Epstein-Barr virus (EBV) alters the regulation and expression of a variety of cytokines in its host cells to modulate host immune surveillance and facilitate viral persistence. Using cytokine antibody arrays, we found that, in addition to the cytokines reported previously, two chemotactic cytokines, CCL3 and CCL4, were induced in EBV-infected B cells and were expressed at high levels in all EBV-immortalized lymphoblastoid cell lines (LCLs). Furthermore, EBV latent membrane protein 1 (LMP1)-mediated Jun N-terminal protein kinase activation was responsible for upregulation of CCL3 and CCL4. Inhibition of CCL3 and CCL4 in LCLs using a short hairpin RNA approach or by neutralizing antibodies suppressed cell proliferation and caused apoptosis, indicating that autocrine CCL3 and CCL4 are required for LCL survival and growth. Importantly, significant amounts of CCL3 were detected in EBV-positive plasma from immunocompromised patients, suggesting that EBV modulates this chemokine in vivo. This study reveals the regulatory mechanism and a novel function of CCL3 and CCL4 in EBV-infected B cells. CCL3 might be useful as a therapeutic target in EBV-associated lymphoproliferative diseases and malignancies.
The objective of this study was to evaluate the antihyperlipidemic and antihyperglycemic effects and mechanism of the extract of Clitocybe nuda (CNE), in high-fat- (HF-) fed mice. C57BL/6J was randomly divided into two groups: the control (CON) group was fed with a low-fat diet, whereas the experimental group was fed with a HF diet for 8 weeks. Then, the HF group was subdivided into five groups and was given orally CNE (including C1: 0.2, C2: 0.5, and C3: 1.0 g/kg/day extracts) or rosiglitazone (Rosi) or vehicle for 4 weeks. CNE effectively prevented HF-diet-induced increases in the levels of blood glucose, triglyceride, insulin (P < 0.001, P < 0.01, P < 0.05, resp.) and attenuated insulin resistance. By treatment with CNE, body weight gain, weights of white adipose tissue (WAT) and hepatic triacylglycerol content were reduced; moreover, adipocytes in the visceral depots showed a reduction in size. By treatment with CNE, the protein contents of glucose transporter 4 (GLUT4) were significantly increased in C3-treated group in the skeletal muscle. Furthermore, CNE reduces the hepatic expression of glucose-6-phosphatase (G6Pase) and glucose production. CNE significantly increases protein contents of phospho-AMP-activated protein kinase (AMPK) in the skeletal muscle and adipose and liver tissues. Therefore, it is possible that the activation of AMPK by CNE leads to diminished gluconeogenesis in the liver and enhanced glucose uptake in skeletal muscle. It is shown that CNE exhibits hypolipidemic effect in HF-fed mice by increasing ATGL expression, which is known to help triglyceride to hydrolyze. Moreover, antidiabetic properties of CNE occurred as a result of decreased hepatic glucose production via G6Pase downregulation and improved insulin sensitization. Thus, amelioration of diabetic and dyslipidemic states by CNE in HF-fed mice occurred by regulation of GLUT4, G6Pase, ATGL, and AMPK phosphorylation.
Objective. To evaluate the efficacy of autologous cytokine-induced killer (CIK) cells in patients with renal cell carcinoma (RCC). Methods. 20 patients diagnosed with TNM stage I or II RCC were randomly divided into two groups, a CIK cell treatment group and a control group. The endpoint was progression-free survival (PFS) evaluated by Kaplan-Meier analyses. Results. CD3+, CD3+/CD8+, CD3+/CD4+, and CD3+/CD56+ levels increased after CIK cell culture (P < 0.01). The median PFS in CIK cell treatment group was significantly longer than that in control group (PFS, 32.2 months versus 21.6 months; log-rank, P = 0.032), all patients were alive during the course of followup, and there are no statistically significant differences between two groups in OS (log-rank, P = 0.214). Grade III or greater adverse events were not observed. Conclusions. CIK cells treatment could prolong survival in patients with RCC after radical nephrectomy and showed acceptable curative effect with potential enhancement of cellular immune function. This trial is registered with Clinicaltrials.gov NCT01799083.
