24 research outputs found
PRODUCTION AND CHARACTERIZATION OF siRNA LOADED MAGNETIC NANOPARTICLES TO BE USED IN CANCER TREATMENT
No full textABSTRACT
PRODUCTION AND CHARACTERIZATION OF siRNA LOADED MAGNETIC NANOPARTICLES TO BE USED IN CANCER TREATMENT
AMINA SELIMOVIC
Master of Science, Bioengineering
Supervisor: Prof. Dr. Emir Baki DENKBAŞ
June 2017, 73 pages
Cancer is one of the leading causes of death in economically developed and developing countries and factors such as smoking, physical inactivity and nutritional habits are increasing the possibility of its development. After decades of extensive research of cancer, today it can be described as a genetic disease of the somatic cell. Genetic changes such as rearrangements of chromosomes (deletion, translocation, insertion), point mutation, gene amplification are affecting kinase inhibitors, growth factors, receptors, a cascade of transcription factors and signal transduction members are leading to an impaired balance of cell proliferation and changes in the function of genes that induce apoptosis. These actions are leading to the abnormal growth of the cells called neoplasia. An approach based on nanotechnology provides a great promise in developing strategies for cancer treatment by helping to improve the safety and efficacy of therapeutic delivery vehicles. Powerful investigational tools with great potential in therapeutics-RNA interference (RNAi) is known as a highly efficient regulatory process in which short double-stranded RNAs are giving rise to sequence-specific posttranscriptional gene silencing. With time it has been proven that specific protein expression can be inhibited. Small interfering RNA (siRNA) as part of RNA interference process has been extensively studied to treat various genetic diseases, cardiovascular diseases and various cancers. However, due to its polyanionic nature siRNAs cannot cross the cellular membrane that is why it needs a carrier to prevent enzymatic degradation and to take siRNA to the specific target inside the cell. Properties such as safety, effectiveness, ease of manufacturing and production are quite important to consider when selecting a proper carrier for siRNA. In this work iron oxide nanoparticles are coated with natural biopolymer gelatin, loaded with mTOR siRNA targeting specific oncogene with the aim to deliver gene silencing complex to colon cancer cells inducing therapeutic effect. For this iron oxide and gelatin coated iron oxide nanoparticles were synthesized, optimized and characterized. Morphological characterization was done using SEM (Scanning Electron Microscopy). Size and surface charge of produced nanoparticles was revealed by Zeta-Sizer (3000 HSA, Malvern, England). To determine the chemical structures of the nanoparticles, molecular bond characterization had been performed using Fourier Transform Infrared Spectroscopy (FTIR) (Nicolet iS10, USA). VSM (vibrating sample magnetometer) and ESR (electro spin resonance) are used to analyse magnetic properties of the prepared particles. siRNA was loaded to gelatin coated iron oxide nanoparticles and its binding efficiency (%) was examined. siRNA loaded nanoparticles were transfected to colon cancer cell line CaCo-2 and mouse fibroblast cell line L929. Cell cytotoxicity test, MTT was performed using different concentrations of siRNA and under different incubation time MTT assay showed that toxic effect in both cell lines was significantly higher when siRNA loaded gelatin coated IONs were used. Also, according to the results obtained, synthesized gelatin coated IONs showed similar anticancer activity as HiperFect which is commercial siRNA carrier. This work showed that gelatin coated iron oxide nanoparticles as cheap and easily synthesized carrier are promissing tool for siRNAs delivery.
