To date, generation of single-chain fragment variable (scFv) has become an established technique used to produce a completely functional antigen-binding fragment in bacterial systems. The advances in antibody engineering have now facilitated a more efficient and generally applicable method to produce Fv fragments. Basically, scFv antibodies produced from phage display can be genetically fused to the marker proteins, such as fluorescent proteins or alkaline phosphatase. These bifunctional proteins having both antigen-binding capacity and marker activity can be obtained from transformed bacteria and used for one-step immunodetection of biological agents. Alternatively, antibody fragments could also be applied in the construction of immunotoxins, therapeutic gene delivery, and anticancer intrabodies for therapeutic purposes. This paper provides an overview of the current studies on the principle, generation, and application of scFv. The potential of scFv in breast cancer research is also discussed in this paper.
The present study was to determine the anticancer potential of Labisia pumila in in vitro models. Results from the study revealed that ethanol extract of L. pumila was more cytotoxic against HM3KO cells while having reduced effects on nonmalignant cells as compared to aqueous and hexane extracts. Thus, ethanol extract was selected to be further separated by using the bioassay-guided fractionation method to give an active fraction, SF2Lp. Results obtained from the flow cytometry analysis showed that SF2Lp was able to arrest the HM3KO cell cycle at the G1 phase, while morphological findings from AO-EB nuclear staining assays along with the Apoptotic Index confirmed the induction of apoptosis by SF2Lp in HM3KO cells. Results from the mechanistic study further revealed that SF2Lp treatment was able to concurrently increase the expression level of p53 and pro-apoptotic protein Bax and also reduce the expression level of anti-apoptotic protein BCl-2 in HM3KO cells, directly contributing to the increase in Bax/Bcl-2 ratio. These findings, therefore, suggested that L. pumila was able to inhibit HM3KO cell growth possibly by arresting the cell cycle at G1 phase and inducing apoptosis in HM3KO cells via the up- and down-regulation of Bax/Bcl-2 protein, mediated through a p53-dependent pathway.
MeSH terms: Ethanol; Biological Assay; Cell Cycle; Flow Cytometry; Hexanes; Staining and Labeling; Down-Regulation; Tumor Suppressor Protein p53; G1 Phase; Apoptosis; Proto-Oncogene Proteins c-bcl-2; Apoptosis Regulatory Proteins; bcl-2-Associated X Protein
We investigated the antioxidant potential, cytotoxic effect, and TNF-α inhibition activity with NF-κB activation response in a chloroform fraction of Centratherum anthelminticum seeds (CACF). The antioxidant property of CACF was evaluated with DPPH, ORAC, and FRAP assays, which demonstrated significant antioxidant activity. The cytotoxicity of CACF was tested using the MTT assay; CACF effective inhibitory concentrations (IC(50)) for A549, PC-3, MCF-7, and WRL-68 cells were 31.42 ± 5.4, 22.61 ± 1.7, 8.1 ± 0.9, and 54.93 ± 8.3 μg/mL, respectively. CACF effectively and dose-dependently inhibited TNF-α release, in vitro and in vivo. CACF inhibited TNF-α secretion in stimulated RAW264.7 macrophage supernatants with an IC(50) of 0.012 μg/mL, without affecting their viability; the highest dose tested reduced serum TNF-α by 61%. Acute toxicity testing in rats revealed that CACF was non-toxic at all doses tested. Matching the cytotoxic activity towards a mechanistic approach, CACF dose-dependently exhibited in vitro inhibitory effects against the activation of NF-κB translocation in MCF-7 cells. Preliminary phytochemical screening with GC/MS analysis detected 22 compounds in CACF, of which morpholinoethyl isothiocyanate was the most abundant (29.04%). The study reveals the potential of CACF in the treatment of breast cancer and in oxidative stress conditions with associated inflammatory responses.
This paper aims to evaluate the effect of splinting during implant impression. A master model with two fixtures at the sites of 45 and 47 was used. 20 impressions were made for all four techniques: (A) indirect; (B) direct, unsplinted; (C) direct, splinted; and (D) direct, splinted, sectioned, and re-splinted. Splinting was undertaken with autopolymerizing acrylic resin (AAR). Horizontal distance between fixtures was compared using a digital caliper. The difference in distance were analysed with one-way ANOVA. Group A showed a significantly lowest accuracy among all techniques (p < or = 0.05). There was no significant difference of accuracy among the groups using direct techniques (p > or = 0.05). Group D was more accurate compared to group B and C. We conclude that splinting of impression copings would be beneficial to obtain an accurate impression.
