Avaliação do metabolismo do monóxido de azoto eritrocitário

Tese de mestrado. Biologia (Biologia Humana e Ambiente). Universidade de Lisboa, Faculdade de Ciências, 2012O monóxido de azoto (NO) é formado nas células endoteliais vasculares, difundindo-se para os eritrócitos, onde regula a deformabilidade e agregação eritrocitárias, e para a parede vascular onde promove o relaxamento do músculo liso e, por conseguinte, o aumento do diâmetro dos vasos sanguíneos levando ao aumento do fluxo sanguíneo e à redução da pressão arterial. A redução da biodisponibilidade de NO e a consequente disfunção endotelial desencadeiam eventos precursores da formação da placa aterosclerótica. Como tal o estudo do metabolismo do NO eritrocitário pode contribuir para a compreensão da disfunção endotelial e patologias associadas. O objectivo deste estudo foi avaliar o metabolismo do NO eritrocitário em condições fisiológicas normais e em condições agudas de disfunção endotelial e inflamação (doentes com enfarte agudo do miocárdio – AMI). Para atingir o objectivo proposto, procurou-se estudar in vitro os efeitos da alteração conformacional da acetilcolinesterase (AChE), e da inibição/activação enzimática da proteína cinase C (PKC) sobre o metabolismo do NO eritrocitário. Ambos os efeitos foram estudados na presença/ausência de acetilcolina (ACh). Foi também realizado um ensaio ex vivo que teve como objectivo avaliar marcadores do metabolismo do NO eritrocitário em doentes na fase aguda do AMI. Os resultados obtidos demonstram que: (1) a AChE tem um papel regulador na mobilização do NO eritrocitário e na concentração intra-eritrocitária de nitrosoglutatião (GSNO); (2) o grau de fosforilação do complexo 4.1 R e da proteína banda 3 influencia o efluxo de NO e a concentração intra-eritrocitária de GSNO; (3) as alterações do grau de fosforilação da proteína banda 3 e do complexo 4.1 R favorecem uma maior concentração intra-eritrocitária de S-nitrosohemoglobina (SNO-Hb), nitratos e nitritos independentemente do efluxo de NO eritrocitário. Em relação ao estudo ex vivo, verificou-se que os eritrócitos de doentes com AMI apresentaram menor tendência no efluxo de NO e menor concentração intra-eritrocitária de GSNO.Formed in the endothelial cells, the nitric oxide (NO) spreads into erythrocytes, where it regulates erythrocyte deformability and aggregation, and into the vascular wall where it causes vascular smooth muscle relaxation and therefore increasing the diameter of blood vessels leading to blood flow rise and arterial pressure decrease. The reduction of NO bioavailability and subsequent endothelial dysfunction trigger events that lead to formation of atherosclerotic plaques. Therefore, the study of erythrocyte NO metabolism can contribute to the understanding of endothelial dysfunction and associated diseases. This research aimed to assess the erythrocyte NO metabolism under normal physiological conditions and under acute conditions of endothelial dysfunction and inflammation (patients with acute myocardial infarction - AMI). To achieve the objective, the effect of conformational change in acetilcolinesterase (AChE) and of enzymatic inhibition/activation of protein kinase C (PKC) on erythrocyte NO metabolism was investigated in vitro. Both effects were studied in the presence/absence of acetylcholine (ACh). Furthermore, an ex vivo assay that aimed to assess various markers of erythrocyte NO metabolism was conducted in patients with AMI. The results demonstrate that: (1) AChE has a regulating role in the mobilization of erythrocyte NO and intra- erythrocyte concentration of S-nitrosoglutathione (GSNO), (2) the degree of phosphorylation of complex 4.1 R and protein band 3 influences the efflux of erythrocyte NO and intra-erythrocyte concentration of GSNO, (3) changes in the degree of phosphorylation of complex 4.1 R and protein band 3 favor a higher intra- erythrocyte concentration of S-nitrosohemoglobin (SNO-Hb) , nitrates and nitrites regardless of erythrocyte NO efflux. Regarding the ex vivo study, the results seem to suggest a trend for lower erythrocyte NO efflux and a lower intra- erythrocyte concentration of GSNO in AMI patients

MAMMALS IN PORTUGAL : A data set of terrestrial, volant, and marine mammal occurrences in P ortugal

