The Role Of Heme Oxygenase-1 In The Pathogenesis Of Diabetic Peripheral Neuropathy And Related Mechanisms

Background:Diabetic peripheral neuropathy(DPN)is the most prevalent chronic complications of diabetes,which is the most important cause of foot ulceration and a major contributor to falls,fracture and non-traumatic amputation.The mechanisms of DPN are complicated,including metabolic factors(such as hyperglycemia,polyol metabolism alteration and glycosylation proteins increase),vascular factor(such as ultrastructural vessel changes and dysfunction of mircovascular)and immunologic or inflammatory factor.Increased oxidative stress and cell apoptosis are two of the most important pathological changes.Despite the recent major advances in elucidating the pathogenesis of diabetic neuropathy,there remains a lack of treatment options that effectively target the natural history of DPN or reverse DPN once established.Heme oxygenase-1(HO-1)is an endogenic enzyme,which degrades heme in an identical stereospecific manner to biliverdin with the concurrent release of CO and iron.HO-1 poses various cellular protective properties,including antioxidant,anti-inflammation,anti-apoptosis,signal transduction and immune-regulation,which is a novel target in neurological research.In central nerve system,HO-1 can protect cells by modulating the activity of soluble guanylate cyclase,opening calcium-activated potassium channels,promoting vasodilation and reducing apoptosis.However,research concerning the role of HO-1 in peripheral neuropathy,especially DPN is limited.Methods:We first collected 1,800 inpatients with type 2 diabetes(including 68 subjects who had 1-year follow-up).DPN was diagnosed by nerve conduction velocities accompanied with neuropathic signs and symptoms.The serum levels of bilirubins(a product of HO-1 and reflect the activity of HO-1)were compared between patients with and without DPN and the role of baseline bilirubin on the progression of DPN was further investigated.Next,we collect the peripheral blood mononuclear cells and compared the expression of HO-1 by Q-PCR directly.Then,we established DPN mice model and detected the dysfunction of peripheral nerve by thermal and mechanical hyperalgesia,nerve conduction velocities,transmission electron microscope and intra-epidermal nerve fiber density.The expression of HO-1 in sciatic nerve was detected by immunofluorescence and Q-PCR.Next,Schwann cells were dealt with hyperglycemia and the expression of HO-1 was examined by Western blot,cellular proliferation and apoptosis were tested by MTT assay and flow cytometry and the function of mitochondria was detected by DCFH-DA staining and ATP measurement.Furthermore,Schwann cells were treated with HO-1 agonist CoPP and inhibitor SnPP to investigated the relationship between HO-1 and Schwann cell apoptosis and mitochondria dysfunction in euglycemia.At last,Schwann cells dealt with hyperglycemia were treated with CoPP to clarify the protection role of HO-1 in Schwann cell function under hyperglycemia and further investigated signaling pathway involving autophagy.Results:The levels of serum bilirubin were significantly decreased in patients with DPN.Lower level of bilirubin was a risk factor for the presence and progression of DPN after multivariate adjustment and can be used as an additional biomarker of DPN.The expression of HO-1 was decreased in DPN patients,which is consistent with bilirubin level.The expression of HO-1 was also decreased in sciatic nerve of DPN mice,which accompanied with decreased peripheral nerve sensation,nerve conduction velocities,intra-epidermal nerve fiber density and the number of mitochondria.When treated with hyperglycemia,Schwann cells exhibited decreased proliferation,increased apoptosis,impaired function of mitochondria and the expression of HO-1 was significantly decreased.The treatment of HO-1 inhibitor,SnPP,caused mitochondria dysfunction and cellular apoptosis in Schwann cell with euglycemia,which was similar with the treatment of hyperglycemia.CoPP,the agonist of HO-1,alleviated the impairment of mitochondria function and cell apoptosis in Schwann cell with hyperglycemia as well as activating the key factors,including LC3 ? and Beclin1 in autophagy pathway.Conclusions:HO-1 played an important role in the pathogenesis of DPN.Lower level of bilirubin,represented lower activity of HO-1,was a risk factor for the presence and progression of DPN and can be used as an additional biomarker of DPN.HO-1 improved the mitochondria dysfunction and cellular apoptosis of Schwann cells with hyperglycemia by activating autophagy pathway.HO-1 can be a potential effective target for the screening and treatment of DPN.