TACO produces robust multisample transcriptome assemblies from RNA-seq.

Accurate transcript structure and abundance inference from RNA sequencing (RNA-seq) data is foundational for molecular discovery. Here we present TACO, a computational method to reconstruct a consensus transcriptome from multiple RNA-seq data sets. TACO employs novel change-point detection to demarcate transcript start and end sites, leading to improved reconstruction accuracy compared with other tools in its class. The tool is available at http://tacorna.github.io and can be readily incorporated into RNA-seq analysis workflows.

Activation of autophagic programmed cell death and innate immune gene expression reveals immuno-competence of integumental epithelium in Bombyx mori infected by a dipteran parasitoid.

In insects, the integument forms the primary barrier between the environment and internal milieu, but cellular and immune responses of the integumental epithelium to infection by micro- and macro-parasites are mostly unknown. We elucidated cellular and immune responses of the epithelium induced through infection by a dipteran endoparasitoid, Exorista bombycis in the economically important silkworm Bombyx mori. Degradative autophagic vacuoles, lamella-like bodies, a network of cytoplasmic channels with cellular cargo, and an RER network that opened to vacuoles were observed sequentially with increase in age after infection. This temporal sequence culminated in apoptosis, accompanied by the upregulation of the caspase gene and fragmentation of DNA. The infection significantly enhanced the tyrosine level and phenol oxidase activity in the integument. Proteomic analysis revealed enhanced expression of innate immunity components of toll and melanization pathways, cytokines, signaling molecules, chaperones, and proteolytic enzymes demonstrating diverse host responses. qPCR analysis revealed the upregulation of spatzle, BmToll, and NF kappa B transcription factors Dorsal and BmRel. NF kappa B inhibitor cactus showed diminished expression when Dorsal and BmRel were upregulated, revealing a negative correlation (R = (-)0.612). During melanization, prophenol oxidase 2 was expressed, a novel finding in integumental epithelium. The integument showed a low level of melanin metabolism and localized melanism in order to prevent the spreading of cytotoxic quinones. The gene-encoding proteolytic enzyme, beta-N-acetylglucosaminidase, was activated at 24 h post-infection, whereas chitinase, was activated at 96 h post-infection; however, most of the immune genes enhanced their expression in the early stages of infection. Thus the integument contributes to humoral immune responses that enhance resistance against macroparasite invasion.

Rheumatoid nodule presenting as Morton's neuroma.

Among 101 feet that presented with symptoms and signs similar to Morton's neuroma, intermetatarsal rheumatoid nodules were found in five feet (three patients). Two patients had bilateral involvement. Histology of the excised tissue showed the presence of a rheumatoid nodule and Morton's neuroma in four feet and a rheumatoid nodule with unremarkable nerve bundles in one. A rheumatoid nodule can coexist with Morton's neuroma, as seen in our patients, and the presentation is often similar to that of a Morton's neuroma. Our patients were rendered asymptomatic with surgical treatment and went on to have appropriate management of rheumatoid arthritis. Rheumatoid nodule should be considered in the differential diagnosis of Morton's neuroma in not only rheumatoid arthritis patients but also asymptomatic patients who have never been tested for rheumatoid antibodies.

Lung Diseases Detection Using Various Deep Learning Algorithms.

The primary objective of this proposed framework work is to detect and classify various lung diseases such as pneumonia, tuberculosis, and lung cancer from standard X-ray images and Computerized Tomography (CT) scan images with the help of volume datasets. We implemented three deep learning models namely Sequential, Functional & Transfer models and trained them on open-source training datasets. To augment the patient's treatment, deep learning techniques are promising and successful domains that extend the machine learning domain where CNNs are trained to extract features and offers great potential from datasets of images in biomedical application. Our primary aim is to validate our models as a new direction to address the problem on the datasets and then to compare their performance with other existing models. Our models were able to reach higher levels of accuracy for possible solutions and provide effectiveness to humankind for faster detection of diseases and serve as best performing models. The conventional networks have poor performance for tilted, rotated, and other abnormal orientation and have poor learning framework. The results demonstrated that the proposed framework with a sequential model outperforms other existing methods in terms of an F1 score of 98.55%, accuracy of 98.43%, recall of 96.33% for pneumonia and for tuberculosis F1 score of 97.99%, accuracy of 99.4%, and recall of 98.88%. In addition, the functional model for cancer outperformed with an accuracy of 99.9% and specificity of 99.89% and paves way to less number of trained parameters, leading to less computational overhead and less expensive than existing pretrained models. In our work, we implemented a state-of-the art CNN with various models to classify lung diseases accurately.

