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International audience; Currently in cancer treatment, one premise is to use antiangiogenic therapies in association with chemotherapy or radiotherapy to augment their efficacy by benefiting from the vascular “normalization” induced by antiangiogenic therapy. This concept defines the time during which the tumor blood vessels adopt normal-like morphology and functionality, i.e. the blood vessels become more mature, the perfusion augments and hypoxia decreases. To date, there is such a diversity of treatment protocols where the type of antiangiogenic to adopt, its dose and duration of administration are different, that knowing when and how to treat is problematic. In this review, we analyzed thoroughly preclinical and clinical studies that use antiangiogenic treatments to benefit from the “normalization” and showed that the effects depend on the type of antiangiogenic administrated (anti-VEGF, anti-VEGFR, Multi-Kinase Inhibitor) and on the duration of treatment. Finally, biomarkers of “normalization” and resistance that could be used in the clinic are presented.

International audience; Glioma initiating cells (GICs) represent a subpopulation of tumor cells endowed with self-renewal and multilineage differentiation capacity but also with innate resistance to cytotoxic agents, a feature likely to pose major clinical challenges towards the complete eradication of minimal residual disease in glioma patients. In this work, GICs were obtained from two patient-derived high-grade gliomas xenograft model, expressing differently EGFR. GICs were exposed to anti-EGFR monoclonal antibody cetuximab during 48h in 1% or 21% oxygen tension. Cell viability and self-renewal capacity were then evaluated as well as their angiogenic properties. GICs were sensitive to cetuximab only in normoxic condition whatever the EGFR status. Nevertheless, under hypoxia cetuximab was able to decrease the self-renewal capacity as well as the expression of CD133 while expression of GFAP increased. Moreover, cetuximab decreased the effect of GICs on endothelial cell migration under hypoxia. Consequently, anti-EGFR therapy can be envisaged to target specifically GICs in order to limit the tumor recurrence.

International audience; Antiangiogenics are widely used in cancer treatment in combination with chemotherapy and radiotherapy for their vascular effects. Antiangiogenics are supposed to induce morphological and functional changes in the chaotic tumor vasculature that would help enhance the therapeutic efficacy of chemotherapy and radiotherapy through the amelioration of the drug delivery or the oxygenation in the tumor, respectively. However, finding the best treatment sequence is not an easy task to achieve and no consensus has yet been established because of the lack of knowledge regarding when and for how long the vascular network is ameliorated. The aim of this work was to develop a dedicated image processing algorithm able to analyze the vascular structures on optical microscopy images of the vascular network and to follow its fine modifications in vivo, over time. We applied this algorithm to follow the evolution of the vascular parameters (vascularized tissue surface, branches, sprouts and length), in response or not to anti-VEGF therapy (10 mg/kg/day) and determine precisely whether there is really a vascular “normalization” with anti-VEGF therapy in comparison with the parameters extracted from healthy vascular networks. We found that for this determination, the choice of region of interest to analyze is critical as it is important to compare only microcirculation areas and avoid areas with arteriole–venule–capillary hierarchy. The algorithm analysis allowed us to define a vascular “normalization” in treated tumors, between 8 and 12 days of bevacizumab treatment that was confirmed by standard immunohistochemical analysis, microvascular permeability assessment and immunohistological blood perfusion assessment.

The idea to develop a blood substitute was stimulated by the need of military in the last two world wars and by transmission of pathogenic germs (Hepatitis B in 1960, HIV in 1980 and Hepatitis C in 1990) during blood transfusion that limited the donor blood transfusion. There are two main groups of blood substitutes: perfluorocarbon emulsions and hemoglobin-based oxygen carriers (HBOC). These latter are of natural origin: human, bovine or recombinant and undergo three modifications types: chemicals (intramolecular cross-linking, polymerisation, conjugation to macromolecules and combination of several chemical modifications), genetics or technological by microencapsulation. HBOCs are in different phases of clinical trials and some of them present side effects (hemodynamic and oxidative). The understanding of these effects and the possibility of correcting them, condition their use on a large scale and the economic consequences, which they can generate.

RESUME Les solutions de remplissage utilisees en therapeutique (albumine, dextranes, gelatines modifiees, hydroxyethylamidons) sont des substituts du plasma. Aucun ne peut assurer le transport de l’oxygene. Or l'importance clinique des globules rouges est telle que depuis plusieurs decennies, ils sont l’objet de nombreuses tentatives pour leur trouver des composes de substitution. Si ces recherches semblaient autrefois utopiques, elles apparaissent aujourd’hui plus realistes puisqu’elles ont abouti a des substituts erythrocytaires potentiels prometteurs, en particulier pour les transporteurs d’oxygene sous forme de solutions d’hemoglobine (Hemoglobin Based Oxygen Carriers ou HBOC). Pourtant depuis 2000 leur developpement se trouve ralenti par l’observation d’effets secondaires multiples lors des essais cliniques : elevation de la pression arterielle induite par un effet vasoconstricteur de l’hemoglobine libre dans le plasma, auto oxydation de l’hemoproteine circulante avec formation de methemoglobine inapte au transport de l’oxygene, generation de radicaux libres responsables d’un stress oxydant.

Human hemoglobin (Hb) conjugated to benzene tetracarboxylate substituted dextran produces a polymeric Hb (Dex-BTC-Hb) with similar oxygen affinity to that of red blood cells ( P 50 =28–29 mm Hg). Under physiological conditions, the oxygen affinity ( P 50 ) of Dex-BTC-Hb is 26 mm Hg, while that of native purified human HbA 0 is 14 mm Hg, but it exhibits a slight reduction in cooperativity ( n 50 ), Bohr effect, and lacks sensitivity to inositol hexaphosphate (IHP), when compared to HbA 0 . Oxygen-binding kinetics, measured by rapid mixing stopped-flow method showed comparable oxygen dissociation and association rates for both HbA 0 and Dex-BTC-Hb. The rate constant for NO-mediated oxidation of the oxy form of Dex-BTC-Hb, which is governed by NO entry to the heme pocket, was reduced to half of the value obtained for HbA 0 . Moreover, Dex-BTC-Hb is only slightly more sensitive to oxidative reactions than HbA 0 , as shown by about 2-fold increase in autoxidation, and slightly higher H 2 O 2 reaction and heme degradation rates. Dextran-BTC-based modification of Hb produced an oxygen-carrying compound with increased oxygen release rates, decreased oxygen affinity and reduced nitric oxide scavenging, desirable properties for a viable blood substitute. However, the reduction in the allosteric function of this protein and the lack of apparent quaternary T→R transition may hinder its physiological role as an oxygen transporter.

