Small extracellular vesicles as microRNA carriers and their role in neural cell regeneration

Acute spinal cord injury is a serious type of injury, the treatment of which still represents a challenge for contemporary medicine. Neural stem cells (NSCs) transplantation is one of many promising ways to contribute to the regeneration of damaged tissue. NSCs communicate with the rest of the tissue by means of small extracellular vesicles (sEVs), in which their regenerative potential is thus assumed. The aim of this thesis is to verify the antiapoptotic effect of sEVs isolated from the culture medium of two types of NSCs in an rat in vitro model of spinal cord injury. To meet this goal, different methods of isolating sEVs from the culture medium were tested. Subsequently, sEVs were characterized according to their size and the presence of surface markers. This thesis includes an PCR analysis of the cargo of sEVs, which showed an abundance of neuroprotective and antiapoptotic miRNAs. Fluorescent staining of sEVs proved that sEVs penetrate into the cytoplasm of stem cells. Finally, sEVs were applied to a rat in vitro model of spinal cord injury and their neuroprotective effect was demonstrated using the immunoblotting method. Keywords Small extracellular vesicles, exosomes, acute spinal cord injury, regeneration of nervous tissue, miRNA, microRNA, neural stem cellsAkutní poranění míchy je závažný typ úrazu, jehož léčba stále představuje výzvu pro současnou medicínu. Transplantace neurálních kmenových buněk (NSC) je jednou z mnoha nadějných cest, jak přispět k regeneraci poškozené tkáně. NSC komunikují se zbytkem tkáně pomocí malých extracelulárních váčků (sEVs), u kterých se tak předpokládá jejich regenerační potenciál. Cílem této diplomové práce je ověřit antiapoptotický účinek sEVs izolovaných z kultivačního média od dvou typů NSC na potkaním in vitro modelu poranění míchy. Pro splnění tohoto cíle byly nejprve otestovány různé způsoby izolací sEVs z kultivačního média. Následně byly sEVs charakterizovány podle jejich velikostí a přítomností povrchových markerů. Součástí práce byla PCR analýza obsahu neuroprotektivních a antiapoptotických miRNA uvnitř sEVs a fluorescenční barvení sEVs, kterým bylo dokázáno, že sEVs pronikají do cytoplazmy kmenových buněk. Nakonec byly sEVs aplikovány na potkaní in vitro model poranění míchy a pomocí metody imunoblotace byl prokázán jejich neuroprotektivní účinek. Klíčová slova Malé extracelulární váčky, exosomy, akutní poranění míchy, regenerace nervové tkáně, miRNA, mikroRNA, neurální kmenové buňkyDepartment of PhysiologyKatedra fyziologiePřírodovědecká fakultaFaculty of Scienc

The role of miRNA in injury and regeneration of spinal cord tissue

MikroRNA jsou malé nekódující molekuly RNA o délce zhruba 20-24 nukleotidů, které post-transkripčně regulují genovou expresi. Interferují s mRNA přes párování bází v komplementárních sekvencích. V posledních letech se ukazuje jejich významná role při poranění a regeneraci nervové tkáně. Cílem této bakalářské práce je popsat možnou úlohu miRNA při poranění centrální nervové soustavy se zaměřením na poranění míchy.MicroRNAs are small non-coding RNA molecules of a length about 20-24 nucleotides, that regulate gene expression post-transcriptionally. They interfere mRNA molecules via base- pairing with complementary sequences. Recently it was shown that they play an important role in injury and regeneration of nervous tissue. The aim of this bachelor thesis is to describe possible role of miRNAs in central nervous system injury with focus on spinal cord injury.Katedra fyziologieDepartment of PhysiologyPřírodovědecká fakultaFaculty of Scienc

The role of miRNA in injury and regeneration of spinal cord tissue

MicroRNAs are small non-coding RNA molecules of a length about 20-24 nucleotides, that regulate gene expression post-transcriptionally. They interfere mRNA molecules via base- pairing with complementary sequences. Recently it was shown that they play an important role in injury and regeneration of nervous tissue. The aim of this bachelor thesis is to describe possible role of miRNAs in central nervous system injury with focus on spinal cord injury

Blue Box

The bachelor's thesis Blue Box follows a long-term project that began in 2013, when I was on my way to the east coast of the USA. I began creating a mystical Blue Box story around the mythology and ideology of xenophobic movements, after a mystical experience in New York's Orthodox Jewish Quarter and attending a Ku Klux Klan public gathering in Gettysburg. In thematic terms, my work is a universal report about the dangers of extremism in any form, focusing on the essence and mechanisms of its origin and spread

