Serum Components Research Articles

Gas chromatography coupled with mass spectrometry for the rapid characterization and screening of volatile oil of euphorbia fischeriana steud

Gas chromatography (GC)-mass spectrometry is a powerful tool which is used in the analysis of volatile components in food and herbal medicine. Euphorbia fischeriana Steud., a traditional Chinese herb, has been used for thousands of years in China. Recently, scientists are exploring more on the pharmacological action of E. fischeriana. However, so far, there is no information regarding the volatile constituents of E. fischeriana. Therefore, in this study, we aimed to investigate and characterize the global chemical ingredients of volatile oil of E. fischeriana under in vitro and in vivo conditions. In this study, an accurate and rapid GC combined with the mass spectrometric method and MassHunter tool were employed to identify the comprehensive constituents and explored the absorbed components in rat serum after oral administration of E. fischeriana. According to our results, a total of 28 compounds were identified via in vitro analysis by matching with the NIST database. Of them, the following 14 compounds were tentatively characterized in the serum samples: (−)-alpha-cedrene; (+/-)-sesquithuriferone; cedrol; acorenone; neocembrene; geranylgeraniol; abietatriene; abieta-7,13-diene; cembrenol; sclareol; ethyl linoleate; isopimara-7,15-dien-3-one; abietadienal; and abietinol. Most of the compounds identified were diterpenes and sesquiterpenes. These results might be useful in various pharmacological and pharmacodynamic research, and the established method could offer valuable chemical information for bioactive volatile components characterization.

Association of serum sialic acid concentration with diabetic complications and cardiovascular risk factors in an Indian population.

IntroductionSialic acid (SA) is an acetylated product of neuraminic acid. It acts as a cofactor of many cell surface receptors (e.g. insulin receptors) and is positively associated with most of the serum acute phase reactants. Sialic acid is an important component of serum, which is elevated in diseases such as diabetes and certain malignancies. Diabetes mellitus (DM) is associated with an increase in SA concentration along with other complications. The present study was undertaken to assess the relationship between serum SA and type 2 diabetes.Material and methodsA total of 200 type 2 DM patients, 145 males and 55 females, were included in the study. Also, 100 healthy individuals served as the control group. Parameters assessed included serum SA, lipid profile, urine microalbumin, LDL-C, lipoprotein(a), and serum fibrinogen. The relationship between serum SA and diabetic complications viz retinopathy, nephropathy, and neuropathy was also assessed.ResultsType 2 DM patients had significantly higher levels (p < 0.01) of SA (77.35 ±4.6 mg%) as compared to the control group (68.23 ±7.9 mg%). Increased levels of serum SA were seen in patients with diabetic nephropathy and retinopathy. No correlation was seen between serum SA and diabetic neuropathy.ConclusionsElevated serum SA concentration is significantly related to type 2 DM and associated cardiovascular risk factors. Further study of acute-phase response markers and mediators as indicators or predictors of diabetic microvascular complications is therefore justified.

Bruch's Membrane and the Choroid in Age-Related Macular Degeneration.

A healthy choroidal vasculature is necessary to support the retinal pigment epithelium (RPE) and photoreceptors, because there is a mutualistic symbiotic relationship between the components of the photoreceptor/retinal pigment epithelium (RPE)/Bruch's membrane (BrMb)/choriocapillaris (CC) complex. This relationship is compromised in age-related macular degeneration (AMD) by the dysfunction or death of the choroidal vasculature. This chapter will provide a basic description of the human Bruch's membrane and choroidal anatomy and physiology and how they change in AMD.The choriocapillaris is the lobular, fenestrated capillary system of choroid. It lies immediately posterior to the pentalaminar Bruch's membrane (BrMb). The blood supply for this system is the intermediate blood vessels of Sattler's layer and the large blood vessels in Haller's layer.In geographic atrophy (GA), an advanced form of dry AMD, large confluent drusen form on BrMb, and hyperpigmentation (presumably dysfunction in RPE) appears to be the initial insult. The resorption of these drusen and loss of RPE (hypopigmentation) can be predictive for progression of GA. The death and dysfunction of CC and photoreceptors appear to be secondary events to loss in RPE. The loss of choroidal vasculature may be the initial insult in neovascular AMD (nAMD). We have observed a loss of CC with an intact RPE monolayer in nAMD, by making RPE hypoxic. These hypoxic cells then produce angiogenic substances like vascular endothelial growth factor (VEGF), which stimulate growth of new vessels from CC, resulting in choroidal neovascularization (CNV). Reduction in blood supply to the CC, often stenosis of intermediate and large blood vessels, is associated with CC loss.The polymorphisms in the complement system components are associated with AMD. In addition, the environment of the CC, basement membrane and intercapillary septa, is a proinflammatory milieu with accumulation of proinflammatory molecules like CRP and complement components during AMD. In this toxic milieu, CC die or become dysfunctional even early in AMD. The loss of CC might be a stimulus for drusen formation since the disposal system for retinal debris and exocytosed material from RPE would be limited. Ultimately, the photoreceptors die of lack of nutrients, leakage of serum components from the neovascularization, and scar formation.Therefore, the mutualistic symbiotic relationship of the photoreceptor/RPE/BrMb/CC complex is lost in both forms of AMD. Loss of this functionally integrated relationship results in death and dysfunction of all of the components in the complex.

