The Effect Of Bortezomib And Associated With5-aza-CdR On The Expression Of NF-κB,SHP-1,JAKs Genes Of T-cell Lymphoma Cells

Objective: Lymphoma is a kind of entities malignant tumour whichoriginated from lymphocyte. According to the World Health OrganizationClassification of Tumor, there are currently nearly70distinguishablepathological types of lymphoma already known, which can be roughlycategorized into Hodgkin’s lymphoma and non-Hodgkin lymphoma. It is alsoknown that in China about90%of all the cases of lymphoma are non-Hodgkinlymphoma and this rate is still growing every year. Specifically, non-Hodgkinlymphoma can be further categorized into B cell and T/NK cell, and T-celllymphoma comprise about25%of all the cases of non-Hodgkin lymphoma.Bortezomib is a highly selective26S borate proteasome inhibitor, whichis currently widely applied to clinical practice to treat Multiple myeloma andMantle cell lymphoma. In a typical tumor cells, bortezomib mainly acts onseveral cellular proteins, such as nuclear transcription factor NF-κB, to beeffective. According to existing research results, by suppressing the activity ofproteasome26S subunit exclusively and reducing degradation of inhibitingfactor iκB significantly, bortezomib can facilitate the process of combinationof iκB and NF-κB and thus suppress activity of NF-κB, reduce the possibilityof NF-κB entering nucleus to induce or raise proliferation-related genes. As aresult, the expression of the genes related with cell proliferation can besuppressed, so that the excessive proliferation will be prevented and thenormal process of apoptosis will also be induced.Demonstrated by previous research,1) there is reduced or even missingexpression of SH2-containing tyrosine phosphatase-1in lymphoma patients’and lymphoma cells.2) Methylation inhibitor5-aza-CdR can suppress thereproduction of tumor cell by inducing SHP-1’s re-expression.3) Proteasome inhibitors bortezomib can strengthen5-aza-CdR’s demethylation to Jurkatcells. It is also verified that SHP-1can negative regulate JAK/STAT signal bysuppressing JAKs kinase activity, or make apoptosis by accelerating thedegradation of JAKs kinase through proteasome pathway.This experiment will focus on1) the expression of NF-κB gene withinT-cell lymphoma Jurkat cell to which bortezomib is applied.2) The expressionof SHP-1, JAK1, JAK2, JAK3and TYK2within Jurkat cell to whichbortezomib with/without5-aza-CdR is applied.3) To know more about therole SHP-1plays in JAK/STAT pathway and T-cell lymphoma.Methods: In this study, the investigation object were the T cell NHL cellline Jurkat. The cells were suspension cultured in RPMI1640with10%calfserum. These cells in exponential phase of growth were transferred to the cellcultivable board, there were1×106/ml cells in each hole. The concentration ofBTZ were10nmol/L,30nmol/L,50nmol/L (all3holes); BTZ10nmol/L and5-aza-CdR3μmol/L(1hole); no drug in1hole. There were two holes cellswith5different treated conditions. These cells with drugs were incubatedunder37℃,5%CO2in24h,48h,72h. After that, cells were harvested, washed,extracted total RNA, detected the expression of NF-κB,SHP-1and JAKsfamily (JAK1,JAK2,JAK3,TYK2) with RQ-PCR. Then process statisticalanalysis.Results:1The expression of NF-κB in Jurkat cells after treated by BTZ:the results of RQ-PCR showed when Jurkat was treated with the sameconcentration of BTZ, with the extension of time, the expression of NF-κBwas reduced, while the same time treated Jurkat, with the increase ofconcentration, the expression of NF-κB was reduced. It had statisticalsignificance.(P<0.05)2The expression of SHP-1in Jurkat cells after treated by BTZ and5-aza-CdR: the results of RQ-PCR showed when Jurkat was treated with thesame concentration of BTZ, with the extension of time, the expression ofSHP-1compared with control group was increased, while the same timetreated Jurkat, with the increase of concentration, the expression of SHP-1was increased. It had statistical significance (P<0.05). When Jurkat was treatedwith BTZ and5-aza-CdR, the increase of expression of SHP-1was obvious,the expression of combine group was higher than the single group (P<0.05).3The expression of JAKs in Jurkat cells after treated by BTZ and5-aza-CdR: the results of RQ-PCR showed when Jurkat was treated with thesame concentration of BTZ, with the extension of time, the expression ofJAKs compared with control group was reduced, while the same time treatedJurkat, with the increase of concentration, the expression of JAKs was reduced.It had statistical significance (P<0.05). When Jurkat was treated with BTZ and5-aza-CdR, the reduce of expression of JAKs was obvious, the expression ofcombine group was lower than the single group (P<0.05).4The dependence of SHP-1and JAKs (JAK1, JAK2, JAK3, TYK2): theexpression of SHP-1and JAKs was a negative correlation. The JAK1declineis more obvious.Conclusions:1Bortezomib can inhibit the expression ofproliferation-related gene and induct apoptosis of the Jurkat cells. This effectmay be through the way of inhibition of NF-κB pathway, reducing the amountof NF-κB into the nucleus.2Bortezomib and5-aza-CdR increase of SHP-1mRNA expression inJurkat cells, reduce the expression of the JAKs family (JAK1, JAK2, JAK3and TYK2), a dose and time dependence. And the two drugs have a synergisticeffect. The expression of tumor suppressor gene SHP-1was elevated, possiblythrough inhibition of JAK/STAT signaling pathway, regulating genesassociated with cell proliferation and differentiation, thus inhibiting tumor cellgrowth and induce apoptosis. The effect of JAK1was more significant.