Research Of Parthenolide On Proliferation And Apoptosis Of KSHV Infected Of Cells

Objective:To investigate the effect of parthnolide?PTL?on the proliferation and apoptosis of Kaposi sarcoma-associated herpesvirus?KSHV?infected cells.Methods:?1?CCK-8 was used to determine the half inhibitory concentration(IC₅₀)of different concentrations of parthenolide-treated KSHV infected cells for 24 h.?2?Observed the morphological changes of KSHV-infected cells treated with different concentrations of parthenolide at 24h and 48h using the microscope.?3?CCK-8 and colony formation experiments were used to determine the proliferation of KSHV-infected cells treated with different concentrations of parthenolide.?4?Cell cycle changes of KSHV-infected cells treated with parthenolide were detected by flow cytometry.And the expression levels of CDK4,CDK6,Cyclin D1 were detected by Western Blot after different concentrations of parthenolide-treated KSHV infected cells.?5?The expression levels of Bax,Bcl-2 were detected by Western Blot.?6?Western Blot was used to determine the expression levels of IKK?,P-p65,and P-IKB-?.?7?The effect of parthenolide on tumor growth in vivo was measured by animal experiment.?8?The effects of parthenolide on CDK6,CyclinD1,Bax,IKK?,P-p65,and P-IKB-?expression were detected by immunohistochemistry.?9?After treatment of KSHV-infected cells with parthenolide,mRNA expressions of latent genes LANA,V-FLIP,V-Cyclin and lysed genes RTA,K8.1,and V-GPCR were detected by real-time fluorescence quantitative PCR.?10?Immunocytochemistry?ICC?was used to detect the expression levels of latent gene LANA,lytic gene K8.1A,and RTA after treatment of KSHV-infected cells with parthenolide.Results:?1?The IC₅₀0 of KSHV latently infected cells iSLK.219 and SK-RG cells were 21.08?mol/L and 10.75?mol/L,respectively.?2?Parthenolide changed the morphology of KSHV infected cells,causing iSLK.219 cells to become swollen,round,and shed.While SK-RG cells became round,and cytoplasmic protrusions were reduced or even disappeared.?3?Parthenolide significantly reduced the cell proliferation ability,and had a significant G1cell cycle arrest in KSHV latently infected cells.At the same time,the expression of cyclin Cyclin D1,CDK6,and CDK4 was reduced by parthenolide in KSHV latently infected cells.?4?Parthenolide promoted the apoptosis,reduced the expression of the survival gene Bcl-2,and increased the expression of the pro-apoptotic gene Bax in KSHV latently infected cells in a concentration-dependent manner.?5?Parthenolide reduced the expression of NF-?B-related proteins IKK?,P-p65,and P-IKB-?in KSHV latently infected cells in a concentration-dependent manner.?6?Intraperitoneal injection of 5 mg/kg of parthenolide had a tendency to reduce tumor volume in nude mice.The expression of apoptosis-associated factor Bax increased,and the expression of cell cycle factors Cyclin D1,CDK6,and NF-?B-related factors IKK?,P-p65,and P-IKB-?decreased in the parthenolide treatment group.?7?Induction of phorbol myristate acetate?PMA?for 48 hours can make SK-RG cells enter the lysing replication phase.?8?The IC₅₀0 of KSHV lystic cells Lytic iSLK.219 and Lytic SK-RG cells were 33.51?mol/L and 28.53?mol/L,respectively.?9?Parthenolide significantly reduced the cell proliferation ability,and promoted apoptosis,reduced the expression of survival gene Bcl-2,and increased the expression of pro-apoptotic gene Bax in of KSHV lytic cells in a concentration-dependent manner.?10?Parthenolide reduced he expression of KSHV virus gene LANA,RTA,K8.1and V-GPCR.Conclusion:?1?Parthenolide inhibits proliferation and promotes apoptosis of both KSHV latent and lytic cells.?2?Parthenolide inhibits the activity of cyclinD1 and CDK4/6 kinase complex,and has a significant G1 phase arrest effect on KSHV latently infected cells.?3?Parthenolide down-regulated the expression of NF-?B signaling pathway-related proteins in KSHV latently infected cells,which may be related with inhibiting cell proliferation and promoting apoptosis.?4?Parthenolide inhibits KSHV virus gene expression and may be a candidate drug for KSHV infection and related diseases.