| BackgroundProstate cancer is the most common cancer, and the incidence ranks six of all malignant tumors and two among men on a global scale.In the mean time, the growth rate of morbidity ranks six in2012compared with eight in2001. The morbidity of Occident is above10/1000000, and in the USA,130000people have the cancer every year. Although the incidence exceed it in our country notablely, the epidemiological investigation shown that the incidence of some metropolitans has got close to the Occident’s. The phenomenon may be closely linked to the aging population,the change of dietary structure, the pollution of environment, and the development of detecting techniques.The research for prostate cancer is increasing as the the significant growth of the incidence of prostate cancer. All of them focuses on the signaling pathway, the mutations of androgen receptor, and the amplification of gene. Some studies have shown that, the activation and upregulation of the original oncogene ras Fos, c-myc, c-met, Bcl-2, the inactivation of tumor suppressor genes Rb, p53, p16, PTEN, CDKN2, the abnormal expression of protein AR,PSCA,the abnormal control of promoter methylation and DNA-damage response pathway may be involved in the process of prostate cancer development. Recent studies have found that receptor for advanced glycation end products (RAGE) and prostate cancer have closely relationship.RAGE encodes genes in the major histocompatibility complex class III region. In the past20years, RAGE is considered to play an important role in diabetes, sepsis, inflammation and development of tumors. RAGE, as a signal transduction receptor, binds in the surface of mononuclear macrophages, endothelial cells, kidney mesangial cells, nerve cells, vascular smooth muscle cellsand a variety of tumor cells with its ligand, actives signal transduction pathway within cells and results in pathological effects.The present study suggests that, the expression of RAGE is upregulated in many tumors in addition to the lung tumor and it’s closely related with the tumorigenesis, development and the poor prognosis of tumors.Rb gene is one of the suppressor gene which has been found originally, and it encodes a nuclear protein product that is110kd with ability of binding DNA. RB protein plays a negative role in the regulation of cell growth cycle. Non-phosphorylation Rb protein plays an anti-cancer role, while the phosphorylation Rb protein doesn’t. Rb protein inhibit the activity of E2F by binding to it, make the cell stay on G1phase, and inhibit cell proliferation and induction of apoptosis finally. Rb protein would disassociate from transcription factor E2F when being phosphorylated. And the DNA transcription of the relative enzymes would start as the disassociation finished which impels the cell from G1phase to S phase and causes cell proliferation. But in prostate cancer, the loss of the most Rb allele leads to the functional defection of the Rb protein. Clinical observation shows that60%prostate cancer case have lost Rb heterozygous. Maddison’s studies suggest that the inactivation of Rb would increase the expression of the E2F and promote the progression and metastasis of prostate cancer. Taneja SS and co-workers found that androgen may involve in the process of degradation of Rb protein in the ubiquitin-proteasome system. And the degradation process could be accelerated by the Rb phosphorylation which induced by cyclin A/Cdk2and cyclin B/Cdkl. Also, this will promote the growth of LNCap cells. On the other hand, some studies show that Rb protein can specifically combine with the androgen receptor protein to enhance the transcription of it, thus promote the progression of prostate cancer.In our previous work, we demonstrated that the interaction between AGE and RAGE can promote the proliferation of prostate cancer cells, and this effect may be related with the expression of RB protein, but the specific mode of action is not yet clear. Ning’s studies have shown that certain proteins related to cell cycle regulation, such as RB may be affected by the AKT signaling pathway, so we hypothesized that the changes of RB protein may be caused by the AKT pathway.Akt is the congener of proto-oncogene v-Akt, that present in the human chromosome homologue, and also a proto-oncogene in essence. It encodes the protein PKB, the serine/threonine protein kinase of about57kd, that is the core protein in PI3K/Akt signal transduction pathway. The three kinds of subtypes of Akt family, AM/PKBa, Akt2/PKBp, Akt3/PKB7, located in14q32.32,19ql3.1-q13.2,lq44respectively. The three subtypes could active the downstream substrates, and Akt2play the most obvious role among them. The three subtypes have not only more than80%homology to the amino acid sequence, but also common structural features. The common structure comprises three different functional areas, the pH region in the amino terminal, the important kinase area of