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Abstract Diabetic Neuropathy (DN) is one of the prevailing micro vascular complications of diabetes which can be characterized by neuropathic pain. Streptozotocin (STZ) induced diabetes in the rat has been increasingly used as a model of painful diabetic neuropathy. STZ injection leads to neurotoxicity of peripheral nerves that leads to development of Peripheral Diabetic Neuropathy in rat model. The present study was aimed at exploring the protective role of Tinospora cordifolia extract in STZ induced neurotoxicity and evaluating mechanisms responsible for attenuating neuropathic pain. Neuropathic pain markers like hyperalgesia, allodynia and motor deficits were assessed before STZ injection and after the treatment with 250 mg/kg and 500 mg/kg dose of Tinospora cordifolia. Oxidative stress markers, NGF expression in sciatic nerve were observed after seven weeks treatment. Our results demonstrated that seven weeks treatment with Tinospora cordifolia leaf extract significantly relieved thermal hyperalgesia and allodynia by increasing the antioxidant enzyme levels, decreasing the lipid peroxidation and by increasing the Nerve growth factor (NGF) expression in diabetic rat sciatic nerves. Our findings highlighted the beneficial effects of oral administration of Tinospora cordifolia extract in attenuating diabetic neuropathic pain, possibly through a strong antioxidant activity and by inducing NGF m RNA in sciatic nerves.
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Animals , Male , Rats , Plants, Medicinal/adverse effects , Plant Extracts/analysis , Menispermaceae/classification , Hyperalgesia/diet therapyABSTRACT
Objective To investigate the effects of peripheral blood mononuclear cells (PBMCs) on spinal motor neurons after end-to-end anastomosis of transected sciatic nerves in rats and further explore the possible mechanisms concerned.Methods A total of 30 SD rats were randomly divided into five groups:blank control group,sciatic nerve transaction group (model group),nerve anastomosis after sciatic nerve transaction (anastomosis group),nerve anastomosis with PBMCs treatment after sciatic nerve transaction (PBMCs group),and nerve anastomosis with solvent control treatment after sciatic nervetransaction (solvent control group),with six rats per group.Except for the rats in blank control group,transection of the left sciatic nerves conducted 0.5 cm under the piriformis muscle was performed in all animals,immediately followed by end-to-end anastomosis of sciatic nerves in anastomosis,PBMCs and solvent control groups.The rats in PBMCs group were given 0.2 ml PBMCs (1 × 107 PBMCs) at anastomosis,and the rats in solvent control group were injected with 0.2 ml PRMI-1640.After rats were on regular diet for 4 weeks,morphological examination was performed by HE staining and the counting of apoptotic cells was evaluated by TdT mediated dUTP nick end labeling (TUNEL) staining.The protein expression of brain derived neurotrophic factor (BDNF) and glial cell derived neurotrophic factor (GDNF) were detected by Western blot.Results In model group,HE staining showed that most of the spinal motor neurons became vacuoles with karyopyknosis and cytomorphosis.In anastomosis group and solvent control group,some spinal motor neurons had contraction,deformation and even plasmolysis,with vacuoles status of cytoplasm,karyopyknosis in some cells.In PBMCs group,some spinal motor neurons were normal with nucleus in the middle while some other spinal motor neurons were seen plasmolysis and vacuoles status of cytoplasm.Through TUNEL,the numbers of apoptotic cells were 0.44 ± 0.10 in blank control group,5.78 ± 1.11 in model group,5.22 ±0.51 in anastomosis group,2.56-± 0.42 in the PBMCs group,and 3.78 ± 0.19 in solvent control group,respectively.Compared with blank control group,the apoptosis of spinal motor neurons was significantly increased in model group (P ＜ 0.05).Compared with model group,the apoptosis of spinal motor neurons was significantly decreased in treatment groups except anastomosis group (P ＜ 0.05).Compared with solvent control group,the apoptosis of spinal motor neurons was significantly decreased in PBMCs group (P ＜ 0.05).The BDNF protein levels of the spinal cord tissue were 1.83 ± 0.72 in blank control group,1.35 ± 0.46 in model group,1.29 ± 0.44 in anastomosis group,1.87 ± 0.55 in PMBCs group,and 1.22 ± 0.50 in solvent control group,respectively.Compared with blank control group,the protein expression of BDNF was decreased in model group (P ＜ 0.05).Compared with model group,the protein levels of BDNF and GDNF were increased in PMBCs group (P ＜ 0.05).There was no significant difference between anastomosis group and solvent control group in regard of protein levels of BDNF and GDNF (P ＞ 0.05).Conclusions The transaction of sciatic nerves may induce injury and apoptosis of the spinal motor neurons.End-to-end anastomosis combined with PMBCs can effectively ameliorate apoptosis of the spinal motor neurons,the mechanism of which may be related to PMBCs that may enhance protein levels of BDNF and GDNF in spinal cord.