AIMS: Glioblastoma (GBM) is the most lethal of all human tumors. Ionizing radiation (IR), as a major therapeutic modality, induces multiple types of DNA lesions in cells, therefore causes cell death. However, IR could also affect neighboring unirradiated cells, inducing Bystander Effects as chromosomal aberrations, increased proliferation, etc. Intercellular communication through the release of different components is involved in the mechanism. GBM cells release different soluble factors as well as tumor microvesicles (TMVs) to modify the phenotype of neighboring cells, thus participating in the tumor progression. The present study was designed to investigate in vitro the impact of IR on the communication between tumor cells and endothelial cells in the tumor microenvironment via soluble factors and TMVs. MATERIALS AND METHODS: Two GBM cell lines (T98G, U87) were grown and sham-irradiated (0Gy) or irradiated (2 or 10Gy) using a Clinac iX linear accelerator. Cell culture media (CM) were collected. Filtrate (containing only soluble factors) and TMVs were separated with successive centrifugations and Pierce concentrator. Cell viability was assessed by cell counting using trypan blue. TMVs quantifications were performed by flow cytometry. The effect of CM/Filtrate/TMVs on the global behavior (proliferation, adhesion) of bystander tumor cells or HUVEC was investigated using the xCELLigence system (ACEA). RESULTS: As expected, irradiation caused a loss of cell number in U87 and T98G: 20% at 2Gy and 60% at 10Gy 48h post-IR as compared to untreated cells. Both CM and Filtrate collected from sham-irradiated tumor cells induced a 50% reduction of bystander tumor cells proliferation, while CM and Filtrate recovered from 10Gy-irradiated cells had different influence on the proliferation: herein, the inhibitory properties of CM were less marked than those of Filtrate. The discordant effects between CM and Filtrate led us to investigate the role of TMVs. When quantified using flow cytometry, two-fold more TMVs were found in 10Gy-irradiated T98G as compared to untreated cells 48h post-IR, while in U87 this stimulation of release was observed at 2h. TMVs were shown to induce a 2-fold increase of HUVEC cell index. Interestingly after 10Gy-irradiation, TMVs from both cell lines slightly also favored both bystander tumor cells and HUVEC proliferation. CONCLUSION: IR inhibits GBM cells proliferation. IR stimulates TMVs release by GBM cells in a time- and line-dependent manner. IR facilitates tumor progression by stimulating the proliferation of bystander tumor cells as well as favoring tumor angiogenesis via TMVs.

BACKGROUND : Hb-based oxygen carriers (HbOCs) have vasoactive effects that are still poorly understood. Factors known to have vasoactive effects, such as plasma, whole-blood viscosity, and the rheologic behavior of RBCs, are modulated by HbOCs in vitro, but few in vivo studies have been performed. STUDY DESIGN AND METHODS : Rabbits were phlebotomized (30%) and resuscitated with unmodified stroma-free Hb (SFHb), dextran-tetracarboxylate-Hb (Dex-BTC-Hb), O-raffinose-polymerized Hb (OrpHb), HSA, or hydroxyethyl starch 200 (HES). Plasma viscosity was assessed with a capillary viscometer and whole-blood viscosity with a rotational viscosimeter. RBC aggregation kinetics were determined by analysis of back-scattered light in a rotating device. RESULTS : As compared to that in the control RBC suspension, resuscitation with SFHb, OrpHb, or HSA decreased plasma and whole-blood viscosity as well as RBC aggregation; resuscitation with Dex-BTC-Hb increased whole-blood viscosity at low shear rates as well as RBC aggregation, whereas that with HES decreased whole-blood viscosity but increased RBC aggregation. CONCLUSION : HbOCs have different rheologic effects in vitro and in vivo. There are marked differences among the Hb solutions in their in vivo effects on viscosity and RBC rheologic behavior (especially at low shear rates encountered in the venous circulation and the microcirculation), which may be related to the chemical modifications applied to hemoprotein. These results could contribute to an understanding of the vasoactive effects of HbOCs.

The in vivo behavior of monomethoxypoly(ethylene oxide)-poly(lactic acid) (MPEO20-PLA45/PLA (75/25)) nanoparticles in comparison with PLA ones was studied in guinea pig. Indeed, the aim of this study was to bring to the fore the in vivo stealth character of these copolymer nanoparticles and to identify the phagocytic circulating cells involved in their uptake. After the intravascular administration of fluorescent nanoparticles (rubrene), their phagocytosis by granulocytes and monocytes was assayed by flow cytometry. At the same time, the evolution of the number of these phagocytic cells was realized in order to identify their function in the nanoparticle uptake. Finally, a histological study of the spleen (30 h after the nanoparticle administration) was investigated to highlight the splenic trapping of these stealth nanoparticles. This study has shown that the phagocytic circulating cells involved in the nanoparticle uptake were mainly neutrophilic granulocytes and some of them were found in the spleen.

The cardiovascular effects of human albumin (Alb) and three human hemoglobin (Hb) solutions, dextran-benzene-tetracarboxylate Hb, αα-crosslinked Hb, and o-raffinose-polymerized Hb were compared in anesthetized rabbits undergoing acute isovolemic hemodilution with Hct reduction from 41.4 ± 2.7 to 28.8 ± 1.6%. The impact of the vasoconstricting properties of Hb was examined by measuring heart rate (HR), mean arterial pressure (MAP), abdominal aortic, and femoral arterial blood flow, vascular resistance (VR), and aortic distension during the first 3 h after hemodilution. The impact of the hemorheological parameters was assessed by measurements of hemodiluted blood viscosity. In contrast to Alb, the Hb solutions elicited an immediate increase in MAP (20–38%). The effects of Alb and Hb solutions on HR, as well as on aortic and femoral arterial blood flow, were similar. VR decreased with Alb (20–28%) and increased with all three Hb solutions (30–90%), but the MAP and VR rising trends were different with each Hb solution. Aortic distension decreased in Hb groups compared with the Alb group for the first 60 min. The viscosity of hemodiluted blood was similar for all groups at high shear rates but was dependent on the viscosity of the solutions at low shear rates. We conclude that the vasoconstriction elicited by the Hb solutions overrides the vasodilation associated with viscosity changes due to hemodilution and would be the major factor responsible to the cardiovascular changes.

Circulating volume expansion for intentional hemodilution and/or resuscitation of hemorrhagic shock can be performed with hemoglobin-based oxygen carriers (HBOC) which, in addition to oxygen transport, have vasoactive effects through poorly documented mechanisms. Among these, the effects of HBOC on red blood cell (RBC) rheology are relatively unknown. The aim of the present in vitro study was to measure the rheological effects of human hemoglobin bound to benzene-tetracarboxylate substituted dextran (Dex-BTC-Hb) as an example of chemically modified hemoglobin. The viscosity was assessed with a capillary and a rotational viscometer for shear rates of 0.5–128 s −1 . Erythrocyte aggregation was determined by analysis of the red light backscattered in a RBC suspension and with a rheoscope. The deformability was determined by the pressure–flow relationship of the RBC suspensions passed through polycarbonate filters. At hematocrit of 0.35 l/l and at low shear rates, the viscosity of RBC was higher in the presence of Dex-BTC-Hb as compared to free Hb, Dex-BTC, Dextran 40 (Plasmacair®), modified fluid gelatin (MFG-Plasmion®) or hydroxyethyl starch (HEA-Elohes®). The effect on erythrocyte aggregation of Dex-BTC-Hb was greater than that of standard solutions, but close to that of MFG or HEA. There was no apparent change in RBC deformability. Dex-BTC-Hb, unlike free Hb, has a hyperaggregating effect on RBC, similar to that of some clinically used volume expanders. This hyperaggregating effect could influence the in vivo rheological behavior of substituted Hb by increasing shear stress.

International audience; Specific phenotypic effects of PTEN in head and neck squamous cell carcinoma (HNSCC) remain poorly defined without a direct causal connection between the loss of PTEN function and the progression of cancer. Here, we describe a potential role for PTEN in cancer progression. Using an shRNA targeting PTEN in HNSCC cells, we show that the loss of PTEN expression is associated with a decrease of cell adhesion, a reduction in E-cadherin expression while cell migration is promoted. Together with the tissue organization and molecular markers expressed in tumors derived from shPTEN cells in vivo, this study indicates that HNSCC cells deficient in PTEN expression undergo an epithelial‑mesenchymal transition (EMT). Additionally, our results suggest that both the low levels of expression and subcellular localization of PTEN are involved in the EMT phenotype, and ultimately in possible locoregional reccurences. We hypothesize that the loss of PTEN expression as well as the subcellular localization could be of interest as a predictive marker of recurrence in HNSCC.

