REVIEW ARTICLE

Oxygen transport in blood at high altitude: role of the hemoglobin–oxygenaffinity and impact of the phenomena related to hemoglobinallosterism and red cell function

Accepted: 10 April 2003 / Published online: 18 September 2003� Springer-Verlag 2003

Abstract Altitude hypoxia is a major challenge to theblood O2 transport system, and adjustments of theblood–O2 affinity might contribute significantly to hy-poxia adaptation. In principle, lowering the blood–O2

affinity is advantageous because it lowers the circulatoryload required to assure adequate tissue oxygenation upto a threshold corresponding to about 5,000 m altitude,whereas at higher altitudes an increased blood–O2

affinity appears more advantageous. However, the ra-ther contradictory experimental evidence raises thequestion whether other factors superimpose on theapparent changes of the blood–O2 affinity. The mostimportant of these are as follows: (1) absolute temper-ature and temperature gradients within the body; (2) theintracapillary Bohr effect; (3) the red cell populationheterogeneity in terms of O2 affinity; (4) control ofaltitude alkalosis; (5) the possible role of hemoglobin asa carrier of the vasodilator nitric oxide; (6) the effect ofvaried red cell transit times through the capillaries.

Keywords Altitude hypoxia Æ Oxygen equilibriumcurve Æ Hemoglobin allosterism Æ Red cell function

Introduction

The blood–O2 equilibrium curve (OEC), i.e., the partialpressure of O2 (PO2) versus hemoglobin (Hb) O2 satu-

ration (SO2) plot, and the P50, i.e., the PO2 at whichSO2=50%, are useful indexes of the blood–O2 affinity.A variety of factors, the most important of which arepH, partial pressure of CO2 (PCO2), the red blood cell(RBC) concentration of 2,3-diphosphoglyceric acid(DPG), temperature and the amount of Hb bound tocarbon monoxide, influence OEC and hence the P50

(Severinghaus 1966; Siggaard-Andersen 1971; Thomas1972; Severinghaus 1979; Winslow et al. 1983; Rovidaet al. 1984; Siggaard-Andersen et al. 1984). Exposure toaltitude is well known to decrease the blood–O2 affinity(P50 rises by 2–5 mmHg, depending on altitude andexposure duration) by increasing the blood DPG con-tent (Lenfant et al. 1971), but Mg2+, ATP and Cl) alsoplay important roles (Mairbaurl et al. 1993).

Under normal conditions, the sigmoidicity of theOEC facilitates the blood O2 transport as virtually allHb is loaded with O2 in the lungs at a relatively lowarterial PO2 (PaO2), while O2 is unloaded in the tissuesat a relatively high venous PO2 (PvO2). Thus, the re-quired amount of O2 is carried at a relatively low flow,thereby optimizing the cardiac output and the circula-tory load. In theory, changes in the blood–O2 affinityshould have important effects on the blood O2 transport.In the lungs, where PaO2 is set by the ventilation rate,the changes in P50 have little effect on arterial O2 satu-ration (SaO2) due to the flatness of the OEC in its upperportion. In contrast, PvO2 and venous O2 saturation(SvO2) are significantly affected (Fig. 1). In healthy hu-mans at sea level, the increased blood–O2 affinity wouldadversely influence the circulation, whereas the reverse isexpected in the case of decreased blood–O2 affinity.Table 1 reports an attempt to quantify the circulatory‘‘advantage’’ of increased P50 under normal conditionsat sea level.