Effects of Improved (Addition of Antimicrobials and Antioxidants) Locally Produced Natural Vitamin Premix on Performance, Hematology and Some Serum Constituents of Broiler Chickens

An experiment was conducted to compare the haematology and some serum constituents and performance of broilers fed diets containing improved (addition of anti-microbial and anti-oxidant) locally produced natural vitamin premix (LPNVP), LPNVP without improvement and commercial vitamin premix (CVMP). Haematology of the birds was not affected by the dietary treatments. However, red blood cell (RBC) count was significantly high (P&lt;0.05) for the birds fed diet containing improved LPNVP The haemoglobin, PCK, RBC and WBC values observed for birds fed improved LPNVP were 13 gm%, 32%, 246 x 1012/L and 3.1 x 1010/L respectively. Serological parameters observed in the study were not significantly different (P&gt;0,.05) for all the dietary conditions studied. Birds fed diet containing improved LPNVP also recorded comparable serum components to control diet containing the commercial vitamin/’mineral premix. Serum calcium, phosphorus, glucose, ureate, creatinine and total protein observed for birds fed diet containing improved LPNVP were 2.32%, 0.70%, 3.60Mmo/L, 2.9Mmo/L, 0.25Mmol/L, 58Mmol/L and 66g/100ml respectively. Feed intake, weight gain and feed utilization were similar (P&gt;0.05) for the various dietary treatments. Birds fed improved LPNVP based diets tended to record better feed intake than did the control birds on CVMP based diets and unimproved LPNVP. Generally, improvement of LPNVP tended to give better performance of broilers than did other dietary treatments.

Pre and post-gestation blood indices of rabbit does fed graded levels of neem (Azadirachta indica) leaf meal

This study was conducted to investigate the effect of neem leaf meal on the blood status of rabbit does before and after gestation. Atotal of twenty-four (24) rabbit does of mixed breeds, with initial average live weight of 1975g were randomly allocated into four (4) experimental dietary treatments; each of which was replicated three (3) times with two (2) rabbits per replicate. The rabbits were fed diets containing 0%, 2.5%, 5.0%, and 7.5% neem (Azadirachta indica) leaf meal. Parameters measured included packed cell volume (PCV), haemoglobin, red blood cells (RBC), white blood cells (WBC) and its differentials for haematology while total protein, globulin, albumin, cholesterol, alkaline phosphate (ALP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured for biochemical indices. All data generated were subjected to one-way Analysis of Variance in a Completely Randomized Design. The findings from this study at pre-gestation showed that neem leaf meal (NLM) significantly (p&lt;0.05) influenced basophil and serum total protein, globulin and glucose. Basophil was significantly reduced in rabbits fed 5.0% NLM (0.33%) while the serum components were affected in rabbits fed 2.5% NLM. The result at post-gestation showed no significant (p&gt;0.05) difference in all the parameters measured exceptneutrophils. Neutrophils were significantly (p&lt;0.05) reduced in rabbits on the control diet (26.67%) while rabbits on NLM diets recorded similar values. From the serum component, cholesterol was significantly (p&lt;0.05) highest in rabbits fed 5.0% NLM (62.33mg/dL) and lowest in those fed 2.5% NLM (49.67mg/dL). It was, therefore, concluded that inclusion of NLM in rabbit does' diet up to 7.5% had no adverse effect on the health status of the animals as the values obtained for neutrophils and cholesterol were still within the normal range of rabbit blood profile.

