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Rak sečnega mehurja je deseto najpogostejše rakavo obolenje na svetu. Uporaba humane amnijske membrane (hAM) za zdravljenje raka je ena od perspektivnih idej, ki se je pojavila v zadnjih letih. Cilj doktorske disertacije je bil proučiti celično-biološke mehanizme protirakavega delovanja homogenata in izvlečka hAM na modelih raka in normalnega sečnega mehurja in vitro. Z uporabo različnih molekularnih in mikroskopskih metod smo ugotovili, da pripravka iz hAM zmanjšata izražanje označevalcev fokalnih stikov, migracije, invazije in proliferacije mišično invazivnih urotelijskih celic T24 in neinvazivnih urotelijskih celic papiloma RT4. Dokazali smo, da je eden izmed mehanizmov protirakavega delovanja pripravkov iz hAM inhibicija FAK in p-FAK ter posledično zmanjšanje izražanja genov in proteinov, ki so del signalne poti PI3K/Akt/mTOR v rakavih urotelijskih celicah T24. Poleg tega smo dokazali, da homogenat hAM v mišično invazivnih urotelijskih celicah T24 in urotelijskih celicah papiloma RT4 zavre izražanje proteinov, ki so vpleteni v reorganizacijo aktinskega citoskeleta. Naša raziskava je potrdila, da pripravka iz hAM nimata škodljivega učinka na normalne urotelijske celice. Ugotovili smo, da proučevana pripravka iz hAM delujeta na več molekularnih tarč karcinogeneze, na osnovi česar menimo, da imata potencial za uporabo, kot dodatni protirakavi učinkovini, v večtarčni intravezikalni terapiji po transuretralni resekciji tumorjev sečnega mehurja. Bladder cancer is the tenth most common cancer. Using human amniotic membrane (hAM) to treat cancer is one of the promising ideas that have emerged in recent years. The aim of the PhD thesis was to investigate the effect of hAM homogenate and extract on different in vitro models of bladder cancer. Using molecular and microscopic methods, we have shown that hAM homogenate and extract reduce the expression of markers of focal adhesion, migration, invasion and proliferation in muscle-invasive urothelial T24 cells and transitional cell papilloma RT4 cells. In addition, we have shown that one of the mechanisms of action is through downregulation of genes and proteins that are part of the FAK/PI3K/Akt/mTOR pathway in T24 cancer cells. We have also shown that the hAM homogenate inhibits the expression of focal adhesion proteins and proteins involved in the reorganization of the actin cytoskeleton of cancer urothelial cells. Our research has also confirmed that the hAM preparations have no harmful effects on normal urothelial cells. In conclusion, we have shown that hAM preparations have multitargeted anticancer effect, which further supports the idea of a possible clinical application of hAM homogenate and extract as additional anti-cancer agents in multi-target intravesical therapy after transurethral resection of bladder tumors.

Urinary tract infections (UTIs) represent a serious global health issue, especially due to emerging multidrug-resistant UTI-causing bacteria. Recently, we showed that the human amniotic membrane (hAM) could be a candidate for treatments and prevention of UPEC and Staphylococcus aureus infections. However, its role against multidrug-resistant bacteria, namely methicillin-resistant S. aureus (MRSA), extended-spectrum beta-lactamases (ESBL) producing Escherichia coli and Klebsiella pneumoniae, vancomycin-resistant Enterococci (VRE), carbapenem-resistant Acinetobacter baumannii, and Pseudomonas aeruginosa has not yet been thoroughly explored. Here, we demonstrate for the first time that the hAM homogenate had antibacterial activity against 7 out of 11 tested multidrug-resistant strains, the greatest effect was on MRSA. Using novel approaches, its activity against MRSA was further evaluated in a complex microenvironment of normal and cancerous urinary bladder urothelia. Even short-term incubation in hAM homogenate significantly decreased the number of bacteria in MRSA-infected urothelial models, while it did not affect the viability, number, and ultrastructure of urothelial cells. The hAM patches had no antibacterial activity against any of the tested strains, which further exposes the importance of the hAM preparation. Our study substantially contributes to basic knowledge on the antibacterial activity of hAM and reveals its potential to be used as an antibacterial agent against multidrug-resistant bacteria.