Keywords: nanooncology, iron oxide nanoparticles, gelatin, siRNA1. INTRODUCTION
Cancer is one of the leading causes of death in economically developed and developing countries. After decades of extensive research today it can be described as a genetic disease of the somatic cell where changes in the sequence of the genome occur. Genetic changes such as rearrangements of chromosomes (deletion, translocation, insertion), point mutation, gene amplifications are affecting kinase inhibitors, growth factors and their receptors, cascade of transcription factors and signal transduction complex members. Today, gene therapy has emerged as one of the promising ways to treat cancer cells. Gene therapy is referring to gene replacement where the right copy (DNA or RNA fragments) of the mutated gene is delivered to the cell without causing toxicity to neighbouring cells and tissues. Gene therapy is providing the possibility to silence the gene that potentially causes hereditary disease. Gene replacement in gene therapy is considered successful if the right delivery vehicle is used to cause gene expression in the target cell. Today, instead of traditional approaches to cancer treatment, great interest is dedicated to targeted therapies because it does not harm healthy cells and have high selectivity. Nanoparticles are having the ability to deliver drugs, vaccines, siRNAs and proteins to different body organs such as lungs, brain, lymphatic system, spleen and lymphatic system for targeting. In this context, a promising approach is RNA interference system where siRNA therapy operates on mRNA level preventing genes from being translated into protein. RNA interference mechanism is a revolutionary process of post transcriptional gene silencing led by double-stranded RNA (dsRNA) who is homologous in the sequence to a target protein inducing sequence-specific degradation of mRNA of interest. Double-stranded RNAs known as siRNA found in the cytoplasm are capable of silencing own complementary mRNA. Dicer is cleaving long dsRNAs into short interfering RNA (siRNA) that is usually RNA double stranded sequence of 21-24 nt in length. Subsequently, obtained siRNAs will be loaded onto activated form of RISC (RNA-induced silencing complex) where protein Ago-2 will cleave dsRNA sequence and release it as ssRNA. Released ssRNA represents guide sequence right toward mRNA of the target with whom it has a high degree of sequence complementarity. All molecular players of RISC complex are yet unknown, however, the susceptibility of siRNA to rapid degradation and its short half-life is an important barrier that restricts therapeutic use.
The efficiency of siRNA delivery in great extent depends on its delivery vehicle and surface modification which gives protection to siRNA complex ones found in the biological environment during circulation.
Basically, in order to avoid systematic elimination of siRNA, chemical modification and selection of nano-sized carrier is crucial. In order to avoid „off-target-effect" and innate immune response there exist several properties in design and synthesis of nanoparticles such as size, shape, surface charge, density, composition, and surface chemistry that gives rise to the amount of efficiency in performing a function. The development and application of nano-oncology have raised as a promising tool for early termination of cancer because coated nanostructures have great potential to carry and deliver oncogene silencing complex to the cells without disturbing its integrity and with no causing damage to healthy tissues. Work of Jiang et al, states that iron metabolism which occurs in the human body through multiple pathways and dextran-coated iron oxide nanoparticles have been clinically tested and approved by the FDA which made it potential drug in cancer treatment but the problem was seen with nanoparticle toxicity and off-target siRNA effects [1, 2]. That is why in order to decrease toxicity biodegradable polymers were used to coat nano carriers to increase results efficiency. In line with a prior study done by of Chen et al, where it is stated that magnetic nanoparticles (IONs) covered with a layer of biodegradable polymer are noted to be effective magnetic drug carrier [3] this study is based on delivering siRNA mTOR to CaCo-2 cell culture by using magnetic nanoparticles covered with a layer of biodegradable polymer gelatin. Gelatin, biological macromolecule except being cheap it is also biodegradable, biocompatible and vastly used in pharmaceutical and nanotechnological approach as drug delivery or vaccine release system. When comparing with other delivery systems, gelatin nanoparticles are a solid delivery system for drug molecules or siRNA they encapsulate. The whole study is composed of three parts where firstly iron nanoparticle synthesis and characterization by the co-precipitation method using iron salts in alkaline medium is done, then same nanoparticles were coated with gelatin. After coating with gelatin, iron oxide nanoparticles were loaded with mTOR siRNA so that interaction and gene silencing performance of produced nano platform could be tested when transfected to CaCO-2 colon cancer cell culture.
2.GENERAL INFORMATION
2.1. Gene therapy
Twenty years after revealing DNA structure in 1953 by Watson and Crick, DNA restriction enzymes popped out as a new tool in DNA recombinational technology. Recombinant DNA technology provides different ways of DNA usage by DNA recombinant expression in bacteria, expressing recombinant DNAs in transgenic animals or just using DNA recombinant as therapy for replacing or silencing genes causing a specific disease. Right after recombinant technology popped out gene therapy strategies started to be a theme of interest in scientific circles. Gene therapy is referring to gene replacement where the right copy (DNA or RNA fragments) of the mutated gene is delivered to the cell without causing toxicity to neighbouring cells and tissues. Also, gene therapy is providing silencing of the gene that potentially can cause hereditary disease. Gene replacement in gene therapy is considered successful if the right delivery vehicle is used to cause gene expression in the target cell. Delivery to the cell can be made throughout either viral or non-viral method. So far, viral vectors that showed promissing results are lentiviral, retroviral, and adeno-associated viral vectors which have the ability to fuse into the human genome. Also, transfection can be done by microinjection, electroporation, ballistic DNA injection or a gene gun, sonoporation or ultrasound, photoporation, magnetofection, jet injection and using many other methods [4, 5].
