Scutellariae Barbatae Herba Extract Promotes Cycle Arrest of Nasopharyngeal Carcinoma Cells by Inhibiting STAT3/SKP2 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the effect of Scutellariae Barbatae Herba extract on on the cycle arrest of nasopharyngeal carcinoma cells and the possible mechanism by adding different concentration of Scutellariae Barbatae Herba extract （0.25， 0.5， 1 g·L-1） in the culture medium， taking CNE1 （nasopharyngeal carcinoma cells） as the research object. MethodAfter the treatment of CNE1 by Scutellariae Barbatae Herba extract， cell counting kit-8 （CCK-8） was used to detect cell proliferation， and Giemsa staining was used to detect the clone formation rate. Flow cytometry was used to detect cell cycle distribution， and reverse transcription-polymerase chain reaction （ RT-PCR） assay and Western blot assay were used to detect the relative expression of messenger ribonucleic acid （mRNA） by small interfering RNA （siRNA） or overexpression. ResultAs compared with the blank group， the proliferation and colony formation rate of CNE1 in the Scutellariae Barbatae Herba extract group significantly decreased （P<0.05， P<0.01） in a dose and time-dependent manner， whereas the percentage of cells in the presynthetic phase （G0/G1） increased （P<0.05， P<0.01）. The expression level of S-phase kinase associated protein 2 （SKP2） in the Scutellariae Barbatae Herba extract group significantly decreased （P<0.01） as compared with the blank group. As compared with the Scutellariae Barbatae Herba extract group， the protein levels of p21 and p27 significantly decreased in the overexpressed SKP2+ Scutellariae Barbatae Herba extract group （P<0.01）. As compared with the blank group， the signal activation and the phosphorylation level of signal transducer and activator of transcription 3 （STAT3） of CNE1 in the S. barbata extract group significantly decreased （P<0.05， P<0.01）. ConclusionScutellariae Barbatae Herba extract effectively inhibits the proliferation of CNE1， and the mechanism may be related to its action on the STAT3/SKP2 pathway.

Scutellariae Barbatae Herba Extract Promotes Cycle Arrest of Nasopharyngeal Carcinoma Cells by Inhibiting STAT3/SKP2 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the effect of Scutellariae Barbatae Herba extract on on the cycle arrest of nasopharyngeal carcinoma cells and the possible mechanism by adding different concentration of Scutellariae Barbatae Herba extract （0.25， 0.5， 1 g·L-1） in the culture medium， taking CNE1 （nasopharyngeal carcinoma cells） as the research object. MethodAfter the treatment of CNE1 by Scutellariae Barbatae Herba extract， cell counting kit-8 （CCK-8） was used to detect cell proliferation， and Giemsa staining was used to detect the clone formation rate. Flow cytometry was used to detect cell cycle distribution， and reverse transcription-polymerase chain reaction （ RT-PCR） assay and Western blot assay were used to detect the relative expression of messenger ribonucleic acid （mRNA） by small interfering RNA （siRNA） or overexpression. ResultAs compared with the blank group， the proliferation and colony formation rate of CNE1 in the Scutellariae Barbatae Herba extract group significantly decreased （P<0.05， P<0.01） in a dose and time-dependent manner， whereas the percentage of cells in the presynthetic phase （G0/G1） increased （P<0.05， P<0.01）. The expression level of S-phase kinase associated protein 2 （SKP2） in the Scutellariae Barbatae Herba extract group significantly decreased （P<0.01） as compared with the blank group. As compared with the Scutellariae Barbatae Herba extract group， the protein levels of p21 and p27 significantly decreased in the overexpressed SKP2+ Scutellariae Barbatae Herba extract group （P<0.01）. As compared with the blank group， the signal activation and the phosphorylation level of signal transducer and activator of transcription 3 （STAT3） of CNE1 in the S. barbata extract group significantly decreased （P<0.05， P<0.01）. ConclusionScutellariae Barbatae Herba extract effectively inhibits the proliferation of CNE1， and the mechanism may be related to its action on the STAT3/SKP2 pathway.