The above considerations were made considering thewhole body as a lumped compartment, with the sub-script ‘‘v’’ denoting the mixed venous return. However,the same change in the blood–O2 affinity is expected tohave different effects in various organs, depending ontheir O2 requirement and the blood flow through them

Eur J Appl Physiol (2003) 90: 351–359DOI 10.1007/s00421-003-0954-8

Michele Samaja Æ Tiziano Crespi Æ Marco Guazzi

Kim D. Vandegriff

M. Samaja (&) Æ T. Crespi Æ M. GuazziDipartimento di Medicina,Chirurgia e Odontoiatria, Ospedale San Paolo,Universita’ Di Milano, via di Rudini’ 8,20142 Milan, ItalyE-mail: Michele.Samaja@unimi.itTel.: +39-028-9179250Fax: +39-028-9179251

K. D. VandegriffSangart, Inc., San Diego,California, USA

(Samaja 1988). In organs with high O2 requirement, suchas the heart and the working muscles, the high O2

extraction diminishes the dependence of that organ on

the changes in the blood–O2 affinity. In an extreme case,should an organ be able to extract all O2 from blood, thechanges in the blood–O2 affinity should have no effect atall on that organ. In contrast, in organs with low O2

extraction, such as the kidney, the changes in the blood–O2 affinity should have important effects. However, forthe sake of clarity, this manuscript concerns the wholebody as a single compartment.

Physiological effects of changes in the blood–O2 affinity

Theory

Exposure to altitude and arterial hypoxemia is a majorchallenge to the blood O2 transport system, and changesin blood–O2 affinity might contribute significantly tohypoxia adaptation. As yet there is no agreement on thequestion of whether a decreased or an increased blood–O2 affinity is preferable. Results from animals adaptedto live at altitude, the great majority of which haveblood P50 values lower than their sea level counterparts,suggest that an increased blood O2 affinity is advanta-geous (Monge and Leon-Velarde 1991). However, withthe exception of early reports, probably compromised bylack of knowledge of thermolabile DPG as an importantcofactor of the Hb–O2 affinity, all subsequent studies inhumans invariably report decreased blood–O2 affinitydue to increased DPG. Thus the question arises ofwhether this represents an adaptive or a maladaptivepattern. A theoretical study (Samaja et al. 1986) sug-gested that decreased blood–O2 affinity is preferable upto 5,500 m altitude, because decreased SvO2 tends to

Table 1 Advantage led by aP50 [i.e., the oxygen pressure (PO2) atwhich oxygen saturation (SO2)=50%] increase of 1 mmHg. Twoblood–O2 equilibrium curves (OEC) are calculated (Winslow et al.1983) varying the pH to simulate the unit change in the P50. In theiso-SO2 model [i.e. decreasing venous oxygen pressure (PvO2) atconstant venous oxygen saturation (SvO2)] (see Fig. 1), thePO2 iscalculated at the indicated arterial oxygen saturation (SaO2) andSvO2 values that were chosen arbitrarily, and the change in arterialto venous oxygen pressure (Da–vPO2) is then calculated. Loweringthe blood–O2 affinity by 1 mmHg P50 increases Da–vPO2 by3.2 mmHg, thereby representing an advantage for the perfusedtissue. In the (iso-PO2) model (i.e. increasingSvO2 at constantPvO2), SO2 is calculated at the indicatedPaO2 and PvO2 values that

were chosen arbitrarily, and the change in arterial to venous oxygensaturation (Da–vSO2) is then calculated. In this case, the unit changein the P50 increases Da-vSO2 by 1.4%. To estimate the physiologicaladvantage of this change, the Fick’s equation is used in the formQ= _VV O2/{[Hb](SaO2)SvO2)+a(PaO2)PvO2)}, where Q is the car-diac output (liters per minute), _VV O2 is the O2consumption(0.25 l min)1 at rest), [Hb] is the Hb concentration (160 g l)1), a isthe O2 solubility in plasma (1.4·10)6 mol l)1 at 37�C; Roughtonand Severinghaus 1973), and the other values are from the table.The cardiac output needed to sustain the assumed _VV O2 decreasesby 5.8% for unitP50 decrease. DPG 2,3-Diphosphoglyceric acid;Hbhemoglobin

OEC ‘‘left’’ OEC ‘‘right’’

High O2 affinity Low O2 affinity

Constant parameters pH 7.445 7.400PCO2, mmHg 40 40DPG Hb)1, mol mol)1 0.8 0.8P50, mmHg 27.8 28.8