Figure 2.1. Direct and cell-based delivery in gene therapy [4].
In so-called cell-based delivery or more known as ex-vivo delivery, cells of interest taken from a patient are genetically modified in the laboratory and packaged with a viral or non-viral delivery vector to be re-transfected to patients. In vivo therapy is relying on delivering vector loaded with genetic material to direct gene modification and permanent silencing of gene expression after entering the cell. In direct delivery or so-called in situ gene therapy, the genetic material after packaging in the delivery vehicle is directly transfected to the target tissue. This method is used in respiratory tract disease treatment using lipids and adenoviruses where the low efficiency of transfection is noted [5]. Gene therapy is reported to have success in treatment primary stages of the disease. To increase the success of therapy proper gene delivery agent has to be chosen to decrease toxicity and increase the cure rate.
2.2. Gene silencing strategies
It is reported that small RNAs, miRNAs, siRNAs are involved in sequence-specific inhibition of specific gene. Gene expression is controlled throughout translational repression, chromatin modification and mRNA degradation.
Figure 2.2. Gene control mechanisms [6].
There are two ways how gene control mechanism is taking place; throughout miRNA found in the nucleus or by siRNA found in the cell cytoplasm. miRNA leads to translational inhibition where on the other side siRNA leads to mRNA cleavage after dsRNAs are formed in the cytoplasm or complementary strand is synthetically injected by a viral agent or transfected using different delivery systems [6].
2.2.1. miRNA Mechanism
In the cell nucleus, transcripts of miRNA genes, mostly known as introns are folding into hairpin shape making miRNAs. These miRNAs before leaving nucleus are reaching maturation stage by the help of RNase III family enzymes; Drosha and Dicer making so-called Drosha-Pasha complex. After being processed with Dicer, siRNAs and miRNAs are associated with Argonaut proteins. Like siRNAs, miRNAs are incorporated into RNA-induced silencing complex (RISC) that eventually leads to sequence-specific inhibition of gene expression. With help of exportin-5 pre-miRNA are transported to the cytoplasm. If siRNA is associated complex is known as RISC, but if miRNA is associated miRNPs complex is formed [6].
2.2.2. siRNA Mechanism and RNAi
RNA interference mechanism is a revolutionary process of post transcriptional gene silencing. This mechanism is led by double-stranded RNAs which are homologous in the sequence to a target protein leading to specific degradation of target mRNA. Double-stranded RNAs (dsRNAs) known as siRNAs found in the cytoplasm are capable of silencing own complementary mRNA. siRNAs are part of the non-coding RNAs family (ncRNAs) which is composed of long ncRNAs (lncRNAs) and small ncRNAs. The most well-known small ncRNAs that have a function in transcriptional regulation and post-transcriptional gene silencing are siRNA and miRNA while on the other side long dsRNAs are precursors for siRNAs. Gene silencing mechanism with help of endoribonuclease Dicer is cleaving long dsRNAs into short interfering RNA which are usually RNA double stranded sequences of 21-24 nt in length as shown in Figure 1. Name Dicer (DCR) is coming from the fact it is having the capacity to digest dsRNA into small RNAs of a proper size. As stated in work of Agrawal N. et al, DCR has shown same function and outcome as nucleases of RNAase III family members. It is reported that RNAase III-like enzyme in Drosophila showed the same outcome giving final product of 22nt fragments [7].
