神经病理性痛大鼠背根神经节TRESK mRNA表达的变化

目的 探讨神经病理性痛大鼠背根神经节(DRG) 孔钾离子通道TRESK mRNA表达的变化.方法 雄性SD大鼠32只,体重22、0～250 g,采用随机数字表法,将大鼠随机分为2组(n=16):假手术组(S组)和神经病理性痛组(NP组).采用坐骨神经分支选择性损伤法制备神经病理性痛模型.S组仅暴露神经,不结扎.于术前1 d和术后1、3、5、7、14 d取8只大鼠,测定左后肢机械缩足反应阈值(MWT)和热缩足潜伏期(TWL).于术前1 d和术后14 d痛阈测定结束后取L4,5术侧DRG,采用RT-PCR法测定TRESK mRNA的表达.结果 与S组比较,NP组MWT明显降低,DRG TRESK mRNA表达明显下调(P<0.05或0.01),TWL差异无统计学意义(P>0.05).结论 神经病理性痛大鼠DRG TRESK mRNA表达下调,该变化可能与神经病理性痛的形成有关.

Abstract：

Objective To evaluate the changes in the expression of diplopore potassium ion channel TRESK mRNA in dorsal root ganlion (DRG) in rats with neuropathic pain (NP) .Methods Thirty-two male SD rats weighing 220-250 g were randomly divided into 2 groups ( n = 16 each) : group sham operation (group S) and group NP. NP was induced by ligation and severance of left tibial and common fibular nerves according to the technique described by Decosterd. Eight rats in each group were sacrificed 1 day before and 14 day after operation and their L4,5 DRGs in the operated side were isolated for determination of TRESK mRNA expression by RT-PCR. In the remaining 8 rats in each group paw withdrawal threshold to mechanical stimuli ( MWT) and paw withdrawal latency to a thermal nociceptive stimulus (TWL) were measured at 1 day before (baseline) and 1, 3, 5, 7, 14 day after operation. Results MWT was significantly lower in group NP than in group S. The TRESK mRNA expression in L4,5 DRGs in the operated side was significantly decreased after operation as compared with the baseline before operation in group NP and was significantly lower in group NP than in group S. Conclusion The development and maintenance of NP may be closely related with down-regulation of TRESK mRNA.     

鞘内注射DREAM-shRNA对神经病理性痛大鼠脊髓背角p-CREB表达的影响

Objective To investigate the effects of intrathecal (IT) DREAM-short hairpin RNA (DREAM-shRNA) on expression of phosphorylated cyclic AMP response element binding protein (p-CREB) in the spinal dorsal horn in a rat model of neuropathic pain. Methods Adult male SD rats weighing 280-320 g were anesthetized with intraperitoneal 10% chloral hydrate. Neuropathic pain was induced by chronic constrictive injury (CCI) to sciatic nerve. IT catheters were placed according to the method described by Yaksh on 3rd day after CCI. Twenty-four rots in which IT catheter was successfully implanted were randomly divided into 4 groups (n = 6 each) : group Ⅰ sham operation (group S) ; group Ⅱ neuropathic pain (group NP) ; group Ⅲ RNA interference (group RNAi) and group Ⅳ blank vector (group BV). Lentivius with DREAM-shRNA 5 μl was injected IT in group RNAi, and blank vector 5 μl in group BV, and once a day for 7 days, starting from the day 8 after CCI. The mechanical pain threshold was measured at day 1 before CCI (T0 ,baseline) and day 7-14 after CCI (T1-8). The animals were killed on 15th day after CCI. The L4-6 lumbar segment of the spinal cord was removed for determination of the expression of green fluorescent protein (GFP) and p-CREB by immuno-fluorescent method.Results The mechanical pain threshold was significantly decreased as compared with the baseline at T0 in all 4 groups and returned to the baseline levels at T5-8 in group S and RNAi, but remained low in group NP and BY. The mechanical pain threshold was significantly lower after CCI/sham operation and significanty higher at T8 in group RNAi than in the other 3 groups. The expression of p-CREB in the spinal dorsal horn was up-regulated in group NP, RNAi and BV as compared with group S, and in group NP and BV as compared with group RNAi. The green fluorescence was observed in group RNAi but not in the other 3 groups. Conclusion IT DREAM-shRNA can ameliorate neuropathic pain in rats through inhibiting the expression of p-CREB in the spinal dorsal horn.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

