RESUMO
Three-dimensional microscopy is increasingly prevalent in biology due to the development of techniques such as multiphoton, spinning disk confocal, and light sheet fluorescence microscopies. These methods enable unprecedented studies of life at the microscale, but bring with them larger and more complex datasets. New image processing techniques are therefore called for to analyze the resulting images in an accurate and efficient manner. Convolutional neural networks are becoming the standard for classification of objects within images due to their accuracy and generalizability compared to traditional techniques. Their application to data derived from 3D imaging, however, is relatively new and has mostly been in areas of magnetic resonance imaging and computer tomography. It remains unclear, for images of discrete cells in variable backgrounds as are commonly encountered in fluorescence microscopy, whether convolutional neural networks provide sufficient performance to warrant their adoption, especially given the challenges of human comprehension of their classification criteria and their requirements of large training datasets. We therefore applied a 3D convolutional neural network to distinguish bacteria and non-bacterial objects in 3D light sheet fluorescence microscopy images of larval zebrafish intestines. We find that the neural network is as accurate as human experts, outperforms random forest and support vector machine classifiers, and generalizes well to a different bacterial species through the use of transfer learning. We also discuss network design considerations, and describe the dependence of accuracy on dataset size and data augmentation. We provide source code, labeled data, and descriptions of our analysis pipeline to facilitate adoption of convolutional neural network analysis for three-dimensional microscopy data.
Assuntos
Bactérias/classificação , Bactérias/ultraestrutura , Imageamento Tridimensional/métodos , Redes Neurais de Computação , Algoritmos , Animais , Biologia Computacional , Bases de Dados Factuais/estatística & dados numéricos , Humanos , Imageamento Tridimensional/estatística & dados numéricos , Intestinos/microbiologia , Microscopia de Fluorescência , Pseudomonas/ultraestrutura , Máquina de Vetores de Suporte , Vibrio/ultraestrutura , Peixe-Zebra/microbiologiaRESUMO
OBJECTIVE: Introduction: The article presents data from literary sources and a statistical analysis of one's own research on the nature, mechanism and prescription of spleen injury in the case of mechanical trauma and the absence of alcohol intoxication. The aim: To study the dynamics of changes in the histological parameters of the spleen injured tissues in case of mechanical trauma depending on the prescription of injury. PATIENTS AND METHODS: Materials and methods: The material of the study was the spleen tissue of 56 males and females aged from 20-60 who died at known and unknown time in the presence and absence of alcohol in the blood. We used histological, histochemical methods, and carried out a statistical analysis of the results. RESULTS: Results: The obtained results showed that during the mechanical injury of spleen there often developed a capsule and a parenchyma with hematoma in the area of injury. Our records showed that during the first 6 hours after injury, there appeared a hematoma in the center of the injury. Hemolysis of the erythrocyte particles was observed in the center of the hematoma. There were isolated leukocytes and fibrin tissues closer to the edge of the hematoma. CONCLUSION: Conclusions: The obtained results indicate that there are several histological changes in the damaged spleen tissues area which directly depend on the time which passed from the moment of injury.
Assuntos
Medicina Legal/métodos , Hematoma/diagnóstico , Técnicas Histológicas , Baço/lesões , Adulto , Feminino , Fibrina , Humanos , Leucócitos , Masculino , Pessoa de Meia-Idade , Ferimentos e Lesões , Adulto JovemRESUMO
The fluids of the intestine serve as a physical barrier to pathogens, a medium for the diffusion of nutrients and metabolites, and an environment for commensal microbes. The rheological properties of intestinal mucus have therefore been the subject of many investigations, thus far limited to in vitro studies due to the difficulty of measurement in the natural context of the gut. This limitation especially hinders our understanding of how the gut microbiota interact with the intestinal space, since examination of this calls not only for in vivo measurement techniques, but for techniques that can be applied to model organisms in which the microbial state of the gut can be controlled. We have addressed this challenge with two complementary approaches. We performed passive microrheological measurements using thermally driven nanoparticles and active microrheology using micron-scale ellipsoidal magnetic microparticles, in both cases using light-sheet fluorescence microscopy to optically access the intestinal bulb of the larval zebrafish, a model vertebrate. We present viscosity measurements in germ-free animals (devoid of gut microbes), animals colonized by a single bacterial species, and conventionally reared animals, and find that in all cases, the mucin-rich intestinal liquid is well described as a Newtonian fluid. Surprisingly, despite known differences in the number of secretory cells in germ-free zebrafish and their conventional counterparts, the fluid viscosity for these two groups is very similar, as measured with either technique. Our study provides, to our knowledge, the first in vivo microrheological measurements of the intestinal space in living animals, and we comment on its implications for timescales of host-microbe interactions in the gut.