Degradation of SARS-CoV-2 receptor ACE2 by the E3 ubiquitin ligase Skp2 in lung epithelial cells / 医学前沿

An unexpected observation among the COVID-19 pandemic is that smokers constituted only 1.4%-18.5% of hospitalized adults, calling for an urgent investigation to determine the role of smoking in SARS-CoV-2 infection. Here, we show that cigarette smoke extract (CSE) and carcinogen benzo(a)pyrene (BaP) increase ACE2 mRNA but trigger ACE2 protein catabolism. BaP induces an aryl hydrocarbon receptor (AhR)-dependent upregulation of the ubiquitin E3 ligase Skp2 for ACE2 ubiquitination. ACE2 in lung tissues of non-smokers is higher than in smokers, consistent with the findings that tobacco carcinogens downregulate ACE2 in mice. Tobacco carcinogens inhibit SARS-CoV-2 spike protein pseudovirions infection of the cells. Given that tobacco smoke accounts for 8 million deaths including 2.1 million cancer deaths annually and Skp2 is an oncoprotein, tobacco use should not be recommended and cessation plan should be prepared for smokers in COVID-19 pandemic.

Skp2 inhibitor SKPin C1 decreased viability and proliferation of multiple myeloma cells and induced apoptosis

Multiple myeloma (MM) is a malignant neoplasm of plasma, and exhibits several harmful effects including osteolytic injuries, hypercalcemia, and immune dysfunction. Many patients with MM succumb to the underlying malignancy. An S-phase kinase-related protein 2 (Skp2) inhibitor, designated SKPin C1, has been developed and confirmed to have an inhibitory effect on metastatic melanoma cells. This study aimed to determine the effect of SKPin C1 on MM. Normal B lymphocytes, THP-1 cells, and MM U266 and RPMI 8226 cells were exposed to various dosages of SKPin C1 for 48 h. Cell proliferation was determined by MTT, EdU staining, and cell cycle assays. Western blot assays were performed to assess intracellular protein levels of Skp2, p27, and cleaved caspase-3. The amount of ubiquitin attached to p27 was determined using an immunoprecipitation assay. The viability of U266 and RPMI 8226 cells was significantly inhibited by 10 μM SKPin C1 and the inhibitory effect was enhanced with increasing doses of SKPin C1. In contrast, 50 μM SKPin C1 only marginally decreased viability of normal B lymphocytes in 12 h. Skp2 and p27 expression in U266 and RPMI 8226 cells was higher and lower, respectively, than that in the normal B lymphocytes. Treatment with SKPin C1 or Skp2 knockdown increased p27 protein levels in U266 and RPMI 8226 cells by preventing p27 from being ubiquitinated, which slowed the cell cycle, inhibited cell proliferation, and triggered apoptosis. Therefore, this study suggested SKPin C1 as a potent inhibitor against aberrant proliferation and immortalization of MM.

Cloning and expression analysis of an F-box protein gene DoSKP2A in Dendrobium officinale / 中草药

Objective: To clone an F-box protein namely S phase kinase-associated protein gene DoSKP2A with full length cDNA, in a rare endangered medicinal orchid species Dendrobium officinale, followed by bioinformatics analysis and expression pattern analysis. Methods: RACE technology were used for gene identification. Characteristics of physiochemical properties, conserved domains and subcellular localization of the protein were determined using a series of bioinformatics tools. The analyses of multiple alignment and phylogenetic tree were performed using DNASTAR 7.0 and MEGA 7.0, respectively. Real time quantitative PCR was used for gene expression analysis. Results: DoSKP2A gene was cloned (GenBank accession KU160472). The full length cDNA of DoSKP2A was 1 507 bp in length, and ORF was 1 101 bp, encoding a 366-aa protein with a molecular weight of 39 590 and an isoelectric point of 7.9. The deduced DoSKP2A protein, without transmembrane or signal peptide residues, contained an F-box core domain (26-88), a leucine-rich repeat (202-226), and multiple conserved motifs. DoSKP2A had high identities (64.6%-72.4%) with SKP2As proteins from various plants. DoSKP2A belonged to the monocotyledons subgroup of the SKP2As evolutionary tree. DoSKP2A gene was differentially expressed in the three included organs. The transcripts were more abundant in the roots, with 6.16 fold, then the stems and the lowest in the leaves. Conclusion: The novel full-length F-box protein gene DoSKP2A was obtained, along with bioinformatics and expression characteristics, which provided molecular basis for the growth and development, signal transduction, and stress resistance of D. officinale.

