Erythrocyte function modifying substance
A technology of erythrocytes and substances, applied in the field of pharmaceutical compositions containing the above-mentioned substances, capable of solving problems such as unclear details and ambiguity of ATP oxygen concentration dependence
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Embodiment 1
[0154] Add heparin to collect 20mL of human peripheral blood, centrifuge at 2000rpm for 7 minutes, collect red blood cell pellet, wash and use Krebs buffer (krebs buffer) to make 1×10 7 cell / ml of erythrocyte suspension. Incubate the sealed cuvette containing 1.8 mL of Krebs buffer solution whose oxygen concentration is adjusted to the required value of 0-150 mmHg at 37°C, and inject 200 μl of erythrocyte suspension into the sealed cuvette with a microsyringe. After 5 minutes, place in ice at 4°C to stop the reaction, sample 200 μl at 4°C, centrifuge, and quantify ATP in the supernatant by fluorescence analysis. Normal red blood cells are at partial pressure of oxygen (PO 2 ) Curve with an inflection point around 50~70mmHg ( figure 1 S-shaped curve). The CO-saturated red blood cells produced by passing 100% CO into the red blood cells for one minute are at a low level, and the ATP release is stable ( figure 1 "CO-Hb" curve); with 10 -4 mol / L (final concentration) treated ...
Embodiment 2
[0156] In this example, the effects of carbon monoxide-treated red blood cells (CO-Hb) and NO-treated red blood cells (αNO-Hb) on ATP release were studied. The method is according to Example 1 above. The function of NO is to completely deoxygenate the collected erythrocytes by argon gas, then add NO-glutathione at a ratio of 1:2 relative to the concentration of Hb, centrifuge for several minutes, and use the recovered erythrocytes for experiments. The hemoglobin contained in the above erythrocytes is expressed as "αNO-Hb".
[0157] Normal-Hb and CO-Hb showed the same result as Example 1 ( figure 2 ), αNO-Hb is higher than the bezafibrate of embodiment 1, and the release of ATP is stable ( figure 2 ). figure 2 The "*" in the middle indicates that there is a significant difference (P<0.05) compared with normal red blood cells (Hb).
Embodiment 3
[0159] Recovery Effect of Human Washed Red Blood Cell Samples on Liver Dysfunction Induced by 40% Hemorrhagic Shock in Rats
[0160]Wistar male rats (250-300 g) were anesthetized by intramuscular injection of 50 mg / kg pentobarbital sodium. The trachea of the anesthetized rat was incised, and the femoral artery was cannulated. The bile duct was then intubated to monitor bile secretion. The left carotid artery of the rat was also intubated as blood vessel removal, and the blood was removed at 1-2 mL / min until the total blood loss reached 40% of the whole body blood. After 15 minutes of shock state after the end of blood removal, each preparation was adjusted to the amount of Hb equivalent to the amount of hemorrhage, and after 60 minutes, the base excess was calculated according to the hepatic microcirculation, bile flow, Hb of arterial blood, and bicarbonate concentration pH ( image 3 ). Usually the base excess is above plus 2.5 for alkalosis, and below minus 2.5 for acid...
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