Weil osteotomy: assessment of medium term results and predictive factors in recurrent metatarsalgia.

BACKGROUND: The angle of the Weil osteotomy is usually referenced relative to the floor irrespective of the plantar angulation of the metatarsal. This study aims to analyse the long term results following the Weil osteotomy and identify the cause of poor outcome. METHODS: This study presents a retrospective review of 61 patients (86 feet), with mean follow-up of 31 months. Each patient underwent clinical, pedobarographic and radiological examination. The radiographs obtained included 'Metatarsal Skyline Views' (MSV), to assess the plantar declination of the metatarsal heads following the osteotomy. The functional scoring was performed using AOFAS and Foot Function Index. RESULTS: Fifty-five patients (80 feet) showed good to excellent results clinically. Six patients had persistent metatarsalgia. All these 6 patients had callosities beneath metatarsal heads. Pedobarography showed peak pressures in the same distribution as callosities and the MSV showed increased plantar declination of the metatarsal heads. This correlation was found to be significant (p<0.05). CONCLUSION: The Weil osteotomy is a safe and effective treatment for metatarsalgia. An MSV radiograph is helpful to identify the plantar prominence of metatarsal which can be associated with poor clinical outcomes.

Thromboprophylaxis in elective foot and ankle patients--current practice in the United Kingdom.

BACKGROUND: The incidence of venous thromboembolism (VTE) is unknown in elective foot and ankle surgery. The National Institute for Health and Clinical Excellence (NICE) recently published guidelines on reducing the risk of venous thromboembolism in surgical patients. This includes patients undergoing elective foot and ankle surgery. METHOD: In March 2010 we surveyed the current practice in VTE prophylaxis in elective foot and ankle surgery amongst members of the British Orthopaedic Foot and Ankle Society (BOFAS). RESULTS: The response rate was 84 (53%). The total number of elective foot and ankle operations performed by the surveyed group was 33,500 per annum. The estimated incidence of DVT, PE and fatal PE was 0.6%, 0.1% and 0.02%. In our study the number of patients needed to treat to prevent a single fatal PE is 10,000 although this figure is open to important bias. CONCLUSION: We question the applicability of the NICE guidelines to patients undergoing elective foot and ankle surgery. We consider that this data justifies the prospective study of the incidence of VTE in patients undergoing elective foot and ankle surgery, without the use of chemical thromboprophylaxis.

Crystallization kinetics and phase transformation in superionic lithium metaphosphate (Li(2)O-P(2)O(5)) glass system.

The ionic conductivity of mol% 50Li(2)O-50P(2)O(5) melt quenched glass shows an anomalous increase after its glass transition temperature (T(g)) around 590 K. On further increasing the temperature gradually, the conductivity decreases owing to the devitrification of Li(2)O-P(2)O(5) glass. The evolution of devitrified crystallites was evidenced by XRD patterns. To understand the devitrification process, isothermal and non-isothermal DSC studies have been carried out on mol% 50Li(2)O-50P(2)O(5) glass. T(g) as well as T(c) values are found to increase monotonically with increasing heating rates. Variation of T(g) as a function of heating rates has been investigated to evaluate the lower limiting temperature of T(g) and the activation energy for structural relaxation. Results of the DSC studies indicate (i) single-stage bulk crystallization of the glass, with DSC traces exhibiting a single [Formula: see text] transition, (ii) an order parameter (Avrami constant) of 2.8 ± 0.1, suggesting internal (bulk) crystallization of the glass, (iii) an activation energy for crystallization equal to 121.7 kJ mol(-1) and (iv) the activation energy for structural relaxation, E(g), to be 558.8 kJ mol(-1). The crystallization mechanism is closely associated with the JMA model and the experimental dataset have been fitted to a non-isothermal Avrami expression and the obtained parameters confirm the experimental results.

Relationship between Dislocation Density and Antibacterial Activity of Cryo-Rolled and Cold-Rolled Copper.