Anti-growth Effects of Imatinib and GNF5 via Regulation of Skp2 in Human Hepatocellular Carcinoma Cells

BACKGROUND: Human hepatocellular carcinoma (HCC) is a common liver tumor and the main cause of cancer-related death. Tyrosine kinase inhibitors, such as imatinib and GNF5 which were developed to treat chronic myelogenous leukemia, regulate the progression of various cancers. The aim of this study was to confirm the anti-tumor activity of tyrosine kinase inhibitors through regulation of S-phase kinase-associated protein 2 (Skp2), an important oncogenic factor in various cancer cells, in human hepatocarcinoma SK-HEP1 cells. METHODS: Cell viability and colony formation assays were conducted to evaluate the effects of imatinib, GNF5 and GNF2 on the growth of SK-HEP1 cells. Using immunoblot analysis, we assessed change of the activation of caspases, PARP, Akt, mitogen-activated protein kinases, and Skp2/p27/p21 pathway by imatinib and GNF5 in SK-HEP1 cells. Using sh-Skp2 HCC cells, the role of Skp2 in the effects of imatinib and GNF5 was evaluated. RESULTS: Imatinib and GNF5 significantly inhibited the growth of SK-HEP1 cells. Treatment of imatinib and GNF5 decreased Skp2 expression and Akt phosphorylation, and increased the expression of p27, p21, and active-caspases in SK-HEP1 cells. In sh-Skp2 HCC cells, cell growth and the expression of Skp2 were inhibited by more than in the mock group treated with imatinib and GNF5. CONCLUSIONS: These results suggest that the anti-growth activity of tyrosine kinase inhibitors may be associated with the regulation of p27/p21 and caspases through Skp2 blockage in HCC cells.

Inhibition of SKP2 Sensitizes Bromocriptine-Induced Apoptosis in Human Prolactinoma Cells / Journal of the Korean Cancer Association, 대한암학회지

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.

Effects of Skp2 overexpression on sensitivity of troglitazone in breast cancer cells / 国际检验医学杂志

Objective To investigate the effects of Skp2 overexpression on the sensitivity of troglitazone (TRG) in breast cancer cells and to devote to develop a novel drug for increasing the patient survival rate and eventually reaching the cure goal .Methods The transcription activities of PPARγ were analyzed on peroxisome proliferators response element(PPRE) luciferase reporter .The flow cytometry analysis and CCK‐8 assay were adopted to study that overexpression of Skp2 was associated with resistance to TRG‐mediated inhibition growth and apoptosis of breast cancer cells .Results Our study found that overexpression of Skp2 inhibi‐ted the transcriptional activity of the endogenous PPARγ and resisted to TRG‐mediated inhibition growth and apoptosis of breast cancer cells .Conclusion Overexpressed Skp2 breast cancer cells is able to be resistant to TRG‐induced sensitivity of breast cancer cells .Furthermore down‐regulating Skp2 will significantly enhance the growth inhibition of TRG on breast cancer cells .

Effect of Skp2 antisense oligodeoxynucleotide on growth and proliferation of colorectal cancer SW 480 cells / 重庆医学

Objective To investigate the effect of S-phase kinase-associated protein 2 antisense oligodeoxynucleotide (Skp2 ASODN) on the growth and proliferation of colorectal cancer cells SW480 and its mechanism .Methods The liposome-mediated dif-ferent concentrations of Skp2 ASODN was adopted to transfect SW480 cells ,the final concentrations of 0 .050 ,0 .125 ,0 .250 ,0 .500 , 1 .000 ,2 .000 ,4 .000 μmol/L were respectively set up with Skp2 nonsense oligodeoxynucleotides(NSODN) and blank group as con-trol .Then the inhibited effect of growth and proliferation of colorectal cancer cells were measured by the inverted microscope obser-vationandmethylthiazolyltetrazolium(MTT),thecellcyclewassurveyedbytheflowcytometry(FCM).TheexpressionofmRNA and protein of Skp2 and P27kip1 were inspected by reverse transcription-polymerase chain reaction(RT-PCR) and immunocytochem-istry methods .Results The inverted microscope observed that the SW480 cells had no change in form and grew at a slower speed . When the Skp2 ASODN concentration reached 0 .125 μmol/L ,the inhibition rate was 14 .48% (P0 .05) ,but its protein expression was elevates obviously (P<0 .01) .Conclusion Skp2 ASODN can inhibit the growth and proliferation of SW480 cells .Its possible mechanism is that the ubiq-uitin degradation of the Skp2 to P27kip1 is decreased by the gene occlusion effect of Skp2 ASODN ,thus the cell cycle progression is arrested .

