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1.
Izolacija in biološki učinki potencialnih protektivnih oziroma probiotičnih bakterij v kozjem mleku in intestinalnem traktu
Martin Trapečar, 2009, diplomsko delo

Opis: Iz vzorcev kozjega mleka in fecesa smo uspešno izolirali 16 bakterijskih vzorcev in jih karakterizirali s pomočjo klasičnih in sodobnih mikrobioloških metod. Izmed izoliranih in karakteriziranih 16 bakterij jih 14 pripada rodu Lactobacillus ter 2 rodu Lactococcus. Dokazali smo pozitiven vpliv bakterij v in vitro celičnem modelu intestinalnega trakta. Rezultati so pokazali dobro stopnjo vezave na intestinalni epitelij, sorazmerno nizko citotoksičnost ter spodbujanje nespecifičnega imunskega odziva. S tem smo dokazali tudi zadostnost naših sevov za selekcijske kriterije. So funkcionalni, predvsem pa ne patogeni. Zanimivi za nadaljnje raziskave so predvsem vzorci M5K1 (Lactobacillus Brevis), F1K1K2 (Lactobacillus Plantarum) ter F1K1K1 (Lactococcus Rafinolactis), ki so najbolj izstopali po učinkovitosti. Navkljub dejstvu, da so vsi izolirani sevi pripadniki mlečnokislinskih bakterij je opazna vrstna specifičnost posameznih izoliranih sevov, ki se med seboj močno razlikujejo glede biološke aktivnosti. Prav tako prihaja do razhajanj znotraj posamezne vrste. Naše delo predstavlja v svetu edinstven eksperimentalni model za tovrstne analize.
Ključne besede: koze / probiotiki / celični model / vezava / imunski odziv
Objavljeno: 20.10.2009; Ogledov: 1815; Prenosov: 185
.pdf Celotno besedilo (1,49 MB)

2.
Biološki učinki naravnih mineralnih vod Radenska z ali brez rastlinskih ekstraktov v humanem celičnem modelu
Martin Trapečar, 2010, magistrsko delo

Opis: Naravne mineralne vode vsebujejo naravno raztopljene mineralne snovi in imajo zaradi tega dokazane številne prehransko-fiziološke učinke na človekov organizem. Ugotavljali smo specifičen učinek naravnih mineralnih vod Radenska Classic in Naturelle v humanih celičnih modelih v kombinaciji z ali brez ekstraktov vrste Malva sylvestris. Določali smo biološko dostopnost posameznih mineralov, antitumorsko in imunomodulatorno delovanje kombinacij naravnih mineralnih vod in rastlinskih ekstraktov kot tudi njihov vpliv na fiziologijo razvoja intestinalnega epitelija. Dokazali smo, da so minerali vod biološko visoko dosegljivi ter, da ugodno vplivajo na razvoj transepitelijske rezistence intestinalnega epitelija. Hkrati ugotavljamo da naravna mineralni vodi Radenska delujeta antiproliferativno in proapoptotično na celice adenokarcinoma debelega črevesa. Naravni mineralni vodi Radenska Naturelle ter Classic z ali brez ekstraktov gozdnega slezenovca dodatno spodbujata antiinflamatorni Th2 imunski odgovor z zaviranjem proinflamatornega TNF-α ter spodbujanjem antiinflamatornih IL-10 ter TGF-β. Prav tako, zaključujemo, da ekstrakti vrste Malva sylvestris zaradi njihove izjemne biološke aktivnosti še dodatno povečujejo biološki učinek naravnih mineralnih vod Radenska in predstavljajo velik potencial kot funkcionalni prehrambni dodatek.
Ključne besede: mineralna voda, Malva sylvestris, biološki učinki, celični modeli
Objavljeno: 23.10.2012; Ogledov: 1503; Prenosov: 77
.pdf Celotno besedilo (1,42 MB)

3.
UPORABA IN VALIDACIJA FUNKCIONALNIH CELIČNIH MODELOV NESPREMENJENIH CELIČNIH LINIJ ZA OCENO BIOLOŠKE AKTIVNOSTI IN ANALIZO TVEGANJA V BIOMEDICINI
Martin Trapečar, 2014, doktorska disertacija

