Methods Blood Samples Research Articles

Cellular immune response to a single dose of live attenuated hepatitis a virus vaccine in obese children and adolescents

BackgroundLimited data are currently available regarding the cellular immune response to a live attenuated hepatitis A virus (HAV) vaccine, especially in children with obesity. The objective of this retrospective cohort study was to compare the activation of antigen-specific interferon (IFN)-γ+ T cells in obese children and adolescents with healthy individuals before and after immunization with a single dose of live attenuated HAV vaccine. MethodsBlood samples were obtained from the 2021 study by Dumrisilp et al. investigating the immunogenicity of the live attenuated hepatitis A vaccine in children and young adults. Prior to enrollment, all 212 subjects had never received any HAV vaccine and tested negative for anti-HAV antibodies. The participants were vaccinated with a freeze-dried, live attenuated HAV vaccine of the H2 strain. In this study, we analyzed the stored peripheral blood mononuclear cells (PBMCs) obtained from a subgroup of 30 obese subjects and 30 normal-weight healthy controls of the same age and sex. PBMCs were collected before and 8–9 weeks after HAV vaccination for further analysis. These cells were stimulated with a recombinant antigen derived from HAV-VP3, and the immune response was evaluated using the IFN-γ enzyme-linked immunospot (ELISpot) assay. ResultsThe between-group analysis indicated that the T-cell response of obese participants was comparable to that of normal-weight controls both before and after vaccination. The change in IFN-γ production from before to after vaccination in the obese group was not significantly different from that of the control group. Additionally, in the obese group, no correlation was found between IFN-γ production and clinical characteristics such as sex, body mass index, waist circumference, and acanthosis nigricans. ConclusionTesting for cellular immune response provides a comprehensive understanding of the overall immune response to vaccination. This study, the first to explore this significant aspect, suggests that obesity does not affect the short-term cellular immune response to live attenuated HAV vaccination.

Significant hemolysis is present during irreversible electroporation of cardiomyocytes in vitro

BackgroundPulsed field ablation (PFA) of atrial fibrillation is a new method in clinical practice. Despite a favorable safety profile of PFA in atrial fibrillation ablation, rare cases of renal failure, probably due to hemolysis, have recently been reported. ObjectiveThe aim of this study was to determine the rate of hemolysis and cardiac cell death during in vitro PFA with different electric field intensities. MethodsBlood samples from healthy volunteers and mouse HL-1 cardiomyocyte cell lines were subjected to in vitro irreversible electroporation using 216 bipolar pulses, each lasting 2 μs with intervals of 5 μs, repeated 20 times at a frequency of 1 Hz. These pulses varied from 500 V to 1500 V. Cell-free hemoglobin levels were assessed spectrophotometrically, and red blood cell microparticles were evaluated by flow cytometry. Cardiomyocyte death was quantified with propidium iodide. ResultsPulsed field energy (1000 V/cm, 1250 V/cm, and 1500 V/cm) was associated with a significant increase in cell-free hemoglobin (0.32 ± 0.16 g/L, 2.2 ± 0.96 g/L, and 5.7 ± 0.39 g/L; P < .01) and similar increase in the concentration of red blood cell microparticles. Significant rates of cardiomyocyte death were observed at electric field strengths of 750 V/cm, 1000 V/cm, 1250 V/cm, and 1500 V/cm (26.5% ± 5.9%, 44.3% ± 6.2%, 55.5% ± 6.9%, and 74.5% ± 17.8% of cardiomyocytes; P < .01). ConclusionThe most effective induction of cell death in vitro was observed at 1500 V/cm. This intensity was also associated with a significant degree of hemolysis.

Vaccine‐induced humoral response of BNT162b2 and mRNA-1273 against BA.1, BA.5, and XBB.1.5. (sub)variants 6 months after a homologous booster: is immunogenicity equivalent?

IntroductionSome studies suggest that the monovalent mRNA-1273 vaccine is more effective than BNT162b2 in producing higher levels of antibodies. However, limited data are available, and the methods used are not directly comparable. Material and methodsBlood samples were obtained before the booster (third dose) and after 14, 90, and 180 days in two similar cohorts who received the original BNT162b2 or mRNA-1273 vaccine designed to target wild type SARS-CoV-2. The aim of our study is to compare their effectiveness by assessing the levels of binding and neutralizing antibodies specifically against each of the BA.1 variant, BA.5 variant, and the XBB.1.5 subvariant. ResultsOnce the peak was reached after two weeks, a drastic decline in binding and neutralizing antibodies was observed up to 6 months after the homologous booster administration. The humoral response was however more sustained with the mRNA-1273 booster, with half-lives of 167, 55, and 48 days for binding, BA.1, and BA.5 neutralizing antibodies compared to 144, 30, and 29 days for the BNT162b2 booster, respectively. Compared to the BA.1 variant, the neutralizing capacity was significantly decreased at 6 months with the BA.5 variant (fold-decrease: 1.67 to 3.20) and the XBB.1.5. subvariant (fold-decrease: 2.86 to 5.48). ConclusionAlthough the decrease in the humoral response was observed with both mRNA vaccines over time, a more sustained response was observed with the mRNA-1273 vaccine. Moreover, the emergence of Omicron-based variants causes a reduced neutralizing capacity, notably with the XBB.1.5. subvariant. The administration of subsequent boosters would therefore be needed to restore a sufficiently high neutralizing response.

