Showing papers in "BioMed Research International in 2007"
TL;DR: The principle mechanisms of nano- and microscale cell-surface interactions in vitro for different cell types onto typical orthopaedic biomaterials such as titanium, cobalt-chrome-molybdenum (CoCrMo) alloys, stainless steel (SS), as well as synthetic polymers are elucidated.
Abstract: Cell-surface interactions play a crucial role for biomaterial application in orthopaedics. It is evident that not only the chemical composition of solid substances influence cellular adherence, migration, proliferation and differentiation but also the surface topography of a biomaterial. The progressive application of nanostructured surfaces in medicine has gained increasing interest to improve the cytocompatibility and osteointegration of orthopaedic implants. Therefore, the understanding of cell-surface interactions is of major interest for these substances. In this review, we elucidate the principle mechanisms of nano- and microscale cell-surface interactions in vitro for different cell types onto typical orthopaedic biomaterials such as titanium (Ti), cobalt-chrome-molybdenum (CoCrMo) alloys, stainless steel (SS), as well as synthetic polymers (UHMWPE, XLPE, PEEK, PLLA). In addition, effects of nano- and microscaled particles and their significance in orthopaedics were reviewed. The significance for the cytocompatibility of nanobiomaterials is discussed critically.
162 citations
TL;DR: In this paper, the effective diameters of gold-DNA conjugates were investigated by a comprehensive gel electrophoresis study based on 1200 gels, where a controlled number of single-stranded DNA of different length was attached specifically via thiol-Au bonds to phosphine-stabilized colloidal gold nanoparticles or by nonspecific adsorption.
Abstract: Gold-DNA conjugates were investigated in detail by a comprehensive gel electrophoresis study based on 1200 gels. A controlled number of single-stranded DNA of different length was attached specifically via thiol-Au bonds to phosphine-stabilized colloidal gold nanoparticles. Alternatively, the surface of the gold particles was saturated with single stranded DNA of different length either specifically via thiol-Au bonds or by nonspecific adsorption. From the experimentally determined electrophoretic mobilities, estimates for the effective diameters of the gold-DNA conjugates were derived by applying two different data treatment approaches. The first method is based on making a calibration curve for the relation between effective diameters and mobilities with gold nanoparticles of known diameter. The second method is based on Ferguson analysis which uses gold nanoparticles of known diameter as reference database. Our study shows that effective diameters derived from gel electrophoresis measurements are affected with a high error bar as the determined values strongly depend on the method of evaluation, though relative changes in size upon binding of molecules can be detected with high precision. Furthermore, in this study, the specific attachment of DNA via gold-thiol bonds to Au nanoparticles is compared to nonspecific adsorption of DNA. Also, the maximum number of DNA molecules that can be bound per particle was determined.
117 citations
TL;DR: The authors investigated the use of microencapsulated bile salt hydrolase (BSH) overproducing Lactobacillus plantarum 80 cells for oral delivery applications using a dynamic computer-controlled model simulating the human gastrointestinal (GI) tract.
Abstract: This study investigated the use of microencapsulated bile salt hydrolase (BSH) overproducing Lactobacillus plantarum 80 cells for oral delivery applications using a dynamic computer-controlled model simulating the human gastrointestinal (GI) tract Bile salt deconjugation rates for microencapsulated BSH overproducing cells were 487 ± 028 μmol/g microcapsule/h towards glycoconjugates and 079 ± 015 μmol/g microcapsule/h towards tauroconjugates in the simulated intestine, a significant (P< 05) increase over microencapsulated wild-type cells Microcapsules protected the encased cells in the simulated stomach prior to intestinal release, maintaining cell viability above 109 cfu/mL at pH 25 and 30 and above 106 cfu/mL at pH 20 after 2-hour residence times In the simulated intestine, encased cell viability was maintained above 1010 cfu/mL after 3, 6, and 12-hour residence times in bile concentrations up to 10% Results show that microencapsulation has potential in the oral delivery of live BSH active bacterial cells However, in vivo testing is required
102 citations
TL;DR: The aim of this article is to discuss the factors that define a high-risk patient and to present some of the issues pertinent to their management.
Abstract: Nonmelanoma skin cancers (squamous cell and basal cell carcinomas) occur at an epidemic rate in many countries with the worldwide incidence increasing. The sun-exposed head and neck are the most frequent sites for these cancers to arise and in most patients diagnosed with a cutaneous squamous cell carcinoma, local treatment is usually curative. However, a subset is diagnosed with a high-risk cutaneous squamous cell carcinoma. High-risk factors include size (>2 cm), thickness/depth of invasion (>4 mm), recurrent lesions, the presence of perineural invasion, location near the parotid gland, and immunosuppression. These patients have a higher risk (>10–20%) of developing metastases to regional lymph nodes (often parotid nodes), and in some cases also of experiencing local morbidity (perineural invasion), based on unfavourable primary lesion and patient factors. Despite treatment, many patients developing metastatic cutaneous squamous cell carcinoma experience mortality and morbidity usually as a consequence of uncontrolled metastatic nodal disease. It is therefore important that clinicians treating nonmelanoma skin cancers have an understanding and awareness of these high-risk patients. The aim of this article is to discuss the factors that define a high-risk patient and to present some of the issues pertinent to their management.
