The rapid detection of procalcitonin in septic serum using immunoaffinity MALDI chips.

BACKGROUND: Sepsis is a common worldwide health condition with high mortality. It is caused by a dysregulated immune response to the pathogen. Severe infections resulting in sepsis can be also determined by monitoring several bloodstream biomarkers, one of them being pro-hormone procalcitonin (PCT). PCT concentration in the bloodstream correlates well with sepsis and in severe cases increases up to a thousand times from the healthy physiological values in a short time. In this study, we developed a rapid technique for PCT detection by MALDI-TOF mass spectrometry, that uses in-situ enrichment directly on the specialized immuno MALDI chips that are utilized as MALDI plates. The method's ability to detect PCT was confirmed by comparing the results with LC-MS bottom-up workflow. The new method detects intact PCT by its m/z and uncovers its alternations in septic serum. METHODS: The MALDI chips used for the detection of PCT were prepared by ambient ion soft landing of anti-PCT antibody on an ITO glass slide. The chips were used for the development of the rapid MALDI-TOF MS method. A parallel method based on affinity enrichment on magnetic beads followed by LC-MS/MS data-dependent peptide microsequencing was used to prove PCT presence in the sample. All samples were also tested by ELISA to determine PCT concentration prior to analyzing them by mass spectrometry methods. RESULTS: The MALDI chip method was optimized using recombinant PCT spiked into the human serum. The PCT detection limit was 10 ng/mL. The optimized method was used to analyze 13 sera from patients suffering sepsis. The PCT results were confirmed by LC-MS/MS. The measurement of the intact PCT by the MALDI chip method revealed that sera of patients with severe sepsis have other forms of PCT present, which show post-processing of the primary sequence by cleavage of PCT, resulting in the formation of N and C termini fragments. CONCLUSIONS: Procalcitonin from human serum was successfully enriched and detected using immunoaffinity MALDI chips. The intact PCT was characterized in 13 septic patients. The method is more specific compared to non-MS-based immunoaffinity techniques and allows observation of different variants of PCT in septic patients.

Critical care for multimorbid patients.

Multimorbidity - the simultaneous presence of several chronic diseases - is very common in the critically ill patients. Its prevalence is roughly 40-85 % and continues to increase further. Certain chronic diseases such as diabetes, obesity, chronic heart, pulmonary, liver or kidney disease and malignancy are associated with higher risk of developing serious acute complications and therefore the possible need for intensive care. This review summarizes and discusses selected specifics of critical care for multimorbid patients.

Medical nutrition therapy and clinical outcomes in critically ill adults: a European multinational, prospective observational cohort study (EuroPN).

BACKGROUND: Medical nutrition therapy may be associated with clinical outcomes in critically ill patients with prolonged intensive care unit (ICU) stay. We wanted to assess nutrition practices in European intensive care units (ICU) and their importance for clinical outcomes. METHODS: Prospective multinational cohort study in patients staying in ICU ≥ 5 days with outcome recorded until day 90. Macronutrient intake from enteral and parenteral nutrition and non-nutritional sources during the first 15 days after ICU admission was compared with targets recommended by ESPEN guidelines. We modeled associations between three categories of daily calorie and protein intake (low: < 10 kcal/kg, < 0.8 g/kg; moderate: 10-20 kcal/kg, 0.8-1.2 g/kg, high: > 20 kcal/kg; > 1.2 g/kg) and the time-varying hazard rates of 90-day mortality or successful weaning from invasive mechanical ventilation (IMV). RESULTS: A total of 1172 patients with median [Q1;Q3] APACHE II score of 18.5 [13.0;26.0] were included, and 24% died within 90 days. Median length of ICU stay was 10.0 [7.0;16.0] days, and 74% of patients could be weaned from invasive mechanical ventilation. Patients reached on average 83% [59;107] and 65% [41;91] of ESPEN calorie and protein recommended targets, respectively. Whereas specific reasons for ICU admission (especially respiratory diseases requiring IMV) were associated with higher intakes (estimate 2.43 [95% CI: 1.60;3.25] for calorie intake, 0.14 [0.09;0.20] for protein intake), a lack of nutrition on the preceding day was associated with lower calorie and protein intakes (- 2.74 [- 3.28; - 2.21] and - 0.12 [- 0.15; - 0.09], respectively). Compared to a lower intake, a daily moderate intake was associated with higher probability of successful weaning (for calories: maximum HR 4.59 [95% CI: 1.5;14.09] on day 12; for protein: maximum HR 2.60 [1.09;6.23] on day 12), and with a lower hazard of death (for calories only: minimum HR 0.15, [0.05;0.39] on day 19). There was no evidence that a high calorie or protein intake was associated with further outcome improvements. CONCLUSIONS: Calorie intake was mainly provided according to the targets recommended by the active ESPEN guideline, but protein intake was lower. In patients staying in ICU ≥ 5 days, early moderate daily calorie and protein intakes were associated with improved clinical outcomes. Trial registration NCT04143503 , registered on October 25, 2019.

