Arterial Oxygen Content Is Precisely Maintained by Graded Erythrocytotic Responses in Settings of High/Normal Serum Iron Levels, and Predicts Exercise Capacity: An Observational Study of Hypoxaemic Patients with Pulmonary Arteriovenous Malformations

Abstract

Background

Oxygen, haemoglobin and cardiac output are integrated components of oxygen transport: each gram of haemoglobin transports 1.34 mls of oxygen in the blood. Low arterial partial pressure of oxygen (PaO2), and haemoglobin saturation (SaO2), are the indices used in clinical assessments, and usually result from low inspired oxygen concentrations, or alveolar/airways disease. Our objective was to examine low blood oxygen/haemoglobin relationships in chronically compensated states without concurrent hypoxic pulmonary vasoreactivity.

Methodology

165 consecutive unselected patients with pulmonary arteriovenous malformations were studied, in 98 cases, pre/post embolisation treatment. 159 (96%) had hereditary haemorrhagic telangiectasia. Arterial oxygen content was calculated by SaO2 x haemoglobin x 1.34/100.

Principal Findings

There was wide variation in SaO2 on air (78.5–99, median 95)% but due to secondary erythrocytosis and resultant polycythaemia, SaO2 explained only 0.1% of the variance in arterial oxygen content per unit blood volume. Secondary erythrocytosis was achievable with low iron stores, but only if serum iron was high-normal: Low serum iron levels were associated with reduced haemoglobin per erythrocyte, and overall arterial oxygen content was lower in iron deficient patients (median 16.0 [IQR 14.9, 17.4]mls/dL compared to 18.8 [IQR 17.4, 20.1]mls/dL, p<0.0001). Exercise tolerance appeared unrelated to SaO2 but was significantly worse in patients with lower oxygen content (p<0.0001). A pre-defined athletic group had higher Hb:SaO2 and serum iron:ferritin ratios than non-athletes with normal exercise capacity. PAVM embolisation increased SaO2, but arterial oxygen content was precisely restored by a subsequent fall in haemoglobin: 86 (87.8%) patients reported no change in exercise tolerance at post-embolisation follow-up.

Significance

Haemoglobin and oxygen measurements in isolation do not indicate the more physiologically relevant oxygen content per unit blood volume. This can be maintained for SaO2 ≥78.5%, and resets to the same arterial oxygen content after correction of hypoxaemia. Serum iron concentrations, not ferritin, seem to predict more successful polycythaemic responses.

About the Authors:

Vatshalan Santhirapala

Affiliations Imperial College School of Medicine, Imperial College, London, United Kingdom, National Heart and Lung Institute (NHLI) Cardiovascular Science, Imperial College, London, United Kingdom, Respiratory Medicine, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

Louisa C. Williams

Affiliation: Respiratory Medicine, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

Hannah C. Tighe

Affiliation: Respiratory Medicine, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

James E. Jackson

Affiliation: Department of Imaging, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

Claire L. Shovlin

* E-mail: [email protected]

Affiliations National Heart and Lung Institute (NHLI) Cardiovascular Science, Imperial College, London, United Kingdom, Respiratory Medicine, Hammersmith Hospital, Imperial College Healthcare NHS Trust, London, United Kingdom

Introduction

The primary function of haemoglobin is to transport oxygen from the alveolar capillaries to the tissues. Blood oxygen content is determined by the haemoglobin concentration, and the partial pressure of oxygen in blood (PaO2) which governs the percentage haemoglobin saturation (SaO2) [1]. In turn, the overall transport of oxygen to the tissues depends upon the oxygen content of arterial blood, and the volume of blood reaching the tissues in any given period (cardiac output) [1]. Profound arterial hypoxaemia can be tolerated if subjects are gradually acclimatised [2],[3],[4],[5]. Mechanisms that help sustain oxygen delivery include higher haemoglobin concentrations through secondary erythrocytosis/polycythaemia [2],[3],[4], and tissue changes that optimise energy metabolism including suppression of mitochondrial biogenesis and oxidative metabolism[5]. Similarly, severe anaemia is tolerated if chronic compensatory mechanisms can be employed: anaemic patients unable to sustain normal haemoglobin concentrations have hyperdynamic circulations, with high cardiac outputs accompanied by lower systemic vascular resistance [6],[7],[8].

Although this is an integrated system of oxygen delivery, and challenged by multiple different pathological states, the problem is that components are generally discussed in a discipline-restricted manner. Oxygen and clinical practice guidelines are based on PaO2 and/or SaO2 measurements: of more than 2,700 PubMed results retrieved using “oxygen” and “guidelines,” only 106 were found by including the term “haemoglobin” [or ”hemoglobin”], with relevant articles predominantly restricted to the anaesthetic/critical care literature. Synthesis of iron-containing haemoglobin is impaired by iron deficiency which affects more than a billion individuals worldwide[9], but in the iron deficiency literature, anaemia is generally discussed without reference to oxygen transport: On 25.11.2013 although there were 14,284 PubMed references for “iron”, “deficiency” and “anaemia/anemia,” only 73 were retrieved including the terms “oxygen” and either “delivery” or “transport.” In addition, it is rarely discussed that hypoxaemia caused by low inspired oxygen concentrations, at altitude [2],[3],[4],[5], or due to alveolar/airways disease [10], is inevitably accompanied by a separate process, namely hypoxic pulmonary vasoconstriction (HPV) [2],[11]. HPV is the physiological response to alveolar hypoxia, responsible for ventilation-perfusion matching [2],[11]. As HPV leads to local or generalised elevation of pulmonary vascular resistance [2],[11], this process has the potential to confound physiological and clinical responses in hypoxaemic subjects.

