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Home , Hemoglobin

O2 Transport Linda Costanzo, Ph.D.

OBJECTIVES:

After studying this lecture, the student should understand:

1. How is carried in . 2. Features of adult and its variants. 3. Features of the oxygen-hemoglobin dissociation curve, including the sigmoidal shape and P50. 4. The meaning of right- and left-shifts of the oxygen-hemoglobin dissociation curve and what causes each. 5. The effect of on the oxygen-hemoglobin dissociation curve. 6. How to calculate the oxygen content of blood.

I. FORMS OF O2 IN BLOOD

In the blood, O2 is carried in two forms: dissolved O2 and O2 bound to hemoglobin (called O2- hemoglobin).

Dissolved O2. Dissolved O2 constitutes 2% of the total O2 content of the blood. The quantity of O2 dissolved in blood is described by Henry’s law, which states that the of dissolved gas is proportional to its ; the proportionality constant is the solubility. The solubility of O2 is 0.003 ml O2/100 ml blood/mm Hg. Thus, for an arterial PO2 of 100 mm Hg, the dissolved O2 content is 0.3 ml O2/100 ml blood. Alone, dissolved O2 is woefully inadequate for O2 delivery to the tissues; we must have O2-hemoglobin!

O2-hemoglobin. The remaining 98% of O2 in blood is O2-hemoglobin, the major topic of this lecture.

II. HEMOGLOBIN

A. Features of hemoglobin

Hemoglobin, a globular , has four subunits. Each subunit has a moiety (an -binding ) and a polypeptide chain. Adult hemoglobin (called HbA) has two α chains and two β chains, thus it is designated α2 β2. Each subunit can bind one molecule of O2; thus a hemoglobin molecule can bind a total of four O2, which is referred to as 100% saturation. In the oxygenated form, hemoglobin is called oxyhemoglobin; in the unoxygenated form, it is deoxyhemoglobin. To 2+ bind O2, iron must be in the ferrous state, Fe .

B. Variants of hemoglobin

1. Adult hemoglobin (HbA). See above. 2. (HbF). In fetal hemoglobin, HbF, the two β chains are replaced by γ chains, thus it is described as α2 γ2. This modification to the hemoglobin molecule results in a higher affinity for O2 (than HbA), which lowers the PO2 (free, dissolved O2) and facilitates movement of O2 from the maternal to the . Within the first year of life HbF is replaced by HbA. 3. . In methemoglobin, iron is in the ferric (Fe+3) state instead of the normal Fe2+ state. Methemoglobin does not 2+ bind O2. is caused by drugs that oxidize Fe to Fe+3, (e.g., sulfonamides) or congenital enzyme deficiency of methemoglobin reductase (the blood enzyme that normally keeps iron in its Fe2+ state). 4. Hemoglobin S (HbS). HbS is the hemoglobin variant that causes . In HbS, the α subunits are normal and the β A S subunits are abnormal, so it is designated α 2 β 2. The A S deoxygenated form of α 2 β 2 forms sickle-shaped rods that distort the shape of the red blood cells (i.e., “sickling”) and causes them to A S occlude small blood vessels. α 2 β 2 has a lower affinity for O2 than does normal HbA.

III. O2-HEMOGLOBIN DISSOCIATION CURVE

Figure 1. O2 binds reversibly to the heme groups on hemoglobin. Therefore, each hemoglobin molecule has the capacity to bind four of O2. The percent saturation (“O2 sats”) tells what percent of heme groups are bound to O2. When all four are bound to O2, there is 100% saturation, when three hemes are bound to O2, there is 75% saturation, etc. The O2-hemoglobin dissociation curve shows the relationship between % saturation and PO2 of the blood. This is one of the most famous and most important curves in all of ! Live it, love it!

For convenience, the table below gives various values of PO2 and the corresponding % saturation for the normal O2-hemoglobin dissociation curve.

PO2 % Saturation 10 25% 20 35% 25 50% (P50) 30 60% 40 75% (mixed venous blood) 50 85% 60 90% 80 96% 100 98% (≈ 100% )

A. Sigmoidal shape

The O2-hemoglobin dissociation curve has a sigmoidal shape. Thus, rather than a linear relationship between % saturation and PO2, the % saturation increases steeply between a PO2 of zero and 40 mm Hg, then increases less steeply between 40 and 60 mm Hg, and then is nearly flat between 60 and 100 mm Hg. (Note that one of the implications of the nearly “flat” portion of the curve between 60 and 100 mm Hg is that changes in arterial PO2 in this range have little effect on the % saturation of hemoglobin and therefore the amount of O2 carried in the blood.)