101 citations
TL;DR: The discrete dynamical system determined by the stroboscopic map is obtained and it is proved that the infection-free periodic solution of the impulsive epidemic system is globally attractive if the vaccination rate is larger enough and that the disease is uniformly persistent if the vaccinations rate is less than some critical value.
Abstract: Pulse vaccination, the repeated application of vaccine over a defined age range, is gaining prominence as an effective strategy for the elimination of infectious diseases. An SIR epidemic model with pulse vaccination and distributed time delay is proposed in this paper. Using the discrete dynamical system determined by the stroboscopic map, we obtain the exact infection-free periodic solution of the impulsive epidemic system and prove that the infection-free periodic solution is globally attractive if the vaccination rate is larger enough. Moreover, we show that the disease is uniformly persistent if the vaccination rate is less than some critical value. The permanence of the model is investigated analytically. Our results indicate that a large pulse vaccination rate is sufficient for the eradication of the disease.
95 citations
TL;DR: The promising clinical potential of quantum dots, their hindrances for clinical use and the current progress in abrogating their inherent toxicity are examined.
Abstract: Advances in nanotechnology have led to the development of novel fluorescent probes called quantum dots. Quantum dots have revolutionalized the processes of tagging molecules within research settings and are improving sentinel lymph node mapping and identification in vivo studies. As the unique physical and chemical properties of these fluorescent probes are being unraveled, new potential methods of early cancer detection, rapid spread and therapeutic management, that is, photodynamic therapy are being explored. Encouraging results of optical and real time identification of sentinel lymph nodes and lymph flow using quantum dots in vivo models are emerging. Quantum dots have also superseded many of the limitations of organic fluorophores and are a promising alternative as a research tool. In this review, we examine the promising clinical potential of quantum dots, their hindrances for clinical use and the current progress in abrogating their inherent toxicity.
87 citations
TL;DR: The concept of using DNA vaccines for both protection and therapy against these protozoan parasites in a variety of mouse models is validated and genome-mining and new antigen discovery strategies may provide new tools for a more rational search of novel vaccine candidates.
Abstract: Over the past 15 years, DNA vaccines have gone from a scientific curiosity to one of the most dynamic research field and may offer new alternatives for the control of parasitic diseases such as leishmaniasis and Chagas disease. We review here some of the advances and challenges for the development of DNA vaccines against these diseases. Many studies have validated the concept of using DNA vaccines for both protection and therapy against these protozoan parasites in a variety of mouse models. The challenge now is to translate what has been achieved in these models into veterinary or human vaccines of comparable efficacy. Also, genome-mining and new antigen discovery strategies may provide new tools for a more rational search of novel vaccine candidates.
67 citations
Journal Article•
TL;DR: Results show that microencapsulation has potential in the oral delivery of live BSH active bacterial cells, however, in vivo testing is required.
Abstract: This study investigated the use of microencapsulated bile salt hydrolase (BSH) overproducing Lactobacillus plantarum 80 cells for oral delivery applications using a dynamic computer-controlled model simulating the human gastrointestinal (GI) tract. Bile salt deconjugation rates for microencapsulated BSH overproducing cells were 4.87 ± 0.28 μmol/g microcapsule/h towards glycoconjugates and 0.79 ± 0.15 μmol/g microcapsule/h towards tauroconjugates in the simulated intestine, a significant (P <.05) increase over microencapsulated wild-type cells. Microcapsules protected the encased cells in the simulated stomach prior to intestinal release, maintaining cell viability above 10 9 cfu/mL at pH 2.5 and 3.0 and above 10 6 cfu/mL at pH 2.0 after 2-hour residence times. In the simulated intestine, encased cell viability was maintained above 10 10 cfu/mL after 3, 6, and 12-hour residence times in bile concentrations up to 1.0%. Results show that microencapsulation has potential in the oral delivery of live BSH active bacterial cells. However, in vivo testing is required.
65 citations
TL;DR: With this method, Mycobacterium tuberculosis in bacterial mixture as well as in spiked sputum was detected and the use of the fluorescent nanoparticles reveals amplified signal intensity and higher photostability than the direct use of conventional fluorescent dye as label.
Abstract: A method of fluorescent nanoparticle-based indirect immunofluorescence microscopy (FNP-IIFM) was developed for the rapid detection of Mycobacterium tuberculosis. An anti-Mycobacterium tuberculosis antibody was used as primary antibody to recognize Mycobacterium tuberculosis, and then an antibody binding protein (Protein A) labeled with Tris(2,2-bipyridyl)dichlororuthenium(II) hexahydrate (RuBpy)-doped silica nanoparticles was used to generate fluorescent signal for microscopic examination. Prior to the detection, Protein A was immobilized on RuBpy-doped silica nanoparticles with a coverage of ∼5.1×102 molecules/nanoparticle. With this method, Mycobacterium tuberculosis in bacterial mixture as well as in spiked sputum was detected. The use of the fluorescent nanoparticles reveals amplified signal intensity and higher photostability than the direct use of conventional fluorescent dye as label. Our preliminary studies have demonstrated the potential application of the FNP-IIFM method for rapid detection of Mycobacterium tuberculosis in clinical samples.
