Immunophenotypic Lymphocyte Profiles in Human African Trypanosomiasis

Human African trypanosomiasis (HAT) is a deadly vector-born disease caused by an extracellular parasite, the trypanosome. Little is known about the cellular immune responses elicited by this parasite in humans. We used multiparameter flow cytometry to characterize leukocyte immunophenotypes in the blood and cerebrospinal fluid (CSF) of 33 HAT patients and 27 healthy controls identified during a screening campaign in Angola and Gabon. We evaluated the subsets and activation markers of B and T lymphocytes. Patients had a higher percentage of CD19+ B lymphocytes and activated B lymphocytes in the blood than did controls, but lacked activated CD4+ T lymphocytes (CD25+). Patients displayed no increase in the percentage of activated CD8+ T cells (HLA-DR+, CD69+ or CD25+), but memory CD8 T-cell levels (CD8+CD45RA−) were significantly lower in patients than in controls, as were effector CD8 T-cell levels (CD8+CD45RA+CD62L−). No relationship was found between these blood immunophenotypes and disease severity (stage 1 vs 2). However, CD19+ B-cell levels in the CSF increased with disease severity. The patterns of T and B cell activation in HAT patients suggest that immunomodulatory mechanisms may operate during infection. Determinations of CD19+ B-cell levels in the CSF could improve disease staging

Apports diagnostiques au cours de la trypanosomose humaine africaine

La trypanosomose humaine africaine (THA), ou maladie du sommeil est une parasitose due à des trypanosomes du groupe Trypanosoma brucei (T. b.). Ces parasites sont transmis par piqûre d une mouche du genre Glossina, la mouche tsé-tsé. La THA pose un réel problème de santé publique en Afrique noire et Subsaharienne. Deux stades sont classiquement décrits dans l'évolution de la maladie, le stade lymphatico-sanguin et le stade nerveux. Le diagnostic et la détermination du stade, dont dépend le traitement, restent difficiles. En effet, les critères couramment utilisés sont peu sensibles ou peu spécifiques et nécessitent une ponction lombaire avec prélèvement de liquide céphalo-rachidien (LCR). C est pourquoi notre travail de thèse avait pour objectif la recherche de nouvelles approches diagnostiques permettant de déterminer le stade de la THA. Pour cela, nous avons étudié diverses sous-populations de lymphocytes T et B, et les cytokines/chémokines les régulant. Nous avons d abord caractérisé les lymphocytes T régulateurs CD4+CD25+Foxp3+ (nTregs), chez des souris infectées par T. b. brucei. L augmentation des nTreg dans le thymus jusqu à 120 jours d infection puis leur diminution à 240 jours, peuvent être dues à un mécanisme de blocage de maturation dans le thymus ou à un blocage de leur sortie en dehors du thymus. Inversement, dans les ganglions mésentériques, nous avons trouvé une constante diminution des nTregs au cours de l infection. Ces résultats suggèrent que la variation de leur nombre est due à l évolution de l infection, mais est aussi âge-dépendante. Notre travail a ensuite permis de préciser le type de cellules lymphocytaires impliquées dans le sang et dans le LCR de patients atteints de THA. Dans le sang, tous stades confondus, la proportion de lymphocytes B (CD19) augmente alors que celle des lymphocytes T diminue confirmant le caractère immunosuppressif de la maladie. Dans le LCR, l augmentation des cellules B CD19 chez les patients en stade 2, pourrait constituer un nouveau critère du stade nerveux. D autre part nous avons cherché à comprendre quels étaient les mécanismes d attraction des lymphocytes dans le système nerveux central par le dosage dans le sérum et le LCR de différentes cytokines/chémokines. Dans le sérum, les taux de cytokines/chémokines étaient uniquement associés à la présence du trypanosome dans le LCR. Dans le LCR, leur expression était associée avec la présence de signes neurologiques, démontrant leur intérêt pour le diagnostic du stade nerveux. Enfin, nous avons montré que T. b. gambiense peut activer l expression de CXCL-13 dans les lignées cellulaires microgliales et endothéliales, suggérant un rôle direct des trypanosomes dans le processus de régulation. Nos travaux sur les cellules lymphocytaires et les cytokines/chémokines impliquées montrent la nécessité de leur évaluation en tant que marqueurs de stade par des études multicentriques de terrain.LIMOGES-BU Médecine pharmacie (870852108) / SudocSudocFranceBrazilFRB

[Criteria for diagnosis of the neurological stage of human African trypanosomiasis: update and perspectives]

International audienceSleeping sickness or human African trypanosomiasis (HAT) is due to parasite infection by a sanguicolous flagellate protozoan of the Trypanosoma brucei genus. The disease is classically divided into two stages, i.e., the hemolymphatic stage and the CNS stage. Disease staging is currently a major challenge for therapeutic decision-making. In the field, diagnosis is based solely on white blood cell (WBC) count and detection of the parasite in the patient's cerebrospinal fluid (CSF). This technique is unreliable and invasive. Numerous studies are now under way to adapt staging to field conditions and to develop a reliable, low-cost, non-invasive test. This article describes the mechanisms underlying CNS involvement during HAT and reviews the different techniques now being studied to simplify and improve diagnosis of the CNS stage

B cell recruitment processes in the central nervous system provide early diagnostic tools for Trypanosoma brucei gambiense meningo-encephalitis.

