Study On The Regulation Of JA And DNA Topoisomerase Ⅱα And The Down - Regulation Of All - Trans Retinoic Acid On DNA Topoisomerase

Objective:To investigate the interaction between JWA and TopoⅡα; to explore if the interaction of JWA and TopoⅡα impact on JWA’s repair capacity of DNA damage repair and related signaling molecules such as Akt, p-Akt, NF-κB.Methods:Over expression of JWA or TopoⅡα was obtained by transfection of JWA/TopoⅡα plasmid in the cells. Knockdown of JWA or TopoⅡα was achieved by transfection of siRNA targeting JWA and TopoIIa. Cells were treated with hydrogen peroxide to form an oxidative damaged cell model; JWA and TopoIIa levels, and Akt, p-Akt, NF-κB levels were detected by Western blot and reverse transcription polymerase chain reaction.Results:(1) In HeLa cells, there was a converse-regulation between JWA and TopoⅡα. Transfection of TopoⅡα decreased JWA protein and mRNA levels. That means TopoⅡα downregulated JWA; knockdown of JWA increased TopoⅡα protein and mRNA levels which means JWA downregulated TopoⅡα;(2) In human gastric cancer BGC823 cells, human ovarian cancer A2780 cells and hunman lung cancer NCI-H460 cells, there also shown a converse-regulation between JWA and TopoⅡα. TopoⅡα downregulated JWA protein level in a dose-dependent manner. Also, JWA downregulated TopoⅡα protein level in a dose-dependent manner;(3) Compared with wild-type mice, the TopoⅡα level in lung tissues in the JWA knockdown mice was higher. Compared with the adjacent normal tissues, the TopoⅡα level in human cervical cancer tumor tissues was higher;(4) Co-immunoprecipitation and immunofluorescence analysis showed that there was no direct interaction between JWA and TopoIIa. They did not affect each other’s cellular localization;(5) In HeLa cells, hydrogen peroxide upregulated p-Akt, NF-κB, but transfection of JWA inhibited the increase of p-Akt and NF-κB. The converse-regulation between JWA and TopoⅡα affect JWA. However, hydrogen peroxide and JWA did not impact Akt obviously.Conclusion:JWA and TopoIIa downregulated each other in several tumor cell lines. There was no direct interaction between JWA and TopoIIa. The converse-regulation between JWA and TopoⅡα affects JWA in reducing p-Akt and NF-κB. In other words, the converse-regulation may affect the ability of JWA in oxidative damage repair.Objective:To explore the mechanism of the downregulation of ovarian cancer tumor marker topoisomerase Ⅱα (TopoⅡα), the upregulation of retinoic acid receptors by all-trans retinoic acid (ATRA) and the relationship between the level of TopoⅡα in ovarian cancer tissues and the level of serum ATRA.Methods:Various concentrations of ATRA from 0-10μM were treated to the human ovarian cancer A2780 and SKOV3 cells. After 24h, TopoⅡα protein level was detected with Western blot; various concentrations of ATRA from 0-2μM were treated to the human cervical cancer HeLa cells. After 24h, retinoic acid receptors (RARα、RARβ、RARγand RXRα) protein level were detected with Western blot; TopoⅡα expression in ovarian cancer tissues were detected by immunohistochemistry; the level of serum ATRA in ovarian cancer patients were detected by High Performance Liquid Chromatography (HPLC).Results:In human ovarian cancer A2780 and SKOV3 cells, ATRA could downregulate TopoⅡα protein level. In human cervical cancer HeLa cells, ATRA could upregulate retinoic acid receptors (RARα、RARβ、RARγ and RXRα) protein level. The expression of TopoIIa in ovarian cancer tissues had a negative correlation with the level of serum ATRA in patients (r=-0.63, P<0.05).Conclusions:ATRA could downregulate TopoⅡα protein expression in human ovarian cancer cells (A2780 and SKOV3). ATRA could upregulate retinoic acid receptors (RARα、RARβ、RARγ and RXRα) protein expression in human cervical cancer cells (HeLa). The carcinogenic effects of ovarian cancer may be related to the decreased level of serum ATRA.