EXPRESSION OF p16 AND Ki67 IN SQUAMOUS INTRAEPITHELIAL LESIONS OF THE UTERINE CERVIX

Vrat maternice predstavlja donju trećinu maternice i povezuje trup maternice s rodnicom preko endocervikalnog kanala. Na vratu maternice nalaze se dvije vrste epitela: cilindrični i pločasti epitel. Pločasti epitel je visokodiferenciran, višeslojan epitel. Područje spoja između pločastog i cilindričnog epitela je od velike važnosti u nastanku raka grlića maternice. Taj spoj poznat je pod nazivom transformacijska zona. Iz tog razloga veliki naglasak je stavljen na uzorkovanje transformacijske zone cervikovaginalnim razmazom. Infekcija humanim papilomavirusom (HPV) danas je najčešća virusom uzrokovana spolno prenosiva bolest. Svi tipovi HPV-a mogu izazvati blage promjene stanica bez ozbiljnih posljedica, ali neki od njih ( onkogeni tipovi) mogu dovesti do karcinoma vrata maternice. Rak vrata maternice danas je drugi najčešći zloćudni tumor ženske populacije u gotovo svim dijelovima svijeta, a po učestalosti ga nadmašuje samo rak dojke. Papa test ili VCE obrisak je jednostavna, brza i neinvazivna medicinska pretraga kojom se uzima obrisak rodnice, vrata maternice i kanala vrata maternice. Služi za otkrivanje raka grlića maternice i promjena koje mu prethode. Također, može otkriti upalne promjene te identificirati uzročnika.Tekućinska citologija (LBC) je novija metoda obrade obrisaka grlića maternice. Glavna razlika između tekućinske citologije i Papa testa je da se stanice uzete s vrata maternice prenose izravno u bočicu s tekućinom za fiksiranje. Stanice su nanešene u jednom sloju na predmetnici bez primjesa krvi, sluzi i/ili upalnih stanica koje bi ometale analizu nakon bojanja po Papanicolaou. Imunocitokemijski test za istodobno kvalitativno otkrivanje proteina p16INK4A i Ki67 u cervikalnim citološkim preparatima indiciran je kao pomoć pri određivanju žena s cervikalnim intraepitelnim lezijama visokog stupnja u populaciji podvrgnutoj probiru i u podskupinama bolesnica s rezultatom citološkog nalaza Papa testa ASC-US i LSIL te u bolesnica pozitivnim na HPV visokog rizika. Budući da velik udio epitelnih stanica u CIN lezijama također pokazuje proliferativnu aktivnost, otkrivanje cervikalnih epitelnih stanica koje istodobno eksprimiraju p16INK4A i Ki67 može se upotrebljavati kao indikator promijenjenog statusa stanice. U reakciji se stvaraju dva različito obojena produkta: smeđi talog na mjestima antigena p16INK4A i crveni talog na mjestima antigena Ki67. Kako bi se održala optimalna osjetljivost i spriječio gubitak jasnoće kromogena, potrebno je provesti postupak uklapanja u dva koraka. Rezultati bojenja se procjenjuju pomoću svjetlosnog mikroskopa. Pozitivnim rezultatom se smatra prisutnost jedne ili više cervikalnih epitelnih stanica koje istodobno pokazuju smeđe obojenu citoplazmu i crveno obojenu jezgru.The uterine cervix is the lower third of uterus and connects uterus with vagina through the endocervical canal. There are two types of epithelium on cervix: cylindrical and squamous epithelium. Squamous epithelium is highly differentiated and multilayered. The connection area between squamous and cylindrical epithelium is of great importance in development of cervical cancer. This juncture is known as the transformation zone. For this reason, great emphasis is placed on sampling cervicovaginal smear of the transformation zone. Infection with human papillomavirus (HPV) is the most common sexually transmitted disease caused by a virus. All types of HPV can cause mild cell changes without serious consequences, but some of them (oncogenic types) can lead to cervical cancer. Cervical cancer is now second most common malignancy of the female population in almost all parts of the world and only breast cancer is more frequent. Pap test or VCE smear is simple, fast and noninvasive medical test that takes a swab of vagina, cervix and cervical canal. It is used to detect cervical cancer and changes that precede it. It can also detect inflammatory changes and identify the cause of disease.Liquid Based Cytology ( LBC) is a newer method of processing cervical smears. The main difference between LBC and Pap test is that the cells from the cervix are transferred directly in the bottle with a liquid preservative. Cells are placed in a single layer on the slides without a tinge of blood, mucus and/or inflammatory cells that would interfere with the analysis after staining it with Papanicolaou method.Immunocytochemistry test for simultaneous qualitative detection of protein p16INK4A and Ki67 in cervical cytology preparations is indicated as help in determining the women with cervical intraepithelial lesions of high grade in a population subjected for screening and in subgroups of patients with a result of Pap test ASC–US or LSIL or in patients positive for high risk HPV. Since a large proportion of epithelial cells in CIN lesions also demonstrate proliferative activity, detection of cervical epithelial cells simultaneously expressing p16INK4A and Ki67 can be used as an indicator of the changed status of the cells. The reaction creates two differently colored product: brown sediment in places of antigen p16INK4A and red sediment in places of Ki67 antigen. To maintain optimal sensitivity and prevent the loss of chromogen clarity, it is necessary to carry out the procedure of fitting in two steps. Result of staining can be estimated using a light microscope. A positive result is considered to be the presence of one or more cervical epithelial cells that show at the same time brown colored cytoplasm and red colored nucleus