Določitev sposobnosti preživetja primarnih celic iz različnih tkiv ortopedskih bolnikov po daljšem obdobju krioprezervacije

Trifkovič, Maja (avtor), Zupan, Janja (mentor), Stražar, Klemen (komentor)

Mezenhimske matične/stromalne celice (MSC) so multipotente celice, ki imajo določene lastnosti, kot so sposobnost samoobnavljanja, proliferacije, diferenciacije in parakrinega delovanja, zaradi katerih so lahko koristne pri zdravljenju različnih bolezni. Izoliramo jih lahko iz različnih tkiv odraslih organizmov. Njihova sposobnost krioprezervacije pa predstavlja pomemben biotehnološki izziv pri pripravi naprednih celičnih terapij s temi celicami. V magistrski nalogi smo preučevali sposobnost preživetja primarnih celic, izoliranih iz različnih tkiv kolka (kostnega tkiva in treh različnih področij sinovijske ovojnice) po daljšem obdobju krioprezervacije. Tkiva so odvzeli 13 ortopedskim bolnikom med artroskopijo kolka (skupno 49 vzorcev), iz njih izolirali primarne celice, nato pa jih sveže zamrznili in shranili v tekočem dušiku približno pet do šest let. Te celice smo odmrznili, gojili na gojilnih ploščah ter pod mikroskopom opazovali njihovo rast in tvorbo kolonij. Ugotovili smo, da so nekatere celice sposobne preživeti večletno shranjevanje v tekočem dušiku, saj je bila skupna uspešnost krioprezervacije 24,5 %. Največjo sposobnost tvorbe kolonij so izkazale celice, izolirane iz kostnega tkiva (60 %), veliko manjšo pa celice, izolirane iz sinovijske ovojnice (iz vseh treh področij skupno 15,4 %). Izkazalo se je, da izvor celic vpliva na uspešnost njihove krioprezervacije (p = 0,03994). V 12 vzorcih, ki smo jih uspešno odmrznili po krioprezrvaciji, smo prešteli med 5000 in 82 500 celic na mL vzorca. Korelacija med številom preštetih celic in trajanjem krioprezervacije ni bila statistično značilna (p = 0,7546, r = -0,1011), zato povezave med številom celic in časom shranjevanja v tekočem dušiku nismo potrdili. Pri večini drugih podobnih raziskav na tem področju so celice shranjevali krajši čas. Naši rezultati pa kažejo, da je primarne celice iz različnih tkiv kolka mogoče shraniti tudi dalj časa. Dokazali smo, da te celice tudi po več letih shranjevanja v tekočem dušiku ohranijo sposobnost tvorbe kolonij, pri čemer so bolj primerne za tako shranjevanje primarne celice iz kostnega tkiva. Naši rezultati pomembno prispevajo k optimizaciji postopkov krioprezervacije primarnih celic pri izdelavi zdravil za napredno zdravljenje.

• Jezik: Slovenski jezik
• Ključne besede: mezenhimske matične/stromalne celice (MSC), kostno tkivo, sinovijska ovojnica, krioprezervacija
• Vrsta gradiva: Magistrsko delo/naloga
• Tipologija: 2.09 - Magistrsko delo
• Organizacija: FFA - Fakulteta za farmacijo
• Založnik: [M. Trifkovič]
• Leto izida: 2026
• PID: 20.500.12556/RUL-183809
• UDK: 615:60:576(043.2)
• COBISS.SI-ID: 282531843
• Datum objave v RUL: 19.06.2026

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Determination of survival of primary cells from various tissues obtained from orthopaedic patients after prolonged cryopreservation

Izvleček

Mesenchymal stem/stromal cells (MSC) are multipotent cells that possess key properties such as self-renewal, proliferation, differentiation and paracrine activity which make them promising for the treatment of various diseases. They can be isolated from different tissues of adult organisms. Their ability to undergo cryopreservation represents an important biotechnological challenge in the development of advanced cell therapies using these cells. In this master’s thesis we studied the survival ability of primary cells isolated from different hip tissues (bone tissue and three different areas of the synovial membrane) following prolonged cryopreservation. These tissues were obtained from 13 orthopedic patients during hip arthroscopy (a total of 49 samples), primary cells were isolated, freshly frozen and stored in liquid nitrogen approximately five to six years. After thawing, the cells were cultured on growth plates, and their proliferation and colony forming ability were evaluated using a microscopic observation. We found that some cells were capable of surviving long-term storage in liquid nitrogen, with an overall cryopreservation success rate of 24,5 %. Cells isolated from bone exhibited the highest colony-forming ability (60 %), whereas cells derived from the synovial membrane showed markedly lower capacity (15,4 % when all three synovium areas were combined). The results indicated that the tissue of origin significantly influences the cryopreservation success (p = 0,03994). In 12 successfully recovered samples, cell counts ranged from 5,000 to 82,500 cells per mL. No statistically significant correlation was observed between cell number and the duration of cryopreservation (p = 0,7546, r = -0,1011), indicating that storage time in liquid nitrogen did not significantly affect cell yield. In most comparable studies, the cells have been cryopreserved for shorter periods. However, our results indicate that these cells can also withstand long-term storage, since they retained their colony-forming abilities even after several years in liquid nitrogen. Notably, primary cells derived from bone tissue appear to be more suitable for extended cryopreservation than those obtained from the synovial membrane. Our results make a significant contribution to the optimization of cryopreservation procedures for primary cells in the production of advanced therapy medicinal products.

• Ključne besede: mesenchymal stem/stromal cells (MSC), bone tissue, synovial membrane, cryopreservation