Thermal Pain and Pain Anticipation Induce a Decrease in Microvascular Perfusion in Sickle Cell and Normal Subjects

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 67-67 ◽  
Author(s):  
Maha Khaleel ◽  
Mammen Puliyel ◽  
John Sunwoo ◽  
Payal Shah ◽  
Roberta Miyeko Kato ◽  
...  
Keyword(s):  
Blood Flow ◽  
Sickle Cell ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Painful Stimulus ◽  
Peripheral Blood Flow ◽  
Heat Pain ◽  
Microvascular Flow ◽  
Blood Disorder ◽  
Cross Correlation Analysis

Abstract INTRODUCTION: Sickle cell disease is an inherited blood disorder characterized by vaso-occlusive crises (VOC). HbS in red blood cells (RBC) polymerizes rapidly after it releases oxygen to tissues, causing RBC to become rigid. Anything that decreases flow in the microvasculature increases the chance that this flexible-to-rigid transformation occurs causing the rigid blood cell to lodge in the microvasculature, therefore increasing the chance of vaso-occlusion and the risk of VOC. Although hypoxia and stress are known risk factors for crises, the exact mechanism that initiates VOC events is not well known. We have previously shown that transient hypoxia causes parasympathetic withdrawal and sighs cause vasoconstriction more frequently in SCD subjects than in normal controls. Pain is the hallmark of SCD and is a consequence of VOC but has not been considered as a possible trigger of vasoconstriction that may lead to VOC. OBJECTIVES: To determine if heat induced pain causes decrease in peripheral blood flow (PBF) in SCD. METHODS: 30 SCD and 30 control subjects (healthy and sickle cell traits) were recruited at Children's Hospital Los Angeles (CHLA). Quasi-periodic pulses of pain were induced on the right forearm using TSA-II neuro analyzer heating thermode. We implemented a technique using cross correlation analysis to detect changes in complex microvascular flow signals measured bilaterally on the hands, using laser-Doppler flowmeter (LDF), Peripheral Arterial Tonometer (PAT) and photo-plethysmography (PPG). We also measured the average drop from baseline in the microvascular flow during the heat pain. Data were analyzed using one- and two- sample Student t-test. RESULTS: Data on 53 subjects were analyzed. There was a significant correlation between heat pain pulses and PBF responses, as well as a significant drop in blood flow in all study participants (PPG signal, both p<0.001), indicating that heat pain pulses lead to vasoconstriction. Males had higher correlation (p<0.005) and stronger vasoconstriction (p<0.05) during heat stimuli compared to females. Other Signals (LDF and PAT) had a similar pattern but were less significant. The vasoconstriction response consisted of two components, the first one occurred prior to administration of the painful stimulus indicating that anxiety or anticipation of pain causes significant vasoconstriction. Application of the painful stimulus causes further vasoconstriction. CONCLUSIONS: The findings demonstrate a significant decrease in PBF in both SCD and controls in response to heat pain and possibly to pain anticipation. The decrease in PBF could play a critical role in the genesis of VOC in SCD by markedly prolonging microvascular transit time, increasing the likelihood of red cell entrapment when sickle red cells transform from flexible to rigid. Furthermore, the potent vasoconstriction response to pain in SCD means that pain resulting from VOC could potentially trigger a cascade effect in which vasoconstriction could lead to even more serious VOC. Since regional blood flow is regulated by the autonomic nervous system (ANS), which has been described to be dysfunctional in SCD, our study calls attention to the ANS as a factor in the genesis of crisis in this disorder. Disclosures No relevant conflicts of interest to declare.

scholarly journals Laboratorial and Genetic Biomarkers Associated to Cerebral Blood Flow Velocity on Hemoglobin SC Disease

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4847-4847
Author(s):  
Rayra Pereira Santiago ◽  
Camilo Vieira ◽  
Corynne Stephanie Ahouefa Adanho ◽  
Sanzio Silva Santana ◽  
Caroline Conceição Guarda ◽  
...  
Keyword(s):  
Blood Flow ◽  
Cerebral Blood Flow ◽  
Sickle Cell ◽  
Methylenetetrahydrofolate Reductase ◽  
Conflicts Of Interest ◽  
Cerebral Blood Flow Velocity ◽  
Direct Bilirubin ◽  
Healthy Population ◽  
Mean Velocity ◽  
Hemoglobin Sc Disease

