BACKGROUND: S Phase Kinase Associated Protein 2 (Skp2), an F-box protein necessary for DNA replication, has recently been demonstrated to be an oncogene. The purpose of this study was to examine the Skp2 expression and to investigate its association with expressions of estrogen receptor (ER), androgen receptor (AR) and HER-2, as well as clinicopathological variables including tumor recurrence. METHODS: The expressions of Skp2, ER and AR were examined by immunohistochemistry and HER-2 amplification by chromogenic in situ hybridization (CISH) in 117 cases of breast carcinoma. RESULTS: Skp2 was expressed in 26 patients (22.2%) and was significantly correlated with tumor type (p=0.031), tumor grade (p=0.017) and ER expression (p=0.038). Twenty four (20.5%) of 117 patients had a tumor recurrence, and 6 patients (5.1%) died of multifocal metastases. Tumor recurrence was significantly correlated with histological grade (p=0.041) and lymph node status (p<0.001). CONCLUSIONS: Although Skp2 expression was statistically insignificant in association with tumor recurrence, it might be useful as a biologic predictor in breast cancer. The simple and reliable immunohistochemical assay presented in this study can be a routine part of breast cancer evaluation and may influence patient management.
Breast Neoplasms* ; Breast* ; DNA Replication ; Estrogens ; Humans ; Immunohistochemistry ; In Situ Hybridization ; Lymph Nodes ; Neoplasm Metastasis ; Oncogenes ; Receptors, Androgen ; Recurrence ; S Phase* ; S-Phase Kinase-Associated Proteins*

PURPOSE: Prolactinoma (prolactin-secreting pituitary adenoma) is one of the most common estrogen-related functional pituitary tumors. As an agonist of the dopamine D2 receptor, bromocriptine is used widely to inhibit prolactinoma progression. On the other hand, it is not always effective in clinical application. Although a dopamine D2 receptor deficiency contributes to the impaired efficiency of bromocriptine therapy to some extent, it is unknown whether there some other underlying mechanisms leading to bromocriptine resistance in prolactinoma treatment. That is the main point addressed in this project. MATERIALS AND METHODS: Human prolactinoma samples were used to analyze the S-phase kinase associated protein 2 (SKP2) expression level. Nutlin-3/adriamycin/cisplatin-treated GH3 and MMQ cells were used to analyze apoptosis in SKP2 overexpression or knockdown cells. SKP2 expression and the interaction partners of SKP2 were also detected after a bromocriptine treatment in 293T. Apoptosis was analyzed in C25 and bromocriptine-treated GH3 cells. RESULTS: Compared to normal pituitary samples, most prolactinoma samples exhibit higher levels of SKP2 expression, which could inhibit apoptosis in a p53-dependent manner. In addition, the bromocriptine treatment prolonged the half-life of SKP2 and resulted in SKP2 overexpression to a greater extent, which in turn compromised its pro-apoptotic effect. As a result, the bromocriptine treatment combined with C25 (a SKP2 inhibitor) led to the maximal apoptosis of human prolactinoma cells. CONCLUSION: These findings indicated that SKP2 inhibition sensitized the prolactinoma cells to bromocriptine and helped promote apoptosis. Moreover, a combined treatment of bromocriptine and C25 may contribute to the maximal apoptosis of human prolactinoma cells.
Apoptosis* ; Bromocriptine ; Half-Life ; Hand ; Humans* ; Pituitary Neoplasms ; Prolactinoma* ; Receptors, Dopamine D2 ; S-Phase Kinase-Associated Proteins

OBJECTIVETo study the effect of all-trans retinoic acid (ATRA) on U937 cell growth and its mechanism.

METHODSCell cycle was detected by flow cytometry (FCM), expressions of cell cycle associated protein and the p27 related protein were detected by Western blot. The binding of P27 and Skp2 was detected by immunoprecipitation.

RESULTSFCM displayed that ATRA could inhibit the proliferation of U937 cells. At 72 h on 1 micromol/L ATRA treatment, 72% of the cells were arrested at G0/G1 phase. Western blot displayed that ATRA could decrease the expression of cyclin A, up-regulate the expression of p21 and p27, and down-regulate the expression of p27 related proteins Skp2. p27 could bind with Skp2 in U937 cells as detected by immunoprecipitation.

CONCLUSIONATRA may arrest the proliferation of U937 cells through the reduction of Skp2 expression, and finally the induction of the accumulation of p27.

Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Humans ; S-Phase Kinase-Associated Proteins ; metabolism ; Tretinoin ; pharmacology ; U937 Cells

Carcinoma, Hepatocellular ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Neoplasm Staging ; Phosphorylation ; S-Phase Kinase-Associated Proteins ; metabolism

OBJECTIVE: To construct the siRNA expression vector of Skp2 and inhibit the expression of Skp2 through RNA interference in laryngeal carcinoma cell line Hep2 cell. METHOD: According to the encoding sequence of mRNA of Skp2, two pieces of oligonucleotide sequences were designed and synthesized. The annealed oligonucleotide fragments were subcloned into pGPU6/Neo siRNA expression vector, and the recombinant plasmid was transformed into strain DH5a. The plasmid identified by restriction enzyme was used for sequencing. After being identified by sequencing, the recombinant plasmids pGPU6Skp2 were transfected into Hep2 cells. Skp2 expression in the transfected cells was assayed by flow cytometry. RESULT: DNA sequencing showed that the oligonucleotide fragments were correctly inserted into pGPU6 vector, and Skp2 expression in the transfected cells was down-regulated significantly by pGPU6Skp2 at the protein level. CONCLUSION The siRNA expression vector of Skp2 was successfully constructed and could inhibit Skp2 expression in Hep2 cells. This result will facilitate further studies of Skp2 in gene therapy for tumors.
Base Sequence ; Cell Line, Tumor ; Genetic Vectors ; Humans ; Laryngeal Neoplasms ; genetics ; Plasmids ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; genetics ; S-Phase Kinase-Associated Proteins ; genetics ; Transfection

OBJECTIVETo study the expression of S phase kinase associated protein 2 (Skp2), phosphatase and tensin homolog deleted on chromosome ten (PTEN) in human laryngeal carcinoma and precancerous lesions, to explore their relations and clinical significance.

METHODSFormalin-fixed and paraffin-embedded tissues from 79 cases of laryngeal carcinoma, 16 cases of atypical hyperplasia of vocal fold,14 cases of adult laryngeal papillomas and 27 cases of vocal cord polyps were evaluated for the expression of Skp2, PTEN by SP immunohistochemistry, the levels of these proteins in tissues with the different types of lesion and their correlation with clinicopathological parameters of laryngeal carcinoma were analyzed.

RESULTSThe expression rates of Skp2 in vocal cord polyps, adult laryngeal papillomas, atypical hyperplasia of vocal cord and laryngeal carcinoma were 11. 11% ,14. 29% ,37. 50% ,39. 24% respectively. There was significant difference among them( Hc = 11. 57, P <0. 01). The expression rates of PTEN protein in vocal cord polyps,adult laryngeal papillomas, atypical hyperplasia of vocal cord and laryngeal carcinoma were 100% ,92. 86% ,75. 00%, 56. 96% respectively . There was significant difference among them (Hc = 62. 86, P<0. 05). There was a negative correlation between the expression of Skp2 and PTEN,and their correlation coefficient was r = -0. 4512(P <0. 01). Patients with Skp2 expression in laryngeal carcinoma revealed poorer five yeas survival rate than patients with negative expression of Skp2 (x2 = 21. 46, P = 0. 000).

CONCLUSIONSThe expression of Skp2 and PTEN took important roles in the tumorigenesis, aggressiveness, metastases of laryngeal carcinoma. The high expression of Skp2 was negative correlation with the lower PTEN in laryngeal carcinoma, which suggested that PTEN may regulate the expression of Skp2.

Adult ; Aged ; Aged, 80 and over ; Carcinoma ; pathology ; Female ; Humans ; Laryngeal Neoplasms ; metabolism ; pathology ; Male ; Middle Aged ; PTEN Phosphohydrolase ; metabolism ; S-Phase Kinase-Associated Proteins ; metabolism

OBJECTIVETo investigate the effect of S-phase kinase-associated protein 2 antisense oligodeoxynucleotide (Skp2 ASODN) on the growth and proliferation of gastric carcinoma SGC-7901 cells and its mechanism.

METHODSThe Skp2 oligodeoxynucleotides (ODNs) were embedded in cationic liposome Lipofectamine 2000 reagent and transfected into SGC-7901 cells. The cell growth and proliferation were observed with light microscopy and MTT assay. Cell cycle was measured by flow cytometry. The expression levels of Skp2 and p27 mRNA were detected by reverse transcription-polymerase chain reaction. The expression levels of Skp2 protein and its substrate p27 protein were detected by Western blot.

RESULTAfter treatment with Skp2 ASODN, the growth and proliferation of SGC-7901 cells were inhibited in a dose-dependent manner with a peak value at 48 h. The inhibition rate of 200 nmol/L group at 48 h was 42.4 % (P<0.01). In cell cycle study the percentage of S phase cells in 200 nmol/L group was significantly higher than that in normal control group (P<0.05). Both Skp2 mRNA and its protein levels in 200 nmol/L group were significantly lower than those in control group and in Skp2 nonsense oligodeoxynucleotide (Skp2 NSODN) group (P<0.05). However, p27 mRNA level remained unchanged although its protein level was significantly higher than that in control group and NSODN group (P<0.05).

CONCLUSIONSkp2 ASODN can inhibit the growth and proliferation of SGC-7901 cells, which may be mediated by interfering with ubiquitin-proteosome pathway and cell cycle regulation.

Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Humans ; Oligodeoxyribonucleotides, Antisense ; genetics ; pharmacology ; S-Phase Kinase-Associated Proteins ; genetics ; pharmacology ; Stomach Neoplasms ; pathology ; Transfection

OBJECTIVETo study the expression of Skp2 gene (s-phase kinase associated protein 2) in the pathological scars and its relationship with p27kip1 level and to investigate its role and its probable mechanism in the pathogenesis of abnormal scars.

METHODSImmunohistochemical technique was performed to detect the expression and distribution of Skp2 and p27kip1 protein in hypertrophic scar (42 cases), keloid (18 cases), normal scar (40 cases) and normal skin (50 cases), statistics was used to analyze the data.

RESULTSThe positive rate of Skp2 and p27kip1 protein expression was not statistically different between the hypertrophic scar and keloid (P > 0.05), while they were all remarkably significant in comparison between normal scar and abnormal scar (P < 0.05). In pathological scar the protein of Skp2 and p27kip1 showed a strong inverse correlation (P < 0.01).

CONCLUSIONThe result indicated that the expression of Skp2 was increased and it may lead to decrease p27kip1 level in the hypertrophic scar and keloid, Skp2 overexpression might play an important role in the proliferation of fibroblasts and in the pathogenesis of pathological scar by adjusting the regulation of cyclins such as p27kip1.

Adolescent ; Adult ; Child ; Child, Preschool ; Cicatrix, Hypertrophic ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p27 ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Keloid ; metabolism ; pathology ; Male ; Middle Aged ; S-Phase Kinase-Associated Proteins ; metabolism ; Young Adult

OBJECTIVETo investigate whether the connection of p27(Kip1) to S-phase kinase-associated protein 2 (Skp2) plays an oncogenic role in intraductal proliferative lesions of the breast.

METHODSHere we investigated the mechanism involved in association of Skp2’s degradation of p27(Kip1) with the breast carcinogenesis by immunohistochemical method through detection of Skp2 and p27(Kip1) protein levels in 120 paraffin-embedded tissues of intraductal proliferative lesions including usual ductal hyperplasia (UDH, n=30), atypical ductal hyperplasia (n=30), flat epithelial atypia (FEA, n=30), and ductal carcinoma in situ (DCIS, n=30). Moreover, the expression status of Skp2 and p27(Kip1) in 30 cases of the normal breast paraffin-embedded tissues were explored.

RESULTSThe DCIS group was with the highest Skp2 level and the lowest p27(Kip1) level, and the UDH group was with the lowest Skp2 level and the highest p27(Kip1) level.Both Skp2 and p27(Kip1) levels in the DCIS group were significantly different from those in the UDH group (all P<0.01).The levels of Skp2 and p27(Kip1) in the FEA group were significantly different from both the DCIS and UDH groups (all P<0.05).p27(Kip1) was negatively correlated with Skp2 in both the UDH group (r=-0.629, P=0.026) and DCIS group (r=-0.893, P=0.000).

CONCLUSIONOverexpression of Skp2 might be the mechanism underlying p27(Kip1) over degradation.

Adult ; Aged ; Breast ; pathology ; Breast Neoplasms ; etiology ; Carcinoma, Intraductal, Noninfiltrating ; etiology ; Cell Proliferation ; Cyclin-Dependent Kinase Inhibitor p27 ; physiology ; Female ; Humans ; Hyperplasia ; Middle Aged ; S-Phase Kinase-Associated Proteins ; physiology

OBJECTIVETo detect the expression of skp2 and p27kip1 in human renal cell carcinoma (RCC) using tissue chip technique, and evaluate the relationship between the proteins and the biological behavior of RCC.

METHODSTissue chip technique and immunohistochemical SP method was used to detect the expression of skp2 and p27kip1 in normal and tumor tissues.

RESULTSThe positivity rate of Skp2 in RCC was significantly higher than that in normal renal tissues (P=0.025). The positivity rate of Skp2 expression in RCC was significantly correlated to poor differentiation of the tumor (P=0.002), and was not associated with the patients gender, age, tumor size, lymph node metastasis and stages of RCC (P>0.05). The positivity rate of p27kip1 in RCC was significantly lower than that in normal renal tissues (P=0.007). The positivity rate of p27kip1 expression was inversely correlated to the malignancy and stage of RCC (P<0.05), but not with the patients' age, gender, lymph node metastasis and tumor size (P>0.05). An inverse correlation was noted between Skp2 and p27kip1 expressions (r= -0.273, P=0.014).

CONCLUSIONOverexpression of Skp2 protein may lead to decreased p27kip1 level in RCC, indicating its involvement in the carcinogenesis and development of RCC.

Adult ; Aged ; Carcinoma, Renal Cell ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p27 ; biosynthesis ; Female ; Humans ; Immunohistochemistry ; Kidney Neoplasms ; metabolism ; pathology ; Male ; Middle Aged ; S-Phase Kinase-Associated Proteins ; biosynthesis ; Tissue Array Analysis ; Young Adult