The Use Of Two Photon Microscopy To Image Vaso-Occulsion In Pulmonary Microvessels Of Living Mice With Sickle Cell Disease

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 976-976
Author(s):  
Natacha Ralainirina ◽  
Ferron Lynn Sonderegger ◽  
Hong Pei ◽  
Grzegorz Chodaczek ◽  
Joel Linden
Keyword(s):  
Endothelial Cells ◽  
Sickle Cell Disease ◽  
Real Time ◽  
Sickle Cell ◽  
Intravital Microscopy ◽  
Blood Cells ◽  
White Blood Cells ◽  
Cell Disease ◽  
Alexa Fluor ◽  
Two Photon

Abstract Although sickle cell anemia is initiated by red cell pathology, it is accompanied by an inflammatory immune response involving platelets and white blood cells that contribute to vaso-occlusvie episodes including painful vaso-occlusive crises (pVOC) and acute chest syndrome (ACS). In order to better understand the cellular and molecular bases of vaso-occlusion we are in the process of developing procedures to image microvessels in the lung, liver and spleen of living mice by 2-photon microscopy, a procedure that is based on excitation of a fluorophore by two photons simultaneously. The two-photon technique utilizes infrared light that efficiently penetrates tissues up to 200 microns with low phototoxicity allowing time-lapse imaging. Two-photon intravital microscopy can be used to study the behavior of intravascular cells during vaso-occlusive events. Mice are prepared for lung intravital microscopy by the intraperitoneal injection of a mixture of ketamine and xylazine. Additional anesthesia is added during experimentation. The trachea is opened and the mouse intubated. The chest is opened to allow access to the left lobe of the lung through a window that is a few millimeters in diameter. PBS is applied to keep the lung moist. A custom built suction device is placed on the lung and covered with a cover glass at the same time pressure is exerted to seal the organ and the glass cover together. Throughout the procedure, the mouse is held at a temperature of 37°C. Once surgery is completed, a mixture of antibodies coupled to fluorophores is given by retro-orbital injection. In order to minimize photobleaching we used antibodies conjugated to Alexa Fluor 488, Alexa Fluor 555 or Alexa Fluor 647. We are able to visualize and quantify interactions between red blood cells, white blood cells, and endothelial cells as well as the expression of adhesion molecules on endothelial cells in real time. During pVOC triggered by hypoxia, cell adhesion of neutrophils, lymphocytes and monocytes to the endothelium is observed that is associated with an increase in endothelial expression of ICAM-1 and V-CAM. We label endothelial cells with anti-CD31, lymphocytes with anti-CD45, monocytes with anti-Ly6C and neutrophils with anti-Ly6G. Platelets are labeled with anti-CD41 or anti-CD62P, NK cells with anti-NKp46, and macrophages with anti-F4/80 and anti-CD1d. We are able to quantify cell shape, rolling, adhesion and movement. Our preliminary results demonstrate that it is possible in real time to image the sequence of events occurring during pulmonary vasoocclusion in sickle cell disease. In conclusion, intravital 2-photon microscopy holds great potential for enabling us to better understand inflammatory responses within the blood vessels of living SCD mice. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The prevalence of leukocyte abnormalities among Sudanese patients with sickle cell disease

2021 ◽  
Vol 9 (1) ◽  
pp. 262-267
Author(s):  
Tarig Osman Khalafallah Ahmed ◽  
Ekhlas Alrasheid Abu Elfadul ◽  
Ahmed A. Agab Eldour ◽  
Omer Ibrahim Abdallah Mohammed
Keyword(s):  
Sickle Cell Disease ◽  
Cell Volume ◽  
Sickle Cell ◽  
Blood Cells ◽  
White Blood Cells ◽  
Blood Film ◽  
Cell Disease ◽  
Mean Cell Volume ◽  
Blood Disorder ◽  
Age Range

Sickle cell disease (SCD) is an inherited blood disorder that affects red blood cells. The study was conducted in Elobied town during the period May 2011 to September 2011. The aim of this study is to detect the abnormalities of leucocytes among sickle cell anemic patients. 40 sickle cell anemic patients; age range between 8 months to 23 years. Blood sample was taken for all patients and the laboratory investigation were performed using automated estimation for: hemoglobin (Hb), Packed cell volume (PCV), red cell count (RBCs), mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell concentration (MCHC), and total white blood cells, comment on blood film using manual methods. The conclusion of this study there is increase in total white blood cells with shift to left in neutrophil precursor in sickle cell patients with complications ,the most immature cells are band form, myelocytes and metamyelocytes, and there also lymphocytosis and neutrophilia which has been increases in response to infections.