Subsequently, obtained siRNAs fragments will be loaded onto activated form of RISC where protein Ago-2 will cleave dsRNA sequence and release it as ssRNA. For proper RISC activation, the passenger strand is removed, while onto Ago-2 guide strand is loaded. Ago-2 cleaves the target mRNA at 5′ end between 10 and 11 base of the siRNA antisense strand. Released ssRNA represents guide sequence right toward mRNA of the target with whom it has a high degree of sequence complementarity. All molecular players of RISC complex are yet unknown, so far it is known that Ago2 protein complex has great role in post transcriptional silencing [8].
Figure 2.3. siRNA mediated gene silencing process [9].
After thermodynamically favourable base pairing of siRNA and mRNA is done mRNA is cleaved and destructed by cellular nucleases. Right after finishing this cycle RISC complex is free to take up additional mRNA molecules of the target.
When it comes to the internalisation process major role play endocytosis enabling molecules internalisation together with cell membrane component. Naked siRNA cannot readily cross anionic cell membrane because of its high molecular weight, large size and negative charge coming from hydrophilic phosphate backbone. It is known that if a carrier is coated with ligands or antibodies specific binding of a carrier to cell membrane enables much easier passage into the cell. In the process of endocytosis, anionic proteoglycans are making interaction with negatively charged siRNAs forming an endocytic vesicle on the cell surface [8]. When RNAi enclosed in endosome is released in cytoplasm it is ready for interaction with RISC. Once RNAi is taken up via endocytosis it is exposed to the degradation by nucleolytic enzymes if successfully avoids it successful delivery is done.
2.2.2.1. Function of Dicer in silencing
Dicer is part of RNase enzyme family composed of PAZ domain, helicase /ATPase, DUF283 domains, a C-terminal dsRBD and two catalytic RNase III domains (RIIIa and RIIIb). Dicer’s main role is to process dsRNA to siRNA and to induce cleavage of pre-miRNA to 20-bp miRNA sequences in the cytoplasm. Dicer protein is known to be involved in assembly of RISC complex and close interaction with Argonaute proteins.
Figure 2.4. Schematic representation of Dicer’s function [6].
Two catalytic sites RIIIa and RIIIb direct cleavage of dsRNA just 20bp from its terminus. PAZ domain recognises pre-miRNA 3’overhang which is coming from Drosha processing. The function of helicase/ATPase in not fully understood. In RNA silencing Dcr-1 is functioning in the processing of miRNA were on the other side Dcr-2 is involved in RNAi processing [6].
2.2.3. Design of siRNA
When designing siRNAs, basic parameters such as internal bps repeats at specific positions at the sense strand, GC content and siRNA length of around 19–22 bps should be considered. According to Draz et al, substitution of 2'-O-methyl ribosyl group at position 2 in the guide strand could reduce silencing of most off-target transcripts which are complementar to the siRNA guide [10].
2.2.4. siRNA and cell interaction
One of the leading challenges when it comes to delivery of siRNA⁄nanoparticle complex to the cell is the way because to yield significant effect on a target cell ratio between siRNAs and delivery reagent must be adjusted. Therapeutic use of siRNA is shortened because it is rapidly degraded by nucleases in the cytoplasm and therefore it has a short half-life. For this, it is a great need for the use of effective and suitable carrier system for siRNA to trigger RNAi mechanism. Although effective viral delivery systems have been developed to address this problem risk of virus recombination and strong immunogenicity are limiting its use. The ability of nanoparticles to pass easily through the cell membrane has offered alternative treatment methods by carrying DNA or RNA oligonucleotides. In order to reach the target in the cytoplasm by freely passing through the cell membrane and escaping from the cytoplasmic vesicle; liposomes, peptides, polymer-based, magnetic nanoparticles were offered as an alternative in transfection.
2.3. Cancer development
Cancer is one of the leading causes of death in economically developed and developing countries. After decades of extensive research, cancer can be described as a genetic disease of the somatic cells where certain changes in the sequence of the gene occur. Genetic changes such as rearrangements of chromosomes (deletion, translocation, insertion), point mutation and gene amplification are affecting kinase inhibitors, growth factors, growth factor receptors, a cascade of transcription factors and signal transduction complex members.