Abstract INTRODUCTION: Clinical complications in hemoglobin SC disease (HbSC) are mild compared to sickle cell anemia (SCA), the most severe type of sickle cell disease. However, HbSC individuals have 100 times greater risk of stroke compared to the healthy population. The lack of studies to determine reference values for transcranial Doppler (TCD) velocities in HbSC is still a limitation considering that we have been used TCD velocities previously determined for the SCA and having as standard the one described by Adams et al. (1992). Other studies described cerebral blood flow velocities (CBFV) for HbSC individuals, and considered as abnormal values that exceed 128 cm/s and 143.5 cm/s respectively. Studies show the association of abnormal TCD with distinct laboratorial biomarkers, despite the controversial results. To our knowledge, there are no published studies based on specific biomarkers and abnormal CBFV for HbSC individuals. Thus, we aim to investigate the association of genetic, hematological and biochemical data with CBFV in HbSC. METHODS: A cross-sectional study comprising 68 HbSC individuals, with an average age of 6.96 ± 3.90 years, whom 40 (58.82 %) were female, attended the Pediatric Cerebrovascular Disease Outpatient Center at the Hospital Universitario Professor Edgar Santos (HUPES) of the Universidade Federal da Bahia (UFBA) was conducted. All procedures were in accordance with the 1964 Helsinki declaration and its later amendments. The CBFV was estimated with TCD, and the time-averaged maximum mean velocity (TAMMV) in the middle cerebral arteries, anterior cerebral artery and distal intracranial internal carotid was assessed. Hematological and biochemical parameters, nitric oxide (NO) measurement and genetic polymorphisms in methylenetetrahydrofolate reductase (MTHFR) 677C>T (rs1801133), vascular cell adhesion molecule (VCAM)-833T>C (rs1041163)and VCAM 1238G>C genes, α3.7Kb thalassemia (-α3.7-thal) and Beta S (βS) haplotypes were investigated. Statistical analyses were performed using SPSS version 18.0 software, Graphpad Prism version 6.0 and JMP software. P values <0.05 were considered significant. RESULTS: The median of TAMMV in the HbSC individuals was 111.50 cm/s, the 25th percentile was 101.50 cm/s and the 75th percentile was 125.75 cm/s. The TAMMV was negatively correlated to red blood cells (RBC) (R= -0.2734; P= 0.0315), hemoglobin (Hb) (R= -0.3390; P= 0.0070), hematocrit (Ht) (R= -0.3470; P= 0.0057) and direct bilirubin (R= -0.2545; P= 0.0363) and positively correlated to monocytes (R= 0.2533; P= 0.0470) and ferritin (R= 0.3044; P= 0.0145). Based on previous protocol for SCA established by Adams and colleagues, we found two individuals with low TCD, 64 with normal TCD, one with abnormal TCD and one with inconclusive TCD. Noteworthy, this classification is not suitable for HbSC individuals. Using a cut off value of 128 cm/s, defined by Deane and colleagues (2008), we found significant decreased RDW (p=0.035) and NO (p=0.017) values in HbSC individuals with TAMMV higher than 128 cm/s. Vieira and colleagues (2014) defined a cut off value of 143.50 cm/s for HbSC individuals in the same population of this study. We found significant decrease of Hb (p=0.008) and Ht (p=0.008) concentration and increased ferritin levels (p=0.023) in HbSC individuals with TAMMV higher than 143.50 cm/s. Value of TAMMV at 75th percentile (125.75 cm/s) was used in order to evaluate the laboratorial markers. We found significant differences in Hb, Ht, RDW and NO levels, which were decreased, and in the ferritin, which was elevated in HbSC individuals with TAMMV above 125.75 cm/s (Table 1). The multivariate analysis adjusted by age and sex showed that Ht (< 33.35%), RDW (> 15.55%) and NO (< 29.86 uM) were independently associated with TAMMV values higher than 125.75 cm/s (75th percentile) (Table 2). CONCLUSIONS: We suggest that RBC, Hb, Ht, RDW, monocyte, direct bilirubin, NO, ferritin, polymorphism on the MTHFR 677C>T gene and the absence of α3.7Kb thalassemia are associated with TAMMV on HbSC individuals. The TAMMV 125.75cm/s is lower than the those proposed by Deane et al. (2008) and Vieira et al. (2014), but have shown to be associated with hematological and biochemical changes found in both velocities 128 cm/s and 143.50 cm/s respectively. Thus, we also suggest that TAMMV of 125.75 cm/s should be investigated in additional studies as a cut-off point for stroke risk in HbSC individuals. Disclosures No relevant conflicts of interest to declare.

