Bronchiolitis obliterans after allogeneic hematopoietic SCT - Nature

Bone Marrow Transplantation (2013) 48, 1224–1229 & 2013 Macmillan Publishers Limited All rights reserved 0268-3369/13 www.nature.com/bmt

ORIGINAL ARTICLE

Bronchiolitis obliterans after allogeneic hematopoietic SCT: further insight—new perspectives? M Ditschkowski, AH Elmaagacli, M Koldehoff, T Gromke, R Trenschel and DW Beelen Bronchiolitis obliterans (BO) is a late non-infectious pulmonary complication after allogeneic hematopoietic SCT. Among 982 patients after myeloablative hematopoietic SCT between January 2000 and October 2010, 68 were diagnosed with BO according to NIH criteria. The median onset of BO was 18 months post transplant, 5-year cumulative incidence was 5.8% and 5-year mortality 41%. BO prevalence rate was 10% among all long-term surviving hematopoietic SCT recipients and 12% among chronic GVHDpatients. Chronic GVHD, peripheral SCT and ABO blood group incompatibility were identified as risk factors associated with BO. IgG levels were significantly decreased at the onset of BO (6.7 g/L±0.7, P ¼ 0.001), the mean exhaled NO concentrations were lower in BO-patients than in stem cell recipients without BO (14 p.p.b.±0.9 vs 20 p.p.b.±2.1) or healthy controls (25 p.p.b.±2.4, Po0.001). Hypoxia-inducible factor 1 alpha (HIF-1a) was significantly elevated in BO as compared with healthy controls or GVHD-patients without lung involvement (340±61 vs 127±22 vs 140±32, P ¼ 0.02). Calculated 5-year survival was superior in female than in male BO-patients (86 vs 45%, P ¼ 0.04). These results emphasize the relevance of BO as serious late complication with substantial mortality and point to essential pathophysiological changes due to regulatory responses to hypoxia. Bone Marrow Transplantation (2013) 48, 1224–1229; doi:10.1038/bmt.2013.17; published online 25 February 2013 Keywords: bronchiolitis obliterans; nitric oxide; allogeneic; GVHD; HIF-1a; SCT

INTRODUCTION Allogeneic hematopoietic SCT has been shown to provide longterm disease-free survival for otherwise fatal malignant or nonmalignant hematological disorders. With increasing survival rates due to toxicity reduced hematopoietic SCT methods, advanced GVHD-management and improved anti-infectious therapy and prophylaxis, a distinct increment in late transplant related complications can be observed. Pulmonary complications develop in 30–60% of allogeneic hematopoietic SCT recipients and are considered as a major cause of morbidity and mortality.1,2 Late-onset non-infectious pulmonary complications3 beginning 3 months after transplantation with an incidence of 10–15% are considered as life-threatening complications significantly limiting the quality of life.4,5 Although late-onset non-infectious pulmonary complications classification is rather undefined, the most common categorizations include bronchiolitis obliterans (BO), bronchiolitis obliterans organizing pneumonia and interstitial pneumonia.6,7 According to the NIH consensus criteria, BO is considered to be the only pulmonary manifestation of chronic GVHD.8 The reported incidence of BO in allogeneic hematopoietic SCT recipients ranges from 2–10% and mortality rates have been assessed between 13 and 23%.9,10 BO is prevalent in 2–5.5% of hematopoietic SCT recipients and in up to 14% of hematopoietic SCT recipients with chronic GVHD.11 Patients diagnosed with BO have a 5-year survival rate of only 45% vs 75% in patients without BO.9 Advanced disease stages at transplantation, preceding GVHD, older donor and recipient age, MTX use, anteceding respiratory infection, serum Ig deficiency and PBSC in contrast to BM are discussed as risk factors for BO.7,11 To better characterize contributing factors to BO development in a contemporary cohort of stem cell recipients after allogeneic hematopoietic SCT we retrospectively analyzed

