The Role Of SET7/9 And LSD1 On Macrovascular Complications In Patients Wtih Type 2Diabetes

Objective:The type 2 diabetic macrovascular complications currently are the main reasons for disability and death in diabetic patients, which have not yet fully understood. Nowadays,the treatments of those are limited,because the pathogenesis of diabetic macrovascular complications is not completely clear yet. The basic pathology of diabetic macrovascular complications considered atherosclerosis (AS) at present, and AS is not only the result of lipid metabolism disorders but also caused by inflammation which plays the key role in endothelial injury and vascular membrane lesion.Nuclear factor-kappa B (NF-κB) is the core factor of modulating inflammatory reaction,and it can be activated by hyperglycemia,glycosylation end products and hyperlipidemia etc,which could induce the transcription of many factors including cell growth factors, chemotactic factors(CF), cell adhesion molecules (CAM) and inhibitory kappa B (IκB), which could enhance the function of mononuclear leucocytes in blood circulation where they participate the early steps of AS through aggregation and retention in blood circulation.This is the main mechanism of diabetic macrovascular diseases. Recent studies have shown that histone methyltransferases and demethylases associated with diabetic metabolic memory possibly play important role in modulating the gene transcription of NF-KB.The epigenetics refers to the alteration of gene expression and function without the change of DNA sequence,including the methylation of DNA and covalent modification of histone etc. Exploring pathogenesis of disease by epigenetics is the hot research in recent years. It has been found that epigenetic mechanism may control the genesis and development of diabetes recently. However, whether histone methyltransferases and demethylases involve in the patients of type 2 diabetic macrovascular complications has not been reported. To explore the role of SET7/9 and Lysine-specific histone demethylase 1 (LSD1) in the diabetic macrovascular diseases and provide theoretical basis to treat them,we detected and compared with the protein level and mRNA expression of SET7/9 and LSD1 in normal person, type 2 diabetics without macrovascular complication and type 2 diabetic macrovascular complications patiens. Methods:1.Research object grouping:(1) Normal control group (NC group):we selected 10 Healthy volunteers randomly.(2) type 2 diabetics without macrovascular complication group (DM group):we randomly selected 10 type 2 diabetes mellitus patiens without macrovascular complication.(3)type 2 diabetic macrovascular complications group (DV group):10 patiens with type 2 diabetic macrovascular complications were brought into the group.2.Draw 3.5ml peripheral blood sample, with 3ml blood used for separation of individual nucleus cells and 0.25ml blood used for extraction of total RNA.3.Western blot was used to detect the protein expression of SET7/9 and LSD1.4.Cellular immune fluorescence was used to observe the expression of LSD1 protien after cell smear through confocal microscope.5.RT-PCR was used to detect the expression of NF-κB, SET7/9 and LSD1 mRNA.6.Statistical Analysis: Continuous data was expressed as means±SEM.Multiple group comparisons were made by one-way ANOVA analysis with Student-Newman-Keuls post hoc analysis.Times in the same group comparison were made by Student's t test. Statistical significance was accepted at P<0.05. Results:1.protein expression detected by Western blot:(1) Expression of SET7/9 protein was not changed in DM group compared with NC group,but expression of SET7/9 protein was increased in DV group compared with NC group(P<0.05) and DM group(P<0.05) (2) Expression of LSD1 protein was decreased in DM group compared with NC group(P<0.05),and the expression was decreased in DV group compared with NC group (P<0.05) and DM group(P<0.05) 2.Intensity of LSD1 protien fluorescence was reduced in DM group compared with NC group,and the intensity was further reduced in DV group.3.RT-PCR detecting mRNA expression:(1)the NF-κB mRNA expression was increased in DM group (P<0.05)and DV group (P<0.05) compared with NC group, and it was increased in DV group (P<0.05) compared with DM group; (2) the SET7/9 mRNA expression was increased in DM group (P<0.05)and DV group (P<0.05) compared with NC group, and it was increased in DV group (P<0.05) compared with DM group; (3) the LSD1 mRNA expression was decreased in DM group (P<0.05)and DV group (P<0.05) compared with NC group, and it was decreased in DV group (P<0.05) compared with DM group. Conclusions: 1. Disorders of histone methyltransferase SET7/9 and demethylase LSD1 protien expression occur in diabetic patients.2. Activated NF-κB gene expression by disorders of SET7/9 and LSD1 may be the important epigenetic mechanism of the occurrence and development of type 2 diabetic macrovascular complications.