| Diabetic peripheral neuropathy(DPN)is the most common complication of diabetes,its pathogenesis is not clear,and there is no effective mitigation measures.Therefore,it is of great significance to explore the accurate pathogenesis of DPN in order to relieve patients’pain and relieve social burden.At present,it is well known that the main cause of DPN is the direct action of hyperglycemia on dorsal root ganglion(DRG)neurons and the early degeneration of neurons.Excess glucose stimulation can motivate multiple signaling pathways,such as PERK pathway and it mediated apoptosis,oxidative stress and endoplasmic reticulum stress,which can further stimulate the production of a series of cytokines and chemokines.Cytokines and chemokines also increase the oxidative stress and nitriding stress of cells and promote the apoptosis of neurons.The protein kinase Piml discussed in this study belongs to Serine/threonine kinase,which is continuously activated in cells and does not require the phosphorylation from upstream kinase.Therefore,the activity of Piml kinase is directly related to its expression level and regulated by the level of transcription and translation.Although other fields of study have reported the role of Piml,we investigated the new role of Piml in the oxidative stress induced by DPN,and its induced apoptosis and diabetic neuropathic pain,and explored the new correlation between Piml and PERX-ATF4-CHOP signaling pathwaysIn this study,we used streptozotocin(STZ)to induce a diabetic mouse model to measure the change in protein expression of Piml,and found that Pim1 kinase was significantly increased in DRG neurons of diabetic neuropathic pain mice.To verify whether Piml is an important molecule for the survival of DRG neurons in vitro,we first used immunofluorescence to detect the expression and localization of Piml in DRG neurons.The results showed that Piml was not only expressed in DRG neurons,but also co-located with DRG neuron stubtypes CGRP(+)and NF200(+)neurons.Subsequently,we found that Piml kinase expression was elevated after exposure to high glucose 12 hours,so we hypothesized that Piml up-regulation was a protective response of neurons to hyperglycemia injurySubsequently,we found that Piml expression was elevated at protein level after hyperglycemia treatment,especially after 48 hours of treatment,so we hypothesized that Piml up-regulation was a protective response of neurons to hyperglycemia injury Based on the above studies,we decided to investigate the effects of Piml on DRG neurons after inhibiting their activity with Piml inhibitor TCS in the following two aspects:cell survival:decreased survival rate of neurons detected by CCK8 and significantly enhanced apoptosis observed by TUNEL staining.The expression levels of pBad and anti-apoptotic proteins bcl-2 and bcl-xl were significantly down-regulated.These results suggest that inhibition of Piml effect may make neurons vulnerable to external noxious stimulusIn order to verify whether protein kinase Piml can inhibit oxidative stress.endoplasmic reticulum stress and cell apoptosis induced by high glucose,we gave DRG neuron Piml inhibitor TCS to inhibit Piml kinase activity to investigate the effect of high glucose on oxidative stress in neurons,while the results showed that high glucose significantly increased ROS level in neurons.We detected that the protein expressions of signaling pathway molecules PERK,CHOP and ATF4 were up-regulated slightly after high glucose treatment,but were significantly up-regulated after TCS treatment,suggesting that Piml inhibition may aggravate ER stress through this pathway.Through the above results,we found that,under the condition of high glucose,DRG neurons within ROS levels and increased apoptosis is not clear,may be associated with Piml kinase levels increase,but in the use of TCS-Piml inhibition activity,cell apoptosis and ROS increased significantly,showed that after inhibition of the Piml kinase activity sensitizing the DRG neurons in response to the high glucose damage. |
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