Iso-SO2 model PO2 at SaO2=95%, mmHg (PaO2) 94.8 99.5PO2 at SvO2=70%, mmHg (PvO2) 39.2 40.7Da–vPO2, mmHg 55.6 58.8

Iso-PO2 model SO2 at PaO2=95 mmHg, %(SaO2) 95.0 94.5SO2 at PvO2=40 mmHg, %(SvO2) 71.0 69.1Da–vSO2, % 24.0 25.4Cardiac output, l min)1 6.51 6.15

Fig. 1 Increasing the blood–O2 affinity [i.e., shifting from thecontinuous curve on the right to the dashed curve on the left withdecreased P50, i.e., the oxygen pressure (PO2) at which oxygensaturation (SO2)=50%] has little effect on arterial oxygensaturation (SaO2) [not shown in the figure, arterial oxygen pressure(PaO2) is assumed constant because it is primarily controlled by theventilatory rate rather than by the blood–O2 affinity]. In the venouscompartment, the same increase in the blood–O2 affinity shouldhave one of the following effects: (1) decreasing venous oxygenpressure (PvO2) at constant venous oxygen saturation (SvO2) (iso-SO2 model); (2) increasing SvO2 at constant PvO2 (iso-PO2 model);(3) a combination of both. Thus, increasing the blood–O2 affinityadversely influences the circulation, because: (1) in the iso-SO2

model, tissues are forced to operate at a lower PO2; (2) in the iso-PO2 model, a higher cardiac output is required to compensate forthe decreased the change in arterial to venous oxygen saturation;(3) a combination of both. Table 1 reports a quantification of theseeffects

352

maximize the change in arterial to venous oxygen satu-ration (Da–vSO2), thereby lowering the circulatory load.In contrast, above this altitude, increased blood–O2

affinity is preferable because the preservation of SaO2

becomes a priority.

Experimental studies

Table 2 summarizes the essential results from some ofthe studies aimed at demonstrating the physiologicaladvantage of changes in the blood–O2 affinity in vivo orex vivo. Despite the majority of them being fundamen-tally in agreement with the theoretical predictions, con-siderable controversy remains. It is therefore legitimateto address the question whether such conflicting exper-imental evidence can be at least in part explained byovercoming the phenomena that tend to mask the effectof the measured changes in the blood–O2 affinity. Ouraim here is to assess some of these phenomena, namelythose that are linked to the blood–O2 affinity, but whichhave not yet been adequately measured or assessedunder hypoxic conditions.

Phenomena linked to Hb allosterism and RBC functionthat might mask the measured changes in theblood–O2 affinity

Temperature

Most often, the blood–O2 affinity is measured at thestandard 37�C temperature, but temperature has beenlong known to be an important allosteric factor of Hbfunction (Barcroft and King 1909). From the averageslope of the log10P50 versus temperature relationship(0.023 mmHg �C)1) (Samaja et al. 1983), P50 is expectedto decrease by 1.5 mmHg �C)1 temperature decrease.There are two aspects of this issue. First, central tem-perature should be taken into account when character-izing the blood–O2 affinity at altitude, but few studies, ifany, report this parameter. Temperature control at alti-tude is subject to many variables, but it is likely thathypothermia is more frequent at altitude than at sea leveland may therefore be one of the factors that mask themeasured changes in the blood–O2 affinity. Second, it islikely that considerable temperature gradients occur inthe body, especially at altitude with high blood hemat-ocrit and impaired circulation through some peripheralareas. Thus, the blood–O2 affinity characteristics may bequite different in the lungs and in some peripheral areasof the body. For example, for a ‘‘central’’ P50 value of28 mmHg at 37�C, the local P50 values in activelyworking muscles and upper joints may be 36.5 mmHgand 8.7 mmHg if temperature in these areas shifts to42�C and 15�C, respectively. Such changes are muchwider than those usually measured in humans at altitude,but we are not aware of studies reporting the existence ofsuch temperature gradients at altitude.

