Btk Mutations Selectively Regulate Btk Expression And Upregulate Monocyte Xbp1 Mrna In Xla Patients.

Mutations in the Bruton agammaglobulinemia tyrosine kinase (BTK) gene are responsible for X-linked agammaglobulinemia (XLA). Unfolded or misfolded proteins can trigger stress pathways in the endoplasmic reticulum (ER), known as unfolded protein response (UPR). The aim was to clarify the involvement of UPR in XLA pathophysiology. By reverse transcription-quantitative PCR, we evaluated the expression of BTK and 12 UPR-related genes in eight patients. Moreover, we assessed the BTK protein expression and pattern in the patients' monocytes by flow cytometry and fluorescence immunocytochemistry. We found a reduced BTK expression in patients with stop codon mutations (P < 0.02). However, missense mutations did not affect BTK expression. Flow cytometry showed a reduction of BTK in patients which was corroborated by an absent or nonfunctional protein synthesis revealed by immunocytochemistry. In contrast with the other UPR-related genes, X-box binding protein 1 (XBP1) was markedly upregulated in the patients (P < 0.01), suggesting Toll-like receptor (TLR) activation since BTK directly interacts with TLRs as a negative regulator and XBP1 can be activated in direct response to TLR ligation. Different BTK mutations can be identified by the BTK expression. Inasmuch as UPR-related genes were downregulated or unaltered in patients, we speculate the involvement of the TLRs-XBP1 axis in the XLA pathophysiology. Such data could be the basis for further studies of this novel pathomechanism concerning XLA.3171-18

Sensitivity of the marine benthic copepod Tisbe biminiensis (copepoda, harpacticoida) to potassium dichromate and sediment particle size

For the future use of the marine benthic copepod Tisbe biminiensis in solid-phase sediment toxicological bioassays, the present study investigated the effect of muddy sediment from the Maracaípe estuary (northeastern Brazil), sediment particle size and the reference toxicant potassium dichromate on the species. Muddy sediment from Maracaípe can be used as control sediment, since it does not interfere in the copepod life-cycle and has metal contamination levels that are unlikely to produce any detrimental biological effects on benthic invertebrates. Neither survival nor fecundity was affected by grain size, suggesting that this species can be used with any kind of sediment from muddy to sandy. The sensitivity of T. biminiensis to K2Cr2O7 in acute tests was similar to that of other organisms. The LC50 (lethal concentration to 50% of the test organisms) medium values for T. biminiensis were 7.51, 4.68 and 3.19 mg L-1 for Cr in 48, 72 and 96 h, respectively. These results suggest that T. biminiensis is a promising organism for use in solid-phase sediment toxicity assessments.Visando o uso futuro do copépodo marinho bentônico Tisbe biminiensis em bioensaios toxicológicos de sedimentos na fase sólida, o presente estudo investigou o efeito do sedimento lamoso do estuário de Maracaípe (Nordeste do Brasil). Foram considerados a granolometria e o tóxico de referência dicromato de potássio sobre a espécie. O sedimento lamoso de Maracaípe pode ser usado como controle, uma vez que não interfere no ciclo de vida do copépodo e possui níveis de contaminação de metais que não causariam efeitos biológicos em invertebrados bentônicos. Nem a sobrevivência ou fecundidade foi afetada pelo tamanho do grão, sugerindo que esta espécie pode ser usada com qualquer tipo de sedimento, de lama a areia. A sensibilidade de T. biminiensis ao K2Cr2O7 em testes agudos foi similar a de outros organismos. Os valores de CL50 (concentração letal a 50% dos organismos) para T. biminiensis foram em média 7,51, 4,68 e 3,19 mg L-1 para o Cr em 48, 72 e 96h, respectivamente. Estes resultados sugerem que T. biminiensis é um organismo promissor para uso em avaliações de toxicidade com a fase sólida do sedimento

Unveiling the Influence of Carbon Nanotube Diameter and Surface Modification on the Anchorage of L-Asparaginase

L-asparaginase (ASNase, EC 3.5.1.1) is an amidohydrolase enzyme known for its anti-cancer properties, with an ever-increasing commercial value. Immobilization has been studied to improve the enzyme’s efficiency, enabling its recovery and reuse, enhancing its stability and half-life time. In this work, the effect of pH, contact time and enzyme concentration during the ASNase physical adsorption onto pristine and functionalized multi-walled carbon nanotubes (MWCNTs and f-MWCNTs, respectively) with different size diameters was investigated by maximizing ASNase relative recovered activity (RRA) and immobilization yield (IY). Immobilized ASNase reusability and kinetic parameters were also evaluated. The ASNase immobilization onto f-MWCNTs offered higher loading capacities, enhanced reusability, and improved enzyme affinity to the substrate, attaining RRA and IY of 100 and 99%, respectively, at the best immobilization conditions (0.4 mg/mL of ASNase, pH 8, 30 min of contact time). In addition, MWCNTs diameter proved to play a critical role in determining the enzyme binding affinity, as evidenced by the best results attained with f-MWCNTs with diameters of 10–20 nm and 20–40 nm. This study provided essential information on the impact of MWCNTs diameter and their surface functionalization on ASNase efficiency, which may be helpful for the development of innovative biomedical devices or food pre-treatment solutionspublishe

Patient-physician discordance in assessment of adherence to inhaled controller medication: a cross-sectional analysis of two cohorts