A current prevalence and relevant risk factors for impaired fasting glucose (IFG) have been reported by various ethnic groups and countries. By contrast, nationwide data for the incidence of IFG in Taiwan have not been presented in the past 15 years. The aim for this manuscript was to estimate the prevalence of IFG and associated risk factors in the population of Taiwan.
For this cross-sectional research, we used a nationally representative sample (N = 2604) obtained from the 2005–2008 Nutrition and Health Survey in Taiwan (NAHSIT), and adopted a stratified multistage sampling design. The tools employed for data collection included questionnaire interviews, anthropometry measurements, and laboratory analysis.
The prevalence of IFG among adults in Taiwan is 35.8% (a fasting glucose level between 100 and 125 mg/dl is considered abnormal). An estimated number of people with IFG is 6.5 million. A higher prevalence of IFG is observed in men younger than 65 years compared to women. However, this trend is reversed for the elderly population. The factors significantly associated with IFG include the following: sex, age, overweight (27 > BMI ≥ 24), obesity (BMI ≥ 27), waist circumference (men ≥ 90 cm, women ≥ 80 cm), hypertension, and hyperlipidemia.
IFG among adults in Taiwan is a health concern that requires attention. We recommend targeting the younger population, especially overweight and obese men between the ages of 19 and 40 years, to provide applicable healthy lifestyle counseling and services. Furthermore, appropriate screening of elderly people is required to detect undiagnosed IFG cases and provide early intervention and treatment.
Impaired fasting glucose (IFG); Prevalence; Adult; Epidemiology; Taiwan
IF (intermediate filament) proteins can be cleaved by caspases to generate proapoptotic fragments as shown for desmin. These fragments can also cause filament aggregation. The hypothesis is that disease-causing mutations in IF proteins and their subsequent characteristic histopathological aggregates could involve caspases. GFAP (glial fibrillary acidic protein), a closely related IF protein expressed mainly in astrocytes, is also a putative caspase substrate. Mutations in GFAP cause AxD (Alexander disease). The overexpression of wild-type or mutant GFAP promotes cytoplasmic aggregate formation, with caspase activation and GFAP proteolysis. In this study, we report that GFAP is cleaved specifically by caspase 6 at VELD225 in its L12 linker domain in vitro. Caspase cleavage of GFAP at Asp225 produces two major cleavage products. While the C-GFAP (C-terminal GFAP) is unable to assemble into filaments, the N-GFAP (N-terminal GFAP) forms filamentous structures that are variable in width and prone to aggregation. The effect of N-GFAP is dominant, thus affecting normal filament assembly in a way that promotes filament aggregation. Transient transfection of N-GFAP into a human astrocytoma cell line induces the formation of cytoplasmic aggregates, which also disrupt the endogenous GFAP networks. In addition, we generated a neo-epitope antibody that recognizes caspase-cleaved but not the intact GFAP. Using this antibody, we demonstrate the presence of the caspase-generated GFAP fragment in transfected cells expressing a disease-causing mutant GFAP and in two mouse models of AxD. These findings suggest that caspase-mediated GFAP proteolysis may be a common event in the context of both the GFAP mutation and excess.