Hypoxaemic patients with pulmonary arteriovenous malformations (PAVMs) [12] provide an opportunity to study integrated oxygen/haemoglobin relationships without confounding hypoxic pulmonary vasoreactivity: PAVMs are abnormal blood vessels that usually develop by teenage years, and provide direct capillary-free communications between the pulmonary and systemic circulations [12]. Hypoxaemia results from deoxygenated pulmonary arterial blood transiting these right-to-left (R-L) shunts and bypassing the alveolar capillary sites of gas exchange. There is good agreement between the calculated R–L shunt using measured SaO2, and the anatomic R-L shunt, confirming that the R-L shunt is the predominant cause of the arterial hypoxaemia [13]. Hypoxaemic PAVM patients are therefore not at risk of hypoxic pulmonary hypertension, and pulmonary vascular resistance at rest is low in patients with severe PAVMs [14],[15]. Arterial PaO2 and SaO2 are inversely related to the proportion of the cardiac output passing through the R–L shunts [13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24], and hypoxaemia may be severe. As at altitude, and in patients with other cardiorespiratory cyanotic disease, secondary polycythaemia [13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24],[25] and increased cardiac outputs [14],[19] help to sustain long term oxygen delivery in hypoxaemic PAVM patients.

PAVM patients provide a particularly good model in which to study integrated oxygen/haemoglobin relationships not only because of the absence of HPV, but also because serial evaluations can be performed before and after correction of hypoxaemia by PAVM embolisation treatments which are recommended to prevent paradoxical embolic strokes and other complications [23],[24],[25],[26],[27]. High proportions of PAVM patients have iron deficiency, due to the presence of underlying hereditary haemorrhagic telangiectasia (HHT), [28],[29], and inadequate replacement of haemorrhagic iron losses [30],[31],[32]. Importantly, such iron deficiency generally occurs in the absence of confounding inflammation: inflammatory markers such as C reactive protein are generally normal [24],[33], and hepcidin concentrations are appropriate for iron stores, exhibiting the same relationships with ferritin as in the general population [30].

Our aim was to evaluate oxygen/haemoglobin relationships in a large cohort of PAVM patients before and after embolisation treatment, and in the presence and absence of iron-restricted erythropoesis, in order to obtain insights not available from other hypoxaemic pathologies or models. This is important for PAVM patients because they are currently managed by oxygen/polycythaemia guidelines extrapolated from evidence in other patient groups [34],[35],[36], and our clinical observations suggested that the extrapolated guidelines were not serving the population well [12],[37].

The study objectives were achieved. Here we present data that provide new insights into the regulation of blood oxygen content through polycythaemia, and add to the evidence base from which clinical guidance may be developed.

Methods

Ethical approved was from the Hammersmith, Queen Charlotte's, Chelsea, and Acton Hospital Research Ethics Committee (LREC 2000/5764: “ Case Notes Review: Hammersmith Hospital patients with pulmonary arteriovenous malformations and hereditary haemorrhagic telangiectasia (HHT).”) The ethics committee approved the review of the case notes for research purposes without seeking individual consents. Individuals were asked for written consent as part of separate ethics approvals for any research protocol (questionnaire, blood test, scan or other study) that was not part of standard clinical practice, but no such data are reported in this manuscript.

Clinical evaluations

Due to the pan-UK referral base to our PAVM/HHT service, patients are initially evaluated in single day assessments (see www.imperial.ac.uk/nhli/hht_pavm_patient). For patients referred with suspected/confirmed HHT [38],[39], PAVM screening or assessment is performed using validated SaO2 measurements [24], and thoracic CT scans if not performed previously/recently. The study cohort represents the 165 consecutive patients with CT-proven PAVMs first seen between June 2005- September 2010, and excludes the May 1999-May 2005 patients whose findings [23] precipitated the current study.

Presentation assessments.

At initial assessment, SaO2 was measured by pulse oximetry (Ohmeda Biox 3900, Boulder, Colorado) while breathing room air. Measurements were made for 10 minutes in the erect posture, recorded at one minute intervals, with the mean value from minutes 7–10 reported, as utilised in previous clinical studies [17],[20],[21],[23],[24],[33],[40] and recently validated [24]. To minimise diurnal variability for iron measurements, timings of blood tests were standardised to mid-late afternoon [33]. Full blood counts measured haemoglobin, haematocrit, red blood cell (RBC, erythrocyte) number, mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC) and red cell distribution width (RDW)) on XE Series Analysers (Sysmex, UK). Biochemical indices including serum iron, transferrin saturation index (TfSI) and ferritin were measured on Ci1600 Architect Analysers (Abbott Diagnostics, Ireland).

Embolisation admissions and follow up.

Embolisation was performed as described elsewhere [12],[37],[40],[41] in 98 patients during standardised 48hr admissions. Pulmonary artery pressures (PAP) were recorded by a centrally-placed catheter prior to contrast injection [12],[37],[40],[41]. Embolisation was not indicated in 66 patients, primarily due to PAVMs with feeding arteries too small for treatment (n = 63 including two who had previous maximal treatment elsewhere). Three patients had contraindications [40],[42], including two with very severe pulmonary hypertension, one of whom was on a liver transplant waiting list. One patient declined embolisation. Twenty patients required more than one planned embolisation session to achieve maximal embolisation; seventeen required two sessions which spanned 1.5–13 (median 4.3) months; three required three sessions spanning 19–24 (median 20) months. Oximetry and blood tests were repeated in the 24hs before each pulmonary angiography/embolisation session, and oximetry the day after embolisation. Oximetry and blood tests were repeated in a standardised follow-up clinic which took place at least 2 months after the final embolisation. In their first post-embolisation follow up clinic, treated patients were asked in a non biased manner whether they had noticed any differences after embolisation, then asked to report what those differences were.

