Complications In Sickle Cell Disease Patients Research Articles

NOS3, ET1, TGF-β1, and THBS1 polymorphisms modulating clinical complications in sickle cell disease patients

Clinical manifestations are highly variable in sickle cell disease (SCD) and polymorphisms (SNP) may modify vascular homeostasis. Because of this, we investigated of SNP of NOS3, ET1, TGF-β1 and THBS1 genes in vascular complications of SCD. High risk of hospitalization was found for genotypes NOS3TC/CC (rs2070744 - RR: 1.94) and THBS1AG/GG (rs1478604 - RR: 1.59) while low risk to NOS3GT/TT (rs1799983 - RR: 0.53). Hydroxyurea treatment was more frequent in the genotypes NOS3TC/CC (rs2070744 - RR: 1.83), ET1GT/TT (rs5370 - RR: 2.67) and ET1TG/GG (rs1800541 - RR: 1.43). Alleles ET1TG/GG (rs1800541) showed correlation for abdominal, arm and leg pain, bone disorders, cholelithiasis and vaso-occlusive crisis, while ET1GT/TT (rs5370) with chest pain, painful episodes, splenic sequestration and stroke. Interestingly, the NOS3GT/TT (rs1799983) alleles had a high stroke risk (RR:4.21), though, protection against bone disorders, cholelithiasis and less risk of hospitalization. We believed that our results indicate polymorphisms that may provide benefits in the treating Clinical Complications in SCD Patients, especially in vascular SCD pathophysiology, modulating vaso-occlusion crisis.

5609968 REAL-WORLD PATIENT-REPORTED OUTCOMES AND WEARABLE METRIC CORRELATIONS: REMOTE DAILY MONITORING FOR EXPLORATION OF QUALITY OF LIFE IN SICKLE CELL DISEASE

Background/Aims: Sickle Cell Disease (SCD) is a debilitating condition that lacks data on its natural history and quality of life (QoL) beyond limited snapshots through infrequent healthcare contacts. This lack of longitudinal targeted QoL measurement and understanding of the physiological factors that may influence it sees a need for more detail on the impacts key metrics have on patient outcomes. The core objective of this study was to generate real-world evidence tracking changes in live wearable metrics and patient QoL, to better characterise a baseline of key correlations in SCD. Methods: Data was collected from 238 patients over 16 months, using a CE and FDA-cleared wearable (Withings ScanWatch) to automatically record metrics including sleep quality, activity, and heart rate. EQ-5D-5L and pain scores (0-10) were manually entered by patients via a digital patient-reported outcomes (PRO) portal, and a daily cohort average across each metric was calculated. Statistically significant correlations were analysed both at an overall day-to-day level and split by weekday vs. weekend to explore the potential impacts of activities such as work on sleep patterns. Patients provided informed e-consent to participate and for analysis of their de-identified data. Results: Increasing daily EQ-5D-5L scores were found to correlate with increasing numbers of sleep disturbances (p=0.024) and decreasing pain scores (p<0.001). In turn, lower pain scores were associated with lower sleep heart rates (p<0.001) and wake-up durations (p<0.001), as well as lower active heart rates (p<0.001) and higher activity levels (p<0.001).While sleep quality (sleep disturbances) appeared to show a contradictory increase with higher EQ-5D scores, the significance of this correlation was lost following a split by weekdays and weekends. Statistical significance was also lost for the link between pain scores and steps during the weekend, whereas an additional correlation between pain and total sleep duration (p=0.023) was found only during weekdays, and between pain and the number of sleep disturbances (p=0.033) during the weekend. Similarly, a correlation between EQ-5D and active heart rate (p=0.022), as well as duration to wakeup (p=0.017), was seen only during the weekend. Summary-Conclusion: Analysis of putative associations between PROs and real-time metrics, as well as the potential influences of activities such as work, were shown by changes in significance both before and after split by weekdays/weekends. Notably, lower EQ-5D scores were found to correlate with higher levels of pain, in turn linking to lower daily steps. With a correlating increase in both active and sleep heart rates despite these lower activity levels, this may indicate higher detectable stress correlating with increased pain, with paralleled impacts on QoL. As such, these metrics may form points of monitoring when identifying or predicting risks of pain and related complications in SCD patients. Future work will explore potential causality, differences across patient groups, and more granular detail of the impact of specific treatments or healthcare events. Additional investigation into changes at an individual patient level will be conducted to better characterise deviations from patients’ own baselines. This will support ongoing work to improve the understanding of baselines in SCD, and what the key factors in pain and quality of life may be in order to support patient outcomes improvement.

Individual Red Blood Cell Fetal Hemoglobin Concentrations Determined By Imaging Flow Cytometry and the Number of F Cells Are Very Good Predictors of Clinical Response to Increased Level of HbF and Are Associated with Hemolysis, Tricuspid Regurgitant Jet Velocity, and Sickle Retinopathy in Adult Sickle Cell Disease Patients

Sickle cell disease (SCD) patients with similar total fetal hemoglobin (HbF) levels may present with different severity, which is attributed to the heterogeneous distribution of HbF among red blood cells (RBCs) with varying percentages of circulating HbF-containing RBCs (% F cells). Steinberg et al. (2014) proposed that an individual HbF concentration per cell (iHbF) above 10pg inhibits the polymerization of HbS, so the development of precise methods to quantify iHbF and the percentage of F cells may allow clinicians to use those parameters as targets to monitor patient treatment. We devised a novel procedure to very accurately measure and quantify iHbF using imaging flow cytometry (Amnis Imagestream) and investigated its correlations with clinical manifestations. Samples from 54 adult SCD patients (22 [41%] female, 39 [72%] on hydroxyurea) were used to acquire images from RBCs labeled with fluorescent anti-HbF antibody and were analyzed using IDEAS software v 6.2. Along with the hematimetric data and HPLC quantification of total HbF of each sample, we modified the formulas by Maier-Redelsperger et al. (1994) and Horiuchi et al. (1995), with minimization of interference from autofluorescence of non-F cells, correction for area and fluorescence intensity, and analyzed RBC subpopulations with varying levels of HbF expression. Statistical analysis was performed using R considering a p value <0.05 to be statistically significant. We did not find associations of iHbF with alpha thalassemia carrier state or beta globin cluster haplotypes Benin or Bantu, the most common in our population. Perpendicular cutoff correlation studies defined two patient populations: one group with lower mean iHbF (9.9pg) and lower % F cells (28.2%) and another group with a higher mean iHbF (15.8pg) and higher mean % F cells (72.5%). The latter group had a lower frequency of vaso-occlusive crises (p=0.011). That was further validated with our finding that the cutoffs of % F cells>30% and iHbF>10pg were strongly associated with lower absolute reticulocyte counts (p<0.001), LDH (p=0.002), and indirect bilirubin (p<0.001). The association between hemolysis and iHbF was further explored regarding pulmonary hypertension, showing that a value of iHbF>10pg was associated with a lower median tricuspid regurgitant jet velocity than in patients with iHbF<10pg (2.72m/s vs. 2.29m/s, p=0.046). We also found an association of a mean iHbF<8pg with the presence of sickle retinopathy (p=0.002). Patients with retinopathy had a lower mean iHbF and % F cells than those without it (7.08pg vs. 8.75pg, p=0.032; 48.5% vs. 71.0%, p=0.043, respectively), suggesting increments in iHbF and % F cells may be protective against sickle retinopathy, a complication not usually associated with high hemolytic rates. Our results suggest that a percentage of F cells above 30% along with individual RBC HbF concentration of 10pg may be protective against complications in SCD patients and, therefore, should be explored as goals for hydroxyurea therapy in this population. In addition, the data presented here suggest further studies on the efficacy of HbF induction therapy against the development of sickle retinopathy may be performed in a large population of sickle cell disease patients.

