The Bd37gene encoding for a glycosyl-phosphatidyl-inositol anchored protein of Babesia divergens displays genetic polymorphisms among isolates. Five major polymorphic groups (clades) were shown by PCR-RFLP among different B. divergens isolates. Each group has been characterized according to a reference Bd37 gene (Rouen87, W8843, Y5, 6303E and 1705B). Recombinant (GST fusion) protein (Bd37r) expressed from the Bd37 gene, was used as antigen in a saponin-based formulation and was able to protect gerbils, after 2 injections at low dose vaccine concentration (1 μg per dose), against a virulent challenge with the B. divergens Rouen87 isolate. In spite of polymorphism of Bd37 gene, Bd37r induced complete immunoprotection against challenges with each of the 5 reference isolate groups defined by PCR-RFLP.
New “all-in-one” theranostic systems, combining a magnetic resonance imaging (MRI) contrast agent with a biphotonic photodynamic therapy (2P-PDT) photosensitiser generating cytotoxic singlet oxygen, were envisaged and synthesised. They are based on azamacrocycles, regiospecifically functionalised by two-photon PDT π-conjugated dibromobenzene-picolinate photosensitisers and acetate, able to complex gadolinium(III) and allow an MRI signal. Our approach was to use two different macrocyclic platforms, tacn and pyclen, for modulating simultaneously the structures, properties and solubility of the complexes. Photophysical properties of the ligands and their gadolinium(III) complexes were fully investigated. The Gd3+-pyclen derivative showed the best water solubility and the greatest value of singlet oxygen generation of the series with φΔ = 0.53 enabling in vitro studies. The biological PDT activity under mono and biphotonic excitation was evaluated in human breast cancer cells (MCF-7). While a very low dark toxicity was observed, an almost total cell death was induced after only 3 successive irradiations of 1.57 sec. Finally, its relaxivity was measured in a DMSO/H2O solvents mixture with r1p = 11.21 and r2p = 24.60 mM-1s -1 at 3.0 T and T1- and T2-weighted phantom MR images were obtained highlighting a first generation of “all-in-one” PDT/MRI theranostic agents.
Abstract On account of its extreme intrinsic resistance to apoptosis and of its strong ability to become chemoresistant after a primary response to drugs, malignant melanoma ( MM ) is still a therapeutic challenge. We previously showed that glutathione S‐transferase mu 1 ( GSTM 1) acts in synergy with multidrug resistance protein 1 ( MRP 1) to protect GSTM 1‐transfected human CAL 1 melanoma cells from toxic effects of vincristine ( VCR ). Herein, we investigated the role of these proteins in the acquired resistance of CAL 1 cells to vinca alkaloids ( VA s). Resistant lines were established by continuous exposure (>1 year) of parental CAL 1‐wt cells to VCR , vindesine ( VDS ), or vinorelbine ( VRB ): CAL 1R‐ VCR , CAL 1R‐ VDS , CAL 1R‐ VRB , respectively. All resistant lines displayed more than 10‐fold increase in resistance to their selection VA , and specifically expressed GSTM 1. Suggesting a direct interaction between this protein and VA s, each VA specifically decreased the GSTM 1‐mediated glutathione conjugation activity in cell lysates. Curcumin ( GSTM 1 inhibitor), BSO (glutathione synthesis inhibitor), and MK 571 ( MRP 1 inhibitor) considerably reversed the acquired resistance to VCR and VDS , but not to VRB . Microarray data analysis revealed similar gene expression patterns of CAL 1R‐ VCR and CAL 1R‐ VDS , and a distinct one for CAL 1R‐ VRB . These data suggest a differential involvement of GSTM 1 and MRP 1 in acquired resistance to VA s. A coordinated expression and activity of GSTM 1 and MRP 1 is required to protect CAL 1 cells from VCR and VDS , while the simple expression of GSTM 1 is sufficient, possibly by a direct drug/protein interaction, to confer resistance against VRB .
Babesia divergens est un parasite intraerythrocytaire membre du phylum des Apicomplexes, proche de Toxoplasma (agent de la toxoplasmose) et Plasmodium (agent du paludisme). B. Divergens est capable de se developper chez le bovin (hote naturel), chez l'homme (hote accidentel) et chez la gerbille (modele de laboratoire). Le parasite est transmis a son hote mammifere lors du repas sanguin de l'arthropode vecteur, la tique Ixodes ricinus. Nous avons mis en evidence deux nouvelles proteines impliquees dans les phases precoces d'adherence / invasion des erythrocytes. La proteine Bd25 est une proteine riche en cysteines. Elle presente une forte homologie avec des domaines d'interaction cellulaire connus chez les Apicomplexes (module PAN : Plasminogen Apple Nematode), ainsi qu'avec le site catalytique de la plasmepsine 2 (impliquee dans la degradation de l'hemoglobine par Plasmodium falciparum). Bd25 est presente au sein d'un complexe lie par des interactions non covalentes. Bd37. 2 est une proteine parasitaire appartenant a une nouvelle famille de proteines ancrees dans la membrane du parasite par un groupement GPI. Les deux membres connus de cette famille, Bd37. 1 et Bd37. 2, ont ete etudies en parallele, afin de caracteriser cette famille et de comprendre son implication dans les phenomenes d'invasion de l'erythrocyte par le parasite. Ces deux antigenes sont co-exprimes a la surface du parasite, et leur capacite d'adherence a ete demontree ; cependant, seule Bd37. 1 est efficace en vaccination chez la gerbille.