The sigmoidal shape results from positive of O2 binding to hemoglobin. As each successive O2 binds, it increases the affinity for the next O2. Binding of the first O2 increases the affinity for the second O2, etc. Affinity for the fourth (last) O2 is the highest, which corresponds to the portion of the curve where % saturation is near or at 100%.

B. P50

By definition, P50 is the PO2 that corresponds to 50% saturation. Changes in P50 reflect changes in the affinity of hemoglobin for O2. Increases in P50 reflect decreased affinity. Decreases in P50 reflect increased affinity.

C. Right- and left-shifts of the O2-hemoglobin dissociation curve

Figure 2.

Changes in affinity of hemoglobin for O2 produce changes in the P50 and shift the O2-hemoglobin dissociation curve to the right or left.

Right-shifts of the O2-hemoglobin dissociation curve occur when there is a decreased affinity of hemoglobin for O2, which produces an increase in P50. When affinity is decreased, unloading of O2 is facilitated. The factors that cause a right-shift make sense in terms of this decreased affinity and include:

1. Increases in PCO2 and decreases in pH, such as those occurring when there is increased metabolic activity in a tissue (e.g., during exercise. Called the . 2. Increases in temperature, such as during exercise. 3. Increases in 2,3-diphosphoglycerate (2,3-DPG). 2,3 DPG binds to the β chains of deoxyhemoglobin and reduces their affinity for O2. During (such as at high altitude), 2,3 DPG production in red cells increases, causing a helpful decrease in the affinity of hemoglobin for O2 (facilitates O2 unloading in tissues).

Left-shifts of the O2-hemoglobin dissociation curve occur when there is increased affinity of hemoglobin for O2, which produces a decrease in P50. When affinity is increased, unloading of O2 is more difficult. The factors causing a left-shift are:

1. Decreases in PCO2 and increases in pH, such as when there is decreased metabolic activity in a tissue. 2. Decreases in temperature. 3. Decreases in 2,3-DPG. 4. Hemoglobin F (HbF). The mechanism of the left-shift with HbF relates to 2,3 DPG, which binds less avidly to the γ chains of HbF than to the β chains of HbA. With less 2,3 DPG bound to HbF, the O2 affinity increases, lowering the PO2 of the and facilitating O2 diffusion from mother to fetus. 5. Carbon monoxide (CO) not only causes a left-shift, but also a decrease in O2-binding capacity (see below).

D. CO poisoning

Figure 3.

Carbon monoxide (CO) binds to hemoglobin (to form ) with an incredibly high affinity, 250 times that of O2! Any heme groups bound to CO cannot bind to O2, so CO poisoning decreases the O2-binding capacity of hemoglobin. In the figure, for illustration, the O2-binding capacity of hemoglobin was reduced to 50% of normal, meaning ½ of the heme sites were occupied by CO. CO also increases the affinity of hemoglobin for whatever O2 is bound (a left-shift of the O2-hemoglobin dissociation curve). Thus, the effects of CO poisoning are devastating for O2 delivery to tissues: less O2 is bound and the O2 that is bound is less readily released.

E. O2 Content of Blood

Blood flow and O2 content of blood are the major factors determining O2 delivery to tissues. O2 content of blood is comprised of dissolved O2 and O2-hemoglobin.

1. Dissolved O2 is easy — it is described by Henry’s law as the PO2 times the solubility of O2 in blood.

Dissolved O2= PO2 x solubility P x 0.003 ml O /100 ml = O2 2 blood/mm Hg

Thus, at the normal arterial PO2 of 100 mm Hg:

100 mm Hg x 0.003 ml O /100 ml Dissolved O = 2 2 blood/mm Hg = 0.3 ml O2/100 ml blood, or 0.3 vol %

2. O2 bound to hemoglobin depends upon three things:

a. Hemoglobin concentration. The normal value is 15 g/100 ml of blood. b. O2-binding capacity. The maximal amount of O2 that can be bound to hemoglobin when there is 100% saturation of heme sites. The O2-binding capacity is normally 1.34 ml O2/g hemoglobin. c. % Saturation. % of heme groups bound to O2. Varies from zero to 100%

Thus, if the hemoglobin concentration of blood is 15 g/100 ml, the O2- binding capacity of that hemoglobin is 1.34 ml O2/g, and the % saturation of arterial blood is 100% (at an arterial PO2 of 100 mm Hg):

O2-hemoglobin = 15 g/100 ml x 1.34 ml O2/g x 100% 20.1 ml O /100 ml blood, or 20.1 vol = 2 %

If the % saturation of hemoglobin is less than 100% then, accordingly, there will be less O2-hemoglobin.

