Profiling of Carnitine Shuttle System Intermediates in Gliomas Using Solid-Phase Microextraction (SPME).

Alterations in the carnitine shuttle system may be an indication of the presence of cancer. As such, in-depth analyses of this pathway in different malignant tumors could be important for the detection and treatment of this disease. The current study aims to assess the profiles of carnitine and acylcarnitines in gliomas with respect to their grade, the presence of isocitrate dehydrogenase (IDH) mutations, and 1p/19q co-deletion. Brain tumors obtained from 19 patients were sampled on-site using solid-phase microextraction (SPME) immediately following excision. Analytes were desorbed and then analyzed via liquid chromatography-high-resolution mass spectrometry. The results showed that SPME enabled the extraction of carnitine and 22 acylcarnitines. An analysis of the correlation factor revealed the presence of two separate clusters: short-chain and long-chain carnitine esters. Slightly higher carnitine and acylcarnitine concentrations were observed in the higher-malignancy tumor samples (high vs. low grade) and in those samples with worse projected clinical outcomes (without vs. with IDH mutation; without vs. with 1p/19q co-deletion). Thus, the proposed chemical biopsy approach offers a simple solution for on-site sampling that enables sample preservation, thus supporting comprehensive multi-method analyses.

Characteristics of selected peripheral blood parameters in polar fox (Alopex lagopus L.) fed diets with inulin.

This study aimed at investigating changes in selected peripheral blood parameters in male polar foxes fed diets with different supplementation of inulin: 0.25% (group El), 0.5% (E2) and 1% (E3). The blood for analysis was sampled from the brachial vein. The study showed that adding 0.25 and 0.5% of inulin to fox feed resulted in a lower content of haemoglobin (Hb) as well as mean mass of Hb in red blood cells in the 0.5% inulin group. The total count of thrombocytes decreased significantly with a higher level of prebiotic, while the total number of white blood cells and the percentage of different leukocytes tested remained invariable. The lowest supplementation of inulin affected the partial pressure of carbon dioxide, however, the remaining acid-base parameters did not change. The present study provides the first preliminary information about the effect of dietary inulin on some haematological indices and acid-base parameters in adult polar foxes. The results may be helpful in practice to improve the health condition of farmed polar foxes.

Changes in blood chemistry in broiler chickens during the fattening period.

The aim of the study was to determine the changes in the selected biochemical serum parameters in male Ross 308 broilers during the fattening period. The birds were kept under standard farm conditions and they were fed on commercial mixtures. The blood for analysis was taken from the jugular vein on the 14th, 21st, and 42nd days of age. The concentration of serum protein (total protein, albumins, uric acid, creatinine), lipid (TG, TCHL, HDL) and mineral (Ca, P(i), Mg, Fe) indices were determined. The measurements were carried out with the use of Epoll 20 photometer. The content of LDL and VLDL lipoprotein fractions was calculated on the basis of the Friedewald equation. Most of the estimated parameters, except for LDL and Pi, were age-dependent (P<0.05). Total protein, albumins and total Ca levels showed a constant increase between the 14th and 42nd days of life. A lower (P<0.05) concentration of TG, TCHL, HDL, VLDL, Mg and Fe was determined at the end of the fattening period compared to 14-day-old broilers. A significant decrease of TG, VLDL, Mg and Fe content was noted already in the first age range (days 14-21) while in the case of TCHL and HDL a significant decrease was found between the 21st and 42nd days of fattening. The obtained results may be helpful in the evaluation of changes in the metabolic profile, health condition and production patterns in growing broiler chickens reared under farm conditions.

One extraction tool for in vitro-in vivo extrapolation? SPME-based metabolomics of in vitro 2D, 3D, and in vivo mouse melanoma models.

