Orita H, Fukasawa M, Hirooka S, Uchino H, Fukui K, Washio M
Second Department of Surgery, Yamagata University School of Medicine, Japan.
Cardiovasc Drugs Ther. 1994 Dec;8(6):851-9. doi: 10.1007/BF00877404.
In this study we evaluated cardiac myocyte viability and function under hypothermic conditions using three types of buffer solutions: phosphate buffer solution (PBS), Krebs-Henseleit bicarbonate buffer solution (KHB), and Hepes buffered minimum salt solution (MSS). As a control, normal saline solution (NSS) was used. Cardiac myocytes were isolated from neonatal rat ventricles. Myocytes (12.5 x 10(5) myocytes/culture flask) were then incubated at 4 degrees C for 6, 12, 18, and 24 hours in various buffer solutions. After each incubation time, CPK and LDH were measured. The myocytes were then incubated for an additional 24 hours at 37 degrees C to evaluate the recovery of the myocyte beating rate. Group MSS had a significantly better beating rate recovery than group NSS (control) after 18 hours (MSS, 32.7%, NSS, 0.0% of control; i.e., beating rate prior to hypothermic incubation). In contrast, group KHB showed a significantly lower recovery ratio than group NSS at 12 hours (41.0%, 78.8%, respectively), and the lowest recovery was observed in group PBS beginning at 6 hours of hypothermic incubation (27.6%). Group MSS significantly suppressed the release of CPK and LDH compared to group NSS at 24 hours (MSS, 246.7 and 440.2 mIU/flask; NSS, 369.7 and 821.3 mIU/flask, respectively). In contrast, groups PBS and KHB showed significantly increased CPK and LDH levels compared to group NSS after 12 hours (PBS, 388.6 and 721.4 mIU/flask; KHB, 340.5 and 540.5 mIU/flask; NSS, 91.5 and 222.7 mIU/flask, respectively). In conclusion, Hepes buffer has cytoprotective characteristics that may be suitable for long-term hypothermic preservation of immature myocardium compared to phosphate or bicarbonate buffer.
在本研究中，我们使用三种缓冲溶液评估了低温条件下心肌细胞的活力和功能：磷酸盐缓冲溶液（PBS）、克雷布斯 - 亨泽莱特碳酸氢盐缓冲溶液（KHB）和羟乙基哌嗪乙磺酸缓冲的最低盐溶液（MSS）。作为对照，使用了生理盐水溶液（NSS）。心肌细胞从新生大鼠心室中分离出来。然后将心肌细胞（12.5×10⁵个心肌细胞/培养瓶）在4℃下于各种缓冲溶液中孵育6、12、18和24小时。在每个孵育时间后，测量肌酸磷酸激酶（CPK）和乳酸脱氢酶（LDH）。然后将心肌细胞在37℃下再孵育24小时，以评估心肌细胞搏动率的恢复情况。18小时后，MSS组的搏动率恢复明显优于NSS组（对照组）（MSS组为对照组的32.7%，NSS组为0.0%；即低温孵育前的搏动率）。相比之下，KHB组在12小时时的恢复率明显低于NSS组（分别为41.0%和78.8%），并且从低温孵育6小时开始，PBS组的恢复率最低（27.6%）。与24小时时的NSS组相比，MSS组显著抑制了CPK和LDH的释放（MSS组分别为246.7和440.2 mIU/培养瓶；NSS组分别为369.7和821.3 mIU/培养瓶）。相比之下，12小时后，PBS组和KHB组的CPK和LDH水平与NSS组相比显著升高（PBS组分别为388.6和721.4 mIU/培养瓶；KHB组分别为340.5和540.5 mIU/培养瓶；NSS组分别为91.5和222.7 mIU/培养瓶）。总之，与磷酸盐或碳酸氢盐缓冲液相比，羟乙基哌嗪乙磺酸缓冲液具有细胞保护特性，可能适用于未成熟心肌的长期低温保存。