61 citations
TL;DR: The role of these two groups of antigens in the immune response observed during the infection is discussed and several laboratorial observations on Leishmania Sir2 and LicTXNPx as excreted/secreted proteins and LmS3arp and LimTX NPx as nonsecreted/panantigens are included.
Abstract: Leishmania infection consists in two sequential events, the host cell colonization followed by the proliferation/dissemination of the parasite. In this review, we discuss the importance of two distinct sets of molecules, the secreted and/or surface and the nonsecreted antigens. The importance of the immune response against secreted and surface antigens is noted in the establishment of the infection and we dissect the contribution of the nonsecreted antigens in the immunopathology associated with leishmaniasis, showing the importance of these panantigens during the course of the infection. As a further example of proteins belonging to these two different groups, we include several laboratorial observations on Leishmania Sir2 and LicTXNPx as excreted/secreted proteins and LmS3arp and LimTXNPx as nonsecreted/panantigens. The role of these two groups of antigens in the immune response observed during the infection is discussed.
55 citations
TL;DR: Cell-cell adhesion of HPC to human mesenchymal stromal cells was impaired and surface antigen detection of CXCR4 was reduced upon treatment with SDF-1α or AMD3100 and it was enhanced by CTCE-9908.
Abstract: Stromal cell-derived factor-1alpha (SDF-1α) has pleiotropic effects on hematopoietic progenitor cells (HPCs). We have monitored podia formation, migration, proliferation, and cell-cell adhesion of human HPC under the influence of SDF-1α, a peptide agonist of CXCR4 (CTCE-0214), a peptide antagonist (CTCE-9908), and a nonpeptide antagonist (AMD3100). Whereas SDF-1α induced migration of CD34+ cells in a dose-dependent manner, CTCE-0214, CTCE-9908, and AMD3100 did not induce chemotaxis in this concentration range albeit the peptides CTCE-0214 and CTCE-9908 increased podia formation. Cell-cell adhesion of HPC to human mesenchymal stromal cells was impaired by the addition of SDF-1α, CTCE-0214, and AMD3100. Proliferation was not affected by SDF-1α or its analogs. Surface antigen detection of CXCR4 was reduced upon treatment with SDF-1α or AMD3100 and it was enhanced by CTCE-9908. Despite the fact that all these molecules target the same CXCR4 receptor, CXCR4 agonists and antagonists have selective effects on different functions of the natural molecule.
TL;DR: In this paper, superparamagnetic iron oxide nanoparticles (SPIONs) were developed for liver specifically and to be dispersed in physiological environment stably, demonstrating the capacities to be delivered in liver specifically.
Abstract: Our goal is to develop the functionalized superparamagnetic iron oxide nanoparticles (SPIONs) demonstrating the capacities to be delivered in liver specifically and to be dispersed in physiological environment stably. For this purpose, SPIONs were coated with polyvinylbenzyl-O-β-D-galactopyranosyl-D-gluconamide (PVLA) having galactose moieties to be recognized by asialoglycoprotein receptors (ASGP-R) on hepatocytes. For use as a control, we also prepared SPIONs coordinated with 2-pyrrolidone. The sizes, size distribution, structure, and coating of the nanoparticles were characterized by transmission electron microscopy (TEM), electrophoretic light scattering spectrophotometer (ELS), X-ray diffractometer (XRD), and Fourier transform infrared (FT-IR), respectively. Intracellular uptake of the PVLA-coated SPIONs was visualized by confocal laser scanning microscopy, and their hepatocyte-specific delivery was also investigated through magnetic resonance (MR) images of rat liver. MRI experimental results indicated that the PVLA-coated SPIONs possess the more specific accumulation property in liver compared with control, which suggests their potential utility as liver-targeting MRI contrast agent.
TL;DR: A detection limit of 18 attomol QDs within the immunoassay is accomplished, an improvement of more than two orders of magnitude compared to commercial systems, as well as detailed decay time analysis of the FRET process.
Abstract: CdSe/ZnS core/shell quantum dots (QDs) are used as efficient Forster Resonance Energy Transfer (FRET) acceptors in a time-resolved immunoassays with Tb complexes as donors providing a long-lived luminescence decay. A detailed decay time analysis of the FRET process is presented. QD FRET sensitization is evidenced by a more than 1000-fold increase of the QD luminescence decay time reaching ca. 0.5 milliseconds, the same value to which the Tb donor decay time is quenched due to FRET to the QD acceptors. The FRET system has an extremely large Forster radius of approx. 100 A and more than 70% FRET efficiency with a mean donor-acceptor distance of ca. 84 A, confirming the applied biotin-streptavidin binding system. Time-resolved measurement allows for suppression of short-lived emission due to background fluorescence and directly excited QDs. By this means a detection limit of 18 attomol QDs within the immunoassay is accomplished, an improvement of more than two orders of magnitude compared to commercial systems.
TL;DR: A model is proposed in which TGF-β1+EGF-dependent MMP-10 elevation directs focalized matrix remodeling events that promote epithelial cell plasticity and tissue invasion, further facilitating epithelial invasive potential.