Correlating the Structure and Activity of Y. pestis Ail in a Bacterial Cell Envelope

Understanding microbe-host interactions at the molecular level is a major goal of fundamental biology and therapeutic drug development. Structural biology strives to capture biomolecular structures in action, but the samples are often highly simplified versions of the complex native environment. Here, we present an Escherichia coli model system that allows us to probe the structure and function of Ail, the major surface protein of the deadly pathogen Yersinia pestis. We show that cell surface expression of Ail produces Y. pestis virulence phenotypes in E. coli, including resistance to human serum, cosedimentation of human vitronectin, and pellicle formation. Moreover, isolated bacterial cell envelopes, encompassing inner and outer membranes, yield high-resolution solid-state NMR spectra that reflect the structure of Ail and reveal Ail sites that are sensitive to the bacterial membrane environment and involved in the interactions with human serum components. The data capture the structure and function of Ail in a bacterial outer membrane and set the stage for probing its interactions with the complex milieu of immune response proteins present in human serum.

Magnetic bead extraction with acid dissociationimmunoassay for the determination of serum CD80-Fc fusion protein.

Background: To detect concentrations of subtherapeutic doses of the CD80-Fc fusion protein FPT155 in serum in Phase I studies, a highly sensitive assay was developed. Materials & methods: FPT155 was purified from human serum using magnetic beads coupled to cytotoxic T-lymphocyte-associated antigen-4. After washing away the serum components, FTP155 was released by acid dissociation and neutralization. The eluted drug was quantified in an ELISA using cytotoxic T-lymphocyte-associated antigen-4 as a capture reagent and biotinylated anti-human Fc for detection. The assay was validated with a calibration range of 5-40ng/ml and a dilutional integrity of up to 100,000ng/ml. Conclusion: A highly sensitive assay to determine serum concentrations of FPT155 using readily available reagents was developed. The results were in conformity with theoretical calculations.

Relationship between Serum Fatty Acids and Components of Physical Frailty in Community-Dwelling Japanese Older Adults.

Polyunsaturated fatty acids help maintain insulin sensitivity, mitochondrial function, and anti-inflammation. It is well known that deterioration in these areas can cause frailty. However, little is known about the differences in serum polyunsaturated fatty acid levels among frailty components. We investigated the cross-sectional relationship between frailty and serum fatty acids in 1,033 community-dwelling older adults aged 60-88 years. Polyunsaturated fatty acid concentrations were measured from fasting blood samples. The modified phenotype criteria defined frailty. Polyunsaturated fatty acid levels were compared among each component using general linear modeling after controlling for sex, age, body mass index, smoking status, household income, and medical history. Lower polyunsaturated fatty acid levels were associated with the modified frailty criteria, including shrinking and weakness (p < 0.05). Our findings suggest that serum polyunsaturated fatty acid levels differ depending on the frailty status of older adults.

Immunoglobulin D Myeloma and Amyloidosis: Immunochemical and Structural Studies of Bence Jones and Amyloid Fibrillar Proteins

Urinary Bence Jones protein and amyloid fibril protein isolated from the subcutaneous tissue of a patient with IgD myeloma and associated amyloidosis were subjected to physicochemical and immunochemical identification. Peptide maps and amino-terminal tetrapeptide composition obtained from the two proteins were comparable. Immunochemical cross-reactivity between the two proteins, with other lambda-type amyloid and Bence Jones proteins, and with a serum component was demonstrated. The results suggest that the source of the amyloid fibril protein is an intact circulating light polypeptide chain as well as smaller amino-terminal fragments.