incidences and outcomes at our center over the past decade and surveyed the influence of transplantation-related features for BO. Measurement of fractional exhaled nitric oxide (FeNO) is widely accepted as a non-invasive marker for airway inflammation in many respiratory conditions but it’s role in BO is not yet examined.12 The transcription factor hypoxia-inducible factor-1 (HIF-1) has a central role in the cellular adaptation to hypoxia and probably even promotes fibrogenesis and interstitial collagen deposition.13 Here, we focus on biological characteristics, risk factors and new technical and molecular approaches with a potential impact on early BO recognition. PATIENTS AND METHODS In a retrospective review 982 adult patients, who underwent myeloablative allogenic hematopoietic SCT between January 2000 and October 2010 and survived more than 100 days post-transplant were evaluated. Exclusively patients conditioned without the use of MoAb like anti-thymocyte globuline or alemtuzumab were considered. Immunoprophylaxis was standardised using CYA and a short course MTX at days 1, 3, 6, 11 after transplantation. Hematopoietic SCT recipients were in obligatory continuous ambulatory aftercare at our center, during which GVHD characteristics were documented. The diagnosis of BO was based on clinical presentation and repeated pulmonary function tests (PFTs) revealing decreased forced expiratory volumes for 1 s (FEV1) corresponding to the NIH consensus criteria for chronic GVHD.8 According to these criteria BO was categorized as mild (mild symptoms like shortness of breath after climbing one flight of steps and/or FEV1 60–79%), moderate (moderate symptoms with shortness of breath after walking on flat ground and/or FEV1 40–59%) and severe (severe symptoms with shortness of breath at rest, requiring O2 and/or FEV1o40%). In two patients surgical lung biopsy was performed for histological confirmation of the diagnosis. High-

Department of Bone Marrow Transplantation, WTZ, Comprehensive Cancer Centre, University Hospital Essen, Essen, Germany. Correspondence: Dr M Ditschkowski, Department of Bone Marrow Transplantation, University Hospital of Essen, University Duisburg-Essen, Hufelandstr. 55, Essen 45147, Germany. E-mail: [email protected] Received 14 September 2012; revised 16 January 2013; accepted 23 January 2013; published online 25 February 2013

BO after hematopoietic cell transplantation M Ditschkowski et al

1225 resolution computed tomography and exclusion of infectious etiological factors by bronchoalveolar lavages or induced sputum analyses were supplements included in the diagnostic routine. PFTs were performed routinely every 6 months after transplantation and additionally outof-band in symptomatic patients with non-resolving symptoms like exertional dyspnea and dry cough. BO treatment was standardized and consisted of inhaled beta mimetics and steroids, increase in the dosage of systemic immunosuppressive treatment (calcineurin inhibitors, corticosteroids, mycophenolate), medication with montelukast and azithromycin, occasional administration of systemic beta agonists and supportive ant-infectious therapy/prophylaxis.

RNA isolation and real-time-PCR for HIF-1a HIF-1a mRNA levels were measured in blood samples of 40 patients with advanced BO, 20 patients with moderate and severe chronic GVHD without BO and 10 healthy controls by quantitative real-time-PCR. RNA was isolated using the RNeasy Mini Kit (Qiagen, Hilden, Germany), according to the manufacturer’s instructions. Real-time PCR (primer and probe) for HIF1a was performed as previously described by Dales et al.14 Abl quantified by real-time PCR as published earlier was used as housekeeping gene.15 Quantification of RNA transcript expression was normalized determining the ratio between expression levels of targets and Abl. Median time of HIF-1a analysis was 48 month post-transplant in BO and 46 months after hematopoietic SCT in non-BO patients.

Table 1. Summary of characteristics describing patient profile, transplantation modalities and transplantation-related features Characteristics

Non-BO patients (survival4100 days) n

Total number of patients Median age at transplant Median donor age Recipient sex Male Female

%

BO-patients

n

914

68

42

42

39

39

%

n.s.

n.s. 533 381

58 42

43 25

37 63

342

37

26

38

Graft source BMT PBSCT

181 733

20 80

8 60

12 88

Donor relation Sibling Unrelated

433 481

47 53

25 43

37 63

HLA-pattern Matched Mismatched

705 190

77 21

53 15

78 22

Sex-mismatch hematopoietic SCT

Significance P

n.s.

n.s.

Measurement of exhaled nitric oxide (NO) and total IgG Online assessment of the FeNO was performed according to European Respiratory Society/American Thoracic Society guidelines16 using the electrochemistry-based NIOX MINO device (Aerocrine AB, Stockholm, Sweden). Constant expiratory flow rate of 50±5 mL/s was ensured at each measurement. FeNO levels were recorded in p.p.b. and quantified serially in a cohort of 45 patients with manifest BO, 30 hematopoietic SCTrecipients without BO (22 with chronic GVHD, 8 without chronic GVHD) and 25 healthy controls. Median time of FeNO measurement was 51 months post-transplant in BO and 48 months after hematopoietic SCT in non-BO patients. Serum levels of IgG were assessed in the context of the diagnostic blood routine in 45 BO patients at the onset of BO and in 30 hematopoietic SCTpatients without BO at corresponding time points between 14 and 18 months after transplant. IgG levels between 7.0 and 16.0 g/L were regarded as reference range. Values were given as mean±s.e.m.