Characterization of esophageal motor activity, gastroesophageal reflux, and evaluation of prokinetic effectiveness in mechanically ventilated critically ill patients: a high-resolution impedance manometry study.

BACKGROUND: Motility disorders of upper gastrointestinal tract are common in critical illness and associated with significant clinical consequences. However, detailed quantitative and qualitative analyses of esophageal motor functions are lacking. Therefore, we aimed to characterize the key features of esophageal motility functions using high-resolution impedance manometry (HRIM) and to evaluate an objective link between esophageal motor patterns, gastric emptying, and gastroesophageal reflux. We also studied the prokinetic effects of metoclopramide. METHODS: We prospectively performed HRIM for 16 critically ill hemodynamically stable patients. Patients were included if they had low gastric volume (LGV; < 100 mL/24 h, n = 8) or high gastric volume (HGV; > 500 mL/24 h, n = 8). The HRIM data were collected for 5 h with intravenous metoclopramide administration (10 mg) after the first 2 h. RESULTS: The findings were grossly abnormal for all critically ill patients. The esophageal contraction vigor was markedly increased, indicating prevailing hypercontractile esophagus. Ineffective propulsive force was observed for 73% of esophageal activities. Panesophageal pressurization was the most common pressurization pattern (64%). Gastroesophageal reflux predominantly occurred with transient lower esophageal sphincter relaxation. The common features of the LGV group were a hyperreactive pattern, esophagogastric outflow obstruction, and frequent reflux. Ineffective motility with reduced lower esophageal sphincter tone, and paradoxically fewer reflux episodes, was common in the HGV group. Metoclopramide administration reduced the number of esophageal activities but did not affect the number of reflux episodes in either group. CONCLUSION: All critically ill patients had major esophageal motility abnormalities, and motility patterns varied according to gastric emptying status. Well-preserved gastric emptying and maintained esophagogastric barrier functions did not eliminate reflux. Metoclopramide failed to reduce the number of reflux episodes regardless of gastric emptying status. Trial registration ISRCTN, ISRCTN14399966. Registered 3.9.2020, retrospectively registered. https://www.isrctn.com/ISRCTN14399966 .

Vagus Nerve Stimulation Attenuates Multiple Organ Dysfunction in Resuscitated Porcine Progressive Sepsis.