Hydrostatic pressure

Hydrostatic pressure decreases the P50 by 0.033 mmHgatm)1 without either altering the RBC DPG content oraffecting the Donnan equilibrium (Reeves and Morin1986). These changes, however, appear too modest tocompete with those attributed to altitude exposure.

Bohr effect

The equilibrium of the reaction HHb+O2��H-bO2+H+ shifts toward the formation of deoxygenatedHb on increasing acidity of the medium, the well knownBohr effect. Whereas arterial pH (pHa) normally fluc-tuates in a narrow range, the end-capillary pH is sub-jected to wide changes as a function of the metabolicstate and changing release of lactic acid. Thus, theblood–O2 affinity is expected to decrease progressivelyalong the capillary, thereby increasing the O2 delivery totissue in proportion to the tissue metabolic needs. Thismechanism may thus act as a signal originated from thetissue and targeted to the RBC to modulate O2 delivery(Wasserman et al. 1991). At sea level, the acidification ofmuscle capillary blood due to lactic acidosis was shownto facilitate the O2 transport during exercise (Stringeret al. 1994). We are not aware of similar studies per-formed at altitude, but class C heart failure patients mayrepresent a reliable model for hypoxia (Perego et al.1996). It was shown that, of the three mechanisms thatcan enhance the O2 delivery to working muscles, i.e.,increasing PaO2, increasing [Hb] and increasing P50 dueto the intracapillary Bohr effect, the last is a majorcontributor in class C patients (Agostoni et al. 1996).Thus, enhanced HbO2 dissociation, secondary to lactateproduction and acidification of capillary blood, helps tooxygenate the tissue above the anaerobic threshold. Ataltitude, however, this model is more complicated be-cause natives and lowlanders generate lower-than-ex-pected amounts of lactate on exercise, the so-called‘‘lactate paradox’’ (Grassi et al. 1996). In principle, thisshould diminish the relevance of the Bohr effect at alti-tude, but we are not aware of capillary blood pH mea-surements in hypobaric hypoxia, either direct orindirect, i.e., femoral vein pH, which is often used as anindex of end-capillary pH in working muscles duringexercise (Stringer et al. 1994). Thus the doubt remainswhether the expected decreased blood buffer power ataltitude, because of lower bicarbonate levels, may partlycompensate for the lower lactate output in maintainingthe physiological significance of lactic acid as a majorcontributory factor for O2 delivery to tissues.

Blood as a non-homogeneous population of RBCs

Lack of nuclei and mitochondria in the mature RBCrenders this cell unable to perform certain functions thatnormally occur in most aerobic cells, including protein

353

Table

2Summary

ofstudiesreportingthephysiologicaladvantageordisadvantageofchanges

intheblood–O

2affinity()

negative;

+positive;

=none).RBC

Red

bloodcell;Cr

creatine;

PCrphosphocreatine;

IHPinositolhexaphosphate;NaOCN

sodium

cyanate

Reference

P50change(m

mHg),methodused

Main

measurement

Main

conclusion,andeff

ectofincreasedHb–O

2affinity

Woodsonet

al.1973

36to

23,Exchangetransfusionwith

DPG-depletedblood

Work

perform

ance

inrats

Perform

ance

decreases,

)

Wranneet

al.1974

26to

24,AcidosisandDPG

depletion

Work

capacity

inman

Noeff

ect,=

Yhapet

al.1975

26to

20,DPG

depletion

_ VVO

2in

isolatedhindlimb

_ VVO

2decreasesdueto

lowered

O2extraction,)