Genetically Corrected Autologous Stem Cells Engraft, But Host Immune Responses Limit Their Utility in Canine α-L-iduronidase Deficiency

Canine alpha-L-iduronidase (alpha-ID) deficiency, a model of the human storage disorder mucopolysaccharidosis type I (MPS I), is an ideal system in which to evaluate the clinical benefit of genetically corrected hematopoietic stem cells. We performed adoptive transfer of genetically corrected autologous hematopoietic cells in dogs with alpha-ID deficiency. Large volume marrow collections were performed on five alpha-ID-deficient dogs. Marrow mononuclear cells in long-term marrow cultures (LTMCs) were exposed on three occasions during 3 weeks of culture to retroviral vectors bearing the normal canine alpha-ID cDNA. Transduced LTMC cells from deficient dogs expressed enzymatically active alpha-ID at 10 to 200 times the levels seen in normal dogs. An average of 32% of LTMC-derived clonogenic hematopoietic cells were provirus positive by polymerase chain reaction and about half of these expressed alpha-ID. Approximately 10(7) autologous gene-modified LTMC cells/kg were infused into nonmyeloablated recipients. Proviral DNA was detected in up to 10% of individual marrow-derived hematopoietic colonies and in 0.01% to 1% of blood and marrow leukocytes at up to 2 to 3 years postinfusion. Despite good evidence for engraftment of provirally marked cells, neither alpha-ID enzyme nor alpha-ID transcripts were detected in any dog. We evaluated immune responses against alpha-ID and transduced cells. Humoral responses to alpha-ID and serum components of the culture media (fetal bovine and horse sera and bovine serum albumin) were identified by enzyme-linked immunosorbent assay. Cellular immune responses to autologous alpha-ID but not neo(r) transduced cells were demonstrated by lymphocyte proliferation assays. To abrogate potential immune phenomena, four affected dogs received posttransplant cyclosporine A. Whereas immune responses were dampened in these dogs, alpha-ID activity remained undetectable. In none of the dogs engrafted with genetically corrected cells was there evidence for clinical improvement. Our data suggest that, whereas the alpha-ID cDNA may be transferred and maintained in approximately 5% of hematopoietic progenitors, the potential of this approach appears limited by the levels of provirally derived enzyme that are expressed in vivo and by the host's response to cultured and transduced hematopoietic cells expressing foreign proteins.

Shear-responsive peptide/siRNA complexes as lung-targeting gene vectors

Particles administrated intravenously will pass through the pulmonary capillary network before being distributed to the body. Therefore, fabrication of vectors sensitive to blood shear and active with blood components should be a practical approach to develop lung-targeting gene carriers self-regulated by circulatory system. In this work, we designed a series of cationic peptides with the same charge density but varying hydrophobicity and capacity to form hydrogen bonds, and investigated their ability to form complexes with siRNA, the behaviours of peptide/siRNA complexes in the presence of serum under shear, and the lung-targeting efficacy of the complexes regulated by blood. The hydrophobic interaction controls the complexation between peptide and siRNA, while the hydrogen bonds are responsible for the binding of peptides to the serum components in blood. In vivo tests show that all the peptide/siRNA complexes can accumulate in lung. However, only the complexes that exhibit weak interaction with serum components and can be broken down by shear avoid the inflammation and death caused by pulmonary embolism. Moreover, the peptide with strong hydrophobicity can retain siRNA in lung without early release of the cargo. Our study provides a step toward the development of adaptive gene carriers under the regulation of circulatory system.

Detection of Glycine as a Model Protein in Blood Serum Using 2D-IR Spectroscopy.

Glycine (Gly) is used as a model system to evaluate the ability of ultrafast two-dimensional infrared (2D-IR) spectroscopy to detect and quantify the low-molecular-weight proteinaceous components of blood serum. Combining data acquisition schemes to suppress absorption bands of H2O that overlap with the protein amide I band with analysis of peak patterns appearing in the off-diagonal region of the 2D-IR spectrum allows separation of the Gly spectral signature from that of the dominant protein fraction of serum in a transmission-mode 2D-IR measurement without any sample manipulation, e.g., filtration or drying. 2D-IR spectra of blood serum samples supplemented with varying concentrations of Gly were obtained, and a range of data analysis methods compared, leading to a detection limit of ∼3 mg/mL for Gly. The reported methodology provides a platform for a critical assessment of the sensitivity of 2D-IR for measuring the concentrations of amino acids, peptides, and low-molecular-weight proteins present in serum samples. We conclude that, in the case of several clinically relevant diagnostic molecules and their combinations, the potential exists for 2D-IR to complement IR absorption methods as the benefits of the second frequency dimension offered by 2D-IR spectroscopy outweigh the added technical complexity of the measurement.