Mammals are threatened worldwide, with 26% of all species being includedin the IUCN threatened categories. This overall pattern is primarily associatedwith habitat loss or degradation, and human persecution for terrestrial mam-mals, and pollution, open net fishing, climate change, and prey depletion formarine mammals. Mammals play a key role in maintaining ecosystems func-tionality and resilience, and therefore information on their distribution is cru-cial to delineate and support conservation actions. MAMMALS INPORTUGAL is a publicly available data set compiling unpublishedgeoreferenced occurrence records of 92 terrestrial, volant, and marine mam-mals in mainland Portugal and archipelagos of the Azores and Madeira thatincludes 105,026 data entries between 1873 and 2021 (72% of the data occur-ring in 2000 and 2021). The methods used to collect the data were: live obser-vations/captures (43%), sign surveys (35%), camera trapping (16%),bioacoustics surveys (4%) and radiotracking, and inquiries that represent lessthan 1% of the records. The data set includes 13 types of records: (1) burrowsjsoil moundsjtunnel, (2) capture, (3) colony, (4) dead animaljhairjskullsjjaws, (5) genetic confirmation, (6) inquiries, (7) observation of live animal (8),observation in shelters, (9) photo trappingjvideo, (10) predators dietjpelletsjpine cones/nuts, (11) scatjtrackjditch, (12) telemetry and (13) vocalizationjecholocation. The spatial uncertainty of most records ranges between 0 and100 m (76%). Rodentia (n=31,573) has the highest number of records followedby Chiroptera (n=18,857), Carnivora (n=18,594), Lagomorpha (n=17,496),Cetartiodactyla (n=11,568) and Eulipotyphla (n=7008). The data setincludes records of species classified by the IUCN as threatened(e.g.,Oryctolagus cuniculus[n=12,159],Monachus monachus[n=1,512],andLynx pardinus[n=197]). We believe that this data set may stimulate thepublication of other European countries data sets that would certainly contrib-ute to ecology and conservation-related research, and therefore assisting onthe development of more accurate and tailored conservation managementstrategies for each species. There are no copyright restrictions; please cite thisdata paper when the data are used in publications.info:eu-repo/semantics/publishedVersio

Mammals in Portugal: a data set of terrestrial, volant, and marine mammal occurrences in Portugal

Mammals are threatened worldwide, with ~26% of all species being included in the IUCN threatened categories. This overall pattern is primarily associated with habitat loss or degradation, and human persecution for terrestrial mammals, and pollution, open net fishing, climate change, and prey depletion for marine mammals. Mammals play a key role in maintaining ecosystems functionality and resilience, and therefore information on their distribution is crucial to delineate and support conservation actions. MAMMALS IN PORTUGAL is a publicly available data set compiling unpublished georeferenced occurrence records of 92 terrestrial, volant, and marine mammals in mainland Portugal and archipelagos of the Azores and Madeira that includes 105,026 data entries between 1873 and 2021 (72% of the data occurring in 2000 and 2021). The methods used to collect the data were: live observations/captures (43%), sign surveys (35%), camera trapping (16%), bioacoustics surveys (4%) and radiotracking, and inquiries that represent less than 1% of the records. The data set includes 13 types of records: (1) burrows | soil mounds | tunnel, (2) capture, (3) colony, (4) dead animal | hair | skulls | jaws, (5) genetic confirmation, (6) inquiries, (7) observation of live animal (8), observation in shelters, (9) photo trapping | video, (10) predators diet | pellets | pine cones/nuts, (11) scat | track | ditch, (12) telemetry and (13) vocalization | echolocation. The spatial uncertainty of most records ranges between 0 and 100 m (76%). Rodentia (n =31,573) has the highest number of records followed by Chiroptera (n = 18,857), Carnivora (n = 18,594), Lagomorpha (n = 17,496), Cetartiodactyla (n = 11,568) and Eulipotyphla (n = 7008). The data set includes records of species classified by the IUCN as threatened (e.g., Oryctolagus cuniculus [n = 12,159], Monachus monachus [n = 1,512], and Lynx pardinus [n = 197]). We believe that this data set may stimulate the publication of other European countries data sets that would certainly contribute to ecology and conservation-related research, and therefore assisting on the development of more accurate and tailored conservation management strategies for each species. There are no copyright restrictions; please cite this data paper when the data are used in publications

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Acetylcholinesterase conformational states influence nitric oxide mobilization in the erythrocyte

© Springer Science+Business Media New York 2015In the human erythrocyte, band 3 protein mediates nitric oxide (NO) translocation and its effects are strongly related to phosphorylated/dephosphorylated intracellular states. The metabolism of NO could change in the presence of acetylcholinesterase (AChE). Therefore, the present study was designed to assess the effect of conformational changes in AChE (via N-19 and C-16 antibodies) and enzymatic inhibition/activation of protein kinase C (PKC) in erythrocyte NO mobilization in vitro. Our results show that by inhibiting PKC with cheletrine, impaired erythrocyte NO efflux and s-nitrosoglutathione (GSNO) levels were verified, while PKC's activation by Phorbol 12-myristate 13-acetate had the opposite effect. Those results demonstrate the influence of 4.1R complex and band 3 protein level of phosphorylation on NO efflux and GSNO concentration mediated by PKC inhibition/activation. In addition, the present study shows evidence that conformational changes in AChE promoted by incubation with N-19 and C-16 antibodies alter the enzyme's functional connection to acetylcholine (ACh) (AChE-ACh complex) in an irreversible manner, resulting in impaired GSNO concentration and NO efflux from the erythrocyte. Novel insight into NO metabolism in the erythrocyte is brought with the presented findings allowing new possibilities of modulating NO delivery, possibly involving PKC and AChE conformational alterations in combination.This study was supported by Fundação para a Ciência e a Tecnologia Grants (SFRM/BPD/6308/2009).info:eu-repo/semantics/publishedVersio