International audience; Driven by a multidisciplinary approach combination (Structure-Based (SB) Three-Dimensional Quantitative Structure-Activity Relationships (3-D QSAR), molecular modeling, organic chemistry and various biological evaluations) here is reported the disclosure of new thienopyrimidines 1-3 as inhibitors of KDR activity and human umbilical vein endothelial cell (HUVEC) proliferation. More specifically, compound 2f represents a new lead compound that inhibits VEGFR-2 and HUVEC at μM concentration. Moreover by the mean of an endothelial cell tube formation in vitro model 2f tartaric acid salt proved to block angiogenesis of HUVEC at μM level.

Resume Les effets vasoactifs de l'hemoglobine-dextrane 10-benzene-tetracarboxylate (Hb-Dex-BTC), transporteur d'oxygene potentiel a base d'hemoglobine modifiee chimiquement, ont ete compares a ceux de solutions de reference, le sang en autotransfusion et une solution d'albumine a 50 g/l, par des mesures de velocite sanguine carotidienne, de pression arterielle moyenne et de frequences cardiaque et respiratoire, dans un modele de choc hemorragique chez le cobaye Hartley anesthesie. L'hemoglobine libre provoque des l'injection une hypertension de 40 % et une augmentation de la velocite sanguine de 110 % par rapport aux valeurs initiales. Trois heures apres l'injection, la velocite sanguine reste encore de 38 % superieure a sa valeur initiale. Le calcul des resistances vasculaires montre un accroissement du tonus carotidien. L'Hb-Dex-BTC induit une hypertension transitoire de 35 %, sans modification significative de la velocite sanguine et des resistances vasculaires. Ses effets vasoactifs sont comparables a ceux de la solution temoin d'albumine. Ces resultats montrent que, contrairement a l'hemoglobine libre, l'Hb-Dex-BTC n'affecte pas significativement le tonus vasculaire. Ceci pourrait etre explique par une moindre interaction de l'hemoglobine modifiee avec les differents facteurs de la regulation du tonus vaculaire.

International audience; Antiangiogenic and vascular disrupting agents are in the current cancer therapeutic armamentarium. A better understanding of the intricate mechanisms ruling neovessel survival within tumors during or after treatment is needed. Refinement of imaging and a growing knowledge of molecular biology of tumor vascularization provide new insights. It is necessary to define suitable methods for monitoring tumor response and appropriate tools to analyze data. This review compares most commonly used preclinical models, considering their recent improvements, and describes promising new approaches such as microfluidics, real-time electrical impedance based technique and noninvasive imaging techniques. The advantages and limitations of the in vitro, ex vivo and in vivo models are discussed. This review also provides a critical summary of emerging approaches using mathematical modeling.

A solution of human hemoglobin bound to benzene tetracarboxylate substituted dextran, whose physicochemical characteristics are defined in part I, was evaluated in vivo as a potential red blood cell substitute. Further experiments show: - the confirmation of a lack of acute toxicity in mice and guinea pigs after injection of 12.5%, 25% and 50% of the blood mass and the absence of death in rabbits having undergone three successive 25% hemorrhagic shocks in three week intervals. A plasma half-life of 9.5 +/- 0.5 hours in 70-75% hemorrhagic shocks on guinea pigs and the absence of dex-BTC-Hb in thoracic and abdominal cavities. No tissue oedema was noticed. Total hemoglobinuria did not exceed 10% of the injected hemoglobin quantity and only involved free hemoglobin. A lack of death in 70-75% hemorrhagic shocks and survival times ranging from 10 hours to 3 days in total exchange transfusions in guinea pig experiments.

Overexpression of EGFR plays a key-role in head and neck squamous cell carcinoma (HNSCC) and justifies the extensive use of cetuximab, a monoclonal anti-EGFR antibody, as well as EGFR-tyrosine kinase inhibitors (EGFR-TKI), which have been reported to inhibit tumor cell growth and the secretion of pro-angiogenic factors by tumor cells, such as VEGF and IL-8. Moreover, vessel normalization in tumors, suggesting a more complex mediation of endothelial cell growth control has also been observed in vivo. The present study was designed to investigate the angiogenic consequences of exposure of HNSCC tumor cell lines to cetuximab and intercellular signaling between tumor and endothelial cells by secretion of pro- and anti-angiogenic mediators in the conditioned media (CM). The results achieved showed that cetuximab decreased the secretion of VEGF by HNSCC cells and that exposure of human umbilical vein endothelial cells (HUVEC) to CM from HNSCC cells exposed to cetuximab induced an increase in endothelial cell network formation. Angiogenesis proteome profiling showed that cetuximab induced a complex alteration of the secretion of pro- and anti-angiogenic factors by HNSCC cells without enabling to identify a unique molecular marker. Expression of endothelial membrane receptors (VEGFR-2, EGFR, PECAM-1 and Notch-4) was investigated and only EGFR expression was found influenced when HUVEC were exposed to CM from cetuximab-exposed HNSCC cells. These results showed that the decrease in the secretion of pro-angiogenic agents like VEGF by HNSCC cells exposed to cetuximab could not be sufficient to justify its anti-angiogenic activity in vitro.

The implication of loss of PTEN expression in resistance to targeted therapy has already been described in many tumor types. The absence of response to anti-EGFR agents in PTEN-deficient tumors relies on persistent activation of signaling pathways downstream of pEGFR. To investigate the role of PTEN loss of expression in head and neck squamous cell carcinoma (HNSCC) response to cetuximab, we used siRNA in Cal 27 cells and then evaluated key signaling protein activation (pAKT and pERK 1/2) as well as cell viability and proliferation. PTEN silencing in Cal 27 cells led to a constitutive activation of signaling pathways evidenced by a strong increase in pAKT and pERK 1/2 expression. Moreover, PTEN-silenced cells did not show any significant changes either in cell viability or proliferation, only slight modifications on cell cycle. Additionally and unpredictably, our results indicated that PTEN silencing, led to a drastic reduction in pEGFR expression whereas total EGFR level did not significantly vary. Strikingly, despite this overactivation of signaling pathways ruling cell survival and proliferation in siPTEN cells, cetuximab fully exerted pAKT and pERK 1/2 inhibition of expression, similarly to its effect in untransfected Cal 27 cells. In conclusion, our study established that in Cal 27 cells, cetuximab keeps full ability to inhibit EGFR-dependent mechanisms, as shown by a decreased pAKT and pERK 1/2 level of expression, despite a strong PTEN silencing-induced overactivation. In Cal 27 cells, loss of PTEN expression does not lead to a loss of cetuximab efficacy in inhibiting EGFR-downstream signaling pathways, contrarily to data shown in previous works conducted in other tumor types.

International audience; Purpose : This article characterizes the vascular effects following vascular-targeted photodynamic therapy with a photosensitizer which actively targets endothelial cells. Methods : This strategy was considered by coupling a chlorin to a heptapeptide targeting neuropilin-1 in human malignant glioma-bearing nude mice. A laser Doppler microvascular perfusion monitor was used to monitor microvascular blood perfusion in tumor tissue. Endothelial cells' ultra structural integrity was observed by transmission electron microscopy. The consequences of photosensitization on tumor vessels, tissue factor expression, fibrinogen consumption, and thrombogenic effects were studied by immunohistochemical staining. Results : Treatment of glioma-bearing mice with the conjugate showed a statistically significant tumor growth delay. Vascular effect was characterized by a decrease in tumor tissue blood flow at about 50% baseline during treatment not related to variations in temperature. This vascular shutdown was mediated by tumor blood vessels' congestion. A pro-thrombotic behavior of targeted endothelial cells in the absence of ultra structural changes led to the induction of tissue factor expression from the earliest times post-treatment. Expression of tissue factor-initiated thrombi formation was also related to an increase in fibrinogen consumption. Conclusion : Using a peptide-conjugated photosensitizer targeting neuropilin-1, induction of tissue factor expression immediately post-treatment, led to the establishment of thrombogenic effects within the vessel lumen.