3. Now, to calculate the total O2 content of blood at a PO2 of 100 mm Hg.

O2 content of blood = O2-hemoglobin + dissolved O2 20.1 ml O /100 ml + 0.3 ml O /100 = 2 2 ml = 20.4 ml O2/100 ml, or 20.4 vol%

**Special note: ml gas/100 ml is also called “volume %.” Thus, an O2 content of 20.4 ml O2/100 ml blood is called “20.4 volume %” for short.

4. Summary of O2 transport

Figure 4.

Humidified tracheal air has a PO2 of 150 mm Hg. Alveolar air has a lower PO2 of 100 mm Hg because O2 has diffused from alveolar gas into pulmonary blood. Pulmonary capillary blood, which becomes systemic arterial blood, equilibrates with alveolar gas, so it too has a PO2 of 100 mm Hg. The PaO2 of 100 mm Hg corresponds to 100% saturation of hemoglobin on the O2-hemoglobin dissociation curve. The O2 content of systemic arterial blood is the sum of dissolved O2 and O2-hemoglobin per our discussion above. Dissolved O2 was 0.3 vol% and O2-hemoglobin was 20.1 vol% for a grand total of 20.4 vol% in systemic arterial blood. In the tissues, O2 diffuses from the to the tissues for aerobic . Thus mixed venous blood has as lower PO2 of 40 mm Hg, a correspondingly lower % saturation of 75% (read it off the O2-hemoglobin curve!), and a correspondingly lower O2 content of 15 vol %. Thus 5 vol % of O2 must have been transferred to the tissues. Mixed venous blood will be re-loaded with O2 in the next pass through the .

IV. PRACTICE QUESTIONS

1. A person who is hypoxemic has an arterial PO2 of 75 mm Hg and a venous PO2 of 30 mm Hg. This person has a normal hemoglobin concentration and a normal O2-binding capacity. Hemoglobin is 85% saturated at 75 mm Hg and 55% saturated at 30 mm Hg. For arterial and venous blood, calculate dissolved O2, O2-hemoglobin, and total O2 content. What was O2 consumption by the tissues (in vol %)?

2. What is the effect of increased PCO2, increased pH, and CO on O2-binding capacity, P50, and affinity of hemoglobin for O2?

3. A right-shift of the O2-hemoglobin curve is:

A. Associated with a decrease in P50. B. Associated with an increase in O2 content of blood at a PO2 of 50 mm Hg. C. Caused by an increase in blood pH. D. Caused by an increase in 2,3 DPG concentration.

4. Which of the following causes a decrease in O2-binding capacity of hemoglobin?

A. Decreased hemoglobin concentration B. CO poisoning C. Decrease in arterial PO2 to 60 mm Hg D. Increase in arterial PO2 to 120 mm Hg E. Left-shift of the O2-hemoglobin curve

EXPLANATIONS

1.

2.

3. Answer = D. Start by drawing a normal O2-Hb curve and a right-shifted curve. Write down what you know: associated with increased P50, decreased affinity, and decreased O2 content at a given PO2; caused by increased PCO2, decreased pH, and increased 2,3 DPG. I hope you see why choice B is incorrect – at a PO2 of 50 mm Hg, the right-shifted curve has a lower % saturation, thus a lower O2 content. BTW: “associated with” means any association, cause or effect.

4. Answer = B. Get oriented! What is O2-binding capacity of hemoglobin? It is how many ml of O2 each gram of hemoglobin can hold at 100% saturation. Decreased hemoglobin concentration does not affect the amount of O2 each gram of hemoglobin can hold. Decreasing or increasing PO2 changes the % saturation of hemoglobin, but does not affect the amount of O2 that hemoglobin can hold at 100% saturation. Same with a left-shift of the curve; it increases the % saturation at a given PO2 and increases affinity, but does not alter the maximum amount of O2 that hemoglobin can hold. CO attaches to binding sites on hemoglobin and prevents binding of O2, which reduces the O2-binding capacity of hemoglobin. I suspect a few people learned something from this question!