1. Experiments have been performed to investigate the cardiovascular actions in the rat of SCA40, a novel potassium channel opener which is a potent relaxant of guinea-pig airway smooth muscle in vivo and in vitro. 2. SCA40 (0.01-30 microM) caused a complete and concentration-dependent relaxation of rat isolated thoracic aorta contracted with 20 mM KCl but failed to inhibit completely the spasmogenic effects of 80 mM KCl. 3. The ATP-sensitive K(+)-channel blocker, glibenclamide (3 microM), failed to antagonize the relaxant action of SCA40 on 20 mM KCl-contracted rat isolated thoracic aorta. 4. SCA40 (0.001-100 microM) had dual effects on rat isolated atria. At low concentrations, SCA40 produced a concentration-dependent decrease in the rate and force of contractions. At higher concentrations (greater than 1 microM) SCA40 induced concentration-dependent increases of atrial rate and force. 5. In vivo, in normotensive Wistar rats, SCA40 elicited a dose-dependent (1-100 micrograms kg-1) decrease in mean arterial pressure which was accompanied by a moderate dose-dependent increase in heart rate. SCA40 (100 micrograms kg-1) had a slightly greater hypotensive effect than cromakalim (100 micrograms kg-1) but the duration of the hypotension was longer with cromakalim than with SCA40. 6. The hypotensive effect of SCA40 was not reduced by propranolol, atropine, NG-nitro-L-arginine methyl ester (L-NAME) or glibenclamide. 7. It is concluded that the mechanism by with SCA40 relaxes vascular smooth muscle in vitro and in vivo involves activation of K(+)-channels distinct from glibenclamide-sensitive ATP-sensitive K(+)-channels.
On account of its strong ability to become chemoresistant after a primary response to drugs, malignant melanoma (MM) remains a therapeutic challenge. This study focuses on acquired resistance to vinca alkaloids (VAs) using VA-resistant MM cell lines (CAL1R-VCR, CAL1R-VDS, and CAL1R-VRB), established by long-term continuous exposure of parental CAL1-wt cells to vincristine (VCR), vindesine (VDS), or vinorelbine (VRB), respectively. Transcriptomic profiling using rma and rdam methods led to distinguish two cell groups: CAL1R-VCR and CAL1R-VDS, CAL1R-VRB, and CAL1-wt. mgsa of the specifically altered genes in the first group evidenced the GO terms 'lysosomal lumen' and 'vacuolar lumen' linked to underexpressed genes, and 'endoplasmic reticulum (ER) stress response' associated with overexpressed genes. A specific reduction of lysosomal enzymes, independent of acidic vacuole organelle (AVO) turnover, was observed (LTG probe) in CAL1R-VCR and CAL1R-VDS cells. It was associated with the specific lowering of cathepsin B and L, known to be involved in the lysosomal pathway of apoptosis. Confirming gene profiling, the same groups (CAL1R-VCR and CAL1R-VDS, CAL1-wt and CAL1R-VRB) could be distinguished regarding the VA-mediated changes on mean size areas and on acidic compartment volumes. These two parameters were reduced in CAL1R-VCR and CAL1R-VDS cells, suggesting a smaller AVO accumulation and thus a reduced sensitivity to lysosomal membrane permeabilization-mediated apoptosis. In addition, 'ER stress response' inhibition by tauroursodeoxycholic acid induced a higher VA sensitization of the first cell group. In conclusion, lysosomes and unfolded protein response could be key determinants of the differential resistance of MM to VAs.
The (Z) and (E)-isomers of an extended tetraphenylethylene-based chromophore with optimized two-photon-induced luminescence properties are separated and functionalized with water-solubilizing pendant polymer groups, promoting their self-assembly in physiological media in the form of small, colloidal stable organic nanoparticles. The two resulting fluorescent suspensions are then evaluated as potential two-photon luminescent contrast agents for intravital epifluorescence and two-photon fluorescence microscopy. Comparisons with previously reported works involving similar fluorophores devoid of polymer side chains illustrate the benefits of later functionalization regarding the control of the self-assembly of the nano-objects and ultimately their biocompatibility toward the imaged organism.