Assuntos
Líquido Extracelular/metabolismo , Larva/citologia , Reologia , Peixe-Zebra , AnimaisRESUMO
In this study, through severe reduced-scale braking tests, we investigate the wear and integrity of organic matrix brake pads against gray cast iron (GCI) discs. Two prototype pad materials are designed with the aim of representing a typical non-metal NAO and a low-steel (LS) formulation. The worn surfaces are observed with SEM. The toughness of the pad materials is tested at the raw state and after a heat treatment. During braking, the LS-GCI disc configuration produces heavy wear. The friction parts both keep their macroscopic integrity and wear appears to be homogeneous. The LS pad is mostly covered by a layer of solid oxidized steel. The NAO-GCI disc configuration wears dramatically and cannot reach the end of the test program. The NAO pad suffers many deep cracks. Compacted third body plateaus are scarce and the corresponding disc surface appears to be very heterogeneous. The pad materials both show similar strength at the raw state and similar weakening after heat treatment. However, the NAO material is much more brittle than the LS material in both states, which seems to favor the growth of cracks. The observations of crack faces suggest that long steel fibers in the LS material palliate the brittleness of the matrix, even after heat damage.
RESUMO
The microbial communities resident in animal intestines are composed of multiple species that together play important roles in host development, health, and disease. Due to the complexity of these communities and the difficulty of characterizing them in situ, the determinants of microbial composition remain largely unknown. Further, it is unclear for many multispecies consortia whether their species-level makeup can be predicted based on an understanding of pairwise species interactions or whether higher-order interactions are needed to explain emergent compositions. To address this, we examine commensal intestinal microbes in larval zebrafish, initially raised germfree, to allow the introduction of controlled combinations of bacterial species. Using a dissection and plating assay, we demonstrate the construction of communities of one to five bacterial species and show that the outcomes from the two-species competitions fail to predict species abundances in more complex communities. With multiple species present, interbacterial interactions become weaker, suggesting that higher-order interactions in the vertebrate gut stabilize complex communities.IMPORTANCE Understanding the rules governing the composition of the diverse microbial communities that reside in the vertebrate gut environment will enhance our ability to manipulate such communities for therapeutic ends. Synthetic microbial communities, assembled from specific combinations of microbial species in germfree animals, allow investigation of the fundamental question of whether multispecies community composition can be predicted solely based on the combined effects of interactions between pairs of species. If so, such predictability would enable the construction of communities with desired species from the bottom up. If not, the apparent higher-order interactions imply that emergent community-level characteristics are crucial. Our findings using up to five coexisting native bacterial species in larval zebrafish, a model vertebrate, provide experimental evidence for higher-order interactions and, moreover, show that these interactions promote the coexistence of microbial species in the gut.
Assuntos
Bactérias/metabolismo , Microbioma Gastrointestinal , Trato Gastrointestinal/microbiologia , Simbiose , Peixe-Zebra/microbiologia , Animais , Bactérias/classificação , Fenômenos Fisiológicos Bacterianos , Larva/microbiologiaRESUMO
Correlating the presence of bacteria and the genes they carry with aspects of plant and animal biology is rapidly outpacing the functional characterization of naturally occurring symbioses. A major barrier to mechanistic studies is the lack of tools for the efficient genetic manipulation of wild and diverse bacterial isolates. To address the need for improved molecular tools, we used a collection of proteobacterial isolates native to the zebrafish intestinal microbiota as a testbed to construct a series of modernized vectors that expedite genetic knock-in and knockout procedures across lineages. The innovations that we introduce enhance the flexibility of conventional genetic techniques, making it easier to manipulate many different bacterial isolates with a single set of tools. We developed alternative strategies for domestication-free conjugation, designed plasmids with customizable features, and streamlined allelic exchange using visual markers of homologous recombination. We demonstrate the potential of these tools through a comparative study of bacterial behavior within the zebrafish intestine. Live imaging of fluorescently tagged isolates revealed a spectrum of distinct population structures that differ in their biogeography and dominant growth mode (i.e., planktonic versus aggregated). Most striking, we observed divergent genotype-phenotype relationships: several isolates that are predicted by genomic analysis and in vitro assays to be capable of flagellar motility do not display this trait within living hosts. Together, the tools generated in this work provide a new resource for the functional characterization of wild and diverse bacterial lineages that will help speed the research pipeline from sequencing-based correlations to mechanistic underpinnings.IMPORTANCE A great challenge in microbiota research is the immense diversity of symbiotic bacteria with the capacity to impact the lives of plants and animals. Moving beyond correlative DNA sequencing-based studies to define the cellular and molecular mechanisms by which symbiotic bacteria influence the biology of their hosts is stalling because genetic manipulation of new and uncharacterized bacterial isolates remains slow and difficult with current genetic tools. Moreover, developing tools de novo is an arduous and time-consuming task and thus represents a significant barrier to progress. To address this problem, we developed a suite of engineering vectors that streamline conventional genetic techniques by improving postconjugation counterselection, modularity, and allelic exchange. Our modernized tools and step-by-step protocols will empower researchers to investigate the inner workings of both established and newly emerging models of bacterial symbiosis.