Bakker

etal.1976

25to

18,DPG

depletion

_ VVO

2andbileflow

inratliver

atvaryingperfusionrates

Bileflow

rate

decreasesin

norm

oxia

andhypoxia._ VVO

2

decreasesin

hypoxia,butnotin

norm

oxia,)/=

Bakker

etal.1976,1977

25to

18,DPG

depletion

_ VVO

2andbileflow

inratliver

atvaryinghem

atocrits

Nochange,

butPVO

2islower

at2%

hem

atocrit,+

Tureket

al.1978a

33to

22,NaOCN

treatm

ent

PVO

2and

D a–vSO

2O

2transport

impaired

innorm

oxia,more

efficientin

hypoxia,+

Tureket

al.1978b

36to

22,NaOCN

treatm

ent

BloodO

2content,cardiacoutputand

flow

toorgansin

hypoxia

Noeff

ectin

hearts,highO

2affinitydisadvantageousfor

other

organsin

mildhypoxia,butadvantageous

inseverehypoxia,+

/=Martin

etal.1979

32to

14,Carbonmonoxide

_ VVO

2in

workingrathearts

Nochangedueto

increasedcoronary

flow

andcapillary

density,=

Malm

berget

al.1979

42to

21,StorageandNaOCN

Compensationto

hem

orrhagic

shock

inrats

Lim

ited

O2deliveryduringhypotension,)

Ross

andHlastala

1981

30to

14,StorageandNaOCN

O2extractionin

isolatedskeletalmuscle

_ VVO

2decreasesdueto

factors

unrelatedto

RBC–O

2

affinity,=

WoodsonandAuerbach

1982

38to

17,Exchangetransfusionwith

DPG-depletedblood

Cardiacoutputandflow

distributionin

sedatedrats

Nochangein

cardiacoutput,increasedcoronary

and

cerebralbloodflow,)

Lister1984

23to

31,Exchangetransfusion

_ VVO

2andO

2transport

inthehypoxic

lamb

Improved

cardiacfunction,)

Apsteinet

al.1985

33to

17,Storage

Cardiacfunctionin

isolatedrabbithearts

Decreasedperform

ance

and

_ VVO

2,)

Stucker

etal.1985

19to

47,IH

Ploading

_ VVO

2in

isolatedrathearts

Increased

_ VVO

2,)

Teisserie

etal.1985

31to

41,IH

Ploading

Bloodgasses

inexchange–tranfusedpiglets

IncreasedPaO

2and

D a–vSO

2,decreasedcardiacoutput,

)Koehleret

al.1986

26to

37,Exchangetransfusionwith

heterologouslow-O

2affinityblood

Cerebralbloodflow

response

tohypoxia

inlambsin

vivo

Reducedcerebralbloodflow

andO

2transport

tobrain,+

Rosenberget

al.1986

17to

32,Exchangetransfusionwith

heterologouslow-O

2affinityblood

Cerebralbloodflow

andbloodgasses

inlambsin

vivo

Noeff

ectontheabilityto

maintain

theO

2transport

duringhypoxia,=

Allanet

al.1986

16to

8,Treatm

entwithBW12C

Functionofisolatedrabbithearts

Decreasedfunction,PvO

2andincreasedlactate

output,

)Baronet

al.1987

21to

43,IH

Ploading

CardiacfunctionandO

2extraction

inperfusedrabbithearts

Increased

_ VVO

2,withincreasedvascularresistance

dueto

IHP,)

Woodsonet

al.1987

32to

22,Bloodmanipulation

Isolatedperfusedrathearts

DecreasedO

2extraction,increasedcoronary

bloodflow,)

Teisserie

etal.1987

26to

80,IH

Ploading

Long-term

effects

oncardiacoutputin

pigs

Increased

D a–vSO

2,)

GutierrezandAndry

1989

31to

21,NaOCN

O2extractionandPCr/Crratioin

hypoxic

hindlimbs

Noeff

ectontissueoxygenationdueto

peripheral

limitationto

O2diffusion,=

Samaja

etal.1991

30to

15,Storage

Cardiacfunctionin

isolatedrathearts

exposedto

low-flow

ischem

iaPerform

ance

insensitiveto

P50changes,=

Khandelwalet

al.1993

Allosteric

modificationofHb

TissueP

O2

IncreasedtissuePO

2,)

Liard

andKunert1993

31to

39,IH

Ploading

Hem

odynamicsin

dogs

DecreasedcardiacoutputandincreasedO

2delivery,)