鞘内注射吗啡加可乐定对切口痛大鼠脊髓背角蛋白激酶A催化亚单位表达的影响

Objective To observe the effect of intrathecal clonidine plus morphine on expression of protein kinase A (PKA) catalytic subunit in the spinal dorsal horn in a rat model of incisional pain. Methods Eighty male Sprague-Dawley rats were divided randomly into five groups: sham group, control group, pre-incisional morphine 2.5 μg group, pro-incisional clonidine 5 μg group and preincisional morphine 2.5 μg plus clonidine 5 lag group (n=16). lntrathecal catheter and the model of incisional pain were pro-duced according to Yaksh and Brennan's described method respectively. Changes of pain behavior were assessed by mechanical with-drawal threshold (MWT) and thermal withdrawal latency(TWL). The expressions of PKA catalytic subunit in the spinal dorsal horn were assessed by immunohistochemical method and western blotting analysis. Results Compared with sham group, MWT and TWL in control group were decreased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were increased significantly in control group (P<0.01). Compared with control group, MWT and TWL in pre-incision morphine 2 μg plus clonidine 5 lag group were increased significantly at 2 h after incision (P<0.01) and the number of positive cells and protein expression of PKA catalytic subunit in the spinal dorsal horn were decreased significantly in pre-incision morphine 2 μg plus clonidine 5 μg group (P<0.01). However, MWT, TWL and the number of positive cells and pro-tein expression of PKA catalytic subunit in the spinal dorsal horn changed with no statistical significance in pre-incisional morphine 2.5 μg group and pre-incisional clonidine 5 μg group compared with control group. Conclusion lntrathecal clonidine significantly enhances the antinociceptive effect of intrathecal morphine in a rat model of incisional pain, which might be associated with inhibi-tion of the increased expression of PKA catalytic subunit in spinal cord.     

神经干细胞移植数量对大鼠神经病理性痛的影响

目的 探讨神经干细胞(NSGs)移植数量对大鼠神经病理性痛的影响.方法 取出生1～3 d的SD大鼠,制备NSCs悬液.清洁级雄性SD大鼠84只,体重150～180 g,采用右侧坐骨神经半切断法制备大鼠神经病理性痛模型.采用随机数字表法,将大鼠随机分为7组(n=12),假手术组(S组):仅暴露坐骨神经,不切断,鞘内注射细胞培养液30 μl;神经病理性痛组(NP组):于模型制备后3d鞘内注射细胞培养液30 μl;NP+NSCs 103组(N1组)、NP+NSCs 104组(N2组)、NP+NSCs 105组(N3组)、NP+NSCs 106组(N4组)和NP+NSCs 107组(N5组):于模型制备后3 d,鞘内注射相应浓度的NSCs悬液.模型制备前1 d和制备后1、3.7、14和21 d时测右足机械缩足阈值(MWT)和热缩足潜伏期(TWL);于模型制备后7和21 d,痛阈测定结束后,取右侧腰膨大脊髓背角及L5背根神经节,测定脑源性神经营养因子(BDNF)mRNA及其蛋白的表达水平.结果 与S组比较,NP组和N1～5组MWT降低,TWL缩短,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF阳性细胞数增多,染色加深;与NP组比较,N2～5组MWT升高,TWL延长,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF 阳性细胞数增多,染色加深;模型制备后7 d N1～5组MWT依次升高,TWL依次延长,脊髓背角和DRG BDNF mRNA表达依次上调(P<0.05),BDNF阳性细胞数依次增多,染色逐渐加深;模型制备后14、21d,N1～3组MWT依次升高,TWL依次延长(P<0.05),模型制备后21 d,N1～3组脊髓背角和DRG BDNF mRNA表达依次上调,BDNF阳性细胞数依次增多,染色逐渐加深;N3～5组上述指标比较差异无统计学意义(P>0.05).结论 NSCs移植减轻大鼠神经病理性痛的适宜移植数量为105个.     