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Técnicas Genéticas , Genoma Bacteriano , Microbiota , Proteobactérias/classificação , Animais , Técnicas de Introdução de Genes , Técnicas de Inativação de Genes , Intestinos/microbiologia , Fenótipo , Plasmídeos , Análise de Sequência de DNA , Simbiose , Peixe-Zebra/microbiologiaRESUMO
Light sheet fluorescence microscopy enables fast, minimally phototoxic, three-dimensional imaging of live specimens, but is currently limited by low throughput and tedious sample preparation. Here, we describe an automated high-throughput light sheet fluorescence microscope in which specimens are positioned by and imaged within a fluidic system integrated with the sheet excitation and detection optics. We demonstrate the ability of the instrument to rapidly examine live specimens with minimal manual intervention by imaging fluorescent neutrophils over a nearly 0.3 mm3 volume in dozens of larval zebrafish. In addition to revealing considerable inter-individual variability in neutrophil number, known previously from labor-intensive methods, three-dimensional imaging allows assessment of the correlation between the bulk measure of total cellular fluorescence and the spatially resolved measure of actual neutrophil number per animal. We suggest that our simple experimental design should considerably expand the scope and impact of light sheet imaging in the life sciences.
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Embrião não Mamífero , Larva , Microscopia de Fluorescência/métodos , Peixe-Zebra , Animais , Imageamento Tridimensional/métodosRESUMO
It is increasingly practical to co-opt many native cellular components into use as elements of synthetic biological systems. We present the design and experimental investigation of the first exogenous genetic construct to be successfully targeted by RNA activation, a phenomenon whereby small double-stranded RNAs increase gene expression from sequence-similar promoters by a mechanism thought to be related to that of RNA interference. Our selection of activating RNA candidates was informed by a custom-written computer program designed to choose target sites in the promoter of interest according to a set of empirical optimality criteria drawn from prior research. Activating RNA candidates were assessed for activity against two exogenously derived target promoters, with successful candidates being subjected to further rounds of validation as a precaution against potential off-target effects. A genetic platform was assembled that allowed activating RNA candidates to be simultaneously screened both for positive activity on the target reporter gene and for possible nonspecific effects on cell metabolism. Several candidate sequences were tested to appraise the utility of this platform, with the most successful achieving a moderate activation level with minimal off-target effects.
Assuntos
Marcação de Genes/métodos , RNA de Cadeia Dupla/metabolismo , Linhagem Celular , Expressão Gênica , Genes Reporter , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Regiões Promotoras Genéticas , Interferência de RNA , RNA de Cadeia Dupla/antagonistas & inibidores , RNA de Cadeia Dupla/genética , Transfecção , Interface Usuário-ComputadorRESUMO
The growth rate and carrying capacity of a cell population are key to the characterization of the population's viability and to the quantification of its responses to perturbations such as drug treatments. Accurate estimation of these parameters necessitates careful analysis. Here, we present a rigorous mathematical approach for the robust analysis of cell count data, in which all the experimental stages of the cell counting process are investigated in detail with the machinery of Bayesian probability theory. We advance a flexible theoretical framework that permits accurate estimates of the growth parameters of cell populations and of the logical correlations between them. Moreover, our approach naturally produces an objective metric of avoidable experimental error, which may be tracked over time in a laboratory to detect instrumentation failures or lapses in protocol. We apply our method to the analysis of cell count data in the context of a logistic growth model by means of a user-friendly computer program that automates this analysis, and present some samples of its output. Finally, we note that a traditional least squares fit can provide misleading estimates of parameter values, because it ignores available information with regard to the way in which the data have actually been collected.