Kunertet

al.1996

36to

52,allosteric

modification

ofHb

TissueP

O2andflow

inratcrem

aster

muscle

IncreasedO

2deliveryandresistance,=

Curtiset

al.1997

30to

42,allosteric

modification

ofHb

TissueP

O2andO

2extractionin

canineskeletalmuscle

Noeff

ectonO

2extraction,increasedtissueP

O2,=

Weiss

etal.1999

Allosteric

modificationofHb

Myocardialhigh-energyphosphatesduring

low-flow

ischem

iain

dogs

Ameliorationofmetabolicandfunctionalconsequences

ofischem

ia,)

Berlinet

al.2002

17to

33,IH

Ploading

Cardiacfunctionin

heartsduringlow-flow

ischem

iaDecreasedfunction,)

354

synthesis and energy generation from processes requir-ing O2. The inability to repair damage leads to unbal-anced metabolism, altered ion homeostasis and a limited120-day life span. In humans, the RBC removal from thecirculation by the reticuloendothelial system is selectivefor the aging RBC (Clark 1988) and is characterized byreduced deformability (Linderkamp and Meiselman1982) and DPG content (Seaman et al. 1980). Inactiva-tion of membrane Na+–K+-ATPase leads to shrinkingand >20% increase in RBC [Hb]. The combined effectof increased [Hb] and decreased [DPG] reduces the[DPG]/[Hb] ratio from 1.0 mol mol)1 in the ‘‘young’’RBC to 0.5 mol mol)1 in the ‘‘old’’ RBC, which corre-sponds to a decrease in P50 from 29 mmHg to 23 mmHg(Samaja and Rovida 1987). The various types of RBCcoexist in blood, and despite differences in rheologicalproperties (Linderkamp and Meiselman 1982), there isno evidence that capillaries select the RBC passagebased on their age. Therefore, capillaries are perfusedsequentially with RBCs that vary widely in their O2

affinity properties. One may wonder whether the chan-ges in whole blood P50 by 2–5 mmHg, as those observedin humans at altitude, still retain biological significancewhen wider changes are already routinely present at themicrocirculatory level.

The majority of the techniques used for determiningthe blood–O2 affinity make use of blood or RBC sampleswith volumes in the microliter-to-milliliter range.Therefore, the measured values represent the average ofthe various RBC types in that sample. It is likely that theinter-individual P50 or OEC variability stems from al-tered distribution in ‘‘young’’ and ‘‘old’’ RBCs ratherthan from intrinsic changes in the Hb–O2 affinity. TheRBC distribution is profoundly altered in several blooddisorders (Nakashima et al. 1973). At altitude, the wholeRBC population undergoes rejuvenation, because in-creased erythropoiesis inputs more ‘‘young’’ RBCs intothe blood (Samaja et al. 1993), but we are not aware ofstudies reporting modifications of the RBC life span ataltitude, making it difficult to assess whether the RBCheterogeneity increases or decreases at altitude.

High-altitude alkalosis

At altitude, hypoxic stimulation of chemoreceptors in-creases alveolar ventilation, thereby increasing CO2

washout, reducing PaCO2 and increasing pHa. Althoughat sea level pHa is allowed to fluctuate within a rathernarrow 7.37–7.43 range, at altitude the mechanisms thattitrate pHa to the normal value are less efficient.Therefore, the blood remains alkaline, perhaps becauseof the slow renal compensation that cannot cope withcontinuous CO2 washout from the lungs. After 7–10 days at 6,450 m, pHa remains at 7.496 (0.006) andbase excess (BE) is )6 mEq l)1, whereas a BE of )10 to)9 mEq l)1 is necessary for complete compensation(Winslow et al. 1984; Samaja et al. 1997). Alkalosis in-creases the blood–O2 affinity, an effect contrasted by