scholarly journals Engagement and Education of the Sickle Cell Disease Community: What Sickle Cell Disease Patients and Parents Want to Know about CRISPR Genome Editing

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5805-5805
Author(s):  
Stacy Desine ◽  
Brittany Hollister ◽  
Anitra Persaud ◽  
Vence L. Bonham
Keyword(s):  
Sickle Cell Disease ◽  
Genome Editing ◽  
Sickle Cell ◽  
Conflicts Of Interest ◽  
Scientific Information ◽  
Educational Video ◽  
Cell Disease ◽  
Genetic Literacy ◽  
Blood Disorder ◽  
A Genome

Abstract Introduction: One of the first therapeutic targets of CRISPR genome editing will likely be sickle cell disease (SCD), the most commonly inherited blood disorder in the U.S.SCD affects 100,000 individuals in the U.S. and more than 250 million individuals globally. Two targets of genome editing in SCD currently under investigation are HBB and BCL11A. The first approach (HBB) focuses on correcting the Glu6Val mutation in patient-derived stem and progenitor cells to promote differentiation into normal erythrocytes. The second approach involves disrupting BCL11A, a repressor of fetal hemoglobin. While preliminary results from research using both mechanisms show promise, risks related to insertional mutagenesis and host immune responses remain. As the research advances, limited knowledge exists regarding SCD stakeholders' (patients, parents and physicians) understanding of genome editing and their educational needs. Methods: We conducted a mixed methods study, which included a CRISPR genome editing educational video, pre- and post-video surveys, and fifteen, moderated focus groups: 6 patient groups, 6 parent groups, and 3 physician groups. The surveys included an assessment of participants' knowledge of CRISPR gene editing and the Genetic Literacy and Comprehension (GLAC) instrument. Eligible participants were patients with a diagnosis of SCD, parents of a child with SCD, and physicians who have delivered care to at least five adult or pediatric patients living with SCD for a minimum of 12 months. Differences between the pre- and post-video score of CRISPR technology knowledge were assessed using paired t-tests. NVivo 11 qualitative software tool was used to assist in data analysis. Results: For each of the eight complex genetic terms in the GLAC, patients and parents were asked to rank their familiarity on a scale from 1-7, and to complete fill-in-the-blank, multiple choice questions aimed at assessing their understanding of the term. Baseline genetic literacy levels were high for both stakeholder groups. Patients and parents had an average familiarity score of 6.34, and 6.88, respectively. 89% of both groups scored >70% on the fill-in-the-blank questions. As shown in Figure 1, the educational video tool was effective at increasing CRISPR genome editing knowledge among patients and parents - the average CRISPR genome editing knowledge scores of both groups increased by over two points (p<0.0001 and p=0.05, respectively). Prior to viewing the video, patients and parents had lower CRISPR genome-editing knowledge scores than physicians (p<0.0002 and p<0.0002, respectively). However, after viewing the video, there were no significant differences in scores between patients, parents, and physicians. Several qualitative themes emerged related to patients' and parents' interest in understanding CRISPR genome editing. These themes included the need for easily accessible scientific information regarding: 1) CRISPR's mechanism of action; 2) potential adverse reactions to CRISPR; 3) effects of genome editing on future generations; and 4) burden of participating in a genome editing clinical trial. Discussion: After watching the video tool, patients' and parents' understanding of CRISPR genome editing improved significantly. This suggests that video education tools may be an effective method to educate individuals living with SCD and parents about genome editing. Furthermore, this study reveals patients and parents were not only familiar with complex genetic terms, but also correctly defined them, suggesting a good baseline genetic literacy. The themes indicate the importance of keeping patients and parents informed and supporting their desires to be educated about the applications of genome editing technology which will further the advancement of clinical trials. Disclosures No relevant conflicts of interest to declare.

scholarly journals Assessment of Erythroid Chimerism in Sickle Cell Patients Undergoing HSCT By Digital Droplet PCR