神经干细胞移植数量对大鼠神经病理性痛的影响

目的 探讨神经干细胞(NSGs)移植数量对大鼠神经病理性痛的影响.方法 取出生1～3 d的SD大鼠,制备NSCs悬液.清洁级雄性SD大鼠84只,体重150～180 g,采用右侧坐骨神经半切断法制备大鼠神经病理性痛模型.采用随机数字表法,将大鼠随机分为7组(n=12),假手术组(S组):仅暴露坐骨神经,不切断,鞘内注射细胞培养液30 μl;神经病理性痛组(NP组):于模型制备后3d鞘内注射细胞培养液30 μl;NP+NSCs 103组(N1组)、NP+NSCs 104组(N2组)、NP+NSCs 105组(N3组)、NP+NSCs 106组(N4组)和NP+NSCs 107组(N5组):于模型制备后3 d,鞘内注射相应浓度的NSCs悬液.模型制备前1 d和制备后1、3.7、14和21 d时测右足机械缩足阈值(MWT)和热缩足潜伏期(TWL);于模型制备后7和21 d,痛阈测定结束后,取右侧腰膨大脊髓背角及L5背根神经节,测定脑源性神经营养因子(BDNF)mRNA及其蛋白的表达水平.结果 与S组比较,NP组和N1～5组MWT降低,TWL缩短,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF阳性细胞数增多,染色加深;与NP组比较,N2～5组MWT升高,TWL延长,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF 阳性细胞数增多,染色加深;模型制备后7 d N1～5组MWT依次升高,TWL依次延长,脊髓背角和DRG BDNF mRNA表达依次上调(P<0.05),BDNF阳性细胞数依次增多,染色逐渐加深;模型制备后14、21d,N1～3组MWT依次升高,TWL依次延长(P<0.05),模型制备后21 d,N1～3组脊髓背角和DRG BDNF mRNA表达依次上调,BDNF阳性细胞数依次增多,染色逐渐加深;N3～5组上述指标比较差异无统计学意义(P>0.05).结论 NSCs移植减轻大鼠神经病理性痛的适宜移植数量为105个.     

神经干细胞移植数量对大鼠神经病理性痛的影响

目的 探讨神经干细胞(NSGs)移植数量对大鼠神经病理性痛的影响.方法 取出生1～3 d的SD大鼠,制备NSCs悬液.清洁级雄性SD大鼠84只,体重150～180 g,采用右侧坐骨神经半切断法制备大鼠神经病理性痛模型.采用随机数字表法,将大鼠随机分为7组(n=12),假手术组(S组):仅暴露坐骨神经,不切断,鞘内注射细胞培养液30 μl;神经病理性痛组(NP组):于模型制备后3d鞘内注射细胞培养液30 μl;NP+NSCs 103组(N1组)、NP+NSCs 104组(N2组)、NP+NSCs 105组(N3组)、NP+NSCs 106组(N4组)和NP+NSCs 107组(N5组):于模型制备后3 d,鞘内注射相应浓度的NSCs悬液.模型制备前1 d和制备后1、3.7、14和21 d时测右足机械缩足阈值(MWT)和热缩足潜伏期(TWL);于模型制备后7和21 d,痛阈测定结束后,取右侧腰膨大脊髓背角及L5背根神经节,测定脑源性神经营养因子(BDNF)mRNA及其蛋白的表达水平.结果 与S组比较,NP组和N1～5组MWT降低,TWL缩短,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF阳性细胞数增多,染色加深;与NP组比较,N2～5组MWT升高,TWL延长,脊髓背角和DRG BDNF mRNA表达上调(P<0.05),BDNF 阳性细胞数增多,染色加深;模型制备后7 d N1～5组MWT依次升高,TWL依次延长,脊髓背角和DRG BDNF mRNA表达依次上调(P<0.05),BDNF阳性细胞数依次增多,染色逐渐加深;模型制备后14、21d,N1～3组MWT依次升高,TWL依次延长(P<0.05),模型制备后21 d,N1～3组脊髓背角和DRG BDNF mRNA表达依次上调,BDNF阳性细胞数依次增多,染色逐渐加深;N3～5组上述指标比较差异无统计学意义(P>0.05).结论 NSCs移植减轻大鼠神经病理性痛的适宜移植数量为105个.