OBJECTIVES: To investigate the potential benefits of vagus nerve stimulation in a clinically-relevant large animal model of progressive sepsis. DESIGN: Prospective, controlled, randomized trial. SETTING: University animal research laboratory. SUBJECTS: Twenty-five domestic pigs were divided into three groups: 1) sepsis group (eight pigs), 2) sepsis + vagus nerve stimulation group (nine pigs), and 3) control sham group (eight pigs). INTERVENTIONS: Sepsis was induced by cultivated autologous feces inoculation in anesthetized, mechanically ventilated, and surgically instrumented pigs and followed for 24 hours. Electrical stimulation of the cervical vagus nerve was initiated 6 hours after the induction of peritonitis and maintained throughout the experiment. MEASUREMENTS AND MAIN RESULTS: Measurements of hemodynamics, electrocardiography, biochemistry, blood gases, cytokines, and blood cells were collected at baseline (just before peritonitis induction) and at the end of the in vivo experiment (24 hr after peritonitis induction). Subsequent in vitro analyses addressed cardiac contractility and calcium handling in isolated tissues and myocytes and analyzed mitochondrial function by ultrasensitive oxygraphy. Vagus nerve stimulation partially or completely prevented the development of hyperlactatemia, hyperdynamic circulation, cellular myocardial depression, shift in sympathovagal balance toward sympathetic dominance, and cardiac mitochondrial dysfunction, and reduced the number of activated monocytes. Sequential Organ Failure Assessment scores and vasopressor requirements significantly decreased after vagus nerve stimulation. CONCLUSIONS: In a clinically-relevant large animal model of progressive sepsis, vagus nerve stimulation was associated with a number of beneficial effects that resulted in significantly attenuated multiple organ dysfunction and reduced vasopressor and fluid resuscitation requirements. This suggests that vagus nerve stimulation might provide a significant therapeutic potential that warrants further thorough investigation.

Proteins adsorbed to a polysulfone hemodialysis membrane under heparin and citrate anticoagulation regimens.

The study aim was to compare molecular-level effects (blood-dialyzer interactions) of heparin and citrate anticoagulation using proteome-wide analysis of biofilm adsorbed to dialysis membrane. Ten patients receiving maintenance hemodialysis were examined in a crossover design under three different anticoagulation regimens, namely citrate, heparin, and anticoagulation-free (control). Following a regular hemodialysis session (4 hours, polysulfone membrane), dialyzers were flushed and the surface biofilm eluted by acetic acid. Protein composition of the eluates was determined by 2-dimensional gel electrophoresis and resulting patterns compared between regimens. Proteins responsible for the difference were identified by mass spectrometry. Citrate anticoagulation was associated with significantly less protein adsorption to the membrane than heparin (2.2 [1.1-2.9] mg vs. 6.5 [2.9-11.6] mg, P = 0.009). Among the proteins identified as major discriminators between citrate and the other regimens, fibrin α-chain fragments of molecular weight below 40 kDa prevailed. In these fragments, an analysis of the amino acid sequence has been performed by comparison with the UniProt database. It showed missing α-chain cross-links. On the contrary, heparin prevented adsorption and cleavage of several heparin-binding proteins; especially complement factor H-related protein 3, insulin-like growth factor binding proteins (2, 4, and 5), and chemerin. Compared to heparin, citrate is associated with less protein adsorption and imperfectly crosslinked fibrin clot formation. Membrane adsorptive properties are significantly modified by the anticoagulation regimen.

Oncologic emergencies.

Oncologic emergencies and life-threatening cancer-related and treatment-related complications are the net effect of gradually increasing incidence of malignant diseases, improvement of therapeutic options and survival of oncologic patients. These complications are relatively specific for such population of patients and they are quite rare within the individuals without malignancy. Selected oncological emergencies are discussed in this review.

Sepsis mimics.

Number of identical pathophysiological mechanisms is shared by sepsis and other clinical conditions and diseases. This could lead to their nearly similar clinical phenotype. However, the early discrimination of them is crucial - treatment of particular diseases differs significantly, and the mortality of the vast majority of them is considerable. The differential diagnostics possibilities together with brief description of selected clinical conditions are discussed within the review.

Extracorporeal removal techniques in toxicology: part 1.