Lipid-Lowering Effects of Lotus Leaf Alcoholic Extract on Serum, Hepatopancreas, and Muscle of Juvenile Grass Carp via Gene Expression.

Compared with wild grass carp (Ctenopharyngodon idellus), intensively cultured fish displayed disordered lipid metabolism, showing excess lipid deposition in the hepatopancreas and muscle. Lotus leaf prevents fat accumulation in humans and may have similar effects on fish. This study explored the regulatory mechanisms by which the dietary addition of an alcoholic extract of lotus leaf (AELL) reduced lipid deposition in the hepatopancreas and muscle of juvenile grass carp. The fish (average initial weight: 34.00 ± 0.40 g) were fed four experimental diets containing different AELL levels (0, 0.07, 0.14, and 0.21%) for 8 weeks. Serum components, lipid droplet size, triacylglycerol (TAG) content, enzymatic activities, and mRNA levels of genes related to lipid metabolism in the hepatopancreas and muscle were analyzed. The results show that dietary AELL supplementation significantly reduced the TAG content and lipid droplet area in the histological sections as well as the fatty acid synthase (FAS) activity in both the hepatopancreas and muscle but enhanced the activities of lipoprotein lipase (LPL) and carnitine palmitoyltransferase I (CPT1) in both tissues. In addition, dietary AELL supplementation decreased the mRNA expression of genes involved in fatty acid uptake (cd36, fatp1/fatp4/fatp6, fabp10/fabp11, acsl1/acsl4) and de novo lipid synthesis (pgd, g6pd, and fasn) as well as the transcription factors pparg and srebf1 in the hepatopancreas and muscle but increased the mRNA levels of genes relating to lipid catabolism (cpt1a, lipe, pnpla2, lpl), lipid transportation (apob), and the transcription factor ppara in both tissues. In conclusion, dietary AELL supplementation reduced lipid accumulation in the hepatopancreas and muscle by affecting the gene expression of proteins with known effects on lipid metabolism in juvenile grass carp.

Relationship between Serum Uric Acid Levels, Insulin Resistance and Components of Metabolic Syndrome in Korean Adults

Relationship between Serum Uric Acid Levels, Insulin Resistance and Components of Metabolic Syndrome in Korean Adults

Electrochemical immunosensor for serum parathyroid hormone using voltammetric techniques and a portable simulator

An electrochemical platform based on a screen-printed carbon electrode (SPCE) is developed to detect parathyroid hormone (PTH). A nanocomposite of multi-walled carbon nanotube (MWCNT) and gold nanoparticles (AuNP) was deposited on the SPCE to immobilize antibodies and horseradish peroxidase (HRP). MWCNT improved the stability and conductivity of the immunosensor because of its good electron-transfer ability and tubular structure. The AuNP not only provided a large surface area for antibody immobilization, but it also enhanced the electrochemical signal for enzyme-linked immunosensing. Cyclic voltammetry showed both electron transfer and the effective surface area were increased on the modified electrode. The characteristics of the modified SPCE were assayed by Raman spectroscopy, scanning electron microscopy, atomic force microscopy, and electrochemical techniques. The linear detection range of this PTH immunosensor was within 1–300 pg/ml, and the electrochemical performance was not affected by interference from protein components in human serum. After storage at 4 °C for 28 days, 85% PTH sensing ability of this immunosensor was maintained compared to the freshly prepared one using the SWV and DPV methods. The relative standard deviations of all measurements were within 3–8% for both voltammetric methods. These results indicated the developed immunosensor had good stability and reproducibility. This PTH immunosensor had a detection limit of 0.886 and 0.065 pg/ml for the differential pulse voltammetry and square wave voltammetry, respectively. We provided a quick analysis of serum PTH which might be used as an electrochemical immunosensing platform for point-of-care testing.