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 563-563
Author(s):  
Lihui Yin ◽  
Rupa A Udani ◽  
Mary Parlow ◽  
Daniel B Bellissimo ◽  
Joel A Brochstein
Keyword(s):  
Sickle Cell ◽  
Conflicts Of Interest ◽  
Hematopoietic Stem ◽  
Post Transplant ◽  
Digital Droplet Pcr ◽  
Blood Disorder ◽  
The Status ◽  
Droplet Pcr ◽  
Allele Specific ◽  
Donor Genotype

Abstract Sickle cell disease (SCD) is an inherited autosomal recessive blood disorder associated with significant morbidity. Nonmyeloablative allogeneic hematopoietic stem cell transplantation (HSCT) is increasingly used in severely affected patients with SCD. These transplants result in mixed hematopoietic chimerism so accurate chimerism testing is important for monitoring the status of the transplant. Reconstitution of the erythroid compartment is essential. Since red cells lack a nucleus, DNA-based chimerism assays do not directly assess the chimerism in the erythroid compartment. Several studies have shown that the chimerism in the white cells and erythroid cells can be very different (W, C., etal. Exp. Hematol. 2003, 31:924; Andreanni, M., etal. Haematologica, 2011, 96:128). We developed a procedure for quantification of chimerism in the erythroid compartment of blood using RNA-based digital droplet PCR (ddPCR). The sensitivity and specificity of the assay was evaluated then tested post-transplant samples from sickle cell patients. Total RNA is reverse transcribed and amplified in a two-step RT-PCR approach. The PCR reaction containing allele-specific hydrolysis probes is partitioned into ~15,000 droplets then amplified. The copy number of HbA and HbS transcripts from cells of the erythroid lineage is determined with ddPCR (BioRad, QX-200). The %HbA and the %Donor can be calculated using the donor genotype (A/A or A/S). The assay is designed to have a sensitivity of 1% with donor genotype of A/A and 5% with donor genotype of A/S. Each post-transplant sample is tested in duplicate along with an AA control, SS control, AS control and 1% sensitivity controls (5 controls total). Thirty AA or SS samples were tested to assess assay specificity. The average %HbA detected in a SS sample was 0.03%; the average %HbS in a AA sample was as 0.02%. The background signal is significantly below the cutoff for 1% sensitivity. Using contrived samples with low, medium, and high %HbA, we demonstrate the assay is accurate and linear to 0-2%HbA across the reportable range of 0%HbA to 100%HbA. The % CV of the minor allele for samples in the range of 10%-90%HbA, were equal or below 15%. A total of 11 post-transplant samples from 7 transplanted sickle cell patients were tested, and the results were compared to DNA-based/FISH chimerism, if available. Our results were comparable with DNA-based chimerism (Table 1). Five of the samples had slightly higher %donor in the erythroid compartment compared to the white cell compartment. The erythroid chimerism reflected changes in chimerism status: the decrease then increase in chimerism (P2), stable chimerism (P5) and graft failure (P7). Abstract 563. Table 1: Erythroid chimerism status of post-transplant SCD patients Patient # Donor Genotype Sample # %HbA %Donor %Recipient %Donor by FISH/STR P1 AS 1 52 100 0 95 P2 AA 1 66 66 34 72 2 44 44 56 50 3 100 100 0 95 P3 AS 1 54 100 0 85 P4 AA 1 100 100 0 100 P5 AS 1 14 28 72 15 2 12 24 76 15 P6 AS 1 57 100 0 95 P7 AA 1 0 0 100 Rejecting graft 2 0 0 100 Rejecting graft Assessment of the chimerism in the erythroid lineage may be a better indicator of donor erythropoiesis. We describe an accurate and sensitive assay for monitoring erythroid chimerism and the effects of post-transplant therapies in sickle cell patients undergoing HSCT. This assay also demonstrates the feasibility measuring erythroid chimerism detection in other hematologic disorders, such as thalassemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals Blood rheology biomarkers in sickle cell disease

2020 ◽  
Vol 245 (2) ◽  
pp. 155-165
Author(s):  
Madeleine Lu ◽  
Minke AE Rab ◽  
Sergey S Shevkoplyas ◽  
Vivien A Sheehan
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Blood Rheology ◽  
Early Mortality ◽  
Microvascular Blood Flow ◽  
Novel Therapies ◽  
Cell Disease ◽  
Clinical Complications ◽  
Blood Disorder