Supporting clearance of a toxic substance by an extracorporeal removal technique is one of the advanced treatment methods applied in poisoned patient management. General indications stem from toxicokinetics of the poison while individual indications are determined by poisoning severity. The first part of this review deals in detail with particular options of extracorporeal treatment in toxicology and also with its specific application when treating lithium and salicylates poisoning or dabigatran overdose. The aim of this review is to facilitate the clinicians and nephrologists decision making whether to indicate this invasive procedure, to communicate and summarize the existing recommendations and to highlight the most important ways of how to treat poisoning by specific toxic substances.

Extracorporeal removal techniques in toxicology: part 2.

The second part of the review deals in detail with the diagnostics and treatment of toxic alcohols poisoning and management and indication of extracorporeal removal techniques in intoxication with other drugs, theophylline, valproic acid, metformin and metformin associated lactic acidosis, respectively. The extracorporeal treatment enhances the clearance of the toxin and corrects patients metabolic disturbances as well. It is necessary to use this treatment in severe intoxications. Indication of this invasive procedure falls within clinicians and nephrologists competence being advised by a toxicologist. This review could help make fast decisions.

Aortic dissection and other acute aortic syndromes in the emergency department.

Acute aortic syndromes are emergent life-threatening conditions affecting the aorta, which actual incidence is difficult to determine. Mortality of untreated patients increases steadily over time, so early diagnosis and initiation of therapy are crucial. Management of patients in Czech Republic follow, similar as in other European countries, the European Society of Cardiology guidelines from 2014, which were updated in 2018. The basis for diagnosis consists of history, physical examination, ECG, determination of vital signs, hemodynamic status and stratification of dia-gnosis probability by ADD-RS (aortic dissection detection risk score). This is followed by a series of laboratory and imaging examinations, of which the D-dimer, CT aortography and echocardiography are the most important. Recent studies show the benefit of combination of ADD-RS with D-dimer or measurement of ascendant aorta diameter by echocardiography. New emerging biomarkers are currently under investigation. Thanks to advances in technology, magnetic resonance imaging could take place as emergent diagnostic tool in the future. Initial therapy depends on the hemodynamic status of the patient. It must be followed by definitive therapy. In this publication we summarize the approach to a patient with acute aortic syndrome in the emergency department focusing on aortic dissection as its most common type.

Nontuberculous mycobacterial disease: a case report-based review.

Nontuberculous mycobacterial infections are rare diseases. However, as number of immunocompromised patients is growing and modern diagnostic tools are available, both the importance and incidence of nontuberculous myco-bacterial infections are gaining clinical importance. Based on a clinical case, this article briefly summarizes the cur-rent knowledge on this issue.

Intravenous fluid therapy in acutely ill patients for non-intensivists.

Intravenous fluid therapy is the most frequent therapeutic intervention in acutely hospitalized patients. They are administered in order to resuscitate the circulation in hypovolemia-associated shock states, to compensate for an impending or existing fluid extracellular deficit, or as a maintenance infusion if the patient is incapable of taking fluid by other means. Any fluid should be prescribed with the same caution as with any other drug. Errors in fluid therapy adversely affect patient - centered outcome. This may be the result of an incorrectly selected amount or inappropriate fluid composition for a given clinical situation. Prescribing intravenous fluids is a complex process involving a decision on the type, composition, dose, rate and possible toxicity of the particular solution. Balanced crystalloid solutions are the first choice for most acute conditions. The need for fluids dynamically changes over time in acutely ill patients. Uncontrolled cumulative positive balance is associated with substantial morbidity and mortality.

Effects of Hyperoxia During Resuscitation From Hemorrhagic Shock in Swine With Preexisting Coronary Artery Disease.