Assuntos
Contagem de Células/métodos , Proliferação de Células , Modelos Biológicos , Acesso à Informação , Algoritmos , Teorema de Bayes , Linhagem Celular Tumoral , Feminino , Humanos , Internet , Análise dos Mínimos Quadrados , Modelos Logísticos , Neoplasias Ovarianas/fisiopatologia , Probabilidade , Software , TempoRESUMO
OBJECTIVE: To investigate the influence of haemorrhagic shock in mice on ex vivo TNF production by whole blood cells (WBC) stimulated through Toll-like receptors (TLR) 4 and 2. STUDY DESIGN AND ANIMALS: Experimental study using BALB/c male mice. METHODS: Haemorrhage (0,026+/-0,003 ml/g) by transparietal cardiac puncture under general anaesthesia. Measurement of left intraventricular pressure through a direct subcostal cardiac puncture. Possible restitution of shed blood volume (SBV) in retroorbital venous plexus, 60 minutes following haemorrhage. Lethal exsanguination 120 minutes following general anaesthesia (Control group), cardiac puncture (Sham group), blood sample (Haemorrhage group), or 60 minutes following SBV retransfusion (SBV group). Cultures (24 hours) of whole blood from the exsanguination, alone or with Escherichia coli endotoxin (LPS, TLR 4) or with heat-killed Staphylococcus aureus Cowan (SAC, TLR 2). Assessment of TNF levels in the cultures supernatant (Elisa). RESULTS: Hemorrhage (approximately 30% of calculated blood volume) resulted in arterial hypotension (-50%) which was reversed by SBV retransfusion. TNF production by LPS-stimulated WBC was reduced by haemorrhage (approximately -50%) with or without SBV retransfusion. TNF production by SAC-stimulated WBC remained unchanged. CONCLUSION: The reduction of proinflammatory cytokines production by WBC stimulated with pathogen-associated molecular patterns is not a generalized phenomenon following murin haemorrhagic shock. It depends on the used stimulus and studied signalling pathways.
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Receptores de Superfície Celular/fisiologia , Choque Hemorrágico/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Animais , Transfusão de Sangue , Células Cultivadas , Hipotensão/etiologia , Hipotensão/fisiopatologia , Leucócitos/metabolismo , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Choque Hemorrágico/fisiopatologia , Infecções Estafilocócicas/fisiopatologia , Receptor 2 Toll-Like , Receptor 4 Toll-LikeRESUMO
HIV-1 innate sensing requires direct contact of infected CD4+ T cells with plasmacytoid dendritic cells (pDCs). In order to study this process, the protocols described here use freshly isolated human peripheral blood mononuclear cells (PBMCs) or plasmacytoid dendritic cells (pDCs) to sense infections in either T cell line (MT4) or heterologous primary CD4+ T cells. In order to ensure proper sensing, it is essential that PBMC are isolated immediately after blood collection and that optimal percentage of infected T cells are used. Furthermore, multi-parametric flow cytometric staining can be used to confirm that PBMC samples contain the different cell lineages at physiological ratios. A number of controls can also be included to evaluate viability and functionality of pDCs. These include, the presence of specific surface markers, assessing cellular responses to known agonist of Toll-Like Receptors (TLR) pathways, and confirming a lack of spontaneous type-I interferon (IFN) production. In this system, freshly isolated PBMCs or pDCs are co-cultured with HIV-1 infected cells in 96 well plates for 18-22 hr. Supernatants from these co-cultures are then used to determine the levels of bioactive type-I IFNs by monitoring the activation of the ISGF3 pathway in HEK-Blue IFN-α/ß cells. Prior and during co-culture conditions, target cells can be subjected to flow cytometric analysis to determine a number of parameters, including the percentage of infected cells, levels of specific surface markers, and differential killing of infected cells. Although, these protocols were initially developed to follow type-I IFN production, they could potentially be used to study other imuno-modulatory molecules released from pDCs and to gain further insight into the molecular mechanisms governing HIV-1 innate sensing.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Técnicas de Cocultura/métodos , HIV-1/imunologia , Linfócitos T CD4-Positivos/citologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Células Dendríticas/virologia , Citometria de Fluxo , Humanos , Interferon Tipo I/imunologia , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologiaRESUMO
The effects of frovatriptan and sumatriptan on internal carotid and coronary vascular haemodynamics were investigated and compared in conscious dogs. Frovatriptan and sumatriptan (0.1 - 100 microg kg(-1)) induced a transient increase in external coronary artery diameter (eCOD) of up to 2.9+/-1.2 and 1.8+/-0.6%, respectively (both P:<0.05). This was followed by a prolonged and dose-dependent decrease in eCOD of up to -5.2+/-1.2 and -5.3+/-0.9% (both P:<0.05), with ED(50) values of 86+/-21 and 489+/-113 micromol kg(-1), respectively. In contrast, only a decrease in the external diameter of the internal carotid artery was observed (-6.0+/-0.6 and -6.2+/-1.4%, both P:<0.05, and ED(50) values of 86+/-41 and 493+/-162 micromol kg(-1), respectively). Frovatriptan was thus 5.7 fold more potent than sumatriptan at the level of both large coronary and internal carotid arteries. After endothelium removal by balloon angioplasty in coronary arteries, the initial dilatation induced by the triptans was abolished and delayed constriction enhanced. The selective antagonist for the 5-HT(1B) receptors SB224289 dose-dependently blocked the effects of sumatriptan on large coronary and internal carotid arteries whereas the selective antagonist for the 5-HT(1D) receptors BRL15572 did not affect any of these effects. In conclusion, frovatriptan and sumatriptan initially dilate and subsequently constrict large coronary arteries in the conscious dog, whereas they directly constrict the internal carotid artery. The vascular endothelium modulates the effects of these triptans on large coronary arteries. Finally, 5-HT(1B) but not 5-HT(1D) receptors are primarily involved in canine coronary and internal carotid vasomotor responses to sumatriptan.