Essential oils obtained by hydrodistillation from the rhizomes of Etlingera pyramidosphaera (K. Schum.) R. M. Sm, E. megalocheilos (Griff.) A.D. Poulsen, comb. nov., E. coccinea (Blume) S. Sakai & Nagam, E. elatior (Jack) R. M. Sm, and E. brevilabrum (Valeton) R. M. Sm were analyzed by GCMS. The highest oil yield was obtained from E. pyramidosphaera (0.45%), followed by E. elatior (0.38%), E. coccinea (0.30%), E. brevilabrum (0.28%) and E. megalocheilos (0.25%). The major constituents of the essential oils were oxygenated monoterpenes, followed by sesquiterpenes, oxygenated sesquiterpenes, oxygenated diterpenes and diterpenes. The essential oils from E. pyramidosphaera and E. brevilabrum exhibited the best cytotoxicity against MCF 7 (LC50: 7.5 +/- 0.5 mg mL(-1)) and HL 60 (LC50: 5.0 mg mL(-1)), respectively. Strong inhibition was also observed for the essential oils of E. coccinea and E. megalocheilos against Staphylococcus aureus (MIC: 8.0 +/- 0.5 mg mL(-1), and 5.0 +/- 0.5 mg mL(-1)) and Streptococcus pyrogenes (MIC: 6.0 +/- 0.5 mg mL(-1) and 8.0 +/- 0.5 mg mL(-1)).
Epidemic foci of Plasmodium knowlesi malaria have been identified during the past ten years in Malaysia, in particular in the States of Sarawak and Sabah (Malaysia Borneo), and in the Pahang region (peninsular Malaysia). Based on a review of the available recent international literature, the authors underline the importance of molecular biology examinations, polymerase chain reactions (PCR), performed with primers specific for P. knowlesi, since the current microscopic examination (haemoscope) may fail to distinguish P. knowlesi from Plasmodium malariae, due to the very similar appearance of the two parasites. P. knowlesi has been described as the causal agent of life-threatening and lethal forms of malaria: its clinical picture is more severe when compared with that of P. malariae, since the disease is characterized by greater parasitaemia, as opposed to that documented in the course of P. malariae disease. The most effective carrier is Anopheles leucosphyrus: this mosquito is attracted by both humans and monkeys. Among primates, the natural hosts of P. knowlesi are Macaca fascicularis and Macaca nemestina, while Saimiri scirea and Macaca mulatta, which cannot become infected in nature, may be useful in experimental models. When underlining the potentially severe evolution, we note the key role played by prompt disease recognition, which is expected to be more straightforward in patients monitored in endemic countries at high risk, but should be carefully implemented for subjects being admitted to hospital in Western countries suffering from the typical signs and symptoms of malaria, after travelling in South-East Asia where they were engaged in excursions in the tropical forest (trekking, and similar outdoor activities). In these cases, the diagnosis should be prompt, and suitable treatment should follow. According to data in the literature, in non-severe cases chloroquine proves very effective against P. knowlesi, achieving the disappearance of signs and symptoms in 96% of cases after only 24 hours after treatment start. In the light of the emerging epidemiological data, P. knowlesi should be added to Plasmodium vivax, Plasmodium ovale, P. malariae, and Plasmodium falciparum, as the fifth aetiological agent of malaria. During the next few years, it will become mandatory to plan an appropriate surveillance program of the epidemiological evolution, paying also great attention to the clinical features of patients affected by P. knowlesi malaria, which are expected to worsen according to the time elapsed; some studies seem to point out greater severity according to increased parasitaemia, paralleling the increased interhuman infectious passages of the plasmodium.
BACKGROUND: Streptococcus pneumoniae is a major cause of bacterial infections resulting in significant morbidity and mortality worldwide. Currently, up to 13 serotypes are included in pneumococcal conjugate vaccines (PCVs). However, the serotype formulation of these vaccines was initially designed to protect children against serotypes most commonly causing invasive disease in North America, and may not reflect the serotype distribution across the world. Data regarding pneumococcal epidemiology from the other parts of the world, in particular South East Asia, has not been reviewed.