Alexander disease; caspase; GFAP; intermediate filament; AD, Alzheimer’s disease; AxD, Alexander disease; C-GFAP, C-terminal GFAP; CNS, central nervous system; DTT, dithiothreitol; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GFAP, glial fibrillary acidic protein; IF, intermediate filament; N-GFAP, N-terminal GFAP OA, okadaic acid; TBS, Tris-buffered saline; TBST, TBS containing 0.1% (v/v) Tween 20
The development of a new vaccine as a substitute for Bacillus Calmette–Guerin or to improve its efficacy is one of the many World Health Organization goals to control tuberculosis. Mycobacterial vectors have been used successfully in the development of vaccines against tuberculosis. To enhance the potential utility of Mycobacterium smegmatis as a vaccine, it was transformed with a recombinant plasmid containing the partial sequences of the genes Ag85c, MPT51, and HspX (CMX) from M. tuberculosis. The newly generated recombinant strain mc2-CMX was tested in a murine model of infection. The recombinant vaccine induced specific IgG1 or IgG2a responses to CMX. CD4+ and CD8+ T cells from the lungs and spleen responded ex vivo to CMX, producing IFN-γ, IL17, TNF-α, and IL2. The vaccine thus induced a significant immune response in mice. Mice vaccinated with mc2-CMX and challenged with M. tuberculosis showed better protection than mice immunized with wild-type M. smegmatis or BCG. To increase the safety and immunogenicity of the CMX antigens, we used a recombinant strain of M. smegmatis, IKE (immune killing evasion), to express CMX. The recombinant vaccine IKE-CMX induced a better protective response than mc2-CMX. The data presented here suggest that the expression of CMX antigens improves the immune response and the protection induced in mice when M. smegmatis is used as vaccine against tuberculosis.
The septin gene belongs to a highly conserved family of polymerizing GTP-binding cytoskeletal proteins. SEPTs perform cytoskeletal remodeling, cell polarity, mitosis, and vesicle trafficking by interacting with various cytoskeletons. Our previous studies have indicated that SEPTIN12+/+/+/− chimeras with a SEPTIN12 mutant allele were infertile. Spermatozoa from the vas deferens of chimeric mice indicated an abnormal sperm morphology, decreased sperm count, and immotile sperm. Mutations and genetic variants of SEPTIN12 in infertility cases also caused oligozoospermia and teratozoospermia. We suggest that a loss of SEPT12 affects the biological function of microtublin functions and causes spermiogenesis defects. In the cell model, SEPT12 interacts with α- and β-tubulins by co-immunoprecipitation (co-IP). To determine the precise localization and interactions between SEPT12 and α- and β-tubulins in vivo, we created SEPTIN12-transgene mice. We demonstrate how SEPT12 interacts and co-localizes with α- and β-tubulins during spermiogenesis in these mice. By using shRNA, the loss of SEPT12 transcripts disrupts α- and β-tubulin organization. In addition, losing or decreasing SEPT12 disturbs the morphogenesis of sperm heads and the elongation of sperm tails, the steps of which are coordinated and constructed by α- and β-tubulins, in SEPTIN12+/+/+/− chimeras. In this study, we discovered that the SEPTIN12-microtubule complexes are critical for sperm formation during spermiogenesis.
spermiogenesis; SEPT12; microtubules
Highly sensitive guaiac-based faecal occult blood (Hemoccult SENSA) and Helicobacter pylori stool antigen testing might help detect upper gastrointestinal lesions when appended to a colorectal cancer screening programme with faecal immunochemical testing. We evaluated the diagnostic accuracies of two stool tests in detecting upper gastrointestinal lesions.
Hospital-based and community-based screening settings.
A hospital-based deviation cohort of 3172 participants to evaluate test performance and a community-based validation cohort of 3621 to verify the findings.
Three types of stool tests with bidirectional endoscopy as the reference standard.
Sensitivity, specificity and positive and negative likelihood ratios.
For detecting upper gastrointestinal lesions in cases with negative immunochemical tests, the sensitivity, specificity, and positive and negative likelihood ratios of the guaiac-based and H pylori antigen tests were 16.3% (95% CI 13.3% to 19.8%), 90.1% (88.9% to 91.2%), 1.64 (1.31 to 2.07), and 0.93 (0.89 to 0.97), respectively, and 52.5% (48.1% to 56.9%), 80.6% (79.0% to 82.1%), 2.71 (2.41 to 3.04) and 0.59 (0.54 to 0.65), respectively. For detecting upper gastrointestinal lesions in cases with normal colonoscopy, the results of the guaiac-based and H pylori antigen tests were 17.9% (14.8% to 21.5%), 90.1% (88.9% to 91.2%), 1.81 (1.45 to 2.26) and 0.91 (0.87 to 0.95), respectively, and 53.1% (48.6% to 57.4%), 80.7% (79.1% to 82.2%), 2.75 (2.45 to 3.08) and 0.58 (0.53 to 0.64), respectively. Within the community, positive predictive values of the immunochemical and H pylori antigen tests were 36.0% (26.0% to 46.0%) and 31.9% (28.3% to 35.5%), respectively, for detecting lower and upper gastrointestinal lesions, which were similar to expected values.