PSIX-13 Influence of an integrated fodder additive on productivity and metabolic homeostasis of dairy cattle in transition period

Abstract The transition period is the most stressful for dairy cows in terms of metabolic rate and characterized by dynamic changes in the hormonal profile, loss of appetite, negative energy balance, nutrient and vitamin deficiencies, compromised immunity and oxidative stress. The aim of the research was to balance the increase in cows’ productivity and the consumption of nutrients from the feed during the transition period through the use of energy feed additives (EFS), high protein feed concentrate (PFC) in combination with biologically active (BA) substances. Thirty-six dairy cows of black-motley breed were divided into 3 groups: (1) The main diet (G0, n = 12), not supplemented with additives, (2) additive 1 (G1; supplemented with 0.5 kg EFS / cow per day, n = 12), (3) additive 2 (G2; supplemented with 0.5 kg EFS; 0.5 kg PFC and 0.06 kg BA / cow per day, n = 12). The experimental part of the feeding lasted for 20 days before calving and the first 100 days after calving (DPP). Blood samples were collected during the first and the third months after calving. The biochemical analyzer was used to measure blood serum components. The milk yield was recorded monthly throughout lactation. BCS losses from 7 days prior to calving to 28 DPP were -0.29 for Gr2, -0.42 for Gr1 and -0.60 for Gr0. Feeding with additive 2 (Gr2) increased milk yield (p≤0.05) in the first 100 DPPs and (p≤0.001) 305 DPPs compared to G0. The number of samples with SCC ≥ 500 000 / ml. per 100 DPP was 51% in the Gr0 group, 27% in Gr1, 25% in Gr2. Also, microbiological control showed an excess of Bacterial cells in the milk of cows from group G0, which correlates with SCC indicators. Cows from Gr2had a lower content of ketone bodies (P≥0.001) and higher (P≥0.001) antioxidant activity of blood serum compared to Gr0. Cows from Gr2 had an increase in ALT activity by1.4 times (P &amp;lt;0.05) from the first to the third months after calving.

The Role of Masks in Mitigating the SARS-CoV-2 Pandemic: Another Piece of the Puzzle.

Bundgaard and colleagues report the results of a unique randomized controlled trial of mask wearing in the community to mitigate SARS-CoV-2 infection. The editorialists discuss how the findings add to what we know about mask wearing in the community and risk for SARS-CoV-2 infection.

Neck adipose tissue accumulation is associated with higher overall and central adiposity, a higher cardiometabolic risk, and a pro-inflammatory profile in young adults

Neck adipose tissue (NAT) volume increases with general adiposity, with fat accumulating in different neck tissue compartments. In patients with certain malignant/benign tumours, the accumulation of NAT, and certain NAT distributions, have been associated with cardiometabolic risk (CMR). However, it is unknown whether the same relationships exist in healthy people, and whether NAT accumulation and distribution are related to the inflammatory status. In this cross-sectional study, 139 young healthy adults (68% women) underwent a computed tomography scan to quantify the volume of compartmental (i.e., subcutaneous, intermuscular and perivertebral) and total NAT at the height of vertebra C5. Anthropometric indicators were measured, and body composition determined using dual energy X-ray absorptiometry. Information on CMR factors (i.e., blood glycaemic and lipid markers, blood pressure and physical fitness) was also gathered, and a CMR score calculated. Several plasma cytokines and serum components of the innate immune system were measured to determine the inflammatory status. Compartmental and total NAT volumes were directly related to body mass index (BMI), and lean, fat, and visceral adipose tissue (VAT) masses (all, P ≤ 0.05). Larger compartmental (especially intermuscular) and total NAT volumes were directly associated with the CMR score, several CMR factors (i.e., glycaemic and lipid markers and blood pressure), and the C3, C4 and leptin concentrations. They were, however, inversely correlated with the CMR factors high density lipoprotein-cholesterol (HDL-C) and physical fitness, and with the adiponectin concentration (all P ≤ 0.05). Several of these associations remained statistically significant (P ≤ 0.05) after adjustment for BMI, body fat percentage or VAT mass. Overall, results did not change after applying false discovery rate correction. NAT volume and its distribution among different tissue compartments is associated with the CMR and inflammatory profile of young healthy adults. Total NAT volume appears to be as valuable as VAT mass in terms of predicting CMR and inflammatory status.

