Clinical image of sepsis-associated encephalopathy midst E. coli urosepsis: Emergency department database study.

Background: Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to infection, which, if untreated, leads to multi-organ failure. One of the severe possible complications is sepsis associated encephalopathy (SAE), a neurological dysfunction occurring secondary to a severe inflammatory response. It manifests as acute cognitive dysfunction and sudden-onset dysfunctions in mental state. Uropathogenic Escherichia coli is the most common pathogen causing bacteremia, responsible for 80% of uncomplicated outpatient urinary tract infections and 40% of nosocomial infections. The study aimed to assess the difference in the severity and the course of urosepsis caused by E. coli in patients with and without septic encephalopathy. Materials and methods: This study presents a retrospective analysis of the population of urosepsis patients admitted to the Emergency Department between September 2019 and June 2022. Inflammatory parameters, urinalysis and blood cultures were performed, along with a clinical evaluation of sepsis severity and encephalopathy. The patients were then stratified into SAE and non-SAE groups based on neurological manifestations and compared according to the collected data. Results: A total of 199 septic patients were included in the study. E. coli-induced urosepsis was diagnosed in 84 patients. In this group, SAE was diagnosed in 31 (36.9%) patients (33.3% in males, 40.5% females). Patients with SAE were found to be hypotensive (p < 0,005), with a higher respiratory rate (p < 0,017) resulting in a higher mortality rate (p = 0.002) compared to non-SAE septic patients. The APACHE II score was an independent risk factor associated with a higher mortality rate. Biochemical parameters between the groups did not show any statistical importance related to the severity of urosepsis. Conclusions: The severity of urosepsis and risk of SAE development increase according to the clinical condition and underlying comorbidities. Urosepsis patients with SAE are at a higher risk of death. Patients should undergo more careful screening for the presence of SAE on admission, and more intense monitoring and treatment should be provided for patients with SAE. This study indicates the need to develop projects aiming to further investigate neuroprotective interventions in sepsis.

Determination of urinary androgens in women with polycystic ovary syndrome using LC-QqQ/MS and the application of thin film solid-phase microextraction (TF-SPME).

Hyperandrogenism is one of the most pronounced symptoms of Polycystic Ovary Syndrome (PCOS) and seems to play a key role in the pathogenesis of this complex disorder. Nevertheless, there is still a lack of consistent results regarding common steroid predictors of PCOS. Therefore, a liquid chromatography tandem mass spectrometry (HPLC-QqQ/MS) method was developed and validated to determine the concentrations of four classic androgens: androstenedione (An-dione), testosterone (T), 5α-dihydrotestosterone (DHT) and androsterone (An) in urine samples obtained from women with PCOS and healthy controls. The limits of detection were between 0.04 and 0.09 ng/mL, while the limits of quantification ranged from 0.1 to 0.3 ng/mL respectively. As a pre-treatment procedure prior to analysis, hydrolysis using ß-glucuronidase and thin film solid-phase microextraction (TF-SPME) was applied. The methodology was employed to perform targeted metabolomics of urinary steroids in women with PCOS and healthy controls. All measured androgens: An-dione (p < 0.0001), T (p = 0.0001), DHT (p < 0.0001) and An (p = 0.0002) showed significantly higher concentrations in the urine of women with PCOS. The largest difference in the mean concentration was found for DHT, which was 2.8 times higher in the PCOS group (13.9 ± 14.1 ng/mg creatinine) in comparison to healthy controls (4.9 ± 3.4 ng/mg creatinine). The results of receiver operating characteristic curve indicated that determination of the panel of three urinary androgens: T+DHT+An-dione with, under the study assumptions, was the best predictor of PCOS diagnosis (AUC of ROC curve = 0.91 (95 % CI: 0.8212-0.9905). The application of an LC-MS/MS-based analysis, together with highly sensitive extraction techniques like TF-SPME, is a suitable approach to perform fast assays and obtain reliable results - crucial in the search for valuable and significant steroids predictors of PCOS.

Molecular signature of renal cell carcinoma by means of a multiplatform metabolomics analysis.

