Stupanj metilacije gena ASC/TMS1 i MyD88 u uzorcima zdravog tkiva i tkiva karcinoma pločastih stanica grkljana

Kovač Bilić, Lana (2018) Stupanj metilacije gena ASC/TMS1 i MyD88 u uzorcima zdravog tkiva i tkiva karcinoma pločastih stanica grkljana. PhD thesis, Sveučilište u Zagrebu.

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Abstract

Laryngeal Cancer is the most frequent Head and Neck Cancer comprising high mortality rate. Almost 90% of all Laryngeal Cancers develop due to genetical and epigenetical changes caused by lifestyle and environmental factors. More than 95% of all laryngeal cancers are Squamous Cell Carcinoma. To date there are no established biomarkers to facilatate the diagnosis, prognosis and treatment of patients suffering from this malignant disease. Recent studies showed that inadequate control of inflammation and presence of infection are tightly related with developing and promotion of the cancer. Innate immue system has a very important role in the control of inflammation. Innate immunity defense mechanisms activate pattern recognition receptors (PRRs) and adaptor molecules play the key role in all signalling pathways. In this study, we hypothesized that the changes in the methylation status of promotor regions of ASC/TMS1 and MyD88 genes, responsible for activation and regulation of inflammation, in healthy and laryngeal cancer tissue might be related with development and progression of cancer. The aim of this study is to investigate is there a difference in the methylation status of this genes in healthy and tumor tissue and does it correlate with protein expression. In recent literature the methylation status of this genes has still not been analyzed in Laryngeal Cancer and the pyrosequencing method in methylation status determination as performed in this study has not been used in other cancers. Results of this study show that the overall methylation level of the promotor region of MyD88 gene is statistically significantly higher in healthy laryngeal tissue in relation to cancer tissue and this finding correlate with protein expression level. The overall methylation level of promotor region of ASC/TMS1 gene is unchanged in Laryngeal Cancer and healthy laryngeal tissue. The protein expression level of ASC/TMS1 is statistically significantly higher in cancer in relation to healthy laryngeal tissue. Taking into account there is no difference in the overall methylation level of the promotor region of ASC/TMS1 gene in healthy and tumor tissue we assume that the observed difference in ASC/TMS1 protein expression (higher expression in tumor tissue) is the outcome of other posttranscriptional regulation mechanisms. We consider that the results of this study in the future may lead to earlier detection, treatment and follow-up of Laryngeal Squamous Cell Carcinoma and reveal new scientific discoveries of the role of ASC/TMS1 and MyD88 genes.

Abstract in Croatian

Karcinom grkljana je najčešći karcinom glave i vrata sa visokom stopom mortaliteta. Gotovo 90% karcinoma grkljana nastaje uslijed genetskih i epigenetskih promjena uzrokovanih načinom života i čimbenicima okoliša. Više od 95% karcinoma grkljana patohistološki su karcinomi pločastih stanica. Do danas ne postoje pouzdani klinički etablirani tumorski biljezi koji bi olakšali dijagnozu, prognozu i liječenje oboljelih od ove maligne bolesti. Novijaistraživanja pokazuju da su poremećena kontrola upalne reakcije i prisutnost infekcije povezani s nastankom i razvojem tumorskih bolesti. Važnu ulogu u kontroli upalne reakcije ima urođena imunost. Obrambeni mehanizam urođene imunosti očituje se aktivacijom receptora odgovornih za prepoznavanje patogena, a ključnu ulogu u tom procesu imaju adaptorne molekule signalnih puteva. U ovom istraživanju pretpostavili smo da bi postojanje promijenjenog statusa metilacije promotorske regije gena ASC/TMS1 i gena MyD88, odgovornih za aktivaciju i regulaciju upale, unutar zdravog i tumorskog tkiva grkljana moglo biti povezano s nastankom i progresijom tumora. Cilj istraživanja je ispitati postoji li razlika u stupnju metilacije ovih gena zdravog i tumorskog tkiva grkljana i očituje li se ta razlika na proteinskoj razini. Status metilacije ovih gena u karcinomu grkljana do sada nije proučavan, niti je u bilo kojem od karcinoma korištena metoda pirosekvenciranja za određivanje statusa metilacije kao u ovom istraživanju. Rezultati ovog istraživanja pokazuju da je ukupna metilacija promotorske regije gena MyD88 statistički značajno povećana u zdravim tkivima grkljana u odnosu na tumorska tkiva i taj nalaz korelira s ekspresijom proteina. Ukupna metilacija promotorske regije gena ASC/TMS1 nepromijenjena je u tumorskom u odnosu na zdravo tkivo grkljana. Ekspresija proteina ASC/TMS1 je statistički značajno povećana u tumorskom tkivu u odnosu na zdravo tkivo grkljana. Obzirom da ne postoji razlika u stupnju ukupne metilacije promotorske regije gena ASC/TMS1 u zdravom i tumorskom tkivu, pretpostavljamo da je uočena razlika u ekspresiji proteina ASC/TMS1 (povećana ekspresija u tumorskom tkivu) posljedica nekih drugim posttranskripcijskih mehanizama regulacije. Smatramo da bi rezultati ovog istraživanja u budućnosti mogli doprinjeti ranijem otkrivanju, terapiji i praćenju karcinoma pločastih stanica grkljana ali i doprinjeti novim spoznajama o ulozi gena ASC/TMS1 i MyD88.

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