Statistics Differences in frequencies of discrete variables were tested using two-sided Fisher’s exact or w2 tests. The Wilcoxon rank–sum test was used to test the differences of continuous variables. In cases where no competing event needed to be considered, the probabilities of events over time were calculated by the product-limit method, and heterogeneity of time-to-event distribution functions was compared using log-rank scores.17 For the analysis of OS in BO and non-BO patients a landmark analysis was performed calculating OS from the post-transplant landmark month (median onset of BO) to death. To determine whether possibly competing events were independent (that is, death without BO) the probabilities of events over time were estimated by cause-specific cumulative incidence rates.18 Differences between the distributions of variables between groups were analyzed by nonparametric testing using the Mann–Whitney-U-test or the one-way analysis of variance according to Kruskal–Wallis. Mean values of independent samples were tested by unpaired t-test. Spearman’s rank coefficient was applied to test statistical dependence. Results were considered statistically significant at Po0.05. Data are presented as mean±s.e.m. For statistical analyses SPSS software version 18 (SPSS Inc., Chicago, IL, USA) and NCSS software 2004 (NCSS, LLC, Kaysville, UT, USA) were used.

RESULTS Incidences of BO and PFT pretransplant and at BO diagnosis The clinical features, demographic profile and transplant related data of the patient cohort are summarized in Table 1. Sixty-eight patients met the diagnostic criteria for BO resulting in a 5-year cumulative incidence of 5.8% (95% confidence interval (CI) 4.5–7.5%) post-hematopoietic SCT (Figure 1). Prevalence rate of BO was 10% among all long-term surviving hematopoietic SCT & 2013 Macmillan Publishers Limited

n.s.

n.s.

ABO incompatibility Major Minor Bidirectional Identical

458

50

46

68

0,028

191 195 73 455

21 21 8 50

18 19 9 23

26 28 13 33

Chronic GVHD Limited Extended

558 312 246

61 34 27

59 10 49

87 15 72

o0,001

Relapse

249

27

7

10

0,002

Abbreviations: BO ¼ bronchiolitis obliterans, HSCT ¼ hematopoietic SCT, PBSCT ¼ PBSC transplantation.

recipients and 12% among chronic GVHD-patients at the time of analysis. Except eight patients (12%) with at most moderately impaired pretransplant PFT due to chronic obstructive pulmonary disease all other BO-patients had no abnormality detected in PFT’s before hematopoietic SCT. The median time of post-transplant BO-diagnosis was 18 (range 6–168) months. According to the NIH criteria 10 patients (15%) were identified as having mild forms of BO, whereas 29 patients (43%) were categorized each with moderate and severe BO. In BO patients mean FEV1 was 44% (±2.4 s.e.m.), mean value for maximal expiratory flow at 25% of forced vital capacity was 28% (±3.1 s.e.m.) and mean FEV1/forced vital capacity ratio was 71% (±2.4 s.e.m.) at time of BO diagnosis. Pretransplant mean FEV1 was 90% (±3.4 s.e.m.) and mean FEV1/ forced vital capacity ratio 99% (±2.5 s.e.m.) in these patients. FeNO concentrations, serum IgG levels and HIF-1a Comparison of HIF-1a, FeNO and IgG between BO and non-BO patients is summarized in Table 2. The mean exhaled NO levels among BO-patients were 14 p.p.b. (±0.9), which was significantly lower compared with 20 p.p.b. (±2.1) in hematopoietic SCTrecipients without BO. Likewise, healthy controls showed with Bone Marrow Transplantation (2013) 1224 – 1229


