Involvement Of Pituitary Tumor Transforming Gene1in Psoriasis And Cutaneous Tumors

[BACKGROUND]Involvement of pituitary tumor transforming gene1(PTTG1), encoding for securin, is a transcription factor and originally isolated from rat pituitary tumor cells. PTTG1was identified in human tissues with low expression in normal cells but elevated overexpression occurs in a wide variety of human invasive tumors, especially in endocrine-related tumors. Accumulating evidence suggest PTTG1as a potential biomarker for cancer malignancy.PTTG1-deficient cells have a significant decrease in proliferative potential after exposure to DNA-damaging agents.PTTG1is a well established cell-cycle regulatory protein, involving in cell replication, DNA damage/repair, organ development, metabolism, cell transformation and cell senescence.Overexpression of PTTG1caused upregulation of cyclin B1, cyclin-dependent kinase1(CDK1), and c-Myc, which resulted in enhanced proliferation and suppression of early differentiation without apparent alterations in terminal differentiation in keratinocytes. This aberrant cell cycle regulation due to overexpression of PTTG1suggests that it may have important roles in hyperproliferative disorders of epidermis, such as psoriasis and squamous cell carcinoma. Enhanced expression of PTTG1in the psoriatic epidermis may result in aberrant regulation of the cell cycle and impaired differentiation via the interplay between PTTG1and TNF-a. The mechanism of PTTG1in the proliferation of epidermal keratinocytes is still unclear.We sought to clarify the subcellular localization of PTTG1and its possible involvement in proliferative skin diseases.[OBJECTIVE]To define the immunolocalization and expression of PTTG1in skin and primary cultured keratinocytes.To clarify the different expression and localization of PTTG1in seborrheic keratosis, BCC and SCC tissues.To explore the possible involvement of PTTG1in proliferative skin diseases.[MTTHODS]Primary cultured keratinocytes and skin samples were investigated in vitro, including skin from psoriasis, seborrheic keratosis (SK) and skin tumors, basal cell carcinoma (BCC) and squamous cell carcinoma (SCC). To define the expression of PTTG1in normal and psoriatic keratinocytes, mRNA extracted from normal and psoriatic epidermis was quantified relatively by real-time PCR. Immunofluorescence and immunohistochemistry of PTTGl were assayed to define the subcellular localization of PTTG1. Cell cycle synchronized by chemicals was done to examine the association of PTTG1with cell cycle in psoriatic keratinocytes. To examine the effect of PTTG1on the invasion of A431cells, cells stablely expressing PTTG1and control siRNA were compared in a transwell invasion assay.[RESULTS] I. Expression of PTTG1in normal and psoriatic skin1.PTTG1was expressed in all the samples from normal and psoriatic skin.2. In normal epidermis, PTTG1was localized predominantly in the basal keratinocytes, and about10%of basal keratinocytes were PTTG1positive.3. In psoriatic epidermis, abundant PTTG1was observed in basal keratinocytes and suprabasal keratinocytes. The PTTG1positive keratinocytes were about30%-40%in basal and suprabasal keratinocytes. 4. PTTG1in psoriatic epidermis was about5-fold of normal ones (P<0.01).II. Subcellular immunolocalization of PTTG1in skin, primary cultured normal and psoriatic keratinocytes and HaCaT cells1. PTTG1was predominantly expressed in the cytoplasm determined by Santa Cruz antibody, but identified both in cytoplasm and nucleus by using Abcam antibody in normal and psoriatic skin.2. In primary cultured normal and psoriatic keratinocytes, PTTG1was localized in cytoplasm.3. PTTG1in HaCaT cells was distributed throughout the cytoplasm of metaphase cells.III. The relationship between the expression of PTTG1and cyclin B1in different cell cycle of psoriatic keratinocytes1. Primary cultured psoriatic keratinocytes without any treatment showed high and low cytoplasmic expression of PTTG1in most cells.2. The PTTG1high keratinocytes also showed a corresponding high expression of cyclin B1, no cyclin B1signals was seen in the PTTG1low cells.3. Cyclin B1increased at G2phase and peaked at M phase, PTTG1was seen both at G2phase and M phase.4. Highest PTTG1expression correlated with highest cyclin B1expression and highest degree of nuclear pleomorphism at M phase.5. Psoriatic keratinocytes at M phase expressed high levels of PTTG1also expressed high levels of cyclin B1, and the expression was spatial concordance at M phase.IV. Immunolocalization and expression of PTTG1in seborrheic keratosis, BCC and SCC tissues1. PTTG1was mainly distributed in cytoplasm, only occasionally in the nucleus.2. In seborrheic keratosis, PTTG1was positive in about10%keratinocytes.3. In BCC cells, PTTG1was about20%positive.4. In SCC tissues, more than80%cells were stained, especially prominently in poorly differentiated tumor cells. V. The effect of PTTG1on the invasion of A431cells1. PTTG1siRNA reduced the expression of PTTG1by about50in A431cells.2. Significantly fewer PTTG1siRNA-expressed A431cells (11±2cells/field) migrated through the Matrigel compared to control cells (24±4cells/field, P<0.01).[CONCLUSION]I. In normal epidermis, PTTG1was localized predominantly in the basal keratinocytes while in psoriatic epidermis PTTG1was observed in both basal keratinocytes and suprabasal keratinocytes. PTTG1in psoriatic epidermis was much more than normal onesII. In primary cultured normal and psoriatic keratinocytes, PTTG1was localized in cytoplasm, in HaCaT cells PTTG1was distributed throughout the cytoplasm of metaphase cells.III. Psoriatic keratinocytes at M phase expressed high levels of PTTG1also expressed high levels of cyclin B1, and their expression was spatial concordance at M phase.IV. In seborrheic keratosis, BCC and SCC tissues,PTTG1was mainly distributed in cytoplasm, only occasionally in the nucleus, PTTG1-positive keratinocytes is about10%,20%and more than80%respectively.V. The invasive capacity was reduced in more than50%of PTTG1siRNA-expressed A431cells.