We aimed to compare patient's and physician's ratings of inhaled medication adherence and to identify predictors of patient-physician discordance.(SFRH/BPD/115169/2016) funded by Fundação para a Ciência e Tecnologia (FCT); ERDF (European Regional Development Fund) through the operations: POCI-01-0145-FEDER-029130 ('mINSPIRERS—mHealth to measure and improve adherence to medication in chronic obstructive respiratory diseases—generalisation and evaluation of gamification, peer support and advanced image processing technologies') cofunded by the COMPETE2020 (Programa Operacional Competitividade e Internacionalização), Portugal 2020 and by Portuguese Funds through FCT (Fundação para a Ciência e a Tecnologia).info:eu-repo/semantics/publishedVersio

Inhibition of Bacterial and Fungal Biofilm Formation by 675 Extracts from Microalgae and Cyanobacteria

Bacterial biofilms are complex biological systems that are difficult to eradicate at a medical, industrial, or environmental level. Biofilms confer bacteria protection against external factors and antimicrobial treatments. Taking into account that about 80% of human infections are caused by bacterial biofilms, the eradication of these structures is a great priority. Biofilms are resistant to old-generation antibiotics, which has led to the search for new antimicrobials from different sources, including deep oceans/seas. In this study, 675 extracts obtained from 225 cyanobacteria and microalgae species (11 phyla and 6 samples belonging to unknown group) were obtained from different culture collections: The Blue Biotechnology and Ecotoxicology Culture Collection (LEGE-CC), the Coimbra Collection of Algae (ACOI) from Portugal, and the Roscoff Culture Collection (RCC) from France. The largest number of samples was made up of the microalgae phylum Chlorophyta (270) followed by Cyanobacteria (261). To obtain a large range of new bioactive compounds, a method involving three consecutive extractions (hexane, ethyl acetate, and methanol) was used. The antibiofilm activity of extracts was determined against seven different bacterial species and two Candida strains in terms of minimal biofilm inhibitory concentration (MBIC). The highest biofilm inhibition rates (%) were achieved against Candida albicans and Enterobacter cloacae. Charophyta, Chlorophyta, and Cyanobacteria were the most effective against all microorganisms. In particular, extracts of Cercozoa phylum presented the lowest MBIC50 and MBIC90 values for all the strains except C. albicans

New endoperoxides highly active in vivo and in vitro against artemisinin-resistant Plasmodium falciparum

Background: The emergence and spread of Plasmodium falciparum resistance to artemisinin-based combination therapy in Southeast Asia prompted the need to develop new endoperoxide-type drugs. Methods: A chemically diverse library of endoperoxides was designed and synthesized. The compounds were screened for in vitro and in vivo anti-malarial activity using, respectively, the SYBR Green I assay and a mouse model. Ring survival and mature stage survival assays were performed against artemisinin-resistant and artemisinin-sensitive P. falciparum strains. Cytotoxicity was evaluated against mammalian cell lines V79 and HepG2, using the MTT assay. Results: The synthesis and anti-malarial activity of 21 new endoperoxide-derived compounds is reported, where the peroxide pharmacophore is part of a trioxolane (ozonide) or a tetraoxane moiety, flanked by adamantane and a substituted cyclohexyl ring. Eight compounds exhibited sub-micromolar anti-malarial activity (IC50 0.3–71.1 nM), no cross-resistance with artemisinin or quinolone derivatives and negligible cytotoxicity towards mammalian cells. From these, six produced ring stage survival < 1% against the resistant strain IPC5202 and three of them totally suppressed Plasmodium berghei parasitaemia in mice after oral administration. Conclusion: The investigated, trioxolane–tetrazole conjugates LC131 and LC136 emerged as potential anti-malarial candidates; they show negligible toxicity towards mammalian cells, ability to kill intra-erythrocytic asexual stages of artemisinin-resistant P. falciparum and capacity to totally suppress P. berghei parasitaemia in mice.info:eu-repo/semantics/publishedVersio

Is axenicity crucial to cryopreserve microalgae?

Large culture collections of microalgae and cyanobacteria such as the Coimbra Collection of Algae (ACOI) hold unialgal cultures consisting of a population of cells/colonies of a certain species. These cultures are usually non-axenic, as other organisms such as bacteria and microfungi are also present in culture due to co-isolation. Attention has been recently given to partner organisms since studies indicate that some bacteria are important for nutrient uptake of the algal cells, acting as simbionts. Despite this benign effect in the actively growing cultures, when cryopreservation is applied for inactive-stage storage, these organisms may recover faster than the algae, thus affecting their recovery and the viability assessments. In this study, a set of mucilaginous ACOI microalgae were selected, cell features known for their relevance in cryopreservation success were recorded and simple two-step cryopreservation tests were applied. Thawed samples were transferred to fresh culture medium for recovery. Viability was assessed and partner organism proliferation (pop) was recorded. Results were analyzed by t-tests. Statistical models allowed us to support the known tendency for small, unicellular algae with no outer structures to be successfully cryopreserved and the negative effect of vacuoles in the cell prior to cryopreservation. On average cryopreservation with MeOH or Me2SO led to the recovery of nearly half the cells. It was found that the cryoprotection step with MeOH is when pop is triggered and that the use of Me2SO can prevent this effect. Progress on understanding the cultured consortia will assist the improvement of cryopreservation and research using microalgal cultures.The authors acknowledge financial support provided by the Portuguese National Science and Technology Foundation (FCT) for the research project PTDC/BIA-QOR/71319/2006 and for the Ph.D. grant SFRH/BD/73359/2010 to Raquel