The H pylori stool antigen test is more accurate than the guaiac-based test in the screening of upper gastrointestinal lesions in a population with high prevalence of H pylori infection and upper gastrointestinal lesions. It is applicable to add the H pylori antigen test to the immunochemical test for pan detection.
Scratching triggers skin flares in atopic dermatitis (AD). We demonstrate that scratching of human skin, and tape stripping of mouse skin, causes neutrophil influx. This influx in mice was largely dependent on the generation of leukotriene B4 (LTB4) by neutrophils and their expression of the LTB4 receptor BLT1. Allergic skin inflammation in response to epicutaneous (EC) application of ovalbumin to tape-stripped skin was severely impaired in Ltb4r1−/− mice, and required expression of BLT1 on both T cells and non-T cells. Co-transfer of WT neutrophils, but not neutrophils deficient in BLT1 or the LTB4 synthesizing enzyme LTA4H, restored the ability of WT CD4+ effector T cells to transfer allergic skin inflammation to Ltb4r1−/− recipients. Pharmacologic blockade of LTB4 synthesis inhibited allergic skin inflammation elicited by cutaneous antigen challenge in previously EC-sensitized mice. Our results demonstrate that a neutrophil-T cell axis reliant on LTB4-BLT1 interaction is required for allergic skin inflammation.
Dengue virus is a mosquito-transmitted virus that can cause self-limiting dengue fever, severe life-threatening dengue hemorrhagic fever and dengue shock syndrome. The existence of four serotypes of dengue virus has complicated the development of an effective and safe dengue vaccine. Recently, a clinical phase 2b trial of Sanofi Pasteur's CYD tetravalent dengue vaccine revealed that the vaccine did not confer full protection against dengue-2 virus. New approaches to dengue vaccine development are urgently needed. Our approach represents a promising method of dengue vaccine development and may even complement the deficiencies of the CYD tetravalent dengue vaccine.
Two important components of a vaccine, the immunogen and immunopotentiator, were combined into a single construct to generate a new generation of vaccines. We selected dengue-2 envelope protein domain III (D2ED III) as the immunogen and expressed this protein in lipidated form in Escherichia coli, yielding an immunogen with intrinsic immunopotentiation activity. The formulation containing lipidated D2ED III (LD2ED III) in the absence of exogenous adjuvant elicited higher D2ED III-specific antibody responses than those obtained from its nonlipidated counterpart, D2ED III, and dengue-2 virus. In addition, the avidity and neutralizing capacity of the antibodies induced by LD2ED III were higher than those elicited by D2ED III and dengue-2 virus. Importantly, we showed that after lipidation, the subunit candidate LD2ED III exhibited increased immunogenicity while reducing the potential risk of antibody-dependent enhancement of infection in mice.
Our study suggests that the lipidated subunit vaccine approach could be applied to other serotypes of dengue virus and other pathogens.
Vaccines are considered a cost-effective way to control infectious diseases. To rationally design vaccines, antigens and, frequently, adjuvants must be selected to trigger appropriate immune responses against a specific pathogen. We selected dengue-2 envelope protein domain III as a dengue vaccine candidate and expressed this candidate in the lipidated form in an Escherichia coli-based system. Dengue envelope protein domain III mediates binding of the dengue virus to the host cellular receptor. The lipid moiety of the bacterial-derived lipoprotein can activate the innate immune system to elicit an appropriate adaptive immune response. We demonstrated that lipidated dengue-2 envelope protein domain III is more immunogenic than nonlipidated dengue-2 envelope protein domain III. Most importantly, the lipidated dengue-2 envelope protein domain III alone triggered a durable neutralizing antibody response with a low risk of severe side effects. Lipidated subunit vaccines are non-replicating and thus may be less susceptible to replication interference than live attenuated vaccines. Our study suggests that the lipidated subunit vaccine approach could be applied to other serotypes of dengue virus as well as other pathogens.