Abstract: The emergence of highly aggressive subtypes of human cutaneous squamous cell carcinoma (SCC) often reflects increased autocrine/paracrine TGF-β synthesis and epidermal growth factor receptor (EGFR) amplification. Cooperative TGF-β/EGFR signaling promotes cell migration and induces expression of both proteases and protease inhibitors that regulate stromal remodeling resulting in acquisition of an invasive phenotype. TGF-β1+EGF stimulation increases the production of several matrix metalloproteinases (MMPs) in human SCC. Among the most prominent is MMP-10 which is known to be elevated in SCC in situ. Activation of stromal plasminogen appears to be critical in triggering downstream MMP activity. Paradoxically, PAI-1, the major physiological inhibitor of plasmin generation, is also up-regulated under these conditions and is an early event in progression of incipient epidermal SCC. A model is proposed in which TGF-β1+EGF-dependent MMP-10 elevation directs focalized matrix remodeling events that promote epithelial cell plasticity and tissue invasion. Increased PAI-1 expression serves to temporally and spatially modulate plasmin-initiated pericellular proteolysis, further facilitating epithelial invasive potential. Defining the complex signaling mechanisms that maintain this elegant balance is critical to developing potential therapeutics for the treatment of human cutaneous malignancies.
TL;DR: This work reports on FRET between streptavidin- (STV-) conjugated CdTe quantum dots, Qdot705-STV, with biotinylated DY731-Bio fluorophores in a donor-acceptor assay and compares the influence of the two QD relaxation routes upon FRET dynamics in the presence of DY 731- Bio.
Abstract: Colloidal quantum dots (QDs) are now commercially available in a biofunctionalized form, and Forster resonance energy transfer (FRET) between bioconjugated dots and fluorophores within the visible range has been observed. We are particularly interested in the far-red region, as from a biological perspective there are benefits in pushing to ∼700 nm to minimize optical absorption (ABS) within tissue and to avoid cell autofluorescence. We report on FRET between streptavidin- (STV-) conjugated CdTe quantum dots, Qdot705-STV, with biotinylated DY731-Bio fluorophores in a donor-acceptor assay. We also highlight the changes in DY731-Bio absorptivity during the streptavidin-biotin binding process which can be attributed to the structural reorientation. For fluorescence beyond 700 nm, different alloy compositions are required for the QD core and these changes directly affect the fluorescence decay dynamics producing a marked biexponential decay with a long-lifetime component in excess of 100 nanoseconds. We compare the influence of the two QD relaxation routes upon FRET dynamics in the presence of DY731-Bio.
TL;DR: The function in both activating prothrombin and catalyzing fibrinogenolysis suggests that Ef P-III-1 may play a role in the balance between procoagulation and anticoagulation.
Abstract: The fibrinolytic function of earthworm protease-III-1 (EfP-III-1) has been studied in recent years Here, we found that EfP-III-1 acted not only in fibrinogenolysis, but also in fibrogenesis We have used EfP-III-1 to hydrolyze fibrinogen, and to activate plasminogen and prothrombin Based on the N-terminal sequences of the hydrolytic fragments, EfP-III-1 was showed to specifically recognize the carboxylic sites of arginine and lysine Analyses by fibrinogenolysis mapping and amino acid sequencing revealed that the isozyme could cleave the alpha, beta, and gamma chains of fibrinogen, showing a high α-fibrinogenase, moderate β-fibrinogenase, and low γ-fibrinogenase activities Interestingly, EfP-III-1 activated plasminogen and released active plasmin, suggesting a tPA-like function Furthermore, EfP-III-1 showed a factor Xa-like function on prothrombin, producing alpha-thrombin The function in both activating prothrombin and catalyzing fibrinogenolysis suggests that EfP-III-1 may play a role in the balance between procoagulation and anticoagulation
TL;DR: UV-induced EGFR activation increases keratinocyte proliferation, suppresses apoptosis, and augments and accelerates epidermal hyperplasia in response to UV, suggesting that the receptor is an appropriate target for the development of molecularly targeted therapies for skin cancer and other pathologies.
Abstract: The epidermal growth factor receptor (EGFR) regulates the proliferation of keratinocytes through multiple mechanisms that differ depending on the localization of the cell within the skin. Ultraviolet (UV) irradiation, the main etiologic factor in the development of skin cancer, also activates the receptor. In this review, we discuss how the UV-induced activation of EGFR regulates the response of the skin to UV. UV-induced EGFR activation increases keratinocyte proliferation, suppresses apoptosis, and augments and accelerates epidermal hyperplasia in response to UV. Pharmacological inhibition of the UV-induced activation of EGFR in a genetically initiated mouse skin tumorigenesis model suppresses tumorigenesis and the activation of mitogen-activated protein (MAP) kinases and phosphatidyl inositol-3-kinase (PI3K)/AKT signaling pathways. EGFR has pleiotropic, complex, and cell-type-specific functions in cutaneous keratinocytes; suggesting that the receptor is an appropriate target for the development of molecularly targeted therapies for skin cancer and other pathologies.