scholarly journals Role of some members of chemokine/cytokine network in the pathogenesis of thalassemia and sickle cell hemoglobinopathies: a mini review

2019 ◽  
Vol 8 (1) ◽  
Author(s):  
Zahra Mousavi ◽  
Zinat Yazdani ◽  
Alireza Moradabadi ◽  
Fatemeh Hoseinpourkasgari ◽  
Gholamhossein Hassanshahi
Keyword(s):  
Endothelial Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
White Blood Cells ◽  
Cell Disease ◽  
Cytokine Network ◽  
Exact Role ◽  
Cytokines And Chemokines ◽  
Wide Range

Abstract The word of hemoglobinopathy is described for an array of disorders that affecting hemoglobin (Hb) functions. Hb is a molecule with 68 kDa molecular weight, serving as oxygen carrying metalloprotein. Hemoglobinopathy includes a wide range of Hb structural deficits varying from thalassemia to sickle cell disease. Cyto-chemokine network members are pivotally involved in the pathogenesis of hemoglobinopathies, however, the exact role of these mediators in the development of these disorders yet to be well addressed. Cytokines and chemokines are generated by inflamed endothelial cells that promote the expression of their respected receptors and further activate NF-κβ, recruit red blood cells (RBCs) and white blood cells (WBCs) toward the inflamed endothelium. Therefore, due to critical roles played by the cyto-chemokine network in several aspects of hemoglobinopathies pathophysiology including apoptosis of endothelial cells, RBC, WBC and etc.…, in the present review, we focused on the critical parts played by this network in the pathogenesis of hemoglobinopathies.

scholarly journals Persistence of Chronic Inflammation Despite Years of Transfusion Program in SCD Patients: Changing Red Blood Cells Is Not Sufficient to Treat Sickle Cell Disease

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 764-764
Author(s):  
Abdoul Karim Dembele ◽  
Patricia Hermand-Tournamille ◽  
Florence Missud ◽  
Emmanuelle Lesprit ◽  
Malika Benkerrou ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Sickle Cell Disease ◽  
Red Blood Cells ◽  
Chronic Inflammation ◽  
Sickle Cell ◽  
Blood Cells ◽  
Healthy Controls ◽  
Cell Disease ◽  
The Impact

Abstract Sickle cell disease (SCD) is a severe hemoglobinopathy due to abnormal hemoglobin S (HbS). Although red blood cell dysfunction is at the core of the SCD pathophysiology, several studies have highlighted the important role of inflammatory cells like neutrophils. One of the most serious complications of SCD is cerebral vasculopathy (CV), due to the occlusion of one or more intracranial or cervical arteries. In 1998, the STOP study demonstrated that monthly blood transfusions could reduce the risk of stroke by 90% in children with CV. However, there is large heterogeneity in the evolution of CV under chronic transfusion, sometimes requiring exchange transfusion (ET) program for years without succeeding in healing the CV. The aim of the study is to investigate the impact of long-term transfusion program on neutrophil dysfunction, in order to understand if persistent inflammation could contribute to the non-healing of CV despite HbS permanently below 40%. In SCD children undergoing ET program for at least 1 year, we analysed i)the phenotype of neutrophils with 8 markers of activation/adhesion/ageing, ii)the plasmatic levels of elastase, witnessing the NETose activity of neutrophils, and iii)the ex-vivo adhesion of neutrophils on activated endothelial cells. One hundred and two SCD children with an ET transfusion program for at least 6 months because of CV were included in the study. ET session, carried out every 5 weeks and most of the time by erythrapheresis, reached their biological objectives with a mean HbS rate after ET session of 14.1%, and 35.4% before the next ET session, which means that these patients globally live at an average HbS level of 24% for at least 1 year. We managed to limit iron overload with a mean ferritinemia of 207 µg/L in the whole cohort. Despite these satisfactory results in terms of HbS reduction, the efficiency in curing the CV was modest in accordance with the previously described efficiency of ET program in SCD children: after a mean ET program duration of 4.4 years only 22% of them had an improvement of their CV since the beginning of the ET program, while 60% of them had a stagnation of their CV, and 18% of them worsened their vascular lesions. Considering inflammatory parameters, the patients had persistence of high leukocytosis and high neutrophils count (respective mean of 9810 G/L and 5742 G/L), significantly not different of neutrophils count before inclusion in the ET program. In a random subgroup of 20 patients, we analysed neutrophils phenotype, NETose and endothelial adhesion and compared them to healthy controls and SCD children without ET, treated or not with Hydroxyurea (HU). Overall, we observed as expected an activated, aged and adherent profile of neutrophils from untreated SCD children compared to healthy controls, characterized by an overexpression of CD18/CD11b (p=0,03), CD18/CD11a (p=0,02), CD162 (p=0,01), CD66a (p=0,01) and the ageing markers CD184 high/CD62Llow (p=0,04) as well as a higher plasmatic level of elastase (p=0. 01) and higher adhesion of neutrophils to endothelial cells. All these parameters were alleviated in SCD patients treated with HU. In SCD patient undergoing ET program, we found a similar profile of activated neutrophils to that of untreated SCD patients with a similar expression of activation molecules, high level of elastase and the same increase of neutrophils adhesion to endothelial cells compared to controls, witnessing a persistence of chronic inflammation despites years of ET. Overall, our study highlights that the replacement of sickle red blood cells, even for years, is not sufficient to reverse the deleterious inflammatory phenotype of neutrophils. Given the major role of inflammation in endothelial dysfunction, these could contribute to the persistence of CV in a majority of patients despite efficient ET programs. This raises the question of systematically combining ET program with anti-inflammatory treatment such as HU or P-selectin inhibitors in children with CV. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Evaluation of Baseline Hematologic Parameters of Steady State Sickle Cell Disease Patients at The Chantal Biya’s Foundation, Yaounde- Cameroon