Neutrophils as drivers of vascular injury in sickle cell disease.

While neutrophils are the main effectors of protective innate immune responses, they are also key players in inflammatory pathologies. Sickle cell disease (SCD) is a genetic blood disorder in which red blood cells (RBCs) are constantly destroyed in the circulation which generates a highly inflammatory environment that culminates in vascular occlusions. Vaso-occlusion is the hallmark of SCD and a predictor of disease severity. Neutrophils initiate and propagate SCD-related vaso-occlusion through adhesive interactions with the activated and dysfunctional endothelium, sickle RBCs, and platelets, leading to acute and chronic complications that progress to irreversible organ damage and ultimately death. The use of SCD humanized mouse models, in combination with in vivo imaging techniques, has emerged as a fundamental tool to understand the dynamics of neutrophils under complex inflammatory contexts and their contribution to vascular injury in SCD. In this review, we discuss the various mechanisms by which circulating neutrophils sense and respond to the wide range of stimuli present in the blood of SCD patients and mice. We argue that the central role of neutrophils in SCD can be rationalized to develop targets for the management of clinical complications in SCD patients.

Incidence of Stroke and Other Brain Imaging Findings in Adult Patients with Sickle Cell Disease: A Retrospective Study

Background: Vaso-occlusion and hemolytic anemia are the main features of sickle cell disease. Central Nervous system complications of sickle cell disease (SCD) can include silent cerebral infarcts (SCI), hemorrhagic or ischemic strokes, seizures and Moyamoya syndrome. SCI is the most common neurological complications in SCD. SCI patients have abnormal Magnetic Resonance Imaging (MRI) of the brain with normal neurological examination and no history of stroke. More precisely, Casella et al in the Silent Cerebral Infarct Multi-Center Clinical trial (SIT) defined infarct as MRI signal abnormality that was at least 3 mm in one dimension and that was visible in two planes on fluid-attenuated inversion recovery T2 -weighted images, as determined by agreement of two of the three study neuroradiologists. SCI in children is associated with lower Intelligence Quotient and increased risk of stroke.Methods: In this retrospective study we reviewed 100 adult patients at our Comprehensive Sickle Cell Center. We looked at the incidence of documented stroke and other neurological complications in SCD patients. We reviewed the hospital records of SCD patients for any incidental brain imaging (MRI brain or Computed Tomography(CT) brain) findings, done as inpatient or in the emergency room as part of hospital visits.Results: Out of the 100 patients, 14 patients had documented stroke. On the review of other 86 patients, 10 patients had MRI brain, 23 patients had CT brain, and 53 patients had no brain imaging record. Out of the 10 patients with MRI brain, 7 patient had significant findings- 3 patients had small vessel ischemic changes, 1 patient with chronic petechial hemorrhage of cerebellar vermis, 1 patient with ischemic changes and Chiari malformation type 1 (CTM 1), 1 patient with CTM 1 and non-specific white matter changes and 1 patient with CTM 1 alone. Out of the 22 CT of the brain and 1 CT angiogram of brain, 2 imaging showed ischemic changes and the angiogram showed large cerebral vessel narrowing. Therefore, on our review, 19 patients out of 100 were found to have stroke and 24 out of 100 had significant neurological findings. However, only 14 out of 19 patients had documented stroke. 7 out the 10 patients with MRI brain had significant incidental neurological findings. The number of adult SCD patients with neurological findings in our cohort could have been higher, if we had brain imaging record for the 53 other patients.Conclusion: This study shows that there is a heavy burden of neurological complications in SCD patients and SCI is under-diagnosed. MRI brain is superior to CT brain in detecting neurological complications in SCD patients. SIT showed that regular blood-transfusion significantly reduced the recurrence of cerebral infarct in children with SCI. Finding of SCI in adults also warrants consideration for follow up brain imaging, neurology referral and discussion of treatment modalities including chronic blood transfusion and hydroxyurea therapy. CTM 1 can also be associated with SCD. In CTM 1, tonsil herniates 5 millimeter or below foramen magnum. SCD may cause increased thickness of calvarium from extramedullary hematopoiesis in skull, which can result in cranio-cephalic disproportion with cerebellar ectopia and subsequent CTM 1. Because of high incidence of neurological complications in SCD patients, routine screening with MRI brain should be considered in adult SCD patients. DisclosuresIdowu: Forma Therapeutics, Inc.: Research Funding; Pfizer: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Ironwood: Research Funding.