increased DPG. As a result, in Caucasians at 6,300 m,and despite an increased [DPG]/[Hb] ratio from 0.80 to1.36 (Samaja et al. 1997), P50 increased from 28 mmHgto only 30 mmHg. At this altitude, this might be con-sidered as a protective mechanism. At 6,450 m, hyper-ventilatory alkalosis increases SaO2 to an extent thatcorresponds to a descent by 1,300 m (Samaja et al.1997), and at 8,848 m, the extreme alkalosis in a personon the summit of Mount Everest should have allowedthis subject to maintain essentially the same SaO2 as thatmeasured at 6,300 m (Winslow et al. 1984). Therefore, atextreme altitudes alkalosis helps maintaining Da–vSO2

despite low PaO2.Prolonged alkalosis, however, is not compatible with

normal body function and one of the effects of accli-matization is to reduce alkalosis, with subjects withacute mountain sickness (AMS) being more alkaloticthan non-AMS subjects (Samaja et al. 1997). As amatter of fact, acetazolamide, a drug used to fight AMS,tends to lower pHa by inhibiting carbonic anhydrase(Bradwell et al. 1987). Comparative analysis of the pH–arterial pressure of CO2 (PaCO2) relationships in othergroups of subjects helps to address this issue. In Sherpas,PaCO2 is higher, and their pHa is lower than in Cauca-sians (Samaja et al. 1997). As BE is comparable, it ispossible that less hypoxic ventilatory drive in Sherpas(Lahiri et al. 1969) prevents excessive PaCO2 fall and riseof alkalosis. Probably, low DPG in Sherpas’ bloodcontributes to a larger extent to keep high SaO2 thanhyperventilation does in Caucasians.

Nitric oxide

Nitric oxide (NO) is an endothelium-derived relaxingfactor with a number of effects in somatic cells as astimulator of soluble guanylate cyclase. The local con-centration of NO produced by endothelial cells(�400 nM) is in excess of that required to activate sol-uble guanylate cyclase (20 nM), but the reaction of NOwith HbO2 is so fast (k=107 M)1 s)1) that virtually allNO should be inactivated by HbO2 (Hobbs et al. 2002).Yet, it is unquestionable that NO mediates a largenumber of effects in the circulation. The apparent par-adox that endothelial cells release NO in a medium, theblood, which is particularly rich in HbO2, a NO inacti-vator, led to the conjecture that NO has a role in thehuman respiratory cycle (McMahon et al. 2002).

The hypothesis that Hb is a transporter of NO arosefrom the observation that NO does not only bind to theheme to form nitrosyl-Hb (HbNO), but also to the thiolgroup at the highly conserved b-cysteine 93 (b-Cys93)residue to form the S-nitrosothiol derivative (HbSNO)(Jia et al. 1996). Total bound NO is essentially constantin oxygenated and deoxygenated blood, but when bloodis in the oxygenated state, HbSNO is predominant,whereas HbNO is higher in the deoxygenated state(McMahon et al. 2000). Thus, Hb binds both O2 andNO in the lungs at two different sites, and in the low PO2

355

environment in the capillaries, some NO bound tob-Cys93 is made free (McMahon et al. 2000) (Fig. 2).The presence of thiol acceptors, one of which is theanion-exchanger-1 ( or band-3 protein) in the RBCmembrane (Pawloski et al. 2001), may mediate NOexport out of the RBC, thereby promoting localvasorelaxation and improving O2 delivery upon Hbdeoxygenation (McMahon et al. 2002). Althoughfavored by low tissue PO2, it is difficult to assess whetherthe Hb-mediated trans-nitrosylation may overcome theblood–O2 affinity changes at altitude. But the depen-dence of this phenomenon on the allosteric state of Hbsuggests that the NO-donor activity of HbSNO mayenhance the O2 delivery properties of blood in a fashiondependent on local hypoxia (Pawloski et al. 2001).

