Analyse automatisée des liquides de ponction et des liquides céphalorachidiens: Evaluation du Sysmex XE-5000
HO, Thi Thanh Giang ; KEUTGENS, Aurore ; HUWART, Aline et al
in Immuno-Analyse & Biologie Spécialisée [=IBS] (2011), 26
Le mode d’analyse des liquides de ponction du Sysmex XE-5000 a été évalué sur 159 liquides de ponction, dont 60 liquides céphalorachidiens. La numération érythrocytaire et leucocytaire a été comparée aux ... [more ▼]
Le mode d’analyse des liquides de ponction du Sysmex XE-5000 a été évalué sur 159 liquides de ponction, dont 60 liquides céphalorachidiens. La numération érythrocytaire et leucocytaire a été comparée aux comptages réalisés en microscopie en chambre de Fuchs-Rosenthal. La différentiation des cellules monucléées et polynucléées par le XE-5000 a été comparée à la formule manuelle obtenue sur lames de cytocentrifugation. Une bonne corrélation a été obtenue entre le XE-5000 et la technique manuelle avec des coefficients de corrélation supérieurs à 0,8 pour la numération des érythrocytes et des leucocytes ainsi que pour la différenciation leucocytaire pour la plupart des liquides de ponction. La limite de sensibilité fonctionnelle (c’est-à-dire une précision intra-série supérieure à 20 %) a été atteinte pour des numérations érythrocytaires inférieures à 400/ L et leucocytaires inférieures à 30/ L. La contamination inter-échantillons est négligeable et la linéarité est satisfaisante jusqu’à 400 érythrocytes/ L et 30 leucocytes/ L. Ces résultats montrent que l’analyse automatisée des liquides de ponction par le XE-5000 est une alternative acceptable à la technique microscopique de référence aussi bien pour la numération érythrocytaire et leucocytaire que pour la différenciation des cellules polymorphonucléées et mononucléées pour la majorité des liquides de ponction y compris les liquides céphalorachidiens. The body fluid mode on the Sysmex XE-5000 automated hematology analyzer was evaluated on 99 body fluids and 60 cerebrospinal fluids. Erythrocyte and leukocyte numeration was compared to microscopic counts on Fuchs-Rosenthal chambers. Differentiation of mononuclear (MN) and polymorphonuclear (PMN) cells on the XE-5000 was compared to manual differential on cytospin slides. A good agreement was found between the XE-5000 and the manual method with correlation coefficients more than 0.8 for erythrocyte counts, leukocyte counts and leukocyte differentiation in most types of body fluids. The functional sensitivity limit (i.e., within run precision more than 20%) was reached for red blood cell counts more than 400/ L and white blood cell counts more than 30/ L. Carry over was negligible and linearity was adequate for as low as 400 erythrocytes/ L and 30 leucocytes/ L. Our results demonstrate that the automated body fluid analysis on the XE-5000 is an acceptable alternative to the microscopic reference method for erythrocyte and leukocyte numeration as well as MN and PMN differential for most body fluids including cerebrospinal fluids. [less ▲]Detailed reference viewed: 142 (19 ULg)
RhoGDI alpha-dependent balance between RhoA and RhoC is a key regulator of cancer cell tumorigenesis
Ho, Thi Thanh Giang ; ; Dubail, Johanne et al
in Molecular Biology of the Cell (2011), 22(17), 3263-75
RhoGTPases are key signaling molecules regulating main cellular functions such as migration, proliferation, survival, and gene expression through interactions with various effectors. Within the RhoA ... [more ▼]
RhoGTPases are key signaling molecules regulating main cellular functions such as migration, proliferation, survival, and gene expression through interactions with various effectors. Within the RhoA-related subclass, RhoA and RhoC contribute to several steps of tumor growth, and the regulation of their expression affects cancer progression. Our aim is to investigate their respective contributions to the acquisition of an invasive phenotype by using models of reduced or forced expression. The silencing of RhoC, but not of RhoA, increased the expression of genes encoding tumor suppressors, such as nonsteroidal anti-inflammatory drug-activated gene 1 (NAG-1), and decreased migration and the anchorage-independent growth in vitro. In vivo, RhoC small interfering RNA (siRhoC) impaired tumor growth. Of interest, the simultaneous knockdown of RhoC and NAG-1 repressed most of the siRhoC-related effects, demonstrating the central role of NAG-1. In addition of being induced by RhoC silencing, NAG-1 was also largely up-regulated in cells overexpressing RhoA. The silencing of RhoGDP dissociation inhibitor alpha (RhoGDIalpha) and the overexpression of a RhoA mutant unable to bind RhoGDIalpha suggested that the effect of RhoC silencing is indirect and results from the up-regulation of the RhoA level through competition for RhoGDIalpha. This study demonstrates the dynamic balance inside the RhoGTPase network and illustrates its biological relevance in cancer progression. [less ▲]Detailed reference viewed: 44 (13 ULg)
The RhoGDIalpha-dependent balance between RhoA and Rho C is a key regulator of cancer cell tumorigenesis.