Sickle cell disease (SCD) is the most common inherited blood disorder, affecting approximately 100,000 patients in the U.S. and millions more worldwide. Patients with SCD experience a wide range of clinical complications, including frequent pain crises, stroke, and early mortality, all originating from a single-point mutation in the β-globin subunit. The RBC changes resulting from the sickle mutation lead to a host of rheological abnormalities that diminish microvascular blood flow, and produce severe anemia due to RBC hemolysis, and ischemia from vaso-occlusion initiated by sticky, rigid sickle RBCs. While the pathophysiology and mechanisms of SCD have been investigated for many years, therapies to treat the disease are limited. In addition to RBC transfusion, there are only two US Food and Drug Administration (FDA)-approved drugs to ameliorate SCD complications: hydroxyurea (HU) and L-glutamine (Endari™). The only curative therapy currently available is allogeneic hematopoietic stem cell transplantation (HSCT), which is generally reserved for individuals with a matched related donor, comprising only 10–15% of the total SCD population. Potentially curative advanced gene therapy approaches for SCD are under investigation in ongoing clinical trials. The ultimate goal of any curative treatment should be to repair the hemorheological abnormalities caused by SCD, and thus normalize blood flow and prevent clinical complications. Our mini-review highlights a set of key hemorheological biomarkers (and the current and emerging technologies used to measure them) that may be used to guide the development of novel curative and palliative therapies for SCD, and functionally assess outcomes. Impact statement Severe impairment of blood rheology is the hallmark of SCD pathophysiology, and one of the key factors predisposing SCD patients to pain crises, organ damage, and early mortality. As novel therapies emerge to treat or cure SCD, it is crucial that these treatments are functionally evaluated for their effect on blood rheology. This review describes a comprehensive panel of rheological biomarkers, their clinical uses, and the technologies used to obtain them. The described technologies can produce highly sensitive measurements of the ability of current treatments to improve blood rheology of SCD patients. The goal of curative therapies should be to achieve blood rheology biomarkers measurements in the range of sickle cell trait individuals (HbAS). The use of the panel of rheological biomarkers proposed in this review could significantly accelerate the development, optimization, and clinical translation of novel therapies for SCD.

scholarly journals The effect of hypnosis on pain and peripheral blood flow in sickle-cell disease: a pilot study

2017 ◽  
Vol Volume 10 ◽  
pp. 1635-1644 ◽  
Author(s):  
Ravi Bhatt ◽  
Sarah Martin ◽  
Subhadra Evans ◽  
Kirsten Lung ◽  
Thomas Coates ◽  
...  
Keyword(s):  
Blood Flow ◽  
Pilot Study ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Peripheral Blood ◽  
Peripheral Blood Flow ◽  
Cell Disease

The Effect Of Microalbuminuria On Duration Of Hospitalization In Adult Sickle Cell Disease Patients With Pain Crisis

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4694-4694
Author(s):  
Mohammed Shaik ◽  
Borys Hrinczenko
Keyword(s):  
Sickle Cell Disease ◽  
Length Of Stay ◽  
Sickle Cell ◽  
Conflicts Of Interest ◽  
Categorical Variables ◽  
Cell Disease ◽  
Exact Test ◽  
Blood Disorder ◽  
Significant Difference ◽  
Pain Crisis