Opis: Celični modeli iz nespremenjenih tkiv človeškega izvora ponazarjajo delovanje živih sistemov in pomembno dopolnjujejo ter nadgrajujejo konvencionalne predklinične raziskovalne metode. Njihova uporabnost sega vse od odkrivanja novih učinkovin, mehanističnih in toksikoloških študij do odkrivanja novih terapevtskih tarč. Poleg tega so in vitro modeli skladno z zakonskimi zaostritvami uporabe testov na živalih, združeni v načelu treh R-jev (Replacement, Reduction, Refinement) po katerem se zavezujemo k iskanju relevantnejših in učinkovitejših raziskovalnih modelov. Cilj doktorske naloge je bil razvoj, ovrednotenje in validacija novih celičnih modelov na podlagi nespremenjenih celičnih linij epitelija tankega črevesa, makrofagov ter skeletnega mišičevja. Uporabnost linij smo dokazovali v 2D modelih primernih za hitropretočne teste in toksikološke analize ter v 3D modelih primernih za natančnejše mehanistične in sistemske študije. Nespremenjeno celično kulturo tankočrevesnega epitelija smo uporabili za določanje varnosti potencialno patogenih sevov B. cereus in kasneje v kombinaciji s celično linijo makrofagov ustvarili 3D model razvijajočega se tankega črevesa v katerem smo preučevali imunomodulatorne lastnosti mlečnokislinskih bakterij. 2D in 3D model tankega črevesa kot primarnega interakcijskega mesta za hranila, mikroorganizme in druge ksenobiotike smo dopolnili s sekundarnim modelom skeletnega mišičevja, ki smo ga v kombinaciji z najnovejšo tehnologijo slikovne pretočne citometrije uporabili za določanje regenerativnega potenciala novih zdravil. Dobljene rezultate smo validirali s primerjavo testov na živalih ter dokazovanjem funkcionalnosti celičnih linij. Na ta način smo v svet in vitro predkliničnih raziskav, kjer prevladuje uporaba umetno transformiranih celičnih linij za ponazarjanje dejanskega in vivo okolja umestili nespremenjene linije, ki so zaradi fenotipske izraženosti in diferenciacije bistveno boljši preiskovalni model. S tem smo v duhu evropske zakonodaje in raziskovalne etike stroki ustvarili alternativo, ki zmanjšuje možnost napak v procesu predkliničnih preskušanj zaradi medvrstne raznolikosti in fenotipske neprimernosti ter odločno prispevali k zmanjševanju testov na živalih.
Ključne besede: Celična linija, Celični model, Ocena biološke aktivnosti, Analiza tveganja
Objavljeno: 11.09.2014; Ogledov: 1025; Prenosov: 67
.pdf Celotno besedilo (19,01 MB)

4.
5.
Application of gut cell models for toxicological and bioactivity studies of functional and novel foods
Martin Trapečar, Avrelija Cencič, 2012, pregledni znanstveni članek

Opis: The concept of functional and novel foods undoubtedly bears great potential as an asset to human health. However, this very same quest for ever new bioactive ingredients calls for reliable and distinct risk assessment as they may be potentially hazardous to human health. Most of today's methodologies still rely on decades old routines of animal trials and use of tumor-derived cell lines. Since such methodologies are not in line with the actual processesin the human body and with the 3R (replacement, reduction, refinement) concept, the results are often unreliable and misleading. Therefore, in this paper we propose the utilization of available untransformedsmall intestinal cell lines derived from human and pig tissue of non-tumor origin and describe several available cell models of the gut that offer a functional, close resemblance with the in vivo environment.
Ključne besede: gut, cell models, risk assessment, toxicology, functional food
Objavljeno: 10.07.2015; Ogledov: 452; Prenosov: 30
URL Povezava na celotno besedilo

6.
Skeletal muscle derived cell cultures as potent models in regenerative medicine research
Robi Kelc, Martin Trapečar, Matjaž Vogrin, Avrelija Cencič, 2013, pregledni znanstveni članek

Opis: Cell cultures have been used extensively by many scientists in the last decades to study various cell and tissue mechanisms. They have many advantagesover in vivo experimental models, but their limitations are to be considered as well. As skeletal muscle-derived cell cultures are becoming increasingly interesting in studies of muscle regeneration processes the question of their relevance in experiments arises according to in vivo experimental models. This article reviews studies that have simultaneously performed in vivo and in vitro experiments on skeletal muscle and discusses the correlation of both results. Although they seem to correlate, no such studies on humans have been performed so far.
Ključne besede: skeletal muscle cell, cell culture, In vivo, In vitro, experimental model
Objavljeno: 10.07.2015; Ogledov: 476; Prenosov: 40
URL Povezava na celotno besedilo

7.
A co-culture model of the developing small intestine offers new insight in the early immunomodulation of enterocytes and macrophages by Lactobacillus spp. through STAT1 and NF-kB p65 translocation
Martin Trapečar, Aleš Goropevšek, Mario Gorenjak, Lidija Gradišnik, Marjan Rupnik, 2014, izvirni znanstveni članek