1226 mean 25 p.p.b. (±2.4) FeNO significantly higher levels than BO patients (Po0.001). The mean FeNO concentration assessed in seven patients at the onset of BO was 10 p.p.b. (±2.5). A cutoff value of less than 15 p.p.b. for FeNO concentration was able to discriminate significantly between BO-patients, patients without BO and healthy controls (Po0.0001) resulting in a specificity of 82% and a sensitivity of 71%. Total serum IgG levels in hematopoietic SCT recipients without BO assessed at median 14 months after transplantation were found to be within the normal range (10.9 g/L±1.3). In contrast, IgG values of BO patients measured at the time of BO diagnosis were significantly lower (6.7 g/L±0.7, P ¼ 0.001) and below the normal range. On the basis of normal or decreased serum IgG levels as a distinctive factor for the risk of BO, specificity was calculated 83% and sensitivity 72%. The mean expression level of HIF-1a was significantly higher in BO patients compared with healthy controls and hematopoietic SCT recipients without BO (340±61 vs 127±22 vs 140±32, P ¼ 0.02). Among BO patients HIF-1a expression levels and FeNO concentrations revealed negative correlation ( 0.58, P ¼ 0.011). Hematopoietic SCT-related risk factors for BO development Clinical risk factors for BO were the presence of chronic GVHD and hematopoietic SCT with major or minor ABO incompatible donors. Both features were significantly more prevalent in patients who developed BO (Table 1) and the cumulative incidence for BO was significantly increased after ABO incompatible hematopoietic SCT resulting in a cumulative BO incidence of 7.4% (95% CI 5.4–10%)

Cumulative incidence

0.20

0.15

0.10

0.05

compared with 4.2% (95% CI 2.7–6.4%, P ¼ 0.032) after ABO identical hematopoietic SCT. Compared with ABO identical hematopoietic SCT BO incidence was significantly higher for major (P ¼ 0.033) and minor (P ¼ 0.048) but not for bidirectional incompatibility. Considering only hematopoietic SCT recipients with chronic GVHD, major incompatibility remained to be overrepresented in BO patients (P ¼ 0.05). Frequencies of ABO identical and incompatible transplantation were significantly (P ¼ 0.02) different with respect to the stem cell source. Hematopoietic SCT recipients with BMT showed 40% identical, 10% bidirectional, 20% major and 30% minor incompatibility, whereas 51% transplanted with PBSC were ABO identical, 8% bidirectional, 21% major and 20% minor incompatible. Significantly higher BO incidences according to major (P ¼ 0.046), minor (P ¼ 0.005) and bidirectional (P ¼ 0.018) ABO incompatibility were only seen after PBSCT. Although there were no differences regarding the transplantation frequency of PBSC or BM among patients with and without BO the 5-year cumulative BO incidence was significantly (P ¼ 0.019) higher following PBSC transplantation (6.7%, 95% CI 5.2–8.7%) than after BMT (2.2%, 95% CI 0.8–5.8%). No impact on BO development was detectable for the covariate patient sex, donor gender, donor relation (sibling or unrelated), donor age, HLA-match, conditioning regimen (with or without TBI) or occurrence of acute GVHD. Outcomes of BO-patients Among BO-patients 5-year OS from the time of BO diagnosis was calculated 59% (95% CI 42–76%) and 10-year OS was calculated 52% (95% CI 32–71%), which was significantly less than the survival of patients without BO (76 and 67%, P ¼ 0.043) by landmark analysis including only transplant recipients beyond 18 months survival post hematopoietic SCT (Figure 2). Although mild BO showed superior mean survival with 156 months (95% CI 119–192) compared with 72 months (95% CI 42–101) for moderate and 84 months (95% CI 61–106) for severe BO, differences were NS. Of 20 cases of death in the BO-cohort eight (40%) were directly BO-related. Other causes of death were relapse (n ¼ 2), sepsis (n ¼ 4), pneumonia (n ¼ 2) and GVHD (n ¼ 4). Of note is that OS was significantly (P ¼ 0.04) higher in female (86% after 5 years) compared with male (45% after 5 years) BO-patients (Figure 3). Post-BO survival was independent from 1.00

0.00 50 100 150 Time after transplantation (months)

200

Figure 1. Cumulative incidence of BO for patients surviving more than 100 days after hematopoietic SCT considering death and relapse as a competing risk. The 95% CI for all values are indicated by the upper and lower line. A full color version of this figure is available at the Bone Marrow Transplantation journal online.

Table 2.

Comparison of blood quantitative HIF-1a measures, serum IgG levels and exhaled NO concentrations in BO and non-BO-patients after allogeneic hematopoietic SCT Feature HIF-1a/ABL ratio IgG (g/L) FeNO (p.p.b.)