Introduction Sickle cell disease (SCD) is a genetic blood disorder of hemoglobin resulting in severe morbidity and early mortality. The prevalence of microalbuminuria and/or proteinuria in children with SCD varies from 18 to 28% and increases with age. However, the exact prevalence in adults in not known. In the general population, the presence of albuminuria has been shown to be associated with all cause mortality. The urine microalbumin to creatinine ratio (MA) is considered to be an early sign of impending sickle cell nephropathy. We sought to investigate the association of MA with various SCD genotypes (HbSS, HbSC, HbS/β-thalassemia) in our clinic and also the length of stay (LOS) in hospitalized SCD patients with acute pain crisis. Methods Twenty-eight consecutive SCD patients diagnosed in our clinic by hemoglobin electrophoresis were included in our study. The patients (pts) age, gender, hemoglobin electrophoresis, and baseline MA were obtained. Based on their MA level they were divided into two groups, abnormal MA (MA ≥ 30 mg/g creatinine) or normal MA (< 30 mg/g creatinine). Eleven of these 28 patients were eventually hospitalized for a sickle cell related pain crisis. Their MA level was obtained within 24-hours of hospital admission and their hospitalization length of stay (LOS) was also recorded. We analyzed the association between the two groups of patients, those with normal or abnormal MA, with the different genetic variants of SCD in both our clinic and hospitalized pts. Furthermore, for hospitalized pts we also assessed an association of MA with their mean LOS. The Chi-square test/Fisher’s exact test was used for categorical variables and the T-test/Mann-Whitney test was used for numerical variables. Results All twenty-eight patients were African American without significant renal impairment, with 11, 10, and 7 pts with SS, SC and S β-thalassemia (Sβ), respectively. Fifteen of these pts had abnormal MA, 12 pts were female. The median age was 35.5 yrs (range, 19 - 59), median LOS was 3.5 days (range, 2-8). The SS pts had higher abnormal MA levels followed by SC and then Sβ (p=0.03) (Table 1). There was no significant difference in gender between the two groups (p=0.2). SCD pts admitted to the hospital for pain crisis with an abnormal MA within 24-hours of admission had a significantly higher mean LOS when compared to pts with normal MA (p=0.0089) (Table 1). Conclusion Microalbuminuria is more prevalent in the severe genotypes of SCD disease such as SS and SC vs. Sβ pts. Thereby, MA might be a useful biomarker of generalized SCD vasculopathy, in addition to a known marker of progressive nephropathy. Furthermore, MA has a significant impact on length of hospitalization. An abnormal MA obtained within the first 24-hrs of hospitalization of a SCD pt in pain crisis was predictive of prolonged hospital duration. Early and more aggressive supportive care symptom management for those patients might be a reasonable option but requires more study. Further large prospective clinical trials are needed to validate our findings. Disclosures: No relevant conflicts of interest to declare.

Targeting Neutrophil Aging and the Microbiota for the Treatment of Sickle Cell Disease

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 279-279
Author(s):  
Dachuan Zhang ◽  
Grace Chen ◽  
Deepa Manwani ◽  
Arthur Mortha ◽  
Chunliang Xu ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Conflicts Of Interest ◽  
Leukocyte Adhesion ◽  
Cell Disease ◽  
Fecal Transplantation ◽  
Penicillin V ◽  
Blood Disorder ◽  
Prophylaxis Therapy