Cardiac transthyretin wild-type amyloidosis (ATTRwt): a prospective study of 400 patients followed at the Italian referral center

Abstract Background Cardiac wild type transthyretin (ATTRwt) amyloidosis, formerly known as senile systemic amyloidosis, is an increasingly recognized, progressive, and fatal cardiomyopathy. Two biomarkers staging systems were proposed based on NT-proBNP (in both cases) and troponin or estimated glomerular filtration rate, that are able to predict survival in this population. The availability of novel effective treatments requires large studies to describe the natural history of the disease in different populations. Objective To describe the natural history of the disease in a large, prospective, national series. Methods Starting in 2007, we protocolized data collection in all the patients diagnosed at our center (n=400 up to 7/2019). Results The referrals to our center increased over time: 5 cases (1%) between 2007–2009, 33 (9%) in 2010–2012, 90 (22%) in 2013–2015 and 272 (68%) in 2016–2019. Median age was 76 years [interquartile range (IQR): 71–80 years] and 372 patients (93%) were males. One hundred and seventy-three (43%) had atrial fibrillation, 63 (15%) had a history of ischemic cardiomyopathy and 64 (15%) underwent pacemaker or ICD implantation. NYHA class was I in 58 subjects (16%), II in 225 (63%) and III in 74 (21%). Median NT-proBNP was 3064 ng/L (IQR: 1817–5579 ng/L), troponin I 0.096 ng/mL (IQR: 0.063–0.158 ng/mL), eGFR 62 mL/min (IQR: 50–78 mL/min). Median IVS was 17 mm (IQR: 15–19 mm), PW 16 mm (IQR: 14–18 mm) and EF 53% (IQR: 45–57%). One-hundred and forty-eight subjects (37%) had a concomitant monoclonal component in serum and/or urine and/or an abnormal free light chain ratio. In these patients, the diagnosis was confirmed by immunoelectron microscopy or mass spectrometry. In 252 (63%) the diagnosis was based on bone scintigraphy. DNA analysis for amyloidogenic mutations in transthyretin and apolipoprotein A-I genes was negative in all subjects. The median survival of the whole cohort was 59 months. The Mayo Clinic staging based on NT-proBNP (cutoff: 3000 ng/L) and troponin I (cutoff: 0.1 ng/mL) discriminated 3 different groups [stage I: 131 (35%), stage II: 123 (32%) and stage III: 127 (33%)] with different survival between stage I and II (median 86 vs. 81 months, P=0.04) and between stage II and III (median 81 vs. 62 months, P&amp;lt;0.001). The UK staging system (NT-proBNP 3000 ng/L and eGFR 45 mL/min), discriminated three groups [stage I: 170 (45%), stage II: 165 (43%) and stage III: 45 (12%)] with a significant difference in survival: between stage I and stage II (86 vs. 52 months, P&amp;lt;0.001) and between stage II and stage III (median survival 52 vs. 33 months, P=0.045). Conclusions This is one of the largest series of patients with cardiac ATTRwt reported so far. Referrals and diagnoses increased exponentially in recent years, One-third of patients has a concomitant monoclonal gammopathy and needed tissue typing. Both the current staging systems offered good discrimination of staging and were validated in our independent cohort. Funding Acknowledgement Type of funding source: None

Key Immune Checkpoint PD-1/PD-L1 Signaling Pathway Components in the Blood Serum from Patients with Bone Tumors

The levels of sPD-1 and sPD-L1 were analyzed in blood serum of 132 patients (age 14-70 years) with primary bone tumors: osteosarcoma (N=39), chondrosarcoma (N=42), Ewing sarcoma (N=9), chordoma (N=12), giant-cell bone tumor (GCBT) (N=16), benign neoplasms (N=14) and in and practically healthy subjects (age 19-58 years; N=27). sPD-L1 levels in all studied bone neoplasms were significantly higher than in the control. Serum sPD-1 level in GCBT patients was significantly higher than in the control, benign neoplasms, chondrosarcoma, and chordoma patients, but did not differ from osteosarcoma group. sPD-1 concentration in Ewing sarcoma was significantly higher than in chordoma and chondrosarcoma, but did not differ from the control. sPD-1 level in chondrosarcoma patients was also lower than in osteosarcoma, Ewing sarcoma, and in the control. Both sPD-1 and sPD-L1 concentrations were not significantly associated with the type of affected bone, process localization, disease stage, tumor histological grade, patients' age and sex. These results suggest the possibility of using these biological markers for preliminary assessment of the character of the process in the bone.