Effects of SKP2 on the bystander effect induced by irradiated esophageal cancer cells / 中华放射医学与防护杂志

Objective To investigate the effect of SKP2 expression on radiation induced bystander effect (RIBE) of esophageal cancer cells.Methods The esophageal cancer cell lines with different SKP2 levels were applied for the study and the SKP2 expression was identified by Western blot.Micronuclei (MN) assay and DNA foci assay were used to evaluate the effect of SKP2 on RIBE.The cells were transfected with SKP2 gene or SKP2 siRNA to further verify the effect of SKP2 on RIBE.Results MN assay showed that the bystander effect induced by the cells with a high level of SKP2 was lower than that induced by the cells with a lower level of SKP2 (t =8.06,P ＜ 0.01).These results were further confirmed by the gene transfection experiments.When the expression of SKP2 was increased,RIBE was decreased (t=11.12,10.16,P ＜ 0.01).Contrarily,when the expression of SKP2 was reduced,RIBE was increased (t =8.39,8.83,P ＜ 0.01).γ-H2AX foci formation assay disclosed that when SKP2 expression in the irradiated cells increased,the repair ability of DNA damage in the bystander cells was higher than the control (t =6.85,7.10,P ＜ 0.01).With the expression of SKP2 decreased,the repair ability of DNA damage was lower than the control (t =7.66,8.47,P ＜ 0.01).Conclusions Over-expression of SKP2 inhibits RIBE of esophageal cancer cells,at least partly through regulating DNA damage repair ability.

Effect of intermittent high glucose on INS-1 cell proliferation and the expression of skp2-p27 / 中华内分泌代谢杂志

Objective To investigate the effect of intermittent high glucose on proliferation, apoptosis, and cell cycle progression of INS-1 cells, and the possible intracellular pathways activated by intermittent high glucose. Methods Cell viability was evaluated by cell counting kit, the cell cycle was determined by flow cytometry,Annexin-V/PI double-labeled cell apoptosis detection kit was used to monitor cell apoptosis. Cell cycle related protein Skp2 and p27 expressions were detected by Western blot. Results ( 1 ) Both intermittent and constant high glucose significantly inhibited the growth of INS-1 cells, and the former effect was more significant. ( 2 ) Intermittent and constant high glucose levels significantly increased apoptosis in INS-1 cells, and the former effect was more significant. (3) Intermittent and constant high glucose levels significantly inhibited the cell process, the G0/G1 cell cycle arrest also was induced by intermittent high glucose, resulting in lowered proportion of the G2/M phase and S phase of INS-1 cells. (4) Intermittent and constant high glucose significantly decreased the level of protein Skp2 and increased the level of cell cycle related protein p27. Conclusion Intermittent high glucose levels affect INS-1 cell growth and proliferation, as well as induce cell apoptosis, probably by decreasing the level of protein Skp2 and increasing the level of p27 in the cells, resulting in arrest of progression through the G1 phase to the S phase of INS1 cells, and thus impairment of cell proliferation.

Clinical significance of SKP2 and MRP-1/CD9 expression in glottic carcinoma and adjacent tissues / 临床耳鼻咽喉头颈外科杂志

Objective:To study the expression of SKP2 and MRP-1/CD9 protein in glottic cancer and adjacent tissues,and to analyze its significance for a safe surgical margin.Method:Thirty-eight cases of glottic squamous cell carcinoma were studied for its cancer tissue, tissue 2 mm, 5 mm , and 10 mm away from cancer ,and 10 cases of vocal cord polyp were served as control. SKP2 and MRP-1/CD9 protein were examined by immunoh istochemical method.Result:The positive expression of SKP2 proteins decreased in sequence of polyp mucosa, those adjacent to carcinoma (10 mm, 5 mm, 2 mm ) and cancer tissue, and there was significant difference between them(P<0.05);On the contrary, the positive expression of the MRP-1/CD9 proteins increased in sequence of polypusmucosa, those adjacent to carcinoma (10 mm,5 mm, 2 mm) and cancer tissue,and there was significant difference between them (P<0.05).Conclusion:SKP2 and MRP-1/CD may act as the reference index for judging the biological speciality of LSCC. It is appropriate to regard 5 mm or above 5 mm away from tumors as a safe margin for surgical treatment of glottic carcinoma.

