, and blood samples of 90 lung cancer patients with malignant pleural effusion. RESULTS: PANAMutyper™ detected more EGFR mutations than PNAClamp™, especially in body fluids (pleural effusion and serum). Patients with additional EGFR mutations detected using PANAMutyper™ had a favorable response to EGFR-tyrosine kinase inhibitor (TKI) treatment. CONCLUSION: The diagnostic performance of PANAMutyper™ was superior to that of PNAClamp™ for the detection of EGFR mutations. It was also better at identifying lung cancer patients with malignant pleural effusion who were likely to benefit from EGFR-TKI treatment.

Hypoxia-induced cancer stemness acquisition is associated with CXCR4 activation by its aberrant promoter demethylation.

BACKGROUND: A hypoxic microenvironment leads to an increase in the invasiveness and the metastatic potential of cancer cells within tumors via the epithelial-mesenchymal transition (EMT) and cancer stemness acquisition. However, hypoxia-induced changes in the expression and function of candidate stem cell markers and their possible molecular mechanism is still not understood. METHODS: Lung cell lines were analyzed in normoxic or hypoxic conditions. For screening among the stem cell markers, a transcriptome analysis using next-generation sequencing was performed. For validation, the EMT and stem cell characteristics were analyzed. To determine whether an epigenetic mechanism was involved, the cell lines were treated with a DNA methyltransferase inhibitor (AZA), and methylation-specific PCR and bisulfite sequencing were performed. RESULTS: Next-generation sequencing revealed that the CXCR4 expression was significantly higher after the hypoxic condition, which functionally resulted in the EMT and cancer stemness acquisition. The acquisition of the EMT and stemness properties was inhibited by treatment with CXCR4 siRNA. The CXCR4 was activated by either the hypoxic condition or treatment with AZA. The methylation-specific PCR and bisulfite sequencing displayed a decreased CXCR4 promoter methylation in the hypoxic condition. CONCLUSIONS: These results suggest that hypoxia-induced acquisition of cancer stem cell characteristics was associated with CXCR4 activation by its aberrant promoter demethylation.

A contact investigation after exposure to a child with disseminated tuberculosis mimicking inflammatory bowel disease.

PURPOSE: Tuberculosis (TB) is one of the most important diseases that cause significant mortality and morbidity in young children. Data on TB transmission from an infected child are limited. Herein, we report a case of disseminated TB in a child and conducted a contact investigation among exposed individuals. METHODS: A 4-year-old child without Bacille Calmette-Guérin vaccination was diagnosed as having culture-proven disseminated TB. The child initially presented with symptoms of inflammatory bowel disease, and nosocomial and kindergarten exposures were reported. The exposed individuals to the index case were divided into 3 groups, namely household, nosocomial, or kindergarten contacts. Evaluation was performed following the Korean guidelines for TB. Kindergarten contacts were further divided into close or casual contacts. Chest radiography and tuberculin skin test or interferon-gamma-releasing assay were performed for the contacts. RESULTS: We examined 327 individuals (3 household, 10 nosocomial, and 314 kindergarten contacts), of whom 18 (5.5%), the brother of the index patient, and 17 kindergarten children were diagnosed as having latent TB infection (LTBI). LTBI diagnosis was more frequent in the children who had close kindergarten contact with the index case (17.1% vs. 4.4%, P=0.007). None of the cases had active TB. CONCLUSION: This is the first reported case of TB transmission among young children from a pediatric patient with disseminated TB in Korea. TB should be emphasized as a possible cause of chronic diarrhea and failure to thrive in children. A national TB control policy has been actively applied to identify Korean children with LTBI.

Clinical Characteristics of False-Positive Lymph Node on Chest CT or PET-CT Confirmed by Endobronchial Ultrasound-Guided Transbronchial Needle Aspiration in Lung Cancer.