Abstract：

Objective To investigate the effect of intrathecal (IT) transplantation of different quantities of neural stem cells (NSCs) on neuropathic pain (NP) in rats.Methods Eighty-four adult pathogen-free male SD rats weighing 150-180 g were randomly divided into 7 groups ( n = 12 each) : group sham operation (S group) , NP group, NP+ NSCs 103 , 104 , 105 , 106 , 107 groups (N1-5 groups) . NP was induced by partial transection of right sciatic nerve. NSCs were transplanted into subarachnoid space in N1-5 groups. Paw withdrawal threshold to mechanical stimulation (MWT) and paw withdrawal latency to nociceptive thermal stimuli (TWL) were measured at 1 day before (baseline) and 1, 3, 7, 14 and 21 days after operation. Brain derived neurotrophic factor ( BDNF) expression in spinal dorsal horn and dorsal root ganglia (DRG) was detected by immuno-histochemistry and RT-PCR on the 7th and 21st day after operation. Results Partial transection of the sciatic nerve gradually reduced MWT and TWL after operation starting from day 1 until day 7 and 14 as compared with the baseline in group NP. IT NSC transplantation significantly increased MWT and TWL and expression of BDNF in spinal dorsal horn and DRG in a dese-dependent manner at day 7 after operation in N1-5 groups as compared with group NP. There were no significant differences in MWT and TWL and BDNF expression among N3, N4 and N5 groups at day 21 after operation.Conclusion The proper quantity of transplanted NSCs which are able to ameliorate NP induced by partial transection of sciatic nerve in rats is 105 .     

神经病理性痛大鼠背根神经节TRESK mRNA表达的变化

目的 探讨神经病理性痛大鼠背根神经节(DRG) 孔钾离子通道TRESK mRNA表达的变化.方法 雄性SD大鼠32只,体重22、0～250 g,采用随机数字表法,将大鼠随机分为2组(n=16):假手术组(S组)和神经病理性痛组(NP组).采用坐骨神经分支选择性损伤法制备神经病理性痛模型.S组仅暴露神经,不结扎.于术前1 d和术后1、3、5、7、14 d取8只大鼠,测定左后肢机械缩足反应阈值(MWT)和热缩足潜伏期(TWL).于术前1 d和术后14 d痛阈测定结束后取L4,5术侧DRG,采用RT-PCR法测定TRESK mRNA的表达.结果 与S组比较,NP组MWT明显降低,DRG TRESK mRNA表达明显下调(P<0.05或0.01),TWL差异无统计学意义(P>0.05).结论 神经病理性痛大鼠DRG TRESK mRNA表达下调,该变化可能与神经病理性痛的形成有关.     

神经病理性痛大鼠背根神经节TRESK mRNA表达的变化

目的 探讨神经病理性痛大鼠背根神经节(DRG) 孔钾离子通道TRESK mRNA表达的变化.方法 雄性SD大鼠32只,体重22、0～250 g,采用随机数字表法,将大鼠随机分为2组(n=16):假手术组(S组)和神经病理性痛组(NP组).采用坐骨神经分支选择性损伤法制备神经病理性痛模型.S组仅暴露神经,不结扎.于术前1 d和术后1、3、5、7、14 d取8只大鼠,测定左后肢机械缩足反应阈值(MWT)和热缩足潜伏期(TWL).于术前1 d和术后14 d痛阈测定结束后取L4,5术侧DRG,采用RT-PCR法测定TRESK mRNA的表达.结果 与S组比较,NP组MWT明显降低,DRG TRESK mRNA表达明显下调(P<0.05或0.01),TWL差异无统计学意义(P>0.05).结论 神经病理性痛大鼠DRG TRESK mRNA表达下调,该变化可能与神经病理性痛的形成有关.     