Study methodology

Pre-study power calculations to determine series length were performed on the HHT thrombotic endpoints under parallel study [23],[24],[33],[43]. Blinded to all other measurements (and subsequent analyses), two investigators (VS, CLS) assigned patient's clinic reports of their presentation exercise capacity to a scale [44] adapted from the Medical Research Council dyspnoea scale [45]. Because many were highly athletic individuals, the “normal” Grade 1 classification was subclassified into Grade 1a (if participating in intense sporting activity such as rowing, football, distance cycling or gym activities at least three times per week), and Grade 1b (representing other normals, who described dyspnoea only on strenuous exertion). Grades 2–5 describe progressively lower exercise tolerance/greater dyspnoea. The grading of individuals in the current cohort was published in Abstract format in 2011 [44]. Treated patients were also stratified as “improved exercise tolerance” or “no improvement” by their reports of changes post embolisation, blinded to other patient variables. Study follow up continued until all feasible first post embolisation assessments had been performed. In order to avoid inadvertent introduction of bias, other than entering data for remaining post embolisation assessments, no additional information was sought, no additional assessments were performed for the purposes of this study, and specifically, groupings generated by exercise tolerance or subsequent statistical analyses were not restudied or reassessed.

For the current study, oxygen content of blood and iron deficiency were generated as new variables by command line entry, blinded to all other parameters. Arterial oxygen content was calculated by SaO2 x haemoglobin x 1.34/100 where SaO2 was expressed as a %, and 1.34mls is the amount of oxygen carried per gram of haemoglobin [1]. Iron deficiency was assigned as absent (“0”) if ferritin, iron and TfSI were all clearly in the normal range (ferritin>20 µg/L, serum iron>11 µmol/L and TfSI>20%). In keeping with current recommendations [46],[47],[48], iron deficiency was assigned as present (“1”) if same-day ferritin was <15 µg/L. As discussed previously by ourselves [24],[33] and others [46],[47],[48], ferritin may be spuriously high in iron deficiency due to concurrent pathologies. Thus following haematinic validations [24], iron deficiency was also assigned as present (“1”) for individuals with both iron and TfSI clearly subnormal (<7 µmol/L and <20% respectively). All other combinations were assigned as intermediate/unknown (“.”).

Statistical Analyses

STATA IC version 12 (Statacorp, Texas) and GraphPad Prism 5 (Graph Pad Software Inc, San Diego) were used to calculate distributions of participant-specific variables, to perform comparisons between groups, and to generate graphs. Two group comparisons were by Spearman rank or Mann Whitney; three group repeated measures comparisons by Kruskal Wallis with post-test Dunns corrections. Univariate and multivariate linear, logistic and quantile regression was performed in STATA IC version 12 (Statacorp, Texas).

Results

Series demographics

The 165 patients were aged 17–87 (median 49) years. Sixty-two (37.6%) were male. 159 (96.4%) patients had underlying hereditary haemorrhagic telangiectasia (HHT) [38]. PAVMs had been diagnosed by a variety of routes, most commonly screening for PAVMs in suspected HHT patients/families (N = 62 [37%]); incidental detection by chest x-rays or thoracic/abdominal CT scans (N = 34 [20.5%]); investigations following strokes, brain abscess or neurological symptoms (N = 18 [12%]); and PAVM respiratory symptoms (N = 17 [10%]). Only 15 (9.1%) patients had evidence of significant co-existing disease, with obstructive spirometry due to either asthma or COPD the most common. One patient was receiving supplementary oxygen therapy at presentation and in follow up. No patients underwent venesection in the course of these studies.

SaO2 at rest ranged from 78.5–99% (median 95%, Table 1). In keeping with the high prevalence of HHT, many of the PAVM population had biochemical and haematinic evidence of iron deficiency (Table 1). Overall, haemoglobin ranged from 7.7 to 20.9 g/dl (median 14.1 g/dl), haematocrit from 0.26 to 0.61 (median 0.43).

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Table 1. Demographics of 165 consecutive unselected PAVM patients.

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Basis of polycythaemia and anaemia responses

As expected, haemoglobin values were higher in patients with lower SaO2 (Figure 1A). On average, for every 1% fall in SaO2, haemoglobin rose by 0.82 g/dl (regression coefficient -0.82 (95% CI −1.12, −0.51, p<0.0001). There was no change in the haemoglobin concentration per red cell (Figure 1B), and the rise in haemoglobin reflected higher red cell counts at lower SaO2 (Figure 1C). In the same population, lower serum iron was associated with lower haemoglobin (Figure 1D), attributable to a reduced haemoglobin concentration per red cell, with no change in red cell count (Figure 1E, 1F).

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Figure 1. Basis of polycythaemia and anaemia responses in PAVM patients.

Univariate associations demonstrating individual patient data (small diamonds), linear regression lines, and 95% confidence intervals (shaded) for two way relationships. A–C): The polycythaemic response to hypoxaemia. A) Lower SaO2 was associated with higher haemoglobin (the ‘polycythaemic response’). B) This polycythaemia was not attributable to increased haemoglobin concentration in red cells (mean corpuscular haemoglobin concentration, MCHC). C) Instead, the polycythaemia reflected increased red cell number (RBC), that is, secondary erythrocytosis, D–F) The anaemic response to iron deficiency: D) Lower serum iron concentrations were associated with lower haemoglobin (the ‘anaemic response’). E) This anaemic response to iron deficiency resulted from reduced haemoglobin concentration in red cells (MCHC), and not a change in red cell number (RBC) (F).

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To portray the inter-relationships graphically, three-way contour plots were generated. These indicated that the polycythaemic response was evident even in the setting of low ferritin concentrations: For haemoglobin (Figure 2A), haematocrit (Figure 2B), and MCHC (Figure 2C), higher values were observed in more hypoxaemic patients across all ferritin values. The erythrocytotic response in hypoxaemic patients was particularly prominent in patients with subnormal serum ferritin (Figure 2D).

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Figure 2. Three-way plots of relationships between haematinic indices and SaO2.

