Modification of partial pressure of oxygen (P50) in mammalian red blood cells by incorporation of an allosteric effector of hemoglobin.

Internalization of inositol hexaphosphate (IHP) in mammalian red blood cells (RBC) produces a modification of the hemoglobin-oxygen affinity, leading to a rightward shift of the dissociation curve. The process of incorporation, based on an osmotic shock, has been tested on RBC of different species. Two dialysis protocols have been defined to transform RBC, the first one for small volumes in a cellulose bag and the other for larger volumes using a commercially available dialysis device. Different optimal conditions must be used for each species. Most of the cellular characteristics of the transformed RBC having encapsulated IHP are similar to those of native cells. For several species, such modified RBC could be reinfused and used for physiological studies.

Cardiac effects of phytic acid induced high P50 with free and limited coronary blood flow.

A blood preparation with a large increase in P50, obtained by a new technique to encapsulate inositol hexaphosphate (IHP) in erythrocytes was evaluated on an isolated heart model. Each heart was alternatively perfused with control stored human blood (P50 = 2.12.+/- 0.9 mmHg) and IHP-treated human blood (P50 = 42.5 +/- 9.33 mmHg). Changes of perfusates were performed when coronary blood flow (CBF) was free and adapted to a constant perfusion pressure (FREE CBF). Changes of perfusates were also performed at a constant but restricted CBF corresponding to 55% of basal value (RESTRICTED CBF). Both bloods were oxygenated and equilibrated to achieve the same acid-base balance, arterial PO2 and O2 content. When CBF was not restricted, switching from control blood to IHP-treated blood, induced a decrease in CBF (-23%), an increase in coronary sinus PO2 (57%) and a decrease in coronary sinus O2 content associated with an increase in myocardial O2 consumption (14%). These metabolic changes were associated with a decrease in left ventricular developed pressure (LVDevP) (-15%) and its maximal positive derivative (-12%). When CBF was restricted, switching from control blood to IHP-treated blood, induced an increase in perfusion pressure (59%). This vasoconstriction was associated with the same changes in the blood gas measurements as those observed when CBF was not restricted, while LVDevP increased significantly (7%). It is concluded that the beneficial effects on myocardial metabolism from the increase in P50 with IHP, are lessened by the cardiodepressive effect of the blood preparation which is predominant when CBF is not restricted.

Long-term physiological effects of enhanced O2 release by inositol hexaphosphate-loaded erythrocytes.

A continuous lysing and resealing procedure with erythrocytes permitted incorporation in these cells of inositol hexaphosphate (InsP6), a strong allosteric effector of Hb. This leads to significant rightward shifts of the HbO2 dissociation curves with in vitro P50 (partial pressure of O2 at 50% Hb saturation), values increasing from 32.2 +/- 1.8 torr for control erythrocytes to 86 +/- 60 torr (pH 7.40; PCO2 40 torr at 37 degrees C; 1 torr = 1.333 X 10(2) Pa). The shape of the dissociation curve was still sigmoidal, although the Hill coefficient was decreased. The life span of InsP6-loaded erythrocytes equaled that of control erythrocytes. The long-term physiological effects of the InsP6-loaded erythrocytes on piglets were increased O2 release and reduced cardiac output. The reduced O2 affinity of the InsP6-loaded erythrocytes was still effective 20 days after transfusion in awake piglets. The electrolyte concentration appeared stable over the 5-day observation period except for a transient, but significant, hyperkalemia immediately after transfusion. The reductions in the O2 affinity of Hb reported here are large compared with previously reported values. Introduction of InsP6 into viable erythrocytes improves tissue oxygenation when, for any reason, normal blood flow is impaired.

Oxygen transport to tissue modified by entrapment of an allosteric effector of haemoglobin in erythrocytes.

Oxygen affinity of haemoglobin is modulated by several parameters such as the allosteric effector 2-3 DPG for most mammalians. Inositol hexaphosphate (I.H.P.) exerts the same effect on haemoglobin. A previously developed new methodology for the entrapment of drugs into erythrocytes has been adapted to I.H.P.; it is based on a reversible osmotic shock. I.H.P. loaded red blood cells have characteristics very similar to those of native cells. The decrease in oxygen affinity is related to the dose of encapsulated I.H.P. In piglets, transfusion of such cells has led to an increase of oxygen extraction from haemoglobin. Normal anesthetized animals regulate their oxygen consumption by reduction of cardiac output.

Coronary response to large decreases of hemoglobin-O2 affinity in isolated rat heart.

In this study, the consequences of large increases of P50 (O2 partial pressure at 50% oxyhemoglobin saturation) on coronary blood flow (CBF) were investigated in isolated Wistar rat heart. Rightward shifts of the O2 dissociation curve (ODC), obtained by lysing and resealing erythrocytes to encapsulate inositol hexaphosphate (IHP), led to a very large increase in P50 without side effects. Each heart was perfused alternatively with control stored human blood [P50 = 18.8 +/- 0.3 (SE) mmHg] and IHP-treated human blood (P50 = 47.1 +/- 1.7 mmHg), according to the technique of Langendorff (mean perfusion pressure 80 mmHg; hematocrit 25%). Arterial PO2 of 180 mmHg was maintained to keep arterial O2 content identical for both types of blood. When hemoglobin affinity was lowered, CBF decreased from 5.32 +/- 0.20 to 3.40 +/- 0.14 ml X min-1 X g-1, coronary sinus PO2 (PcsO2) rose from 39.9 +/- 0.9 to 69.9 +/- 4.2 mmHg, and myocardial O2 consumption (MVO2) rose slightly from 0.125 +/- 0.005 to 0.149 +/- 0.010 ml O2 X min-1 X g-1 (P less than 0.05). A significant negative correlation was found between CBF and P50 (r = -0.90; n = 32) and a significant positive correlation between PcsO2 and P50 (r = +0.84; n = 28). The coronary blood flow response to high P50 values was not abolished when maximal dilation was induced by adenosine, so this response seems independent of metabolic needs. These experiments have demonstrated that if O2 uptake by erythrocytes remains constant, in the presence of a high P50, sufficient O2 supply may be achieved with substantially less blood flow.(ABSTRACT TRUNCATED AT 250 WORDS)