The role of miRNA in injury and regeneration of spinal cord tissue

MicroRNAs are small non-coding RNA molecules of a length about 20-24 nucleotides, that regulate gene expression post-transcriptionally. They interfere mRNA molecules via base- pairing with complementary sequences. Recently it was shown that they play an important role in injury and regeneration of nervous tissue. The aim of this bachelor thesis is to describe possible role of miRNAs in central nervous system injury with focus on spinal cord injury

Nickel alloys in aerospace

Letecký a kosmický průmysl je v dnešní době jedním z nejrychleji se vyvíjejících průmyslových odvětví. Zvyšující se nároky na efektivitu a spotřebu paliva proudových motorů lze uspokojit mimo jiné za použití kvalitnějších materiálů, mezi které jednoznačně patří superslitiny na bázi niklu, jež tyto požadavky dokáží splnit. Tato práce podává přehled právě o těchto materiálech: a to jak o stavebním kameni celé struktury – niklu, superslitinách obecně, tak především těch na niklové bázi. Pojednává o složení mikrostruktury tvořené austenitickou matricí ( fází) nejčastěji vytvrzovanou karbidickými nebo substitučními precipitáty, jedinečných mechanických vlastnostech, ale také vlivu vysokých teplot a creepu na degradaci těchto vlastností. Práce také obsahuje srovnání vybraných superslitin z různých hledisek, a to především podle typu částic, jež vytvrzují matrici, nebo vlivu zatížení na superslitinu v přítomnosti tepelně-oxidačního prostředí. Dle oblasti použití jsou tyto materiály předurčeny pro další budoucí vývoj a zdokonalování, protože jejich unikátní vlastnosti v oblastech s extrémními podmínkami jsou prozatím nenahraditelné.The aerospace industry is one of the fastest growing industries today. Increasing demands on the efficiency and fuel consumption of jet engines can be met, among other things, by using higher quality materials, which clearly include nickel-based superalloys, which can meet these demands. This work provides an overview of these materials: both the building block of the whole structure - nickel, superalloys in general, and especially those nickel-based. It deals with the composition of the microstructure formed by the austenitic matrix ( phases) most often cured by carbide or substitution precipitates, unique mechanical properties, but also the influence of high temperatures and creep on the degradation of these properties. The work also includes a comparison of selected superalloys from different perspectives, especially according to the type of particles that cure the matrix or the effect of loading on the superalloy in the presence of thermal oxidation environment. Depending on the area of application, these materials are predestined for further future development and improvement, because their unique properties in areas with extreme conditions are irreplaceable for the time being.

Small extracellular vesicles as microRNA carriers and their role in neural cell regeneration

Acute spinal cord injury is a serious type of injury, the treatment of which still represents a challenge for contemporary medicine. Neural stem cells (NSCs) transplantation is one of many promising ways to contribute to the regeneration of damaged tissue. NSCs communicate with the rest of the tissue by means of small extracellular vesicles (sEVs), in which their regenerative potential is thus assumed. The aim of this thesis is to verify the antiapoptotic effect of sEVs isolated from the culture medium of two types of NSCs in an rat in vitro model of spinal cord injury. To meet this goal, different methods of isolating sEVs from the culture medium were tested. Subsequently, sEVs were characterized according to their size and the presence of surface markers. This thesis includes an PCR analysis of the cargo of sEVs, which showed an abundance of neuroprotective and antiapoptotic miRNAs. Fluorescent staining of sEVs proved that sEVs penetrate into the cytoplasm of stem cells. Finally, sEVs were applied to a rat in vitro model of spinal cord injury and their neuroprotective effect was demonstrated using the immunoblotting method. Keywords Small extracellular vesicles, exosomes, acute spinal cord injury, regeneration of nervous tissue, miRNA, microRNA, neural stem cell

Nickel alloys in aerospace

The aerospace industry is one of the fastest growing industries today. Increasing demands on the efficiency and fuel consumption of jet engines can be met, among other things, by using higher quality materials, which clearly include nickel-based superalloys, which can meet these demands. This work provides an overview of these materials: both the building block of the whole structure - nickel, superalloys in general, and especially those nickel-based. It deals with the composition of the microstructure formed by the austenitic matrix ( phases) most often cured by carbide or substitution precipitates, unique mechanical properties, but also the influence of high temperatures and creep on the degradation of these properties. The work also includes a comparison of selected superalloys from different perspectives, especially according to the type of particles that cure the matrix or the effect of loading on the superalloy in the presence of thermal oxidation environment. Depending on the area of application, these materials are predestined for further future development and improvement, because their unique properties in areas with extreme conditions are irreplaceable for the time being