OBJECTIVES: Investigation of the effects of hyperoxia during resuscitation from hemorrhagic shock in swine with preexisting coronary artery disease. DESIGN: Prospective, controlled, randomized trial. SETTING: University animal research laboratory. SUBJECTS: Nineteen hypercholesterolemic pigs with preexisting coronary artery disease. INTERVENTIONS: Anesthetized, mechanically ventilated, and surgically instrumented pigs underwent 3 hours of hemorrhagic shock (removal of 30% of the calculated blood volume and subsequent titration of mean arterial blood pressure ≈40 mm Hg). Postshock resuscitation (48 hr) comprised retransfusion of shed blood, crystalloids (balanced electrolyte solution), and norepinephrine support. Pigs were randomly assigned to "control" (FIO2 0.3, adjusted for arterial oxygen saturation ≥ 90%) and "hyperoxia" (FIO2 1.0 for 24 hr) groups. MEASUREMENTS AND MAIN RESULTS: Before, at the end of shock and every 12 hours of resuscitation, datasets comprising hemodynamics, calorimetry, blood gases, cytokines, and cardiac and renal function were recorded. Postmortem, organs were sampled for immunohistochemistry, western blotting, and mitochondrial high-resolution respirometry. Survival rates were 50% and 89% in the control and hyperoxia groups, respectively (p = 0.077). Apart from higher relaxation constant τ at 24 hours, hyperoxia did not affect cardiac function. However, troponin values were lower (2.2 [0.9-6.2] vs 6.9 [4.8-9.8] ng/mL; p < 0.05) at the end of the experiment. Furthermore, hyperoxia decreased cardiac 3-nitrotyrosine formation and increased inducible nitric oxide synthase expression. Plasma creatinine values were lower in the hyperoxia group during resuscitation coinciding with significantly improved renal mitochondrial respiratory capacity and lower 3-nitrotyrosine formation. CONCLUSIONS: Hyperoxia during resuscitation from hemorrhagic shock in swine with preexisting coronary artery disease reduced renal dysfunction and cardiac injury, potentially resulting in improved survival, most likely due to increased mitochondrial respiratory capacity and decreased oxidative and nitrosative stress. Compared with our previous study, the present results suggest a higher benefit of hyperoxia in comorbid swine due to an increased susceptibility to hemorrhagic shock.

Molecular differences in susceptibility of the kidney to sepsis-induced kidney injury.

BACKGROUND: Septic acute kidney injury affects 40-50% of all septic patients. Molecular differences between septic patients with and without acute kidney injury (AKI) are only poorly understood. Here, we investigated gene expression changes that differentiated the subjects who developed septic AKI from those who did not and coupled this approach with traditional parameters of renal physiology. METHODS: In 15 anesthetized, mechanically ventilated and instrumented pigs, progressive sepsis was induced either by peritonitis or by continuous intravenous infusion of Pseudomonas aeruginosa. Animals received standard intensive care including goal-directed hemodynamic management. Analyses were performed on kidneys from sham operated animals, septic pigs without AKI, and pigs with septic AKI. Before, and at 12, 18 and 22 h of progressive sepsis, systemic and renal hemodynamics, cortex microcirculation and plasma IL-6 and TNF-α were measured. At 22 h whole kidney expression of pre-selected genes was analyzed by quantitative Real Time PCR. RESULTS: Animals with septic AKI had systemic hemodynamic phenotype (normo- or hyperdynamic) comparable with non-AKI subjects, but demonstrated higher plasma levels of cytokines, an increase in renal vascular resistance and early fall in cortical microcirculatory blood flow. The genes whose expression discriminated septic AKI from non-AKI included Toll like receptor 4 (up-regulated 2.7-fold, P = 0.04); Cyclooxygenase-2 (up-regulated 14.6-fold, P = 0.01), Angiotensin II Receptor (up-regulated 8.1-fold, P = 0.01), Caspase 3 (up-regulated 5.1-fold, P = 0.02), Peroxisome Proliferator-Activated Receptor Gamma, Coactivator 1 Alpha (down-regulated 2-fold, P = 0.02). CONCLUSIONS: In this preliminary experimental study, kidney gene expression was profoundly different in animals that developed septic AKI as opposed to septic animals that did not. The biological functions of the genes differentially expressed support a role of inflammatory overstimulation coupled with metabolic and apoptotic molecular responses in early septic AKI. Cyclooxygenase-2 and angiotensin type 2 receptor-dependent downstream mechanisms appear fruitful targets for future mechanistic research.