Renal cell carcinoma (RCC) is a disease with no specific diagnostic method or treatment. Thus, the evaluation of novel diagnostic tools or treatment possibilities is essential. In this study, a multiplatform untargeted metabolomics analysis of urine was applied to search for a metabolic pattern specific for RCC, which could enable comprehensive assessment of its biochemical background. Thirty patients with diagnosed RCC and 29 healthy volunteers were involved in the first stage of the study. Initially, the utility of the application of the selected approach was checked for RCC with no differentiation for cancer subtypes. In the second stage, this approach was used to study clear cell renal cell carcinoma (ccRCC) in 38 ccRCC patients and 38 healthy volunteers. Three complementary analytical platforms were used: reversed-phase liquid chromatography coupled with time-of-flight mass spectrometry (RP-HPLC-TOF/MS), capillary electrophoresis coupled with time-of-flight mass spectrometry (CE-TOF/MS), and gas chromatography triple quadrupole mass spectrometry (GC-QqQ/MS). As a result of urine sample analyses, two panels of metabolites specific for RCC and ccRCC were selected. Disruptions in amino acid, lipid, purine, and pyrimidine metabolism, the TCA cycle and energetic processes were observed. The most interesting differences were observed for modified nucleosides. This is the first time that the levels of these compounds were found to be changed in RCC and ccRCC patients, providing a framework for further studies. Moreover, the application of the CE-MS technique enabled the determination of statistically significant changes in symmetric dimethylarginine (SDMA) in RCC.

Untargeted Metabolomics Study of Three Matrices: Seminal Fluid, Urine, and Serum to Search the Potential Indicators of Prostate Cancer.

The simultaneous determination of metabolites from biological fluids may provide more accurate information about the current body condition. So far, the metabolomics approach has been successfully applied to study the mechanism of several disorders and to search for novel biomarkers. Urine and plasma are widely accepted matrices for the evaluation of several pathologies, while prostate cancer (CaP) development is still unknown. For this reason, an alternative matrix, the seminal fluid, was proposed to expand the knowledge about the CaP pathomechanism. The main aim of this study was to develop and optimize the sample preparation protocol to ensure the highest coverage of the metabolome of ejaculate samples. Parameters like the type and composition of the solvent mixture, time of extraction, and applied volume of the solvent were tested. The optimized method was applied for the untargeted metabolomics profiling of seminal fluid samples obtained from CaP patients. Moreover, urine and serum samples were also prepared for untargeted metabolomics analysis. Analyses were carried out with the use of two complementary analytical techniques: GC-EI-QqQ/MS and LC-ESI-TOF/MS. Finally, the metabolic signature of seminal fluid (n = 7), urine (n = 7), and plasma (n = 7) samples was compared. Furthermore, the hypothesis of the increased level of metabolites in ejaculate samples related to the CaP development was evaluated. The results indicated that the developed and optimized sample preparation protocol for seminal fluid may be successfully applied for metabolomics study. Untargeted analysis of ejaculate enabled to determine the following classes of compounds: fatty acids, sphingolipids, phospholipids, sugars, and their derivatives, as well as amino acids. Finally, a comparison of the three tested matrices was carried out. To our best knowledge, it is the first time when the metabolic profile of the three matrices, namely, urine, plasma, and seminal fluid, was compared. Based on the results, it can be pointed out that ejaculate comprises the metabolic signature of both matrices (polar compounds characteristic for urine, and non-polar ones present in plasma samples). Compared to plasma, semen samples revealed to have a similar profile; however, determined levels of metabolites were lower in case of ejaculate. In case of urine samples, compared to semen metabolic profiles, the levels of detected metabolites were decreased in the latter ones.

The potential role of fatty acids in prostate cancer determined by GC-MS analysis of formalin-fixed paraffin-embedded tissue samples.

Prostate cancer (PCa) is one of the leading types of cancer in men. Although the diagnosis of this disease is currently quite effective, there is still a need to search for noninvasive diagnostic and monitoring methods. Consequently, identifying the mechanisms underlying the development and progression of PCa is crucial. It has been confirmed that the hallmarks of PCa include changes in metabolism, particularly that of fatty acids. Therefore, the application of lipidomics with an accurate histopathological assessment can provide the necessary information and reveal the metabolites that are characteristic of the disease. The use of formalin-fixed, paraffin-embedded (FFPE) tissue samples as an alternative matrix in retrospective research makes this approach highly innovative. The main goal of this study was to perform an untargeted lipidomic analysis of FFPE PCa tissue samples (n = 52) using gas chromatography coupled with mass spectrometry (GC-MS), in comparison to controls (n = 50). To our knowledge, this study is the first to simultaneously conduct a metabolic analysis and histopathological assessment. In the latter, the samples were evaluated based on Gleason grading score and pTNM stage. The obtained results were evaluated by univariate (Student's t-test or Mann-Whitney U-test) as well as multivariate statistical analysis (principal component analysis, partial least squares-discriminant analysis, variable importance into projection, and selectivity ratio) in order to select the metabolites with the most discriminative power. Additionally, the correlation between the level of metabolites and pathological characteristics was determined. The results of the analyses confirmed the changes in the lipid metabolism pathway in PCa. It can be assumed that PCa is linked with elevated de novo biosynthesis of steroid hormone-related fatty acids and beta-oxidation of fatty acids. An increased level of three fatty acids, namely 9-octadecanoic acid, 9,12-octadecadienoic acid, and 5, 8, 1,14-eicosatetraenoic acid, was observed in the PCa samples. These fatty acids were assigned as metabolites with the best discriminative power for the two tested groups. In practice, these compounds could be considered as specific biochemical factors that may be implemented in the diagnosis of PCa, but their significance should be validated on a more extensive set of samples. Undoubtedly, these results are valuable as they provide important information on prostate cancerogenesis in the context of a metabolic switch.

