2016, vol. 66, br. 2, str. 222-233
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Ekspresija KIT receptora na kutanim mastocitomima pasa bez c-kit mutacija
KIT receptor expression in canine cutaneous mast cell tumors (CMCTS) without c-kit mutation
Projekat: Molekularno-genetička i ekofiziološka istraživanja u zaštiti autohtonih animalnih genetičkih resursa, očuvanju dobrobiti, zdravlja i reprodukcije gajenih životinja i proizvodnji bezbedne hrane (MPNTR - 46002)
Sažetak
Histopatološko ispitivanje, gradacija, imunohistohemijsko bojenje i molekularno genetska ispitivanja su predloženi kriterijumi za klasifikaciju kutanih mastocitoma pasa. Prisustvo aberantne ekspresije CD117 i mutacija c-kit protoonkogena moglo bi da bude indikativni parameter za finalnu histopatološku klasifikaciju mastocitoma. Glavni cilj ispitivanja je određivanje povezanosti mesta ekspresije KIT receptora i histološkog gradusa mastocitoma bez c-kit mutacija. Ispitivanjima su obuhvaćena 24 mastocitoma i 6 kontrolnih isečaka kože uzorkovanih od ukupno 30 odraslih pasa različitog uzrasta, rase i pola. Uzorci fiksirani u formalinu i ukalupljeni u parafinske kalupe su bojeni hematoksilin-eozinom, toluidin plavo metodom i imunohistohemijski primenom CD117 antitela. DNK je izolovana iz istih parafinskih kalupa, a PCR amplifikacija je izvršena primenom specifičnih prajmera PE1 i PE2. Stepen malignosti je utvrđen na osnovu prisustva mitotskih figura, multinuklearnih ćelija, ćelija sa bizarnim jedrom i kariomegalije na 10 polja velikog uveličanja. Na osnovu histopatoloških osobina, 14 od 24 mastocitoma su svrstani u visoki stepen maligniteta, dok je 10 mastocitoma niskog stepena maligniteta. Citoplazmatska ekspresija CD117 je uočena kod 9 od 14 mastocitoma visokog stepena maligniteta, što potvrđuje vezu između aberantne ekspresije CD117 i povećane ćelijske proliferacije.
Abstract
Histopathological examination, grading, immunohistochemical staining and molecular genetic examinations are the proposed criteria that should be used for cutaneous mast cell tumors (CMCTs) classification. The presence of aberrant CD117 expression and mutations of the c-kit proto-oncogene could be an indicative parameter for final histological grading. Determination of the connection between the localization of KIT receptor expression and the histological grade of CMCTs without c-kit proto-oncogene mutations was the main goal of this study. The study included twenty four CMCTs and six control skin samples from 30 dogs of different ages, breed and sex. Formalinfixed and paraffin-embedded tissue samples were stained with hematoxylin-eosin and toluidine blue and immunohistochemically tested for CD117 expression. DNA was extracted from the same paraffin blocks and subsequent polymerase chain reaction amplification was performed using PE1 and PE2 primers. Degree of malignancy was determined based on the presence of mitotic figures, multinucleated cells, bizarre nuclei and karyomegaly in 10 high power fields. Based on histological features, fourteen of 24 CMCTs were of a high histological grade, while ten were classified as a lowgrade malignancy. CD117 cytoplasmic expression was observed in nine of fourteen high-grade malignancy CMCTs, which confirms the link between the aberrant CD117 expression and increased cell proliferation.
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