Assuntos
Carbazóis/farmacologia , Artéria Carótida Interna/efeitos dos fármacos , Vasos Coronários/efeitos dos fármacos , Agonistas do Receptor de Serotonina/farmacologia , Sumatriptana/farmacologia , Resistência Vascular/efeitos dos fármacos , Animais , Artéria Carótida Interna/fisiologia , Vasos Coronários/fisiologia , Cães , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiologia , Hemodinâmica/efeitos dos fármacos , Hemodinâmica/fisiologia , Receptor 5-HT1B de Serotonina , Receptores de Serotonina/efeitos dos fármacos , Receptores de Serotonina/fisiologia , Triptaminas , Resistência Vascular/fisiologia , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologia , Vasodilatação/efeitos dos fármacos , Vasodilatação/fisiologiaRESUMO
Onset of adult T-cell leukemia (ATL) usually follows a long period of viral latency. Strongyloides stercoralis infection has been considered a cofactor of leukemogenesis. Hypereosinophilia (HE) is also observed and could be associated with either the presence of parasites or the leukemic process. In non-Hodgkin's lymphoma, eosinophilia may or may not affect prognosis. To determine whether infection with S stercoralis and therefore eosinophilia has a significant effect on the development of ATL, we studied two variables in 38 patients: age at onset and median survival rate. Infected (Ss+) patients (n = 19) were younger (P = .0002) and survived longer (P = .0006) than uninfected (Ss-) patients (n = 19) (median age, 39 vs 70 years; median survival, 167 vs 30 days). Mean survival of patients with hypereosinophilia (HE+) was not significantly different from that of patients without hypereosinophilia (HE-) (P = .57). However, overall survival was longer for Ss + HE + patients than for Ss-HE-patients (P = .01; 180 vs 30 days) or Ss-HE + patients (P = .03; 180 vs 45 days). Among patients with mean survival more than 180 days, Ss + HE + patients survived longer (P = .028). Our data confirm that cofactors related to the environment, such as S stercoralis and hypereosinophilia associated with S stercoralis or human T-cell leukemia virus, type 1 (HTLV-1) might be important in HTLV-1-associated leukemogenesis and suggest that hypereosinophilia affects the prognosis of HTLV-1-associated leukemia.
Assuntos
Eosinofilia/parasitologia , Leucemia-Linfoma de Células T do Adulto/parasitologia , Strongyloides stercoralis , Estrongiloidíase/etiologia , Adulto , Idade de Início , Idoso , Idoso de 80 Anos ou mais , Animais , Eosinofilia/mortalidade , Feminino , Humanos , Leucemia-Linfoma de Células T do Adulto/mortalidade , Masculino , Martinica/epidemiologia , Pessoa de Meia-Idade , Prognóstico , Estrongiloidíase/mortalidade , Taxa de SobrevidaRESUMO
OBJECTIVE: To determine the pharmacokinetics and absolute bioavailability of ciprofloxacin in 12 critically ill patients receiving continuous enteral feeding. DESIGN: a prospective, cross-over study. SETTING: 12-bed surgical intensive care unit in a University Hospital. PATIENTS: 12 stable critically ill patients on mechanical ventilation and receiving continuous enteral feeding (Normoreal fibres) without diarrhea or excessive residual gastric contents ( < 200 ml/4 h). None had gastro-intestinal disease, renal insufficiency (estimated creatinine clearance > or = 50 ml/min) or was receiving medications that could interfere with ciprofloxacin absorption or metabolism. MEASUREMENTS AND MAIN RESULTS: The study was carried out after the fourth (steady state) b. i. d. intravenous (i. v.) 1-h infusion of 400 mg and the second b. i. d. nasogastric (NG) dose of 750 mg (crushed tablet in suspension). Plasma concentrations were measured by high-performance liquid chromatography. The median (range) peak concentration after i. v. infusion was 4.1 (1.5-7.4) mg/l, and that after NG administration was 2.3 (0.7-5.8) mg/l, occurring 1.25 (0.75-3.33) h after dosing. The median [range] areas under plasma concentration-time curves were similar for the two administration routes (10.3 [3.3-34.6] and 8.4 [3.6-53.4] for i.v. infusion and NG administration, respectively). Ciprofloxacin bioavailability ranges from 31 to 82 % (median, 44%). CONCLUSIONS: In tube-fed critically ill patients, a switch to the NG ciprofloxacin after initial i. v. therapy to simplify the treatment of severe infections is restricted to those for whom serial assessments of ciprofloxacin levels are routinely available.