International audience

Volume loading solutions used therapeutically (albumin, dextrans, modified gelatins and hydroxylstarches) are simple plasma substitutes and cannot ensure oxygen transport. The search for erythrocyte substitutes initially seemed utopian, but this goal is now within our reach, in the form of hemoglobin-based oxygen carriers (HBOC) for example. The clinical development of HBOC has been slowed by adverse effects, including an increase in arterial pressure due to the vasoconstrictive effect of cell-free hemoglobin in plasma, haemoprotein autoxidation leading to methemoglobin formation, and free radical generation that creates oxidative stress.

Background The hypertension induced by haemoglobin-based oxygen carriers could be a result of different pharmacological and physicochemical factors. Objective To investigate whether production of superoxide anion (O 2 ·- ) and release of endothelin could be the factors responsible. Methods We studied the variation in mean arterial pressure (MAP) in guinea pigs by carrying out a 50% isovolaemic exchange transfusion with conjugated oxyhaemoglobin (non-oxidized form) or conjugated methaemoglobin (fully oxidized form) in the presence or absence of BQ-788 (5 nmol/l), an endothelin receptor type B (ETR-B) antagonist. At key timepoints of variation in MAP, the plasma concentrations of O 2 ·- were measured. The presence of conjugated oxyhaemoglobin and increases in ETR-B concentrations inside the vascular wall were investigated in different vessels, using western blotting. Results We found that the administration of conjugated oxyhaemoglobin induced a significant increase in MAP, whereas conjugated methaemoglobin had no significant haemodynamic effect. Pretreatment with BQ-788 attenuated the increase in MAP induced by conjugated oxyhaemoglobin. This haemoglobin induced the production of high concentrations of O 2 ·- that declined towards control values after 120 min and decreased in the presence of BQ-788. Western blot analysis showed that the presence of conjugated oxyhaemoglobin inside the vascular wall was time-dependent and correlated with increased ETR-B. Conclusion These results show that the release of O 2 ·- during auto-oxidation of conjugated oxyhaemoglobin is associated with the observed increase in MAP, which may be a result of the vasoconstriction caused by an increase in activation of ETR-B. This activation may be caused by the massive release of endothelin induced by the production of O 2 ·-.

Haemoglobin initiates free radical chemistry. In particular, the interactions of peroxides with the ferric (met) species of haemoglobin generate two strong oxidants: ferryl iron and a protein-bound free radical. We have studied the endogenous defences to this reactive chemistry in a rabbit model following 20% exchange transfusion with cell-free haemoglobin stabilized in tetrameric form [via cross-linking with bis-(3,5-dibromosalicyl)fumarate]. The transfusate contained 95% oxyhaemoglobin, 5% methaemoglobin and 25 μM free iron. EPR spectroscopy revealed that the free iron in the transfusate was rendered redox inactive by rapid binding to transferrin. Methaemoglobin was reduced to oxyhaemoglobin by a slower process (t1/2=1 h). No globin-bound free radicals were detected in the plasma. These redox defences could be fully attributed to a novel multifunctional role of plasma ascorbate in removing key precursors of oxidative damage. Ascorbate is able to effectively reduce plasma methaemoglobin, ferryl haemoglobin and globin radicals. The ascorbyl free radicals formed are efficiently re-reduced by the erythrocyte membrane-bound reductase (which itself uses intra-erythrocyte ascorbate as an electron donor). As well as relating to the toxicity of haemoglobin-based oxygen carriers, these findings have implications for situations where haem proteins exist outside the protective cell environment, e.g. haemolytic anaemias, subarachnoid haemorrhage, rhabdomyolysis.

The hemoglobin-based oxygen carriers (HBOC), like dextran-benzene-tetracarboxylate-hemoglobin (Dex-BTC-Hb), which are present at high concentrations in plasma disturb arterial pressure and induce hypertension. To study if the increase of mean arterial pressure (MAP) is due to the presence of cell-free hemoglobin (Hb) inside abdominal aortic wall, we followed on a model of 50% isovolemic exchange transfusion (IET) in anesthetized guinea pigs, the kinetic of Dex-BTC-Hb distribution inside abdominal aortic wall and we investigated the relationship between arterial pressure modifications and modified Hb distribution. The administration of Dex-BTC-Hb induced instantaneously an increase of MAP that reached its maximum (53% of hypertension from baseline) at 17 min after the end of the IET and was maintained maximally up to 30 min. A significantly decrease of MAP (45% of hypertension from baseline) was observed after 60 min and the baseline level was recovered at 180 min. The investigation of tissue at 17 min by confocal microscopy showed the presence of free Hb in or upon endothelial cells (EC) in intima and in vasa vasorum. At 180 min, the free Hb was found in or upon EC and inside all abdominal aortic wall meanwhile MAP recovered its basal value. These results suggest for the first time that Hb in intima seems to induce the hypertension observed upon IET but can not sustain it even if Hb stayed present in intima and in abdominal aortic wall.

Infusion of hemoglobin-based oxygen-carrying solutions (HBOCs) produce an immediate rise in blood pressure with most solutions, both in animals and humans, as a result of systemic and pulmonary vasoconstriction. Autoregulation of the O2 supply by the microvasculature has been proposed as a phenomenon involved in the vasoconstriction elicited by HBOCs. Nevertheless, little is known about the ability of various HBOCs to induce constriction in the microcirculation according to their specific physicochemical properties (viscosity, molecular weight, P50, etc.). This study was therefore designed to assess the effects of three HBOCs, that is, bis(3.5-dibromosalicyl) fumarate-crosslinked hemoglobin (alphaalpha-Hb), dextran-benzene-tetracarboxylate-conjugated hemoglobin (Hb-Dex-BTC) and o-raffinose-oligomerized hemoglobin (o-raffinose-Hb), on the vascular tone of rat mesenteric arterioles (diameter, 15-25 microm) viewed microscopically in moderate hemodilution conditions. The effects of HBOCs were compared to those elicited by a reference solution of hydroxyethyl starch (HES-200) infused in the same conditions. In each experimental group, a fall in arteriolar diameter was observed 2 min and 5 min after infusion of the solution. The maximum changes were observed in Hb-Dex-BTC and o-raffinose-Hb groups, in which diameter decreased from 6.9 +/- 0.5% and 5.2 +/- 0.7%, respectively, 2 min after infusion. The changes in arteriolar diameter induced by Hb-Dex-BTC and o-raffinose-Hb were significantly higher than those elicited by HES-200 and alphaalpha-Hb. In conclusion, our data indicate that moderate hemodilution with HBOCs induces instantaneous constriction in rat mesenteric arterioles, with amplitudes depending on both pharmacological and physicochemical properties of the hemoglobin solution infused.