METHODS: This systematic literature review analyses published serotype data regarding S. pneumoniae isolates from South East Asian countries (defined as countries belonging to the Association of South East Asian Nations, ASEAN): Brunei, Cambodia, Indonesia, Laos, Malaysia, Myanmar, Philippines, Singapore, Thailand and Vietnam up to 3rd of March 2012.
RESULTS: Analysis of data from six ASEAN countries, from which information on pneumococcal serotypes was available, showed that the most common disease causing serotypes (in rank order) were 19F, 23F, 14, 6B, 1, 19A and 3. Serotype distribution of pneumococcal isolates was similar across the ASEAN region. Serotype level data was more commonly reported for pneumococcal isolates causing invasive pneumococcal disease than for those from non-invasive disease. Studies from Malaysia, Thailand and Singapore contributed the largest proportion of pneumococcal isolates, and serotype data, when compared to other ASEAN countries.
CONCLUSION: This review demonstrates that the majority of IPD causing serotypes in SE Asia are included in currently licensed PCVs. However, PCV's are included in the routine childhood immunisation schedule of only one of the ten countries included in this analysis. Our findings demonstrate the scarcity of information available on serotype prevalence and distribution of pneumococci in SE Asia.
Fistula formation following pelvic surgery and radiotherapy, including ureteric-arterial fistulas (UAF), is well documented, however, ureteric-arterial-enteric fistula is extremely rare. Conventional autopsy is usually required for the definitive diagnosis of pelvic vascular fistulas although an accurate diagnosis can still be complicated and challenging. The role of post-mortem computed tomography (PMCT) as an adjunct to conventional autopsy is well documented in the literature. One of the limitations of PMCT is the diagnosis of vascular conditions. Post-mortem computed tomography angiography (PMCTA) is a recently introduced technique that can assist in detecting such pathology. We present a case of post-radiotherapy ureteric-arterial-enteric fistula presenting as massive rectal and vaginal bleeding diagnosed prior to autopsy on PMCTA. The role of PMCTA in the diagnosis of such a UAF has not previously been reported in the literature.
To determine the diffusion coefficient, water sorption and solubility of various types of restorative dental composites and to evaluate the effect of acidic media (orange juice and coke) on their characteristics.
Toxoplasma gondii is a public health risk in developing countries, especially those located in the tropics. Widespread infection may inflict a substantial burden on state resources, as patients can develop severe neurological defects and ocular diseases that result in lifelong loss of economic independence. We tested sera for IgG antibody from 493 eye patients in Malaysia. Overall age-adjusted seroprevalence was estimated to be 25% (95% CI: [21%, 29%]). We found approximately equal age-adjusted seroprevalence in Chinese (31%; 95% CI: [25%, 38%]) and Malays (29%; 95% CI: [21%, 36%]), followed by Indians (19%; 95% CI: [13%, 25%]). A logistic regression of the odds for T. gondii seroprevalence against age, gender, ethnicity and the occurrence of six types of ocular diseases showed that only age and ethnicity were significant predictors. The odds for T. gondii seroprevalence were 2.7 (95% CI for OR: [1.9, 4.0]) times higher for a patient twice as old as the other, with ethnicity held constant. In Malays, we estimated the odds for T. gondii seroprevalence to be 2.9 (95% CI for OR: [1.8, 4.5]) times higher compared to non-Malays, with age held constant. Previous studies of T. gondii seroprevalence in Malaysia did not explicitly adjust for age, rendering comparisons difficult. Our study highlights the need to adopt a more rigorous epidemiological approach in monitoring T. gondii seroprevalence in Malaysia.
A new dienamide, (2E,4E)-7-(3',4'-dimethoxyphenyl)-N-ethyl-6-(R)-hydroxyhepta- 2,4-dienamide, named (-)-kunstleramide (1), were isolated from the bark of Beilschmiedia kunstleri Gamble together with one neolignan: (+)-kunstlerone (2) and seven known alkaloids: (+)-nornuciferine (3), (-)-isocaryachine (4), (+)-cassythicine (5), (+)-laurotetanine (6), (+)-boldine (7), noratherosperminine (8), (+)-N-demethylphyllocaryptine (9). Their structures were established from spectroscopic techniques, most notably 1D- and 2D-NMR, UV, IR, OR, circular dichroism (CD) spectra and LCMS-IT-TOF. (-)-Kunstleramide (1) exhibited very poor dose-dependent inhibition of DPPH activity, with an IC₅₀ value of 179.5 ± 4.4 μg/mL, but showed a moderate cytotoxic effect on MTT assays of A375, A549, HT-29, PC-3 and WRL-68 with EC₅₀ values of 64.65, 44.74, 55.94, 73.87 and 70.95 µg/mL, respectively.