Early urinary biomarkers of diabetic nephropathy in type 1 diabetes mellitus show involvement of kallikrein-kinin system.

BACKGROUND: Additional urinary biomarkers for diabetic nephropathy (DN) are needed, providing early and reliable diagnosis and new insights into its mechanisms. Rigorous selection criteria and homogeneous study population may improve reproducibility of the proteomic approach. METHODS: Long-term type 1 diabetes patients without metabolic comorbidities were included, 11 with sustained microalbuminuria (MA) and 14 without MA (nMA). Morning urine proteins were precipitated and resolved by 2D electrophoresis. Principal component analysis (PCA) and Projection to latent structures discriminatory analysis (PLS-DA) were adopted to assess general data validity, to pick protein fractions for identification with mass spectrometry (MS), and to test predictive value of the resulting model. RESULTS: Proteins (n = 113) detected in more than 90% patients were considered representative. Unsupervised PCA showed excellent natural data clustering without outliers. Protein spots reaching Variable Importance in Projection score above 1 in PLS (n = 42) were subjected to MS, yielding 33 positive identifications. The PLS model rebuilt with these proteins achieved accurate classification of all patients (R2X = 0.553, R2Y = 0.953, Q2 = 0.947). Thus, multiple earlier recognized biomarkers of DN were confirmed and several putative new biomarkers suggested. Among them, the highest significance was met in kininogen-1. Its activation products detected in nMA patients exceeded by an order of magnitude the amount found in MA patients. CONCLUSIONS: Reducing metabolic complexity of the diseased and control groups by meticulous patients' selection allows to focus the biomarker search in DN. Suggested new biomarkers, particularly kininogen fragments, exhibit the highest degree of correlation with MA and substantiate validation in larger and more varied cohorts.

Renal Hemodynamics in AKI: In Search of New Treatment Targets.

Novel therapeutic interventions are required to prevent or treat AKI. To expedite progress in this regard, a consensus conference held by the Acute Dialysis Quality Initiative was convened in April of 2014 to develop recommendations for research priorities and future directions. Here, we highlight the concepts related to renal hemodynamics in AKI that are likely to reveal new treatment targets on investigation. Overall, we must better understand the interactions between systemic, total renal, and glomerular hemodynamics, including the role of tubuloglomerular feedback. Furthermore, the net consequences of therapeutic maneuvers aimed at restoring glomerular filtration need to be examined in relation to the nature, magnitude, and duration of the insult. Additionally, microvascular blood flow heterogeneity in AKI is now recognized as a common occurrence; timely interventions to preserve the renal microcirculatory flow may interrupt the downward spiral of injury toward progressive kidney failure and should, therefore, be investigated. Finally, development of techniques that permit an integrative physiologic approach, including direct visualization of renal microvasculature and measurement of oxygen kinetics and mitochondrial function in intact tissue in all nephron segments, may provide new insights into how the kidney responds to various injurious stimuli and allow evaluation of new therapeutic strategies.

Effects of Hyperoxia and Mild Therapeutic Hypothermia During Resuscitation From Porcine Hemorrhagic Shock.