Hb-based O2-carrying solutions (HbOCs) have been developed as red blood cell substitutes for use in patients undergoing hemodilution. Variously modified Hb with diverse solution properties have been shown to produce variable hemodynamic responses. We examined, through pulsed-Doppler velocimetry, the systemic and renal hemodynamic effects of dextran-benzene-tetracarboxylate-conjugated (Hb-Dex-BTC), bis(3,5-dibromosalicyl)fumarate cross-linked (αα-Hb), and o-raffinose-polymerized ( o-raffinose-Hb) Hb perfused in rabbits after moderate hemodilution (30% hematocrit), and we compared the effects of these Hb solutions with the effects elicited by plasma volume expanders. In addition, vascular hindrance (resistance/blood viscosity at 128.5 s− 1) was calculated to determine whether a moderate decrease in the viscosity of blood mixed with HbOCs may impair vasoconstriction as a result of autoregulation after infusion of cell-free Hb. No changes were observed in renal hemodynamics after hemodilution with reference or Hb solutions. Increase in blood pressure and vascular resistance was found with Hb-Dex-BTC and αα-Hb (for 180 min) and, to a lesser extent, with o-raffinose-Hb (for 120 min). Furthermore, Hb-Dex-BTC (high viscosity) and o-raffinose-Hb (medium viscosity) induced comparable increases in vascular hindrance (from 0.091 to 0.159 and from 0.092 to 0.162 cm− 1, respectively) but far less than that produced by αα-Hb (low viscosity, from 0.092 to 0.200 cm− 1). These results suggest that maintaining the viscosity of blood by infusing solutions with high viscosity makes it possible to limit vasoconstriction due to autoregulation mechanisms and mainly caused by hemodilution per se.

Hemoglobin solutions are potential resuscitative fluids with volume expanding and oxygen delivery abilities developed to reduce the use of blood transfusion. Most hemoglobin solutions in clinical trials increase transiently arterial pressure by inhibiting nitric oxide-dependent vasodilation. Our objective was to compare the effects on central hemodynamics and carotid blood flow of two hemoglobin solutions after resuscitation from hemorrhage in anesthetized guinea pigs. After anesthesia and instrumentation, severe hemorrhage was induced by withdrawing 50% of the blood volume. Resuscitation was performed after 15 min of hypovolemia with 5% albumin, stroma-free hemoglobin, or hemoglobin conjugated to dextran-benzenetetracarboxylate (Dex-BTC-Hb). The mean arterial pressure (MAP), carotid blood flow (CBF), vascular resistance index and heart rate (HR) were monitored for 3 hours after resuscitation. After hemorrhage, MAP and CBF dropped to 57.6 +/- 4.4% and 58.9 +/- 3.7% of control values respectively. Albumin failed to maintain hemodynamics in the decompensatory phase of shock. Both hemoglobin solutions gave rise to a transient increase in MAP (35%); stroma-free hemoglobin increased the CBF (150%) and resistance index (24%) whereas Dex-BTC-Hb had no effect on CBF and vascular resistances. None of the solutions affected the HR. Modified hemoglobin has attenuated effects on CBF and resistance index compared to stroma-free hemoglobin. This may be due to a balance between the stimulation of nitric oxide synthesis by shear-stress and the inhibition of vasodilation by nitric oxide trapping.

The endothelium is the production site of several potent vasoactive factors that contribute to the modulation of the vascular tone. Because hemoglobin-based oxygen carriers (HBOC) have been demonstrated to cause vasoconstriction and thereby increase arterial pressure by interacting with endothelium-derived factors such as nitric oxide and endothelin-1, we hypothesized that hemoglobin could penetrate into the endothelial cells. Therefore, we investigated the presence of hemoglobin into guinea pig aortic endothelial cells by immunohistochemical staining after exchange transfusion with a hemoglobin-based oxygen carrier. Despite the large molecular size of HBOC due to chemical modifications designed to prevent hemoglobin subunit dissociation and extravascular leakage, hemoglobin was detectable by immunohistochemical staining into the endothelial cells. These findings suggest that the vascular endothelial cells could uptake hemoglobin by endocytosis mechanisms or could help hemoglobin to cross the endothelial barrier toward media by transcytosis mechanisms. These findings are very important to lead future investigations to the mechanisms by which HBOC cause vasoconstriction.

The amount of circulating methemoglobin in healthy humans is the result of a balance between methemoglobin production (from autooxidation and oxidation) and hemoglobin reduction. Hemoglobin autooxidation and oxidation are very complex and are not well understood. This article analyses the literature on hemoglobin autooxidation, oxidation and reduction and sets out a sequence of reactions for the oxidation of hemoglobin and the ways in which the percentage of methemoglobin is regulated or methemoglobin production prevented. Most of the information concerns erythrocyte hemoglobin, but plasma extracellular hemoglobin (from hemolysis or hemoglobin-based blood substitutes) is also considered where possible.

While conducting pharmacological investigations into oxygen carriers, it is important to study the in vitro and in vivo rheological behavior of blood cells in the presence of such preparations. With regard to the original nature of human hemoglobin bound to benzene tetracarboxylate substituted dextran (Dex-BTC-Hb), it seemed necessary to study its rheological effect in a simulated in vitro hemorrhagic shock compensated by a blood substitute. The viscosity of substitutes was determined as well as several rheological parameters after 0, 3 and 6 hours incubation periods of red blood cells with substitutes: viscosity of blood-substitute mixtures at different levels of plasma substitution erythrocyte aggregation of blood-substitute mixtures by determining the velocity of rouleau formation and the cohesion of rouleau network. This work yielded several observations: The viscosity of Dex-BTC-Hb was slightly higher than those of solutions of native Hb, Dex-BTC T10, Dextran 40 (Plasmacair, modified fluid gelatin (Plasmion and hydroxyethyl starch 200 (Elohes). The substitution of a blood volume with Dex-BTC-Hb, corresponding to a compensated 45% hemorrhagic shock, slightly increased the viscosity of hemodiluted blood as compared to other substitutes. In the presence of Dex-BTC-Hb, the aggregation of erythrocytes appears to be increased as compared to standard solutions. Yet, the effect was close to that of Plasmion or Elohes.

With the aim of assessing of dextran-benzene-tetracarboxylate hemoglobin as an oxygen carrier, we studied histological changes in the intestinal loop in anesthetized guinea pig. The intestinal tissue being very sensitive to hypoxia, an innervated loop was vascularly perfused with open-flow during one hour at zero hematocrit. To estimate the capacity of hemoglobin solution to oxygenate this tissue, we observed the mechanical and histological changes in the organ and the arterio-venous difference in PO2, oxyhemoglobin, deoxyhemoglobin and we compared them with human albumin, Tyrode and non-modified hemoglobin. The PO2 arteriovenous differences were 51.9 +/- 7.1 torr (m +/- SEM) for Tyrode, 40.2 +/- 6.4 torr for albumin solution, 113.7 +/- 6.5 torr for non-modified hemoglobin and 132.7 +/- 6.8 torr for dex-BTC-Hb. Compared to albumin and Tyrode solutions, hemoglobin solutions transferred more oxygen to tissues. The desaturation of dex-BTC-Hb was significantly superior (p0.05) to the one non-modified hemoglobin. With Hb solutions, this desaturation increased with time and it depended on the perfusion flow. The structure of jejunal villi when perfused with a hemoglobin solution, remained almost normal and the loop was still active. Nevertheless, non-modified hemoglobin leaked from the vessels to the lumen and caused edema and a rupture of overlapping epithelium at the tip of the villi. With dex-BTC-Hb, such histological modifications were less significant. With albumin and Tyrode, all villi were totally necrosed and the loop was completely inert. We have demonstrated that dextran-benzene-tetracarboxylate hemoglobin had the ability to maintain the tissue alive thanks to its good capacity to release oxygen and its satisfactory vascular persistence. Dex-BTC-Hb solution can answer to needs of tissue.