2019 ◽  
Vol 7 (07) ◽  
Author(s):  
Bih Adelaide ◽  
Ngala Solange Mudih ◽  
Alima Yanda ◽  
Akaba Fergus Ambe ◽  
Jutcha Florent ◽  
...  
Keyword(s):  
Steady State ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Reference Values ◽  
Blood Cells ◽  
Healthy Subjects ◽  
White Blood Cells ◽  
Great Decrease ◽  
Cell Disease ◽  
Mean Values

Introduction Sickle cell disease is a genetic abnormality involving the hemoglobin. Although it is primarily a red blood cell disorder, the white blood cells and platelets are also affected by the mutation. The consequence hemoglobin S causing polymerization of hemoglobin, results in hemolysis and anemia. This study aims to provide baseline hematologic parameters in steady state sickle cell disease patients compared with the reference values of normal healthy subjects used at the Chantal Biya Foundation (CBF), in order to monitor other sicklers in Cameroon. Methodology A comparative analysis of sickle cell hematologic parameters with control hematologic parameters of normal healthy subjects (reference values of healthy subjects used at the Chantal Biya Foundation) was carried out. Results A total of 62 sickle cell disease patients in steady state who complied with the selection criteria were recruited. Of the 62, 29 were females and 33 were males. The age range was from 1 year to 19 years and an average age of 6 ± 4.19 SD. Results from sickle cell patients showed an increase in white blood cells (WBCs), neutrophils and lymphocytes and a great decrease in the mean values of hematocrit Hct. as well as RBC indices, but no great or slight difference in the values of basophils compared with the reference values of normal healthy subjects in the CBF Yaoundé, 2015. Conclusion Sickle cell disease patients in steady state have lower values of red cells parameters, but higher values of white cells and platelets count when compared with the reference values of normal healthy subjects used at the Chantal Biya Foundation.

scholarly journals Management of Hodgkin Lymphoma in a Sickle Cell Patient: A Case Report

2019 ◽  
Vol 12 (1) ◽  
pp. 224-227
Author(s):  
Farah Ashraf ◽  
Pragnan Kancharla ◽  
Mendel Goldfinger
Keyword(s):  
Quality Of Life ◽  
Case Report ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Blood Cells ◽  
Patient Population ◽  
White Blood Cells ◽  
Modern Medicine ◽  
Cell Disease

Sickle cell disease (SCD) is an inherited disorder of hemoglobin mutation in red blood cells, with a patient population that is increasing in age in recent decades due to advances in modern medicine. Hodgkin’s lymphoma (HL) is a cancer of white blood cells, and while concomitance of SCD and Hodgkin’s has been reported, a discussion of treatment for HL in SCD is lacking from the literature. We present a case of effectively treated HL in SCD and put forth that the regimen used is a practical choice, and as it was completed fully as outpatient, it improved the patient’s quality of life compared to an inpatient regimen.