Patients; with BO

Patients w/o BO

Significance P

340 (n ¼ 40)

140 (n ¼ 20)

0.007

5,5 (n ¼ 45) 14 (n ¼ 45)

9,1 (n ¼ 32) 20 (n ¼ 30)

0.033 0.002

Abbreviations: FeNO ¼ fraction of exhaled nitric oxide, HIF-1a ¼ hypoxiainducible factor 1 alpha, p.p.b. ¼ parts per billion.

Bone Marrow Transplantation (2013) 1224 – 1229

Survival estimate

0

0.75

0.50

0.25

0.00 0

50 100 150 Time after transplantation (months)

200

Figure 2. Predicted OS of patients with and without BO from landmark month 18 after transplantation. Kaplan–Meier analysis was used to predict OS in all 68 patients with BO (solid lines) and 581 hematopoietic SCT-recipients without BO (dashed lines). P ¼ 0.043. The 95% CI for all values are indicated by upper and lower solid and dashed lines. A full color version of this figure is available at the Bone Marrow Transplantation journal online. & 2013 Macmillan Publishers Limited


1227

Survival estimate

1.00

0.75

0.50

0.25

0.00 0

50 100 150 Time after transplantation (months)

200

Figure 3. Predicted OS of female and male patients with BO. Kaplan– Meier analysis was used to predict OS in 25 female patients with BO (solid lines) and 43 male patients with BO (dashed lines). P ¼ 0.04. The 95% CI for all values are indicated by upper and lower solid and dashed lines. A full color version of this figure is available at the Bone Marrow Transplantation journal online.

donor-recipient sex constellation. The median follow-up after diagnosis of BO was 25 (range 4–118) months and the overall median follow-up of surviving hematopoietic SCT recipients was 64 (range 6–126) months. Of seven relapses detected among BO patients five occurred before and only two after BO manifestation. Five-year cumulative incidence of relapse was 26% (95% CI 24–29%) in non-BO and 10% (95% CI 5–20%) in BO patients. DISCUSSION BO remains difficult to diagnose and once clinical symptoms develop airway obstruction and small airway disease often are significant and irreversible. As we could have shown that the occurrence of BO is influenced by T-cell depletion our analysis included only hematopoietic SCT recipients conditioned myeloablatively without MoAb.19 In this homogenous patient cohort with comparatively long patient follow-up, we found a BO prevalence of 10% which was twice as high as recently reported, whereas the prevalence of 12% among patients with chronic GVHD nearly was identical.10 The 5-year cumulative incidence of 5.8% for BO was consistent with previous studies as well as the median time of BOonset after hematopoietic SCT inclusive of occasional late-onset BO-diagnoses.7,9,20 Using landmark analysis we could corroborate findings by Nakaseko et al.,9 who reported on a significantly impaired survival probability for patients with BO after hematopoietic SCT. Furthermore, we could confirm published data on diminished relapse rates among hematopoietic SCT-recipients who developed BO3 supporting the concept of a sufficient GVL effect. As suggested by an Center for International Blood and Marrow Transplant Research study reporting a threefold increase for BOrisk after PBSC transplantation20 we also found significantly higher incidence rates for BO after PBSC transplantation compared with BMT. This might be explained by a higher frequency of chronic GVHD after PBSC transplantation.21 Chronic GVHD again was demonstrated in the present study to be a major risk factor for the development of BO as described in nearly all studies on BO before.3,5,7–9,20,22 In contrast, ABO blood group incompatibility, a pretransplant factor that remained largely disregarded up to now could be confirmed to be associated with BO development in the present analysis. Different patterns of ABO compatibility according to the graft source are reproducible. For PBSC transplantation no ABO & 2013 Macmillan Publishers Limited