Inhibition of Factor XII Attenuates Prothrombotic Complications in Sickle Cell Mice

Sickle Cell Disease (SCD) is the most common inherited hemoglobinopathy, affecting millions worldwide. Although characterized by chronic hemolytic anemia and recurrent vaso-occlusive episodes, SCD is increasingly recognized as a hypercoagulable state. Indeed, SCD patients have an 11-25% incidence of venous thromboembolism at a median age of 30 years, associated with a 3-fold increased risk of mortality. Moreover, ischemic stroke and silent cerebral infarctions occur in 7-13% of SCD patients. We have previously shown that tissue factor, an initiator of the extrinsic coagulation pathway, contributes to thrombo-inflammation and microvascular cerebral thrombosis in mouse models of SCD . Recently, the intrinsic coagulation pathway, including Factor XII (FXII), has received significant attention because targeting components of this pathway reduces thrombosis without affecting primary hemostasis. We have shown that FXII deficiency reduces plasma markers of thrombin generation and inflammation in sickle mice. However, the contribution of FXII to thrombosis and prothrombotic complications in SCD is not known. In this study we evaluated the effects of blocking FXII activity on venous thrombosis and ischemia/reperfusion (IR)-induced brain injury in SCD mice.First, Townes HbSS mice (SS) and non-sickle Townes HbAA controls (AA) (male and female, 16 weeks) received anti-FXII antibody or control IgGκ1 (10 mg/kg, IV) 30 minutes prior to subjecting them to venous thrombosis, initiated by applying positive current (3 volts, 90 sec) to the femoral vein. To visualize platelet and fibrin accumulation, mice were injected with rhodamine 6G and anti-fibrin antibody 59D8 labeled with Alexa Fluor 647, respectively. The femoral vein thrombi were imaged by intravital fluorescence microscopy using time-lapse capture every 10 seconds, to acquire images of fibrin and platelets over 60 min. The accumulation of platelets and fibrin was quantified for relative intensity of each fluorophore over the region of the observed thrombus. As previously shown, thrombi of SS/IgG mice showed an increased fibrin and platelet accumulation compared to AA/IgG group. Importantly, 15D10 treatment significantly attenuated both fibrin (p<0.001) and platelet (p<0.05) deposition over time in SS mice compared to SS/IgG group. The same effect of 15D10 treatment was observed in AA mice. At the end of experiment, clots were collected and stained with hematoxylin and eosin, and clot volume was assessed histomorphometrically (Nikon Ti-2, FIJI Software). Surprisingly, despite higher fibrin content, clots from SS/IgG mice had significantly smaller volume than clots from AA/IgG group (0.32 ± 0.04 versus 0.60 ± 0.11 mm 3, p<0.05). Importantly, administration of 15D10 significantly reduced clot volume in both SS (0.086 ± 0.01 mm 3, p<0.05) and AA mice (0.1 ± 0.02 mm 3, p<0.05).Next, AA and SS mice (male and female, 8-10 weeks) were subjected to brain IR injury induced by middle cerebral artery occlusion for 60 minutes followed by 24 hours of reperfusion (mouse model of ischemic stroke). 15D10 or control IgGκ1 (10 mg/kg, IV) were injected 30 minutes before occlusion and again at 6 hours into the reperfusion period to generate 3 experimental groups: AA/IgG, SS/IgG and SS/15D10. All analyzed parameters of brain IR injury were significantly worse in the SS/IgG group compared to the AA/IgG group. Compared to IgG, pre-treatment of SS mice with 15D10 significantly attenuated neuronal damage determined by volume of brain infarction (11.7 ± 3.7 vs 24.9 ± 2.4%, p<0.001) and improved behavioral deficit assessed by mean stroke score (9.0 ± 0.9 vs 14.6 ± 0.9, p<0.01). These changes were accompanied by a significant increase in leukocytes rolling (1978.0 ± 93.5 vs 1517.0 ± 180.3 rolling leukocytes/sec/mm 2, p<0.001), and significant reduction in the number of adherent leukocytes (367.2 ± 49.0 vs 723.4 ± 48.5, adherent leukocytes/mm 2, p<0.001) observed in the brain microvasculature of SS mice treated with 15D10 compared to SS/IgG group.Together, our data indicates that in the mouse model of SCD FXII contributes to the experimental venous thrombosis and ischemic stroke. Given that targeting the intrinsic pathway can reduce thrombosis without affecting hemostasis, our data suggest that targeting FXII might be a beneficial treatment in reducing inflammatory and thrombotic complications in SCD patients without a risk of bleeding. DisclosuresWallisch: Aronora Inc,: Current Employment. Key: Grifols: Research Funding; Takeda: Research Funding; BioMarin: Honoraria, Other: Participation as a clinical trial investigator; Sanofi: Consultancy; Uniqure: Consultancy, Other: Participation as a clinical trial investigator. Gruber: Aronora Inc.: Current Employment, Current equity holder in publicly-traded company; Oregon Health and Science University: Current Employment.

Aggravated Dental and Periodontal Status in Patients with Sickle Cell Disease and Its Association with Serum Ferritin

The present study aimed to evaluate the possible consequences of sickle cell disease (SCD) on dental and periodontal health in middle-aged patients and to examine the association of certain cardiovascular parameters and serum ferritin with the dental and periodontal status. Thirty-seven patients (mean age 43.2 years old) with SCD and 30 non-SCD and otherwise healthy individuals (mean age 38.9 years old) were examined for caries experience and periodontal status in addition to cardiovascular characteristics and ferritin level in serum. Compared to controls, SCD patients exhibited higher plaque and gingival bleeding scores, higher prevalence of periodontal diseases, and higher caries experience. Multiple stepwise linear regression analysis showed that caries experience was predominantly determined by the presence of SCD and the age, while major determinants of periodontitis were the ferritin levels and the male gender. The results reveal an aggravation of oral health in SCD patients regarding both caries and periodontal diseases. A potential role of the increased central inflammatory response, reflected by the elevated ferritin level in serum, is suggested for the impaired periodontal health of SCD patients. Compliance with precautionary dental checks and early management of dental complications is of great importance in order to improve oral health status and prevent general health complications in SCD patients.

Predictors of dental complications post-dental treatment in patients with sickle cell disease.

ObjectiveOur aim was to explore potential medical or dental indicators associated with dental complications and the utilization of emergency services in sickle cell disease (SCD), especially that clinical reports on adverse outcomes post‐dental treatment are scarce.Materials and methodsA retrospective analysis of dental treatments of 47 eligible adults with confirmed SCD between May 2016 and October 2019. Logistic regression analysis was used whether clinical outcomes, course of dental treatment, and regularity of dental care are associated with dental complications after dental procedures and/or resulted in emergency care or hospital admissions.ResultsWe identified a new, statistically significant association (p‐value = .01) between the number of prescription medications taken and complications (10%) after dental procedures. The most frequent dental procedures were tooth extractions (36%) and pain management (28%) during a non‐scheduled dental encounter (68%). The majority of cases did not participate in regular recall exams and periodical oral hygiene maintenance.ConclusionsA higher number of prescription medications was associated with an increased risk of post‐dental complications in SCD patients. A thorough medical history, including a list of prescribed medications, and collaboration with the patient medical team are important to assess the risk of complications post‐dental procedures and the need for antibiotic prophylaxis according to the case complexity.

