Objective To study the protective effects of tBHQ on type 2 diabetic rats retina and its related mechanism.Methods Sixty SD rats were divided into normal control group (NC group),model group (diabetes mellitus,DM group) and tBHQ group.After feeding with high fat and high sugar diets for 4 weeks,the rats in model group were induced by intraperitoneal injection of STZ for the model of type 2 diabetes mellitus.1％ tBHQ were added into the high fat and high sugar feed 1 week later after successfully modeled in the tBHQ group.Fasting plasma glucose (FPG) and fasting serum insulin (FINs) were detected at 4 and 12 weeks after modeled.Immunohistochemical method and real-time fluorescent quantitative PCR (qRT-PCR) were respectively used to detect the distributions and relative expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2),heme oxygenase 1 (HO-1),B-cell lymPhoma 2 (Bcl-2) and vascular endothelial growth factor (VEGF) in retinas of the rats.Results FPG levels totally comparative differences were statistically significant between each group (F =78.531,P =0.000).The level of FPG in DM and tBHQ group were obviously higher than that in NC group,the 12 weeks was higher than 4 weeks in DM group,and the 12 weeks was lower than 4 weeks in tBHQ group (all P ＜ 0.05).Totally comparative differences in the level of FINs were statistically significant (F=22.480,P =0.000),NC group was lower than DM and tBHQ group,12 weeks was higher than 4 weeks (all P ＜ 0.05).Immunohistochemical detection showed that each factor in each group were expressed in the retina of rat,and the relative expressions of Nrf2,HO-1,Bcl-2 and VEGF protein in retina have totally statistically differences in different time points after modeled (all P ＜0.05).The expressions of each factor in DM group were more than those in the NC group.The Nrf2,HO-1 and Bcl-2 in tBHQ group were more than those in the DM group,but the VEGF was lower.At 12 weeks,the expressions of VEGF and Nrf2 in DM group were more than those at 4 weeks,but the HO-1 was lower;the Nrf2,HO-1 and Bcl-2 in tBHQ group were more than those at 4 weeks (all P ＜ 0.05).PCR tests revealed that the relative expressions of Nrf2,HO-1,Bcl-2 and VEGF mRNA in retina have totally statistically differences in different time points after modeled(all P ＜ 0.05).The expressions of each factor in DM group were more than those in the NC group.The Nrf2,HO-1 and Bcl-2 mRNA in tBHQ group were more than those in the DM group,but the VEGF mRNA was lower.At 12 weeks,VEGF mRNA in DM group and Nrf2,HO-1,Bcl-2 in tBHQ group were more than those at 4 weeks (all P ＜ 0.05).Conclusion tBHQ maybe have protection for islet function on diabetic rat,and can induce the expressions of Nrf2,HO-1,Bcl-2 in retina of diabetic rats and reduce the expression of VEGF,inhibit the oxidative stress of retinal tissue damage,reduce cell apoptosis and inhibit the proliferation of retinal blood vessels.tBHQ may protect the retina of diabetic rats through the Nrf2/HO-1/VEGF and Nrf2/Bcl-2 way.