Effects of IGF-1 on Proliferation of Human Non-small Cell Lung Cancer Cells / 中国医科大学学报

Objective To investigate the effects of insulin-like growth factor-1（IGF-1）on the cell proliferation of human non-small-cell lung cancer（NSCLC） and the possible molecular mechanism.Methods MTT assay was used to examine the effects of IGF-1 （0.1,1,10,100 ng/mL）on the cell proliferation of NSCLC cell lines（A549,LK2,H460）,Flow cytometry（FCM）and Western blot to ana-lyze the cell cycles and the protein expression of S-Phase Kinase-Associated Proteins 2（Skp2）and CDC20 homolog 1（CDH1）,respectively.Results The cell proliferation of NSCLC cell lines（A549,LK2,H460）could be promoted by the IGF-1 at different concentrations and the proliferation rate peaked when the cells were treated with 1 ng/mL IGF-1.Compared with control,the percentage of the S-phase cell population was significantly increased after the treatment of IGF-I（P 〈 0.01）and the protein expression of SKP2 also increased obviously（P 〈0.05）.However,there was no change in the CDH1 protein expression（P 〉 0.05）.Conclusion IGF-1 may accelerate the cell-cycle pro-gression of NSCLC cells by negatively modulating p27 protein via the up-regulation of SKP2 protein expression.

Correlation of Expression of galectin-3, skp2, p27 and cyclin D1 in Benign and Malignant Thyroid Lesions

BACKGROUND: The overexpression of cyclin D1 and galectin-3 and the loss of p27 in thyroid cancers have recently been reported by many studies. The S-phase kinase associated protein 2 (skp2) plays an important role in the degradation of p27. We compared the correlation of the expressions of galectin-3, p27, cyclin D1 and skp2 in thyroid lesions. METHODS: Sixty five cases were included in this study and immunohistochemical staining for galectin-3, skp2, p27 and cyclin D1 was performed. RESULTS: The expression of galectin-3 increased in the order of nodular hyperplasia, follicular adenoma, follicular carcinoma and papillary carcinoma (p<0.01). The expression rate of skp2 was 0% for nodular hyperplasia, 16.7% for follicular adenoma, 33.3% for follicular carcinoma and 16.7% for papillary carcinoma. The loss of the expression of p27 was more frequently detected in papillary carcinoma as compared with nodular hyperplasia (p<0.01). The increased expression of cyclin D1 was noted in follicular adenoma and carcinoma as compared with nodular hyperplasia (p=0.043). The expression of galectin-3 was related with the loss of a p27 expression (p<0.01), and the expression of skp2 was related with the expression of the cyclin D1 (p=0.022). CONCLUSIONS: Galectin-3 appears to be the most useful marker for making the diagnosis of thyroid lesions. The loss of a p27 expression can help differentiate nodular hyperplasia and papillary carcinoma, and the determining the expression of cyclin D1 may be helpful for the differential diagnosis of nodular hyperplasia and follicular neoplasm.

Expression of p27~(kip1),skp2,p53 gene in colorectal adenocarcinoma and its significance / 重庆医科大学学报

Objective:To investigate the expression of p27kip1,skp2 and p53 in colorectal carcinoma. Methods:The expression of p27kip1,skp2,p53 gene mRNA and protein in the progression of colorectal adenocarcinoma were investigated by reverse transcription polymerase chain reaction(RT-PCR) and immunohistochemistry. Results:The mRNA expression of p27kip1 and p53 in colorectal adenocarcinoma were lower than that in normal colon tissue(P

Prognostic Significance of p27(kip1) Expression in Node Negative Colorectal Carcinoma

PURPOSE: Dukes' A & B colorectal cancer patients are often excluded from adjuvant chemotherapy following potentially curative surgery because they are expected to have good long-term survival. However, actually 20 ~ 30% of these patients suffer from recurrent disease, so it would be helpful for these patients of recurrent disease to be able to select a high risk group. METHODS: In 78 Dukes' A & B colorectal cancers, we investigated by immunohistochemistry the role of molecular markers, such as p27(kip1), p53, Ki-67, and Skp2, in identifying high-risk patients. RESULTS: Patients with low p27(kip1) expression showed poor overall survival compared to those with high p27(kip1) expressions (55.3 versus 66.7 months, P=0.018). The only significant factor associated with p27(kip1) expression was p53 expression. The low p27(kip1) expression and positive p53 expression group had poor overall survival (54.3 months, P=0.036). CONCLISIONS: In a node-negative colorectal carcinoma, the molecular marker p27(kip1) does not play an independent prognostic role, but it may have prognostic significance in correlation with other markers such as p53, Ki-67, and Skp2. The assessment of molecular alterations may be useful to node-negative colorectal patients in identifying the high risk group that may benefit from adjuvant chemotherapy.

