The pyruvate kinase activity in peripheral and uterine blood in women with atypical endometrial hyperplasia and endometrial cancer

IntroductionPyruvate kinase in an enzyme that catalyzes the production of pyruvate and ATP as the final step of the glycolysis. Potential role of Pyruvate kinase in tumorigenesis was previously suggested, due to its altered activity in several tumors.Material and methodsThis study looks at M2 isozyme of pyruvate kinase activity (M2-PK) measured in peripheral and uterine blood plasma in various stages of endometrial cancer (EC) as well as in precancerous state of atypical endometrial hyperplasia (AEH). Measurements were performed using spectrophotometric method in citrate plasma samples from peripheral and uterine blood. Study group included 84 patients with endometrial cancer, 28 patients with atypical endometrial hyperplasia and 23 non-cancerous controls.ResultsPyruvate kinase activity in EC group was 3-fold higher than in control group both in peripheral and uterine blood samples. Pyruvate kinase activity was also 3-fold higher in uterine blood when compared to peripheral blood samples. We also found statistically significant correlation between FIGO staging and detected activity with the M2-PK activity being 2-fold higher for FIGO1 than for FIGO3. We also describe a paradox in which the M2-PK activity in patients with atypical endometrial hyperplasia is lower than M2-PK activity in control group in peripheral blood samples, but higher in uterine blood samples.ConclusionsThe measurement of citrate plasma pyruvate kinase metabolic activity varies greatly between samples collected from different sites and samples collected from patients with varied tumor staging. Further studies are needed in order to elucidate molecular pathways that are responsible for observed differences.

A Comparison of Serum and Plasma Blood Collection Tubes for the Integration of Epidemiological and Metabolomics Data

Blood is a rich biological sample routinely collected in clinical and epidemiological studies. With advancements in high throughput -omics technology, such as metabolomics, epidemiology can now delve more deeply and comprehensively into biological mechanisms involved in the etiology of diseases. However, the impact of the blood collection tube matrix of samples collected needs to be carefully considered to obtain meaningful biological interpretations and understand how the metabolite signatures are affected by different tube types. In the present study, we investigated whether the metabolic profile of blood collected as serum differed from samples collected as ACD plasma, citrate plasma, EDTA plasma, fluoride plasma, or heparin plasma. We identified and quantified 50 metabolites present in all samples utilizing nuclear magnetic resonance (NMR) spectroscopy. The heparin plasma tubes performed the closest to serum, with only three metabolites showing significant differences, followed by EDTA which significantly differed for five metabolites, and fluoride tubes which differed in eleven of the fifty metabolites. Most of these metabolite differences were due to higher levels of amino acids in serum compared to heparin plasma, EDTA plasma, and fluoride plasma. In contrast, metabolite measurements from ACD and citrate plasma differed significantly for approximately half of the metabolites assessed. These metabolite differences in ACD and citrate plasma were largely due to significant interfering peaks from the anticoagulants themselves. Blood is one of the most banked samples and thus mining and comparing samples between studies requires understanding how the metabolite signature is affected by the different media and different tube types.

A simple method for measuring fibrinogen and evaluating its functionality

Актуальность. Определение фибриногена (ФГ) по Клауссу является чувствительным к гипо-, дис- и гипер-фибриногенемии, а также к продуктам деградации фибрина, которые влияют на процесс коагуляции и могут быть причиной занижения или повышения уровня фибриногена. Цель настоящей работы - разработка простого турбидиметрического теста для определения фибриногена и оценка его функциональности. Материалы и методы. В работе определяли фибриноген в цитратных плазмах 96 пациентов по методу Клаусса, модифицированным методом Рутберга и новым турбидиметрическим тестом. Результат. Разработан простой турбидиметрический тест определения фибриногена, основанный на определении коагулированного фибриногена при рекальцификации цитратной плазмы по формуле: ФГ (г / л) = ΔА450 × 5,12, где ΔА450 - изменение оптической плотности в опытной пробе; 5,12 - переводной коэффициент ΔА450 в г/л ФГ. Корреляционный анализ показал высокую степень сходимости результатов (R = 0,843). При разнице ФГ более 10% по Клауссу и по разработанному методу констатируют как нарушенную функциональность ФГ. Заключение. Использование простого, доступного турбидиметрического метода определения фибриногена в рутинных исследованиях позволяет оценивать функциональность фибриногена. Background. The Clauss fibrinogen assay is sensitive to hypo-, dys- and hyper-fibrinogenemia, as well as to fibrin degradation products, which may affect coagulation and cause underestimation of or increase the fibrinogen level. The aim of this study was to develop a simple turbidimetric test to measure fibrinogen and evaluate its functionality. Materials and methods. In this study, fibrinogen was measured in citrate plasma from 96 patients using the Clauss assay, a modified Rutberg method, and a new turbidimetric test. Result. A simple turbidimetric test was developed for measuring fibrinogen. This test is based on measuring coagulated fibrinogen during recalcification of citrate plasma according to the formula, FG (g/l) = ΔА450 × 5.12, where ΔА450 is a change in optical density of the experimental sample; 5.12 is a conversion coefficient ΔА450 in g/l FG. Correlation analysis showed a high degree of agreement between the results (R = 0.843). With a FG difference of more than 10% by the Clauss assay and the developed method, impaired functionality of the FG is stated. Conclusion. Using a simple, affordable turbidimetric method to determine fibrinogen in routine studies allows accessing the fibrinogen functionality.