OBJECTIVE: Hemorrhagic shock-induced tissue hypoxia induces hyperinflammation, ultimately causing multiple organ failure. Hyperoxia and hypothermia can attenuate tissue hypoxia due to increased oxygen supply and decreased demand, respectively. Therefore, we tested the hypothesis whether mild therapeutic hypothermia and hyperoxia would attenuate postshock hyperinflammation and thereby organ dysfunction. DESIGN: Prospective, controlled, randomized study. SETTING: University animal research laboratory. SUBJECTS: Thirty-six Bretoncelles-Meishan-Willebrand pigs of either gender. INTERVENTIONS: After 4 hours of hemorrhagic shock (removal of 30% of the blood volume, subsequent titration of mean arterial pressure at 35 mm Hg), anesthetized and instrumented pigs were randomly assigned to "control" (standard resuscitation: retransfusion of shed blood, fluid resuscitation, norepinephrine titrated to maintain mean arterial pressure at preshock values, mechanical ventilation titrated to maintain arterial oxygen saturation > 90%), "hyperoxia" (standard resuscitation, but FIO2, 1.0), "hypothermia" (standard resuscitation, but core temperature 34°C), or "combi" (hyperoxia plus hypothermia) (n = 9 each). MEASUREMENTS AND MAIN RESULTS: Before, immediately at the end of and 12 and 22 hours after hemorrhagic shock, we measured hemodynamics, blood gases, acid-base status, metabolism, organ function, cytokine production, and coagulation. Postmortem kidney specimen were taken for histological evaluation, immunohistochemistry (nitrotyrosine, cystathionine γ-lyase, activated caspase-3, and extravascular albumin), and immunoblotting (nuclear factor-κB, hypoxia-inducible factor-1α, heme oxygenase-1, inducible nitric oxide synthase, B-cell lymphoma-extra large, and protein expression of the endogenous nuclear factor-κB inhibitor). Although hyperoxia alone attenuated the postshock hyperinflammation and thereby tended to improve visceral organ function, hypothermia and combi treatment had no beneficial effect. CONCLUSIONS: During resuscitation from near-lethal hemorrhagic shock, hyperoxia attenuated hyperinflammation, and thereby showed a favorable trend toward improved organ function. The lacking efficacy of hypothermia was most likely due to more pronounced barrier dysfunction with vascular leakage-induced circulatory failure.

Mitochondrial proteomes of porcine kidney cortex and medulla: foundation for translational proteomics.

BACKGROUND: Emerging evidence has linked mitochondrial dysfunction to the pathogenesis of many renal disorders, including acute kidney injury, sepsis and even chronic kidney disease. Proteomics is a powerful tool in elucidating the role of mitochondria in renal pathologies. Since the pig is increasingly recognized as a major mammalian model for translational research, the lack of physiological proteome data of large mammals prompted us to examine renal mitochondrial proteome in porcine kidney cortex and medulla METHODS: Kidneys were obtained from six healthy pigs. Mitochondria from cortex and medulla were isolated using differential centrifugation and proteome maps of cortical and medullar mitochondria were constructed using two-dimensional gel electrophoresis (2DE). Protein spots with significant difference between mitochondrial fraction of renal cortex and medulla were identified by mass spectrometry. RESULTS: Proteomic analysis identified 81 protein spots. Of these spots, 41 mitochondrial proteins were statistically different between renal cortex and medulla (p < 0.05). Protein spots containing enzymes of beta oxidation, amino acid metabolism, and gluconeogenesis were predominant in kidney cortex mitochondria. Spots containing tricarboxylic acid cycle enzymes and electron transport system proteins, proteins maintaining metabolite transport and mitochondrial translation were more abundant in medullar mitochondria. CONCLUSION: This study provides the first proteomic profile of porcine kidney cortex and medullar mitochondrial proteome. Different protein expression pattern reflects divergent functional metabolic role of mitochondria in various kidney compartments. Our study could serve as a useful reference for further porcine experiments investigating renal mitochondrial physiology under various pathological states.

[Sepsis - how to recognize and what to focus on - back to basics in the light of the new definition]. / Sepse - jak ji poznat, na co se zamerit aneb zpet k základum ve svetle nové definice.

UNLABELLED: Sepsis is the primary cause of death from infection. However, its early recognition remains a fundamental challenge in clinical practice. In February 2016, a newly revised sepsis definition has been published (SEPSIS-3). Sepsis has been redefined as life-threatening organ dysfunction caused by a dysregulated host response to infection. In this article, we introduce the updated definition of sepsis, discuss its pros and cons and suggest practical implications. The emphasis is put on basic and comprehensive clinical assessment. KEY WORDS: definition - early recognition of sepsis - infection - sepsis - septic shock.