attention deficit hyperactivity disorder; bioequivalence; generic drugs; methylphenidate; pAUC
Highly variable (HV) drugs are defined as those for which within-subject variability (%CV) in bioequivalence (BE) measures is 30% or greater. Because of this high variability, studies designed to show whether generic HV drugs are bioequivalent to their corresponding HV reference drugs may need to enroll large numbers of subjects even when the products have no significant mean differences. To avoid unnecessary human testing, the US Food and Drug Administration’s Office of Generic Drugs developed a reference-scaled average bioequivalence (RSABE) approach, whereby the BE acceptance limits are scaled to the variability of the reference product. For an acceptable RSABE study, an HV generic drug product must meet the scaled BE limit and a point estimate constraint. The approach has been implemented successfully. To date, the RSABE approach has supported four full approvals and one tentative approval of HV generic drug products.
bioequivalence; generic drugs; highly variable drugs; reference-scaled average bioequivalence; US Food and Drug Administration
Curcuminoids are well known for their capabilities to combat risk factors that are associated with ageing and cellular senescence. Recent reports have demonstrated that curcuminoids can extend the lifespan of model organisms. However, the underlying mechanisms by which these polyphenic compounds exert these beneficial effects remain unknown. In this study, t-BHP-induced premature senescence model in human fibroblasts was chosen to explore the protective effects of a curcuminoid, bisdemethoxycurcumin (BDMC), on cellular senescence. The results demonstrated that BDMC attenuated oxidative stress-induced senescence-like features which include the induction of an enlarged cellular appearance, higher frequency of senescence-associated β-galactosidase staining activity, appearance of senescence-associated heterochromatic foci in nuclei, decrease in proliferation capability, and alteration in related molecules such as p16 and retinoblastoma protein. Notably, we found that BDMC treatment activated Sirt1/AMPK signaling pathway. Moreover, downregulating Sirt1 by the pharmacological inhibitor nicotianamine or small interfering RNA blocked BDMC-mediated protection against t-BHP-mediated decrease in proliferation. These results suggested that BDMC prevented t-BHP-induced cellular senescence, and BDMC-induced Sirt1 may be a mechanism mediating its beneficial effects.
Phthalates and bisphenol A are environmental endocrine-disrupting chemicals used widely in common consumer products. There is increasing concern about human exposure to phthalates and bisphenol A due to the potential adverse effects related to the anti-androgenic activity of phthalates and estrogenic activity of bisphenol A. In assessing environmental exposure to phthalates and bisphenol A, it is essential to have a validated analytical method that can quantify trace concentrations of phthalate metabolites and bisphenol A in humans. In this study, we developed and validated an accurate, sensitive, and robust LC-MS/MS method to simultaneously quantify 5 phthalate monoester metabolites, including mono-methyl phthalate, mono-ethyl phthalate, mono-butyl phthalate, mono-benzyl phthalate, mono-2-ethylhexyl phthalate, and bisphenol A in human urine. In this method, the phthalate metabolites and bisphenol A, along with their isotope labeled internal standards, were extracted from 200 μl of human urine using automated off-line solid phase extraction. The analytes were quantitatively determined using LC-MS/MS operated in negative electrospray ionization multiple reaction-monitoring mode. The limit of quantification was 0.3 ng/ml for mono-methyl phthalate, mono-ethyl phthalate, mono-benzyl phthalate and bisphenol A, and 1 ng/ml for mono-butyl phthalate and mono-2-ethylhexyl phthalate. The precision and accuracy were well within the acceptable 15% range. This validated method has been used successfully in assessing exposure to phthalates and bisphenol A in humans.
