Novel mechanisms for regulation of cell survival and migration by ceramide 1-phosphate

201 p. : gráf.Ceramide 1-phosphate is a bioactive sphingolipid that regulates important cell functions, including cell proliferation, apoptosis, migration or inflammation. Cell homeostasis is essential for the normal development of any organism and alteration of any of these pro¬cesses can lead to metabolic dysfunction or cause illnesses such as autoimmune diseases, chronic inflammation, neural degeneration, cardiovascular disorders, or cancer. The molecular mechanisms involved in the regulation of these cell responses by C1P have only begun to be understood. The aim of this Doctoral Thesis was to study the cellular signaling processes involved in C1P-stimulated cell proliferation and migration in immune cells, mainly macrophages. Concerning the regulation of cell proliferation, this Thesis has contributed to the elucidation of three major mechanisms by which C1P exerts its mitogenic actions: 1) Stimulation of vascular endothelial growth factor (VEGF) secretion, an action that involves activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. 2) Induction of translocation and activation of protein kinase C alpha (PKCα). 3) Activation of NADPH oxidase and the subsequent formation of reactive oxygen species (ROS). With regards to cell migration, we have demonstrated that C1P stimulates the release of macrophage chemoattractant protein-1 (MCP-1), which is essential for the induction of macrophage migration. In addition, we have shown that C1P-stimulated cell migration requires the interaction of C1P with a putative C1P receptor, and the subsequent activation of the PI3K/Akt, MAPK/Erk1-2, p38 and JNK signalling pathways. Finally, this Thesis also shows that C1P increases the expression of CD69 in spleenic mononuclear cells, and that it protects these cells from entering apoptosis. A major pathway involved in this action is the mitogen-activated protein kinase kinase (MEK)/ERK1-2/ NF-кB pathway

Incorporation of Antibiotics into Solid Lipid Nanoparticles: A Promising Approach to Reduce Antibiotic Resistance Emergence

Antimicrobial resistance is one of the biggest threats to global health as current antibiotics are becoming useless against resistant infectious pathogens. Consequently, new antimicrobial strategies are urgently required. Drug delivery systems represent a potential solution to improve current antibiotic properties and reverse resistance mechanisms. Among different drug delivery systems, solid lipid nanoparticles represent a highly interesting option as they offer many advantages for nontoxic targeted drug delivery. Several publications have demonstrated the capacity of SLNs to significantly improve antibiotic characteristics increasing treatment efficiency. In this review article, antibiotic-loaded solid lipid nanoparticle-related works are analyzed to summarize all information associated with applying these new formulations to tackle the antibiotic resistance problem. The main antimicrobial resistance mechanisms and relevant solid lipid nanoparticle characteristics are presented to later discuss the potential of these nanoparticles to improve current antibiotic treatment characteristics and overcome antimicrobial resistance mechanisms. Moreover, solid lipid nanoparticles also offer new possibilities for other antimicrobial agents that cannot be administrated as free drugs. The advantages and disadvantages of these new formulations are also discussed in this review. Finally, given the progress of the studies carried out to date, future directions are discussed.This work was supported by grants from the University of the Basque Country grant number GIU18/229 and grant number COLAB19/08) and the Industry Department of the Basque Government grant number ELKARTEK2020 KK-2020/00007)

Application of Solid Lipid Nanoparticles to Improve the Efficiency of Anticancer Drugs

Drug delivery systems have opened new avenues to improve the therapeutic effects of already-efficient molecules. Particularly, Solid Lipid Nanoparticles (SLNs) have emerged as promising nanocarriers in cancer therapy. SLNs offer remarkable advantages such as low toxicity, high bioavailability of drugs, versatility of incorporation of hydrophilic and lipophilic drugs, and feasibility of large-scale production. Their molecular structure is crucial to obtain high quality SLN preparations and it is determined by the relationship between the composition and preparation method. Additionally, SLNs allow overcoming several physiological barriers that hinder drug delivery to tumors and are also able to escape multidrug resistance mechanisms, characteristic of cancer cells. Focusing on cell delivery, SLNs can improve drug delivery to target cells by different mechanisms, such as passive mechanisms that take advantage of the tumor microenvironment, active mechanisms by surface modification of SLNs, and codelivery mechanisms. SLNs can incorporate many different drugs and have proven to be effective in different types of tumors (i.e., breast, lung, colon, liver, and brain), corroborating their potential. Finally, it has to be taken into account that there are still some challenges to face in the application of SLNs in anticancer treatments but their possibilities seem to be high.This work was supported by grant GIU (2018) GIU18/229

