ABSTRACT
Objective To investigate the effect of evodiamine(EVO)on retinal injury in diabetic rats by regulating the cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA)signaling pathway.Methods Totally 96 Sprague-Dawley rats(192 eyes)were divided into the negative control(NC)group,Model group,low-dose EVO(EVO-L)group,medium-dose EVO(EVO-M)group,high-dose EVO(EVO-H)group,calcium dobesilate(CD)group,SQ22536 group and EVO-H+SQ22536 group,with 12 rats in each group.Rats in the NC group were intraperitoneally injected with physiological saline instead of streptozotocin,and diabetic retinopathy models were established in all other groups.After successful mod-eling,the drug was administered once a day for 4 weeks.The blood glucose level of rats in each group was measured by blood glucose meter;HE staining was applied to detect the pathological changes of the retina of rats;TUNEL staining was adopted to detect the apoptosis of ganglion cells in the retina of rats;the levels of superoxide dismutase(SOD),malondial-dehyde(MDA),tumor necrosis factor-α(TNF-α),interleukin(IL)-6 and cAMP in the retina of rats were detected;West-em blot was used to detect the protein expressions of Bcl-2 associated X protein(Bax),P53 and phosphorylated protein ki-nase A(p-PKA)in the retina of rats.Results Compared with the NC group,the pathological injury of the retina in the Model group was more serious;the blood glucose,apoptosis rate of retinal ganglion cells,MDA,TNF-α,IL-6,Bax and P53 protein expressions increased,and the SOD,cAMP and p-PKA/PKA protein expression decreased,with statistically significant differences(all P＜0.05).Compared with the Model group,the pathological injury of the retina was relieved,the blood glucose,apoptosis rate of retinal ganglion cells,MDA,TNF-α,IL-6,Bax and P53 protein expressions decreased,and the SOD,cAMP and p-PKA/PKA protein expression increased in the EVO-L group,EVO-M group,EVO-H group and CD group,with statistically significant differences(all P＜0.05).Compared with the Model group,the retinal tissue of the SQ22536 group was severely damaged,the blood glucose,apoptosis rate of retinal ganglion cells,MDA,TNF-α,IL-6,Bax and P53 protein expressions increased,and the SOD,cAMP,p-PKA/PKA protein expression decreased,with statistically significant differences(all P＜0.05).Compared with the EVO-H group,the EVO-H+SQ22536 group showed more serious pathological injury of retinal tissue,increased blood glucose,apoptosis rate of retinal ganglion cells,MDA,TNF-α,IL-6,Bax and P53 protein expressions,and decreased SOD,cAMP and p-PKA/PKA protein expression,and the differences were statistically significant(all P＜0.05).Conclusion EVO may alleviate retinal injury in diabetic rats by activating the cAMP/PKA signaling pathway.
ABSTRACT
Objective To explore the roles of spinal microglial Src-family kinases (SFKs) in diabetic neuropathic pain. Methods Male Sprague-Dawley rats, weighing 200 ~ 220 g, were used in the experiments. The level of p-SFKs in spinal dorsal horn was detected by single immunofluorescence staining on day 7th , 14th and 28th after intraperitoneally injection of Streptozotocin (STZ) and its location was detected by double immunofluorescence staining. The changes of 50% paw-withdrawal thresholds of rat were detected by behavioral tests when PP2 , a specific inhibitor of SFKs , was intrathecally administered before intraperitoneally injection of STZ. Results Compared with vehicle group, the blood glucose level increased on day 1 (P < 0.001) and the hyperglycemia persisted at least for 28 days (P < 0.001) after intraperitoneally administered of STZ (50 mg/kg). Paw-withdrawal threshold (PWT) decreased gradually from day 1 (P < 0.05) and reached the minimum on day 28 (P < 0.001) after STZ administration. Meanwhile, the expression of p-SFKs in spinal dorsal horn markedly increased on day 7 (P < 0.01), day 14 (P < 0.01) and day 28 (P < 0.01). The p-SFKs was mainly co-localized only with microglia , but not with neurons or with astrocytes. Intrathecally administered of PP2 before STZ reversed STZ-induced mechanical hyperalgesia. Conclusion Microglial SFKs in spinal dorsal horn maybe play a pivotal role in diabetic neuropathic pain.
ABSTRACT
AIM:To investigate the effects of P2X4 receptor on peri-sciatic administration of recombinant rat TNF-α(rrTNF)-induced mechanical allodynia.METHODS:Male Sprague-Dawley rats (180~200 g) were used in the experiments.The levels of P2X4 receptor on day 3, day 7 and day 14 after peri-sciatic administration of rrTNF were exam-ined by Western blot, and the location of P2X4 receptor in the spinal dorsal horn was observed by double immunofluores-cence staining.The changes of 50%paw-withdrawal thresholds of the rat were detected by behavioral test, and the level of TNF-αin the spinal dorsal horn was also examined by Western blot when TNP-ATP was intrathecally injected before the ad-ministration of rrTNF.RESULTS:Compared with control group, the expression of P2X4 receptor in the spinal dorsal horn on the ipsilateral side significantly increased on day 3, day 7 and day 14 (P<0.01) after rrTNF (100 ng/L) administra-tion.P2X4 receptor was co-localized only with microglia, but not with neurons or astrocytes.Intrathecal injection of TNP-ATP before rrTNF administration prevented mechanical allodynia induced by rrTNF and inhibited the upregulation of TNF-αin the spinal dorsal horn.CONCLUSION:P2X4 receptors in microglia may be involved in rrTNF-induced mechanical allodynia by the upregulation of TNF-αin the spinal dorsal horn.