TL;DR: The data suggest that HHcy is strongly and independently associated to CAD risk increase; and MTHFR C677T polymorphism and smoking habits were the main predictors of tHcy levels.
Abstract: Increased plasma total homocysteine (tHcy) levels have been shown to be a risk factor for coronary artery disease (CAD). The common methylenetetrahydrofolate reductase C677T (MTHFR C677T) polymorphism has been reported to be a strong predictor of mild hyperhomocysteinaemia (HHcy). We assessed whether this mutation was associated with increased risk of CAD and plasma levels of tHcy. We also evaluated interactions between this polymorphism, mild elevated tHcy levels and conventional risk factors of CAD. Method. Using PCR-RFLP analysis, we studied the frequency of the C677T genotypes and its effect on CAD and on tHcy concentrations in 400 subjects without and with CAD angiographically confirmed. There were 210 subjects with CAD and 190 subjects without CAD. Results. The frequencies of the C677T genotypes were 53% (59.5% in controls versus 48.1% in cases), 34.8% (32.1 in controls versus 37.1 in cases), and 11.8% (8.4% in controls versus 14.8% in cases), respectively, for 677CC, 677CT, and 677TT. The genotype frequencies were significantly different between case and control groups (P < .05). The 677T allele enhances the risk of CAD associated to HHcy (P < .01). In multivariate analysis models, MTHFR C677T polymorphism effect on CAD was masked by other risk factors. HHcy was only and independently influenced by MTHFR polymorphism and smoking habits, and it is a strong predictor of CAD independently of conventional risk factors. Conclusion. Our data suggest that HHcy is strongly and independently associated to CAD risk increase; and MTHFR C677T polymorphism and smoking habits were the main predictors of tHcy levels. The CAD risk increase is mainly associated with mild HHcy in 677TT, whereas in 677CT and 677CC it is mainly associated with the conventional risk factors.
TL;DR: In conclusion, in addition to p53 mutations, silencing of the CDKN2A gene might play a significant role in SCC development.
Abstract: p53 tumor suppressor gene is the most commonly mutated gene in human and mouse cancers. Disruption of the p53 and Rb pathways is a fundamental trend of most human cancer cells. Inactivation of CDKN2A can lead to deregulation of these two pathways. Genetic abnormalities in CDKN2A gene have been well documented in human melanoma but their involvement in human nonmelanoma skin cancer (NMSC) and in particular in squamous cell carcinoma (SCC) is less clear. Several studies have shown that human SCCs harbour unique mutations in the p53 gene as well as inactivation of the CDKN2A gene. While mutations in the p53 gene are induced by UV radiation and represent tumor initiating events, the majority of alterations detected in the CDKN2A gene do not appear to be UV-dependent. In conclusion, in addition to p53 mutations, silencing of the CDKN2A gene might play a significant role in SCC development.
TL;DR: The design and synthesis of a tetraethylene glycol- (TEG-) based bidentate ligand functionalized with dihydrolipoic acid and biotin to promote biocompatibility of luminescent quantum dots (QD's).
Abstract: We report the design and synthesis of a tetraethylene glycol- (TEG-) based bidentate ligand functionalized with dihydrolipoic acid (DHLA) and biotin (DHLA—TEG—biotin) to promote biocompatibility of luminescent quantum dots (QD's). This new ligand readily binds to CdSe—ZnS core-shell QDs via surface ligand exchange. QDs capped with a mixture of DHLA and DHLA—TEG—biotin or polyethylene glycol- (PEG-) (molecular weight average ∼600) modified DHLA (DHLA—PEG600) and DHLA—TEG—biotin are easily dispersed in aqueous buffer solutions. In particular, homogeneous buffer solutions of QDs capped with a mixture of DHLA—PEG600 and DHLA—TEG—biotin that are stable over broad pH range have been prepared. QDs coated with mixtures of DHLA/DHLA—TEG—biotin and with DHLA—PEG600/DHLA—TEG—biotin were tested in surface binding assays and the results indicate that biotin groups on the QD surface interact specifically with NeutrAvidin-functionalized microtiter well plates.
TL;DR: This minireview briefly analyzes some of the current data related to the regulatory cells and molecules derived from parasites that affect cellular function and contribute to the polarization of the immune response of the host.
Abstract: Parasitic infections are prevalent in both tropical and subtropical areas. Most of the affected and/or exposed populations are living in developing countries where control measures are lacking or inadequately applied. Although significant progress has been made in our understanding of the immune response to parasites, no definitive step has yet been successfully done in terms of operational vaccines against parasitic diseases. Evidence accumulated during the past few years suggests that the pathology observed during parasitic infections is in part due to deregulation of normal components of the immune system, mainly cytokines, antibodies, and immune effector cell populations. A large number of studies that illustrate how parasites can modify the host immune system for their own benefit have been reported in both metazoan and protozoan parasites. The first line of defense against foreign organisms is barrier tissue such as skin, humoral factors, for instance the complement system and pentraxin, which upon activation of the complement cascade facilitate pathogen recognition by cells of innate immunity such as macrophages and DC. However, all the major groups of parasites studied have been shown to contain and/or to release factors, which interfere with both arms of the host immune system. Even some astonishing observations relate to the production by some parasites of orthologues of mammalian cytokines. Furthermore, chronic parasitic infections have led to the immunosuppressive environment that correlates with increased levels of myeloid and T suppressor cells that may limit the success of immunotherapeutic strategies based on vaccination. This minireview briefly analyzes some of the current data related to the regulatory cells and molecules derived from parasites that affect cellular function and contribute to the polarization of the immune response of the host. Special attention is given to some of the data from our laboratory illustrating the role of immunomodulatory factors released by protozoan parasites, in the induction and perpetuation of chronic disease.