Ceramide and ceramide 1-phosphate in health and disease

Sphingolipids are essential components of cell membranes, and many of them regulate vital cell functions. In particular, ceramide plays crucial roles in cell signaling processes. Two major actions of ceramides are the promotion of cell cycle arrest and the induction of apoptosis. Phosphorylation of ceramide produces ceramide 1-phosphate (C1P), which has opposite effects to ceramide. C1P is mitogenic and has prosurvival properties. In addition, C1P is an important mediator of inflammatory responses, an action that takes place through stimulation of cytosolic phospholipase A2, and the subsequent release of arachidonic acid and prostaglandin formation. All of the former actions are thought to be mediated by intracellularly generated C1P. However, the recent observation that C1P stimulates macrophage chemotaxis implicates specific plasma membrane receptors that are coupled to Gi proteins. Hence, it can be concluded that C1P has dual actions in cells, as it can act as an intracellular second messenger to promote cell survival, or as an extracellular receptor agonist to stimulate cell migration

Antibiotikodun Lipido Solidozko Nanopartikulak bakterioen aurkako eraginkortasuna hobetzeko

Antibiotikoen aurkako erresistentzia duten bakterioak mundu-mailako osasun-arriskua dira gaur egun, infekzioak tratatzeko ditugun terapien eraginkortasunaren galera ekar dezaketelako. Mehatxu horri aurre egiteko modu bat eskuartean ditugun farmakoak garraio-sistema egokietan txertatzea izan daiteke, eraginkortasunaren hobekuntza eta erresistentzien agerpenaren atzeratzea ekar dezaketelako. Farmakoen garraio-sistema ezberdinen artean, Lipido Solidozko Nanopartikulek ezaugarri ezin hobeak eskaintzen dituzte, zeren eta, farmakoen disolbagarritasuna, iragazkortasuna, askapen-zinetika eta egonkortasuna bezalako ezaugarri garrantzitsuak hobetzeaz gain, toxikotasuna murrizten laguntzen baitute. Lipido Solidozko Nanopartikulak matrize lipidiko solido batez eta emultsionatzaile geruza batez osaturiko nanopartikulak dira, konposatu fisiologikoekin osatzen direnak. Dena den, farmakoen garraio-sistema efizienteen garapenak nanopartikulen eta sistema biologikoen arteko elkarrekintzen ezaguera eskatzen du, eta horretarako ezinbestekoak dira ekoizpen- eta karakterizazio-metodologia egokiak. Lan honetan, Lipido Solidozko Nanopartikulen osagaiak, haiek garatzeko metodologia eta haien ezaugarrien determinazioa burutzeko modua deskribatzen dira. Gainera, orain arte argitaratutako lanei buruzko berrikuspen bat egingo da antibiotikoak Solido Lipidozko Nanopartikuletan txertatzeak bakterioen aurkako terapien eraginkortasuna nola hobetu dezakeen azaltzeko.; Antibiotic resistant bacteria are a global threat because they can contribute to the loss of the efficiency of actual therapies against infectious diseases. Maintenance of the efficiency of antibacterial drugs is crucial to overcome this problem and loading antibiotics into drug delivery systems can be a possible solution because they can improve drug efficiency and delay resistance emergence. Among all drug delivery systems, Solid Lipid Nanoparticles present optimal characteristics because apart from improving drug solubility, permeability, drug-release kinetics and stability they also help reducing drug toxicity. Solid Lipid Nanoparticles are composed of a solid lipid matrix surrounded by a surfactant monolayer that can be obtained using physiologic molecules. In order to obtain efficient drug delivery systems, it is important to understand interactions between nanoparticles and biological systems. Therefore, correct Solid Lipid Nanoparticles synthesis and characterization methods are essential. In this work, materials, production and characterization methods for Solid Lipid Nanoparticles development have been described. Besides, the state of the art of antibiotic-loaded Solid Lipid Nanoparticles is described in order to highlight the advantages of applying Solid Lipid Nanoparticles to improve antibacterial efficacy of actual antibiotic therapies

