| Objective To establish a stable and strong repeatable murine bonecancer pain model which is extremly similar with human and toinvestigate the role of NR1 and NOS in the generation and mainterance ofbone cancer pain.Methods The first part of the experiment---Establishment andevaluation of the murine model of bone cancer pain: Thirty maleC57BL/6 mice, weighing about 18-20g, were randomly divided into fourgroup: cancer cell inoculated group (15 mice), 2×10~6 cell in 10 ul PBSwere injected into murine left femur. heat-killed group (5 mice), the samenumber of heat-killed cancer cells in 10 ul PBS were injected into murineleft femur. PBS group(5 mice), no cells were injected, but 10 ul PBSinjected into murine left femur, normal control group(5 mice), give notreatment. Spontaneous lifting time and mechanical allodynia threshold ofmice hind paw were measured in alternative days during throughoutexperiment. The structural damage of the left femur was monitored byradiological analysis and observed respectively on the 7th, 15th, 23rd day.Destruction of bone in mice of cancer cell inoculation group was stainedwith HE, and observed on the same day, but mice in other group onlyobserved on 23rd day.The second part of the experiment---The expression and significanceof NR1 and NOS in the murine bone cancer pain model: Based on thesucessful murine model of bone cancer pain in the first part of experiment,another 30 clear male C57BL/6 mice were randomly divided into a modelgroup (A group): 2×10~6 cell in 10 ul PBS were injected into the left femur,a control group(B group): 10 ul PBS injected the left femur and a blankcontrol group(C group): give no treatment. There were 10 mice in eachgroup. 23 days after inoculation, segments of lumbar spinal cord weretaken to detect NR1 by immunohistochemistry and to mensure the NOSby using NOS kit produced in Nanjing Jiancheng BioengineeringInstitute.Result 1) Mice following intra-femur injections of Lewis lungcarcinoma cells displayed the gradual development of spontaneous pain, beginning on day 11 after injection, then mechanical allodynia camealong. The course of flinch last more time in later experimental session;The mechanical allodynia displayed 50％von Frey threshold issignificantly decreasing on day 13 after injection and lasting the wholeexperimental period. On the 23rd day after operation, X-ray showed thatmedullary cavity of low femur on ipsilateral side disappeared and muchcortical bone lost,or ever abscised. Meanwhile, tumor cells penetratedthrough the medullary canal and invaded peripheral muscles. This modelwere successful established and confirmed by all of ethology,radiologyand histology.2) The immunohistochemistry result of NR1 and the expressionlevel of NOS in spinal cord: The actual staining gray scale and IOD ofNR1 in model group were 95.400±6.076, 11.383±2.277 respectively, thecontrol group were 58.257±9.781, 6.283±1.900 respectively and theblank control group were 52.689±10.290, 5.972±1.352 respectively, therewere significant differences in the actual staining gray scale and IODbetween model group and blank control group, also between model groupand control group (P＜0.05), but there were no differences betweencontrol group and blank control group(P＞0.05). The expression level ofNOS in model group was 2.842±0.663, in control group was 1.770±0.149,in blank control group was 1.445±0.134. There also were obviousdifferences either between model group and blank control group, orbetween model group and control group (P＜0.05), but no differencesbetween control group and blank control group(P＞0.05).Conclusion Inoculating with Lewis lung carcinoma cell intomedullary canal of femur in C57BL/6 mice can succeed to establish amurine bone cancer pain model, the murine model we have developedappears to share many similarities with human cancer-induced bone pain.On the dorsal horn of murine spinal cord, the number of NR1 wasincreased and the expression of NOS was elevateded increased, so theNR1 and NOS in spinal cord may participate and mediate in transmittingalgesthesia message and the forming of hyperalgesia in bone cancer pain.
|
PDF Full Text Request