Three-way plots of relationships between SaO2 (y axis), ferritin/iron (x axis), and haematinic index (z axis). The most hypoxaemic patients are at the bottom of each graph, and patients with the lowest ferritin/iron levels to the left. The z axis effectively provides a 3 dimensional plot in which contours range from blue (lowest value of modelled haematinic variable) to red (highest values). A–D) Serum ferritin/SaO2 stratifications for the 105 PAVM patients with serum ferritin measurements: A) SaO2/ferritin/haemoglobin. B) SaO2/ferritin/haematocrit. C) SaO2/ferritin/MCHC. D) SaO2/ferritin/red cell count. Note that higher haemoglobin, haematocrit and MCHC are seen across the normal range for ferritin (10–150 or 20–300 µg/L, according to gender), but higher RBC number is only seen in patients with low or subnormal ferritin. E–H) Serum iron/SaO2 stratifications for the 141 PAVM patients with serum iron measurements. E) SaO2/iron/haemoglobin. F) SaO2/iron/haematocrit. G) SaO2/iron/MCHC. D) SaO2/iron/red cell count. Although serum iron and serum ferritin were correlated (Spearman rho 0.34, p = 0.006), serum iron did not show the same relationships with hematinic indices as serum ferritin (contrast A and E; B and F; C and G; D and H). The highest haematinic indices were observed with serum iron above the normal range (7–27 µmol/L).

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Different haematinic/SaO2 inter-relationships were observed with serum iron. Three-way contour plots indicated that for patients with lower SaO2, polycythaemic responses (higher haemoglobin (Figure 2E), higher haematocrit (Figure 2F), higher MCHC (Figure 2G) and higher red cell count (Figure 2H)) were evident only in the setting of high-normal serum iron concentrations.

Preservation of arterial oxygen content by polycythaemic response

Due to the higher haemoglobin in more hypoxaemic patients, oxygen content per unit blood volume was similar across all degrees of hypoxaemia (bold black line, Figure 3). SaO2 explained only 0.1% of the variance in arterial oxygen content per unit blood volume (p = 0.83). In non iron deficient patients, the median arterial oxygen content was 18.8 [IQR 17.4, 20.1]ml/dl. Iron deficient patients had similar SaO2 to non iron deficient patients, but haemoglobin was ∼2 g/dl lower. As a result, arterial oxygen content in iron deficient patients either defined conventionally by ferritin <15 µg/L (median 16.0 [IQR 14.9, 17.4]ml/dl) or by “low iron state” which included patients with serum iron <11 µmol/L irrespective of ferritin (median 16.3 [IQR 14.8, 18.1]ml/dl), were both substantially lower than in non iron deficient patients (p values<0.0001, Figure 3).

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Figure 3. Arterial oxygen content stratified by oxygen saturation (SaO2).

Oxygen content across all degrees of hypoxaemia, calculated by SaO2 x haemoglobin x 1.34/100, where SaO2 was expressed as a %, and 1.34mls is the amount of oxygen carried per gram of haemoglobin.[1] The bold black line represents the regression line for all patients, irrespective of iron status (p = 0.69). Grey diamonds/dotted line/shaded 95% confidence interval represent patients without iron deficiency (p = 0.33). Red diamonds/dotted line/shaded 95% confidence interval represent patients with iron deficiency (p = 0.97).

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Consequences of reduced oxygen content

The median values for arterial oxygen content (18.8 ml/dl versus 16.0 ml/dl, see Figure 3) imply that per unit blood volume, oxygen content was reduced by approximately 15% in the iron deficient group. Graded self-reported exercise tolerance [44] was used as a crude indicator of whether compensating for reduced blood oxygen content was achievable.

Figure 4A demonstrates that there was no clear relationship between SaO2 (the most commonly measured index of oxygenation in clinical practice [34],[35],) and graded exercise tolerance: Seven individuals regularly participated in intense sporting activity despite marked resting hypoxaemia (SaO2<90%, Figure 1). In contrast there was a very clear inverse correlation between exercise grade and arterial oxygen content, which incorporates both SaO2 and haemoglobin. As shown in Figure 4B, exercise tolerance was worse in patients with lower oxygen content (coefficient −0.0012, p<0.0001), with the trend evident across all grades of exercise tolerance.

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Figure 4. 165 PAVM patients graded according to self-reported exercise tolerance.

A) SaO2 relationships. Error bars represent mean and standard deviation. Similar trends were observed for median and IQR (data not shown). B) Oxygen content, calculated by SaO2 x haemoglobin x 1.34/100, where SaO2 was expressed as a %, and 1.34mls is the amount of oxygen carried per gram of haemoglobin [1]. Error bars represent mean and standard deviation, but a similar trend was observed for median and IQR (data not shown).

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Oxygen content and iron relationships in athletes

Figure 4B suggested that the blood oxygen content may be higher in the pre-defined athletic group than the other Grade 1 normals. Haemoglobin tended to be higher for the degree of hypoxaemia in the athletes, compared to non-athletic normals (Figure 5A). This could not be ascribed to differential use of iron tablets: excluding the 25 iron tablet users (6 athletes, 19 non athletic normals) the higher ratio of [100*haemoglobin]/SaO2 in the athletic group persisted (median 16.6 [IQR 14.7, 20.0] vs 15.2 [IQR 12.2, 17.7], p = 0.016). This would be in keeping with the limited evidence suggesting that exercise training can increase total haemoglobin and red cell mass, attributed to stimulated bone marrow erythropoiesis [49].

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Figure 5. Boxplot comparisons of athletes and other individuals with normal exercise tolerance.

