ABSTRACT
The expression of Rap1 GAP protein was detected in 69 cases of papillary thyroid carcinoma and adjacent normal thyroid tissues with immunohistochemistry method.Methylation of Rapl GAP gene was analyzed by methylation-specific-PCR (MSP) in these tissues.The immunohistochemistry results indicated that the Rap1GAP protein was down-regulated in tumor tissues of 54 ( 78％ ) cases compared with normal thyroid tissues.Statistical analysis demonstrated that the decreased level of Rap1 GAP protein expression was significantly correlated with the tumor stage T according to American Joint Committee on Cancer ( AJCC,P =0.043 ).The MSP results demonstrated that 46 cases of Rap1GAP gene methylation were detected in 54 cases with down-regulated expression of Rap1GAP (66.7％),but only 1 case of methylation was found in 15 cases without obvious change of Rap1GAP expression (6.67％),showing significant difference ( P＜0.01 ).There was no methylation in normal thyroid tissues.These results suggest that raised methylation of promoter region may contribute to the low expression of Rap1GAP protein in papillary thyroid carcinoma.
ABSTRACT
Rap 1 is expressed in human umbilical vein endothelial cells (HUVECs).Rap1-GTPase activating protein (Rap 1 GAP),with its specific target,Rap 1,has been shown to be important in the regulation of many physiological and certain pathological processes.In this study,we investigated the effect of RaplGAP expression on endothelial cell function,or,more specifically,proliferation and migration of endothelial cells.HUVECs were transfected with pcDNA3.1 (empty vector),pcDNA3.1 containing Flag-tagged-Rap1GAP or Myc-tagged-Rap1N17.The proliferation,migration and tube formation were examined and compared among the 3 groups.Expression of Rap1,Rap1GAP,extracellular signal-regulated kinase (ERK),phospho-ERK,Akt,phosphor-Akt was detected by Western blotting.The results showed that the proliferation,migration and tube formation were significantly reduced in Rap1GAP- and Rap1N17-transfected HUVECs as compared with empty vector-transfected control.These changes were coincident with increased expression of Rap 1 GAP and decreased expression of activated Rap 1,phospho-ERK and -Akt.After treatment of Rap 1 GAP-transfected HUVECs with a stimulator of Rapl guanine-nucleotide-exchange factor (Rap1GEF) 8CPT-2'OMe-cAMP,it was found that Rap1 activity was decreased as compared with empty vector-transfected control.Pretreatment of HUVECs with an ERK inhibitor PD98059 or a PI3K inhibitor LY294002 prior to stimulation not only blocked 8CPT-2'OMe-cAMP-induced phosphorylation of ERK and Akt,but also significantly reduced cell proliferation and migration.Finally,we examined the effect of vascular endothelial growth factor (VEGF) on HUVECs overexpressing Rap1GAP.VEGF-stimulated Rap1 activity,phosphorylation of ERK and Akt,cyclin D1 expression and cell proliferation were repressed in HUVECs overexpressing Rap 1 GAP as compared to empty vector-transfected control.Taken together,our findings demonstrate that Rap1GAP/Rap1 and their downstream effectors regulate proliferation and migration of HUVECs via ERK and Akt pathways.
ABSTRACT
To find the underlying causes of primary myelodysplastic syndrome (MDS), the gene expression profiling of both CD34+ cells and bone marrow mononuclear cells from MDS patients was performed using oligonucleotide microarray and cDNA microarrays, respectively. Several candidate genes which were differentially expressed in MDS patients versus normal controls were selected and confirmed in expanding samples by quantitative real-time reverse transcription-polymerase chain reaction after clustering and bioinformatics analysis. one of these genes, RAP1GAP, was found to be expressed at a significantly higher level in patients with MDS in comparison with those suffering from other hematopoietic diseases including leukemia (P < 0.01). We propose that over-expression of RAP1GAP gene may play a role in the pathogenesis of MDS.