Walsh M J, Blum J J
Department of Cell Biology, Duke University Medical Center, Durham, NC 27710.
Mol Biochem Parasitol. 1992 Feb;50(2):205-14. doi: 10.1016/0166-6851(92)90217-8.
This study further explores the effects of hypoxia and acute osmotic stress on intermediary metabolism of Leishmania major and Leishmania donovani. Late log phase promastigotes were washed and incubated with glucose as the sole exogenous carbon source, and rates of glucose consumption and product formation were measured as a function of osmotic strength (610, 305, and 167 mOsm kg-1) and pO2 (95, 10, and 0% O2) in the presence of 5% CO2. Very mild hypoxia dramatically altered flux through the pathways of intermediary metabolism and increased the rates of production of the major metabolites, thus confirming the presence of a low-affinity O2 sensor which was active under all osmolalities tested. The data also require that as pO2 is lowered towards anoxia an endogenous carbohydrate source is mobilized. Under aerobic conditions, acute hypo-osmotic stress had little effect on product formation, whereas acute hyperosmotic stress altered metabolism in a manner similar to mild hypoxia, with the exception of decreasing the rates of acetate and succinate production. It was also shown in L. donovani promastigotes that the effects of anoxia and hyperosmolality were not additive. Thus, separate sensors with partially overlapping actions are involved in the metabolic responses to hypoxia and hyperosmolality. There was no apparent species-specificity for the responses to pO2 and osmotic stress. Uncoupling with carbonyl cyanide p-trifluoromethoxyphenylhydrazone caused changes in metabolite flux patterns which differed from the changes caused by either hypoxia or acute osmotic stress, while rotenone and calcium ionophore A23187 had no significant effects. The identity of the sensors responsive to pO2 and osmolality, and the mechanisms by which they regulate flux through the pathways of intermediary metabolism, require further study.
本研究进一步探讨了缺氧和急性渗透应激对硕大利什曼原虫和杜氏利什曼原虫中间代谢的影响。将对数后期的前鞭毛体洗涤后，以葡萄糖作为唯一的外源碳源进行孵育，并在5%二氧化碳存在的情况下，测定葡萄糖消耗速率和产物形成速率，作为渗透强度（610、305和167 mOsm kg-1）和pO2（95%、10%和0% O2）的函数。非常轻微的缺氧显著改变了中间代谢途径的通量，并增加了主要代谢产物的产生速率，从而证实存在一种低亲和力的O2传感器，该传感器在所有测试的渗透压下均有活性。数据还表明，随着pO2降至缺氧水平，内源性碳水化合物源会被调动。在有氧条件下，急性低渗应激对产物形成影响不大，而急性高渗应激以类似于轻度缺氧的方式改变代谢，但乙酸盐和琥珀酸盐的产生速率降低。在杜氏利什曼原虫前鞭毛体中还显示，缺氧和高渗的影响不是相加的。因此，在对缺氧和高渗的代谢反应中涉及具有部分重叠作用的不同传感器。对pO2和渗透应激的反应没有明显的物种特异性。用羰基氰化物对三氟甲氧基苯腙解偶联导致代谢物通量模式的变化，这与缺氧或急性渗透应激引起的变化不同，而鱼藤酮和钙离子载体A23187没有显著影响。对pO2和渗透压作出反应的传感器的身份，以及它们调节中间代谢途径通量的机制，需要进一步研究。
