고혈압백서(1-kidney, 1-clip-hypertensive rat)의 적혈구에서 노화 과정에 따른 Na, K-ATPase의 변동을 관찰하고자 노화적혈구를 분리한 다음 세포막에서의 Na-pump 활성도 및 ouabain의 결합실험과 Rb의 세포내 유입실험을 시행하여 다음과 같은 결과를 얻었다. 1. 본 실험에 사용한 고혈압 백서의 혈압은 수축기 및 이완기 혈압이 165.5/119.0 mmHg로 유의하게 증가 하였다. 노화 적혈구의 평균용적(MCV)과 세포막 단백질 함량은 감소되고 혈색소치는 증가되었다. 2. 110 mM NaCl 및 10 mM KCI 존재하에서의 적혈구 세포막 Na, K-ATPase활성도는 대조군에 비해 고혈압군에서 억제 되었으며 양군 모두에서 노화에 의해 그 활성도가 감소되었다. 3. 4 mM RbCl 존재 하에서 Ouabain에 의해 억제되는 Rb의 유입은 정상 및 고혈압군의 노화적혈구에서 약간 감소되었으며 고혈압군의 young erythrocyte에서는 오히려 약간 증가 되었다. 4. 16 mM RbCl 존재 하에서 Ouabain에 의해 억제되는 Rb의 유입은 양군의 노화 적혈구에 서는 각군의 young erythrocyte에 비해 약 30-50% 감소되었으며, 고혈압군에서는 특히 young erythrocyte에서 정상군의 young erythrocyte에 비해 유의하게 감소되었다. 5. 0.13 × 10-6M과 1 × 10-6M에서의 ouabain binding은 정상군의 노화적혈구에서는 young erythrocyte에 비해 약간 감소되었으나 고혈압군의 노화적혈구에서는 유의하게 감소되었다. 6. 6 × 10-6M과 64 × 10-6M 에서의 ouabain binding은 양군의 노화 적혈구에서는 약간 감소되었지만 유의성은 없었으며 고혈압군의 young erythrocyte 및 노화적혈구에서는 정상군의 young erythrocyte및 노화 적혈구에 비해 유의하게 감소되었다. 이상의 결과로부터 (1) 고혈압쥐의 young erythrocyte에서는 low affinity의 Na-pump수의 감소 및 molecular activity의 증가, (2) 정상쥐의 노화 적혈구에서는 molecular activity의 저하, (3) 고혈압쥐의 노화적혈구에서는 molecular activity의 저하 및 high affinity와 low affinity의 Na-pump수의 저하등에 의하여 Na-pump의 기능이 변동될 수 있을 것으로 추측된다.
To study the age dependent change of Na, K-ATPase in the erythrocyte of hypertensive rat, 1-kidey 1-clip hypertensive rat was made by the removal of right kidney and partial ligation of left renal artery. After 4 weeks, aged erythrocyte fraction was separated by density gradient centrifugation, and Na, K-ATPase activity and 3H-ouabain binding with ghost cell membrane and ouabain sensitive Rb-uptake with whole cell were measured. 1) In the hypertensive rats, blood pressure was significantly increased to 165.5/119.0 mmHg (systolic/diastolic). Mean corpuscular volume and membrane protein(mg) per 109RBC were decreased and hemoglobin content was increased in the aged erythrocyte. 2) Na, K-ATPase activity in the solution containing 110 mM NaCl and 10 mM KCI, was decreased in hypertensive rat, and decreased in aged erythrocyte of both group. 3) Ouabain sensitive Rb-uptake by low RbCl concentration(4 mM) was slightly decreased in aged erythrocyte compared to that in young erythrocyte of each group, but slightly increased in young erythrocyte in hypertensive rat compared to that in normotensive rat. 4) Ouabain sensitive Rb-uptake by high RbCl concentration(16 mM) was decreased about 30% to 50 % in aged erythrocyte in both group. And in hypertensive rat, especially in young erythrocyte it was significantly decreased compared to that in normotensive rats. 5) 3H-ouabain binding at 0.13 or 1 × 10-6M ouabain concentration was slightly decreased in aged erythrocyte of normotensive rat, and significantly decreased in aged erythrocyte of hypertensive rats. 6) 3H-ouabain binding at 6 or 64 × 10-6M ouabain concentration is slightly decreased in aged erythrocyte of both group, but significantly decreased in young and aged erythrocyte of hypertensive rats compared to that of normotensive rats. The present results suggest that (1) in the young erythrocyte of hypertensive rat, the alterations of Na-pump activity that slightly increased in weak stimulation and inhibited in strong stimulation, may be related to increased molecular activity and the decrease in the number of low affinity site without change in high affinity site, (2) in the aged erythrocyte of normotensive rat, inhibited Na-pump may be related to the change in molecular activity of pump. (3) And in the aged erythrocyte of hypertensive rat, it may be related to the decrease in the number of high and low affinity site as well as the change in molecular activity.