TL;DR: By modulating human DC reactivity, LAB and intestinal epithelium might modify T cell immune response and regulate the development of allergic reaction.
Abstract: Lactic acid bacteria (LAB) are Gram positive nonpathogenic commensal organisms present in human gastrointestinal tract. In vivo, LAB are separated from antigen-presenting cells such as dendritic cells (DC) by the intestinal epithelial barrier. In this study, the impact of one LAB strain (Lactobacillus casei ATCC393) on human monocyte-derived DC from allergic and healthy donors was assessed by using a polarized epithelium model. Confocal and flow cytometer analyses showed that immature DC efficiently captured FITC-labelled L. casei through the epithelial layer. After interaction with L. casei, DC acquired a partial maturation status (i.e., CD86 and CD54 increase) and increased their interleukin (IL)-10 and IL-12 production. Interestingly, after activation by L. casei in the presence of experimental epithelium, DC from allergic patients instructed autologous naive CD4(+) T cells to produce more interferon-gamma than without the epithelium. Thus by modulating human DC reactivity, LAB and intestinal epithelium might modify T cell immune response and regulate the development of allergic reaction.
TL;DR: The RDB may be a suitable alternative method to study mechanisms of tumor progression and invasiveness in vitro, under more complex physicochemical conditions that may approximate natural tissue environments.
Abstract: The present study represents an investigation of a novel stirred bioreactor for culture of a transformed cell line under defined hydrodynamic conditions in vitro. Cell colonies of the EL-4 mouse lymphoma cell line grown for the first time in a rotating disc bioreactor (RDB), were observed to undergo changes in phenotype in comparison to standard, static flask cultures. RDB cultures, with or without agitation, promoted the formation of adherent EL-4 cell plaques that merged to form contiguous tumor-like masses in longer-term cultures, unlike the unattached spheroid aggregates of flask cultures. Plaques grown under agitated conditions were further altered in morphology and distribution in direct response to fluid mechanical stimuli. Plaque colonies growth in RDBs with or without agitation also exhibited significant increases in production of interleukin-4 (IL-4) and lactate, suggesting an inducible “Warburg effect.” Increases in cell biomass in RDB cultures were no different to flask cultures, though a trend toward a marginal increase was observed at specific rotational speeds. The RDB may therefore be a suitable alternative method to study mechanisms of tumor progression and invasiveness in vitro, under more complex physicochemical conditions that may approximate natural tissue environments.
TL;DR: The challenges of intracellular uses of NCs are focused on and the potential cytotoxicity of cytoplasmically loaded NCs must be evaluated.
Abstract: Semiconductor nanocrystals (NCs) are increasingly being used as photoluminescen markers in biological imaging. Their brightness, large Stokes shift, and high photostability compared to organic fluorophores permit the exploration of biological phenomena at the single-molecule scale with superior temporal resolution and spatial precision. NCs have predominantly been used as extracellular markers for tagging and tracking membrane proteins. Successful internalization and intracellular labelling with NCs have been demonstrated for both fixed immunolabelled and live cells. However, the precise localization and subcellular compartment labelled are less clear. Generally, live cell studies are limited by the requirement of fairly invasive protocols for loading NCs and the relatively large size of NCs compared to the cellular machinery, along with the subsequent sequestration of NCs in endosomal/lysosomal compartments. For long-period observation the potential cytotoxicity of cytoplasmically loaded NCs must be evaluated. This review focuses on the challenges of intracellular uses of NCs.
TL;DR: Modifications of bipartite leukotoxins by two different processes, using fusion with glutathione S-transferase and bridging of the N-terminal extremity to the adjacent β-sheet via disulphide bridges, are not deleterious for biological activity and illustrate a microheterogeneity of the structural organizations between bipartites and α-toxin.
Abstract: The staphylococcal bipartite leukotoxins and the homoheptameric α-toxin belong to the same family of β-barrel pore-forming toxins despite slight differences. In the α-toxin pore, the N-terminal extremity of each protomer interacts as a deployed latch with two consecutive protomers in the vicinity of the pore lumen. N-terminal extremities of leukotoxins as seen in their three-dimensional structures are heterogeneous in length and take part in the β-sandwich core of soluble monomers. Hence, the interaction of these N-terminal extremities within structures of adjacent monomers is questionable. We show here that modifications of their N-termini by two different processes, using fusion with glutathione S-transferase (GST) and bridging of the N-terminal extremity to the adjacent β-sheet via disulphide bridges, are not deleterious for biological activity. Therefore, bipartite leukotoxins do not need a large extension of their N-terminal extremities to form functional pores, thus illustrating a microheterogeneity of the structural organizations between bipartite leukotoxins and α-toxin.
