Izražanje transkripcijskega dejavnika NANOG pri predrakavih spremembah in ploščatoceličnem raku ustne votline

Grubelnik, Gašper (avtor), Zidar, Nina (mentor), Boštjančič, Emanuela (komentor)

Ploščatocelični rak (PR) ustne votline se razvije preko ploščatoceličnih intraepitelijskih sprememb (PIS) kot posledica večstopenjskega procesa kopičenja somatskih mutacij in spremenjenega izražanja različnih genov, med katerimi je tudi NANOG. NANOG je transkripcijski dejavnik, ki je ključen v embrionalnem razvoju, kasneje pa sodeluje v razvoju različnih malignomov. Raziskali smo vlogo in diagnostični pomen ter regulacijo NANOG na proteinskem, RNA in DNA nivoju v PIS in PR ustne votline. V raziskavo smo vključili 367 biopsijskih tkivnih vzorcev 172 bolnikov s PR in PIS ustne votline ter 30 vzorcev normalne ustne sluznice. Uporabili smo imunohistokemijo, qPCR in sekvenciranje po Sangerju. Na nivoju mRNA je bil NANOG izražen v vseh vzorcih, izražanje je bilo najnižje v normalni sluznici in najvišje v PIS visoke stopnje, v PR se je izražanje ponovno znižalo. Protein v normalni sluznici ni bil izražen, ponovno se je aktiviral v procesu kancerogeneze, intenzivnost je naraščala od blede reakcije v PIS nizke stopnje do močnega izražanja v PIS visoke stopnje in PR. Regulatorne miRNA in lncRNA so pokazale korelacijo z izražanjem NANOG. Spremenjenega števila kopij NANOG in spremenjene metilacije promotorja NANOG nismo zaznali. Naši rezultati kažejo, da se izražanje NANOG na nivoju proteina utiša pred rojstvom in se ponovno izrazi v PIS visoke stopnje in PR, zato bi ga lahko uporabili kot diagnostični označevalec za natančnejšo opredelitev PIS. V procesu kancerogeneze se je izražanje NANOG mRNA razlikovalo od izražanja proteina NANOG, kar kaže na pomembno vlogo regulacije na posttranskripcijskem nivoju, z miRNA in lncRNA, poleg mRNA proteinskih regulatorjev kot ključnih regulatornih mehanizmov. Metilacija promotorja in spremenjeno število kopij imata pri regulaciji NANOG manjšo vlogo. Ugotovili smo tudi, da morfološko normalna sluznica ob raku že kaže spremembe v izražanju NANOG in njegovih regulatorjev in se torej pomembno razlikuje od morfološko normalne sluznice zdravih oseb. Z raziskavo smo prispevali nova spoznanja o vlogi NANOG v procesu razvoja PR ustne votline.

• Jezik: Slovenski jezik
• Ključne besede: ploščatocelični rak ustne votline, ploščatocelične intraepitelijske spremembe ustne votline, Nanog, transkripcijski dejavnik, diagnostični označevalec
• Vrsta gradiva: Doktorsko delo/naloga
• Organizacija: MF - Medicinska fakulteta
• Leto izida: 2021
• PID: 20.500.12556/RUL-127279
• COBISS.SI-ID: 65913347
• Datum objave v RUL: 02.06.2021

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Expression of transciption factor NANOG in precancerous lesions and squamous cell carcinoma of oral cavity

Izvleček

Oral squamous cell carcinoma (OSCC) develops from normal epithelium via squamous intraepithelial lesions (SILs) as result of a multistep process, characterized by accumulation of altered gene expression, including NANOG. NANOG is a transcription factor, playing crucial role in embryonal development, later it can contribute to cancer development. We analysed diagnostic significance and regulation of NANOG on protein, RNA and DNA levels in oral SILs and OSCC. Our study included 367 biopsy tissue samples from 172 patients with oral SCC and SILs, and 30 samples of normal oral mucosa. Immunohistochemistry, qPCR and Sanger sequencing were used. At mRNA level, NANOG was expressed in all samples, expression was low in normal mucosa and intensive in high grade SILs, in OSCC it was decreased. At protein level, NANOG was not found in normal mucosa, it was re-expressed in cancerogenesis, ranging from mild staining in low grade SILS to strong staining in high grade SILs and OSCC. Regulatory miRNAs and lncRNAs correlated with expression of NANOG, whereas copy number variation and promoter methylation were not changed. Our results suggest that expression of NANOG protein is silenced before birth and re-expressed in high grade SILs and OSCC and could be thus used as a diagnostic marker for classifying SILs. The expression of NANOG mRNA was different than the expression of NANOG protein, indicating an important role of post-transcription regulation, with miRNA and lncRNA as the key regulatory mechanisms beside protein regulators. Methylation of its promoter and copy number variation probably have a minor role in the regulation of NANOG expression. We also observed that morphologically normal mucosa adjacent to cancer showed an altered expression of NANOG and its regulators, in contrast to morphologically normal mucosa of healthy persons. Our results thus contributed new insights into the role of NANOG in the development of OSCC.

• Ključne besede: oral squamous cell carcinoma, oral squamous intraepithelial lesions, Nanog, transcription factor, diagnostic marker