ABSTRACT
OBJECTIVE： To study the mechanism of analgesic effect of 8-O-acetyl-safalinoside （8-OaS） on chronic inflammatory pain model rats. METHODS ：Totally 30 male SD rats were divided into sham operation group （normal saline ）， model group （normal saline ），8-OaS low-dose ，medium-dose and high-dose groups （3，10，30 μg/kg），with 6 rats in each group. Except for sham operation group ，other groups were given planter injection of Freund ’s complete adjuvant to induce chronic inflammatory pain model. After successful modeling ，the rats in each group were given corresponding drugs intrathecally ，once a day，for 7 consecutive days. Then Von-Frey filaments were used to detect the planter pain threshold of the rats in each group ；the area under the planter pain threshold curve of each group and the half effective dose （ED50）of 8-OaS were calculated. Another 36 male SD rats were divided into sham operation group （normal saline ），model group （normal saline ）and 8-OaS group （dose of ED50），and the modeling method and administration route were the same as above. Immunofluorescence histochemical staining was used to observe the positive expression of ionized calcium binding adapter molecule 1（Iba-1）and signal molecule phosphorylated p38 mitogen-activated protein kinase （p-p38 MAPK）；Western blotting assay was used to determine the expression of Iba- 1，p-p38 MAPK，IL-1β，IL-6 and TNF-α in spinal dorsal horn of rats. RESULTS：Compared with sham operation group ，plantar pain threshold and area under the curve in model group were reduced significantly （P＜0.01）. Compared with model group ，plantar pain threshold increased significantly after 5，6，7 days of administration in 8-OaS low-dose group （P＜0.05），plantar pain threshold and area under the curve in 8-OaS medium-dose and high-dose groups were increased significantly （P＜0.05 or P＜0.01）. Most of above indexes in each dose group of 8-OaS were signifficantly different ，and ED 50 of 8-OaS was 18.87 μ g/kg. Results of immunohistochemistry staining and Western blotting showed that p-p 38 MAPK was mainly expressed in Iba- 1 positive cells. Compared with sham operation group ，the fluorescence density of Iba- 1 and p-p 38 MAPK in spinal dorsal horn ，the expression of Iba-1，p-p38 MAPK，IL-6，IL-1β and TNF-α were significantly increased in model group（P＜0.05 or P＜0.01）. Compared with model group ，the fluorescence density of Iba- 1 and p-p 38 MAPK in spinal dorsal horn ，the expression of Iba- 1，p-p38 MAPK， IL-6，IL-1β and TNF-α were decreased significantly in 8-OaS group （P＜0.05）. CONCLUSIONS ：Intrathecal administration of 8-OaS can effectively alleviate chronic inflammatory pain in rats. The mechanism may be related to the inhibition of the phosphorylation of p 38 MAPK and the expression of IL- 6，IL-1β and TNF-α.
ABSTRACT
OBJECTIVE： To study the effects of 8-O-acetyl-shanzhiside methylester （8-OaS） on the expression of histone deacetylase 1-5 （HDAC1-5） in the spinal dorsal horn of chronic inflammatory pain model rats， and its relationship with Janus-activated kinase 2-signal transductions and activators of transcription 3 （JAK2-STAT3） signaling pathway. METHODS： SD rats were randomly divided into normal control group， sham operation group （normal saline）， complete Freund’s adjuvant （CFA） group （normal saline）， 8-OaS low-dose， medium-dose and high-dose groups （2， 20， 200 μg/kg）， with 6 rats in each group. Except for normal control group and sham operation group， chronic inflammatory pain model was induced by subcutaneous injection of CFA into the left hind toe of rats in other groups； after modeling， those groups were given relevant medicine intraperitoneally， once a day， for consecutive 7 d. Thermal radiation method was used to detect the latency of paw withdraw in rats on the 1st， 2nd， 3rd， 4th， 5th， 6th， 7th， 8th， 11th and 15th day of administration. Rats were grouped and given medicine according to the above method of the latter 5 groups. The protein expression of HDAC 1-5， phosphorylated JAK2 （pJAK2） and phosphorylated STAT3 （pSTAT3） in the spinal dorsal horn of lumbar enlargement segment in rats were detected by Western blot method after last medication. Rats were randomly divided into sham operation group （normal saline）， CFA group （normal saline）， 8-OaS group （20 μg/kg） and JAK2-STAT3 inhibtor AG490 group （8 mg/kg）， with 6 rats in each group； IP model was established by same method as above and then were given relevant medicine intraperitoneally， once a day， for consecutive 7 d. The expression of HDAC5 and glial fibrillary acidic protein （GFAP） in spinal dorsal horn of rats were detected by immunofluorescence histochemical staining. RESULTS： Compared with normal control group and sham operation group， the latency of paw withdraw was shortened significantly in other groups （P＜0.05）. Compared with CFA group， the latency of paw withdraw was prolonged significantly in 8-OaS low-dose， medium-dose and high-dose groups （P＜0.05）， in dose-dependent manner. Compared with sham operation group， the protein expression of HDAC 1-5， pJAK2 and pSTAT3 in spinal dorsal horn of rats were increased significantly in CFA group （P＜0.05）. Compared with CFA group， the protein expression of HDAC5， pJAK2 and pSTAT3 in spinal dorsal horn of rats were decreased significantly in 8-OaS low-dose， medium-dose and high-dose groups （P＜0.05）， but there was no statistical significance in the protein expression of HDAC 1-4 （P＞0.05）. HDAC5 was expressed on astrocytes in the spinal dorsal horn； compared with sham operation group， the expression of GFAP and HDAC 5 were increased significantly in spinal dorsal horn of rats in CFA group （P＜0.05）. Compared with CFA group， the expression of GFAP and HDAC5 in spinal dorsal horn of rats were decreased significantly in 8-OaS group and AG490 group （P＜0.05）. CONCLUSIONS： 8-OaS can effectively relieve CFA-induced chronic inflammatory pain， the mechanism of which may be associated with the down-regulation of HDAC5 expression and the phosphorylation levels of JAK2 and STAT3.