Analytical performance comparing siemens whole blood point of care Atellica VTLi to the central laboratory plasma Atellica IM high-sensitivity cardiac troponin I assays.

INTRODUCTION: This study examined the analytical performance of a whole blood (WB) point of care (POC) hs-cTnI assay compared to a plasma central laboratory hs-cTnI assay in patients presenting with ischemic symptoms to a US emergency department. METHODS: Fresh WB specimens collected at 0 and 2 h from 1089 consecutive patients (2152 total from 1076 matched specimens) were analyzed for hs-cTnI using WB on POC Siemens Atellica VTLi assay and plasma on central laboratory Siemens Atellica IM assay. Concordances were determined based on concentrations ranging from < limit of detection (LoD), LoD to overall and sex specific 99th percentiles from both the IFCC manufacturer package inserts and Universal Sample Bank (USB) data, and > 99th percentiles. Method comparisons were calculated using Passing Bablok regression and Bland Altmann plots, and linear regression determined by Pearson correlation coefficient. RESULTS: Baseline concentration comparisons showed: POC VTLi < LoD 4-5 %, ≥ LoD 95 %; Atellica IM < LoD 5-7 %, and ≥ LoD 94-95 %. From the 2152 paired 0 and 2-hour samples, based on 99th percentiles, overall concordance was 91-92 % (kappa 0.72-0.77) and discordance 8 %. Passing Bablok regression analysis using 1924 specimens between LoD to 500 ng/L showed: slopes 0.469-0.490; y-intercepts 1.753-2.028; r values 0.631-0.817. Pearson correlation coefficient showed moderate to strong correlation strength, even with up to 53 % cTnI concentrations variance (Passing Bablok slopes) vs 27.0-40.1 % (Bland-Altmann plots). CONCLUSIONS: Up to 95 % of measured samples were > LoD for both the POC (Atellica VTLi) and central laboratory (Atellica IM) hs-cTnI assays. Moderate to strong concordance and correlation were observed between assays, despite up to 53 % variances in cTnI concentration.

Platelet Response to Allergens, CXCL10, and CXCL5 in the Context of Asthma.

Asthma is a chronic respiratory disease initiated by a variety of factors, including allergens. During an asthma attack, the secretion of C-X-C-motif chemokine 10 (CXCL10) and chemokine ligand 5 (CCL5) causes the migration of immune cells, including platelets, into the lungs and airway. Platelets, which contain three classes of chemical messenger-filled granules, can secrete vasodilators (adenosine diphosphate and adenosine triphosphate), serotonin (a vasoconstrictor and a vasodilator, depending on the biological system), platelet-activating factor, N-formylmethionyl-leucyl-phenylalanine ((fMLP), a bacterial tripeptide that stimulates chemotaxis), and chemokines (CCL5, platelet factor 4 (PF4), and C-X-C-motif chemokine 12 (CXCL12)), amplifying the asthma response. The goal of this work was threefold: (1) to understand if and how the antibody immunoglobulin E (IgE), responsible for allergic reactions, affects platelet response to the common platelet activator thrombin; (2) to understand how allergen stimulation compares to thrombin stimulation; and (3) to monitor platelet response to fMLP and the chemokines CXCL10 and CCL5. Herein, high-pressure liquid chromatography with electrochemical detection and/or carbon-fiber microelectrode amperometry measured granular secretion events from platelets with and without IgE in the presence of the allergen 2,4,6-trinitrophenyl-conjugated ovalbumin (TNP-Ova), thrombin, CXCL10, or CCL5. Platelet adhesion and chemotaxis were measured using a microfluidic platform in the presence of CXCL10, CCL5, or TNP-OVA. Results indicate that IgE binding promotes δ-granule secretion in response to platelet stimulation by thrombin in bulk. Single-cell results on platelets with exogenous IgE exposure showed significant changes in the post-membrane-granule fusion behavior during chemical messenger delivery events after thrombin stimulation. In addition, TNP-Ova allergen stimulation of IgE-exposed platelets secreted serotonin to the same extent as thrombin platelet stimulation. Enhanced adhesion to endothelial cells was demonstrated by TNP-Ova stimulation. Finally, only after incubation with IgE did platelets secrete chemical messengers in response to stimulation with fMLP, CXCL10, and CCL5.

Characterization of the Presence and Function of Platelet Opioid Receptors.

