P2X4受体在大鼠胫骨癌痛中的变化及可能机制

CHANGES OF P2X4 RECEPTOR EXPRESSION IN RAT TIBIAL BONE PAIN AND ITS POSSIBLE MECHANISM

目的:观察骨癌痛大鼠脊髓P2X4受体表达的变化,并探讨其可能机制。方法:72只SD大鼠随机分为两部分(6组),第一部分:空白对照组(K组,n=14)、假手术组(J组,n=14)、模型组(M组,n=20)。K组不给予任何处理,J组和M组分别在左胫骨上端注射PBS液和Walker256乳腺癌细胞5μl(2×107/ml)。各组分别于术前1天,术后3、6、9、12、15、18天时随机各取8只大鼠测定左后肢机械缩足反射阈值(MWT)。K组和J组于术后12天,M组于术后6、12、18天时各处死6只大鼠,取L4～6脊髓组织,用免疫组化法检测P2X4受体表达;第二部分:溶媒对照组、TNP-ATP组、PPADS组,每组8只。各组从骨癌痛造模术后第7天起,连续5天分别鞘内给予双蒸水(ddH2O)、PPADS(100nmol)、TNP-ATP(30nmol)各10μl,并于建模后第12天取L4～6脊髓,用免疫组织化学方法检测脊髓P2X4受体及p-ERK的表达。结果:与K组和J组比较,M组大鼠术后第6～18天,左后肢MWT进行性下降,P2X4受体(P2X4R)阳性细胞数明显增加(P<0.01)。连续5天鞘内注射TNP-ATP可抑制由胫骨癌痛引起的脊髓P2X4受体及p-ERK表达增加(P<0.05),而PPADS组和ddH2O溶剂组却没有此种效应(P>0.05)。结论:脊髓P2X4受体表达上调可能参与了大鼠胫骨癌痛的产生和维持,其机制可能与激活下游ERK通路有关。

Objective： To observe the changes of P2X4 receptor expression in rat tibial bone pain and explore its possible mechanisms.Methods： SD rats are randomly divided into two parts,in the first part are Control group（K,n=14）,Sham-operated group（J,n=14） and Model group（M,n=20）.K group is not given any treatments,J and M groups are injected 5μl（2×107/ml）PBS solution or Walker 256 breast cancer cells into the left proximal tibia,respectively.On the day before and 3,6,9,12,15,18d after the operation,eight rats selected from K,J and M groups were detected by MWT filaments on the left hind-limb,respectively.Six rats of K and J groups were executed at the 12th days after the operation and the 6th,12th and 18th days after the operation of M group,to observe the P2X4 receptor and p-ERK expression in L4～6 spinal cord tissue by immunohistochemical methods,respectively.In second part,twenty-four rats are randomly divided into 3 groups（n=8）：Solvent control group,PPADS group and TNP-ATP group.10μl of ddH2O、PPADS（100nmol）and TNP-ATP（30nmol） were intrathecally injection on the seventh day after operation for 5 days,respectively.The P2X4 receptor and ERK expressions in L4～6 spinal cord tissue were observed at the twelfth day after modeling by immunohistochemical methods.Results： Compared with K and J groups,MWT of the left hind-limb declined,while P2X4 receptor-positive cells significantly increased on the 6th to 18th day after operation of M group（P0.01）.The P2X4 receptor and p-ERK expressions of spinal cord caused by tibial bone pain were inhibited by intrathecal injection of TNP-ATP for 5 days（P0.05）,while PPADS and ddH2O groups had no such effects in PPADS and ddH2O groups.Conclusion： P2X4 receptor expression in spinal cord might be involved in the generation and maintenance of rat tibial bone cancer pain,which might be related to the activation of downstream ERK pathway.