Mutations in germ and somatic cells increase the likelihood for cancer-causing random tumour development. The final result increases in tumour cell number which is coming either by stimulation of cell division or dysregulation in cell death program. Impaired balance of cell proliferation and changes in genes that regulate cell death are giving rise to the abnormal growth of the cells. Cells, the simplest building blocks of all living things are taking control of their life cycle. If something goes wrong in cell cycle cells are starting to grow irregularly. Irregular cell growth is giving harm to functions and organs in the body. Cell proliferation is regulated through cell cycle which is comprised of four check phases known as G1 (gap 1), S (DNA synthesis), G2 (gap 2) and M (mitosis). The cell cycle is controlled at two checkpoints at G1 and G2. RB tumour suppressor protein is controlling G1 checkpoint, whereas p53 activation triggers cell cycle arrest at G1 or G2 point once DNA damage or some other abnormal behaviour is sensed. There are three types of genes which are known to be affected by gene alterations. These are tumor-suppressor genes, oncogenes and stability genes (caretakers). Mutations in tumour suppressor and oncogenes are leading to uncontrolled cell growth that is having the potential for constant replication. Stability genes or caretakers promotes tumour development in a way that mistakes in control points of DNA replication are made. These are mismatch repair (MMR), nucleotide-excision repair (NER) and base-excision repair (BER) genes [11]. Normal cells, as part of their life span, are falling into the process of senescence where cells are reaching a critical number of division and stop growing. Although, most tumour cells do not undergo senescence and tend to replicate constantly. Cancer cells are known to have limitless replicative potential, sustained angiogenesis, insensitiveness to anti-growth signals, developed a system to evade apoptosis leading to tissue invasion and metastasis. The metastatic tumour is formed when the malignant cells are gaining the ability to leave the primary tumour and migrating to the circulative system. Tumour cells are settling and gaining the ability to proliferate in new microenvironment [12].
Figure 2.5. Schematic representation of cell cycle [12].
In cancer development process, both genetic alterations and epigenetic changes are having influence. These changes are leading to activation or silencing of genes which are in charge of cancer development. Genetic changes such as mutations, polymorphism and translocation are triggering cancer development. Various genetic alterations can be found within one single tumour. Alterations in DNA structure, nucleotide point mutations where specific gene responsible for cancer development is affected, chromosome tr
”Jonas, min vän” : Persuasiva motstrategier för att dekonstruera en stark ethosposition, exemplet Selimovic mot Khemiri i debatten om REVA
No full textIn the year of 2012, a police project called “REVA” was established in Sweden. The goal was to increase efficiency in deporting undocumented migrants from Sweden. Two articles relating to the project were published in Swedish newspaper Dagens Nyheter. One was an open letter to Attorney General Beatrice Ask, written by the famous author Jonas Hassen Khemiri. The other one was a response to Khemiri’s letter, written by Secretary of state and drama director Jasenko Selimovic. The objective of this essay is to perform a rhetorical analysis of these two articles in order to examine how they are constructed to achieve their persuasive goals. Mainly, I am interested in exploring if Selimovic’s persuasive strategies can be regarded as direct counter-strategies to those of Khemiri. I am also interested in exploring to what extent Selimovic’s strategies deal with the specific problems he faces: countering an effective text with a demonstrable rhetorical power, authored by a writer with a contextually strong ethos - and doing so without harming his own ethos. Through using a theory of argumentation as well as a specially designed theoretical model for persuasive strategies “in both directions”, the analysis shows that Selimovic’s main approach was to directly confront the persuasive strategies used by Khemiri in order to make the latter appear too closely focused on subjective emotional experiences and blind to a wider perspective. Theoretically, this approach could be expected to work well in deconstructing an uneven ethos position, but due to Selimovic’s overly confrontational argumentation style, it rather had the opposite effect, making Selimovic appear too skeptical, critical and unwilling to stay on topic.