In the present work, cold rolling and cryo-rolling were performed on 99% commercially pure copper substrates. Both cold and cryo-rolling processes caused severe plastic deformation that led to an increase in dislocation density by 14× and 28× respectively, as compared to the pristine material. Increases in average tensile strengths, by 75% (488 MPa) and 150% (698 MPa), were observed in the two rolled materials as the result of the enhancement in dislocation density. In addition to strength, enhanced antibacterial property of cryo-rolled copper was observed in comparison to cold rolled and pristine copper. Initial adhesion and subsequent proliferation of bio-film forming Gram-positive bacteria Staphylococcus aureus was reduced by 66% and 100% respectively for cryo-rolled copper. Approximately 55% protein leakage, as well as ethidium bromide (EtBr) uptake, were observed confirming rupture of cell membrane of S. aureus. Inductively coupled plasma-mass spectroscopy reveals higher leaching of elemental copper in nutrient broth media from the cryo-rolled copper. Detailed investigations showed that increased dislocation led to leaching of copper ions that caused damage to the bacterial cell wall and consequently killing of bacterial cells. Cryo-rolling enhanced both strength, as well as antibacterial activity, due to the presence of dislocations.

Indications for and technique of first metatarsophalangeal joint arthroscopy.

BACKGROUND: Arthroscopy of the great toe metatarsophalangeal joint has been used for a variety of indications, ranging from synovitis to osteochondral defects. The purpose of the present study was to define the indications for arthroscopy, assess its efficacy, and demonstrate the limitations of this procedure. METHODS: Hallux metatarsophalangeal joint arthroscopy was used in 20 patients (25 feet). Indications included degenerative disease with early osteophytosis, chondromalacia, osteochondral defects, loose bodies, arthrofibrosis, synovitis, gouty arthritis, first metatarsophalangeal joint pain with no obvious findings clinically and radiographically in young adults, and intra-articular fracture of the first metatarsophalangeal joint. All patients had a minimal followup of 2 years and were evaluated clinically and radiographically. RESULTS: Arthroscopic surgery resulted in pain free first metatarsophalangeal joints in 95% (19 of 20 patients). Patients with degenerative disease had a pain-free joint for a minimum of 2 years. The patients with gouty arthritis and intra-articular fracture had good functional outcomes. Arthroscopy also helped in identifying the pathology in painful joints with no obvious radiographic features that included conditions such as 'meniscoid' and other impingement lesions. CONCLUSION: Arthroscopy of the first metatarsophalangeal joint is not suitable for patients with extensive degenerative changes and large osteophytes and those that require cheilectomy or arthrodesis. Arthroscopic management of certain painful hallucal metatarsophalangeal joints is a specialized technique, which if performed for the right indications, gives a favorable outcome with minimal complications.

The induction of cytotoxic T cells and tumor regression by soluble antigen formulation.

CTLs specific for tumor antigens play a major role in the immunity against cancer. We have shown that class I-restricted CTLs can be induced by injecting soluble antigens mixed in an antigen formulation (AF) that consists of squalane, Tween 80, and Pluronic L121 (S. Raychaudhuri et al., Proc. Natl. Acad. Sci. USA, 89: 8308-8312, 1992). In this study, using ovalbumin and the ovalbumin-expressing transfectoma (EG7) as a tumor model system, we examined the in vivo antitumor effect of antigen-AF mixture. Vaccination of mice with ovalbumin in AF 2 or 3 days after EG7 tumor challenge showed significant inhibition of tumor growth compared to mice vaccinated with ovalbumin in alum or in saline. Depletion of CD8+ cells at the time of immunization completely abrogated the AF-induced tumor protection, indicating that CD8+ T cells are the major effectors in tumor protection in vivo. Depletion of CD4+ cells led to a marginal loss of tumor protection, which may be the result of inhibition of ovalbumin-specific CTL response due to the lack of T-helper activity. Our results demonstrate that AF can be used in subunit vaccines to stimulate CTLs and tumor regression in vivo.

The landscape of long noncoding RNAs in the human transcriptome.