Carbapenemase-producing Klebsiella pneumoniae intra-abdominal infection successfully treated with ceftazidime/avibactam plus tigecycline

Ceftazidime/avibactam combines ceftazidime with a new beta-lactam that successfully that inhibits Amber Class A and D carbapenemases.We report a clinical case of a 61 year-old man with a carbapenemase-producing Klebsiella pneumoniae intra-abdominal infection after an elective abdominal hernia repair. The infection was successfully managed with multiple abdominal surgeries, drainage and combined antibiotic therapy with ceftazidime/avibactam plus tigecycline

Routine CD4 monitoring in HIV patients with viral suppression: Is it really necessary? A Portuguese cohort

Purpose: CD4 cell-count has been regarded as the key surrogate marker for prognostic staging and therapeutic monitoring of HIV-infected individuals. Our purpose was to assess the probability of maintaining a CD4 count >200 cells/μL in patients with continuous viral suppression and CD4 cell counts >200 cells/μL. Methods: Retrospective cohort study of HIV-infected patients, treatment naïve, who started antiretroviral therapy between 2007 and 2011. We estimated the probability of maintaining CD4 counts >200 cells/μL during continuous viral suppression using the Kaplan–Meier method. The hazard ratios of a CD4 count 200 cells/μL was 40.5 months. Ninety-three percent of patients maintained CD4 counts ≥200 cells/μL during the period of continuous viral suppression. Compared with those with an initial CD4 count ≥350 cells/μL, patients with initial CD4 count <300 cells/μL had a significantly higher risk of a CD4 count <200 cells/μL. Patients with viral suppression and CD4 counts ≥350 cells/μL had a 97.1% probability of maintaining CD4 cell counts ≥200 cells/μL for 48 months. Conclusions: The probability of a CD4 count <200 cells/μL in an HIV-infected patient with viral suppression and CD4 ≥350 cells/μL was very low. These data suggests less frequent monitoring of CD4 counts in these patients. Keywords: CD4 count monitoring, HIV infection, Viral suppression, CD4 count

Characterization of HIV-1 subtypes in a Portuguese cohort

Introduction: Distribution of HIV-1 subtypes is variable around the world, with the most common subtype in western Europe being subtype B. The aim our study was to describe the prevalence of different HIV-1 subtypes in newly diagnosed patients and identify demographic and epidemiological characteristics related with different subtypes. Materials and Methods: Retrospective single-centre study of patients newly diagnosed with HIV-1 infection between 2006 and 2012. Epidemiological data was gathered and genotyping was performed in each patient identified. Demographic and epidemiological characteristics were compared between patients with subtype B and other subtypes. Continuous variables were summarized by mean and standard deviation whereas categorical variables were presented as proportions. Comparison of groups was performed using the Chi square, Fisher exact test and Student T test. Statistical significance was assumed when p<0.05. Results: In the period of the study, 624 patients newly diagnosed with HIV-1 infection were submitted to genotypic testing but information about subtype was available only for 592 patients. General characteristics of the patients are summarized in Table 1. The distribution of the identified subtypes was the following: 286 (48.3%) patients had subtype B, 157 (26.5%) had subtype G, 54 (9.1%) had subtype C, 36 (6.1%) had subtype A, 32 (5.4%) had subtype F and 25 (4.2%) had CRF's. Patients with subtype B were more commonly male (p=0.001) and younger (p<0.0001) than those with subtypes other than B. Subtype B was more common in MSM patients, while non-B subtypes were more common in heterosexual patients and in injecting drug users (p=0.001). CD4-cell count, viral load and AIDS at presentation were not significantly different between subtypes. Resistance associated mutations were significantly more common in patients with non-B subtypes (15.4% vs 9.8%; p=0.048). Conclusions: The most commonly identified subtype was B in accordance with previous reports from other western European countries. However, in our cohort the proportion of non-B subtypes is higher than that reported for other European countries, probably reflecting the influence of strong bonds with Portuguese speaking African countries. Knowledge about HIV subtypes distribution may help understanding transmission dynamics and can be an important tool in the design of preventive measures