”Jonas, min vän” : Persuasiva motstrategier för att dekonstruera en stark ethosposition, exemplet Selimovic mot Khemiri i debatten om REVA
No full textIn the year of 2012, a police project called “REVA” was established in Sweden. The goal was to increase efficiency in deporting undocumented migrants from Sweden. Two articles relating to the project were published in Swedish newspaper Dagens Nyheter. One was an open letter to Attorney General Beatrice Ask, written by the famous author Jonas Hassen Khemiri. The other one was a response to Khemiri’s letter, written by Secretary of state and drama director Jasenko Selimovic. The objective of this essay is to perform a rhetorical analysis of these two articles in order to examine how they are constructed to achieve their persuasive goals. Mainly, I am interested in exploring if Selimovic’s persuasive strategies can be regarded as direct counter-strategies to those of Khemiri. I am also interested in exploring to what extent Selimovic’s strategies deal with the specific problems he faces: countering an effective text with a demonstrable rhetorical power, authored by a writer with a contextually strong ethos - and doing so without harming his own ethos. Through using a theory of argumentation as well as a specially designed theoretical model for persuasive strategies “in both directions”, the analysis shows that Selimovic’s main approach was to directly confront the persuasive strategies used by Khemiri in order to make the latter appear too closely focused on subjective emotional experiences and blind to a wider perspective. Theoretically, this approach could be expected to work well in deconstructing an uneven ethos position, but due to Selimovic’s overly confrontational argumentation style, it rather had the opposite effect, making Selimovic appear too skeptical, critical and unwilling to stay on topic.
“Sharia on a Plate?” A critical discourse analysis of halal food in two Norwegian newspapers
No full textMarianne Hafnor Bøe og Mona Helen Farstad. Islam. Tradisjon, tilhørighet og praksis. Cappelen Damm Akademisk, 2023
No full textSelimović's peaceful war (Bosnia's memory, fortified)
No full textIn this thesis, I will discuss The Fortress, a novel written by the Bosnian and Herzegovinian author Mehmed Selimovic. With the help of Tolstoy's War and Peace, I will reflect upon the subject of history and questions that so influenced Selimovic: what is history, who writes it and what purpose does it serve. Additionally, I will also look at the author's understanding of history as an incomplete truth and simply, as a reaction to our present situation. Selimovic's use of Bosnia's architecture and folklore further illustrate how cultural identities are born in reaction to historical events. The author's message is one of peace and love. More than that, The Fortress explains the author's own belief in the power of choice, with a larger message that, as historical actors, we can choose peace and love. Ultimately, Selimovic is proposing that even in the most oppressive of societies, there exists the idea of a personal choice
Magnetic gelatin nanoparticles as a biocompatible carrier system for small interfering RNA in human colorectal cancer: Synthesis, optimization, characterization, and cell viability studies
No full text© 2022 Elsevier LtdIron oxide-based nanoparticles have gained tremendous attention in developing next-generation personalized medicine modalities. Gelatin can be a good alternative for encapsulating iron oxide nanoparticles with its biocompatibility, biodegradability, low immunogenicity, and richness of functional groups. Herein, magnetic iron oxide nanoparticles (MNPs) were synthesized, coated with gelatin (Gel-MNPs), and loaded with mammalian target of rapamycin (mTOR)-silencing siRNA to induce the in vitro therapeutic effect in colorectal cancer (CRC) cells. To the best of our knowledge, this study is the first report using Gel-MNPs as siRNA carriers. We first optimized several experimental conditions for the preparation of MNPs and Gel-MNPs and the resulting optimized nanoparticles showed a narrow size and size distribution. Gelatin-coating increased the storage stability by preventing the aggregation of MNPs and did not alter the magnetic characteristics of MNPs significantly. siRNA encapsulation abilities of Gel-MNPs were determined in the range of 18.4% and 41.5% in varying siRNA amounts. Bare Gel-MNPs were highly cytocompatible against CRC cells, Caco-2, while Gel-MNPs-mTOR-siRNA exhibited a significant anticancer effect to kill these cells. Comparison with HiPerFect, a commercial siRNA transfection reagent, demonstrated that Gel-MNPs-mTOR-siRNA inhibited cell viability almost similar to or better than HiPerFect-mTOR-siRNA. Taken together, our data indicated that Gel-MNPs could potentially be used in further gene silencing approaches as a safe and multifunctional siRNA carrier candidate