Development of a high-throughput platform to measure plasmid transfer frequency

Antibiotic resistance represents one of the greatest threats to global health. The spread of antibiotic resistance genes among bacteria occurs mostly through horizontal gene transfer via conjugation mediated by plasmids. This process implies a direct contact between a donor and a recipient bacterium which acquires the antibiotic resistance genes encoded by the plasmid and, concomitantly, the capacity to transfer the acquired plasmid to a new recipient. Classical assays for the measurement of plasmid transfer frequency (i.e., conjugation frequency) are often characterized by a high variability and, hence, they require many biological and technical replicates to reduce such variability and the accompanying uncertainty. In addition, classical conjugation assays are commonly tedious and time-consuming because they typically involve counting colonies on a large number of plates for the quantification of donors, recipients, and transconjugants (i.e., the bacteria that have received the genetic material by conjugation). Due to the magnitude of the antibiotic resistance problem, it is critical to develop reliable and rapid methods for the quantification of plasmid transfer frequency that allow the simultaneous analysis of many samples. Here, we present the development of a high-throughput, reliable, quick, easy, and cost-effective method to simultaneously accomplish and measure multiple conjugation events in 96-well plates, in which the quantification of donors, recipients, and transconjugants is estimated from the time required to reach a specific threshold value (OD600 value) in the bacterial growth curves. Our method successfully discriminates different plasmid transfer frequencies, yielding results that are equivalent to those obtained by a classical conjugation assay

Incorporation of Antibiotics into Solid Lipid Nanoparticles: A Promising Approach to Reduce Antibiotic Resistance Emergence

Antimicrobial resistance is one of the biggest threats to global health as current antibiotics are becoming useless against resistant infectious pathogens. Consequently, new antimicrobial strategies are urgently required. Drug delivery systems represent a potential solution to improve current antibiotic properties and reverse resistance mechanisms. Among different drug delivery systems, solid lipid nanoparticles represent a highly interesting option as they offer many advantages for nontoxic targeted drug delivery. Several publications have demonstrated the capacity of SLNs to significantly improve antibiotic characteristics increasing treatment efficiency. In this review article, antibiotic-loaded solid lipid nanoparticle-related works are analyzed to summarize all information associated with applying these new formulations to tackle the antibiotic resistance problem. The main antimicrobial resistance mechanisms and relevant solid lipid nanoparticle characteristics are presented to later discuss the potential of these nanoparticles to improve current antibiotic treatment characteristics and overcome antimicrobial resistance mechanisms. Moreover, solid lipid nanoparticles also offer new possibilities for other antimicrobial agents that cannot be administrated as free drugs. The advantages and disadvantages of these new formulations are also discussed in this review. Finally, given the progress of the studies carried out to date, future directions are discussed

Novel mechanisms for regulation of cell survival and migration by ceramide 1-phosphate

201 p. : gráf.Ceramide 1-phosphate is a bioactive sphingolipid that regulates important cell functions, including cell proliferation, apoptosis, migration or inflammation. Cell homeostasis is essential for the normal development of any organism and alteration of any of these pro¬cesses can lead to metabolic dysfunction or cause illnesses such as autoimmune diseases, chronic inflammation, neural degeneration, cardiovascular disorders, or cancer. The molecular mechanisms involved in the regulation of these cell responses by C1P have only begun to be understood. The aim of this Doctoral Thesis was to study the cellular signaling processes involved in C1P-stimulated cell proliferation and migration in immune cells, mainly macrophages. Concerning the regulation of cell proliferation, this Thesis has contributed to the elucidation of three major mechanisms by which C1P exerts its mitogenic actions: 1) Stimulation of vascular endothelial growth factor (VEGF) secretion, an action that involves activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. 2) Induction of translocation and activation of protein kinase C alpha (PKCα). 3) Activation of NADPH oxidase and the subsequent formation of reactive oxygen species (ROS). With regards to cell migration, we have demonstrated that C1P stimulates the release of macrophage chemoattractant protein-1 (MCP-1), which is essential for the induction of macrophage migration. In addition, we have shown that C1P-stimulated cell migration requires the interaction of C1P with a putative C1P receptor, and the subsequent activation of the PI3K/Akt, MAPK/Erk1-2, p38 and JNK signalling pathways. Finally, this Thesis also shows that C1P increases the expression of CD69 in spleenic mononuclear cells, and that it protects these cells from entering apoptosis. A major pathway involved in this action is the mitogen-activated protein kinase kinase (MEK)/ERK1-2/ NF-кB pathway