鞘内注射GDNF对神经病理性痛大鼠脊髓背角p38MAPK蛋白表达的影响

目的 评价鞘内注射胶质细胞源性神经营养因子(GDNF)对神经病理性痛大鼠脊髓背角p38丝裂原活化蛋白激酶(p38MAPK)蛋白表达的影响.方法 取鞘内置管成功的健康雄性SD大鼠120只,周龄6周,体重180～200 g,随机分为4组(n=30):对照组(C组)、假手术组(S组)、神经病理性痛组(P组)和GDNF组.采用结扎L5.6脊神经的方法建立大鼠神经病理性痛模型.C组不给予任何处理;S组只暴露脊神经,但不结扎;P组脊神经结扎后鞘内注射生理盐水10μl,隔日1次,连续14 d;GDNF组脊神经结扎后鞘内注射GDNF 2μg,用生理盐水稀释至10μl,隔日1次,连续14 d.分别于脊神经结扎后3、7和14 d时,取10只大鼠,测定机械痛阈,然后处死,取脊髓背角,分别采用免疫组化法和蛋白质印迹法测定p38MAPK蛋白的表达水平.结果 与S组比较,P组和GDNF组机械痛阈降低,脊髓背角p38MAPK蛋白表达上调(P＜0.05或0.01);与P组比较,GDNF组机械痛阈升高,脊髓背角p38MAPK蛋白表达下调(P＜0.05或0.01).结论 鞘内注射GDNF可通过抑制脊髓背角p38MAPK蛋白的表达减轻大鼠神经病理性痛.     

胫骨癌痛大鼠脊髓背角5-羟色胺水平的变化

目的 评价胫骨癌痛大鼠脊髓背角5-羟色胺(5-HT)水平的变化.方法 雌性SD大鼠60只,体重160～180 g,采用随机数字表法,将其随机分为3组(n=20):对照组(C组)、假手术组(S组)和胫骨癌痛组(P组).C组不做任何处理;S组于右侧胫骨上段骨髓腔注射D-hank液10 μl;P组于右侧胫骨上段骨髓腔接种Walker 256大鼠乳腺癌细胞悬液10μl制备胫骨癌痛模型.于接种前1d、接种后3、5、7、9、11、14、16、18和21 d时测定机械痛阈.于接种前1 d、接种后7、14和21 d时,每组痛阈测定结束后随机处死大鼠4只,取脊髓组织,采用高效液相色谱法测定脊髓背角5-HT含量;接种后14 d取接种侧胫骨组织,光镜下观察胫骨破坏情况.脊髓背角5-HT含量与机械痛阈进行直线相关分析.结果 与C组和S组比较,P组接种后7～21 d时机械痛阈降低,接种后7、14和21 d时脊髓背角5-HT含量升高(P＜0.05),且5-HT含量与机械痛阈呈负相关(r=-0.973,P＜0.05).C组和S组各时点机械痛阈和脊髓背角5-HT含量比较差异无统计学意义(P＞0.05).光镜下P组大鼠术侧胫骨骨质严重破坏.结论 大鼠胫骨癌痛的形成与维持可能与脊髓背角5-HT水平升高有关.