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Anti-Infecciosos/administração & dosagem , Anti-Infecciosos/farmacocinética , Ciprofloxacina/administração & dosagem , Ciprofloxacina/farmacocinética , Nutrição Enteral/métodos , Intubação Gastrointestinal , Adulto , Idoso , Anti-Infecciosos/sangue , Disponibilidade Biológica , Cromatografia Líquida de Alta Pressão , Ciprofloxacina/sangue , Estado Terminal , Estudos Cross-Over , Monitoramento de Medicamentos , Feminino , Humanos , Infusões Intravenosas , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Fatores de TempoRESUMO
OBJECTIVE: To investigate the effects of low-dose dopamine (Dop) on renal hemodynamics and function in patients with brain trauma receiving norepinephrine (NE). DESIGN: Prospective clinical study. SETTING: Surgical intensive care unit of a university hospital. PATIENTS: 20 stable, non-septic, mechanically ventilated, sedated patients with brain trauma and normal renal function treated with intravenous NE (0.11-0.65 microg/kg per min) to maintain an adequate cerebral perfusion pressure (> 60 mmHg). INTERVENTIONS: Two successive 1-h study periods with NE alone then NE + Dop (2 microg/kg per min). During each period, creatinine (Cl(CREAT)), sodium (Cl(Na)), potassium (Cl(K)), osmolar (Cl(OSM)) and free water (Cl(H2O)), clearances were measured in all the patients. Effective renal blood flow (ERBF, para-aminohippurate clearance) and glomerular filtration rate (GFR, inulin clearance) were measured in 7 of the 20 patients. RESULTS: Dop during NE infusion induced increases in urine flow and natriuresis which were not correlated with possible changes in arterial pressure. Cl(CREAT), GFR and their difference remained unchanged, whereas ERBF tended to increase. Fractional sodium excretion [100 x (Cl(Na)/Cl(CREAT)] and C1(K) increased during Dop infusion. CONCLUSION: The mechanism of Dop-induced natriuresis during NE infusion in brain trauma patients seems mainly related to a direct tubular effect of the drug.
Assuntos
Lesões Encefálicas/fisiopatologia , Cardiotônicos/farmacologia , Dopamina/farmacologia , Hipertensão Intracraniana/tratamento farmacológico , Rim/efeitos dos fármacos , Vasoconstritores/farmacologia , Adolescente , Agonistas alfa-Adrenérgicos/administração & dosagem , Agonistas alfa-Adrenérgicos/farmacologia , Adulto , Lesões Encefálicas/terapia , Cardiotônicos/administração & dosagem , Dopamina/administração & dosagem , Feminino , Escala de Coma de Glasgow , Hemodinâmica/efeitos dos fármacos , Humanos , Escala de Gravidade do Ferimento , Pressão Intracraniana , Rim/fisiopatologia , Masculino , Norepinefrina/administração & dosagem , Norepinefrina/farmacologia , Estudos Prospectivos , Vasoconstritores/administração & dosagemRESUMO
OBJECTIVE: To describe the evolution of systemic and regional blood flows during and after hypovolemia in humans. DESIGN: Simulation of hypovolemia by a prolonged application of lower body negative pressure (LBNP). SETTING: Laboratory of Clinical Research, Surgical Intensive Care Unit of an University Hospital. PARTICIPANTS: 8 healthy male volunteers. INTERVENTIONS: 3 successive and increasing 15 min-levels of LBNP were followed by a progressive return (10 min) to atmospheric pressure, then a 60 min-recovery period. MEASUREMENTS AND MAIN RESULTS: Simulated hypovolemia induced a parallel one-third decrease in cardiac output (bioimpedance), musculocutaneous (venous plethysmography) and splanchnic (ICG clearance) blood flows. Adrenergic-mediated peripheral vasoconstriction prevented any change in mean arterial pressure. The decrease in renal blood flow (PAH clearance) was limited, glomerular filtration rate (inulin clearance) unchanged and thus filtration fraction increased. All the cardiovascular and biological variables returned to pre-LBNP values during the recovery period except for splanchnic blood flow which remained below control values 60 min after the return to atmospheric pressure. CONCLUSIONS: Since a sustained splanchnic vasoconstriction follows a transient normotensive hypovolemia in healthy men despite adequate treatment considering arterial pressure and cardiac output, the therapeutic goals of fluid resuscitation after hypovolemic shock might be revisited and a supranormal value of cardiac output proposed.