Genero trataeraren azterketa natura zientzien alorreko testuliburu eta ikasgaietan.

Gender bias exist in many aspects of our society, and science is not an exception. The experts in this field indicate that education can be a key factor to end with gender bias and therefore, co-education should be guaranteed. Thus, educational materials and methodology must be reviewed in order to ensure their suitability. The appropriate educational material represents women and men in an equivalent manner, avoiding stereotypes and sexist language, hence, providing proper models to young students. In this research, 14 textbooks from two different high schools have been analyzed in order to evaluate gender treatment. Apart from textbook analysis, teachers of these subjects have been interviewed to know their point of view and compare it to the obtained data. From the results of this study, it can be concluded that our textbooks do not respect gender equality because women are underrepresented and some stereotypes are still present. Nonetheless, some textbooks present exercises related to the integration of female scientists’ works. Hence, there is still a room for improvement and sensitization of educational system is essential for the disappearance of gender bias from the classrooms.; Gizarteko beste hainbat alorretan bezala, zientziaren alorrean ere genero-bereizkeria gertatzen da. Adituek diotenez, hezkuntza izan daiteke egoera honi bukaera emateko giltza eta hezkidetzaren bermakuntza garrantzitsua da horretarako. Honela, ikasgelan erabiltzen diren metodologia eta ikasmaterialak egokiak ote diren ezagutzea ezinbestekoa da. Hezkidetzaren ikuspuntutik, gizon eta emakumeen trataera orekatu eta ez estereotipatu bat eskaini beharko litzaieke nerabeei, hizkuntza sexista saihestuz eta ikasle guztien garapena bermatzeko eredu egokiak eskainiz. Lan honetan, Donostiako bi ikastetxetan erabilitako 14 liburu aztertu dira aipatutako baldintzak betetzen ote diren zehazteko. Gainera, ikastetxe hauetako zientzietako ikasgaietako irakasleei galdeketa bat pasa zaie beraien ikuspuntuak bildu eta jasotako datuekin alderatzeko asmoz. Lortutako emaitzetatik ondoriozta daitekenez, testuliburuek ez dute genero trataera orekatu bat mantentzen eta oraindik hainbat estereotipo aurki daitezke. Dena den, aipatzekoa da testuliburu gutxi batzuetan emakume zientzialarien integraziorako jarduerak proposatzen direla. Beraz, asko dago hobetzeko eta hezkuntza-sistemaren sentsibilizazioa ezinbestekoa da genero bereizkeria ikasgeletatik ezabatzeko..GAKO-HITZAK: genero-bereizkeria, testuliburuak, emakume zientzialariak, hezkidetza, batxilergoa. ABSTRACT: Gender bias exist in many aspects of our society, and science is not an exception. The experts in this field indicate that education can be a key factor to end with gender bias and therefore, co-education should be guaranteed. Thus, educational materials and methodology must be reviewed in order to ensure their suitability. The appropriate educational material represents women and men in an equivalent manner, avoiding stereotypes and sexist language, hence, providing proper models to young students. In this research, 14 textbooks from two different high schools have been analyzed in order to evaluate gender treatment. Apart from textbook analysis, teachers of these subjects have been interviewed to know their point of view and compare it to the obtained data. From the results of this study, it can be concluded that our textbooks do not respect gender equality because women are underrepresented and some stereotypes are still present. Nonetheless, some textbooks present exercises related to the integration of female scientists’ works. Hence, there is still a room for improvement and sensitization of educational system is essential for the disappearance of gender bias from the classrooms.KEYWORDS: gender bias, textbooks, female scientist, co-education, bachillerat