Full-genome based molecular characterization of encephalitis-associated bovine astroviruses
No full textAbstractNovel types of astrovirus have been identified recently in association with neurological disease in cattle. Among those viruses is bovine astrovirus CH13 (BoAstV CH13) that has been identified in Switzerland in a cow with encephalitis. Molecular testing by a combination of reverse transcription (RT-) PCR and in situ hybridization (ISH) indicated that astrovirus infection accounts for around one quarter of viral encephalitis cases of unknown etiology in cattle. Yet, it remained to be explored whether these animals were infected by BoAstV CH13 or other astrovirus species. In the present study we sequenced the entire astrovirus genome in brain tissues of eight RT-PCR and/or ISH positive cattle. Phylogenetic comparison of the genomic RNA and the encoded non-structural and structural proteins revealed that all these astrovirus strains were very similar to BoAstV CH13 as well as to a bovine encephalitis strain reported from the USA (BoAstV NeuroS1), and clearly distinct from other previously reported astroviruses. Conserved 5′ and 3′ untranslated regions (UTRs) were predicted to display distinct secondary RNA structures, which likely play a role in viral RNA replication and/or protein translation. Based on these data we propose that BoAstV CH13/NeuroS1 represents a new genotype species within the genus Mammastrovirus. The high degree of similarity between the strains and their relative distance to other genotype species suggest that during evolution some astroviruses acquired factors that specifically contribute to neuroinvasion
A PREDICTIVE VALUE OF EARLY CLINICAL PARAMETERS FOR ABNORMAL BRAIN MRI SCAN IN NEONATES TREATED WITH THERAPEUTIC HYPOTHERMIA
No full textIntroduction: Brain MRI scans can predict neurodevelopmental outcomes in neonates treated with therapeutic hypothermia. It is a common clinical practice to perform brain MRI before discharge, but brain MRI scans performed at around four months of age have a better prognostic value for a long-term neurological outcome in asphyxiated neonates.
Aim: To identify which of three selected clinical parameters (oral feeding ability, muscle tone, history of seizure) evaluated 10 days after therapeutic hypothermia could predict the primary outcome of an abnormal brain MRI.
Methods: We reviewed the medical records of neonates ≥ 36 completed weeks of gestation consecutively treated with therapeutic hypothermia who underwent brain MRI. Clinical parameters on day 10 after therapeutic hypothermia were correlated with brain MRI findings in the first 7-14 days of life. Logic regression analysis was performed using all three covariates of the clinical status, with an abnormal MRI as the primary outcome.
Results: Brain MRI was abnormal in 42 (51.85 %) neonates with the following distribution of brain injury patterns: abnormal signal in the basal nuclei in 6, an abnormal signal in the cortex in 16, an abnormal signal both in the cortex and basal nuclei in 20 neonates. Out of three analyzed clinical parameters, feeding difficulty (P < 0.001, OR 8.3, 95% CI 2.9 - 28.9) and a history of seizures (P < 0.001, OR 11.95, 95% CI 3 - 44.5) were significantly associated with an abnormal MRI.
Conclusion: Neonates who were capable of full oral feeding by day 10 after therapeutic hypothermia and had no history of seizures were unlikely to have an abnormal MRI. This may be used in selective planning of pre-discharge MRI in asphyxiated neonates
Evaluation of Diagnostic Efficiency of Alpha-Fetoprotein in Patients with Liver Cirrhosis and Hepatocellular Carcinoma: Single-Center Experience
No full textBACKGROUND: AFP serum levels are considered as diagnostic and specific for hepatocellular carcinoma (HCC) in patients with liver cirrhosis (LC).
AIM: This study aimed to examine the diagnostic value of AFP in the distinguishing of patients with HCC from patients with LC, and to analyse the potential correlation between AFP levels and liver disease stages.
MATERIAL AND METHODS: Fifty patients with LC and fifty patients with HCC were included in this study. The majority of the patients were males, while the HBV aetiology was dominant.
RESULTS: Significant differences between LC and HCC patients were detected for AST, ALT, GGT, bilirubin, AFP and AP. Patients with HCC had higher AFP values compared to LC. There was no significant correlation between the size of the tumour lesion and serum AFP levels. A positive correlation between AFP concentration and GGT activity was determined, as was the negative correlation between AFP and age of the subjects. The AFP value of 23.34 ng/m showed high sensitivity (84%) and specificity (82%).
CONCLUSION: The size of the surface below the ROC curve (AUC) was 0.877 (0.80-0.95), which makes AFP a good biomarker and this diagnostic test is sufficient to separate patients with HCC and LC