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Objective To observe the behavioral and the histopathology changes and expression of NR2B subunit in spinal dorsal horn in chronic constriction injury (CCI) rats after pulsed radiofrequency (PRF). Methods Forty-eight adult Sprague-Dawley rats were randomly divided into Sham-Sham (SS), Sham-PRF (SP), CCI-Sham (CS) and CCI-PRF (CP) groups. The right sciatic nerves (SNs) of the CS and CP groups were ligated to create the CCI model. For the SS and SP groups, the right SNs were separated without ligation. The CP and SP groups accepted PRF at the ligation site 15 days after modeling, while the electrode was placed in rats in the SS and CS groups without elec-tricity. The hindpaw withdrawal threshold (HWT) was measured before and 3, 7, 11, 15 days after modeling, and 1, 3, 7, 11, 15 days after treatment. The right SNs at ligation sites were assessed with optical microscopic score 15 days after treatment, and the NR2B expression in the L4-6 spinal dorsal horn were determined with Western blotting. Results HWT was significantly shorter in the CS and CP groups than in the SS and SP groups after modeling, and was more in the CP group than in the CS group. Under the optical microscope, the axonal diame-ter and myelin sheath thickness increased in the CP group compared to those in the CS group (P<0.01), the NR2B expression was less in the CP group than in the CS group after treatment (F=10.769, P<0.05). Conclusion PRF may reduce hyperalgesia and repair damaged SNs in CCI-induced neuropathic pain, which may associate with inhibition of the NR2B subunit expression in spinal dorsal horn.
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OBJECTIVE:To investigate the effects of Tongxinluo capsules on inflammatory cytokines of mice with diabetic pe-ripheral neuropathy. METHODS:40 KK/Upj-Ay mice were randomly divided into model group(pure water),and Tongxinluo low, medium and high dose groups [1,2 and 4 g (medicinial materials)/kg]. 10 C57BL/6 mice were selected into the control group (pure water). The drugs were given once a day for consecutive 12 weeks,ig. The pain perception threshold,motor nerve conduc-tion velocity(MNCV)and sensory nerve conduction velocity(SNCV)were detected for the mice 1 h after the last administration of drugs. The flow cytometry was used to detect the levels of tumor necrosis factor α(TNF-α),interleukin-1β(IL-1β),cyclooxy-genase 2(COX-2)and monocyte chemoattractant protein 1(MCP-1)in serum. Real-time fluorescent quantitative polymerase chain reaction(RT-PCR)and Western blot were respectively employed to detect the mRNA and protein expressions of TNF-α,IL- 1β, COX-2 and MCP-1 in the mice’s sciatic nerves. RESULTS:Compared with control group,pain perception threshold in model group was decreased;MNCV and SNCV were slowed;the levels of TNF-α,IL-1β,COX-2 and MCP-1 in serum were increased;the expressions of mRNA and protein of TNF-α,IL-1β,COX-2,MCP-1 were increased,with significant differences(P<0.01 or P<0.05). Compared with model group,pain perception threshold in Tongxinluo medium and high dose groups were increased;MNCV and SNCV were increased;the levels of TNF-α,IL-1β,COX-2 and MCP-1 in serum were decreased;the expressions of mRNA and protein of TNF-α,IL-1β,COX-2,MCP-1 in sciatic nerves were decreased;the expressions of mRNA of COX-2, MCP-1 and protein of IL-1β,COX-2,MCP-1 in sciatic nerves in Tongxinluo low dose group were decreased,there were statistical significant difference(P<0.01 or P<0.05). CONCLUSIONS:Tongxinluo capsules have certain protective effects on the mice with peripheral nerve in case of diabetes by a mechanism that may be associated with the inhibition of inflammatory cytokines.
