心力衰竭患儿血清β3受体自身抗体水平对离体成鼠心肌细胞的负性变力作用

Negative Inotropic Effects of β_3-Adrenoceptor Autoantibodies from Children with Heart Failure on Isolated Cardiomyocytes of Adult Rats

目的探讨心力衰竭患儿血清抗β3肾上腺素受体(β3AR)自身抗体水平对离体成年大鼠心肌细胞收缩效应的影响。方法采用ELISA筛查心力衰竭和健康儿童血清抗β3AR自身抗体,利用亲和层析法提纯抗β3AR自身抗体阳性血清免疫球蛋白G(IgG);利用动缘探测系统观察提纯的抗β3AR自身抗体阳性IgG对离体成年大鼠心肌细胞收缩效应的影响。结果50例心力衰竭患儿血清抗β3AR自身抗体的阳性率和平均几何滴度为分别30%(15/50例)和1∶(48.32±2.58),明显高于健康儿童[12%(6/50例),P<0.05;1∶(13.72±1.49),P<0.001];与对照组比较,该抗体可降低成年大鼠心肌细胞最大收缩幅度/初长度(3.86±0.31)%、最大收缩速率[(-0.47±0.07)μm/s]和最大舒张速率[(0.17±0.02)μm/s](Pa<0.001)。该作用不能被纳多洛尔阻断,但可被布拉洛尔或β3AR细胞外第二环合成抗原阻断。结论心力衰竭患儿血清抗β3AR细胞外第二环自身抗体的阳性率和平均几何滴度均明显高于健康儿童,且该自身抗体具有类似β3AR激动剂样负性变力效应。

Objective To explore the effects of the autoantibody against the second extracellular loop of β3 - adrenoceptor （ β3 AR） from children with heart failure on isolated cardiomyocytes of adult rats. Methods The synthetic peptide corresponding to the sequence of the second extracellular loop （ 176 - 202 amino acid residues） of the human β3AR was used as the antigen to screen the anti - β3AR autoantibodies from the sera of the 50 children with heart failure and 50 healthy children by enzyme linked immunosorbent assay（ELISA） ;IgG in the children with heart failure of positive autoantibodies sera was purified. The effects of purified IgG per each group on contractile response of isolated cardiomyocytes of adult rats were observed using motion - edge detection system and double - provocation fluorescence photomultiplier system. Results Among the 50 children with heart failure, the anti - β3 AR autoantibodies positive rate was 30% （ 15/50 cases）, and ob-viously higher than that of healthy children [ 12% （6/50） cases, P 〈 0.05 ], and the mean geometric antibody titer of patients group was [ 1 ： （48.32 ± 2.58）]significantly higher than that of control group [ 1 ： （ 13.72± 1.49 ）, P 〈 0. 001 ]. Compared with control group, the autoantibodies （0. 1 μmol/L） against β3 AR from the patients group could reduce cell shortening/cell length （3.86% ± 0.31% ）, the maximal velocity of shortening[ （ -0.47 ±0.07） μm/s] and relengthening[ （0.17 ±0.02） μm/s] （Pa 〈0. 001 ） of adult isolated cardiomyocytes,which could not be modified by pretreating myocytes with Nadolol （ β3 AR and β1AR antagonist ）, but nearly be prevented by Bupranolol （non-selective β3AR antagonist） or β3AR specific antigen. Conclusions The positive rate and mean geometric antibody titer of the autoantibodies against the second extracellular loop of β3AR in the sera of the children with heart failure were obviously higher than that of homogeneous healthy children, and the autoantibodies can induce β3AR agonist -like negative inotropic effects, which may play a role in pathophysiological mechanism during the development of children heart failure.