Inpatient Outcomes and 60-Day Unplanned Readmissions after Hip Arthroplasty in Sickle Cell Patients with Avascular Necrosis

Background: Avascular necrosis (AVN) afflicts up to 10% of patients with sickle cell disease (SCD), and results in significant impairment in mobility and quality of life. The femoral head is the most common site for involvement of AVN in SCD patients. Total hip arthroplasty has a higher rate of perioperative complications in SCD patients, when compared to the general population. In this study, we aimed to evaluate the rates of readmissions in SCD patients undergoing hip arthroplasty as well as examine the causes behind these readmissions. Methods: The Nationwide Readmissions Database (NRD) was queried to include all adults who underwent hip arthroplasty in the years 2016 and 2017. SCD patients with AVN of the hip were compared to those undergoing hip arthroplasty for other reasons. T-test was used to compare means of continuous variables and Chi-square test to compare proportions of categorical variables. Multivariable logistic regression was used to evaluate risk factors for inpatient mortality, increased length of hospital stays, and hospitalization charges. We also examined risk factors for readmissions and the most common causes of readmissions in both sickle cell and non-sickle cell cohorts. Results: A total of 1,167,675 hip arthroplasties were performed in 2016 and 2017, of which 0.1% were for SCD patients with AVN. As expected, those with SCD were younger (mean age 40.6 vs 68.2 years, p&amp;lt;0.0001), with an equal gender distribution (57.8% female) in the two groups. Sickle cell patients were more likely to be from lower income neighborhoods and were also more likely to be treated at teaching hospitals. After adjusting for demographic variables and comorbidity burden, sickle cell disease was associated with higher risk of inpatient mortality (aOR 9.50, 95% CI 2.39-37.77, p&amp;lt;0.01), longer length of hospital stay by 3.2 days (95% CI 2.5-3.9 days, p&amp;lt;0.001), and higher hospitalization costs by roughly 20,000 USD per admission (95% CI 12,000-29,000 USD, p&amp;lt;0.001). On multivariate analysis, sickle cell patients were more likely to be readmitted within 60 days (aOR 5.25, 95% CI 4.01-6.87, p&amp;lt;0.001). 65% of these readmissions were due to SCD itself. Procedural complications resulted in a 60-day readmission rate of 3% in both the SCD and non-SCD groups, whereas, infectious complications accounted for a higher rate of readmission in those with SCD (2.4% vs 1.3%). Conclusions: Hip arthroplasty is associated with inferior inpatient outcomes and higher resource utilization in those with sickle cell disease. This can be attributed to high incidence of sickle cell crises post-procedure, as well as, intraoperative surgical complications (femoral fractures and perforations) and high rates of postoperative infections in the functionally asplenic sickle cell disease population. Disclosures No relevant conflicts of interest to declare.

Homocysteine is associated with severity of microvasculopathy in sickle cell disease patients.

The pathophysiology of sickle cell disease (SCD) includes vasculopathy as well as anaemia. Elevated plasma homocysteine is a risk factor for vascular disease and may be associated with increased risk of vascular complications in SCD patients. In the present study, microvascular characteristics were assessed in the bulbar conjunctiva of 18 paediatric and 18 adult SCD patients, using the non-invasive technique of computer-assisted intravital microscopy. A vasculopathy severity index (SI) was computed to quantify the degree of microvasculopathy in each patient. Plasma homocysteine and several of its determinants [serum folate and vitamin B12, plasma pyridoxal-5'-phosphate (vitamin B6 status) and creatinine (kidney function)] were measured. Age was strongly correlated with microvasculopathy in the SCD patients, with the SI increasing about 0·1 unit per one-year increase in age (P<0·001). After adjusting for age, gender, B-vitamin status and creatinine, homocysteine concentration was directly correlated with severity index (P<0·05). Age and homocysteine concentration were independent predictors of microvasculopathy in SCD patients. It remains to be determined whether lowering homocysteine concentrations using appropriate B-vitamin supplements (folate and vitamins B12 and B6) - particularly if started early in life - could ameliorate microvasculopathy and its associated complications in SCD patients.

Frontal bone infarctions masquerading as bilateral orbital cellulitis in a patient with sickle cell disease

Repeated vaso-occlusive crises (VOCs) are the hallmark of sickle cell disease (SCD). These repeated crises can lead to bone infarcts, necrosis, and, over time, degenerative changes in the bone marrow. Orbital complications in SCD patients are infrequent and usually present as orbital cellulitis. We report the appearance of orbital bone infarction intraoperatively in the case of an 18-year-old Saudi male patient who has been diagnosed with SCD and presented with severe headaches and generalized body aches. He was admitted with a case of SCD with acute VOC and started on the hospital sickle cell protocol. During the admission, the patient developed bilateral periorbital swelling and left inferior dystopia secondary to bilateral frontoparietal bone infarction, which was evident on the magnetic resonance imaging.

Spontaneous Healing of Avascular Necrosis of the Femoral Head in Sickle Cell Disease