In 1990, McGown demonstrated in vitro a limitation of extracellular methemoglobin (metHb) formation by releasing and recycling of ascorbic acid by red blood cells. In order to investigate the autoxidation of free or modified hemoglobin in plasma and the possibility of reproducing McGown's phenomenon in vivo, we performed a 50% blood mass exchange in guinea-pigs with a 70 +/- 5 g/l dex-BTC-Hb solution (metHb5%). Methemoglobin was determined according to Evelyn-Malloy's method. We observed a clear but limited oxidation of plasmatic hemoglobin (MetHb approximately 30-40% at t = 12 hrs up to t = 24 hrs). A similar blood mass exchange was performed with the same hemoglobin solution which was previously totally oxidized into metHb. 40% of this methemoglobin was found to be reduced after 12 hrs. These results demonstrated a marked reducing activity by residual blood as shown by others. The addition of potentially protective compounds such as ascorbic acid (non enzymatic intraerythrocytar reduction pathway), methylene blue or riboflavin (enzymatic intraerythrocytar pathway), allowed a significant drop in the methemoglobin level. On the contrary, we didn't observe any reducing effect with reduced glutathione.

Histocompatibility assessment of 70 g/l stroma-free hemoglobin solutions pyridoxylated or not, and purified or not, was carried out using vascular perfusion of the intestine of rats. Mechanical and ultrastructural changes in the organ and the arteriovenous difference of PO2 were compared to those obtained with albumin, gelatin and dextran. Overlapping epithelium conserves its structure in the presence of hemoglobin, whereas it is partially or totally destroyed with the plasma substitutes. Nevertheless, with non pyridoxylated hemoglobin there is a strong detachment of the epithelium from the lamina propria. The intestine, irrigated by the hemoglobin solutions shows efficient peristalsis, but this totally disappears with plasma substitutes. For similar arterial PO2, for all the solutions, the arteriovenous difference was of 100 mmHg for the hemoglobin solutions, whereas they never exceeded 60 mmHg for the plasma substitutes. Hemoglobin pyridoxylation led to an arteriovenous difference significantly superior (p less than 0.001) to those obtained using non modified hemoglobin. With their O2 supply hemoglobin solutions appear able to limit the development of hypoxia in the tissue. The continuation of peristalsis and the weak ultrastructural modifications confirm the slight histological improvement gained when using pyridoxylated hemoglobin. Nevertheless an extravasation appeared constantly, as well as flow reduction during perfusion with the hemoglobin solutions.

Free hemoglobin (Hb) present at high concentration in plasma--in case of hemolysis, hemoglobin based oxygen carrier (HBOC) administration or in case of subarachnoid hemorrhage--induce hypertension as result of vasoconstriction. In this context, we studied on an exchange transfusion (ET) model at 50% of hematocrit with a HBOC, the distribution of this Hb inside abdominal aortic wall in guinea pigs in relation with mean arterial pressure (MAP) evolution. MAP was monitored during 180 min after ET and rings of abdominal aorta were taken at different times, when modifications of MAP were important, and analyzed by immunohistochemistry and confocal microscopy. Gelofusine 4%, used as control, did not modify MAP while free Hb increased MAP that reached its maximum (53% of hypertension) at t=17 min after the end of ET. MAP started to decrease (45% of hypertension) at t =60 min after ET, and recovered its baseline value at t=180 min. Confocal analysis of the vessel showed that: at 17 min (when hypertension was maximal), free Hb was present in endothelial cells (EC) and in vasa vasorum; at t=60 min (when hypertension decreased) and at t =10 min (when hypertension disappeared), free Hb was detected still in EC, but inside all abdominal aorta wall too. These results suggest in the first time that free Hb could induce a hypertension by direct interaction with EC but would also be unable to maintain this hypertension in spite of its massive tissue distribution.

Le retour veineux est un élément essentiel du système cardiovasculaire, il permet le maintien du débit cardiaque. Son altération est de plus en plus fréquente dans les pays industrialisés et entraine des symptômes variés qui, s'ils ne sont pas pris en charge, peuvent aller jusqu'à l'ulcère variqueux ou vers des complications graves comme la thrombophlébite profonde et l'embolie pulmonaire. Il existe néanmoins des moyens pour protéger son retour veineux, en particulier l'activité physique et la compression. La course à pied est pratiquée par un français sur trois, elle permet de mobiliser tous les muscles et les articulations des membres inférieurs, d'améliorer la capacité du triceps sural et de la semelle plantaire à pomper le sang du bas du corps vers le coeur droit. La compression est utilisée dans le traitement de tous les stades de l'affection veineuse, souvent confondue avec la contention, notamment à cause du terme « bas de contention », elle applique une pression constante sur le membre inférieur, au repos ou à l'effort. Bien qu'ils soient initialement conçus pour traiter et prévenir la maladie veineuse, les vêtements de compression sont de plus en plus retrouvés chez les coureurs de fond. De nombreuses études ont été menées sur les effets du port de compression pendant et après l'effort mais les protocoles étaient très hétérogènes et les résultats discordants. Pourtant, grâce à quelques méta-analyses, nous avons pu distinguer les effets sur les performances et sur les paramètres physiologiques. Ces effets, dans l'ensemble, n'étaient pas très convaincants. Ceux sur la récupération, notamment sur la diminution des douleurs musculaires d'apparition retardée (DOMS) étaient intéressants mais présentaient peu de changements sur les produits métaboliques. Forts de ces découvertes, nous avons mené une enquête sur l'usage de la compression par des coureurs de tout niveau, afin de comprendre pourquoi et comment ils utilisaient la compression. L'étude nous a permis de constater l'ampleur de l'utilisation des vêtements de compression. Les sportifs portaient la compression majoritairement pendant l'effort, et cela même s'ils recherchaient en particulier l'amélioration de la récupération. La préservation du retour veineux était un effet recherché particulièrement par les femmes et les athlètes masters, pourtant le risque de développer une affection veineuse est de 20% dans la population générale. Le pharmacien, en tant que professionnel de santé de proximité, est amené dans sa pratique quotidienne à rencontrer à la fois des sportifs en quête de performances ou des patients à risque de développer une maladie veineuse qui souhaitent pratiquer la course à pied. Pour ces deux types de patients, il pourrait, grâce à ses connaissances sur la compression médicale, conseiller des vêtements de compression adaptés à leurs pratiques. Le pharmacien d'officine pourrait alors prendre un rôle dans l'accompagnement des bonnes pratiques sportives.

Le maintien à domicile de la personne âgée est un enjeu de santé publique important. Le pharmacien d'officine est un professionnel de santé de proximité pour ses patients : il a ainsi un rôle important en conseil en plus de son expertise sur le matériel médical. Dans un premier temps, nous ferons un rappel épidémiologique sur la population âgée en France ainsi que sur les effets du vieillissement sur l'autonomie. Ensuite, nous présenterons les différents professionnels et acteurs du maintien à domicile. Après cela, nous étudierons via une enquête de terrain comment les pharmaciens d'officine de la Meuse abordent le domaine du maintien à domicile de la personne âgée dans leur pratique professionnelle. Une présentation des matériels et autres aides techniques disponibles à l'officine ainsi que leur règlementation (prescription, prise en charge) sera exposée. Enfin, des perspectives d'avenir pour optimiser le rôle du pharmacien d'officine dans la prise en charge du patient en situation de maintien à domicile seront présentées.

Avec l'avènement des AOD à la fin des années 2000, le choix en terme d'anticoagulation s'est trouvé subitement diversifié après plusieurs décennies d'hégémonie des antagonistes de la vitamine K et des héparines. Ce changement brusque a nécessité une refonte des habitudes de prescription et de suivi. Environ 10 ans après, nous pouvons faire le bilan des évolutions en terme de recommandations de bonnes pratiques. Les évaluations des différentes spécialités et recommandations de prise en charge émis par les autorités de santé ont plusieurs fois été bouleversées avant d'aboutir à l'équilibre actuel. L'avènement de nouvelles indications entrainera sans aucun doute de nouveaux bouleversements … Le rôle du pharmacien a évolué en parallèle, avec une implication de plus en plus forte dans l'éducation du patient et depuis peu la possibilité d'acte plus « techniques ». A voir si sa place dans le suivi des patients anticoagulés évoluera dans l'avenir pour tendre vers le modèle Québécois où le pharmacien d'officine gère les prescriptions et le suivi en collaboration étroite avec le reste de l'équipe médical. Cette multiplication des choix entraine aussi un questionnement : quelle est la molécule la plus adéquate pour un patient donné ? Ou plutôt : quelle molécule sera la mieux tolérée ? Ce questionnement est d'autant plus essentiel chez les populations les plus fragiles peu représentées dans les essais cliniques.