Individuals with normal exercise tolerance (Grade 1) were subclassified into Grade 1a (athletes, grey symbols/lines, if participating in intense sporting activity such as rowing, football, distance cycling or gym activities at least three times per week), and Grade 1b (other normals, red symbols/lines, if they described dyspnoea only on strenuous exertion). All assignments were made blinded to physiological parameters [44]. A) Haemoglobin (Hb) adjusted for SaO2, presented as (100*haemoglobin)/SaO2. Mann Whitney p value  =  0.0059. P values were also calculated by Kruskal Wallis across all exercise grades, when Dunn's post test correction comparing the athletic and non athletic normals gave a p value of<0.05. B) Serum iron. Mann Whitney p value  =  0.010. P values were also calculated by Kruskal Wallis across all exercise grades, when Dunn's post test correction comparing the athletic and non athletic normals gave a p value of<0.05.

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The current study cohort suggests a potentially relevant change in iron handling: While there was no difference in serum ferritin values between the athletes (median 38 [IQR 19, 77]µg/L) and non-athletic normals (median 38 [IQR 16, 73]µg/L), the athletic group had higher serum iron concentrations (median 19.5 [IQR 13,28]µmol/l compared to 14 [IQR 9,17]µmol/l, p = 0.01 (Figure 5B)). Again, this could not be ascribed to differential use of iron tablets: excluding the 25 iron tablet users, median serum ferritin values in athletes and other normals were very similar at 43.5 [IQR 19, 86]µg/L and 40 [IQR 25, 78]µg/L respectively. However the differences in serum iron concentrations persisted (athletes: median 21 [IQR 13, 28]µmol/l; non athletic normals: 14 [IQR 10,18]µmol/l, p = 0.043).

Effect of embolisation and correction of hypoxaemia

In the 98 patients whose PAVMs were treated by embolisation, SaO2 improved immediately (median increase +2.5% (IQR +0.75, +5.0%, Table 2). Improvements were sustained on long term follow up 2–24 (median 7) months later (Table 2). Haemoglobin and haematocrit fell in the follow up period: for haemoglobin, the value at a median of 7 months after final embolisation was 0.60 g/dl lower (IQR −1.3 g/dl, +0.1 g/dl) than at presentation (Table 2). As a result, oxygen content in mls/dl generally returned to pre-embolisation values (Table 2, Figure 6).

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Figure 6. Blood oxygen content pre and post embolisation.

Post embolisation data were obtained at clinic follow up at a median of 7 months (range (2–24) months after the final embolisation. Shaded areas represent 95% confidence interval for quadratic regression line for all 52 patients with pre and post embolisation haemoglobin measurements (pseudo r2 0.44, p<0.0001). Open diamonds represent individuals with pre embolisation serum iron <4 µmol/L.

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Table 2. Changes in SaO2, haemoglobin, and oxygen content following PAVM embolisation.

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In these 98 patients, 86 (87.8%) reported no change in exercise tolerance at their first post-embolisation follow up clinic at a median of 7 months (range 2–24, [IQR 6, 9]) post final embolisation. A common comment was feeling better initially after embolisation, then returning to normal and the individuals not being sure if this was because they had ‘got worse’ again, or become more used to an improved state. We interpreted this as reflecting an interim increase in arterial oxygen content before the fall in haemoglobin documented in Table 2.

Twelve patients (12.2%) did report sustained improvements in exercise tolerance, particularly on stairs, on hills, or during specific sports. Three commented on an improved breathing pattern during swimming (n = 2) or yoga (n = 1). Using logistic regression, there was a trend for improvement in exercise tolerance to be reported more commonly by patients with lower oxygen content at presentation, but no relationship with degree of improvement in oxygen content post embolisation. Improved exercise tolerance was reported more commonly by patients with lower serum albumin, which is a marker of illness/inflammation (attributed to reduced hepatic synthesis, increased albumin catabolism and vascular permeability [50]). An improvement in exercise tolerance was also reported more commonly by patients with higher PAP (Table 3), especially PAP(diastolic) which most closely reflects pulmonary vascular resistance [51]. We concluded that co-existing diseases were associated with less successful compensation to initial hypoxaemia, and improved exercise tolerance post embolisation.

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Table 3. Demographics, and univariate associations with improvement in exercise capacity post embolisation for the 98 PAVM patients.

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Discussion

In this study, which examines chronic compensatory changes in an unselected cohort of 165 consecutive patients with PAVMs, we demonstrate that the adaptive polycythaemic/secondary erythrocytotic response not only maintains arterial oxygen content for SaO2 ≥78·5%, but also resets to the same arterial oxygen content after correction of hypoxaemia. Impairment of haemoglobin synthesis due to iron deficiency was the key determinant of reduced oxygen content, and reflected the high proportion of PAVM patients who had underlying HHT. Serum iron concentrations rather than ferritin were more predictive of a successful polycythaemic response. Future studies will be required to test if higher serum iron:ferritin ratios are a consequence of athletic training [49],[52], or prerequisite. The preliminary data on improved exercise tolerance following embolisation treatment of PAVMs suggest factors unrelated to iron deficiency are also important in preventing optimal compensations to hypoxaemia.

Strengths of the current study include the large number of patients studied with consistent methodologies, the extremely reduced SaO2 present long term, the general absence of confounding hypoxic pulmonary vasoreactivity and inflammation, and the high proportion with significant iron deficiency. The absence of arterial blood gas measurements of PaO2 may be considered a weakness, but the reproducibility of the replicate pulse oximetry SaO2 measurements [24] render this criticism less important. Not all patients had ferritin measured, but in view of the similar cohorts identified by low serum iron and the ferritin values conventionally used to define iron deficiency [24],[46],[47],[48], we concluded that for the parameters under examination, it was appropriate to use serum iron <11 µmol/L as a marker of an iron insufficient state. Although a single centre study, there was little bias introduced by geography (UK-wide referrals), or type of presentation.