Background. Avascular necrosis (AVN) is a serious complication of sickle cell disease (SCD) that can lead to significant morbidity including chronic pain and physical impairment with treatment depending on the patient's pain severity and functional limitation. Patient age, hemoglobin levels, and abnormal rheology, particularly high blood viscosity, or thickness, and percentage of dense red blood cells, defined as cells >1.11 mg/mL have been identified as risk factors for the development of AVN, but their contributions to the subsequent clinical course has not been described.The natural history of AVN has been described as chronic and progressive without significant surgical or physical therapy based intervention. However, this description was established before widespread MRI use in diagnosis, or hydroxyurea (HU). A new evaluation of the natural history of AVN is needed.Objective: Review the natural history of AVN at our institution in patients for whom repeat imaging is available, and determine if patient demographics, treatment type, and laboratory data, including rheological values, predict of clinical course.Methods. We identified all patients with a diagnosis of AVN with diagnostic and follow-up imaging studies (pelvic X-rays or MRI of the hip), receiving care at Texas Children's Hospital Hematology Center 2006 to 2018, and collected demographic, treatment, rheological and clinical laboratory data. Patient radiographic images at diagnosis and follow up were reviewed and scored by a board certified pediatric radiologist using the Steinberg criteria. Laboratory values for each patient were collected as available from time of diagnosis through to the date of follow up imaging. Longitudinal patient blood samples were collected for rheological measurements under an IRB approved protocol. Whole blood viscosity was measured on a cone and plate viscometer (DV3T Rheometer, AMETEK Brookfield, Middleboro, MA, USA) at 45 and 225 s-1 at 37oC within 4 hours of sample collection in an EDTA vacutainer tube. Percent dense red blood cells (%DRBCs) were measured by an ADVIA 120 Hematology System (Siemens Medical Solutions USA, Inc., Malvern, PA, USA). Statistical analysis comparing groups with and without radiographic improvement was performed using a two-tailed Student's t-test.Results. 16 patients had repeat imaging, allowing for assessment of radiographic improvement (Table1). All were diagnosed via X ray or MRI of the pelvis. Five of 16 patients had resolution or significant radiographic improvement of AVN without orthopedic intervention or prolonged physical therapy, while 11 patients had stable or worsening disease. In the group with radiographic improvement, the mean age was 9.9 years (SD=2.5) and 14.6 years (SD=2.2) in the group without radiographic improvement (p= 0.002). Five patients underwent core decompression with bone marrow aspirate concentrate injection; one progressed to complete femoral head collapse while the rest had stable or worsening disease. Mean hemoglobin (Hb), fetal hemoglobin (HbF), percent DRBCs, whole blood viscosity, and extent of AVN at diagnosis by Steinberg criteria were not statistically different between the patients that showed improvement and those that did not.Conclusions. Our results challenge the paradigm of untreated AVN in SCD as an inexorably progressive disease. Here we present 5 cases of significant radiographic improvement of AVN without surgical or significant physical therapy interventions. Conversely, 5 AVN cases treated with core decompression remained stable or worsened. Given our small cohort, we were unable to establish laboratory or rheological predictors of spontaneous resolution. However, patients in the radiographic improvement group were younger than patients with progressive disease. HU has shown promise in improving vascular complications in SCD patients however more research is needed to delineate its role as a modifier of AVN natural history as it was not correlated with the improved group in our study (all our study patients were on HU either prior to diagnosis or started during the follow up period). Given the findings that significant radiographic improvement is possible with minimal or no intervention, providers should closely weigh the risks and benefits of observation versus surgical intervention, particularly in young patients with SCD. DisclosuresNo relevant conflicts of interest to declare.

Specific Inhibition of AKT with ARQ 092, an Orally-Available Selective AKT Inhibitor, Attenuates Acute Vaso-Occlusive Events in Sickle Cell Disease

▪Previous studies have identified the Ser/Thr protein kinase, AKT, as a therapeutic target for thromboinflammatory diseases. Recently, we have shown that the basal levels of AKT phosphorylation were significantly increased in neutrophils and platelets isolated from sickle cell disease (SCD) patients compared with those cells from healthy donors and that specific inhibition of AKT2 impaired neutrophil-endothelial cell and neutrophil-platelet interactions in venules of Berkeley (SCD) mice (Li et al. J Clin Invest 2014; Barazia et al. Blood 2015). Although our results suggest that AKT2 inhibition may be a novel therapy to treat a vaso-occlusive crisis in SCD patients, no AKT2 specific inhibitor is available in clinic. Using ARQ 092, an orally-available, allosteric AKT inhibitor that is currently in Phase I clinical trials in oncology indications and AKT-driven overgrowth disease, Proteus syndrome, here we report that specific inhibition of AKT attenuates the adhesive function of neutrophils and platelets from sickle cell disease (SCD) patients in vitro and heterotypic cell-cell interactions in the Berkeley mouse model of SCD in vivo. Studies using neutrophils and platelets isolated from SCD patients revealed that treatment with 50-500 nM ARQ 092 significantly blocks αMβ2 integrin function in neutrophils and reduces P-selectin exposure and glycoprotein Ib/IX/V-mediated agglutination in platelets. Consistently, treatment of isolated platelets and neutrophils with ARQ 092 inhibited heterotypic cell-cell aggregation under shear conditions. Using in vivo real-time fluorescence intravital microscopy, we demonstrated that short-term oral administration of ARQ 092 (100 mg/kg body weight) or hydroxyurea (HU, 250 mg/kg body weight), a main therapy for the treatment of SCD, diminishes neutrophil-endothelial cell and neutrophil-platelet interactions in venules of Berkeley mice challenged with TNF-α to induce a vaso-occlusive crisis. Co-administration of HU and ARQ 092 further reduced the adhesive function of neutrophils in venules and neutrophil transmigration into alveoli, inhibited the expression of E-selectin and intercellular adhesion molecule-1 (ICAM-1) in the cremaster vessel wall, and improved survival in these mice. Ex vivo studies in Berkeley mice suggested that co-administration of HU and ARQ 092 efficiently blocks neutrophil and platelet activation and reactive oxygen species generation. The synergistic effects of both inhibitors resulted from increased NO production by HU and specific inhibition of AKT by ARQ 092. Our results provide important evidence that ARQ 092 has potential as a novel drug to be developed to prevent or treat the acute vaso-occlusive complications in SCD patients. DisclosuresAbbadessa:ArQule, Inc: Employment. Yu:ArQule, Inc: Employment. Schwartz:ArQule, Inc: Employment.

Central Venous Access Device (CVAD) Use and Complications in Sickle Cell Disease Patients from Medicaid and Commercially-Insured U.S. Populations