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Increased CXCL-13 levels in human African trypanosomiasis meningo-encephalitis.

International audienceSummary Objectives To determine the role of the B-cell attracting chemokine CXCL-13, which may initiate B-cell trafficking and IgM production in diagnosing HAT meningo-encephalitis. Methods We determined CXCL-13 levels by ELISA on paired sera and CSF of 26 patients from Angola and of 16 controls (six endemic and ten non-endemic). Results were compared to standard stage determination markers and IgM intrathecal synthesis. Results CXCL-13 levels in patients' sera had a median value of 386.6 pg/ml and increased levels were associated with presence of trypanosomes in the CSF but not with other stage markers. CXCL-13 levels in patients' CSF had a median value of 80.9 pg/ml and increased levels were associated with all standard stage determination markers and IgM intrathecal synthesis. Conclusion CXCL-13 levels in CSF increased significantly during the course of HAT. Hence the value of CXCL-13 for diagnosis, follow-up or as a marker of disease severity should be tested in a well-defined cohort study

A link between chemokine levels and disease severity in human African trypanosomiasis.

International audienceTrypanosoma brucei gambiense infection is an important public health challenge in sub-Saharan Africa. This parasitic disease is difficult to diagnose due to insidious clinical signs and transient parasitaemias. The clinical course is marked by two stages of increasing disease severity. An early systemic parasitic invasion is followed by the development of a progressive meningo-encephalitis. During this latter stage, a broad spectrum of neurological signs appears, which finally lead to a demyelinating and fatal stage if untreated. Treatment is toxic and difficult to administer when the CNS is invaded. Therefore, accurate diagnostic methods for stage determination are needed. The classically used criteria are not sufficiently specific and mechanisms of parasite invasion through the blood-brain barrier remain poorly understood. As cytokines/chemokines are involved in the early recruitment of leukocytes into the CNS, this study has focused on their potential value to define the onset of CNS involvement. Levels of monocyte chemoattractant protein-1/CCL-2, macrophage inflammatory protein-1alpha/CCL-3, IL-8/CXCL-8, regulated upon activation T cell expressed and secreted (RANTES)/CCL-5 and IL-1beta were measured in paired sera and CSF from 57 patients and four controls. Patients were classified into three groups (stage 1, intermediate and stage 2) according to current field criteria for stage determination (CSF cell count, presence of trypanosomes in CSF and neurological signs). In sera, cytokine/chemokine levels were poorly related to disease stage. Only CXCL-8 was higher in stage 1 patients when compared with stage 2 and CCL-5 was higher in controls when compared with patients. In contrast, in CSF the expression of the selected cytokines, except CCL-5, was associated with the presence of neurological signs, demonstrating their diagnostic value. We observed a relationship between the presence of trypanosomes or trypanosome-related compounds in CSF and levels of IL-1beta, CXCL-8, CCL-2 and CCL-3. These cytokines and chemokines may be triggered by the parasite and hence are potential markers of CNS invasion

IL-1 et chémokines dans le diagnostic de l'invasion du système nerveux central au cours de la Trypanosome humaine africaine.

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Distribution of B cells (CD19⁺), T cells (CD3⁺), and T-cell subpopulations (CD3⁺CD4⁺ or CD3⁺CD8⁺) in the blood of 33 HAT patients and 27 controls (surveys 1 and 2 combined).

<p>For each box plot, the features limits correspond, from bottom to top, to the 90^th, 75^th, 50^th (median), 25^th and 10^th percentiles.</p

Clinical and epidemiological features of the subjects studied.

<p>WBC = White Blood Cell Count in CSF.</p

B cells (CD19<b>⁺</b>) in the CSF of HAT patients and controls.

<p>HAT Patients were at stage 1 (S-1), intermediate stage (S-int) or stage 2 (S-2). B cells (CD 19⁺) are expressed as means±standard deviation of absolute values and as a percentage of total CSF cell count. Note: <i>P</i><0.05 for statistical differences.</p>*<p>S-2 patients <i>vs</i> control.</p>¤<p>S-1 <i>vs</i> S-2 patients.</p>•<p>S-int <i>vs</i> S-2 patients.</p