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Role of Exosomes in the Progression, Diagnosis, and Treatment of Brain Tumors
Vaníková, Lucie ; Zíková, Martina (advisor) ; Lacina, Lukáš (referee)
Recent studies have confirmed the importance of extracellular vesicles, particularly exosomes, in the development of brain tumors. Considerable attention has been paid mainly to the influence of exosomes on biological processes in brain tumors. Exosomes mediate intercellular communication in the tumor microenvironment by transporting biomolecules. Most often they transmit various types of ribonucleic acids, specifically microRNAs, which affect the signalling pathways related to tumour growth in target cells. Thus, exosomes play an important role in tumor cell proliferation and differentiation, metastasis, and tumor resistance to chemotherapy or radiation. Due to their small size, exosomes can cross the blood-brain barrier and thus promote tumor progression. The topic of the bachelor thesis is a summary of the current knowledge on the role of exosomes in brain tumor progression, diagnosis and treatment.
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Study of exosomes as drug delivery system in therapy of glioblastoma
Tomášková, Lucia ; Šimůnek, Tomáš (advisor) ; Macháček, Miloslav (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Lucia Tomášková Supervisor: prof. PharmDr. Tomáš Šimůnek, Ph.D. Title of diploma thesis: Study of exosomes as a drug delivery system in the treatment of glioblastoma Central nervous system disorders are among the most serious diseases affecting humans. They affect not only the patient's life, but also his/her surroundings. Therefore, their therapy, whether at the level of complete cure or alleviation of accompanying symptoms, is a challenge for scientific research. In our research, we focused on glioblastoma multiforme, a brain cancer not yet treatable. The main drawback in therapy is overcoming the blood-brain barrier. Exosomes, such as the body's natural nano-vesicles, have been shown to be a suitable system for delivering drugs to brain tissue. Our research has shown that by a suitable method we are able to obtain sufficient quality exosomes from macrophage and fill them very efficiently with antitumor agents paclitaxel, doxorubicin and temozolomide, while the delivered substances show higher efficacy and fewer side effects than the free form.
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Exosomes in viral infection and cancer
Sekavová, Alžběta ; Španielová, Hana (advisor) ; Hirsch, Ivan (referee)
Exosomes facilitate intercellular communication and transport of cellular cargo. Understanding the mechanisms underlying the cargo sorting to exosomes and the transport itself is crucial for vaccine development and diagnostic research. Exosome-mediated transfer contributes to immune response as well as progression of several diseases, including cancer and viral infections. Research on exosomes and their role in life cycles of tumorigenic viruses links already known mechanisms of viral carcinogenesis to the transport mechanisms of both cellular and viral proteins and nucleic acids. Epstein-Barr virus employs exosomes for transmission of the LMP1 oncoprotein and regulatory RNAs, whereas human immunodeficiency virus exploits cellular exosomal pathway for hijacking its membrane during budding, which helps it evade the immune system. It has been discovered that hepatitis C virus transfers its infectious virions between cells in exosomes. Exosomes containing oncoproteins and viral RNAs are also released from cells infected with other human tumorigenic viruses. However, mechanisms and implications of such events remain to be discovered. Keywords: exosome, cancer, viral infection, tumorigenic viruses, immunity, in- tercellular communication, hepatitis C virus, Epstein-Barr virus, human immuno- deficiency virus
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Rab-2A protein: its localization and function in human spermatozoa and fertilization
Sadílková, Lucie ; Postlerová, Pavla (advisor) ; Lánská, Eva (referee)
The Rab-2A protein belongs to the Rab family of monomeric G-proteins involved in the regulation of intracellular vesicular transport. Although several studies describing the role of Rab-2A protein in mammalian sperm have been published, the exact localization and function of this protein in male gametes have not been fully understood yet. In the diploma thesis, we were the first to describe the presence of Rab-2A in human sperm using 5C5, produced by the Laboratory of Reproductive Biology. Using the 5C5 antibody, we were able to identify several Rab-2A isoforms with molecular weights of 26, 24, 22, and 18 kDa in the human sperm lysate, confirmed by mass spectrometry analysis. Using the indirect immunofluorescence method, we localized Rab-2A in the acrosome area of both ejaculated and capacitated sperm. We noticed that Rab-2A leaves the sperm during the acrosomal exocytosis. We also found a reduced Rab-2A protein level in sperm of patients with various pathologies of ejaculate. Based on studies describing the role of Rab-2A in acrosome biogenesis and the presence of Rab-2A in bovine sperm perinuclear theca, we tried to detect Rab-2A in this protein layer in human sperm. However, the chosen method is apparently not applicable to human sperm and isolation has failed. Additionally, we demonstrated the...