Central venous access devices (CVADs), such as tunneled central venous catheters (tCVADs) and peripherally inserted central catheters (PICCs), help provide essential care for some patients with sickle cell disease (SCD). CVADs facilitate administration of multiple intravenous (IV) medications and blood products, as well as blood draws for laboratory analysis. Understanding CVAD use and complications is particularly relevant for SCD patients because of their high risk of having insufficient peripheral IV access. Prior studies describing CVAD use and complications in SCD patients were limited by small sample sizes, typically including 15-20 SCD patients in a single treatment center. The resulting estimates for CVAD use and complications in SCD patients vary widely, are insufficient for development of evidence-based guidelines, and may not be representative of the treatment burden in the broader SCD population. The purpose of this study was to describe the frequency of CVAD use and CVAD-associated complications among SCD patients in a large US population sample.We used data from two large U.S. insurance claims databases from Truven Health MarketScan® Research to examine both Medicaid-insured and commercially-insured SCD patients. From January 2009 through December 2013, these databases encompassed over 14 million Medicaid-insured and over 116 million commercially-insured patients. SCD patients were defined as patients with at least two International Classification of Disease-9 (ICD-9) diagnosis codes for SCD (282.41-42, 282.6x) on separate dates in excess of sickle cell trait codes, or one ICD-9 code for an inpatient (emergency department or hospitalization) visit with sickle cell crisis. CVAD insertions were identified using relevant procedure codes for tCVADs or PICCs. We conducted two sets of analyses for each database: (1) per patient among those with at least one CVAD insertion, and (2) per CVAD insertion. We conducted descriptive analyses on the frequency of CVAD-related procedures (e.g., repair, replacement, removal of obstructive material, repositioning), complications, and infections, thromboses, or phlebitis.A total 17,119 Medicaid-insured SCD patients and 21,342 commercially-insured SCD patients were observed for an average of 3-4 years, during which 1,945 (11.4%) and 1,316 (6.2%) patients, respectively, had at least one CVAD insertion. Most SCD patients (80%) were aged >18 y at time of first CVAD insertion; 18% of adult Medicaid SCD patients had at least one CVAD inserted. The mean number of CVAD insertions per patient was 3.1 (Medicaid) and 2.4 (commercially-insured). In the per CVAD analysis, complication claims were frequent, including infection (31-37%), thrombosis (4-5%), and phlebitis/thrombophlebitis (12-15%). The mean time to removal of CVADs (duration) was 31-34 days (PICC lines) and 102 days (tCVADs). In the per patient analysis, 54.3% had infection claims, 24.3% had thrombosis claims, and 10.2% had phlebitis/thrombophlebitis claims in Medicaid (see Table and Figure; additional results to be provided, also stratified by age).Both tCVADs and PICCs were commonly used in SCD patients, particularly adults, with high occurrence of infection, thrombosis and phlebitis/thrombophlebitis, as well as repeated CVAD insertions. Determinants of CVAD use and complications warrant further investigation to inform practice standards. These findings from a large observational study indicate that device-related risks of administering IV treatments are limitations of IV treatment options and may add to SCD treatment burden.Table 1Medicaid-InsuredCommercially-InsuredAny CVAD (Overall)PICC OnlytCVAD OnlyBoth PICC and tCVADAny CVAD (Overall)PICC OnlytCVAD OnlyBoth PICC and tCVADN (%)1,945681 (35.0%)893 (45.9%)371 (19.1%)1,316450 (34.2%)664 (50.5%)202 (15.3%)Age, mean (sd) y30.0 (16.3)34.2 (14.9)26.9 (17.5)30.1 (13.6)32.6 (17.1)34.1 (15.8)31.3 (18.3)33.2 (15.9)CVAD insertions per patient, mean (sd)3.1 (3.9)3.1 (3.9)1.8 (1.3)6.2 (5.8)2.4 (2.3)2.5 (2.6)1.6 (1.0)4.5 (2.9)CVAD insertions total6,1073,6512,456n/a3,0821,6361,446n/aDuration per CVAD, median days4431102n/a5334102n/aCVAD complications, % patients with CVADComplication, general28.8%13.2%31.8%50.1%22.3%9.1%25.9%39.6%Removal of obstructive material11.2%3.8%15.3%14.6%9.0%2.4%11.4%15.3%Replacement8.3%3.7%10.2%12.4%6.5%3.8%6.5%12.9% [Display omitted] DisclosuresMaserejian:Biogen: Employment, Equity Ownership. Hayflinger:Biogen: Consultancy, Employment. Eaton:Biogen: Employment, Equity Ownership. Madigan:Biogen: Employment, Equity Ownership. Hobbs:Biogen: Employment, Equity Ownership.

Blood Antioxidant Defenses and Sickle Cell Disease: Analyzing Cytochrome b Reductase 1 Protein Levels and Structure in Erythrocyte Membranes

Introduction: The pathology of sickle cell disease (SCD) includes ischemia / reperfusion events in the vasculature, compromised blood antioxidant defenses and elevated risk of cardiovascular complications. Vitamin C (ascorbate) is a major blood antioxidant; ascorbate inside red blood cells (RBC) furnishes an electron (e-) to recycle plasma ascorbate to its active, reduced state (Fig. 1). A key step, transmembrane e- transfer, involves cytochrome b reductase 1 (CYBRD1). Thus, a deficiency or defect in red cell CYBRD1 could plausibly increase oxidative stress and interconnected inflammatory processes, contributing to cardiovascular complications in SCD patients. Testing for CYBRD1 deficiency requires a quantitative assay for the protein in RBC, so we developed a suitable assay and used it to screen a small number of SCD patients and healthy individuals.Materials and methods: Blood samples were collected from steady state adult SCD patients, healthy African American adults, and steady state pediatric SCD patients before and 2-5 months on hydroxyurea (HU) therapy. Protocols were approved by institutional review.The CYBRD1 assay workflow is shown in Fig. 2. RBC membrane proteins were separated on two SDS-PAGE gels, along with a mixed standard (1.0 µg bovine serum albumin (BSA) + 1.9 ng of His6-tagged recombinant human CYBRD1 (rCYBRD1; PMID 21501687)). One gel was Coomassie-stained for quantitation of total membrane protein minus membrane-bound hemoglobin (Hb) (standard: BSA). Proteins in the other gel were transferred to a nitrocellulose filter, probed with CYBRD1 antibody (GeneTex #104793) and visualized with secondary antibody and chromogenic substrate for quantitation of CYBRD1 (standard: rCYBRD1), which was normalized to membrane protein. Gels, blots and calibrated density step tablets were scanned (transmittance mode) with an Epson V550; digitized images were analyzed using ImageJ. Two independent samples and paired t tests were used to compare between- and within-group mean differences in RBC CYBRD1 levels, respectively.Results: The Coomassie-stained gels exhibited the usual profile of RBC membrane proteins (Fig. 2). In the immunoblots, the rCYBRD1 standard showed the expected narrow monomer band at ~30 kDa and faint dimer band at ~60 kDa (Fig. 2). However, RBC membrane samples revealed several CYBRD1 monomer bands, indicating the presence of previously unsuspected isoforms of CYBRD1. Further, the CYBRD1 isoform pattern in RBC from healthy controls was quite different from those seen in SCD patients. The functional impact of these differences remains to be investigated.Assay of progressively diluted control RBC membranes produced a linear response for CYBRD1 from 0.7-7.2 ng (r2 = 0.975) and for total protein from 1 to 10 µg (r2 = 0.993). The sub-nanogram detection of CYBRD1 protein makes the assay usable with as little as 0.25 ml of blood. The coefficient of variation was ~15%, quite reasonable precision for quantitative immunoblotting.CYBRD1 levels in RBC from healthy African American adults averaged 0.68 ± 0.13 ng/µg membrane protein (N1 = 10; Fig. 3). CYBRD1 levels in RBC of adult SCD patients averaged 0.35 ± 0.14 ng/µg membrane protein (N2=7). This difference in means was highly statistically significant by two-tailed t-test (p < 0.0002), supporting our hypothesis that a deficiency in RBC CYBRD1 contributes to elevated oxidative stress in SCD patients.RBC CYBRD1 levels in pediatric SCD patients not on HU therapy averaged 0.27 ± 0.14 ng/µg protein (N3=12; Fig. 3). Pediatric patients on HU therapy averaged 0.41 ± 0.18 ng CYBRD1/µg membrane protein (N4=7). For the five pediatric patients examined both before and during HU therapy there was a marginally significant increased mean CYBRD1 level on HU (p < 0.099 by paired, two-tailed t test).Conclusions: Our newly developed assay allows quantitation of the RBC CYBRD1 protein level and analysis of the CYBRD1 isoform pattern using a small blood sample. The pilot screening results suggest that adult SCD patients have lower levels of CYBRD1 in their RBC, and a different pattern of CYBRD1 isoforms. Analysis of the CYBRD1 protein may be useful in assessing the state of individual SCD patients' blood antioxidant defenses and the antioxidant effects of HU. Thus, it will be worthwhile to examine the RBC CYBRD1 protein and activity in larger numbers of patients and controls, and to explore the origins of any differences between the groups. [Display omitted] [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Thrombospondin-1 Stimulates Calcium Influx and Echinocytosis in Sickle Cell-Derived Red Blood Cells