Can citrate plasma be used in exceptional circumstances for some clinical chemistry and immunochemistry tests?

Background The use of alternative sample matrices may be an advantageous perspective when the laboratory falls short of serum or lithium-heparin plasma for performing clinical chemistry and/or immunochemistry testing. This study was aimed at exploring whether some tests may be performed in citrate plasma as an alternative to lithium-heparin plasma. Methods Paired lithium-heparin and citrate plasma samples collected from 55 inpatients were analyzed on Roche Cobas 8000 for 28 different clinical chemistry and immunochemistry parameters. Data obtained in citrate plasma were adjusted for either the dilution factor or using an equation corresponding to the linear regression calculated by comparing unadjusted lithium-heparin and citrate plasma values. Results Except for magnesium (+17%) and sodium (+11%), unadjusted values of all remaining analytes were significantly lower in citrate than in lithium-heparin plasma, with bias ranging between −6.4% and −25.9%. The correlation between lithium-heparin and citrate plasma values was generally excellent (i.e. >0.90). The adjustment of citrate plasma values for the dilution factor (i.e. 1.1) was only effective in harmonizing the results of albumin and lipase, whilst the concentration of all other analytes remained significantly different between the two sample matrices. The adjustment of plasma citrate values using corrective formulas was instead effective in harmonizing all parameters, with no results remaining statistically different between the two sample matrices. Conclusions Citrate plasma may be used in exceptional circumstances for clinical chemistry and immunochemistry testing as a replacement for lithium-heparin plasma, provided that citrate plasma values are adjusted by using validated corrective equations.

Hibrid biomaterials based on hydroxyapatite and blood components

Hybrid biomaterials based on amorphous hydroxyapatite and blood components (fibrin, citrate plasma) were developed by chemical precipitation of hydroxyapatite in a biopolymer matrix (pH 11; Ca/P ratio 1.67) and by mixing 6–14 wt.% of hydroxyapatite gel (pH 7.0–7.2) with bipolymers. Chemically precipitated hydroxyapatite in biopolymer matrices is single phase or contains ticalcium phosphate impurity up to 30 %, mainly α-modification in fibrin matrix and β-modification in citrate plasma. The interaction of hydroxyapatite gel into the fibrin leads to significant amorphization of hydroxyapatite and an increase in its bioresorbability. Holding the composites with hydroxyapatite obtained by chemical precipitation in the Simulated Body Fluid model solution for 75 days leads to their partial resorption and simultaneous increase of biomimetic apatite, with its greater weight gain on composites with a fibrin. Hybrid biomaterials based on a fibrin obtained from the patient’s blood and hydroxyapatite gel showed positive result during implantation, allowing to form an adequate configuration of the defect, expanding the possibilities of ENT surgery.

IL8 and IL16 levels indicate serum and plasma quality

Background: Longer pre-centrifugation times alter the quality of serum and plasma samples. Markers for such delays in sample processing and hence for the sample quality, have been identified. Methods: Twenty cytokines in serum, EDTA plasma and citrate plasma samples were screened for changes in concentration induced by extended blood pre-centrifugation delays at room temperature. The two cytokines that showed the largest changes were further validated for their “diagnostic performance” in identifying serum or plasma samples with extended pre-centrifugation times. Results: In this study, using R&D Systems ELISA kits, EDTA plasma samples and serum samples with a pre-centrifugation delay longer than 24 h had an IL16 concentration higher than 313 pg/mL, and an IL8 concentration higher than 125 pg/mL, respectively. EDTA plasma samples with a pre-centrifugation delay longer than 48 h had an IL16 concentration higher than 897 pg/mL, citrate plasma samples had an IL8 concentration higher than 21.5 pg/mL and serum samples had an IL8 concentration higher than 528 pg/mL. Conclusions: These robust and accurate tools, based on simple and commercially available ELISA assays can greatly facilitate qualification of serum and plasma legacy collections with undocumented pre-analytics.