BACKGROUND: Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is a standard procedure to evaluate suspicious lymph node involvement of lung cancer because computed tomography (CT) and 18F-fluorodeoxyglucose positron emission tomography-CT (PET-CT) have limitations in their sensitivity and specificity. There are a number of benign causes of false positive lymph node such as anthracosis or anthracofibrosis, pneumoconiosis, old or active tuberculosis, interstitial lung disease, and other infectious conditions including pneumonia. The purpose of this study was to evaluate possible causes of false positive lymph node detected in chest CT or PET-CT. METHODS: Two hundred forty-seven patients who were initially diagnosed with lung cancer between May 2009 and December 2012, and underwent EBUS-TBNA to confirm suspicious lymph node involvement by chest CT or PET-CT were analyzed for the study. RESULTS: Of 247 cases, EBUS-TBNA confirmed malignancy in at least one lymph node in 189. The remaining 58 patients whose EBUS-TBNA results were negative were analyzed. Age ≥65, squamous cell carcinoma as the histologic type, and pneumoconiosis were related with false-positive lymph node involvement on imaging studies such as chest CT and PET-CT. CONCLUSION: These findings suggest that lung cancer staging should be done more carefully when a patient has clinically benign lymph node characteristics including older age, squamous cell carcinoma, and benign lung conditions.

Histogram analysis of volume-based apparent diffusion coefficient in breast cancer.

Background Breast cancer is a heterogeneous disease. Recent studies showed that apparent diffusion coefficient (ADC) values have various association with tumor aggressiveness and prognosis. Purpose To evaluate the value of histogram analysis of ADC values obtained from the whole tumor volume in invasive ductal cancer (IDC) and ductal carcinoma in situ (DCIS). Material and Methods This retrospective study included 201 patients with confirmed DCIS (n = 37) and IDC (n = 164). The IDC group was divided into two groups based on the presence of a DCIS component: IDC-DCIS (n = 76) and pure IDC (n = 88). All patients underwent preoperative breast magnetic resonance imaging (MRI) with diffusion-weighted images at 3.0 T. Histogram parameters of cumulative ADC values, skewness, and kurtosis were calculated and statistically analyzed. Results The differences between DCIS, IDC-DCIS, and pure IDC were significant in all percentiles of ADC values, in descending order of DCIS, IDC-DCIS, and pure IDC. IDC showed significantly lower ADC values than DCIS, and ADC50 was the best indicator for discriminating IDC from DCIS, with a threshold of 1.185 × 10-3 mm2/s (sensitivity of 82.9%, specificity of 75.7%). However, multivariate analysis of obtained ADC values showed no significant differences between DCIS, IDC-DCIS, and pure IDC ( P > 0.05). Conclusion Volume-based ADC values showed association with heterogeneity of breast cancer. However, there was no additional diagnostic performance in histogram analysis for differentiating between DCIS, IDC-DCIS, and pure IDC.

Development of an analytical method for detecting nitrofurans in bee pollen by liquid chromatography-electrospray ionization tandem mass spectrometry.

Bee pollen collected by honeybees, which is in powdered form, is a good nutritional supplement. Nitrofuran antibiotics are assumed not to be present in bee pollen, which is important as the level of antibiotics in bee pollen is strongly regulated in many countries. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to detect nitrofurans in honey has been developed, but this method is not suitable for bee pollen because of it being in powdered form. During preparation of bee pollen samples, the dispersal of powder particles in an aqueous solution often makes them susceptible to forming an emulsion with solvent components such as hexane and ethyl acetate. This may reduce the reproducibility and sensitivity of analyses of nitrofuran levels in bee pollen. Therefore, we attempted to optimize the sample preparation conditions to detect nitrofurans in bee pollen by determining three nitrofuran residues, namely, 3-amino-2-oxazolidinone (AOZ), 3-amino-5-methyl-morpholino-2-oxazolidinone (AMOZ), and 1-aminohydantoin (AHD), using LC-MS/MS. The optimized method prevented the formation of powder-induced emulsion. To verify the reproducibility and sensitivity of this method, it was validated using nitrofuran-free bee pollen spiked with analytes with different side chains at 1.0, 2.0, and 5.0µgkg-1. The accuracy levels were 94.1%-104.0% and the coefficients of variation were less than 12%. The limits of detection for AOZ, AMOZ, and AHD were 0.18, 0.25, and 0.30µgkg-1, respectively, while their limits of quantitation were 0.59, 0.83, and 1.00µgkg-1. The LC-MS/MS method developed to analyze nitrofuran in bee pollen should contribute to the quality control of bee pollen and food safety.