HO, Thi Thanh Giang ; Stultiens, Audrey ; Dubail, Johanne et al
Poster (2011)Detailed reference viewed: 26 (7 ULg)
The RhoGDI-dependent balance between RhoA and Rho Cis a key regulator of cancer cell tumorigenesis.
HO, Thi Thanh Giang ; Stultiens, Audrey ; Dubail, Johanne et al
Conference (2011)Detailed reference viewed: 30 (4 ULg)
Analytical validation of the Liaison hGH assay (DiaSorin)
Keutgens, Aurore ; Ho, Thi Thanh Giang ; Cavalier, Etienne
in European NeuroEndocrine Association (2010, September)Detailed reference viewed: 51 (4 ULg)
Corrélation aldostérone et parathormone en cas d'hyperparathyroïdie primaire vs secondaire?
Ho, Thi Thanh Giang ; Keutgens, Aurore ; Chapelle, Jean-Paul et al
in Annales de Biologie Clinique (2010), 68(6), 703-4Detailed reference viewed: 60 (8 ULg)
RhoA-GDP regulates RhoB protein stability. Potential involvement of RhoGDIalpha.
Ho, Thi Thanh Giang ; ; et al
in Journal of Biological Chemistry (2008), 283(31), 21588-98
RhoA plays a significant role in actin stress fibers formation. However, silencing RhoA alone or RhoA and RhoC did not completely suppress the stress fibers suggesting a residual "Rho-like" activity. RhoB ... [more ▼]
RhoA plays a significant role in actin stress fibers formation. However, silencing RhoA alone or RhoA and RhoC did not completely suppress the stress fibers suggesting a residual "Rho-like" activity. RhoB, the third member of the Rho subclass, is a shortlived protein barely detectable in basal conditions. In various cell types, the silencing of RhoA induced a strong up-regulation of both total and active RhoB protein levels that were rescued by re-expressing RhoA and related to an enhanced half-life of the protein. The RhoA-dependent regulation of RhoB does not depend on the activity of RhoA but is mediated by its GDP-bound form. The stabilization of RhoB was not dependent on isoprenoid biosynthesis, Rho kinase, extracellular signal-regulated kinase, p38 mitogen-activated kinase, or phosphatidylinositol 3'-OH kinase pathways but required RhoGDIalpha. The forced expression of RhoGDIalpha increased RhoB half-life, whereas its knock-down antagonized the induction of RhoB following RhoA silencing. Moreover, a RhoA mutant (RhoAR68E) unable to bind RhoGDIalpha was significantly less efficient as compared with wild-type RhoA in reversing RhoB up-regulation upon RhoA silencing. These results suggest that, in basal conditions, RhoGDIalpha is rate-limiting and the suppression of RhoA makes it available to stabilize RhoB. Our results highlight RhoGDIalpha-dependent cross-talks that regulate the stability of RhoGTPases. [less ▲]Detailed reference viewed: 99 (23 ULg)
Establishment of stable human fibroblast cell lines constitutively expressing active Rho-GTPases.