Long noncoding RNAs (lncRNAs) are emerging as important regulators of tissue physiology and disease processes including cancer. To delineate genome-wide lncRNA expression, we curated 7,256 RNA sequencing (RNA-seq) libraries from tumors, normal tissues and cell lines comprising over 43 Tb of sequence from 25 independent studies. We applied ab initio assembly methodology to this data set, yielding a consensus human transcriptome of 91,013 expressed genes. Over 68% (58,648) of genes were classified as lncRNAs, of which 79% were previously unannotated. About 1% (597) of the lncRNAs harbored ultraconserved elements, and 7% (3,900) overlapped disease-associated SNPs. To prioritize lineage-specific, disease-associated lncRNA expression, we employed non-parametric differential expression testing and nominated 7,942 lineage- or cancer-associated lncRNA genes. The lncRNA landscape characterized here may shed light on normal biology and cancer pathogenesis and may be valuable for future biomarker development.

Steroids in idiopathic pulmonary fibrosis: a prospective assessment of adverse reactions, response to therapy, and survival.

PURPOSE: We evaluated the risk and potential benefit of high-dose corticosteroid therapy in patients with idiopathic pulmonary fibrosis. SUBJECTS AND METHODS: We prospectively studied 41 patients with previously untreated, biopsy-proven idiopathic pulmonary fibrosis. Before treatment, we calculated clinical, radiographic, and physiologic severity-of-illness scores for each patient. We scored high-resolution computerized tomographic (CT) scans for ground glass and interstitial opacity. We determined the extent of cellular infiltration, interstitial fibrosis, desquamation, and granulation in open lung biopsy samples. Patients were monitored monthly for steroid-related side effects, response to therapy at 3 months, and mortality. RESULTS: All patients experienced at least one steroid-induced side effect. Eleven (27%) patients were nonresponders, 11 (27%) were responders, and 19 (46%) remained stable. Of the 19 patients who died during a mean (+/- SD) follow-up of 3.3 +/- 2.3 years, 8 (42%) lost weight during the initial 3 months of steroid therapy; only 3 (14%) of the 22 patients still living (P = 0.08) experienced weight loss. In a multivariate analysis, greater fibrosis (hazard ratio [HR] = 1.4 per unit increase; 95% confidence interval [CI]: 1.0 to 1.9; P = 0.03) and cellularity (RR = 1.9 per unit increase; 95% CI: 1.3 to 2.8; 3, P <0.001) in the biopsy sample and whether a patient was classified as a responder (RR = 0.4 versus nonresponder; 95% CI: 0.2 to 1.0; P = 0.05) or stable (RR = 0.2 versus nonresponder; 95% CI: 0.1 to 0.6, P <0.001) after steroid therapy were associated with mortality. CONCLUSION: Corticosteroid treatment for idiopathic pulmonary fibrosis is associated with substantial morbidity. Patients who remain stable or respond to corticosteroid therapy have better survival than those who fail to respond. Whether this difference reflects an effect of treatment or less severe disease can be determined only in a randomized trial.

Induction of neutralizing antibodies to transmissible gastroenteritis virus by anti-idiotypic antibodies.

The potential of anti-idiotypic antibodies (anti-ids) as immunogens against transmissible gastroenteritis virus (TGEV) was tested in a heterologous system. A month-old pig was immunized with a neutralizing murine monoclonal antibody (MAb, 5A5) of the IgG2a isotype, specific for the E2 glycoprotein of TGEV. The anti-ids were isolated from the serum of the immunized pig by affinity chromatography, initially on a 5A5-Sepharose column, followed by repeated adsorption on a mouse IgG2a column. The swine anti-ids thus obtained bound to the MAb 5A5 (the idiotype), but not to MAbs of the same isotype IgG2a but of different idiotypes. The anti-ids also inhibited the binding of 5A5 to TGEV in a concentration-dependent manner. Mice immunized with the anti-ids produced antibodies to TGEV. These antibodies, neutralized TGEV in vitro and inhibited the binding of 5A5 to TGEV.

Analysis of B cell repertoire specific to the neutralizing epitopes of glycoprotein 120 in HIV-infected individuals.

