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Objective:To observe any effect of electroacupuncture (EA) on the expression of phosphorylated extracellular signal-regulated protein kinase (p-ERK1/2) and phosphorylated cyclic adenosine monophosphate response element binding protein (p-CREB) in the spinal dorsal horns of diabetics experiencing neuropathic pain.Methods:Eight rats were randomly selected from 30 healthy male Sprague-Dawley rats as the normal group (N), and the remaining twenty-two rats were treated with a single high-dose intraperitoneal injection of streptozotocin (STZ) to establish a neuropathic pain model. The rats modeled successfully were randomly divided into a model group (M, n=8) and an EA group ( n=8). In the EA group, electroacupuncture was applied at the bilateral Hou san li and Kunlun acupoints starting on the 15th day after the STZ injection. The daily sessions lasted 30 minutes for 1 week. Body weight (BW), fasting blood glucose (FBG) and paw withdrawal latency (PWL) were observed before the STZ injection and on the 7th, 14th, and 21st days afterward. The expression of p-ERK1/2 and p-CREB in the dorsal horns of the rats′ spinal cords was detected using western blotting. The count of p-CREB-positive cells in the dorsal horns and their co-localization with neurons was detected using immunofluorescence. Results:In comparison with the N group, the average BW of the M group on the 7th, 14th and 21st days after the STZ injection was significantly lower, while the average FBG was significantly higher. There was no significant difference between the M and N groups in the average PWL on the 7th day after the STZ injection, but it had decreased significantly in the M group on the 14th and 21st days. Compared with the M group, the average PWL of the EA group was significantly longer on the 21st day after the injection. The expression of p-ERK1/2 and p-CREB protein in the spines of the M group was significantly higher than in the N group. p-CREB positive cells were more numerous in the M group compared with the N group, while in the EA group they were fewer. P-CREB was co-located with neurons in the spinal dorsal horn.Conclusion:EA can alleviate neuropathic pain effectively, perhaps by inhibiting the expression of p-ERK1/2 and p-CREB in the dorsal horns of the spinal cord.
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Objective This study aims to evaluate the changes of Cdk5 expression at the time of 3 hours to 10 days after moderate brain injury by blunt force impact in a rat model,and to demonstrate its forensic significance.Methods To establish a rat model of blunt focal brain contusion,and to observe the changes of Cdk5 expression in brain tissue at different timepoints after brain injury by immunohistochemistry and Western blot.Results A low expression level of Cdk5 was observed in the brain tissue of both normal and sham control groups.The expression of Cdk5 increased after 3 and 6 hours,remarkably increased at 12 hours,and reached the maximal level at 24 hours after focal brain injury.The Cdk5 level gradually decreased 3 days,5 days,7 days,and 10 days and reached the normal level 7 and 10 days after the injury,with no statistical difference (P＞0.05) compared with the normal and sham control groups.Conclusion The expression of Cdk5 increased in the peripheral area of contusion tissue after blunt brain injury in rats,showing single peak change,and dropped to normal level with the time extension.The change of Cdk5 expression may provide a new reference index for the prediction of early brain contusion.
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Objective To evaluate the effect of endoplasmic reticulum stress in trophocytes,in patients with intrahepatic cholestasis of pregnancy (ICP).Methods Sixty-one pregnant women who were hospitalized in Women's Hospital,School of Medicine,Zhejiang University from January to December 2015 were recruited.Thirty-one women who were diagnosed as ICP were defined as the ICP group and 30 healthy pregnant women were defined as the control group.The localization and expression intensity of glucose regulated protein 78 (GRP-78) in placental tissues were detected by immunohistochemistry technique.Electronic microscope was used to observe ultra-microstructure change of the endoplasmic reticulum in trophocytes and cell line Swan71.Reverse transcription (RT)-PCR and western blot were used to investigate the expression of GRP-78 mRNA and protein in Swan 71 cell.Results (1) GRP-78 protein was mainly expressed in the cytoplasm of cytotrophoblasts and syncytiotrophoblasts.The protein expression of GRP-78 in placentas of the ICP group (13.2±2.4) was significantly higher than that in the control group (7.8±1.3,P＜0.01).(2) The volume of endoplasmie reticulum did not increase and the microvilli developed well,with no swelling and no expansion of endoplasmic reticulum in the control group.In the ICP group,microvilli injury,endoplasmic reticulum edema were found;the volume of endoplasmic reticulum increased,with dilation,vacuolation and significant degranulation.After treated with 100 μmol/L cholyglycine for 24 hours,universal dilatation of the endoplasmic reticulum were seen in the Swan71 cells.(3) In Swan71 cells,cholylglycine displayed a concentration-dependent up-regulation on the expression of GRP-78.The expressions of GRP-78 mRNA in 0,25,50,100 μmol/L cholylglycine experimental group were 1.01±0.17,2.17±0.16,5.47±0.36,5.65 ± 0.82,respectively.The expression of GRP-78 protein in 0,25,50,100 μmol/L cholylglycine experimental group were 1.01±0.04,1.17±0.15,1.33±0.13,1.73±0.13,respectively.The expression of GRP-78 mRNA and protein in 100 and 50 μ mol/L cholylglycine experimental group were significantly higher than 0 μmol/L (all P＜0.01).Conclusion The obvious expansion of endoplasmic reticulum and the increased expression of GRP-78 in trophocytes indicated that endoplasmic reticulum stress of trophocytes may be involved in the pathogenesis of ICP.
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Objective To study the expression of p35 and p25 in rat after focal cerebral contusion and to provide experimental data for estimating brain injury time. Methods Fifty adult male SD rats were randomly divided into 0 h, 6 h, 12 h, 24 h, 3 d, 5 d, 7 d, 10 d after focal cerebral contusion, control and sham-operated groups (5 rats each group). The focal cerebral contusion rat model was established. The expression of p35 and p25 protein of the damage peripheral zone in brain were detected by HEstain-ing, immunohistochemistry and western blotting at different injury time. Results Alarge number of p35 protein and a small amount of p25 protein were expressed in control group and sham-operated group. After focal cerebral contusion, p35 presented unimodal change with time and p25 presented bimodal changes with time. Conclusion Expression of p35 and p25 showed different regularity with good time correlation, which could help to estimate the brain injury time.
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Objective To explore the fluorescent distribution, and the relation between fluorescent intensity and time after injecting rear-thigh muscles with DiI. Methods Sixty-five new born mice were equally and randomly divided into 13 groups. Fluorescent distribution and intensity were investigated at 2, 4, 6, 8, 12, 24 h, and 2, 4, 7, 14, 28 d and 2, 3 months after injecting the left posterior limb rear thigh muscles with 1.5 ?l 4 mg/ml DiI for one mouse. Results The labeling neurons were scattered from L2 level to S2 level and associated dorsal root ganglion (DRG), but the most were located at L4 to L6 section. The faint red fluorescence neurons were observed at dorsal root ganglion and cornu anterius medullae spinalis 6 h post-injection. The labeling neurons increased up to the 4th day. The fluorescent intensity enhanced gradually from 6 h to 24 h, then kept the intensity for 3 months. Conclusion It is a quick, precise, persistent method to trance and label the dorsal root ganglions sensory neuron and spinal motoneuron by injecting rear-thigh muscles with DiI, and the rate of labeling neurons can be improved by prolonging the tracing time properly. It is also provide basic data for clinical or experimental neuron label and location.