phthalate metabolite; Bisphenol A; SPE; LC-MS/MS
This work represents the first evaluation of the effects of water extract of C. nuda (WE-CN), an edible mushroom, on murine bone marrow-derived dendritic cells (BMDCs) and the potential pathway through which the effects are mediated. Our experimental results show that WE-CN could induce phenotypic maturation of DCs, as shown by the increased expression of MHC and costimulatory molecules. In addition, it also induced the proinflammatory cytokines expression on DCs and enhanced both the proliferation and IFN-γ secretion of allogenic T cells. Therefore, since WE-CN did not induce maturation of DCs generated from mice with mutated TLR-4 or TLR-2, suggesting that TLR4 and TLR2 might function as membrane receptors for WE-CN. Moreover, the mechanism of action of WE-CN may be mediated by increased phosphorylation of ERK, p38, and JNK mitogen-activated protein kinase (MAPK) and increased NF-κB p65 activity, which are important signaling molecules downstream of TLR-4 and TLR-2. Finally, coimmunization of mice with WE-CN and a HER-2/neu DNA vaccine induced a HER-2/neu-specific Th1 response that resulted in significant inhibition of HER-2/neu overexpressing mouse bladder tumor (MBT-2) growth. These data suggest that WE-CN induces DC maturation through TLR-4 and/or TLR-2 and that WE-CN can be used as an adjuvant in cancer vaccine immunotherapy.
Whilst data recognise both myeloid cell accumulation during choroidal neovascularisation (CNV) as well as complement activation, none of the data has presented a clear explanation for the angiogenic drive that promotes pathological angiogenesis. One possibility that is a pre-eminent drive is a specific and early conditioning and activation of the myeloid cell infiltrate. Using a laser-induced CNV murine model, we have identified that disruption of retinal pigment epithelium (RPE) and Bruch’s membrane resulted in an early recruitment of macrophages derived from monocytes and microglia, prior to angiogenesis and contemporaneous with lesional complement activation. Early recruited CD11b+ cells expressed a definitive gene signature of selective inflammatory mediators particularly a pronounced Arg-1 expression. Accumulating macrophages from retina and peripheral blood were activated at the site of injury, displaying enhanced VEGF expression, and notably prior to exaggerated VEGF expression from RPE, or earliest stages of angiogenesis. All of these initial events, including distinct VEGF + Arg-1+ myeloid cells, subsided when CNV was established and at the time RPE-VEGF expression was maximal. Depletion of inflammatory CCR2-positive monocytes confirmed origin of infiltrating monocyte Arg-1 expression, as following depletion Arg-1 signal was lost and CNV suppressed. Furthermore, our in vitro data supported a myeloid cell uptake of damaged RPE or its derivatives as a mechanism generating VEGF + Arg-1+ phenotype in vivo. Our results reveal a potential early driver initiating angiogenesis via myeloid-derived VEGF drive following uptake of damaged RPE and deliver an explanation of why CNV develops during any of the stages of macular degeneration and can be explored further for therapeutic gain.
Cardiovascular growth must balance stabilizing signals required to maintain endothelial connections and network integrity with destabilizing signals that enable individual endothelial cells to migrate and proliferate. The cerebral cavernous malformation (CCM) signaling pathway utilizes the adaptor protein CCM2 to strengthen endothelial cell junctions and stabilize vessels. Here we identify a CCM2 paralogue, CCM2L, that is expressed selectively in endothelial cells during periods of active cardiovascular growth. CCM2L competitively blocks CCM2-mediated stabilizing signals biochemically, in cultured endothelial cells, and in developing mice. Loss of CCM2L reduces endocardial growth factor expression and impairs tumor growth and wound healing. Our studies identify CCM2L as a molecular mechanism by which endothelial cells coordinately regulate vessel stability and growth during cardiovascular development as well as postnatal vessel growth.
