Ruthenium(
ii
) complexes are attracting significant research attention as a promising class of photosensitizers (PSs) in photodynamic therapy (PDT). Having previously reported the synthesis of two ...novel Ru(
ii
)-polypyridyl-1,8-naphthalimide Tröger's base compounds
1
and
2
with interesting photophysical properties, where the emission from either the Ru(
ii
) polypyridyl centres or the naphthalimide moieties could be used to monitor binding to nucleic acids, we sought to use these compounds to investigate further and in more detail their biological profiling, which included unravelling their mechanism of cellular uptake, cellular trafficking and cellular responses to photoexcitation. Here we demonstrate that these compounds undergo rapid time dependent uptake in HeLa cells that involved energy dependent, caveolae and lipid raft-dependent mediated endocytosis, as demonstrated by confocal imaging, and transmission and scanning electron microscopy. Following endocytosis, both compounds were shown to localise to mostly lysosomal and Golgi apparatus compartments with some accumulation in mitochondria but no localisation was found to the nucleus. Upon photoactivation, the compounds increased ROS production and induced ROS-dependent apoptotic cell death. The photo-activated compounds subsequently induced DNA damage and altered tubulin, but not actin structures, which was likely to be an indirect effect of ROS production and induced apoptosis. Furthermore, by changing the concentration of the compounds or the laser used to illuminate the cells, the mechanism of cell death could be changed from apoptosis to necrosis. This is the first detailed biological study of Ru(
ii
)-polypyridyl Tröger's bases and clearly suggests caveolae-dependent endocytosis is responsible for cell uptake - this may also explain the lack of nuclear uptake for these compounds and similar results observed for other Ru(
ii
)-polypyridyl complexes. These conjugates are potential candidates for further development as PDT agents and may also be useful in mechanistic studies on cell uptake and trafficking.
Ru(
ii
)-Polypyridyl-1,8-naphthalimide Tröger's bases have undergone detailed biological profiling to unravel their mechanism of cellular uptake, cellular trafficking and cellular responses to photoexcitation.
The early stages of clot formation in blood vessels involve platelet adhesion–aggregation. Although these mechanisms have been extensively studied, gaps in their understanding still persist. We have ...performed detailed in vitro experiments, using the well-known Impact-R device, and developed a numerical model to better describe and understand this phenomenon. Unlike previous studies, we took into account the differential role of pre-activated and non-activated platelets, as well as the three-dimensional nature of the aggregation process. Our investigation reveals that blood albumin is a major parameter limiting platelet aggregate formation in our experiment. Simulations are in very good agreement with observations and provide quantitative estimates of the adhesion and aggregation rates that are hard to measure experimentally. They also provide a value of the effective diffusion of platelets in blood subject to the shear rate produced by the Impact-R.
The emergence of antibiotic-resistant
has become a major public health concern, necessitating the discovery of new antimicrobial compounds. Given that the skin microbiome plays a critical role in the ...host defence against pathogens, the development of therapies that target the interactions between commensal bacteria and pathogens in the skin microbiome offers a promising approach. Here, we report the discovery of two bacteriocins, cerein 7B and cerein B4080, that selectively inhibit
without affecting
, a commensal bacterium on the skin. Our study revealed that exposure of
to these bacteriocins resulted in mutations in the
two-component system, leading to a thickening of the cell wall visible by transmission electron microscopy and subsequent decreased sensitivity to vancomycin. Our findings prompt a nuanced discussion of the potential of those bacteriocins for selective targeting of
on the skin, given the emergence of resistance and co-resistance with vancomycin. The idea put forward implies that by preserving commensal bacteria, selective compounds could limit the emergence of resistance in pathogenic cells by promoting competition with remaining commensal bacteria, ultimately reducing chronical infections and limiting the spread of antibiotic resistance.
Insight into plant annexin function Baucher, Marie; Pérez-Morga, David; El Jaziri, Mondher
Plant signaling & behavior,
04/2012, Letnik:
7, Številka:
4
Journal Article
Recenzirano
Odprti dostop
The multifunctionality of plant annexins and their importance for coordinating development and responses to biotic and abiotic environment have been largely reviewed. We recently described a tobacco ...annexin, named Ntann12, which is mainly localized in the nucleus of root cells when the plant is grown under light conditions. We also found that auxin and polar auxin transport are essential for Ntann12 accumulation in root cells. Under dark condition, Ntann12 is no longer detected in the root system. In the present addendum, light, regulating auxin signaling, is evidenced as an essential determinant for the synchronization of growth and development between the shoot and the root during light/dark cycle. A speculative model for Ntann12 is described and discussed with regards to relevant literature data.
