ISOLATION OF EGG DROP SYNDROME VIRUS AND ITS MOLECULAR CHARACTERIZATION USING SODIUM DODECYL SULPHATE POLYACRYLAMIDE GEL ELECTROPHORESIS

Six isolates of egg drop syndrome (EDS) virus were recovered from five different outbreaks of EDS in commercial laying hens in and around Faisalabad. The aberrant eggs were fed to the susceptible laying hens for experimental induction of infection. The samples from infected birds (egg washing, cloacal swabs, oviducts and spleens) were collected, processed and inoculated into 11-day old duck embryos. The presence of virus in harvested allanto-amniotic fluid was monitored by spot and microhaemagglutination tests and confirmed by haemagglutination inhibition and agar gel precipitation tests. The EDS virus grew well in duck embryos and agglutinated only avian but not mammalian red blood cells. These isolates were purified through velocity density gradient centrifugation. Protein concentration was determined through Lowry method and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) was conducted by loading 300 µg protein concentration on 12.5% gel using discontinuous buffer system. All the six isolates showed 13 polypeptides, which were identical to those described in the referral EDS-76 virus (strain-127). The molecular weights of the polypeptides ranged from 6.5 KDa to 126 KDa

MDR1 siRNA loaded hyaluronic acid-based CD44 targeted nanoparticle systems circumvent paclitaxel resistance in ovarian cancer.

Development of multidrug resistance (MDR) is an almost universal phenomenon in patients with ovarian cancer, and this severely limits the ultimate success of chemotherapy in the clinic. Overexpression of the MDR1 gene and corresponding P-glycoprotein (Pgp) is one of the best known MDR mechanisms. MDR1 siRNA based strategies were proposed to circumvent MDR, however, systemic, safe, and effective targeted delivery is still a major challenge. Cluster of differentiation 44 (CD44) targeted hyaluronic acid (HA) based nanoparticle has been shown to successfully deliver chemotherapy agents or siRNAs into tumor cells. The goal of this study is to evaluate the ability of HA-PEI/HA-PEG to deliver MDR1 siRNA and the efficacy of the combination of HA-PEI/HA-PEG/MDR1 siRNA with paclitaxel to suppress growth of ovarian cancer. We observed that HA-PEI/HA-PEG nanoparticles can efficiently deliver MDR1 siRNA into MDR ovarian cancer cells, resulting in down-regulation of MDR1 and Pgp expression. Administration of HA-PEI/HA-PEG/MDR1 siRNA nanoparticles followed by paclitaxel treatment induced a significant inhibitory effect on the tumor growth, decreased Pgp expression and increased apoptosis in MDR ovarian cancer mice model. Our findings suggest that CD44 targeted HA-PEI/HA-PEG/MDR1 siRNA nanoparticles can serve as a therapeutic tool with great potentials to circumvent MDR in ovarian cancer

CIDI-Lung-Seg: A Single-Click Annotation Tool for Automatic Delineation of Lungs from CT Scans

Accurate and fast extraction of lung volumes from computed tomography (CT) scans remains in a great demand in the clinical environment because the available methods fail to provide a generic solution due to wide anatomical variations of lungs and existence of pathologies. Manual annotation, current gold standard, is time consuming and often subject to human bias. On the other hand, current state-of-the-art fully automated lung segmentation methods fail to make their way into the clinical practice due to their inability to efficiently incorporate human input for handling misclassifications and praxis. This paper presents a lung annotation tool for CT images that is interactive, efficient, and robust. The proposed annotation tool produces an "as accurate as possible" initial annotation based on the fuzzy-connectedness image segmentation, followed by efficient manual fixation of the initial extraction if deemed necessary by the practitioner. To provide maximum flexibility to the users, our annotation tool is supported in three major operating systems (Windows, Linux, and the Mac OS X). The quantitative results comparing our free software with commercially available lung segmentation tools show higher degree of consistency and precision of our software with a considerable potential to enhance the performance of routine clinical tasks.Comment: 4 pages, 6 figures; to appear in the proceedings of 36th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC 2014

PREPARATION OF CONJUGATE FOR USE IN AN ELISA FOR HUMORAL IMMUNE RESPONSE AGAINST EGG DROP SYNDROME VIRUS IN LAYER CHICKS

An indirect enzyme-linked immunosorbent assay (ELISA) was performed for the detection of antibodies against Egg Drop Syndrome (EDS) virus. Virus identification was done through haemaggluti- nation inhibition (HI) test using known antisera. Antichicken immunoglobulins were raised in goats and purified by ammonium sulphate precipitation technique. These goat-antichicken immunoglobulins were conjugated with horseradish peroxidase. Twenty-seven serum samples were collected from a layers flock vaccinated against EDS and specific antibodies were determined by using a horseradish conjugate

TBX2 represses PTEN in rhabdomyosarcoma and skeletal muscle.