Thrombospondin-1 (TSP1) is a secreted matricellular protein found preformed in platelet α-granules and upregulated in the plasma of patients with sickle cell disease (SCD). In preclinical models TSP1 enhances sickle red blood cell (RBC) binding to endothelial cells, while in murine sickle mice TSP1 promotes RBC echinocytosis and vaso-occlusion. Recently we reported elevated plasma TSP1 was associated with vaso-occlusive complications in SCD patients. Herein we tested the hypothesis that TSP1 compromises RBC membrane integrity via enhanced calcium influx leading to morphological changes, such as echinocytosis. RBC were collected via centrifugation of heparinized blood from human HbAA donors (n=3), SCD patients with homozygous HbSS SCD (n=4), and wild type C57BL/6J mice (n=2). Cells were then re-suspended in glucose and calcium-supplemented PBS, and treated with several concentrations of exogenous TSP1 (2.75 nM, 11 nM, 22 nM and 44 nM) for 2 hours at 37°C. RBC morphology was assessed by both light and electron microscopy. TSP1 modulation of RBC calcium influx was studied using the calcium fluorophore Fluo-3, AM. TSP1 treatment, in a dose-dependent manner, stimulated echinocytosis in human SCD and non-SCD RBC and in murine cells. In some instances electron microscopy showed echinocyte membrane projections with a string-of-beads appearance, suggestive of imminent microparticle shedding. TSP1 treatment, again in a dose-dependent manner, also significantly increased calcium influx in SCD (Figure, right panel) and non SCD RBC (Figure, left panel) and murine RBC (not shown). Interestingly, SCD RBC displayed more sensitivity to TSP1-mediated increases in cellular calcium as compared to normal cells. Our results show for the first time that TSP1, at doses found in patient plasma, stimulates significant echinocytosis and calcium influx in both normal and SCD RBC. These data identify a novel role for plasma TSP1 in promoting RBC pathology. Future studies are underway to elucidate this mechanism and its pathogenicity in humans with and without SCD. [Display omitted] DisclosuresIsenberg:Vasculox: Membership on an entity’s Board of Directors or advisory committees; Radiation Control Technologies: Membership on an entity’s Board of Directors or advisory committees.

Paper-Based Assay for Quantification of HbS in Blood of Sickle Cell Disease Patients

Introduction: Sickle cell disease (SCD) is a common inherited blood disorder caused by sickle hemoglobin (HbS) which, unlike normal adult hemoglobin (HbA), becomes insoluble and polymerizes under hypoxic conditions. Patients with SCD experience chronic hemolytic anemia, episodic pain crises and abnormal blood flow to critical organs that cumulatively result in significant illness and shortened lifespans for many. The severity of SCD varies significantly between patients, but for individuals the rate of adverse events is strongly correlated with intraerythrocytic concentration of HbS (%HbS). High per test costs and long turnaround times make conventional laboratory methods (e.g. Hb electrophoresis, HPLC, IEF) impractical for quantifying %HbS in real-time (e.g. during transfusion therapy). The objective of this study was to demonstrate that %HbS in blood could be quantified using our recently developed rapid, low-cost paper-based SCD assay [1].Methods: Blood samples were obtained from SCD and sickle cell trait (SCT) patients at the Texas Children’s Hematology Center (Houston, TX). To perform the SCD assay a 20μL droplet of blood mixed with Hb solubility buffer (1:10 by volume) was dropped on chromatography paper. The resulting blood stain was digitized with a flatbed scanner (Canon USA Inc, Melville, NY) and analyzed using a custom image analysis code (The MathWorks Inc, Natick, MA). Conventional Hb electrophoresis was performed with the semi-automated Sebia Hydrasys 2 Scan system (Sebia Inc, Norcross, GA).Results: The difference in transport of Hb through the paper produced a blood stain with two parts: the area of the initial drop where polymerized HbS is retained (center spot) and the area where soluble Hb is wicked laterally (peripheral ring). The relative color intensity of the center spot and peripheral ring is related to the blood sample %HbS (Fig. 1). The image analysis code automatically isolates and calculates the ratio of the average color intensities of each area (S-index). A series of reconstituted blood samples with artificially adjusted %HbS from 0 to 60% was used to calibrate the assay so that %HbS could be estimated based on blood stain color intensities (Fig. 2a). The values of %HbS estimated for patient samples using our paper-based SCD assay and actual values measured using conventional Hb electrophoresis were highly correlated with R2 = 0.898 (Fig. 2b). The estimated and actual %HbS values also showed strong agreement with the standard deviation of the difference between the two measurements = 5.5 %HbS (Fig. 2c). The majority of the differences between actual and estimated %HbS (96.67%) are within 2 standard deviations of the mean of the differences. The assay could be performed in under 35 minutes and multiple assays could be performed and analyzed in parallel. The cost of consumable materials and reagents for the paper-based SCD assay is less than $0.03.Conclusions: This study demonstrates the feasibility of using our recently developed paper-based assay to quantify %HbS in blood samples in real-time. The ability to rapidly, inexpensively measure %HbS will be particularly useful for monitoring the effectiveness of chronic transfusion or hydroxyurea therapy for long-term control of HbS content in blood of SCD patients. The ability to measure %HbS in real-time could also potentially facilitate more aggressive prophylactic therapy to intervene rapidly and significantly reduce the rate of life-threatening complications in SCD patients, including stroke. [Display omitted] [Display omitted] Acknowledgments: This work was supported in part by a 2012 NIH Director’s Transformative Research Award (NHLBI R01HL117329, PI: SSS).