Abstract：

Objective To investigate the change in 5-hydroxytryptomine (5-HT) content in spinal dorsal horn in a rat model of tibial bone cancer pain (BCP). Methods Sixty female SD rats weighing 160-180 g were randomly divided into 3 groups ( n = 20 each): control group (group C), sham operation group (group S) and BCP group. BCP was induced by intra-tibial inoculation of 10 μl Walker 256 breast cancer cell suspension in group BCP, while group S received intra-tibial inoculation of 10 μl D-hank solution. Paw withdrawal threshold to mechanical stimulation with yon Frey filaments (MWT) was measured 1 d before (baseline) and at 3, 5, 7, 9, 11,14, 16, 18 and 21 d after breast cancer cell inoculation. At 1 d before and 7, 14 and 21 d after breast cancer cell inoculation, four animals in each group were sacrificed after measurement of MWT. Their lumber segments of the spinal cord were removed for assay of 5-HT content in spinal dorsal horn using HPLC with fluorescence detector.HE staining was used to detect the damage to the tibia. Correlation between the 5-HT content and MWT was analyzed. Results MWT was significantly decreased after breast cancer cell inoculation in group BCP ( P ＜ 0.05).Microscopic examination showed serious bone destruction of tibia at the injection site in group BCP, while no bone destruction was found in groups C and S. 5-HT content in spinal dorsal horn was significantly higher in group BCP than in groups C and S (P ＜ 0.05). There was strong negative linear correlation between 5-HT content in spinal dorsal horn and MWT ( r = - 0.973, P ＜ 0.05 ). Conclusion The 5- HT content in spinal dorsal horn is significantly increased in rats with tibial BCP and is involved in the development of BCP.     

NGF和BDNF在炎性痛大鼠背根神经节和脊髓背角的表达

目的 观察神经生长因子(NGF)和脑源性神经营养因子(BDNF)在福尔马林致痛大鼠外周、背根神经节(DRG)和脊髓背角的表达变化及相互关系，探讨神经营养因子在疼痛发生机制中的作用。方法 采用大鼠后足掌皮下注射福尔马林溶液的外周炎性痛模型。成年SD大鼠24只随机分为四组(每组6只)：对照组(A组)；致痛组根据取材时间不同分为，致痛后2h取材(B组)、致痛后24h取材(C组)、致痛后48h取材(D组)。观察致痛1h内疼痛行为变化，应用免疫组织化学方法结合计算机图像分析技术检测致炎足底皮肤、DRG和脊髓背角浅层内NGF的表达，及DRG和脊髓背角浅层表达BDNF的变化。结果 所有致痛组大鼠都表现出明显的两期伤害性反应。在致痛后2h患侧足底皮肤、DRG和24h时脊髓背角浅层表达的NGF开始增加，致痛24h后BDNF在患侧DRG表达开始增加，且与NGF在患侧DRG神经元的表达变化成正相关。48h时BDNF在患侧脊髓背角浅层表达开始增加。结论 NGF不仅是产生炎性痛外周机制的重要介质之一，还可通过促进BDNF在脊髓背角的表达，共同参与中枢敏感化的产生。     

脊髓背角卡配因在大鼠足底炎性痛形成中的作用

目的 探讨脊髓背角卡配因在大鼠足底炎性痛形成中的作用.方法 雄性SD大鼠48只,6周龄,体重160～200 g,采用随机数字表法,将其随机分为3组:正常对照组(C组,n=8)、PBS组(n=16)和酵母多糖诱发足底炎性痛组(Z组,n=24).Z组于大鼠左侧后足足底皮下注射酵母多糖1.25 mg,制备酵母多糖诱发足底炎性痛模型,PBS组给予等容量PBS 100μl.分别于给药前(T0)、给药后30 min(T1)、1 h(T2)、2 h(T3)、4 h(T4)、8 h(T5)、24 h(T6)和48 h(T7)时测定左侧后足机械刺激缩足阈值(MWT)、热缩足反应潜伏期(PWTL)和左侧后足足底最大厚度.PBS组于T4时处死8只大鼠,Z组分别于T4、T6和T7时各处死8只大鼠,取左侧脊髓L4～6节段,采用Western blot法测定脊髓背角spectrin αⅡ降解产物、IκBα、环氧化酶-2(COX-2)的表达和NF-κB活性.结果 与C组比较,Z组MWT降低,PWTL缩短,足底最大厚度增厚,脊髓背角spectrin αⅡ降解产物和COX-2的表达上调,IκBα表达下调,NF-κB活性升高(P＜0.05或0.01),PBS组上述指标差异无统计学意义(P＞0.05).结论 脊髓背角卡配因活化参与了大鼠足底炎性痛的形成,其机制与激活NF-κB,上调COX-2表达有关.