TL;DR: The authors focus in particular on mutations and chromosomal rearrangements flanking the insertion of the T-DNA, and suggest extended backcrossing for elimination of genome-wide mutations, and sequencing of flanking DNA of 50 kbp at each side of the insertion.
Abstract: Latham et al. [1] and Wilson et al. [2] reviewed the long known phenomenon that plant transformation may cause mutations. Mutations can occur at any position in the genome, due to the tissue culture phase or other factors. Furthermore, insertion mutations may be induced by Agrobacterium-mediated transformation or particle bombardment. The authors focus in particular on mutations and chromosomal rearrangements flanking the insertion of the T-DNA. In their view, these mutations pose a risk regarding biosafety. The transgenic plant should in their opinion be as identical to its parent as possible. Therefore, they recommend extended backcrossing for elimination of genome-wide mutations, and sequencing of flanking DNA of 50 kbp at each side of the insertion, and discarding of plants that show any mutation in the flanking DNA compared to the parent plant. These and other precautions should ensure that transformation-induced mutations will not impact on biosafety.
Apparently, the authors suggest that in addition to the inserted transgene, mutations have their own contribution to uncertainties regarding biosafety. If the transgenic plant was to be released in the environment, genotypes with mutations should be discarded.
The authors do not mention mutation breeding of plants. Since the discovery of X-ray induced mutations in barley nearly 80 years ago (Stadler [3]), plant breeders and geneticists have realized how DNA mutations can be induced for widening the genetic variation in their germplasm. During the past seventy years, mutation breeding led to more than 2250 plant varieties (Maluszynski et al. [4]; Ahloowalia et al. [5]). 70% of these varieties were released as directly induced mutants, and the other 30% from crosses with induced mutants. The use of chemical treatments was relatively infrequent, but gamma rays were frequently used (64%), followed by X-rays (22%) (Ahloowalia et al. [5]).
The freely accessible FAO/IAEA website contains a database of plant varieties derived from induced mutations (http://www-infocris.iaea.org/MVD/default.htm). This list has been composed on the basis of official information from plant breeders and authorities. The composers of the database mention that the list is far from complete, as frequently it is not published how new varieties have been obtained. In spite of that, the list contains now (August 2007) already 2543 released plant varieties. In reality, the number of induced mutant varieties is much larger. If spontaneous mutants would also be included, the list would further expand strongly.
The induced mutant varieties have been developed in 175 plant species, including rice, wheat, barley, cotton, rapeseed, sunflower, grapefruit, apple, banana, and many other species. They are released in Europe, Asia, North America, South America, and Australia. Dozens of these varieties are grown at large scales (Ahloowalia et al. [5]). Many millions of people eat and use products of these varieties.
The authors also do not mention the experiences in wheat breeding with gene transfer from wild species using induced translocations. In this approach, a resistance gene was introgressed by means of an interspecific cross followed by repeated backcrosses. This led to addition of an alien chromosome, containing the resistance gene. For adoption of the resistance gene into the wheat chromosomes, the addition lines were irradiated for mutation induction. During repair of the breakages of the wheat chromosomes, sometimes a part of the alien chromosome was incorporated, leading to normal Mendelian inheritance of the resistance gene in the wheat genome (Friebe et al. [6]; Mukai et al. [7]). Many wheat varieties have been released that contain one or more of these kinds of induced translocations.
The authors did not mention that in conventional breeding, traits from wild germplasm are introduced into cultivars by means of crosses and backcrosses with an elite cultivar. During this process, chromosomal parts from the wild germplasm are introduced. These chromosomal parts may harbour hundreds of unknown “wild” alleles and thousands of deviations in the DNA sequence compared to the original elite cultivar. These thousands of natural deviations can be regarded as thousands of mutations. We all use and eat such cultivars for many decades.
The mutant varieties and those originating from translocation events or backcrosses usually are not molecularly characterized by DNA sequencing, nor compared with their parents at the DNA sequence level. Sometimes cytogenetic or genetic marker studies are performed to locate introduced chromosomal parts in the recipient genome. However, the plant breeders of mutant varieties usually do not know the number of mutations or changes, the kinds of mutations, nor the number of rearrangements in their varieties. Neither they know whether new open reading frames or fused genes have been created by the mutations, nor whether expression levels of genes have changed due to mutation, or due to introgression of DNA from wild germplasm. However, the cultivars generally have been phenotyped thoroughly by the breeders and compared phenotypically to their parents and contemporary cultivars in view of their commercial value. In spite of the absence of molecular characterization, the cultivars, either from induced mutation, spontaneous mutation, or introgression breeding, have been widely accepted, grown, and used.
The precautionary measures proposed by Latham et al. [1] and Wilson et al. [2] for genetically engineered plants regarding detection of mutations are not in balance at all with common practice in conventional plant breeding as described above. It is unscientific to propose screening flanking DNA of 50 kbp at each side of the insertion, requiring discarding of plants that show any changes there, but simultaneously accepting plants with tens or hundreds or thousands of unknown but probably more dramatic DNA changes after irradiation or introgression.
