Vpliv mikrookolja, izoliranega z lasersko mikrodisekcijo, na izražanje izbranih molekularnih dejavnikov kronične vnetne črevesne bolezni na mišjem modelu

Unkovič, Ana (avtor), Perše, Martina (mentor)

Kljub intenzivnemu raziskovanju kronične vnetne črevesne bolezni (KVČB) so v literaturi pomanjkljive in nasprotujoče informacije o izražanju molekularnih dejavnikov KVČB v odvisnosti od lokacije (po plasteh), histološke slike, mikrookolja in faze vnetja v črevesni steni. Namen naše raziskave je bil ovrednotiti vpliv teh dejavnikov v steni debelega črevesa pri KVČB na mišjem modelu. Preveriti smo želeli razlike v izražanju izbranih molekularnih dejavnikov med histološko zdravo črevesno mukozo in mukozo pri KVČB in razlike med posameznimi plastmi histološko zdrave črevesne stene. Zanimalo nas je tudi izražanje izbranih molekularnih dejavnikov v/med posameznimi plastmi črevesne stene pri različni histološki sliki bolezenskega procesa. V raziskavo smo vključili parafinske tkivne bloke debelega črevesa miši, ki so imele z dekstran natrijevim sulfatom povzročen kolitis. Odrezali smo cele rezine vzorcev ali z lasersko mikrodisekcijo izrezali področja znotraj posameznih plasti črevesne stene. Preverili smo izražanje mRNA tarčnih genov (TNF?, TNFR, DUOX2, NOS2) ter miRNA (mi-223-3p, miR-181a-5p ter miR3473b) in z multivariatno analizo preverili njihove medsebojne korelacije. Rezultati so pokazali, da se izražanje izbranih molekularnih dejavnikov lahko razlikuje v odvisnosti od lokacije (med posameznimi plastmi črevesne stene), od faze vnetja (se razlikuje v histološko zdravi črevesni mukozi med zdravimi in bolnimi mišmi), ter od histološke slike (med posameznimi plastmi stene debelega črevesa v različni histološki sliki bolezenskega procesa). Pokazali smo tudi, da izražanje, določeno v celih rezinah, ne odraža nujno izražanja v mukozi ali drugih plasteh črevesne stene ter da je vzorec izražanja specifičen za tarčno RNA. Kot prvi smo podali vpogled v izraženost izbranih molekularnih dejavnikov glede na mikrookolja debelega črevesa, s poudarkom na preiskovanju posameznih plasti z različno histološko sliko sprememb ter fazo vnetja. S tem smo pridobili vpogled v medsebojna razmerja preiskovanih molekularnih dejavnikov in ponudili razloge za razhajanje podatkov v literaturi.

• Jezik: Slovenski jezik
• Ključne besede: kronična vnetna črevesna bolezen, laserska mikrodisekcija, mikrookolje, mišji model, dekstran natrijev sulfat.
• Vrsta gradiva: Doktorsko delo/naloga
• Organizacija: MF - Medicinska fakulteta
• Leto izida: 2023
• PID: 20.500.12556/RUL-151747
• Datum objave v RUL: 19.10.2023

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Influence of the microenvironment isolated by laser microdissection on the expression of selected molecular factors of inflammatory bowel disease in a mouse model

Izvleček

Despite intensive research of inflammatory bowel disease (IBD), there is insufficient and conflicting information in the literature about the expression of IBD molecular factors depending on the location (individual layers), histological picture, microenvironment and stage of inflammation in the intestinal wall. We wanted to check the differences in the expression of selected molecular factors between the histologically healthy intestinal mucosa and the mucosa in IBD and the differences between the individual layers of the histologically healthy intestinal wall. We were also interested in the expression of selected molecular factors in/between individual layers of the intestinal wall in different histological patterns of the disease process. We used paraffin-embedded samples of mice colon with dextran sodium sulfate-induced colitis in the study. Whole slices of samples were cut or areas within individual layers of the intestinal wall were excised using laser microdissection. We checked the mRNA expression of target genes (TNFα, TNFR, DUOX2, NOS2) and miRNAs (mi-223-3p, miR-181a-5p and miR3473b) and checked their mutual correlations using multivariate analysis. Results showed that the expression of selected molecular factors can vary depending on the location (between individual layers of the intestinal wall), on the stage of inflammation (histologically healthy intestinal mucosa between healthy and diseased individuals), and on the histological picture (a different histological picture of the disease process). We also showed that the expression determined in whole slices does not necessarily reflect the expression in the mucosa or other layers of the intestinal wall and that the expression pattern is target RNA specific. As first we provided an insight into the expression of selected molecular factors in relation to the colon microenvironments, with an emphasis on the investigation of individual layers with different histological picture and the stage of inflammation. This gave us an insight into the relations of the investigated molecular factors and offered reasons for the divergence of data in the literature.

• Ključne besede: inflammatory bowel disease, lasser microdisection, microenvironment, mouse model, dextran sodium sulfat.