Opis: The early establishment of a complete microbiome has been shown to play an integral part in the development and maintenance of an intact intestine and its immune system, although much remains unknown about the specific mechanisms of immune modulation in newborns. In our study we show in a co-culture model of the undeveloped small intestine that members of Lactobacillus spp. influence STAT1 and NF-kB p65 nuclear translocation in both intestinal epithelial cells as well as underlying macrophages. Moreover, by using imaging flow cytometry we were able to monitor each individual cell and create a framework of the percentage of cells in which translocation occurred in challenged versus control cell populations. We also observed a significant difference in baseline translocation in intestinal cells when cultured alone versus those in a co-culture model, underpinning the importance of 3D models over monolayer set-ups in epithelial in vitro research. In conclusion, our work offers new insights into the potential routes by which the commensal microbiome primes the early immune system to fight pathogens, and shows how strain-specific these mechanisms really are.
Ključne besede: microbiome, Lactobacillus, immune system, pathogens
Objavljeno: 19.06.2017; Ogledov: 206; Prenosov: 109
.pdf Celotno besedilo (2,86 MB)
Gradivo ima več datotek! Več...

8.
Platelet-rich plasma, especially when combined with a TGF-ß inhibitor promotes proliferation, viability and myogenic differentiation of myoblasts in vitro
Robi Kelc, Martin Trapečar, Lidija Gradišnik, Marjan Rupnik, Matjaž Vogrin, 2015, izvirni znanstveni članek

Opis: Regeneration of skeletal muscle after injury is limited by scar formation, slow healing time and a high recurrence rate. A therapy based on platelet-rich plasma (PRP) has become a promising lead for tendon and ligament injuries in recent years, however concerns have been raised that PRP-derived TGF-β could contribute to fibrotic remodelling in skeletal muscle after injury. Due to the lack of scientific grounds for a PRP -based muscle regeneration therapy, we have designed a study using human myogenic progenitors and evaluated the potential of PRP alone and in combination with decorin (a TGF-β inhibitor), to alter myoblast proliferation, metabolic activity, cytokine profile and expression of myogenic regulatory factors (MRFs). Advanced imaging multicolor single-cell analysis enabled us to create a valuable picture on the ratio of quiescent, activated and terminally committed myoblasts in treated versus control cell populations. Finally high-resolution confocal microscopy validated the potential of PRP and decorin to stimulate the formation of polynucleated myotubules. PRP was shown to down-regulate fibrotic cytokines, increase cell viability and proliferation, enhance the expression of MRFs, and contribute to a significant myogenic shift during differentiation. When combined with decorin further synergistc effects were identified. These results suggest that PRP could not only prevent fibrosis but could also stimulate muscle commitment, especially when combined with a TGF-β inhibitor.
Ključne besede: muscles, skeletal, injuries, TGF-beta, plasma, thrombocytes, myoblasts, fibrosis, prevention, regeneration
Objavljeno: 19.06.2017; Ogledov: 283; Prenosov: 39
.pdf Celotno besedilo (2,07 MB)
Gradivo ima več datotek! Več...

9.
Single-cell analysis reveals IGF-1 potentiation of inhibition of the TGF-ß/Smad pathway of fibrosis in human keratocytes in vitro
Tomislav Šarenac, Martin Trapečar, Lidija Gradišnik, Marjan Rupnik, Dušica Pahor, 2016, izvirni znanstveni članek

Opis: Corneal wound healing is often affected by TGF-β–mediated fibrosis and scar formation. Guided fibrosis with IGF-1 and antifibrotic substances might maintain corneal transparency. Primary human corneal keratocytes under serum-free conditions were used as a model of corneal stromal wounding, with markers of corneal fibrosis and opacity studied under TGF-β2 stimulation. Single-cell imaging flow cytometry was used to determine nuclearization of Smad3, and intracellular fluorescence intensity of Smad7 and the corneal crystallin aldehyde dehydrogenase 3A1. Extracellular matrix proteoglycans keratocan and biglycan were quantified using ELISAs. On the TGF-β2 background, the keratocytes were treated with IGF-1, and suberoylanilidehydroxamic acid (SAHA) or halofuginone ± IGF-1. IGF-1 alone decreased Smad3 nuclearization and increased aldehyde dehydrogenase 3A1 expression, with favorable extracellular matrix proteoglycan composition. SAHA induced higher Smad7 levels and inhibited translocation of Smad3 to the nucleus, also when combined with IGF-1. Immunofluorescence showed that myofibroblast transdifferentiation is attenuated and appearance of fibroblasts is favored by IGF-1 alone and in combination with the antifibrotic substances. The TGF-β/Smad pathway of fibrosis and opacity was inhibited by IGF-1, and further with SAHA in particular, and with halofuginone. IGF-1 is thus a valid aid to antifibrotic treatment, with potential for effective and transparent corneal wound healing.
Ključne besede: cornea, wounds, treatment, antifibrotic treatment, keratocytes
Objavljeno: 23.06.2017; Ogledov: 361; Prenosov: 42
.pdf Celotno besedilo (1,89 MB)
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