One could react with a proposal to put also plants from induced mutations under strict safety regulations. This would make sense only if the mentioned 2543 varieties would have induced more frequently biosafety problems then varieties from cross breeding. However, we are not aware of any biosafety problem caused by an induced mutation of a released variety or by an induced translocation. Apparently, the common thorough evaluation of induced mutants at the phenotypic level by the breeders suffices.
Proposing a ban on mutations caused by gene transformation for the sake of biosafety indicates a blind spot for the safety of numerous mutations induced by conventional breeders for more than 70 years, and introgression of unknown chromosomal parts from wild germplasm since centuries.
TL;DR: This work quantifies spectral properties of fluorescence quenching and intermittency on a single-particle and ensemble basis in order to evaluate their usefulness as a tool for indicating QD uptake, breakdown, and processing in living cells.
Abstract: CdSe/ZnS quantum dots (QDs) conjugated to biomolecules that quench their fluorescence, particularly dopamine, have particular spectral properties that allow determination of the number of conjugates per particle, namely, photoenhancement and photobleaching In this work, we quantify these properties on a single-particle and ensemble basis in order to evaluate their usefulness as a tool for indicating QD uptake, breakdown, and processing in living cells This creates a general framework for the use of fluorescence quenching and intermittency to better understand nanoparticle-cell interactions
TL;DR: A systematic evaluation of two commercial CPB tubings found that discrimination between biomaterial coatings can be made only after several hours of blood-biomaterial contact, which is believed to be relevant with respect to the bioengineering of extracorporeal devices that should function in contact with blood for extended time.
Abstract: Extended use of cardiopulmonary bypass (CPB) systems is often hampered by thrombus formation and infection. Part of these problems relates to imperfect hemocompatibility of the CPB circuitry. The engineering of biomaterial surfaces with genuine long-term hemocompatibility is essentially virgin territory in biomaterials science. For example, most experiments with the well-known Chandler loop model, for evaluation of blood-biomaterial interactions under flow, have been described for a maximum duration of 2 hours only. This study reports a systematic evaluation of two commercial CPB tubings, each with a hemocompatible coating, and one uncoated control. The experiments comprised (i) testing over 5 hours under flow, with human whole blood from 4 different donors; (ii) measurement of essential blood parameters of hemocompatibility; (iii) analysis of the luminal surfaces by scanning electron microscopy and thrombin generation time measurements. The dataset indicated differences in hemocompatibility of the tubings. Furthermore, it appeared that discrimination between biomaterial coatings can be made only after several hours of blood-biomaterial contact. Platelet counting, myeloperoxidase quantification, and scanning electron microscopy proved to be the most useful methods. These findings are believed to be relevant with respect to the bioengineering of extracorporeal devices that should function in contact with blood for extended time.
TL;DR: A new, simplistic method of obtaining the system for controlled delivery of the ascorbic acid using physical method with solvent/nonsolvent systems where obtained solutions were centrifuged and the spherical shape, size, and uniformity of DLPLG particles are preserved.
Abstract: This paper is covering new, simplistic method of obtaining the system for controlled delivery of the ascorbic acid. Copolymer poly (D,L-lactide-co-glycolide) (DLPLG) nanoparticles are produced using physical method with solvent/nonsolvent systems where obtained solutions were centrifuged. The encapsulation of the ascorbic acid in the polymer matrix is performed by homogenization of water and organic phases. Particles of the DLPLG with the different content of ascorbic acid have different morphological characteristics, that is, variable degree of uniformity, agglomeration, sizes, and spherical shaping. Mean sizes of nanoparticles, which contain DLPLG/ascorbic acid in the ratio 85/150%, were between 130 to 200 nm depending on which stereological parameters are considered (maximal diameters Dmax, feret X, or feret Y). By introducing up to 15% of ascorbic acid, the spherical shape, size, and uniformity of DLPLG particles are preserved. The samples were characterized by infrared spectroscopy, scanning electron microscopy, stereological analysis, and ultraviolet spectroscopy.
TL;DR: It is demonstrated that it is possible to understand hematopoietic stem cell organization without assumptions on unidirectional developmental hierarchies, preprogrammed asymmetric division events or other assumptions implying the existence of a predetermined stem cell entity.
Abstract: Many experimental findings on heterogeneity, flexibility, and plasticity of tissue stem cells are currently challenging stem cell concepts that assume a cell intrinsically predefined, unidirectional differentiation program. In contrast to these classical concepts, nonhierarchical self-organizing systems provide an elegant and comprehensive alternative to explain the experimental data. Here we present the application of such a self-organizing concept to quantitatively describe the hematopoietic stem cell system. Focusing on the analysis of individual-stem-cell fates and clonal dynamics, we particularly discuss implications of the theoretical results on the interpretation of experimental findings. We demonstrate that it is possible to understand hematopoietic stem cell organization without assumptions on unidirectional developmental hierarchies, preprogrammed asymmetric division events or other assumptions implying the existence of a predetermined stem cell entity. The proposed perspective, therefore, changes the general paradigm of thinking about stem cells.