Rhabdomyosarcoma (RMS) is the most frequent soft tissue sarcoma in children that shares many features of developing skeletal muscle. TBX2, a T-box family member, is highly upregulated in tumor cells of both major RMS subtypes where it functions as an oncogene. TBX2 is a repressor that is often overexpressed in cancer cells and functions in bypassing cell growth control, including the repression of the cell cycle regulators p14 and p21. We have found that TBX2 directly represses the tumor-suppressor phosphatase and tensin homolog (PTEN) in both RMS and normal muscle. Exogenous expression of TBX2 in normal muscle cells downregulates PTEN, and depletion or interference with TBX2 in RMS cells upregulates PTEN. Human RMS tumors show high levels of TBX2 and correspondingly low levels of PTEN. The expression of PTEN in clinical RMS samples is relatively uncharacterized, and we establish that suppression of PTEN is a frequent event in both subtypes of RMS. TBX2 represses PTEN by directly binding to the promoter and recruiting the histone deacetylase, HDAC1. RMS cells have high levels of activated AKT owing to the deregulation of phosphoinositide-3 kinase (PI3K) signaling, and depletion or interference with TBX2, which upregulates PTEN, results in a reduction of phospho-AKT. We have also found that the highly related T-box family member TBX3 does not repress PTEN in the muscle lineage. This work suggests that TBX2 is a central component of the PTEN/PI3K/AKT signaling pathway deregulation in RMS cells and that targeting TBX2 in RMS tumors may offer a novel therapeutic approach for RMS.Oncogene advance online publication, 21 December 2015; doi:10.1038/onc.2015.486

The response of sugarcane (Saccharum officinarum L) genotypes to callus induction, regeneration and different concentrations of the selective agent (geneticin -418)

Two commercial cultivars (CPF-245 and CPF-237) and three advanced lines (CSSG-668, S-2003US633, S-2003US114) of sugarcane (Saccharum officinarium) grown in Punjab, Pakistan were evaluated for their potential to induce callus, embryogenic callus and regeneration. Cultivar CSSG-668 was found to be the best genotype yielding maximum embryogenic callus and regeneration whereas cultivar CPF- 245 exhibited lowest callus induction frequency. Five different concentrations (0, 20, 40, 60, and 80mg/L) of the selective agent (geneticin-418) were used to optimize selection conditions with nontransformed embryogenic calli. The geneticin concentration 60 mg/L was found to be the optimal dose to select the embryogenic calli of genotypes CSSG-668, CPF-245 and S-2003US63, while 35 mg/L geneticin was found to be the best concentration for S-2003US-114. Similarly, 60 mg/L geneticin was optimum dose to select regenerated plantlets of the cultivars CSSG-668 and CPF-245 while it was 40, 25 mg/L for the cultivars S-2003US-114 and S-2003US-633, respectively. It is concluded from the present study that geneticin concentration in the range of 25 to 60 mg/L can be effectively used for the selection of transformed embryogenic calli and regenerants of different sugarcane cultivars.Keywords: Callus induction, embryogenic callus, regeneration, Saccharum officinarum L., selection, geneticin

Physicochemical and functional properties of cassava flour grown in different locations in Sabah, Malaysia

The tuber of cassava is used as raw materials in the bakery, food, pharmaceutical and garment industries. The nutritional value of cassava roots is important because they are the main part of the plant consumed in developing countries. However, there is much variation in the nutrient quality of the cassava root depends on the several factors, such as geographic location, variety, age of the plant, and environmental conditions. This study was performed to compare and provide information on physicochemical and functional properties of cassava flour planted in two different districts in Sabah, Malaysia, namely Tawau and Semporna. Proximate analysis showed significant differences (p<0.05) in crude protein (2.07 and 2.69%), crude fat (0.55 and 0.68%) and dietary fibre contents (2.38 and 2.09%). Determinations on physicochemical and functional characteristics of the cassava flour showed significant differences (p<0.05) in bulk density (0.57 and 0.79 g/ cm3 ), pH (6.75 and 6.72), colour and foam capacity (3.66 and 7.33%) while there was no significant difference shown in water and oil absorption capacities as well as emulsion capacity. Cassava planted in Semporna was observed to have high values of all pasting property parameters relative to the one planted in Tawau except for the setback viscosity. Gelatinization properties of flours showed significant differences (p<0.05) in onset (70.59 and 68.99°C) and end temperatures (79.81 and 80.03°C)