Les EHPAD sont des structures accueillant de plus en plus de personnes âgées polymédiquées. La gestion de leur traitement, depuis la prescription par les médecins jusqu'à l'administration des médicaments par le personnel de soin ne possédait, jusqu'il y a peu, aucun professionnel de santé encadrant ce circuit du médicament. La loi HPST parue en 2009 a introduit une nouvelle mission pour le pharmacien en officine : le pharmacien référent en établissement médico-social sans PUI. Le pharmacien référent est le gestionnaire du circuit du médicament dans ces établissements. Il veille à la sécurisation de ce circuit et participe activement à la prise en charge médicamenteuse du résident. Le circuit du médicament en EHPAD comprend plusieurs étapes : la prescription, la dispensation des médicaments, leur livraison à l'établissement, la préparation des doses à administrer, le stockage des médicaments et leur administration aux résidents. Ces étapes font intervenir plusieurs professionnels de santé. La plupart des EHPAD présentent de grandes difficultés dans la gestion du circuit du médicament avec de nombreux problèmes mis en avant : les ruptures dans les traitements, la nécessité de retranscription des ordonnances dans le logiciel de soin de l'établissement, les difficultés de communication interprofessionnelles, etc. Les missions du pharmacien référent résumées dans « La politique du médicament » écrit en 2013 par Philippe Verger permettent de répondre aux problèmes rencontrés par les EHPAD. Il conseille également de développer l'optimisation du système informatique des EHPAD afin de favoriser la sécurisation du circuit du médicament. Le pharmacien référent s'avérant un atout majeur pour un établissement médico-social, l'EHPAD de Ligny en Barrois a signé une convention avec le pharmacien titulaire d'une de ses pharmacies dispensatrices pour qu'il devienne pharmacien référent. Celui-ci a procédé à de nombreuses améliorations du circuit du médicament en collaboration avec les autres professionnels de santé. Il a également permis le développement de nouveaux outils de sécurisation : les interfaces informatisées. Ces interfaces, lien entre les logiciels des différents professionnels de santé externes à l'EHPAD et le logiciel de soin de l'établissement sont la réponse à de nombreux problèmes rencontrés dans le développement du circuit du médicament et au défaut de communication entre ces intervenants.

Thèse confidentielle; De 1850 à nos jours, la consommation de glucose a considérablement augmenté. Ceci s'explique principalement par l'omniprésence de glucose dans l'alimentation et le rapport que l'être humain a vis-à-vis de ce dernier. Cette surconsommation provient essentiellement d'habitudes alimentaires ou d'un comportement addictif visant à stimuler le système de récompense ou à atténuer une situation de stress. La surcharge en glucose entraîne dans l'organisme un mésusage cellulaire et des complications qu'il convient de prévenir et de traiter. Cette situation pathologique crée un terrain propice au développement de nombreuses pathologies. De l'infection virale, bactérienne, mycologique, en passant par le syndrome métabolique, le diabète et ses complications jusqu'aux cancers : cet apport excessif nocif concerne tout l'organisme. La découverte de nouvelles voies métaboliques pathologiques telles que la voie des polyols, de la glycation ou encore celle de la voie de la protéine kinase C, a permis l'émergence de nouvelles molécules voire l' approfondissement des propriétés de certaines bien connues. Toutefois, l'alimentation semble être la solution thérapeutique première, comme le soulignait Hippocrate. Bien que des mesures hygièno-diététiques soient recommandées, il nous est de plus en plus difficile de les respecter en raison de la surexposition au glucose. Nous pouvons alors nous interroger sur le rôle de l'industrie agro-alimentaire dans cette surexposition et les moyens mis en oeuvre par le gouvernement pour faire face à ce fléau.

La prise en charge par orthèses du petit appareillage des pathologies du rachis thoraco-lombal − aiguës ou chroniques, bénignes ou graves, communes ou symptomatiques,… − appartient au domaine de compétence du pharmacien orthésiste titulaire du diplôme universitaire d'orthopédie. La thèse recense les possibilités et les moyens d'intervention, par orthèses, du pharmacien dans les pathologies thoraciques et lombales les plus fréquemment rencontrées à l'officine. Sont tout d'abord rappelées les notions anatomiques ndispensables à connaître afin de comprendre la physiopathologie. Après un exposé de l'aspect réglementaire, est abordée l'étude des différentes orthèses qui comprennent : les ceintures lombaires de série, les corsets de série, les orthèses thoraciques et les redresse-dos. Un recensement exhaustif et précis de ce type d'orthèses présentes sur le marché a été effectué afin de présenter toutes les possibilités thérapeutiques orthètiques existantes. Enfin sont abordés le rôle du pharmacien et sa capacité à répondre aux attentes des patients, du corps médical et des organismes de santé publique dans le domaine de l'orthopédie. Il résulte de ce travail l'élaboration d'une méthode de dispensation des ceintures lombaires de série destinée à guider, d'une façon claire et précise, l'orthésiste dans sa pratique quotidienne. Ainsi, cette thèse a cherché à donner les moyens au pharmacien orthésiste de répondre aux attentes de santé publique et d'obtenir de tous la reconnaissance de ses compétences.

The main aim of this work was to characterize the effects of the anti-VEGF Bevacizumab (Avastin) on the tumor vascular network, in vivo, over time, thanks to the skin fold chamber model on the nude mouse. Images of the vascular network obtained using intravital microscopy were analyzed par a dedicated image processing algorithm developed within our research team, allowing to highlight the morphological modifications induced by the treatment and to isolate discriminating parameters of the vascular "normalization", by comparison to healthy vascular networks. Le vascular "normalization" period detected with our tool was comforted by the analysis of the functionality of the blood vessels over time, in vivo and by an immunohistochemical analysis of the blood vessels and of the tumor tissue. In preliminary in vivo experiments, we tried to verify the hypothesis of the benefits of an anti-VEGF treatment prior to photodynamic therapy (PDT) on glioblastoma xenografts implanted subcutaneously or in the skin fold chamber. The efficacy of PDT is described as being dependent on tumor oxygenation and on the distribution of the photosensitizing agent within the tumor. In paralel to this work, we tried as a pluridisciplinary team to develop a mathematical model of the tumor response to bevacizumab using biological data obtained on the same in vivo model et that will allow in the future to simulate the response for different doses and different treatment durations, for the optimization of therapeutic protocols; L’objectif principal de ce travail de thèse a été de caractériser les effets de l’anti-VEGF Bevacizumab (Avastin) sur le réseau vasculaire tumoral in vivo, au cours du temps, à l’aide du modèle de la chambre dorsale chez la souris nude. Les images du réseau vasculaire tumoral acquises par microscopie intravitale ont été analysées par un algorithme de traitement d’images développé au sein de notre équipe, permettant de mettre en évidence les modifications morphologiques induites par le traitement et d’isoler des paramètres discriminants de la « normalisation » vasculaire, par comparaison à un réseau vasculaire sain. La période de « normalisation » vasculaire détectée par notre outil a été confortée par l’analyse de la fonctionnalité des vaisseaux sanguins au cours du temps, in vivo et par une analyse immunohistochimique des vaisseaux sanguins tumoraux et du tissu tumoral. A travers des essais préliminaires in vivo, en regard des résultats de ce travail concernant une fenêtre de "normalisation", nous avons cherché à vérifier l'hypothèse d'un bénéfice d'un traitement anti-VEGF préalablement à la thérapie photodynamique (PDT) sur des tumeurs de glioblastome xénogreffées en sous-cutané et en chambre dorsale. L'efficacité de la PDT est décrite comme étant dépendante d'une d'oxygénation tumorale suffisante et d'une distribution maximale de l'agent photosensibilisant au coeur des tumeurs. Parallèlement à ces travaux, nous avons cherché en équipe pluridiscilinaire à développer un modèle mathématique de la réponse au bevacizumab à partir de données biologiques réelles obtenues sur le même modèle in vivo et permettant pour l'avenir de simuler les réponses à différentes doses et différentes durées de traitement, toujours à des fins d'optimisation des protocoles thérapeutiques