Expression of Skp2 and P21 in the Process of Breast Hyperplasia Cancerating and its Significance / 医学研究杂志

Objective To investigate the expression and role of Skp2 and P21 in the process of breast hyperplasia cancerizing.Methods The expression of Skp2 and P21 protein in normal breast tissue,adenosis of breast,breast atypical hyperplasia,breast hyperplasia cancerization,and breast cancer were detected by immunohistochemical staining SP method.Results(1) The expression of Skp2 and P21 in normal breast tissue and adenosis of breast was negative,the positive staining rate of Skp2 and P21 in breast atypical hyperplasia was 25.71% and 37.14%,in breast hyperplasia cancerization 63.16% and 73.68%,breast cancer,75% and 100% respectively.The contradistinction between breast atypical hyperplasia and breast hyperplasia cancerization was of significant statistically.But there was no statistical significance between breast hyperplasia cancerization and breast cancer.(2) The expression of P21 could lead to accelerated expression of Skp2.(?2= 10.024,P=0.002).There was a significant positive correlation between Skp2 and P21(r=0.563,P=0.000).Conclusion Skp2 and P21 are involed in the early stage of breast cancer and contribute to the maligant phenotype without affecting infiltration.Both of them are danger factors for breast hyperplasia cancerizing and contribute to malignant process from normal breast tissue and atypical hyperplasia to breast cancer.They are effective indicators for prognosis of breast hyperplasia.

Construction of Skp2 shRNA and its effect on lung cancer cell growth / 第三军医大学学报

Objective To construct and identify the RNAi eukaryotic vector of Skp2 gene and to observe its interfering effect on the growth of SPC-A-1 lung cancer cells.Methods The specific shRNA sequence was designed and synthesized according to the Skp2 cDNA sequence in GenBank.The sequence was cloned into plasmid pGenesil-1.Then recombinant vector was transfected into SPC-A-1 lung cancer cells by Lipofectamine 2000.The expressions of Skp2 mRNA were analyzed by RT-PCR and the levels of Skp2 protein were detected by Western blot.The cell growth suppression was analyzed by MTT assay.Distribution of cell cycle was assessed by flow cytometry.Results The sequence of template and specific siRNA was correct by sequence analysis.Obvious decrease was observed in the levels of Skp2 mRNA and Skp2 protein after Skp2 shRNA transfection(P

The effect of HGF-mediated Skp2 down-regulation on suppression of proliferation of human hepatocellular carcinoma cells / 中国普通外科杂志

Objective To investigate the effect of hepatocyte growth factor(HGF)on the proliferation of human hepatocellualr carcinoma cells,and the mechanism of HGF-induced proliferation inhibition.Methods Human hepatocellualr carcinoma cell line HepG2 were treated with different concentrations of HGF for different time periods,and the proliferation of these cells was examined by colorimetric BrdU cell proliferation enzyme-linked immunosorbent assay.The expression of S-phase kinase associated protein 2(Skp2)was examined using Western blot and RT-PCR.Plasmids pcDNASkp2 was introduced into HepG2 cells,then the clones showing up-regulation of Skp2 were selected,and the effect of HGF on the proliferation in these clones was investigated.Results HGF inhibited the proliferation of hepatoma cells in a dose and time dependent manner.The expression of Skp2 was significantly suppressed by HGF.Furthermore,HGF did not suppress the proliferation of HepG2 cells transfected with Skp2.Conclusions This study suggests that HGF could inhibit HepG2 cell proliferation,and the down-regulation of Skp2 could be closely related to this suppressed proliferation.

Expression and Clinical Significance of SKP2 and P27~(KIP1) Proteins in Human Gastric Cacinoma / 中国医师杂志

Objective To investigate the clinical significance of P27~ KIP1 and S-phase kinase-associated protein 2 (SKP2) expression in human gastric carcinoma. Methods The expression of P27~ KIP1 and SKP2 was determined by SP immunohistochemical method in 69 specimens of gastric carcinoma. Results The positive rates of P27~ KIP1 and SKP2 expression were 46.38% and 33.33%, respectively. The positive rate of P27~ KIP1 expression in gastric carcinoma decreased with the poor differentiation,deep invasion and progression of pathological grade (P0.05). There was a negative correlation between the P27~ KIP1 and SKP2 expression(P