MARCKSL1 exhibits anti-angiogenic effects through suppression of VEGFR-2-dependent Akt/PDK-1/mTOR phosphorylation.

Myristoylated alanine-rich C kinase substrate-like 1 (MARCKSL1) plays a pivotal role in the regulation of apoptosis and has been shown to maintain antitumor and metastasis-suppressive properties. In the present study, we examined the effects of MARCKSL1 as a novel anti-angiogenic agent on the inhibition of angiogenesis-mediated cell migration. MARCKSL1 also reduced vascular endothelial growth factor (VEGF)-induced human umbilical vein endothelial cell (HUVEC) proliferation, as well as capillary-like tubular structure formation in vitro. MARCKSL1 disrupted phosphorylation of vascular endothelial growth factor receptor-2 (VEGFR-2) in ovarian tumorigenesis. In addition, MARCKSL1 showed potent anti-angiogenic activity and reduced the levels of VEGF and hypoxia-inducible factor 1α (HIF-1α) expression, an essential regulator of angiogenesis. Consistently, MARCKSL1 decreased VEGF­induced phosphorylation of the PI3K/Akt signaling pathway components, including phosphoinositide-dependent protein kinase 1 (PDK-1), mammalian target of rapamycin (mTOR), tuberous sclerosis complex 2 (TSC-2), p70 ribosomal protein S6 kinase (p70S6K), and glycogen synthase kinase 3ß (GSK-3ß) protein. Collectively, our results provide evidence for the physiological/biological function of an endothelial cell system involved in angiogenesis through suppression of Akt/PDK-1/mTOR phosphorylation by interaction with VEGFR-2.

Comparison of automated breast ultrasonography to handheld ultrasonography in detecting and diagnosing breast lesions.

BACKGROUND: Automated breast ultrasonography (ABUS) is increasingly used as a screening tool. Several studies have demonstrated a similar diagnostic performance for ABUS compared with handheld ultrasonography (HHUS), but the overall results have been controversial. PURPOSE: To compare the clinical utility of ABUS and HHUS for detection and diagnosis of breast lesions. MATERIAL AND METHODS: ABUS and HHUS images of suspicious breast lesions were obtained for 173 consecutive women scheduled to undergo ultrasonography (US)-guided or stereotactic biopsy. There were a total of 206 lesions, 46 of which were malignant and 160 benign. Three breast radiologists took part in this study: two reviewed the ABUS images, and the third reviewed all of the images, ABUS and HHUS, as well as the patients' medical records. The biopsied-lesion-detection rates were obtained. Using the Breast Imaging Reporting and Data System (BI-RADS), the images of the biopsied lesions were evaluated. Factors affecting ABUS detectability were analyzed. RESULTS: The overall detection rates were 83.0% for ABUS and 94.2% for HHUS. Ten lesions were not detected on either HHUS or ABUS and these were microcalcifications (one malignancy and nine benign lesions). Of the 194 HHUS-detected lesions, 169 were detected by ABUS and 25 benign were not. ABUS less frequently detected lesions of smaller size as well as those of benign appearance and lower final-assessment category (P = 0.011 and P < 0.0001, respectively). CONCLUSION: ABUS detected all of the malignant lesions that were detected on HHUS. ABUS missed several smaller benign lesions.