Le conseil à l’officine fait partie intégrante de la profession de pharmacien. En effet, nombre de patients viennent trouver à l’officine des réponses à leurs maux. Le pharmacien est donc un acteur de santé de première ligne. En effet, de par sa position de proximité, le pharmacien d’officine est souvent le premier professionnel de santé sollicité par les patients afin d’obtenir des conseils concernant la prise en charge médicamenteuse ou non, les soins locaux ou encore les conseils hygiéno-diététiques à mettre en oeuvre pour palier leurs maux. Ce travail est voulu utile pour fournir à l’équipe officinale mais aussi à l ‘étudiant en pharmacie un outil pratique, recueil de 39 fiches conseils sur demande spontanée du patient concernant des pathologies diverses très fréquemment rencontrées à l’officine. Il sera abordé dans un premier temps une rétrospective sur les études en pharmacie pour aboutir sur l’ensemble des moyens mis à la disposition de l’équipe officinale pour se former et ainsi maintenir un conseil de qualité au comptoir. Les fiches conseils élaborées seront ensuite présentées ainsi que la méthodologie retenue pour leur conception. A travers ce travail, nous pouvons voir que le pharmacien a un rôle prédominant dans la prise en charge des maux du quotidien des patients. Il doit donc pouvoir être en capacité de renvoyer vers le médecin le cas échéant, trouver le traitement optimal adapté à chaque patient, le but étant l’amélioration de la qualité de vie du patient et sa satisfaction.

L’identité nutritionnelle, caractérisée par le goût, la culture, les gênes, l’éducation etc.., représente la ‘carte d’identité alimentaire’ de chaque être humain. Ces dernières années, le sujet de l’alimentation et le respect de l’identité alimentaire ont été remis en question par l’évolution des produits alimentaires proposés et par la hausse de maladies chroniques comme l’hypertension artérielle et le diabète qui sont directement liés à l’alimentation. C’est pour cela que dans cette thèse nous parlerons des différentes facettes de l’alimentation, comme les différents événements qui ont marqué son évolution dans le temps et dans la vie de l’homme, de l’anatomie comparée de l’appareil digestif de l’être humain avec des autres mammifères qui pourrait lui définir un régime particulier, des différents régimes et des différents facteurs extérieurs qui ont pu influencer notre identité nutritionnelle. Parmi toutes ces facettes, se trouve aussi le professionnel de santé qui a un rôle à jouer, en particulier le pharmacien d’officine par sa proximité avec les patients. Nous verrons par la suite, comment le pharmacien peut apporter un conseil alimentaire adapté tout en respectant l’identité nutritionnelle de chaque patient par l’intermédiaire de questions.

La leucémie lymphoïde chronique B (LLC-B) est l’hémopathie maligne la plus fréquente chez l’adulte. Malgré cela, elle reste encore peu connue à l’officine. La sortie de la réserve hospitalière de certains médicaments, l’émergence croissante des thérapies orales ont fait évoluer la prise en charge thérapeutique de la maladie ainsi que la place du pharmacien dans le parcours de soins du patient. En effet, la LLC-B est une pathologie complexe, dont la prise en charge nécessite l’intervention et la coordination de plusieurs professionnels de santé. Le pharmaciend’officine, acteur de santé publique de proximité se place comme un des maillons majeurs du parcours de soins, un intermédiaire entre l’équipe soignante et le patient. Après avoir rappelé la pathogenèse de la maladie, les moyens thérapeutiques conventionnels et innovants, seront développés le rôle du pharmacien dans cette pathologie, les outils nécessaires lui permettant la compréhension de celle-ci, des traitements, de leurs effets indésirables et des soins de support associés afin de pouvoir accompagner au mieux le patient.

The main aim of this work was to characterize the effects of the anti-VEGF Bevacizumab (Avastin) on the tumor vascular network, in vivo, over time, thanks to the skin fold chamber model on the nude mouse. Images of the vascular network obtained using intravital microscopy were analyzed par a dedicated image processing algorithm developed within our research team, allowing to highlight the morphological modifications induced by the treatment and to isolate discriminating parameters of the vascular "normalization", by comparison to healthy vascular networks. Le vascular "normalization" period detected with our tool was comforted by the analysis of the functionality of the blood vessels over time, in vivo and by an immunohistochemical analysis of the blood vessels and of the tumor tissue. In preliminary in vivo experiments, we tried to verify the hypothesis of the benefits of an anti-VEGF treatment prior to photodynamic therapy (PDT) on glioblastoma xenografts implanted subcutaneously or in the skin fold chamber. The efficacy of PDT is described as being dependent on tumor oxygenation and on the distribution of the photosensitizing agent within the tumor. In paralel to this work, we tried as a pluridisciplinary team to develop a mathematical model of the tumor response to bevacizumab using biological data obtained on the same in vivo model et that will allow in the future to simulate the response for different doses and different treatment durations, for the optimization of therapeutic protocols, L’objectif principal de ce travail de thèse a été de caractériser les effets de l’anti-VEGF Bevacizumab (Avastin) sur le réseau vasculaire tumoral in vivo, au cours du temps, à l’aide du modèle de la chambre dorsale chez la souris nude. Les images du réseau vasculaire tumoral acquises par microscopie intravitale ont été analysées par un algorithme de traitement d’images développé au sein de notre équipe, permettant de mettre en évidence les modifications morphologiques induites par le traitement et d’isoler des paramètres discriminants de la « normalisation » vasculaire, par comparaison à un réseau vasculaire sain. La période de « normalisation » vasculaire détectée par notre outil a été confortée par l’analyse de la fonctionnalité des vaisseaux sanguins au cours du temps, in vivo et par une analyse immunohistochimique des vaisseaux sanguins tumoraux et du tissu tumoral. A travers des essais préliminaires in vivo, en regard des résultats de ce travail concernant une fenêtre de "normalisation", nous avons cherché à vérifier l'hypothèse d'un bénéfice d'un traitement anti-VEGF préalablement à la thérapie photodynamique (PDT) sur des tumeurs de glioblastome xénogreffées en sous-cutané et en chambre dorsale. L'efficacité de la PDT est décrite comme étant dépendante d'une d'oxygénation tumorale suffisante et d'une distribution maximale de l'agent photosensibilisant au coeur des tumeurs. Parallèlement à ces travaux, nous avons cherché en équipe pluridiscilinaire à développer un modèle mathématique de la réponse au bevacizumab à partir de données biologiques réelles obtenues sur le même modèle in vivo et permettant pour l'avenir de simuler les réponses à différentes doses et différentes durées de traitement, toujours à des fins d'optimisation des protocoles thérapeutiques