Opioids are typically used for the treatment of pain related to disease or surgery. In the body, they enter the bloodstream and interact with a variety of immune and neurological cells that express the µ-, δ-, and κ-opioid receptors. One blood-borne cell-like body that is not well understood in the context of opioid interactions is the platelet. The platelet is a highly sensitive anucleate cell-like fragment responsible for maintaining hemostasis through shape change and the secretion of chemical messengers. This research characterizes platelet opioid receptors, how specific receptor agonists impact platelet exocytosis, and the role of the κ-and µ-receptors in platelet function. Platelets were found to express all three opioid receptors, but upon stimulation with their respective agonist no activation was detected. Furthermore, exposure to the opioid agonists did not impact traditional platelet secretion stimulated by thrombin, a natural platelet activator. In addition, data collected from knockout mice suggest that the opioid agonists may be interacting nonspecifically with platelets. Dark-field images revealed differences in activated platelet shape between the κ- and µ-knockout platelets and the control platelets. Finally, κ-knockout platelets showed variations in their ability to adhere and aggregate compared to control platelets. Overall, these data show that platelet function is not likely to be heavily affected by blood-borne opioids.

Plasmodium chabaudi Affects Mast Cell Degranulation as Measured by Carbon-Fiber Microelectrode Amperometry.

Mast cells (MCs) are effector cells of the immune system commonly known for their role in asthma and allergy. They are present throughout biological systems in various tissues, serving as an interface between the biological system and environment. Previous work characterizing the impact of malaria on MCs revealed contradictory results, showing minimal to strong correlation between MC degranulation and disease progression. This work seeks to reveal how MC degranulation is impacted in the presence of malaria, induced by Plasmodium chabaudi, using a mouse model and a single cell measurement technique that reveals exquisite biophysical detail about any impacts to the degranulation process. It was hypothesized that the malaria parasites would impact MC degranulation response during live infection, and the differences would be revealed via carbon-fiber microelectrode amperometry. In fact, the data collected show that different stages of malaria infection affect MC degranulation differently, affirming the importance of considering different infection stages in future studies of malarial immune response. Furthermore, a comparison of MC degranulation response to that measured from platelets under similar circumstances shows similar trends in quantitative degranulation, suggesting that MC and platelet exocytosis machinery are affected similarly despite their distinct biological roles. However, based on the small number of mouse replicates, the studies herein suggest that there should be further study about cellular and disease processes. Overall, the work herein reveals important details about the role of MCs in malaria progression, relevant during treatment decisions, as well as a potentially generalizable impact on chemical messenger secretion from cells during malarial progression.

Antimalarial drugs impact chemical messenger secretion by blood platelets.

BACKGROUND: Advances in antimalarial drug development are important for combating malaria. Among the currently identified antimalarial drugs, it is suggested that some interact directly with the malarial parasites while others interact indirectly with the parasites. While this approach leads to parasite elimination, little is known about how these antimalarial drugs impact immune cells that are also critical in malarial response. METHODS: Herein, the effects of two common antimalarial drugs, chloroquine and quinine, on platelets were explored at both the bulk level, using high performance liquid chromatography, and the single cell level, using carbon-fiber microelectrode amperometry, to characterize any changes in chemical messenger secretion. RESULTS: The data reveal that both drugs cause platelet activation and reduce the number of platelet exocytosis events as well as delay fusion pore opening and closing. CONCLUSIONS: This work demonstrates how chloroquine and quinine quantitatively and qualitatively impact in vitro platelet function. GENERAL SIGNIFICANCE: Overall, the goal of this work is to promote understanding about how antimalarial drugs impact platelets as this may affect antimalarial drug development as well as therapeutic approaches to treat malarial infection.

Insight into the Effects of Plasmodium chabaudi on Platelets Using Carbon-Fiber Microelectrode Amperometry.

Platelets are anuclear circulating cell bodies within the bloodstream commonly known for their roles in clot formation during vascular injury to prevent blood loss. They also have significant impact in a range of diseases, including malaria. However, the role of platelets in malaria is controversial, with contradicting evidence suggesting either that they assist in destruction of malarial parasites or facilitate a severe form of malaria. Precedent work suggests that the timing of infection is critical in determining whether platelets switch roles from being protective to deleterious. As such, the work herein makes use of the unique mechanistic perspective offered by carbon-fiber microelectrode amperometry (CFMA) to understand how platelet secretion is impacted in malarial infection stages (ascending parasite count versus descending parasite count). Malarial platelet behavior was compared to platelets from noninfected control mice by probing their exocytotic function. Results suggest that mouse malaria caused by the parasite Plasmodium chabaudi, during both ascending and descending infection stages, reduces platelet exocytotic events and delays platelet granule fusion; in addition, platelets are more impacted by the disease early in the infection stages. In all, understanding platelet behavior in the malarial context may present new therapeutic routes to treat or cure malaria.