Metabolomic Signature of Early Vascular Aging (EVA) in Hypertension.

Arterial stiffening is a hallmark of early vascular aging (EVA) syndrome and an independent predictor of cardiovascular morbidity and mortality. In this case-control study we sought to identify plasma metabolites associated with EVA syndrome in the setting of hypertension. An untargeted metabolomic approach was used to identify plasma metabolites in an age-, BMI-, and sex-matched groups of EVA (n = 79) and non-EVA (n = 73) individuals with hypertension. After raw data processing and filtration, 497 putative compounds were characterized, out of which 4 were identified as lysophosphaditylcholines (LPCs) [LPC (18:2), LPC (16:0), LPC (18:0), and LPC (18:1)]. A main finding of this study shows that identified LPCs were independently associated with EVA status. Although LPCs have been shown previously to be positively associated with inflammation and atherosclerosis, we observed that hypertensive individuals characterized by 4 down-regulated LPCs had 3.8 times higher risk of EVA compared to those with higher LPC levels (OR = 3.8, 95% CI 1.7-8.5, P < 0.001). Our results provide new insights into a metabolomic phenotype of vascular aging and warrants further investigation of negative association of LPCs with EVA status. This study suggests that LPCs are potential candidates to be considered for further evaluation and validation as predictors of EVA in patients with hypertension.

Are Alpha-2D Adrenoceptor Subtypes Involved in Rat Mydriasis Evoked by New Imidazoline Derivatives: Marsanidine and 7-Methylmarsanidine?

The imidazoline compounds may produce mydriasis after systemic administration to some species (rats, cats, and mice). In mydriatic activity of imidazolines, α2D-adrenoceptors subtype(s) seems to be involved. In this study, the pupil dilatory effect evoked by 2 newly synthesized imidazoline derivatives-α2-adrenoceptor agonists: marsanidine and 7-methylmarsanidine-was compared. The compounds were tested alone as well as in the presence of α2-adrenoceptor antagonists (nonselective, yohimbine, and selective toward the following α2-adrenoceptor subtypes-α2A-2-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-2,3-dihydro-1-methyl-1H-isoindole maleate (BRL44408), α2B-2-[2-(4-(2-methoxyphenyl)piperazin-1-yl)ethyl]-4,4-dimethyl-1,3-(2H,4H)-isoquinolindione dihydrochloride (ARC239), α2C-JP1302, α2D-2-(2,3-dihydro-2-methoxy-1,4-benzodioxin-2-yl)-4,5-dihydro-1H-imidazole hydrochloride [RX821002]). The agonists were studied in male Wistar rats and were administered intravenously in cumulative doses. The antagonistic compounds were given in a single dose before the experiment with marsanidine or 7-methylmarsanidine. Pupil diameter was measured with stereoscopic microscope equipped in green light filter. Marsanidine and 7-methylmarsanidine exerted marked mydriatic effects. BRL44408, JP1302, and ARC239 did not cause significant parallel shift to the right of the dose-effect curves obtained for both imidazolines. In case of yohimbine and RX821002, the marked parallel shifts of dose-response curves were observed, with the antagonistic effects of RX821002 more pronounced. In vivo pharmacodynamics experiment suggests that α2D-adrenoceptor subtype is mainly engaged in mydriatic effects evoked in rats by imidazoline derivatives, in particular by clonidine.

Comparative pharmacodynamic analysis of imidazoline compounds using rat model of ocular mydriasis with a test of quantitative structure-activity relationships.