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Analysis of chemotherapeutics in extracellular vesicles
Kožnarová, Simona ; Hložková, Michaela (referee) ; Vašinová Galiová, Michaela (advisor)
Extracellular vesicles are a newly discovered way of cell-to-cell communication. The issue is still significantly unexplored, especially in the case of cancer. The goal of this pilot study was to attempt to detect platinum contained in extracellular vesicles by ICP-MS. In this study, samples of ovarian cancer cell lines treated with platinum derivatives, a type of chemotherapeutic drug, were used. This method was proved as able to detect platinum, even as able to determine its concentration reliably. The quantity of platinum in vesicles was around 1–2 % of the total platinum added to the system. The results varied according to the used platinum derivative, the cell line and the number of cells releasing the vesicles. Most platinum was determined in vesicles of the SK-OV-3 line, which is naturally resistant to this treatment, for all platinum derivatives. From these results it can be concluded that the use of ICP-MS is also advisable for additional research on this issue.
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Study of exosomes as drug delivery system in therapy of glioblastoma
Tomášková, Lucia ; Šimůnek, Tomáš (advisor) ; Macháček, Miloslav (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Lucia Tomášková Supervisor: prof. PharmDr. Tomáš Šimůnek, Ph.D. Title of diploma thesis: Study of exosomes as a drug delivery system in the treatment of glioblastoma Central nervous system disorders are among the most serious diseases affecting humans. They affect not only the patient's life, but also his/her surroundings. Therefore, their therapy, whether at the level of complete cure or alleviation of accompanying symptoms, is a challenge for scientific research. In our research, we focused on glioblastoma multiforme, a brain cancer not yet treatable. The main drawback in therapy is overcoming the blood-brain barrier. Exosomes, such as the body's natural nano-vesicles, have been shown to be a suitable system for delivering drugs to brain tissue. Our research has shown that by a suitable method we are able to obtain sufficient quality exosomes from macrophage and fill them very efficiently with antitumor agents paclitaxel, doxorubicin and temozolomide, while the delivered substances show higher efficacy and fewer side effects than the free form.
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First-trimester screening of pregnancy-related complications using plasma exosomal C19MC microRNAs
Špačková, Kamila ; Hromadníková, Ilona (advisor) ; Daňková, Pavlína (referee)
Pregnancy-related complications such as gestational hypertension, preeclampsia, fetal growth restriction, gestational diabetes mellitus, spontaneous preterm birth, and preterm premature rupture of membranes may have severe consequences for both the mother and the child. The development of reliable early screening methods for pregnancy-related complications has therefore been a long-term goal of obstetrics. New possibilities for prenatal diagnostics have opened with the discovery of circulating microRNAs in maternal plasma. MicroRNAs are short, noncoding, 21 to 23 nucleotides long, single-strand RNAs whose main function is to regulate gene expression. During pregnancy, both common and unique miRNAs are expressed by the placenta, amongst them the miRNAs of the C19MC cluster. Several C19MC miRNAs have been shown to display a different expression profile associated with certain pregnancy-related complications. This thesis identifies the plasma exosomal profiles of six C19MC miRNAs (miR-516-5p, miR-517-5p, miR-518b, miR-520a-5p, miR-520h, and miR-525-5p) in patients in their first trimester of gestation who later developed pregnancy-related complications, and compares them with profiles in patients with normal pregnancies.