Assuntos
Pressão Negativa da Região Corporal Inferior , Fluxo Sanguíneo Regional , Choque/fisiopatologia , Adulto , Débito Cardíaco , Hidratação/métodos , Taxa de Filtração Glomerular , Humanos , Masculino , Circulação Renal , Ressuscitação/métodos , Choque/terapia , Circulação Esplâncnica , Fatores de Tempo , VasoconstriçãoRESUMO
OBJECTIVES: To describe the initial evolution of serum procalcitonin (PCT) and C-reactive protein (CRP) in previously healthy adult trauma patients and to compare the relationship of the expression of these two proteins with indicators of trauma severity. DESIGN: Prospective, descriptive, longitudinal study. SETTING: Surgical ICU in an university hospital. PATIENTS: Twenty-one patients admitted during the first posttraumatic 3 h exhibiting an Injury Severity Score (ISS) between 16 and 50 were enrolled. MEASUREMENTS: Blood sampling was performed on admission and on posttraumatic days 0.5, 1, 2 and 3 to assess serum levels of PCT and CRP. Total creatine kinase (CKtot) and lactate dehydrogenase (LDHtot) activities in the serum were used as tissue damage indicators. RESULTS: PCT exhibited an early and transient increase in serum levels similar to a more delayed change of CRP levels. Peak PCT and peak CRP were related to the ISS, the extent of tissue damage and the amount of fluid replacement during the first day. During the first 3 posttraumatic days, 90% of the patients exhibited a generalized inflammatory syndrome without infection. CONCLUSIONS: An early and transient release of PCT into the circulation was observed after severe trauma and the amount of circulating PCT seemed proportional to the severity of tissue injury and hypovolemia, yet unrelated to infection. The predictive value of both PCT and CRP for a forthcoming multiple organ failure still remains to be clarified.
Assuntos
Proteína C-Reativa/metabolismo , Calcitonina/sangue , Precursores de Proteínas/sangue , Ferimentos e Lesões/imunologia , Adulto , Peptídeo Relacionado com Gene de Calcitonina , Creatina Quinase/sangue , Feminino , Humanos , Escala de Gravidade do Ferimento , L-Lactato Desidrogenase/sangue , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Ferimentos e Lesões/enzimologiaRESUMO
OBJECTIVE: To verify whether the determination of the percentage of cells recovered by bronchoalveolar lavage and containing fat inclusions is a useful diagnostic tool of posttraumatic pulmonary fat embolism. DESIGN: Prospective study. SETTING: Surgical Intensive Care Units in two university hospitals. PATIENTS: 56 successive trauma patients needing prolonged postinjury mechanical ventilation, including 4 with clinical definite fat embolism syndrome, 5 in whom the diagnosis had been clinically suspected but was impossible to confirm or exclude before bronchoscopy, and 47 with no clinical evidence of the syndrome. Control groups included 8 patients without previous trauma who developed ARDS and 6 healthy surgical patients. METHODS: Bronchoalveolar lavage was performed within the first post-traumatic 3 days in trauma patients, at the beginning of the pulmonary disease in non trauma ARDS patients and just after anesthesic induction in healthy ortopedic patients. The magnitude of lipid content in alveolar cells was compared with the clinical pattern of the pulmonary fat embolism syndrome retrospectively evaluated at the seventh day postinjury in trauma patients. RESULTS: All the patients with definite fat embolism syndrome had more than 70% of lavage cells containing fat droplets. The group of patients in whom the diagnosis of the fat embolism syndrome was suspected had percentages of fat cells above 30% in 4 out of 5 patients. A percentage of fat cells above 30% was only observed in 7 out of the 47 patients without clinical evidence of the syndrome. The percentage varied between 0% to 35% in the group of non trauma ARDS patients and between 0 to 5% in healthy surgical patients. CONCLUSION: Lipid inclusions in alveolar cells are common during traumatic and non-traumatic respiratory failure. Determination of the percentage of cells recovered by bronchoalveolar lavage and containing fat droplets may contribute to the diagnosis of the fat embolism syndrome in mechanically-ventilated trauma patients with respiratory failure provided that the significant threshold would be 30%.