The current data are important for oxygen/polycythaemia guidance for PAVM patients, currently extrapolated from data in other patient groups, particularly patients with COPD where HPV also operates. The guidelines [34],[35],[36] are interpreted as stating that PAVM patients who are hypoxaemic and polycythaemic should have venesection (to reduce plasma viscosity [36]), and oxygen supplementation [34],[35],[36]. For HHT/PAVM patients, it is not high haematocrit/haemoglobin that is associated with venous thromboembolism (VTE) [33],[43], but markers of iron deficiency [33] which would be exacerbated by venesection [36],[53],[54]. Venesection will inevitably result in rebound polycythaemia unless iron deficiency is induced [53],[54]. Supplementary oxygen is generally prescribed to hypoxaemic patients acutely to reduce risks of hypoxic tissue injury [34],[35], and longer term to reduce the risks of developing hypoxic pulmonary hypertension [11],[34],[35] and/or hyperviscosity states [36]. The current study reminds of the normal long-term compensations that help prevent tissue hypoxia. Additionally, multiple studies demonstrate that low SaO2/PaO2 are tolerated well by PAVM patients at rest [13],[14],[15],[16],[17],[19],[20],[22],[24],[25],[26], on exercise [14],[15],[18],[19], and even in flight when barometric pressure and alveolar oxygen tension fall further [55]. The data in the current manuscript suggest the PAVM patients most likely to benefit from oxygen supplementation would be iron deficient patients with less successful polycythaemic compensatory responses, and this could be tested in future trials.

Taken more broadly, the data reemphasise the importance of combining haemoglobin and SaO2 to evaluate the oxygen content of arterial blood. Using a term such as OaHb for arterial oxygen content per unit volume on air, calculated by SaO2 x haemoglobin x 1.34/100, may serve as an aide memoire. The haemodynamic implications of impaired haemoglobin synthesis are not currently emphasised, most likely attributable to the discrepancy identified by Hébert et al [8], between the copious data in the old physiological literature (summarised in [7]) and paucity in more recent clinical studies. In the current study, average arterial oxygen content was 15% lower in the iron deficient group, implying that unless there were local tissue compensations, cardiac output would need to be approximately 15% higher to deliver the same amount of oxygen to the tissues [1],[7]. The incremental increase would be greater in higher output states, for example on exercise, during sepsis, or for HHT patients with hepatic and/or other visceral AVMs with left-to-right shunting. The crude exercise data presented here suggest limitations in cardiac/circulatory compensatory capacity in the setting of lower blood oxygen content, but this will clearly need to be addressed in targeted prospective studies.

In conclusion, discipline-restricted discussions of haemoglobin or commonly measured oxygen parameters in isolation do not address the more physiologically relevant arterial oxygen content (mls of oxygen per unit blood volume). For hypoxaemic patients, secondary erythrocytosis maintains the oxygen content of blood, provided the bone marrow has sufficient time, function, and iron to respond, and this may be enhanced by athletic training.

Acknowledgments

The authors want to express their thanks to JMB Hughes for setting up the replicate SaO2 assessments in the late 1980s, to JMB Hughes and Luke Howard for helpful discussions, and to their clinical colleagues for their support of the clinical service, particularly in the pulmonary function laboratory, CT scanner suite, and service pathology laboratories.

Author Contributions

Conceived and designed the experiments: VS CLS. Performed the experiments: LCW HT JEJ CLS. Analyzed the data: VS CLS. Contributed reagents/materials/analysis tools: LCW HCT JEJ CLS. Wrote the paper: CLS. Generated the database: VS. Assigned exercise tolerance grades: VS CLS. Performed pulmonary function measurements: LCW HCT. Advised on oxygen guidelines: LCW HCT. Performed embolisations and associated measurements: JEJ. Reviewed all patients: CLS. Performed statistical analyses presented: CLS. Generated the figures: CLS. Reviewed and approved the final article: VS LCW HT JEJ CLS. Guarantor of the data: CLS.

Citation: Santhirapala V, Williams LC, Tighe HC, Jackson JE, Shovlin CL (2014) Arterial Oxygen Content Is Precisely Maintained by Graded Erythrocytotic Responses in Settings of High/Normal Serum Iron Levels, and Predicts Exercise Capacity: An Observational Study of Hypoxaemic Patients with Pulmonary Arteriovenous Malformations. PLoS ONE 9(3): e90777. https://doi.org/10.1371/journal.pone.0090777

References

1. Pittman RN (2011) Oxygen Transport. Chapter 4 in: Regulation of Tissue Oxygenation. San Rafael (CA): Morgan & Claypool Life Sciences. Available: http://www.ncbi.nlm.nih.gov/books/NBK54103. Accessed 2014 Feb 10.

2. West JB (1998) High Life: a history of high altitude medicine and physiology. Am Physiol Soc and Oxford Univ Press, New York Oxford, 1998.ISBN 978-0-1951219-4-0

3. Grocott MP, Martin DS, Levett DZ, McMorrow R, Windsor J, et al. (2009) Arterial blood gases and oxygen content in climbers on Mount Everest. N Engl J Med 360: 140–9.

4. Ou LC, Salceda S, Schuster SJ, Dunnack LM, Brink-Johnsen T, et al. (1998) Polycythaemic responses to hypoxia: molecular and genetic mechanisms of chronic mountain sickness. J Appl Physiol 84: 1242–51.

5. Levett DZ, Radford EJ, Menassa DA, Graber EF, Morash AJ, et al. (2012) Acclimatization of skeletal muscle mitochondria to high-altitude hypoxia during an ascent of Everest. FASEB 26: 1431–41.

6. Anand IS, Chandrashekhar Y, Wander GS, Chawla LS (1995) Endothelium-derived relaxing factor is important in mediating the high output state in chronic severe anemia. J Am Coll Cardiol 25: 1402–7.