This study describes the clonotypic analysis of neutralizing anti-gp120 antibodies elicited in HIV-infected individuals by a panel of anti-idiotype monoclonal antibodies (anti-Id MAbs). Sera from 80 HIV-infected individuals at various clinical stages of HIV-infection were tested for reactivity to 19 anti-Id MAbs in ELISA. Anti-idiotype MAbs reacted with between 0 and 26% of sera. Among the 13 idiotypes specific for anti-CD4 site antibodies, 4 were expressed in 15 to 20% of individuals, whereas 2 of 4 idiotypes specific for anti-V3 antibodies were expressed in 15 to 26% of the cases. These data suggest that each HIV-infected individuals has a diverse B cell repertoire to a given neutralizing epitope cluster and that certain clonotypes are more prevalent than others. To correlate the binding activity in ELISA with anti-gp120 specificity, the idiotype-positive antibodies (Id+ Abs) from representative serum samples were isolated by anti-Id MAb-Sepharose affinity columns. In most cases, the epitope specificity and the neutralizing properties of the isolated Id+ Abs correlated with that of anti-gp120 antibodies used for the generation of anti-Id MAbs. We propose that these anti-Id MAbs may be used to identify and measure neutralizing anti-gp120 antibodies of defined specificity in the sera of HIV-infected individuals, HIV-vaccinated individuals, and in HIV-infected mother-infant pairs.

Tumor regression in mice following vaccination with human papillomavirus E7 recombinant protein in PROVAX.

Induction of CD8+ cytotoxic T lymphocytes (CTLs) specific for human papillomavirus (HPV) antigens provides an attractive strategy for immunotherapy of HPV-related cancers in humans. In this study, we investigated the potential of utilizing soluble E7 protein of HPV 16 in an adjuvant formulation, PROVAX as a vaccine against a progressively growing E7 transfected K1735-X21 (H-2k) metastatic melanoma cells (HOPE2) in a mouse model. Vaccination of HOPE2 tumor bearing mice (C3H) with E7 protein in PROVAX resulted in significant inhibition of tumor growth, compared to mice vaccinated with E7 in Alum or saline. In vivo depletion of CD8+ or CD4+ cells indicated that CD8+ cells are the major effector cells in mediating the anti-tumor activity in this model. Furthermore, E7-specific CTL activity in vitro was detected in tumor bearing mice vaccinated with E7-PROVAX. Our studies suggest that recombinant HPV antigens in combination with PROVAX could serve as an effective subunit vaccine to stimulate tumor specific CD8+ T cell mediated immunity against HPV-related cancers.

A humanized anti-human CD154 monoclonal antibody blocks CD154-CD40 mediated human B cell activation.

Humanized anti-CD154 antibody, IDEC-131, had a slightly, but reproducibly, better binding affinity for CD154 (Kd = 5.6 nM), compared to the parent antibody 24-31 (Kd = 8.5 nM). Otherwise it was indistinguishable from the murine parent antibody in its ability to bind to CD154, block CD154 binding to CD40 and inhibit T cell-dependent B cell differentiation. The latter activity was independent of FcR binding as the Fab'1 fragment of IDEC-131 had an equivalent biological activity to that of the whole antibody. IDEC-131 blocked soluble CD154 from inducing proliferation of purified B cells, and blocked T cell dependent anti-tetanus toxoid specific antibody production by human B cells in vitro. IDEC-131, gamma1, kappa, had strong Fc gammaRI, Fc gammaRII and C1q binding, but was unable to induce complement dependent (CDC) or antibody dependent cell-cytotoxicity (ADCC) of activated peripheral blood T cells, which express relatively low levels of CD154. IDEC-131 antibody inhibited both primary and secondary antibody responses to ovalbumin in cynomolgus monkeys at a dose of 5 mg/kg. In non-immunized animals, treatment with IDEC-131 at 50 mg/kg weekly for 13 weeks induced no change in any of the measured lymphocyte subsets, including B cells, CD4+ and CD8+ T cells. Similarly, a safety study in chimpanzees showed no discernible safety related issues at 20 mg/kg, including B and T cell subsets. These results show that the humanized anti-CD154 antibody, IDEC-131, has retained the affinity and functional activity of its murine parent antibody, is unlikely to deplete CD154 positive lymphocytes in humans, and is safe and effective in blocking antibody production in monkeys. Based on its safety and efficacy profile, IDEC-131 is being developed for therapy of systemic lupus erythematosus.

Chimeric anti-CD20 (IDEC-C2B8) monoclonal antibody sensitizes a B cell lymphoma cell line to cell killing by cytotoxic drugs.