the central region and the regulatory region in carboxyl terminal. Three functional areas play important roles in PI3K/Akt signaling pathway respectively.The PH region contains about100amino acid sequence and the binding domain for3-phosphoinositides, and mainly regulates the interactions between protein-lipid and protein-protein. The the important kinase area of the central region has a high degree of similarity with the PKA, PKC, and Thr-308site, necessary for the activation of Akt, locates in this region. The regulatory region of the carboxyl terminal composes of40amino acid sequence, and it contains a characteristic hydrophobic motif FXXF/YS/TY/F (X is any amino acid) of protein kinase and the serine/threonine residues that its phosphorylation is necessary for the full activation of Akt.Studies have shown that activation of Akt is involved in the development of tumors. The possible mechanism is that the amplification of the PI3K protein due to coding genes PIK3CA and various other factors have led to the excessive activation of Akt, or the missed fuction of some regulatory elements, such as PTEN. There is excessive activation of Akt in prostate cancer, ovarian cancer, endometrial cancer, pancreatic cancer, breast cancer, lung cancer, follicular thyroid cancer, multiple myeloma, malignant melanoma. There is also missed fuction of PTEN in about50%to80%of patients with prostate cancer, results in overexpression of a second messenger (PIP3).After that, Akt could be excessively active and enhance the survival of prostate cancer cells finally. Gao confirmed Aktl could interfer the combination between the APC/Cdhl and Skp2and promote the tumor progression in prostate cancer patients.Akt could inhibite the apoptosis of cells, that is also importment in tumor progression. Aktinhibit the apoptosis of cells in various maners.And there is interaction between AKT and proteins related with apoptosis, such as Bcl-2family, Foxo family, IAPs (inhibitor of apop-tosis proteins) and caspase-9. In short, AKT, the important molecular in PI3K/Akt pathway, play an importment role in growth and proliferation, invasion and infiltration, and inhibition of apoptosis of prostate cancer cells.We have initially proved the relationship between RAGE and Rb in prostate cancer. But whether the interaction is direct or indirect? What kind of manner the interaction rely on? What is the significance of this interaction in prostate cancer?After these question being answered, We will not only be able to know more about functions of RAGE and Rb, but also can provide theoretical basis for the prevention and cure for prostate cancer and even other related diseases due to the new ideas of the this disease.MethodsTo work these out, we detected the expression of RAGE in the cytoplasm and nuclei of PC-3cells by Western Blott in prostate cancer cell lines to clear the positioning of RAGE, and to speculate the way of interaction between RAGE and RB protein; Moreover, prostate cancer cell lines were stimulated with AGEs in different concentration and timing to observe the variation of Rb protein and the phosphorylation of its Ser-807/811sites. The variation of AKT protein and the phosphorylation of its Ser473, Thr308sites were then detected under the same conditions. After that, we use siRNA to block the expression of RB protein in PC-3cells, for laying the foundation for the further experiments. Finally, we constructed eukaryotic expression vectors for RAGE without V or VC1domain in order to determine the conditions of the interaction between RAGE and Rb protein.ResultsThrough the research above, we have got the following results:1, RAGE mainly located in the cytoplasm of prostate cancer cells;2, AGEs could downregulate the expression of RB protein and also enhanced the phosphorylation of it in prostate cancer. These effect are possibly made by the AKT protein, activated by the interaction between RAGE and AGEs3, The upregulation of phosphorylation of AKT protein was also observe in prostate cancer under the same condition. The interaction between RAGE and AGEs may mediate these effect.4, The expression of RB protein successfully have been blocked in prostate cancer with the siRNA;5The eukaryotic expression vectors for RAGE without V or VC1domain have successfully been constructed.ConclusionsIn this study, a preliminary study of the interaction between RAGE and the Rb protein was made. And AGEs could upregulate the phosphorylation of AKT protein, this result in downregulating of the expression of RB protein and the upregulating of phosphorylation of it in prostate cancer. Finally this effect promoted the proliferation of prostate cancer. These findings will not only deepen our understanding of the biological function of RAGE and Rb protein, but also fundamentally affect and change the understanding and treatment options for prostate cancer on clinical. |
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