7. Porter WB, Watson JG (1953) The Heart in Anemia. Circulation 8: 111–116.

8. Hébert PC, Van der Linden P, Biro G, Hu LQ (2004) Physiologic aspects of anemia. Crit Care Clin 20: 187–212.

9. McLean E, Cogswell M, Egli I, Wojdyla D, Benoist B (2009) Worldwide prevalence of anaemia, WHO Vitamin and Mineral Nutrition Information System, 1993-2005. Public Health Nutr 12: 444–54.

10. Boyer L, Chaar V, Pelle G, Maitre B, Chouaid C, et al. (2011) Effects of polycythemia on systemic endothelial function in chronic hypoxic lung disease. J Appl Physiol 110: 1196–203.

11. Evans AM, Hardie DG, Peers C, Mahmoud A (2011) Hypoxic pulmonary vasoconstriction: mechanisms of oxygen-sensing. Curr Opin Anaesthesiol 24: 13–20.

12. Shovlin CL, Jackson JE (2010) Pulmonary arteriovenous malformations and other vascular abnormalities. In: Mason RJ, Broaddus VC, Martin T, et al.. editors. Murray and Nadel's Textbook of Respiratory Medicine. 5th Edn. Philadelphia, Elsevier-Saunders. pp. 1261–1282.

13. Ueki T, Hughes JMB, Peters AM, Bellingan GJ, Mohammed MAM, et al. (1994) Oxygen and 99mTc-MAA shunt estimations in patients with pulmonary arteriovenous malformations: effects of changes in posture and lung volume. Thorax 49: 327–331.

14. Whyte MK, Hughes JM, Jackson JE, Peters AM, Hempleman SC, et al. (1993) Cardiopulmonary response to exercise in patients with intrapulmonary vascular shunts. J Appl Physiol 75: 321–328.

15. Terry PB, White RI Jr, Barth KH, Kaufman SL, Mitchell SE (1983) Pulmonary arteriovenous malformations. Physiologic observations and results of therapeutic balloon embolization. N Engl J Med 308: 1197–200.

16. Chilvers ER, Peters AM, George P, Hughes JMB, Allison DJ (1989) Quantification of right to left shunt through pulmonary arteriovenous malformations using 99Tcm albumin microspheres. Clin Radiol 39: 611–614.

17. Dutton JA, Jackson JE, Hughes JM, Whyte MK, Peters AM, et al. (1995) Pulmonary arteriovenous malformations: results of treatment with coil embolization in 53 patients. AJR Am J Roentgenol 165: 1119–1125.

18. Pennington D, Gold W, Gordon R, Steiger D, Ring E, et al. (1992) Treatment of pulmonary arteriovenous malformations by therapeutic embolization. Am Rev Resp Dis 145: 1047–1051.

19. Whyte MK, Peters AM, Hughes JM, Henderson BL, Bellingan GJ, et al. (1992) Quantification of right to left shunt at rest and during exercise in patients with pulmonary arteriovenous malformations. Thorax 47: 790–796.

20. Thompson RD, Jackson J, Peters AM, Doré CJ, Hughes JM (1999) Sensitivity and specificity of radioisotope right-left shunt measurements and pulse oximetry for the early detection of pulmonary arteriovenous malformations. Chest 115: 109–13.

21. Gupta P, Mordin C, Curtis J, Hughes JM, Shovlin CL, et al. (2002) Pulmonary arteriovenous malformations: effect of embolization on right-to-left shunt, hypoxemia and exercise tolerance in 66 patients. AJR Am J Roentgenol 179: 347–355.

22. Mager JJ, Zanen P, Verzijlbergen F, Westermann CJ, Haitjema T, et al. (2002) Quantification of right-to-left shunt with (99m)Tc-labelled albumin macroaggregates and 100% oxygen in patients with hereditary haemorrhagic telangiectasia. Clin Sci (Lond) 102: 127–34.

23. Shovlin CL, Jackson JE, Bamford KB, Jenkins IH, Benjamin AR, et al. (2008) Primary determinants of ischaemic stroke/brain abscess risks are independent of severity of pulmonary arteriovenous malformations in hereditary haemorrhagic telangiectasia. Thorax 63: 259–66.

24. Shovlin CL, Chamali B, Santhirapala V, Livesey JA, Angus G, et al. (2014) Ischaemic strokes in patients with pulmonary arteriovenous malformations and hereditary hemorrhagic telangiectasia: associations with iron deficiency and platelets. PLOS ONE 2014 Feb 19;9(2):e88812

25. Cottin V, Chinet T, Lavolé A, Corre R, Marchand E, et al. (2007) Pulmonary arteriovenous malformations in hereditary hemorrhagic telangiectasia: a series of 126 patients. Medicine (Baltimore) 86: 1–17.

26. Hewes RC, Auster M, White RI Jr (1985) Cerebral embolism-first manifestation of pulmonary arteriovenous malformation in patients with hereditary hemorrhagic telangiectasia. Cardiovasc Intervent Radiol 8: 151–5.

27. Moussouttas M, Fayad P, Rosenblatt M, Hashimoto M, Pollak J, et al. (2000) Pulmonary arteriovenous malformations: cerebral ischemia and neurologic manifestations. Neurology 55: 959–64.

28. Faughnan ME, Palda VA, Garcia-Tsao G, Geisthoff UW, McDonald J, et al. (2011) International guidelines for the diagnosis and management of hereditary haemorrhagic telangiectasia. J Med Genet 48: 73–87.

29. Shovlin CL (2010) Hereditary haemorrhagic telangiectasia: Pathogenesis, diagnosis and treatment. Blood Rev 24: 203–19.

30. Finnamore H, Le Couteur J, Hickson M, Busbridge M, Whelan K, et al. (2013) Hemorrhage-adjusted iron requirements, hematinics and hepcidin define hereditary haemorrhagic telangiectasia as a model of hemorrhagic iron deficiency. PLoS ONE 8(10): e76516

31. Kjeldsen AD, Kjeldsen J (2000) Gastrointestinal bleeding in patients with hereditary hemorrhagic telangiectasia. Am J Gastroenterol 95: 415–8.