More than 50% of patients with aggressive B lymphomas and the majority of patients with low grade lymphomas are not cured by current therapeutic strategies. The lymphomas express the B cell antigen CD20 on the cell surface and this antigen serves as target for antibody-directed therapies. Clinical studies with encouraging results have been underway with the use of a chimeric anti-CD20 antibody (IDEC-C2B8), consisting of human IgG1-6 constant regions and variable regions from the murine monoclonal anti-CD20 antibody IDEC-2B8. This study investigated the potential anti-tumor therapeutic value of combination treatment with anti-C2B8 and cytotoxic drugs. The in vitro study examined the sensitizing effect of C2B8 antibody on the DHL-4 B lymphoma line to various cytotoxic agents. Cytotoxicity was determined by the MTT assay. Surface and cytoplasmic proteins were determined by flow cytometry. Pretreatment of DHL-4 with C2B8 resulted in inhibition of cell proliferation and cell death and a fraction of the cells underwent apoptosis. While the DHL-4 tumor cells were relatively resistant to several cytotoxic drugs, pretreatment with C2B8 rendered the cells sensitive to TNF-alpha, ricin, diphtheria toxin (DTX), adriamycin and cisplatin but not to VP-16. Chemosensitization of DHL-4 tumor cells was not due to downmodulation of either the MDR-1 or bcl-2 gene products. However, treatment of DHL-4 with C2B8 inhibited TNF-alpha secretion. These findings demonstrate that C2B8 antibody potentiates the sensitivity of DHL-4 tumor cells to several cytotoxic agents. Further, the findings suggest that combination treatments with C2B8 antibody and drugs may be of clinical benefit in the treatment of patients with resistant aggressive B lymphomas.

Quantitation of bovine immunoglobulins in culture fluids by use of sandwich radioimmunoassay with monoclonal antibodies.

Bovine immunoglobulin isotype-specific murine monoclonal antibodies were used in sandwich radioimmunoassays to detect and quantitate bovine IgG1, IgG2, IgM, and IgA in culture fluids. The concentrations of bovine immunoglobulins in unknown samples were extrapolated from standard curves generated with bovine monoclonal immunoglobulins. The lowest detection limits for the bovine immunoglobulin isotypes ranged from 65 to 270 ng/ml.

Influence of partial replacement of butter fat with peanut oil (in infant formula) on erythrocyte fatty acids in infants.

Erythrocyte fatty acid composition was studied in infants fed with three different formulae: formula I containing 20% butter fat; formula II containing 10% butter fat and 10% peanut oil; and formula III containing 10% butter fat and 5% peanut oil with a fat content itself reduced to 15%. The linoleic acid levels were 2.5, 18 and 13% in formula I-III, respectively. Analysis of fatty acids at the time of birth, and 3 and 6 months thereafter, indicated that linoleic acid levels could be improved by supplementation with peanut oil. Arachidonic acid levels (20:4, n-6) did not show a proportional relationship with respect to linoleic acid intake. The other ratio such as triene/tetraene, oleic/linoleic, linoleic/arachidonic and arachidonic/linoleic were all within the normal range, indicating normal desaturase and elongase activity. Thus, our present study suggests that peanut oils could be used for enhancing the linoleic acid levels in infants.

The 100 most cited papers in foot and ankle surgery.

BACKGROUND: The number of citations of a paper gives an indication of an article's merit and importance within a medical specialty. We identify and analyse the 100 most cited papers in foot and ankle surgery. METHOD: The Science Citation Index Expanded was searched for citations in 15 respected journals containing foot and ankle articles. Papers were analysed for subject, authorship, institution, country and year of publication. The average yearly citation was compared to total number of citations. RESULTS: 3501 foot and ankle papers were returned. The maximum number of citations was 1084 and the mean was 104. The top 100 papers were published between 1979 and 2007, with the majority published in the last decade. The ankle was the most important anatomical region discussed, and basic science and degenerative disease were popular topics. We found a large discrepancy between the total number of citations with average yearly citation. CONCLUSION: Foot and ankle surgery is a young and rapidly developing sub-specialty within orthopaedics. Recently there has been a significant increase in influential papers published. Certain topics are popular indicating their importance within the field. This study highlights important papers in foot and ankle surgery giving an insight into readership.