Differential inflammatory profiles in carriers of reciprocal 22q11.2 copy number variants

BackgroundGenetic copy number variants (CNVs; i.e., a deletion or duplication) at the 22q11.2 locus confer increased risk of neuropsychiatric disorders and immune dysfunction. Inflammatory profiles of 22q11.2 CNV carriers can shed light on gene-immune relationships that may be related to neuropsychiatric symptoms. However, little is known about inflammation and its relationship to clinical phenotypes in 22q11.2 CNV carriers. Here, we investigate differences in peripheral inflammatory markers in 22q11.2 CNV carriers and explore their relationship with psychosis risk symptoms and sleep disturbance. MethodsBlood samples and clinical assessments were collected from 22q11.2 deletion (22qDel) carriers (n=45), 22q11.2 duplication (22qDup) carriers (n=29), and typically developing (TD) control participants (n=92). Blood plasma levels of pro-inflammatory cytokines, including interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ), and anti-inflammatory cytokine interleukin-10 (IL-10) were measured using a MesoScale Discovery multiplex immunoassay. Plasma levels of C-reactive protein (CRP) were measured using Enzyme-linked Immunosorbent Assay (ELISA). Linear mixed effects models controlling for age, sex, and body mass index were used to: a) examine group differences in inflammatory markers between 22qDel, 22qDup, and TD controls, b) test differences in inflammatory markers between 22qDel carriers with psychosis risk symptoms (22qDelPS+) and those without (22qDelPS-), and c) conduct an exploratory analysis testing the effect of sleep disturbance on inflammation in 22qDel and 22qDup carriers. A false discovery rate correction was used to correct for multiple comparisons. Results22qDup carriers exhibited significantly elevated levels of IL-8 relative to TD controls (q<0.001) and marginally elevated IL-8 levels relative to 22qDel carriers (q=0.08). There were no other significant differences in inflammatory markers between the three groups (q>0.13). 22qDelPS+ exhibited increased levels of IL-8 relative to both 22qDelPS- (q=0.02) and TD controls (p=0.002). There were no relationships between sleep and inflammatory markers that survived FDR correction (q>0.14). ConclusionOur results suggest that CNVs at the 22q11.2 locus may have differential effects on inflammatory processes related to IL-8, a key mediator of inflammation produced by macrophages and microglia. Further, these IL-8-mediated inflammatory processes may be related to psychosis risk symptoms in 22qDel carriers. Additional research is required to understand the mechanisms contributing to these differential levels of IL-8 between 22q11.2 CNV carriers and IL-8’s association with psychosis risk.

Role of Donor-derived Cell-free DNA In Predicting Short-term Allograft Health In Liver Transplant Recipients

BackgroundPredicting allograft dysfunction prior to clinical or biochemical evidence remains one of the challenges in transplantation, and a preclinical detection and early management of its cause allows for improved post-transplant outcomes. Donor derived cell-free DNA (ddcfDNA) has been proposed as an important biomarker of allograft injury and has shown to predict dysfunction prior to any biochemical derangements. We aimed to investigate the diagnostic performance of ddcfDNA in detecting and differentiating the causes of early pre-biochemical detection of graft injury and in predicting the short-term outcomes of graft health using a patented protocol and proprietary set of single nucleotide polymorphisms. MethodsBlood samples were collected on defined postoperative days (1-, 3-, 7- and at 3-months) and were analysed through relatively economical patented protocol (TrunomeTM). Biopsy, biochemical tests, and clinical criteria were analysed between various subgroups. ResultOf a total 50 patients, percentage ddcfDNA (%ddcfDNA) levels were significantly elevated in the rejection group (n=8) as compared to the non-rejection group (n= 42; median elevation 12.8% vs 4.3% respectively) with a significant correlation (r = 0.92, p<0.0001). AUC-ROC analysis revealed that the %ddcfDNA levels can predict graft health more precisely when compared to the conventional liver function tests (AUC for %ddcfDNA- 0.86; p<0.001; AUC for AST-0.65, p=0.08; AUC for ALT- 0.75, p<0.01). Moreover, %ddcfDNA levels (with a threshold >10.2%) on post-operative day 7 accurately predicted short-term (3-months) health status of the graft with 93.33% sensitivity, 94.44% specificity, 87.50% positive predictive value, 97.14% negative predictive value and 94.12% accuracy. ConclusionA single time point ddcfDNA on postoperative day 7 accurately predicts graft health and improves risk stratification in the short-term.