Nuclear export is an important process that not only regulates the functions of cellular factors but also facilitates the assembly of viral nucleoprotein complexes. Chromosome region maintenance 1 (CRM1) that mediates the transport of proteins bearing the classical leucine-rich nuclear export signal (NES) is the best-characterized nuclear export receptor. Recently, several CRM1-independent nuclear export pathways were also identified. The nuclear export of the large form of hepatitis delta antigen (HDAg-L), a nucleocapsid protein of hepatitis delta virus (HDV), which contains a CRM1-independent proline-rich NES, is mediated by the host NES-interacting protein (NESI). The mechanism of the NESI protein in mediating nuclear export is still unknown. In this study, NESI was characterized as a highly glycosylated membrane protein. It interacted and colocalized well in the nuclear envelope with lamin A/C and nucleoporins. Importantly, HDAg-L could be coimmunoprecipitated with lamin A/C and nucleoporins. In addition, binding of the cargo HDAg-L to the C terminus of NESI was detected for the wild-type protein but not for the nuclear export-defective HDAg-L carrying a P205A mutation [HDAg-L(P205A)]. Knockdown of lamin A/C effectively reduced the nuclear export of HDAg-L and the assembly of HDV. These data indicate that by forming complexes with lamin A/C and nucleoporins, NESI facilitates the CRM1-independent nuclear export of HDAg-L.
We develop an automated method to determine the foveola location in macular 3D-OCT images in either healthy or pathological conditions. Structural Support Vector Machine (S-SVM) is trained to directly predict the location of the foveola, such that the score at the ground truth position is higher than that at any other position by a margin scaling with the associated localization loss. This S-SVM formulation directly minimizes the empirical risk of localization error, and makes efficient use of all available training data. It deals with the localization problem in a more principled way compared to the conventional binary classifier learning that uses zero-one loss and random sampling of negative examples. A total of 170 scans were collected for the experiment. Our method localized 95.1% of testing scans within the anatomical area of the foveola. Our experimental results show that the proposed method can effectively identify the location of the foveola, facilitating diagnosis around this important landmark.
This study compared the effects of ten types of traditional Chinese medicines (TCMs) and six different antibiotics on E. coli O157:H7 Shiga toxin gene (stx2) mRNA expression level based on real-time PCR and the expression level of Stx toxin using an ELISA quantitative assay. We also compared their effects on the induction of the SOS response. The results clearly indicated that all ten TCMs had negative results in the SOS response induction test, while most TCMs did not increase the levels of stx2 mRNA and the Stx toxin. Some TCMs did increase the mRNA levels of the stx2 gene and the Stx toxin level, but their increases were much lower than those caused by antibiotics. With the exception of cefotaxime, the six antibiotics increased the Stx toxin level and increased the stx2 gene mRNA level. With the exceptions of cefotaxime and tetracycline, the antibiotics increased the SOS induction response. These results suggest that TCMs may have advantages compared with antibiotics, when treating E. coli O157:H7; TCMs did not greatly increase Stx toxin production and release.
Bisphenol A (BPA) is one of the environmental endocrine-disrupting chemicals used widely in common consumer products. There is an increasing concern about human exposure to BPA, particularly in fetuses, due to the potential adverse effects related to the estrogenic activity of BPA. In assessing environmental exposure to BPA, it is essential to have a sensitive, accurate and specific analytical method, particularly for low BPA levels in complex sample matrices. In this study, we developed and validated an accurate, sensitive, and robust liquid chromatography-mass spectrometry (LC-MS) method for determining BPA concentrations in human amniotic fluid. In this method, BPA and the internal standards 13C12-BPA were extracted from 500 μL of human amniotic fluid using solid phase extraction. Calibration curves were linear over a concentration range of 0.3 to 100 ng/mL for BPA. The analytes were quantitatively determined using LC-MS operated in negative electrospray ionization selected ion monitoring mode. This validated method has been used successfully in clinical sample analysis of BPA in second-trimester amniotic fluid specimens.
SPE; LCMS; Quantitation; Bisphenol A; Amniotic fluid
Etiological epidemiology and diagnosis are important issues in adult community-acquired pneumonia (CAP), and identifying pathogens based on patient clinical features is especially a challenge. CAP-associated main pathogens in adults include viruses as well as bacteria. However, large-scale epidemiological investigations of adult viral CAP in China are still lacking. In this study, we analyzed the etiology of adult CAP in Beijing, China and constructed diagnostic models based on combinations of patient clinical factors.
