Expression And Correlation Of P73 And P63 In Middle Ear Cholesteatoma

Background The middle ear cholesteatoma is a cyst in middle ear,its intinemake up of stratified squamous epithelium. The capsular space is fullof caducous epithelium, cornification materials, cholesterol crystal.There is fibrous tissue out of the cyst, whose thickness is uneven. Thefibrous tissues tightly connect the cyst with the bone or tissue nearby.At present the middle ear choles-teatoma is still a common chronicdisease in Otology and do great harm to health such as suppuration,deafness, dinus and intracranial complications. Pathology ischaracterized by highly proliferations of the tabular epithelium in themiddle ear cavity, destruction of adjacent bone and apoptosis of thekeratinocyte. The biological character is similar to tumor, such asinvasive, aggressive, migratory, altered differenation and recidivisticcharacters, but it has been proved to be a benign lesion. Theetiopathogenesis and nosogenesis is not clear yet. Recently, manyresearches in molecular biology has confirmed that cholesteatomaepithelium cells are both hyperprolifetative and overapoptosis,whichwere confirmed by normal expression p53 anti-oncogene in themiddle ear cavity.p53 ,p63 and p73 are FAM-gene and 63 %calmodulin binding domain (CAM) of p73 and p63 is similar to p53 init, who can transcribe into many isomerides. There are MDM2calmodulin binding sequence in p73 TAD. Its function is similar top53. when p73 is overexpressed,it can activate the promoter of p21,activate the transcription of target gene p21WAF1, increase itsexpresstion and promote apoptosis. p63 isomer has central DNAbinding structural domain and oligomerization structural domain,which can be binding with p53DNA binding site, activate p53correlated promoter,p21,MDM2,bax and so on, trans-activate thetranscription of target gene p53 and induce apoptosis. p73 and p63have different effects during many tumor intermediate stage.(oncogene or anti-oncogene)ObjectiveStudy the expression and correlation of p73 and p63 in middleear choleseatoma and discuse their effects in cholesteatoma.Methods23 samples of middle ear cholesteatoma we studied came fromthe patients who were operated in the First Hospital of Jilin Universityduring March. 2004 to Dec.2005, and all of them had been proved tobe cholesteatoma by pathology. Clinical data were integrated. 10samples were normal skin of the external ear canal acquired frometympanoplasty at the same time. The wax specimens were sliced into4μm section continuously. The concentrate antibodies of rabbitanti-human p73 polyclonal antibody, the instant anti-bodies of mouseanti-human p63 monocional antibody, the streptavidin-peroxidaseimmunohistochemistry supersensitivity kit, the citrate buffer solution,PBS and DAB were purchased from MAI XIN biotechnologycompany in Fuzhou We used the immunohistochemical technologywith streptavidin-peroxidase method and all procedures wereperformed strictly according to the description. PBS was used fornegative control instead of the primary antibody at the same time.Assessment standard: p73 positive reaction cells: Dark brown or deepyellow-brown gross pellets were found at karyon and cytopasmic;p63positive reaction cells: yellow-brown pellets were found at karyon.Under high power microscopic observationm, we selected five randomvisual fields and calculated the positive cells, the number of whichdevided by that of the total cells in each field, then we calculated theaverage number. No positive cells is negative (-);less than 25% isweakly positive (+);26%-50% is midrange positive (++);more than50% is strong positive (+++). Datas were dealed with significance testand correlation analysis with fourfold table precise probabilisticmethod of x2 test. P<0.05 was considered statistically significant.Results:1. The result showed that there almost no p73 positive cells innormal ear cananl shin,positive rate was 65.2% in middle earcholesteatoma, whose intensity was strong to weak. 2 samples wereweak positive expression, 8 samples were medium positiveexpression,5 samples were strong positive expression and 8 sampleswere negative expression. p73 positive cells were detected in eachlayers, whose intensity was high to weak, and the positive cells mostlyin spine layer and in granular cell layer. The exact probabilities in 2×2table test shows obvious difference, (P=0.00052;P<0.005). 2. p63positive rate was 70% in basal layer of normal ear canal shin,lowintensive;And p63 positive cells were detected in each layers ofmiddle ear cholesteatoma, p63 positive rate was highest in basal layerand lowest in horny layer, positive rate was 87%, whose intensity wasstrong positive expression, medium positive expression and weakpositive expression.Among them, 1 sample was weak positiveexpression, 4 samples were medium positive expression, 15 sampleswere strong positive expression. Through exact probabicities in 2×2table (P=0.3364;P>0.05), they didn't have significant difference bystatistics. 3. Compared with p63 positive expression and p73 positiveexpression in middle ear cholesteatoma,14 cases were p63 and p73positive ,2 cases were p63 positive in the cases of p73 negativespecimens,and 1 case was p63 negative in the cases of p73 positivespecimens. They have significant difference by statistics test (x2 =4.46;P=0.037;P<0.05).Conclusion :The following aspects can be showed through the result: 1. Highexpression of p73 in the middle ear cholesteatoma indicated that theapoptosis capacity was increased in the cholesteatoma epithelium,which makes the choles-teatoma cutin cells accumulate. Theincreased apoptosis inhibit the proliferation of the cholesteatoma cells,which indicates that middle ear cholesteatoma is different frommalignant tumor. 2. p63 was expressed both in the cholesteatomaepithelium and in the normal external auditory canal skin,but they aremore expressed in the middle ear cholesteatoma. They expressed onlyin the basal layer in the external auditory canal skin, but in thecholesteatoma epithelium, they expressed in all layers. So we supposethat the basal cells in the cholesteatoma epithelium have the potentialof proliferation. 3. p63 and p73 are expressed in middle earcholesteatoma, and they have dependability,so we suppose they havesignificant difference in middle ear cholesteatoma, Proliferation islarger than apoptosis.