Abstract Sickle cell disease (SCD) is a genetic blood disorder characterized by repeated episodes of vaso-occlusion, in which neutrophils play a primary function through their recruitment in inflamed venules and interactions with sickle red blood cells (sRBCs) via activated αMβ2 integrin (Mac-1). Intravital microscopy analyses revealed considerable heterogeneity in Mac-1 activation on adherent neutrophils, suggesting that subsets of neutrophils differ markedly in their pro-inflammatory activity (Nat Med 2009; 15:384). Recently, we found that aged neutrophils, marked by CD62Llo CXCR4hi, represented an overly active subset exhibiting enhanced Mac-1 activation (Blood 2013; 122:324). To further characterize aged neutrophils, we compared their transcriptome with control and in vivo- activated neutrophils. Gene set enrichment analyses revealed that aged neutrophils up-regulated several pathways that were also enhanced during activation, including integrin, leukocyte adhesion, toll-like receptor (TLR), and NFκB signalling pathways (P < 0.05), suggesting a contribution by exogenous inflammatory mediators. Based on the expression profile, we hypothesized that microbiota-derived signals may influence neutrophil aging in the circulation. To test this idea, we analyzed aged neutrophil numbers in antibiotics (ABX)-treated and germ-free (GF) mice, in which the gut microbiota was largely depleted or deficient, respectively. We found significant reductions of aged neutrophil numbers in both models, and the numbers could be significantly restored by either lipopolysaccharide (LPS) gavage or fecal transplantation (Ctrl / ABX / ABX+LPS: 98±14 / 36±11 / 107±25 cells per μl blood; SPF / GF / GF-FT: 56±12 / 4±1 / 15±4 cells per μl blood; P < 0.05). We next investigated whether microbiota-driven neutrophil aging was mediated by TLRs and Myd88 signalling using chimeric mice reconstituted with a mixture of WT and Myd88, TLR4 or TLR2-deficient hematopoietic cells. Interestingly, we observed significant reductions in the aged subset of Myd88, TLR4 or TLR2-deficient neutrophils compared to WT neutrophils in the same mice, and in their capacity to capture fluosphere beads that specifically bound to activated Mac-1 (WT / LysM-cre/Myd88-flox: 10.4±0.3 / 7.0±0.6 %, 1.0±0.1 / 0.4±0.1 bead per cell; WT / Tlr4-/-: 10.3±0.2 / 6.0±0.2 %, 1.0±0.1 / 0.4±0.1 bead per cell; WT / Tlr2-/-: 10.0±0.4 / 6.8±0.5 %, 1.3±0.2 / 0.8±0.1 bead per cell; P < 0.01). Analysis of SCD mice (BERK) revealed that the aged neutrophil population was significantly increased (by >10-fold) compared to hemizygous (SA) mice, and the expansion was completely abrogated by microbiota depletion (SA / SCD (SS)-Ctrl / SS-ABX: 71±11 / 981±261 / 124±27 cells per μl blood, P < 0.01). We then challenged SA, untreated and ABX-treated SCD mice with TNF-α to induce acute vaso-occlusion. SCD mice exhibited significant increases in neutrophil adhesion, Mac-1 integrin activation and heterotypic interactions with RBCs compared to SA mice, all of which were markedly reduced by microbiota depletion (SA / SS-Ctrl / SS-ABX: 6.4±0.6 / 18.1±1.0 / 11.2±1.0 adherent neutrophil per vessel; 0.5±0.1 / 1.0±0.2 / 0.3±0.1 bead per cell; 0.1±0.1 / 0.5±0.1 / 0.2±0.1 RBC interaction per vessel per min; P < 0.05), resulting in significantly enhanced blood flow and prolonged survival in ABX-treated SCD mice (P < 0.05). Most interestingly, the splenomegaly of SCD mice was significantly reduced, and liver damage including fibrosis, necrosis and inflammation was dramatically alleviated in ABX-treated SCD mice (P < 0.05). Finally, we evaluated whether the numbers of circulating aged neutrophils were altered in patients with SCD. As Penicillin V antibiotic prophylaxis therapy is recommended for children < 5 years or older patients with immune defects to prevent life-threatening infections, we determined aged neutrophil numbers in this patient population. We found that SCD patients exhibited a marked increase in the numbers of circulating aged neutrophils compared to healthy controls, which were significantly reduced in patients on Penicillin V prophylaxis (Ctrl / SS / SS-PV: 261±40 / 1175±205 / 439±87 cells per μl blood, P < 0.05). Our results raise the possibility that targeting neutrophil aging and the microbiota may have broad implications in our understanding of sickle cell disease pathogenesis and its management that should be further studied in clinical trials. Disclosures No relevant conflicts of interest to declare.

scholarly journals Vaso-Occlusion in Sickle Cell Disease: Is Autonomic Dysregulation of the Microvasculature the Trigger?

2019 ◽  
Vol 8 (10) ◽  
pp. 1690 ◽  
Author(s):  
Saranya Veluswamy ◽  
Payal Shah ◽  
Christopher Denton ◽  
Patjanaporn Chalacheva ◽  
Michael Khoo ◽  
...  
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Red Blood Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
Critical Role ◽  
Microvascular Blood Flow ◽  
Cell Disease ◽  
Hemoglobin S

Sickle cell disease (SCD) is an inherited hemoglobinopathy characterized by polymerization of hemoglobin S upon deoxygenation that results in the formation of rigid sickled-shaped red blood cells that can occlude the microvasculature, which leads to sudden onsets of pain. The severity of vaso-occlusive crises (VOC) is quite variable among patients, which is not fully explained by their genetic and biological profiles. The mechanism that initiates the transition from steady state to VOC remains unknown, as is the role of clinically reported triggers such as stress, cold and pain. The rate of hemoglobin S polymerization after deoxygenation is an important determinant of vaso-occlusion. Similarly, the microvascular blood flow rate plays a critical role as fast-moving red blood cells are better able to escape the microvasculature before polymerization of deoxy-hemoglobin S causes the red cells to become rigid and lodge in small vessels. The role of the autonomic nervous system (ANS) activity in VOC initiation and propagation has been underestimated considering that the ANS is the major regulator of microvascular blood flow and that most triggers of VOC can alter the autonomic balance. Here, we will briefly review the evidence supporting the presence of ANS dysfunction in SCD, its implications in the onset of VOC, and how differences in autonomic vasoreactivity might potentially contribute to variability in VOC severity.

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