A multicenter cohort was established with 500 adult CAP outpatients enrolled in Beijing between November 2010 to October 2011. Multiplex and quantitative real-time fluorescence PCR were used to detect 15 respiratory viruses and mycoplasma pneumoniae, respectively. Bacteria were detected with culture and enzyme immunoassay of the Streptococcus pneumoniae urinary antigen. Univariate analysis, multivariate analysis, discriminatory analysis and Receiver Operating Characteristic (ROC) curves were used to build predictive models for etiological diagnosis of adult CAP.
Pathogens were detected in 54.2% (271/500) of study patients. Viruses accounted for 36.4% (182/500), mycoplasma pneumoniae for 18.0% (90/500) and bacteria for 14.4% (72/500) of the cases. In 182 of the patients with viruses, 219 virus strains were detected, including 166 single and 53 mixed viral infections. Influenza A virus represented the greatest proportion with 42.0% (92/219) and 9.1% (20/219) in single and mixed viral infections, respectively. Factors selected for the predictive etiological diagnostic model of viral CAP included cough, dyspnea, absence of chest pain and white blood cell count (4.0-10.0) × 109/L, and those of mycoplasma pneumoniae CAP were being younger than 45 years old and the absence of a coexisting disease. However, these models showed low accuracy levels for etiological diagnosis (areas under ROC curve for virus and mycoplasma pneumoniae were both 0.61, P < 0.05).
Greater consideration should be given to viral and mycoplasma pneumoniae infections in adult CAP outpatients. While predictive etiological diagnostic models of viral and mycoplasma pneumoniae based on combinations of demographic and clinical factors may provide indications of etiology, diagnostic confirmation of CAP remains dependent on laboratory pathogen test results.
Community-acquired pneumonia; Etiology; Epidemiology; Diagnosis; Pneumonia; Virus; Polymerase chain reaction; ROC curve
Phase contrast, a noninvasive microscopy imaging technique, is widely used to capture time-lapse images to monitor the behavior of transparent cells without staining or altering them. Due to the optical principle, phase contrast microscopy images contain artifacts such as the halo and shade-off that hinder image segmentation, a critical step in automated microscopy image analysis. Rather than treating phase contrast microscopy images as general natural images and applying generic image processing techniques on them, we propose to study the optical properties of the phase contrast microscope to model its image formation process. The phase contrast imaging system can be approximated by a linear imaging model. Based on this model and input image properties, we formulate a regularized quadratic cost function to restore artifact-free phase contrast images that directly correspond to the specimen's optical path length. With artifacts removed, high quality segmentation can be achieved by simply thresholding the restored images. The imaging model and restoration method are quantitatively evaluated on microscopy image sequences with thousands of cells captured over several days. We also demonstrate that accurate restoration lays the foundation for high performance in cell detection and tracking.
Phase contrast optics; microscopy image analysis; imaging model; image restoration; image segmentation; cell tracking
Preventing diabetic foot problems (DFP) and their associated consequences is a critical in rural regions. The objective is to present an association of non-invasive DFP assessment tools and physiological indicators for early detection among rural cases of diabetes in Taiwan.
Secondary data analysis of 387 participants previously diagnosed with type 2 diabetes was used. The Michigan Neuropathy Screening Instrument (MNSI), Ankle Brachial Index (ABI), optimal scaling combination (OSC) of MNSI, and age were used to examine peripheral neurovascular function. The King’s College classification (KC) and Texas risk classification (TRC) were used to understand diabetic foot complications.
The findings indicated that MNSI was negatively correlated with ABI, but positively with diabetes duration, age, KC, TRC, fasting blood glucose, low density of lipoprotein cholesterol, body mass index and waist circumference. The area under the receiver operating characteristic curves for assessing the risk of ABI based on OSC was larger than for MNSI, KC, and TRC.
It is shown that using OSC, MNSI, and ABI as community screening tools is useful in detecting early neurovasculopathy. In addition, where an ABI machine is unavailable, primary healthcare providers that perform MNSI or OSC may be cost-effective. The study was approved by the institutional review board of the ethical committee (No 98-2224-B).
Diabetic foot problems; Michigan neuropathy screening instrument; Ankle brachial index; Receiver operating characteristic curves