| Background:Diabetic peripheral neuropathy is the most common and debilitating complication of diabetes, more than half of all individuals with diabeties eventually develop neuropathy, which seriously affecting the patient’s quality of life and physical and psychological health. Chronic hyperglycemia triggers nerve damage through multiple pathways, among which oxidative stress and inflammation plays a central role. Glycemia not only causes oxidative stress and inflammatory cascades directly by increasing the number of reactive oxygen and reactive nitrogen in the mitochondria, but also induces a variety of pathways which regulates the activation of transcriptional factors and expression of inflammatory cytokines or enzymes, leading to the occurence and development of neuroinflammation. These damage neuron function and nerve blood directly or indirectly, which is the central mechanism of neurons sensitization and pain. Hydrogen is a nearly discovered novel antioxidant. Apart from its direct protection against oxidative stress and quenching of free radicals, hydrogen gas can also up-regulate antioxidant protein expression and down-regulate inflammatory protein expression. Our previous study found that inhaling molecular hydrogen can significantly reduce the levels of reactive oxidant species in serum and tissue, enhance the activity of antioxidant enzymes, and protect sepsis mice from multiple organ damage, indicating that hydrogen has dual effect of antioxidation and anti-inflammation. So we hypothesized that HRS may provide an effective method against DNP, its mechanism may be related to inhibit oxidative stress induced inflammation directly or indirectly.In this study, we investigated the effect of hydrogen-rich saline on STZ-induced diabetic rats and explored the underlying mechanisms, to indicate if molecular hydrogen plays a protective role in diabetic neuropathy by regulating the expression of inflammation related transcriptional factors.Part1:effect of different ways of hydrogen-rich water injection on diabetic neuropathic pain.Objective:to observe the effect of6different ways of hydrogen-rich water injection on painful diabetic neuropathy. Methods:Diabetes was induced by single dose of STZ (65mg/kg, i.p.). After diabetic models was established successively, hydrogen rich saline (5mg/kg, i.p.) was administered daily, separately from3rd-4th,3rd-6th,3rd-8th,5th-6th,5th-8th,7th-8th after diabetes induction. Nerve functional parameter, behavioral changes and proinflammatory cytokines (TNF-a and IL-6) levels were measured to investigate the effect of hydrogen saline and to select the optimum regimen as treatment group in the following study. Results:Both MNCV and behavioral parameters significantly decreased in diabetic rats from the second week to the eighth week in a time-dependent way, indicating the successful building of DNP model. Hydrogen-rich saline treatment significantly reversed the nerve conduction deficit and corrected the reduction of both thermal and mechanical threshold. In accordance with above results, the elevated proinflammatory cytokines in sciatic nerves of diabetic rats were significantly suppressed by HRS treatment. The group received treatment from7th-8th after diabetes induction was selected as the best one remaining as treatment group. Conclusion:this part demonstrated that DNP is closely related to inflammation, and intraperitoneal injection of hydrogen-rich water can improve the motor nerve conduction velocity, hypersensitivity symptoms and neuroinflammation.The group received treatment from7th-8th after diabetes induction was selected as the best one remaining as treatment group.Part2:Hydrogen-rich saline plays a protective effect on DNP by downregulating NF-kB pathway and upregulating Nrf2pathway.Objective:To observe the effect of HRS injection to NF-kB pathway and Nrf2pathway, and to explore its possible mechanism. Method:Six SD rats with the same month were randomly selected as normal control group. Another12rats were selected from successful diabetic models and randomly divided into STZ-D group and D+H2group (n=6). D+H2group received a HRS treatment from7th-8th after diabetes induction, which isselected as the best protocol in part1. Sciatic nerves were collected after the last behavirol test. Immunohistochemistry was administrated to determing the translocation of NF-κB p65. Western blot was used to using Western blot method NF-kB pathway and Nrf2/HO-1protein expression. Results: Immunohistochemistry and Western blot results supported that,8weeks after STZ injection, NF-kB (P65subunit) positive cells as well as the expression of NF-kB、 IκB、pIκB in sciatic nerves was significantly increased (P<0.01) when compared with control group. HRS injection significantly reversed the elevated NF-kB、IkB and pIκB expression. Moreover, compared with control group, Nrf2and HO-1expression in sciatic nerves of diabetic rats exhibits a significant decreas, while HRS treatment can improve their expression (P<0.01). Conclusion:H2can inhibit the activation of NF-kB pathway and its downstream inflammatory cascade in diabetic rats, and faciliate Nrf2activation, thus strengthening the body’s own antioxidant defense system, restoring redox balance damaged by hyperglycemia.Above all, these results suggest that hydrogen rich saline has the neuroprotective potential of ameliorating various deficits associated with diabetic peripheral neuropathy. Its anti-inflammatory activity arises from the inhibition of NF-kB pathway and downstream mediators of inflammation, while its anti-oxidative capacity from up-regulation of Nrf2. |
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