Solid phase microextraction (SPME) in combination with high-resolution mass spectrometry was employed for the determination of metabolomic profile of mouse melanoma growth within in vitro 2D, in vitro 3D, and in vivo models. Such multi-model approach had never been investigated before. Due to the low-invasiveness of SPME, it was possible to perform time-course analysis, which allowed building time profile of biochemical reactions in the studied material. Such approach does not require the multiplication of samples as subsequent analyses are performed from the very same cell culture or from the same individual. SPME already reduces the number of animals required for experiment; therefore, it is with good concordance with the 3Rs rule (replacement, reduction, and refinement). Among tested models, the largest number of compounds was found within the in vitro 2D cell culture model, while in vivo and in vitro 3D models had the lowest number of detected compounds. These results may be connected with a higher metabolic rate, as well as lower integrity of the in vitro 2D model compared to the in vitro 3D model resulting in a lower number of compounds released into medium in the latter model. In terms of in vitro-in vivo extrapolation, the in vitro 2D model performed more similar to in vivo model compared to in vitro 3D model; however, it might have been due to the fact that only compounds secreted to medium were investigated. Thus, in further experiments to obtain full metabolome information, the intraspheroidal assessment or spheroid dissociation would be necessary.

New chemical biopsy tool for spatially resolved profiling of human brain tissue in vivo.

It is extremely challenging to perform chemical analyses of the brain, particularly in humans, due to the restricted access to this organ. Imaging techniques are the primary approach used in clinical practice, but they only provide limited information about brain chemistry. Solid-phase microextraction (SPME) has been presented recently as a chemical biopsy tool for the study of animal brains. The current work demonstrates for the first time the use of SPME for the spatially resolved sampling of the human brain in vivo. Specially designed multi-probe sampling device was used to simultaneously extract metabolites from the white and grey matter of patients undergoing brain tumor biopsies. Samples were collected by inserting the probes along the planned trajectory of the biopsy needle prior to the procedure, which was followed by metabolomic and lipidomic analyses. The results revealed that studied brain structures were predominantly composed of lipids, while the concentration and diversity of detected metabolites was higher in white than in grey matter. Although the small number of participants in this research precluded conclusions of a biological nature, the results highlight the advantages of the proposed SPME approach, as well as disadvantages that should be addressed in future studies.

Comparison of Metabolomic Profiles of Organs in Mice of Different Strains Based on SPME-LC-HRMS.

Given that the extent to which genetics alters the metabolomic profile of tissues is still poorly understood, the current study aimed to characterize and investigate the metabolite profiles of brain, liver, kidney and skeletal muscle of two common mouse inbred strains (BALB/c, C57BL/6) and one outbred stock (CD1) for strain-specific differences. Male mice (n = 15) at the age of 12 weeks were used: BALB/c (n = 5), C57BL/6 (n = 5) and CD1 (n = 5). Solid phase microextraction (SPME) was applied for the extraction of analytes from the tissues. SPME fibers (approximately 0.2 mm in diameter) coated with a biocompatible sorbent (4 mm length of hydrophilic-lipophilic balanced particles) were inserted into each organ immediately after euthanasia. Samples were analyzed using liquid chromatography coupled to a Q-Exactive Focus Orbitrap mass spectrometer. Distinct interstrain differences in the metabolomic patterns of brain and liver tissue were revealed. The metabolome of kidney and muscle tissue in BALB/c mice differed greatly from C57BL/6 and CD1 strains. The main compounds differentiating all the targeted organs were alpha-amino acids, purine nucleotides and fatty acid esters. The results of the study indicate that the baseline metabolome of organs, as well as different metabolic pathways, vary widely among general-purpose models of laboratory mice commonly used in biomedical research.

The level of selected hormones in peripheral blood in female polar foxes (Alopex lagopus L.) in relation to age.

The objective of the study was to determine the concentration profile of selected hormones in the blood serum of blue polar fox vixens at various ages during the non-mating period, three months after lactation. The investigation was performed on 50 clinically healthy female polar foxes derived from a domestic reproductive farm. The animals were divided into 5 age groups (n = 10) ranging from the 1 to 5 years of life. In the blood serum the contents of insulin, triiodothyronine (total and free), thyroxin (total and free), leptin and ghrelin (total and active) were determined. No significant, female age-dependent differences were found in the contents of insulin, total and free triiodothyronine and total ghrelin in the blood serum. Significant (P < 0.05) differences were observed in the concentration of total and free thyroxin (the highest in the blood of 4-year old females) as well as leptin and active ghrelin (the lowest and the highest content in 3-year old vixens). However, no distinct, female age-dependent tendencies for change in the content of these hormones in the blood serum were observed.