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Objective To study the characteristics of the sciatic nerve injuries inflicted by highvelocity triangle fragments in swine.Methods According to the injury position,14 swines weighing (34.29±5.2)kg were divided into two groups.All swines were shot by 0.37 g triangle fragments at (773.1±12.4)m/s aimed at the midpoint of the lateral body surface projection of the right sciatic nerve in Group A(n=7)and at 2 cm aside from sciatic nerve in Group B(n=7).The entrance,exit and length of wound tract,distance between nerve and wound tract were detected,and the pathology of sciatic nerve and wound tract at 48 hours after injury were observed under light microscope and electron microscope.Results The course of the fragments was deviated to different extent.Six sciatic nerves were in primary wound tract,four of which were lacerated and ruptured at different degrees.The light microscope showed pathological changes including severe hemorrhage and edema,leukocytic infiltration,neurotmesis,axonotmesis and light myelin coloration;while electron microscope showed severe degeneration of myelin sheath and neuraxon distortion.Six nerves(one in Group A and five in Group B)were in the concussion zone,with the distance between sciatic nerve and primary wound tract for(2.07±0.45)cm.Hyporrhea and hyperemia under perilemma and perineurium,partial nerve fiber disrupt and axonotmesis were observed by light microscope and partly myelin sheath delamination by electron microscope.Two nerves (one in Group A and another in Group B)were in the contusion zone,with moderate pathological changes.Conclusions The track of high velocity triangle fragments is unstable in the tissues.High disrupt rate,severe and wide extent lesion,early Wallerian degeneration are the main characteristics of direct injuries of swine's sciatic nerves.Indirect injuries can induce pathologic changes too,for which the lesion severity is related to the distance between nerves and primary wound tract.
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Objective To observe the histomorphological changes of sciatic nerves following pasteurizing warm temperature treatment. Methods The 12 mm left sciatic nerve from healthy adult male Wistar rats was pasteurized at 60℃ for 30 minutes and observed at the first week (1-week group) and the sixth week (6-week group), 15 rats per group. Number, diameter and area ratio of the myelinated axons in the control (proximal to the treated part), treatment and peripheral nerves (distal to the treated part) were calculated. The histomorphological changes were examined with electrophysiological and electron microscopies. Results (1) In the peripheral part, the number of the myelinated axons was significantly decreased in the 1-week group compared with the control group (P
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Objective To investigate the effect of nerve growth factor (NGF) and ciliary neurotrofic factor (CNTF) on the regeneration and functional recovery of sciatic nerves following sciatic nerve injuries (SNI). Methods The left sciatic nerves for 6 mm in 120 Wistar ratswere removed and divided randomly into 4 groups. Target muscular injection of NGF at concentration of 100 ng/kg and CNTF at 100 ng/kg was given to Group NGF+CNTF; CNTF at 100 ng/kg to Group CNTF; NGF at 100 ng/kg to Group NGF and equal quantity of normal saline to Group NS. Sciatic nerve functional index (SFI) measurement, morphometric analysis, S-100? and NF200 immunohistochemistry were performed for all the animals after operation. Results SFI, diameter and number of myelinated axons, and axonal quantity of S-100? and NF200 were significantly higher in Group NGF+CNTF than that in other groups. Conclusions Combined application of NGF and CNTF can enhance regeneration and functional recovery of sciatic nerves posterior to SNI.