Epistasis (gene-gene interaction) is a ubiquitous component of the genetic architecture of complex traits such as susceptibility to common human diseases. Given the strong negative correlation between circulating adiponectin and resistin levels, the potential intermolecular epistatic interactions between ADIPOQ (SNP+45T > G, SNP+276G > T, SNP+639T > C and SNP+1212A > G) and RETN (SNP-420C > G and SNP+299G > A) gene polymorphisms in the genetic risk underlying type 2 diabetes (T2DM) and metabolic syndrome (MS) were assessed. The potential mutual influence of the ADIPOQ and RETN genes on their adipokine levels was also examined. The rare homozygous genotype (risk alleles) of SNP-420C > G at the RETN locus tended to be co-inherited together with the common homozygous genotypes (protective alleles) of SNP+639T > C and SNP+1212A > G at the ADIPOQ locus. Despite the close structural relationship between the ADIPOQ and RETN genes, there was no evidence of an intermolecular epistatic interaction between these genes. There was also no reciprocal effect of the ADIPOQ and RETN genes on their adipokine levels, i.e., ADIPOQ did not affect resistin levels nor did RETN affect adiponectin levels. The possible influence of the ADIPOQ gene on RETN expression warrants further investigation.
This study performed an assessment on the beneficial of the Clean Development Mechanism (CDM) application on waste treatment system in a local palm oil industry in Malaysia. Life cycle assessment (LCA) was conducted to assess the environmental impacts of the greenhouse gas (GHG) reduction from the CDM application. Calculations on the emission reduction used the methodology based on AM002 (Avoided Wastewater and On-site Energy Use Emissions in the Industrial Sector) Version 4 published by United Nations Framework Convention on Climate Change (UNFCC). The results from the studies showed that the introduction of CDM in the palm oil mill through conversion of the captured biogas from palm oil mill effluent (POME) treatment into power generation were able to reduce approximate 0.12 tonnes CO2 equivalent concentration (tCO2e) emission and 30 kW x hr power generation per 1 tonne of fresh fruit bunch processed. Thus, the application of CDM methodology on palm oil mill wastewater treatment was able to reduce up to 1/4 of the overall environment impact generated in palm oil mill.
MeSH terms: Air Pollutants/chemistry; Carbon Dioxide/chemistry; Humans; Industrial Waste; Malaysia; Plant Oils/chemistry*; Waste Disposal, Fluid*; Water Pollutants/chemistry*; Greenhouse Effect
The spinal facet joints are known to be an important component in the kinematics and the load transmission of the spine. The articular cartilage in the facet joint is prone to degenerative changes which lead to back pain and treatments for the condition have had limited long term success. There is currently a lack of information on the basic biomechanical properties of the facet joint cartilage which is needed to develop tissue substitution or regenerative interventions. In the present study, the thickness and biphasic properties of ovine facet cartilage were determined using a combination of indentation tests and computational modelling. The equilibrium biphasic Young's modulus and permeability were derived to be 0.76±0.35 MPa and 1.61±1.10×10⁻¹⁵ m⁴/(Ns) respectively, which were within the range of cartilage properties characterised from the human synovial joints. The average thickness of the ovine facet cartilage was 0.52±0.10 mm, which was measured using a needle indentation test. These properties could potentially be used for the development of substitution or tissue engineering interventions and for computational modelling of the facet joint. Furthermore, the developed method to characterise the facet cartilage could be used for other animals or human donors.
This paper presents solid waste bin level detection and classification using gray level co-occurrence matrix (GLCM) feature extraction methods. GLCM parameters, such as displacement, d, quantization, G, and the number of textural features, are investigated to determine the best parameter values of the bin images. The parameter values and number of texture features are used to form the GLCM database. The most appropriate features collected from the GLCM are then used as inputs to the multi-layer perceptron (MLP) and the K-nearest neighbor (KNN) classifiers for bin image classification and grading. The classification and grading performance for DB1, DB2 and DB3 features were selected with both MLP and KNN classifiers. The results demonstrated that the KNN classifier, at KNN = 3, d = 1 and maximum G values, performs better than using the MLP classifier with the same database. Based on the results, this method has the potential to be used in solid waste bin level classification and grading to provide a robust solution for solid waste bin level detection, monitoring and management.
