| Objective To investigate the Manifestation of ethology and the immunohistochemistry result of NR2B in spinal cord dorsal horn and DRG of bone cancer pain model in mice together with the correlation between the ethology and the immunohistochemistry result of NR2B,then to discuss the role of NR2B in the generation and maintenance of bone cancer pain.Methods Forty-five male C57BL/6 mice,weighing about 18-20g, were randomly divided into three group:cancer cell inoculated group(15 mice),2×10~6 cell in 10 ul D-HANK'S were injected into mice left femur. sham group(15 mice),no cells were injected,but 10 ul D-HANK'S injected into mice left femur,normal control group(15 mice),give no treatment.Spontaneous lifting duration,ambulatory score and mechanical withdrawal threshold of mice hind paw were measured in alternative days during throughout experiment.Destruction of bone in mice of cancer cell inoculation group,sham group and normal group were stained with HE, and observed on the 7th,15th,23rd day.the tissues of cancer distal metastasis also were stained with HE including lung and kidney.The expression and significance of NR2B in the mice bone cancer pain model:Based on the sucessful mice model of bone cancer pain, There were 5 mice in each group.7,15,23 days after inoculation, segments of lumbar spinal cord and L4 DRG were taken to detect NR2B by immunohistochemistry.Result1)Manifestation of ethology:mice following intra-femur injections of Lewis lung carcinoma cells displayed the gradual development of spontaneous pain,beginning on day 7 after injection,then mechanical withdrawal came along,ambulatory score started to lower. The course of flinch last more time in later experimental session;The mechanical withdrawal displayed von Frey hair threshold is significantly decreasing on day 15 after inoculation and lasting the whole experimental period.On the 23rd day after operation,bone tissue of HE staining showed that medullary cavity of low femur on ipsilateral side disappeared and much cortical bone lost,or ever abscised.Meanwhile, tumor cells penetrated through the medullary canal and invaded peripheral muscles.This model were successful established and confirmed by all of ethology and histology.2)The immunohistochemistry result of NR2B in spinal cord dorsal horn and L4 DRG:The immunohistochemical positive indexs of NR2B in model group ipsilateral spinal cord doral horn 7d,15d and 23d were 3.8±1.48,5.6±1.19,5.64±1.04 respectively,the sham group were 1.60±0.55,1.75±0.79,1.8±0.84 respectively and the control group were 1.66±0.41,1.64±0.40,1.60±0.44 respectively;The immunohistochemical positive indexs of NR2B in model group ipsilateral DRG 7d,15d and 23d were 4.60±1.34,9.60±2.51,10.2±2.58 respectively,the sham group were 2.04±1.22,1.80±1.10,2.0±1.22 respectively and the control group 2.1±1.52,1.80±1.10,1.90±1.24 respectively,there were significant differences in the immunohistochemical positive indexs between model group and control group,also between model group and sham group (P＜0.05),but there were no differences between control group and sham group(P＞0.05).In the ipsilateral spinal cord doral horn and L4 DRG of model group There also were obvious differences either between model group 7d and model group 15d,or between model group 7d and model group 23d(P＜0.05),but no differences between model group 15d and model group 23d(P＞0.05).on post-inoculation Day 23,the immunohistochemical positive indexs of NR2B in model group ipsilateral spinal cord doral horn and L4 DRG were positively and significantly correlated with Spontaneous lifting duration of ipsilateral hindpaw respectively(r=0.976,P＜0.001;r=0.882,P＜0.001),the immunohistochemical positive indexs of NR2B in model group ipsilateral spinal cord doral horn and L4 DRG were negatively and significantly correlated with the mechanical withdrawal threshold of ipsilateral hindpaw respectively(r =-0.879,P＜0.001;r=-0.760,P=0.001).Conclusion In the dorsal horn of murine spinal cord and L4 DRG, the immunohistochemical positive indexs of NR2B were increased and significantly correlation with the manifestation of ethology,so the NR2B in the spinal cord and DRG may participate and mediate in transmitting algesthesia message,forming and persisting the hyperalgesia in bone cancer pain. |
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