\u3cem\u3eMDR1\u3c/em\u3e siRNA Loaded Hyaluronic Acid-Based CD44 Targeted Nanoparticle Systems Circumvent Paclitaxel Resistance in Ovarian Cancer

Development of multidrug resistance (MDR) is an almost universal phenomenon in patients with ovarian cancer, and this severely limits the ultimate success of chemotherapy in the clinic. Overexpression of the MDR1 gene and corresponding P-glycoprotein (Pgp) is one of the best known MDR mechanisms. MDR1 siRNA based strategies were proposed to circumvent MDR, however, systemic, safe, and effective targeted delivery is still a major challenge. Cluster of differentiation 44 (CD44) targeted hyaluronic acid (HA) based nanoparticle has been shown to successfully deliver chemotherapy agents or siRNAs into tumor cells. The goal of this study is to evaluate the ability of HA-PEI/HA-PEG to deliver MDR1 siRNA and the efficacy of the combination of HA-PEI/HA-PEG/MDR1 siRNA with paclitaxel to suppress growth of ovarian cancer. We observed that HA-PEI/HA-PEG nanoparticles can efficiently deliver MDR1 siRNA into MDR ovarian cancer cells, resulting in down-regulation of MDR1 and Pgp expression. Administration of HA-PEI/HA-PEG/MDR1 siRNA nanoparticles followed by paclitaxel treatment induced a significant inhibitory effect on the tumor growth, decreased Pgp expression and increased apoptosis in MDR ovarian cancer mice model. Our findings suggest that CD44 targeted HA-PEI/HA-PEG/MDR1 siRNA nanoparticles can serve as a therapeutic tool with great potentials to circumvent MDR in ovarian cancer

Phase II study of TP300 in patients with advanced gastric or gastro-oesophageal junction adenocarcinoma

Background TP300, a recently developed synthetic camptothecin analogue, is a highly selective topoisomerase I inhibitor. A phase I study showed good safety and tolerability. As camptothecins have proven active in oesophago-gastric adenocarcinomas, in this phase II study we assessed the efficacy and safety of TP300 in patients with gastric or gastro-oesophageal junction (GOJ) adenocarcinomas. Methods Eligible patients had metastatic or locally advanced gastric or Siewert Types II or III GOJ inoperable adenocarcinoma. Patients were chemotherapy naïve unless this had been administered in the perioperative setting. TP300 was administered as a 1-h intravenous infusion every 3 weeks (a cycle) for up to 6 cycles at a starting dose of 8 mg/m2 with intra-patient escalation to 10 mg/m2 from cycle 2 in the absence of dose-limiting toxicity. Tumour responses (RECIST 1.1) were assessed every 6 weeks. Toxicity was recorded by NCI-CTCAE version 3.0. Using a modified two-stage Simon design (Stage I and II), a total of 43 patients were to be included providing there were 3 of 18 patients with objective response in Stage I of the study. Results In Stage I of the study 20 patients (14 males, 6 females), median age 67 years (range 40 − 82), performance status ECOG 0/1, with GC [14] or GOJ carcinoma [6] were enrolled. Of the 16 evaluable patients, 11 received the planned dose increase to 10 mg/m2 at cycle 2, 2 decreased to 6 mg/m2, and 3 continued on 8 mg/m2. There were no objective responses after 2 cycles of treatment. Twelve patients had stable disease for 1 − 5 months and 4 had progressive disease. Median progression free survival (PFS) was 4.1 months (CI [1.6 − 4.9]), median time to progression (TTP) was 2.9 months (CI [1.4 − 4.2]). Grade 3/4 toxicities (worst grade all cycles) included 7 patients (35 %) with neutropenia, 4 patients (20 %) with anaemia, 2 patients (10 %) with thrombocytopenia, and 3 patients (15 %) with fatigue. This study was terminated at the end of Stage I due to a lack of the required (3/18) responders. Conclusions This study of TP300 showed good drug tolerability but it failed to demonstrate sufficient efficacy as measured by radiological response. Trial registration EU-CTR 2009-012097-12 2009-09-0