In this article, modified κ-carrageenan hydrogel nanocomposites were synthesized to increase the release ability of carrageenan hydrogels under gastrointestinal conditions. The effect of MgO nanoparticle loading in a model drug (methylene blue) release is investigated. Characterization of hydrogels were carried out using Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), Field Emission Scanning Electron Microscope (FESEM) and Differential Scanning Calorimetry (DSC). Genipin was used to increase the delivery performance in gastrointestinal tract delivery by decreasing release in simulated stomach conditions and increasing release in simulated intestine conditions. It is shown that the amount of methylene blue released from genipin-cross-linked nanocomposites can be 67.5% higher in intestine medium and 56% lower in the stomach compared to κ-carrageenan hydrogel. It was found that by changing the nanoparticle loading and genipin concentration in the composite, the amount of drug released can be monitored. Therefore, applying nanoparticles appears to be a potential strategy to develop controlled drug delivery especially in gastrointestinal tract studies.
MeSH terms: Carrageenan/metabolism*; Carrageenan/chemistry*; Delayed-Action Preparations/chemistry; Drug Carriers/chemistry; Hydrogen-Ion Concentration; Magnesium Oxide/chemistry; Hydrogel/metabolism; Hydrogel/chemistry*; Gastrointestinal Tract/metabolism; Nanocomposites/ultrastructure; Nanocomposites/chemistry*; Metal Nanoparticles/chemistry*
The availability of highly active homologous promoters is critical in the development of a transformation system and improvement of the transformation efficiency. To facilitate transformation of green microalga Ankistrodesmus convolutus which is considered as a potential candidate for many biotechnological applications, a highly-expressed native promoter sequence of ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (AcRbcS) has been used to drive the expression of β-glucuronidase (gusA) gene in this microalga. Besides the determination of the transcription start site by 5'-RACE, sequence analysis revealed that AcRbcS promoter contained consensus TATA-box and several putative cis-acting elements, including some representative light-regulatory elements (e.g., G-box, Sp1 motif and SORLIP2), which confer light responsiveness in plants, and several potential conserved motifs (e.g., CAGAC-motif, YCCYTGG-motifs and CACCACA-motif), which may be involved in light responsiveness of RbcS gene in green microalgae. Using AcRbcS promoter::gusA translational fusion, it was demonstrated that this promoter could function as a light-regulated promoter in transgenic A. convolutus, which suggested that the isolated AcRbcS promoter was a full and active promoter sequence that contained all cis-elements required for developmental and light-mediated control of gene expression, and this promoter can be used to drive the expression of heterologous genes in A. convolutus. This achievement therefore advances the development of A. convolutus as an alternative expression system for the production of recombinant proteins. This is the first report on development of gene manipulation system for unicellular green alga A. convolutus.
P. minus is an aromatic plant, the leaf of which is widely used as a food additive and in the perfume industry. The leaf also accumulates secondary metabolites that act as active ingredients such as flavonoid. Due to limited genomic and transcriptomic data, the biosynthetic pathway of flavonoids is currently unclear. Identification of candidate genes involved in the flavonoid biosynthetic pathway will significantly contribute to understanding the biosynthesis of active compounds. We have constructed a standard cDNA library from P. minus leaves, and two normalized full-length enriched cDNA libraries were constructed from stem and root organs in order to create a gene resource for the biosynthesis of secondary metabolites, especially flavonoid biosynthesis. Thus, large-scale sequencing of P. minus cDNA libraries identified 4196 expressed sequences tags (ESTs) which were deposited in dbEST in the National Center of Biotechnology Information (NCBI). From the three constructed cDNA libraries, 11 ESTs encoding seven genes were mapped to the flavonoid biosynthetic pathway. Finally, three flavonoid biosynthetic pathway-related ESTs chalcone synthase, CHS (JG745304), flavonol synthase, FLS (JG705819) and leucoanthocyanidin dioxygenase, LDOX (JG745247) were selected for further examination by quantitative RT-PCR (qRT-PCR) in different P. minus organs. Expression was detected in leaf, stem and root. Gene expression studies have been initiated in order to better understand the underlying physiological processes.