; ; Lambert, Charles et al
in Protoplasma (2006), 229(2-4), 215-20
Small GTP-binding proteins of the Rho family (RhoA, Cdc42, Rac1) regulate the organisation and the turnover of the cell's cytoskeleton and adhesion structures. A significant function of these cellular ... [more ▼]
Small GTP-binding proteins of the Rho family (RhoA, Cdc42, Rac1) regulate the organisation and the turnover of the cell's cytoskeleton and adhesion structures. A significant function of these cellular structures is to translate and counterbalance forces applied to, or generated by, cells in order to maintain homeostasis and control cell movement. We therefore hypothesised that Rho-GTPases are directly involved in cellular gravity perception and may participate in the alterations induced in microgravity. To define an adequate cellular model allowing to investigate this issue, we have established stable cell lines constitutively expressing active forms of either RhoA, Cdc42, or Rac1. The three cell lines differ by morphology and by their ability to form filopodia, lamellipodia, and bundles of actin stress fibers. Overexpression of the active form of either RhoA, Cdc42, or Rac1 is compatible with cell viability and does not affect cell population doubling time. Thus, our series of mutant cells appear well suited to gain further knowledge on the molecular mechanisms of cellular gravity perception. [less ▲]Detailed reference viewed: 48 (4 ULg)
Cdc42 downregulates MMP-1 expression by inhibiting the ERK1/2 pathway.
Deroanne, Christophe ; ; Ho, Thi Thanh Giang et al
in Journal of Cell Science (2005), 118(Pt 6), 1173-83
The small GTPases of the Rho family are key intermediates in cellular signalling triggered by activated cell-adhesion receptors. In this study, we took advantage of RNA interference (RNAi) using small ... [more ▼]
The small GTPases of the Rho family are key intermediates in cellular signalling triggered by activated cell-adhesion receptors. In this study, we took advantage of RNA interference (RNAi) using small interfering RNAs (siRNAs) to define the roles of the best-characterized members of the RhoGTPase family, RhoA, Rac1 and Cdc42, in the control of MMP-1, MMP-2 and type-I-collagen expression in normal human skin fibroblasts (HSFs). A specific and long-lasting repression, up to 7 days after transfection, of the three GTPases was achieved by transient transfection of specific siRNA. The silencing of Cdc42, but not that of RhoA or Rac1, induced a 15-fold increase in MMP-1 secretion. This upregulation was confirmed at the mRNA level and observed with two different siRNAs targeting Cdc42. Such a regulation was also observed in various human cell lines and was rescued by re-expressing wild-type Cdc42 encoded by a construct bearing silent mutations impeding its recognition by the siRNA. By contrast, MMP-2 and type-I-collagen expression was not affected by the individual silencing of each Rho GTPase. Cytokine protein array, enzyme-linked immunosorbent assays and reverse-transcription PCR measurements revealed that ablation of Cdc42 induced an overexpression of interleukin 8 and MCP-1. Although these cytokines are known to induce the expression of MMP-1, we showed that they were not involved in the Cdc42-mediated upregulation of MMP-1. Silencing of Cdc42 also induced an increased phosphorylation of ERK1/2 and p38 MAP kinase. The use of chemical inhibitors on Cdc42-ablated cells revealed that the upregulation of MMP-1 is dependent on the ERK1/2 pathways, whereas the p38 MAP kinase pathway displayed an inhibitory role. Simultaneous knock-down of two or three Rho GTPases allowed us to demonstrate that the RhoA-ROCK pathway was not involved in this regulation but that the silencing of Rac1 reduced the effect of Cdc42 suppression. These data suggest that, in vivo, when cell/extracellular-matrix interactions via integrins induce cytoskeleton organization, MMP-1 expression is maintained at a low level by Cdc42 via a repression of the Rac1 and ERK1/2 pathways. Therefore, Cdc42 contributes to ECM homeostasis and connective tissue integrity. [less ▲]Detailed reference viewed: 61 (6 ULg)
Role of the RhoGTPases in the cellular receptivity and reactivity to mechanical signals including microgravity
Nusgens, Betty ; ; et al
in Journal of Gravitational Physiology : A Journal of the International Society for Gravitational Physiology (2005), 12(1), 269-270
The small G proteins of the Rho Family (RhoGTPases) are key operators in the signaling arising from extracellular matrix through integrin receptors and from membrane receptors for soluble ligands. FLight ... [more ▼]
The small G proteins of the Rho Family (RhoGTPases) are key operators in the signaling arising from extracellular matrix through integrin receptors and from membrane receptors for soluble ligands. FLight data show that microgravity affects cell architecture and gene expression leading us to assume that the signaling pathways(s) involving the RhoGTPAses might disturbed in a weightlessness environment. TO test this hypothesis in microgravityu, we created genetically engineered human fibroblasts that will be used in Biolab on the ISS. [less ▲]Detailed reference viewed: 35 (10 ULg)