Background Apoptosis is a primary clinical pathological mechanism of diabetic retinopathy (DR).Oxidative stress and high glucose can activate cell apoptosis pathway and thus leads to cellular damage.It is confirmed that tert-butyl hydroquinone (tBHQ) plays an antioxidation effect,however,whether it has a protective role on retinal cells in DR is still unelucidated.Objective This study was to investigate the effect of tBHQ on vascular endothelial growth factor (VEGF) and bcl-2 expressions in retina of type 2 diabetic rats and its possible mechanism via nuclear factor erythroid 2-related factor 2/antioxidant response element (Nrf2/ARE) signal pathway.Methods Fifty clean healthy male SD rats were included in this experimental study.Ten rats were fed with normal diet as the normal control group,and other rats were fed with high fatty and high sugar food for 4 weeks.After 12 hours of fasting,streptozotoin (STZ) (30 mg/kg) was intraperitoneally injected to induce the type 2 diabetic models.The model rats were randomly divided into the diabetic control group and tBHQ group and 1％ tBHQ was added into the high fatty and sugar food 1 week after modeling in the tBHQ group.Fasting plasma glucose (FPG) level,blood total cholesterol (TC) level,blood triglyceride (TG) level,high density lipoprotein-cholesterol (HDL-C),low density lipoproteincholesterol (LDL-C) and fasting serum insulin (FINs) were detected 4 and 12 weeks after modeling,respectively,and radio immunoassay was used to detect the FIN levels of the rats.The relative expression of VEGF and bcl-2 in retinas of the rats were assayed by immunohistochemistry and fluorescence real-time quantitative PCR (qRT-PCR).The use of the animals complied with the Regulations for the Administration of Affairs Concerning Experimental Animals by State and Technology Commission.Results Type 2 diabetic models were successfully established in 35 rats with successful rate 92.1％.The FIN levels were significantly different among different groups and time points (Fgroup =22.480,P =0.000;Ftime =7.636,P =0.008).The FPG,TC,TG and LDL-C levels were significantly different among the groups (FPG:Fgroup =78.531,P =0.000;TC:Fgroup =28.049,P =0.000;TG:Fgroup =13.108,P =0.000;LDL-C:Fgroup =6.804,P＜0.05).Immunohistochemistry showed that VEGF and bcl-2 were mainly expressed in retinal ganglion cell layer,inner plexiform layer and outer plexiform layer.The expressions of VEGF and bcl-2 proteins were significantly different among different groups (VEGF:Fgroup =11.805,P =0.000;bcl-2:Fgroup =22.943,P =0.000);the expression level of bcl-2 protein was higher in 12 weeks after modeling than that in 4 weeks after modeling in the tBHQ group (P＜0.05).The expressions of VEGF and Bcl-2 mRNA in rat retinas were significantly different among different groups and time points (VEGF:Fgroup =79.220,P =0.000;Ftimo =6.090,P＜0.05;Bcl-2:Fgroup =105.000,P=0.000;Ftime =13.170,P=0.001).Four and eight weeks after modeling,the expressions of VEGF and Bcl-2 mRNA in the diabetic control group and tBHQ group were significantly higher than that in the normal control group,and the expressions of Bcl-2 mRNA in the tBHQ group were significantly higher than that in the model control group (all at P＜0.05);the expression of Bcl-2 mRNA was higher at 12 weeks after modeling than that at 4 weeks in the tBHQ group (P＜0.05).Conclusions tBHQ produces anti-oxidative-damage and anti-apoptosis effects on retinal cells by up-regulating VEGF expression and down-regulating bcl-2 expression in DR rats.In addition,tBHQ may have effects on lowering high blood sugar,regulating insulin and blood lipid levels.

OBJECTIVETo determine the genotypes of three blood samples suspected as B subtype through DNA sequencing.

METHODSThe samples were first genotyped with PCR-SSP. Exons 6 and 7 of the ABO genes were subjected to PCR, direct sequencing, and cloned sequencing to determine the genotypes.

RESULTSSerological results of the three samples were similar, with red cells being weakly agglutinatable by anti-B and serum containing anti-B. The samples were preliminarily genotyped as B/O1. Sequencing analysis showed that all three samples contained an O allele and a 905A>G mutation of the B gene, which was previously defined as Bx02.

CONCLUSIONThrough sequencing analysis, the three samples typed as B subtype with serological testing were identified as Bx phenotype. The genotype of samples 1 and 2 was Bx02/O101, and that of sample 3 was Bx02/O102.

ABO Blood-Group System ; genetics ; Adult ; Alleles ; Base Sequence ; Blood Grouping and Crossmatching ; methods ; Exons ; genetics ; Female ; Genotype ; Humans ; Male ; Mutation ; Phenotype ; Polymerase Chain Reaction ; methods ; Sequence Analysis, DNA ; methods ; Young Adult