Although suggestive, this hypothesis requires confir-mation. First, a recent study was unable to confirm theallosteric effect underlying NO release by HbSNO, andshowed dominance of the Hb scavenging over the NO-donor properties due to the high NO–heme affinity thatprecludes reaction of NO with b-cys93 (Deem et al.2001). Second, the low P50 of purified HbSNO(<10 mmHg) should require very severe hypoxia toaccomplish NO release (McMahon et al. 2000). Third,the decrease in P50 upon in vitro incubation with NOand NO donors correlates with methemoglobin forma-tion, suggesting that this species, and not HbSNO, is thefinal product of the NO reaction with Hb (Hrinczenkoet al. 2000). Finally, it is likely that the Hb function as aNO transporter is linked to the effects of hypoxia on NOproduction, but it is not yet clear whether NO produc-tion increases or decreases at altitude. Whereas somedata show that hypoxia increases NO production

(Nathan 1992; Agvald et al. 2002), others suggest thatthe reverse is true (Nelin et al. 1996; Fike et al. 1998;Guzel et al. 2000). Also, although NO production rate iselevated during exercise, exhaled NO is not related todecreased SaO2 during heavy exercise (Sheel et al. 2000).

RBC transit time

The RBC might not have enough time in the capillary torelease all O2. To evaluate this possibility, the Fick dif-fusion equation has been used in a simple boundaryanalysis to estimate the external resistance to O2 diffu-sion for RBCs (Vandegriff and Olson 1984a, b,c). TheO2 flux at the RBC interface can be calculated using asimple diffusion equation for PO2 values across a dif-fusion barrier (P1–P2):

d O2½ �=dt � D P1 � P2ð Þ=Dr ð1Þ

where D is the O2 diffusion constant between phases P1

and P2, and Dr is the diffusion distance between P1 andP2.

Using this analysis, we can evaluate the effects on O2

transport in the capillaries of the lung, assuming con-stant Dr. The rate of O2 uptake by the RBCs at sea levelis determined by the PO2 in alveolar gas spaces (i.e.,alveolar PO2�100 mmHg = P1 in Eq. 1), which makesthe O2 concentration gradient very steep. The time toreach equilibrium between the gas space and PaO2 isvery fast (Fig. 3), and the O2 binding affinity of RBCshemoglobin (i.e., P2 in Eq. 1) is not a critical parameter.This does not hold true at high altitude, where alveolarO2 is much lower. As alveolar PO2 decreases, the dif-fusion gradient becomes narrower as a function of thepulmonary O2 concentration (i.e., decreasing P1 inEq. 1) (Fig. 3). In addition, if the RBC has a high P50

(P2 in Eq. 1), the gradient will be narrowed even further(flP1)›P2). Thus, the hemoglobin O2 binding charac-teristics will be a critical factor at high altitude (Van-degriff and Winslow 1995).

Conclusion

At altitude, the modulation of the O2 delivery to tissuesby fine tuning of the blood–O2 affinity is of greatimportance, because relatively small changes in the

Fig. 2 Putative molecular mechanism by which nitric oxide (NO) iscarried by oxyhemoglobin and exported out of the red blood cell(RBC) upon hemoglobin (Hb) deoxygenation. In the lung, Hbbinds O2 at the heme and NO at the b-Cys93 residue, which lies in apocket of the Hb b chain (left panel). Thus, in the oxygenated state,the NO groups are hidden from the environment. In thecorrespondence of the oxy-to-deoxy transition, the a-threonine41residue interacts with b-Cys93, thereby exposing the NO groups(right panel). Some NO groups are autocaptured by the heme, butothers are transferred to thiol acceptors such as the anionexchanger on the surface of the RBC membrane that pumps NOout of the RBC. As a result, the low PO2 in peripheral tissues forcesthe allosteric changes that destabilize the b-Cys93–NO bond. TheNO released upon deoxygenation promotes vasodilatation andenhances the O2 delivery

356

blood–O2 affinity are capable to maximize Da–vSO2 andoptimize the O2 transport to tissue keeping the circula-tory load to a minimum. However, other factors inde-pendent of the measured P50, and linked to Hballosterism and RBC function, may have effects thatpotentially mask those related to the changes in blood–O2 affinity.

Acknowledgement We gratefully acknowledge the helpful advice byProfessor Robert M. Winslow, Sangart, Inc., for the discussion ofthe RBC transit time.

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