Trypanosoma cruzi is a protozoan parasite transmitted by a triatomine insect, and causing human Chagas disease in South America. This parasite undergoes a complex life cycle alternating between ...non-proliferative and dividing forms. Owing to their high energy requirement, replicative epimastigotes of the insect midgut display high endocytic activity. This activity is mainly restricted to the cytostome, by which the cargo is taken up and sorted through the endosomal vesicular network to be delivered to reservosomes, the final lysosomal-like compartments. In African trypanosomes tomato lectin (TL) and ricin, respectively specific to poly-N-acetyllactosamine (poly-LacNAc) and β-D-galactose, allowed the identification of giant chains of poly-LacNAc in N-glycoproteins of the endocytic pathway. We show that in T. cruzi epimastigote forms also, glycoproteins of the endocytic pathway are characterized by the presence of N-linked glycans binding to both ricin and TL. Affinity chromatography using both TL and Griffonia simplicifolia lectin II (GSLII), specific to non-reducing terminal residue of N-acetylglucosamine (GlcNAc), led to an enrichment of glycoproteins of the trypanosomal endocytic pathway. Incubation of live parasites with TL, which selectively bound to the cytostome/cytopharynx, specifically inhibited endocytosis of transferrin (Tf) but not dextran, a marker of fluid endocytosis. Taken together, our data suggest that N-glycan modification of endocytic components plays a crucial role in receptor-mediated endocytosis of T. cruzi.
Human innate immunity to Trypanosoma brucei involves the trypanosome C-terminal kinesin TbKIFC1, which transports internalized trypanolytic factor apolipoprotein L1 (APOL1) within the parasite. We ...show that TbKIFC1 preferentially associates with cholesterol-containing membranes and is indispensable for mammalian infectivity. Knockdown of TbKIFC1 did not affect trypanosome growth in vitro but rendered the parasites unable to infect mice unless antibody synthesis was compromised. Surface clearance of Variant Surface Glycoprotein (VSG)-antibody complexes was far slower in these cells, which were more susceptible to capture by macrophages. This phenotype was not due to defects in VSG expression or trafficking but to decreased VSG mobility in a less fluid, stiffer surface membrane. This change can be attributed to increased cholesterol level in the surface membrane in TbKIFC1 knockdown cells. Clearance of surface-bound antibodies by T. brucei is therefore essential for infectivity and depends on high membrane fluidity maintained by the cholesterol-trafficking activity of TbKIFC1.
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•The TbKIFC1 kinesin preferentially transports cholesterol-containing membranes•TbKIFC1 is absolutely required for parasite growth in mice•Cholesterol trafficking by TbKIFC1 confers high fluidity to the plasma membrane•High membrane fluidity allows antibody clearance necessary for parasite infectivity
Immunology; Microbiology Parasite; Cell Biology
The primate-specific serum protein apolipoprotein L1 (APOL1) is the only secreted member of a family of cell death promoting proteins
. APOL1 kills the bloodstream parasite Trypanosoma brucei brucei, ...but not the human sleeping sickness agents T.b. rhodesiense and T.b. gambiense
. We considered the possibility that intracellular members of the APOL1 family, against which extracellular trypanosomes could not have evolved resistance, could kill pathogenic T. brucei subspecies. Here we show that recombinant APOL3 (rAPOL3) kills all African trypanosomes, including T.b. rhodesiense, T.b. gambiense and the animal pathogens Trypanosoma evansi, Trypanosoma congolense and Trypanosoma vivax. However, rAPOL3 did not kill more distant trypanosomes such as Trypanosoma theileri or Trypanosoma cruzi. This trypanolytic potential was partially shared by rAPOL1 from Papio papio (rPpAPOL1). The differential killing ability of rAPOL3 and rAPOL1 was associated with a distinct dependence on acidic pH for activity. Due both to its instability and toxicity when injected into mice, rAPOL3 cannot be used for the treatment of infection, but an experimental rPpAPOL1 mutant inspired by APOL3 exhibited enhanced trypanolytic activity in vitro and the ability to completely inhibit T.b. gambiense infection in mice. We conclude that pH dependence influences the trypanolytic potential of rAPOLs.
The contribution of the B isoform of inositol 1,4,5-trisphosphate Ins(1,4,5)P₃ 3-kinase (or Itpkb) and inositol 1,3,4,5-tetrakisphosphate Ins(1,3,4,5)P₄, its reaction product, to B cell function and ...development remains unknown. Here, we show that mice deficient in Itpkb have defects in B cell survival leading to specific and intrinsic developmental alterations in the B cell lineage and antigen unresponsiveness in vivo. The decreased B cell survival is associated with a decreased phosphorylation of Erk1/2 and increased Bim gene expression. B cell survival, development, and antigen responsiveness are normalized in parallel to reduced expression of Bim in Itpkb⁻/⁻ Bim⁺/⁻ mice. Analysis of the signaling pathway downstream of Itpkb revealed that Ins(1,3,4,5)P₄ regulates subcellular distribution of Rasa3, a Ras GTPase-activating protein acting as an Ins(1,3,4,5)P₄ receptor. Together, our results indicate that Itpkb and Ins(1,3,4,5)P₄ mediate a survival signal in B cells via a Rasa3-Erk signaling pathway controlling proapoptotic Bim gene expression.
Highlights ► We design a novel adjuvant system based on aluminum salts. ► Cationic lipid diC14 amidine improves innate detection of alum. ► Mixed preparation outperforms sequential addition of both ...immunomodulators. ► Processing of alum-adsorbed antigen benefits from coupling with diC14 amidine. ► diC14 amidine enhances antigen-specific humoral and cellular immunity induced by alum.
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