barriers exist, whereas major and bidirectional incompatible transplantation of BM is only possible after isoagglutinin reduction or graft processing by erythrocyte depletion. This different modus operandi has not only impact on donor selection but could also explain why ABO incompatibility is of relevance for BO development after PBSC transplantation but not after BMT. It is conceivable that ABO-incompatible transplantation might result in several immunohematological interactions due to Abmediated responses.23 These might initiate allogenic-immune mechanisms, which finally lead to structural changes and tissue fibrosis. In this context, BO-promoting post-transplant IgGdeficiency could be caused by such deregulated humoral immune responses. Chronic GVHD-associated dysgammaglobulinemia24 might additionally be suggested as an etiological factor for decreased serum IgG. However, although the association between lower circulating IgG levels and BO has been well known for nearly 25 years, very few findings with respect to the pathomechanisms and possible causes exist.25 The fact that the severity of BO did not significantly differentiate with respect to the survival probability can be explained by the low number of patients classified as having mild BO. As our survival analyses among BO patients reveal superior outcome for female hematopoietic SCT recipients with BO compared with male BO patients this observation may have significant clinical implications. As a consequence particularly male patients who are to receive PBSC transplantation from ABO blood group incompatible donors should be considered to be preferentially conditioned with anti-thymocyte globuline in order to prevent severe chronic GVHD with lung involvement.26 Regarding survival advantage for female BO patients we can only speculate on reasons. However, there are certain findings on gender differences in pulmonary diseases discussing an involvement of female hormones.27 Among chronic obstructive pulmonary disease patients, a beneficial effect of inhaled corticosteroids was especially noticeable in women28 and estrogen receptor-alpha was identified as a potential drug target preventing from lung inflammation.29 Even changes in exhaled NO concentrations due to cyclic estrogen and progesterone changes were reported.30 NO is a small molecule synthesized by NO synthases that is present in human breath.31 Many physiological and pathological factors can determine exhaled NO levels.32,33 Concentrations of exhaled NO are easily detectable by non-invasive measures and as an association between BO syndrome after lung transplantation and the levels of fractional exhaled NO was reported34 we tried to identify any changes regarding FeNO in BO after hematopoietic SCT. Surprisingly, in contrast to the Munich Lung Transplant Group we found decreased levels of FeNO in our BO-patients and regardless of the time point during the course of the disease it was possible to differentiate between BO- and non-BO-patients post-hematopoietic SCT by using a threshold level for FeNO. This clearly demonstrates that despite clinical analogies BO syndromes differ considering the underlying pathomechanisms. NO release, increasing severity of hypoxia due to edema, malperfusion and increased oxygen demand are considered as physiological responses to an inflammatory setting. BO after allogeneic hematopoietic SCT obviously is not characterized by acute inflammatory processes, in particular not by eosinophilic airway inflammation because otherwise FeNO levels would have been assessed higher than in the control groups. Previous studies could highlight NO having a critical role in preserving an epithelial phenotype and in attenuating epithelial-mesenchymal transition in alveolar epithelial cells.35 Furthermore, several studies have suggested a beneficial impact of NO on pulmonary disorders.36–38 It might be concluded that vice versa a decrease of NO may result in an elevated susceptibility to lung fibrosis. In the present study, we observed a significant negative correlation between HIF-1a expression and FeNO concentrations. Bone Marrow Transplantation (2013) 1224 – 1229


1228 The relationship between NO and HIF-1a is complex and still not completely understood. Increased expression of HIF-1a indicates a response either to hypoxia or chronic NO deficiency in normoxia. Recently, Cattaneo et al.39 concluded that NO deficiency may affect the activity of prolyl hydroxylases, which are inhibited under hypoxic conditions leading to HIF-1a accumulation. HIF-1a is the main factor involved in the regulatory transcriptional responses to hypoxia.40 However, chronic hypoxia itself may promote tissue fibrosis via HIF-1 stimulation, at least in renal fibrosis due to chronic kidney disease.13 As HIF-1a is a key regulator in the cellular adaptation to hypoxia it can be suggested that HIF-1 accumulates during initial BO manifestation consecutively leading to enhanced tissue fibrosis and further aggravation of BO. In conclusion, the present study verified increasing incidences and prevalence rates for BO among long-term survivors after hematopoietic SCT. Peripheral hematopoietic cells as stem cell source, chronic GVHD and low IgG levels were reconfirmed as risk factors associated with BO-development. In addition, for the first time ABO incompatible hematopoietic SCT could be demonstrated to be of impact for BO. Furthermore, survival among hematopoietic SCT recipients with BO was shown to be superior in female patients. Finally, we found evidence for a HIF-1a/NO system involved in the origination or preservation of BO post allogenic hematopoietic SCT. These findings may establish new approaches in diagnosing, monitoring, treating and exploring an insidious late complication after hematopoietic SCT that significantly deteriorates and reduces life expectancy.

CONFLICT OF INTEREST The authors declare no conflict of interest.

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