32. Silva BM, Hosman AE, Devlin HL, Shovlin CL (2013) A questionnaire-based study suggests lifestyle and dietary factors influencing nosebleed severity in hereditary hemorrhagic telangiectasia (HHT). Laryngoscope 123: 1092–9.

33. Livesey JA, Manning RA, Meek JH, Jackson JE, Kulinskaya E, et al. (2012) Low serum iron levels are associated with elevated plasma levels of coagulation factor VIII and pulmonary emboli/deep venous thromboses in replicate cohorts of patients with hereditary haemorrhagic telangiectasia. Thorax 67: 328–33.

34. British Thoracic Society Standards of Care Committee (2006) Clinical component for the home oxygen service in England and Wales. Available: https://www.brit-thoracic.org.uk/Portals/0/Clinical%20Information/Home%20Oxygen%20Service/clinical%20adultoxygenjan06.pdf. Accessed 2014 Feb 10.

35. O'Driscoll BR, Howard LS, Davison AG (2008) BTS guideline for emergency oxygen use in adult patients. British Thoracic Society. Thorax 63 Suppl 6vi1–68.

36. McMullin MF, Bareford D, Campbell P, Green AR, Harrison C, et al. (2005) Guidelines for the diagnosis, investigation and management of polycythaemia/erythrocytosis. Br J Haematol 130: 174–95.

37. Shovlin CL Wilmshurst P, Jackson JE (2011) Pulmonary arteriovenous malformations and other pulmonary aspects of hereditary haemorrhagic telangiectasia. Eur Respir Mon 54: 218–245.

38. Shovlin CL, Guttmacher AE, Buscarini E, Faughnan ME, Hyland RH, et al. (2000) Diagnostic criteria for hereditary hemorrhagic telangiectasia (Rendu-Osler-Weber syndrome). Am J Med Genet 91: 66–7.

39. van Gent MW, Velthuis S, Post MC, Snijder RJ, Westermann CJ, et al. (2013) Hereditary hemorrhagic telangiectasia: how accurate are the clinical criteria? Am J Med Genet A 161A: 461–6.

40. Shovlin CL, Tighe HC, Davies RJ, Gibbs JS, Jackson JE (2008) Embolisation of pulmonary arteriovenous malformations: no consistent effect on pulmonary artery pressure. Eur Resp J 32: 162–9.

41. Hart JL, Aldin Z, Braude P, Shovlin CL, Jackson JE (2010) Embolization of pulmonary arteriovenous malformations using the Amplatzer vascular plug: successful treatment of 69 consecutive patients. Eur J Radiology 20: 2663–70.

42. Shovlin CL, Gibbs JS, Jackson JE (2009) Management of pulmonary arteriovenous malformations in pulmonary hypertensive patients: a pressure to embolise? Eur Respir Rev 18: 4–6.

43. Shovlin CL, Sulaiman NL, Govani FS, Jackson JE, Begbie ME (2007) Elevated factor VIII in hereditary haemorrhagic telangiectasia (HHT): association with venous thromboembolism. Thromb Haemost 98: 1031–9.

44. Santhirapala V, Tighe HC, Springett JT, Wolfenden H, Hughes JMB, et al.. (2011) When hypoxaemia is not the predominant cause of dyspnea. Lessons from a single-centre 2005-2010 cohort of patients with pulmonary arteriovenous malformations. Chest 140: 681A, 10.1378/chest.1114540

45. Fletcher CM (1960) Standardised questionnaire on respiratory symptoms: a statement prepared and approved by the MRC Committee on the aetiology of chronic bronchitis (MRC breathlessness score). BMJ 2: 1665.

46. Smellie WS, Forth J, Bareford D, Twomey P, Galloway MJ, et al. (2006) Best practice in primary care pathology: review 3. J Clin Pathol 59: 781–789.

47. NICE Clinical Knowledge Summaries: Interpreting ferritin levels. Available http://cks.nice.org.uk/anaemia-iron-deficiency#!diagnosisadditional:1/A-487719_1. Accessed 2014 Feb 10

48. Valberg LS (1980) Plasma ferritin concentrations: their clinical significance and relevance to patient care. Can Med Assoc J 122: 1240–8.

49. Hu M, Lin W (2012) Effects of exercise training on red blood cell production: implications for anemia. Acta Haematol 127: 156–64.

50. Friedman AN, Fadem SZ (2010) Reassessment of albumin as a nutritional marker in kidney disease. J Am Soc Nephrol 21: 223–30.

51. Honda S, Sunagawa H, Tasaki H, Hirose M, Fukushige J (1975) Correlation between pulmonary vascular resistance and pulmonary arterial diastolic pressure in congenital heart diseases with left to right shunt. Jpn Heart J 16: 629–38.

52. Phillips BE, Williams JP, Gustafsson T, Bouchard C, Rankinen T, et al. (2013) Molecular networks of human muscle adaptation to exercise and age. PLoS Genet 9: e1003389.

53. Sondel PM, Tripp ME, Ganick DJ, Levy JM, Shahidi NT (1981) Phlebotomy with iron therapy to correct the microcytic polycythaemia of chronic hypoxia. Pediatrics 67: 667–70.

54. Van de Pette JE, Guthrie DL, Pearson TC (1986) Whole blood viscosity in polycythaemia: the effect of iron deficiency at a range of haemoglobin and packed cell volumes. Br J Haematol 63: 369–75.

55. Mason CG, Shovlin CL (2012) Flight-related complications are infrequent with hereditary haemorrhagic telangiectasia/pulmonary arteriovenous malformations, despite low oxygen saturations and anaemia. Thorax 67 80–81.