Diurnal variations of C-reactive protein in trait and sickle cell disease patients

Sickle cell disease (HbSS) is the most common variant of β-globin gene, resulting from the replacement of glutamic acid by valine at position 6 in the β chain. Under the action of certain factors, such as cold, fever, dehydration, and infections, sickle cell crisis is usually precipitated. The crisis predisposes to aggregation of sickle cells, which may produce vaso-occlusion of capillaries, resulting in pain, swelling, and infarction. In response to inflammation, the liver releases a variety of acute phase proteins. C-reactive protein (CRP) is pentameric hepatocyte protein and is the measure marker of the “acute phase response,” or the formation of plasma proteins in response to an inflammatory stimulus in human. This study is an attempt to document diurnal variations in the level of CRP in sickle cell disease (SCD) patients in steady or crisis state. Saliva samples from SCD, sickle cell trait (SCT), and control subjects were collected continuously over a period of three days. C-reactive protein level was detected using RHELAX CRP Latex kit with the sensitivity of 0.6 mg/dl. When pooled data were subjected for Cosinor rhythmometry, it failed to detect sinusoidality among SCD patients suggesting a constant elevated level of CRP level due to inflammation. However, a statistically significant circadian variation among control and SCT in CRP levels was discerned. Result suggests that although the level of CRP in SCT is comparatively higher than the normal individuals, the CRP contents significantly fluctuate in the absence of chronic inflammation among traits. The rhythm-adjusted mean was found to be significantly higher in SCD (p ≤ 0.005) patients when compared to control and trait subjects. A constant higher level of CRP can also be an indicator of future cardiac complications in SCD patients.

Molecular Link between Intravascular Hemolysis and Vascular Occlusion in Sickle Cell Disease

Intravascular hemolysis is a major component of anemia in sickle cell disease (SCD). Plasma extracellular hemoglobin (ECHb) liberated by intravascular hemolysis has deleterious effects on the vasculature. ECHb scavenges nitric oxide (NO) and promotes the pathogenesis of several clinical events including pulmonary hypertension, priapism and non-hemorrhagic strokes. ECHb reduces the bioavailability of NO which down-regulates platelet activation, leading to platelet aggregation and vascular clot formation. Recently we have identified an additional mechanism whereby increased hemolysis can lead to a prothrombotic state in SCD by increasing the activity of von Willebrand factor (VWF), a multimeric plasma glycoprotein secreted by the endothelium. Our studies show that ECHb binds to the A2-domain on VWF at the proteolytic site of the metalloprotease, ADAMTS13, and blocks VWF cleavage in vitro. Elevated ECHb is associated with high levels of ultralarge and hyperactive VWF multimers in SCD patients' plasma. A sub-population of VWF multimers, bound to ECHb is hyperactive, and exists in greater quantity in SCD patients' plasma compared to normal controls. These results suggest a possible role for plasma ECHb in the accumulation of hyperactive VWF multimers in vivo that may mediate thrombotic and vasoocclusive complications in SCD patients.

Is Incentive Spirometry Effective in Preventing Post Surgical Acute Chest Syndrome in Sickle Cell Disease Patients?

Persons with sickle cell disease (SCD) are more likely to undergo cholecystectomy and splenectomy than are the general population. Even with meticulous care, surgical complications are seen in approximately 25% to 30% of patients. A chart review of SCD patients who had splenectomies and cholecystectomies and who were treated in 3 centers between1987–2007 was undergone. Data pertaining to clinical events, surgery, perioperative care and outcome were collected and analyzed. Since 1995, laparoscopic surgery was the modality used. Management consisted of preoperative transfusions in 83% of patients, general anesthesia, adequate hydration, temperature conservation, non sedating analgesia and supervised incentive spirometry exercises (after 1995) in all patients. The total number of patients was 120: 81 underwent splenectomy, 59 cholecystectomy and 20 both procedures. Of patients who underwent splenectomy, 43.2% were females and 56.8% males; 64.2% had sickle cell anemia (SCA) and 35.8% sickle-beta thalassemia (ST). For those undergoing cholecystectomy, 39.0% were females and 61.0% males; 79.7% had SCA and 20.3% ST. Patients undergoing splenectomy were significantly younger (mean age 9.28 years) than those undergoing cholecystectomy (mean age 15.28 years) (p=0.037). Median operative time was 50 minutes, and median hospitalization duration was 2 1/2 days. No major intra operative complications or fatalities were noted. Postoperative complications included acute chest syndrome (ACS) in 5 patients (4% of surgeries). The mean time to onset of symptoms of ACS was 36 hrs after surgery (range, 24–96 hr). All patients who developed this complication did not receive incentive spirometry or were noncompliant with this therapy. As for the relation between clinical events and splenectomy/cholecystectomy, bivariate analysis showed a significant association between splenectomy and regular blood transfusion (p=0.005). Borderline significance was found for the association of cholecystectomy with ACS, joint necrosis and stroke (0.05<p<0.10). Predictors of splenectomy were regular blood transfusion (OR=2.38, [1.36–4.17] 95%CI], ACS (OR= 3.42, [1.64–7.15] 95%CI) and ST as compared to SS (OR=1.89, [1.10–3.27] 95%CI). Predictors of cholecystectomy were regular blood transfusion (OR=2.02, [1.10–3.71] 95%CI and sepsis (OR=2.59, [1.04–6.45] 95%CI). Sensitive predictors of postoperative complications could not be studied due to the small number of complications. This survey shows that splenectomy and cholecystectomy can be performed in SCD patients with minimal morbidity and no mortality. The current rate (4%) of post surgical ACS is lower than previously reported and is probably due to strict adherence to meticulous perioperative care. The absence of post surgical ACS in patients treated with supervised incentive spirometry is suggestive of a beneficial effect of this therapy in preventing this serious complication. The small number of patients with post surgical ACS prevents us, however, from drawing definite conclusions about the preventive role of this treatment. Nevertheless, early use of this simple, available and inexpensive tool most likely minimizes surgical morbidity and mortality from ACS, decreases hospitalization time, cuts down expenses and can be helpful particularly in countries with limited resources. Prospective controlled studies are, however, warranted to evaluate the efficacy of incentive spirometry and other perioperative interventions in preventing complications in SCD patients undergoing surgery.