Imidazol(in)e derivatives, having the chemical structure similar to clonidine, exert diverse pharmacological activities connected with their interactions with alpha2-adrenergic receptors, e.g. hypotension, bradycardia, sedation as well as antinociceptive, anxiolytic, antiarrhythmic, muscle relaxant and mydriatic effects. The mechanism of pupillary dilation observed after systemic administration of imidazol(in)es to rats, mice and cats depends on the stimulation of postsynaptic alpha2-adrenoceptors within the brain. It was proved that the central nervous system (CNS)-localized I1-imidazoline receptors are not engaged in those effects. It appeared interesting to analyze the CNS-mediated pharmacodynamics of imidazole(in)e agents in terms of their chromatographic and calculation chemistry-derived parameters. In the present study a systematic determination and comparative pharmacometric analysis of mydriatic effects in rats were performed on a series of 20 imidazol(in)e agents, composed of the well-known drugs and of the substances used in experimental pharmacology. The eye pupil dilatory activities of the compounds were assessed in anesthetized Wistar rats according to the established Koss method. Among twenty imidazol(in)e derivatives studied, 18 produced diverse dose-dependent mydriatic effects. In the quantitative structure-activity relationships (QSAR) analysis, the pharmacological data (half maximum mydriatic effect - ED50 in µmol/kg) were considered along with the structural parameters of the agents from molecular modeling. The theoretically calculated lipophilicity parameters, CLOGP, of imidazol(in)es, as well as their lipophilicity parameters from HPLC, logkw, were also considered. The attempts to derive statistically significant QSAR equations for a full series of the agents under study were unsuccessful. However, for a subgroup of eight apparently structurally related imidazol(in)es a significant relationship between log(1/ED50) and logkw values was obtained. The lack of "predictive" QSAR for the whole series of the structurally diverse agents is probably due to a complex mechanism of the ligand-alpha2-adrenergic receptor interactions, which are predominantly of a highly structurally specific polar nature. Such interactions are difficult to quantify with the established chemical structural descriptors, contrary to the less specific, molecular bulkiness-related interactions.

Selective determination of cocaine and its metabolite benzoylecgonine in environmental samples by newly developed sorbent materials.

Sewage epidemiology, as compared to crime statistics, health, medical reports or population surveys, is becoming the most objective and realistic approach to estimate drug consumption and trends in local communities. In this study we proposed newly synthesized sorbent materials for selective extraction of cocaine and benzoylecgonine from wastewater samples. The molecular modeling calculations were conducted to provide the choice of proper template and functional monomer for synthesis of extraction materials. The physicochemical properties of synthesized sorbents were studied using various techniques. The newly developed sorbent materials were applied for selective extraction of cocaine and benzoylecgonine from wastewater samples collected from different wastewater treatment plants in Poland. The obtained recoveries values in wastewater samples were 83.6(±7.1)% and 72.1(±4.8)%, for cocaine and benzoylecgonine, respectively. The newly developed sorbents comprise an alternative to conventional ones, which are not entirely suitable for highly efficient purification of environmental samples due to the presence of contaminants.

New sorbent materials for selective extraction of cocaine and benzoylecgonine from human urine samples.

An increase in cocaine consumption has been observed in Europe during the last decade. Benzoylecgonine, as a main urinary metabolite of cocaine in human, is so far the most reliable marker of cocaine consumption. Determination of cocaine and its metabolite in complex biological samples as urine or blood, requires efficient and selective sample pretreatment. In this preliminary study, the newly synthesized sorbent materials were proposed for selective extraction of cocaine and benzoylecgonine from urine samples. Application of these sorbent media allowed to determine cocaine and benzoylecgonine in urine samples at the concentration level of 100ng/ml with good recovery values as 81.7%±6.6 and 73.8%±4.2, respectively. The newly synthesized materials provided efficient, inexpensive and selective extraction of both cocaine and benzoylecgonine from urine samples, which can consequently lead to an increase of the sensitivity of the current available screening diagnostic tests.

Identification of lipid fraction constituents from grasshopper (Chorthippus spp.) abdominal secretion with potential activity in wound healing with the use of GC-MS/MS technique.

In recent years biologically active compounds isolated from insects call special interest of drug researchers. According to some Polish etnopharmacological observations, secretion from the grasshopper's abdomen (Orthoptera family) is believed to speed up the process of wound healing. In the present work we focused on determination of main components of the lipid fraction of material from grasshopper abdomen using GC-MS/MS. Samples were qualitatively analyzed using gas chromatography coupled with mass spectrometry. Both liquid-liquid extraction and solid-phase extraction pretreatment methods were used to concentrate and fractionate the compounds from the insect. In the derivatized fractions ca. 350 compounds were identified, including substances of known biological activity. The potential agents affecting wound healing have been indicated. A set of compounds characteristic for all the studied Chorthippus spp., have been identified. Data analysis revealed different lipidomic profiles of grasshoppers depending on the insects origin and collection area.

Mydriasis model in rats as a simple system to evaluate α2-adrenergic activity of the imidazol(in)e compounds.

Imidazol(in)e compounds show the diversity of pharmacological effects including mydriasis, hypotension, sedation, bradycardia and hypothermia. At first it was postulated that these effects are mediated via α2-adrenoceptors exclusively. Clonidine is well known as a model agent to produce pupillary dilation in rats. However, it became obvious later that clonidine-like imidazol(in)e adrenoceptor agonists which produced mydriasis in rats, exhibit also a high affinity for imidazoline I1-receptors. That short report attempts to review the present status of studies to confirm that the mydriasis model in rats can be a selective system to evaluate the α2-adrenergic activity of potential pharmacologically active compounds of imidazol(in)e structure.