Ropivacaine Disrupt Autophagy, Inhibit Tumor Growth, And Relieve Mechanical Pain

Background: New functions of old drugs,especially for cancer treatment,are rise increasing concerns,but few of them can simultaneously relieve caner pain.Autophagy,a crucial biological degradation process,is closely associated with the pathogenesis of cancer.Autophagy mediates the deregulation of defective organelles,misfolded or aggregated proteins,and certain long-lived molecules to regulate cellular homeostasis.Autophagy plays a dual role in cancer,inhibiting apoptosis and promoting the growth of a given tumor,and inhibiting the accumulation of damaged proteins and organelles to inhibit tumor growth.The microenvironment in which the tumor is located is lacking in nutrients.Under the pressure of nutrient deficiency,cells can gain energy through autophagy,and they can get through the "difficulty".Therefore,by regulating autophagy,it may inhibit the development and metastasis of cancer,kill cancer cells,and enhance the anti-tumor effect of chemotherapy drugs.Ropivacaine is a clinically used aminoamide local anesthetic,mainly used for surgical regional block and epidural anesthesia and epidural postoperative or labor analgesia.In this study we mainly discusses the new role of ropivacaine in cancer treatment and its relationship with autophagy.Methods: Hela cells and B16 cells were divided into control group,starvation group,ropivacaine group,chloroquine group,starvation + ropivacaine group,starvation+chloroquine group,and ropivacaine + chloroquine group,were treated for 0,4,12,24 hours.The 6-8 weeks old male C57BL/6 mice were randomly divided into control group,caloric restriction group,ropivacaine group,and ropivacaine + caloric restriction group,treated for one week,then mechanical pain was detected and tumor tissues weretaken.TUNEL staining,Hoechst 33342/PI staining and MTT were used to detect cell viability and apoptosis,and immunofluorescence labeling LC3,LAMP1 and clathrin were used to observe the autophagy accumulation level,morphology of autophagy-lysosome and autophagic lysosome reformation.Western-blot was used to detect the expression of LC3?,p62,Cath D.Results:Compared with the control group,ropivacaine and starvation group caused autophagosome formation,autophagy marker LC3 II expression and LC3 fluorescence point aggregation increased;In starvation group,most autophagosomes produce new lysosomes,whereas in the ropivacaine or ropivacaine + starvation group,autophagosomes continue to exist,as indicated by the overlapping signals of LAMP1 and LC3 and the colocalization between clathrin fluorescence and LAMP1 fluorescence was no significant change;compared with the control group,the ropivacaine or ropivacaine + starvation group destroyed autophagic degradation,the p62,LC3 II levels increased significantly and Cath D levels decreased.In the in vivo experiment,compared with the control group,the tumor volume of the ropivacaine + caloric restriction group were significantly reduced,and the mechanical stimulation threshold was significantly increased.Conclusion: Ropivacaine impaired the atuophagic lysosome reformation and disrupted the autopahgic degradation,which is involved in its killing of cancer cells in starved environment.In a mouse model of melanoma inoculated near the sciatic nerve,ropivacaine administration cooperated with calorie restriction efficiently suppressed tumor growth.Simultaneously,at early stage of tumor growth,ropivacaine administration provisionally relieved tumor-induced mechanical pain;and at later stage,a synergistic therapy of caloric restriction and ropivacaine enduringly reduced the mechanical pain.