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標題: 犬乳腺組織與惡性乳腺腫瘤的p63蛋白表現
Expression of p63 in canine mammary gland and malignant mammary gland tumors
作者: 周明賢
關鍵字: canine;犬;mammary gland tumor;p63;乳腺瘤
出版社: 獸醫學系
摘 要
p63基因位於人類第三對染色體(3q27)上,其產物至少有6種不同的isoforms。在人類與犬正常乳腺中,只有肌上皮細胞會表現出ΔNp63α的活性。而在人類的原發性肺癌與頭和頸複層扁平上皮細胞癌 (head and neck squamous cell carcinoma,HNSCC)細胞株中,亦可偵測到ΔNp63α的過度表現,因此,ΔNp63α被認為會造成某些人類腫瘤的發生。
本研究在小白鼠的乳腺腫瘤中,發現核苷酸p63序列的變異,將該部份核苷酸序列轉譯成胺基酸序列後,其第17個胺基酸序列與GenBank(AF075438)所登錄的胺基酸序列不同(Tyr to Cys),而這個序列與由小白鼠正常組織中增幅出的mRNA所轉譯出來的胺基酸序列相較時,並沒有發現與GenBank(AF075438)所登錄的胺基酸序列有相異之處。
本實驗利用western blotting的方式來了解在犬正常乳腺,犬乳腺腫瘤細胞株(DMGT cell line)與臨床惡性乳腺腫瘤病例中,ΔNp63α蛋白質的表現量是否有差異。實驗結果發現,在65 kD位置上出現清晰條帶,分子量約等於人類ΔNp63α (68kD)蛋白質。除了犬正常乳腺組織含有豐富的ΔNp63α蛋白質外,DMGT cell line與犬惡性乳腺腫瘤組織內ΔNp63α的含量並不高。另外,SDS-PAGE分析發現在約15kD的位置有清晰的條帶,經N端胺基酸定序的結果發現為犬之α-hemoglobin,而該條帶的出現是否與乳腺的病變有絕對的關連性,仍待進一步實驗釐清。

The p63 gene, mapping to human chromosome 3q27-29, encodes at least six protein isoforms. The ΔNp63 protein expression is restricted to myoepithelial cells in both normal human and canine breast tissue. Additionally, it was observed overexpression of ΔNp63α isoform in head and neck cancer cell lines and primary lung cancers, so some results support that ΔNp63α may play an oncogenic role in human cancer. Our research showed that one missense variant was observed and this involved a change from Tyr to Cys at codon 17. This mutation was not noted in GenBank (AF075438).
We use Western blotting to identify the expression of ΔNp63α isoforms in canine normal breast tissue, canine mammary gland tumor cell line (DMGT cell line) and clinical malignant mammary gland tumor samples. The Western blot showed a clear band of approximately 65 kD in canine normal breast tissue and the band was corresponding to ΔNp63α protein in human tissue. Expression of ΔNp63α protein in canine breast tissue was abundant, but DMGT cell line and clinical malignant mammary gland tumor samples had low levels of ΔNp63α protein. Furthermore, we found a clear band at approximately 15 kD in SDS-PAGE in canine normal breast tissue. After N-terminal sequence analysis, we found this protein was α-hemoglobin, but the significance of the α-hemoglobin protein in the pathogenesis of breast tissue was not known yet.
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