Morrison A D, Berwick L, Orci L, Winegrad A I
J Clin Invest. 1976 Mar;57(3):650-60. doi: 10.1172/JCI108321.
An in vitro preparation of rabbit aortic "intima-media" previously shown to exhibit stable rates of respiration and glucose metabolism and the high rate of aerobic glycolysis considered characteristic of the metabolism of this tissue was subjected to electron microscopic examination. In samples examined immediately after the aortae were dissected free of adipose tissue and adventitia, under conditions similar to those now in common use, marked and widespread alterations in endothelial cell structure were present, including loss of cell integrity. The vascular smooth muscle cells retained a normal electron microscopic (EM) appearance. During subsequent incubation with 5 mM glucose in Krebs-Ringer bicarbonate (KRB), pH 7.4, under the conditions usually employed in studies of this preparation, large zones of the luminal surface were rapidly denuded of endothelium, and the remaining endothelial cells exhibited a wide range of ultrastructural alterations. The smooth muscle cells, however, continued to maintain a normal EM appearance. A method was developed to prepare segments of rabbit aortic intima-media which retained an intact layer of endothelium resembling that observed in tissue fixed in situ. During a 1-h incubation with 5 mM glucose in KRB, pH 7.4, gas phase 5% CO2/95% O2, containing 6% bovine serum albumin, the intact aortic intima-media preparation retains an essentially unmodified EM appearance and exhibits linear rates of respiration. Under these conditions the intact aortic intima-media preparation exhibits significantly higher rates of O2 uptake and glucose uptake than those observed in our previous preparation or in other reported aortic intima-media preparations. The intact aortic intima-media does not exhibit the high rate of aerobic glycolysis during in vitro incubation that has been considered characteristic of the metabolism of rabbit, rat, and swine aortic intima-media. In addition, the magnitude of the Pasteur effect was far greater than that observed in other aortic intima-media preparations. The data suggest that component cells of the aortic intima-media may derive a major fraction of their energy requirements from respiration; they raise further questions concerning the significance of the high rate of aerobic glycolysis observed when aortic intima-media preparations are incubated in vitro, and they suggest that documentation of the EM appearance of the endothelium in such preparations is desirable.
先前已证明兔主动脉“内膜 - 中膜”的体外制剂表现出稳定的呼吸速率和葡萄糖代谢速率，且具有该组织代谢特有的高有氧糖酵解速率，对其进行了电子显微镜检查。在将主动脉从脂肪组织和外膜中分离出来后立即进行检查的样本中，在与目前常用条件相似的情况下，内皮细胞结构出现了明显且广泛的改变，包括细胞完整性丧失。血管平滑肌细胞保持正常的电子显微镜（EM）外观。在随后于pH 7.4的 Krebs - Ringer碳酸氢盐（KRB）中用5 mM葡萄糖进行孵育期间，在该制剂研究通常采用的条件下，管腔表面的大片区域迅速内皮剥脱，剩余的内皮细胞表现出广泛的超微结构改变。然而，平滑肌细胞继续保持正常的EM外观。开发了一种制备兔主动脉内膜 - 中膜片段的方法，该片段保留了完整的内皮细胞层，类似于在原位固定组织中观察到的情况。在含有6％牛血清白蛋白的pH 7.4的KRB中用5 mM葡萄糖进行1小时孵育期间，气相为5％CO2 / 95％O2，完整的主动脉内膜 - 中膜制剂保持基本未改变的EM外观，并表现出线性呼吸速率。在这些条件下，完整的主动脉内膜 - 中膜制剂表现出比我们之前的制剂或其他报道的主动脉内膜 - 中膜制剂中观察到的显著更高的氧气摄取和葡萄糖摄取速率。完整的主动脉内膜 - 中膜在体外孵育期间未表现出被认为是兔、大鼠和猪主动脉内膜 - 中膜代谢特征的高有氧糖酵解速率。此外，巴斯德效应的幅度远大于在其他主动脉内膜 - 中膜制剂中观察到的幅度。数据表明，主动脉内膜 - 中膜的组成细胞可能从呼吸中获取其大部分能量需求；它们进一步引发了关于在体外孵育主动脉内膜 - 中膜制剂时观察到的高有氧糖酵解速率的意义的问题，并且它们表明在这种制剂中记录内皮的EM外观是可取的。