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Study of exosomes in polyomavirus infection
Hyka, Lukáš ; Šroller, Vojtěch (advisor) ; Saláková, Martina (referee)
Exosomes are extracellular vesicles of endosomal origin. It was thought, that exosomes are used by cells only as carriers for cellular waste, but it was found out, that exosomes serve in the cellular communication and have a role in viral infections. Exosomes are exploited by viruses for example for the transport of viral protein or viral RNA/DNA. One of the viruses, where the mechanism of exploitation is unknown (if any exists) is murine polyomavirus. Murine polyomavirus belongs to the family Polyomaviridae, to which other human viruses belong for example, JC virus or virus of Merkel cell carcinoma. Murine polyomavirus codes for small, large and middle T antigen and three capsid proteins. Middle T antigen is known to bind to cellular membranes. Exosomes are membrane derived structures, so we investigated a possible transfer of middle T antigen. To this goal the successful isolation of exosomes and their characterization was necessary. Exosomes were isolated by ultracentrifugation and further purified by the density gradient OptiPrep. Exosomes were characterized by electron microscopy, NanoSight and by protein exosomal markers. These markers are for example Alix and flotillin-1. The cells were transfected in order to produce middle T antigen. It was shown, that exosomes isolated from these cells...
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Microvesicle and exosome detection in immune-related diseases
Šťastná, Evelína ; Drbal, Karel (advisor) ; Fabišik, Matej (referee)
Exosomes (ES) and microvesicles (MV), collectively called extracellular vesicles (EV), are submicroscopic vesicles encapsulated by a phospholipid bilayer. Smaller ES (40 - 100 nm) originate in endosomal compartment, while larger MV (50 - 1000 nm) shed from cell plasma membrane. EV are secreted by all types of cells. They consist of lipids and proteins, but their composition varies according to the cell they originate from. In addition, they differ in the cargo they transport (DNA, RNA and proteins). They occur in every bodily fluid in much higher amounts compared to the original cells themselves, what makes them an attractive and accessible biomarker of autoimmunity diseases, cardiovascular diseases or tumours. For detection of EV, sensitive flow cytometry (FCM) is used, which I am going to compare to alternative methodologies. Part of this work will be description of EV biogenesis and then I will focus on the role of EV in coagulation and inflammation related to autoimmune diseases, more specifically in rheumatoid arthritis (RA).
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Exosomes in viral infection and cancer
Sekavová, Alžběta ; Španielová, Hana (advisor) ; Hirsch, Ivan (referee)
Exosomes facilitate intercellular communication and transport of cellular cargo. Understanding the mechanisms underlying the cargo sorting to exosomes and the transport itself is crucial for vaccine development and diagnostic research. Exosome-mediated transfer contributes to immune response as well as progression of several diseases, including cancer and viral infections. Research on exosomes and their role in life cycles of tumorigenic viruses links already known mechanisms of viral carcinogenesis to the transport mechanisms of both cellular and viral proteins and nucleic acids. Epstein-Barr virus employs exosomes for transmission of the LMP1 oncoprotein and regulatory RNAs, whereas human immunodeficiency virus exploits cellular exosomal pathway for hijacking its membrane during budding, which helps it evade the immune system. It has been discovered that hepatitis C virus transfers its infectious virions between cells in exosomes. Exosomes containing oncoproteins and viral RNAs are also released from cells infected with other human tumorigenic viruses. However, mechanisms and implications of such events remain to be discovered. Keywords: exosome, cancer, viral infection, tumorigenic viruses, immunity, in- tercellular communication, hepatitis C virus, Epstein-Barr virus, human immuno- deficiency virus
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