Changes in the content of major proteins and selected hormones in the blood serum of piglets during the early postnatal period.

The aim of the study was to determine the changes in the content of major proteins, glucose and selected hormones in the blood of piglets during the first 7 days of neonatal life. The study involved an entire litter of eight newborn piglets of F1 hybrids (Polish White Large x Polish Landrace) delivered from one sow in the second gestation. In blood samples collected directly after parturition (before colostrum intake), in the 12th, 24th and 48th hour and in 7th day of life, the content of total protein and its fractions, glucose concentration and the level of insulin, T3 (total and free), T4 (total and free), leptin, resistin and ghrelin (total and active) was determined. In the blood serum of newborn piglets a low content of total protein, albumins, gamma globulins and a high share of alpha- and beta globulins was found. In the 12th hour of life, after colostrum intake, a significant (P<0.05) increase in the content of total protein, albumins, beta-globulins and a rapid increase of gamma globulins as well as decrease of alpha-globulins level were observed. In the consecutive periods of postnatal life a significant (P<0.05) decrease of total protein, beta- and gamma globulins as well as a steady increase in the content of albumins in the blood serum of piglets was observed. The content of glucose, insulin, leptin, resistin and ghrelin in the blood serum of neonates increased significantly (P<0.05) after colostrum intake. During the successive experimental periods a progressive increase (P<0.05) of glucose and T3 as well as systematic decrease of insulin, T4, ghrelin and resistin in the blood serum was observed as compared to the 12th hour of life.

Morphological and mineral characteristics of peripheral blood in female polar fox in relation to age.

The aim of the study was to determine the values of selected haematological and biochemical parameters in peripheral blood of female polar fox in relation to the age of the animals. The research involved 50 polar fox females three months after the lactation period (i.e. the non-mating period). Animals were divided into 5 age groups (n = 10), ranging from 1 to 5 years of age. In blood samples the following parameters were determined: RBC, Ht, Hb, WBC, PLT, red blood cell parameters (MCV, MCH, MCHC) and the percentage of respective kinds of white blood cells in the total number of leukocytes. The content of Ca, Pi, Na, K, Cl, Mg, Fe, Cu and Zn as well as the ALP and ACP activity was determined in blood serum. In comparison with one year-old females, in peripheral blood of females from the remaining age groups an increase in RBC, Ht and Hb content was observed as well as a significant (P < 0.05) decrease in WBC level together with a lower number of lymphocytes and an increase in the relative content of granulocytes and monocytes. No distinct relationship between the content of Ca, Na, Cl, Mg, Cu, Zn, the activity of ALP and ACP and the age of the animals was observed. The highest concentration of Pi and K was found in the blood serum of one year-old females. The content of Fe decreased with age and was lowest in 5 year-old animals (P < 0.05).

One extraction tool for in vitro-in vivo extrapolation?SPME-based metabolomics of in vitro 2D,3D,and in vivo mouse melanoma models / 药物分析学报

Solid phase microextraction(SPME)in combination with high-resolution mass spectrometry was employed for the determination of metabolomic profile of mouse melanoma growth within in vitro 2D,in vitro 3D,and in vivo models.Such multi-model approach had never been investigated before.Due to the low-invasiveness of SPME,it was possible to perform time-course analysis,which allowed building time profile of biochemical reactions in the studied material.Such approach does not require the multiplication of samples as subsequent analyses are performed from the very same cell culture or from the same individual.SPME already reduces the number of animals required for experiment;therefore,it is with good concordance with the 3Rs rule(replacement,reduction,and refinement).Among tested models,the largest number of compounds was found within the in vitro 2D cell culture model,while in vivo and in vitro 3D models had the lowest number of detected compounds.These results may be connected with a higher metabolic rate,as well as lower integrity of the in vitro 2D model compared to the in vitro 3D model resulting in a lower number of compounds released into medium in the latter model.In terms of in vitro-in vivo extrapolation,the in vitro 2D model performed more similar to in vivo model compared to in vitro 3D model;however,it might have been due to the fact that only compounds secreted to medium were investigated.Thus,in further experiments to obtain full metabolome infor-mation,the intraspheroidal assessment or spheroid dissociation would be necessary.