scholarly journals Elevated Urinary Leukotriene E4 Levels Are Associated with Hospitalization for Pain in Children with Sickle Cell Disease.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3405-3405
Author(s):  
Jeanine E. Jennings ◽  
Thiruvamoor Ramkumar ◽  
Jingnan Mao ◽  
Jessica Boyd ◽  
Mario Castro ◽  
...  
Keyword(s):  
Arachidonic Acid ◽  
Sickle Cell Disease ◽  
Incidence Rate ◽  
Sickle Cell ◽  
Blood Cells ◽  
Medical Records ◽  
White Blood Cells ◽  
Cell Disease ◽  
Inflammatory Stimuli ◽  
Inflammatory State

Abstract Introduction: Sickle cell disease (SCD) is associated with an inflammatory state. Luekotrienes are inflammatory mediators derived from arachidonic acid and produced by white blood cells in response to inflammatory stimuli. We tested two hypotheses among children with SCD: Baseline leukotriene levels are elevated in SCD when compared to controls children without SCD, Baseline leukotriene levels are associated with an increased incidence rate of hospitalization for pain. Methods: During routine clinical visits, baseline urinary leukotriene E4 (LTE4) levels were measured in children with SCD (cases) and age, and ethnic, matched children without SCD (controls). Medical records of the cases were reviewed to assess the frequency of hospitalization for pain within three years of entering the study. Results: LTE4 levels were obtained in 71 cases and 22 controls. LTE4 levels were higher in the cases compared to controls (median LTE4: 100 vs.57 pg/mg creatinine); P < 0.001. After adjustment for age and diagnosis of asthma, a greater incidence rate of hospitalization for pain was observed among children with SCD in the highest tertile group when compared to the lowest tertile group (114 vs. 52 episodes per 100 patient-years, P=0.038). Conclusion: LTE4 levels are elevated in children with SCD when compared to controls without SCD and are associated with an increased rate of hospitalizations for pain.

Sphingosine 1-Phosphate (S1P)/S1P Receptor 1 Pathway Has an Essential Role for Sickle Cell Disease

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4063-4063
Author(s):  
Yujin Zhang ◽  
Shushan Zhao ◽  
Hongyu Wu ◽  
Xia Hu ◽  
Renna Luo ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Inflammatory Cytokines ◽  
Sickle Cell ◽  
Blood Cells ◽  
Genetic Disorder ◽  
White Blood Cells ◽  
High Morbidity ◽  
Cell Disease ◽  
S1p Receptors ◽  
Sphingosine 1 Phosphate

Abstract Sickle cell disease (SCD) is a devastating hemolytic genetic disorder associated with high morbidity and mortality. In order to understand the pathogenesis of this disease, we have conducted non-biased metabolomic screening and found that circulating sphingosine-1-phosphate (S1P) was significantly elevated in mice and patients with SCD. S1P is an important bioactive lipid signaling molecule known to regulate inflammation. Our previous study demonstrated that reduced S1P level in plasma and erythrocytes by treatment with sphingosine kinase 1 (SPHK1) inhibitor, PF-543, significantly decreased sickling cells, hemolysis and inflammation in SCD mouse model, which indicated that S1P may play an important role in an SCD complication, especially in inflammation. S1P engages five G-protein coupled receptors known as S1PR1-5. Targeting S1P signaling has been successfully applied in the treatment of the autoimmune disease-multiple sclerosis with the compound named FTY720. In order to understand the roles of S1P/S1PRs signal pathway in pathophysiology of SCD, we treated SCD mice with S1P receptors antagonist FTY720. The results showed that FTY720 successfully inhibited S1P receptors, especially S1P1 expression on immune cells from thymus and lymph node (P<0.05). Circulating white blood cells and inflammatory cytokines, such as CRP, IL-1β, TNF-α and IL-6, also decreased significantly measured by ELSIA kit. Additionally, FTY720 treatment significantly ameliorated organ damage. To investigate the roles of S1P1 receptor in SCD, we treated SCD mice with S1P1 specific antagonist, SEW2871. The results demonstrated that circulating white blood cells and inflammatory cytokines reduced significantly. Histologic studies revealed that the necrosis and congestion of multiple organs including kidney, lung and spleen were substantially reduced by SEW2841.Our studies demonstrate the elevated circulating S1P signaling via its receptor (likely S1PR1) directly contributes to inflammation and multiple tissue damage. Thus, it provides strong evidence that S1P/S1P1 pathway is likely a therapeutic target for SCD. Disclosures No relevant conflicts of interest to declare.

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