Assuntos
Líquido da Lavagem Broncoalveolar/citologia , Embolia Gordurosa/diagnóstico , Embolia Pulmonar/diagnóstico , Ferimentos e Lesões/complicações , Adulto , Embolia Gordurosa/etiologia , Feminino , Humanos , Hipóxia/etiologia , Escala de Gravidade do Ferimento , Metabolismo dos Lipídeos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Alvéolos Pulmonares/metabolismo , Embolia Pulmonar/etiologia , Síndrome do Desconforto Respiratório/etiologia , Fatores de TempoRESUMO
OBJECTIVES: To describe the evolution and the diagnostic value of cardiac troponin I (cTnI) and to relate its concentrations with the indicators of injury in trauma patients. DESIGN: Prospective, observational study of 17 young, previously healthy, mechanically-ventilated patients during the early post-traumatic period in the Surgical ICU of a University Hospital. METHODS: Serial measurements of serum cTnI, total creatine kinase activity (CKtot) and its isoenzyme MB (CK-MB) (on admission, 12 h later, then daily for 7 days), clinical data and repeated electrocardiographic (ECG) and transesophageal echocardiographic (TEE) recordings. RESULTS: Rhabdomyolysis was observed in all the patients with a significant relationship between CK-MB and CKtot. Despite the fact that no patient demonstrated ECG or TEE signs of myocardial contusion, elevated serum levels of cTnI were observed in six patients (35%) without obvious dilutional interference. As compared with the others, these patients exhibited a more frequent arterial hypotension (83% vs 18%, p = 0.035), required greater volume expansion on day 1 (22,000 vs 8,500 ml, p = 0.027) and usually demonstrated early (83% vs 9%, p = 0.005) and late (66% vs 9%, p = 0.028) multiple organ dysfunction syndrome. CONCLUSIONS: Taking into account the high reported sensitivity and specificity of cTnI dosage, the present results suggest cTnI can play a role in the evaluation of indirect myocardial injury following traumatic shock.
Assuntos
Traumatismos Cardíacos/sangue , Traumatismos Cardíacos/diagnóstico , Miocárdio/química , Índices de Gravidade do Trauma , Troponina I/sangue , Adulto , Biomarcadores/sangue , Creatina Quinase/sangue , Cuidados Críticos/métodos , Feminino , Traumatismos Cardíacos/enzimologia , Hemodinâmica/fisiologia , Humanos , Isoenzimas , Masculino , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos/sangue , Insuficiência de Múltiplos Órgãos/fisiopatologia , Traumatismo Múltiplo/sangue , Traumatismo Múltiplo/diagnóstico , Traumatismo Múltiplo/fisiopatologia , Prognóstico , Estudos Prospectivos , Análise de Regressão , Rabdomiólise/sangue , Rabdomiólise/diagnóstico , Fatores de TempoRESUMO
OBJECTIVE: To determine the pharmacokinetic parameters of cefpirome, a new so-called fourth-generation cephalosporin, in previously healthy trauma patients with posttraumatic systemic inflammatory response syndrome (SIRS) and to compare them to parameters obtained in matched, healthy volunteers. DESIGN: A prospective study. SETTING: 12-bed surgical intensive care unit in a university hospital. PATIENTS: 9 severe [Injury Severity Score, median (range) 29 (16-50)] trauma patients on mechanical ventilation with proven or suspected cefpirome-susceptible nosocomial infection, with no renal or hepatic failure, and healthy volunteers matched for age (+/- 5 years), sex, and body surface area (+/- 10%) were enrolled. All were men. INTERVENTIONS: Cefpirome (2 g twice daily) was continuously infused over a 0.5 h period alone or concomitantly with ciprofloxacin (400 mg over 1 h, twice daily). MEASUREMENTS AND MAIN RESULTS: Antibiotic concentrations in plasma were measured by high-performance liquid chromatography; their pharmacokinetic parameters were evaluated at 12 time points after the first drug administration using a noncompartmental model. Cefpirome pharmacokinetic parameters for the two groups were similar despite a wider variation for trauma patients. Specifically, the median (range) time during which the cefpirome concentration in plasma remained over 4 mg/l (corresponding to the French lower cutoff determining cefpirome susceptibility) was 9.5 (7- > 12) and 9 (8-12) h for trauma patients and healthy volunteers, respectively. In the group of five patients receiving combined antibiotic therapy, the interindividual variability of pharmacokinetics was wider for ciprofloxacin than for cefpirome. CONCLUSION: No major pharmacokinetic modification was noted when cefpirome was given to trauma patients with posttraumatic SIRS without significant organ failure, indicating that no dosage adjustment seems required in this population. However, larger studies including determination of antibiotic levels in tissues are warranted to confirm these results.