Blood Sampling and Hormone Measurement for Determining the Stage in the Ovarian Cycle in Marmosets.

Common marmosets are small New World monkeys. Since many of their biological mechanisms are similar to those of humans, marmosets are potentially useful for medical and human biology research across a range of fields, such as neuroscience, regenerative medicine, and development. However, there is a lack of literature describing methods for many basic experiments and procedures. Here, detailed methods for determining the levels of sex hormones (progesterone, estradiol, and chorionic gonadotropin) in marmosets are described. The measurement of these hormones enables the prediction of the stage in the ovarian cycle, which is typically 26-30 days in marmosets; accurate determination is essential for the harvesting of oocytes/zygotes at the correct time point and for the preparation of host females for the generation of genetically modified marmosets. Additionally, the measurement of sex hormone levels is useful for endocrinology, ethology, early development, and reproductive biology studies. This protocol provides a detailed description of the methods for blood sampling from the femoral vein, separation of plasma for hormone measurement, measuring chorionic gonadotropin levels using urine and plasma, resetting the ovarian cycle using injections of a prostaglandin F2α analog to shorten and synchronize the cycle, and promoting follicular growth and ovulation by injecting follicle-stimulating hormone and chorionic gonadotropin. Using these protocols, the stages in the ovarian cycle can be determined for the timely collection of oocytes/zygotes.

P-522 Impact of TGFß/BMP/SMAD Pathway on Premature Ovarian Insuffiency (POI)

Abstract Study question Is TGFß/BMP/SMAD pathway altered in women with POI and can results be detected from peripheral blood samples? Summary answer The TGFß/BMP/SMAD pathway mRNA-expression-alterations are detectable in leukocytes of women with Premature Ovarian Insufficiency (POI) paving the way to use it as predictive tool. What is known already A proper ovarian function relies on a complex system of signalling molecules including several BMP/TGFß-superfamily-members expressed in the ovary at different stages of folliculogenesis acting via two types of Serine/Threonine Kinase receptors. The downstream located SMAD proteins are classified into three subclasses according to their function as activated R-SMADs1,2,3,4,9, common-partner SMAD4 or inhibitory SMADs6,7. Mutations in BMPR2 and its ligands have been shown to be related to POI. Recently neurologic studies demonstrated that FMRP(Fragile-X-Mental-Retardation-1-Protein) represses BMPR2 and its gene FMR1(Fragile X Mental-Retardation-1) is one major regulator of follicular development and the most frequent genetic cause of POI if premutated. Study design, size, duration RT-q-PCR was performed to measure the mRNA-levels of SMAD 1,3,4,5, BMPR2 and FMR1. cDNA was synthesized using RNA obtained from leucocytes of patients with POI and of the control group with a normal ovarian reserve. 163 patients diagnosed with POI and 51 women in the control group were recruited between 2008 and 2022 at Heidelberg University’s Women’s Hospital. Participants/materials, setting, methods Blood samples were drawn from every participant to extract leucocyte-RNA and synthesize non-specific cDNA. RT-q-PCR was performed with TaqMan reagents to quantify relative mRNA-expression levels of SMAD 1,3,4, 5, BMPR2 and FMR1. Statistical analysis was performed with Microsoft Excel and GraphPad Prism 10 using the 2–ΔΔCt method and Mann-Whitney-Test. Statistical significance was set to p = 0.05. Main results and the role of chance In our investigation, we observed a consistent and statistically significant decrease in the expression levels of all tested SMADs—SMAD1, SMAD3, SMAD4, and SMAD5—in leukocytes of women diagnosed with POI compared to unaffected controls. The p-values amounted to p &amp;lt; 0.0001 for SMAD1, p = 0.0340 for SMAD3, p &amp;lt; 0.0001 for SMAD4, and p = 0.0076 for SMAD5. However, we did not identify notable differences in the expression levels of FMR1 and BMPR2 between the two groups. These results suggest an alteration in the regulation of the activated downstream elements SMAD 1, 3, and 5 and their common partner SMAD4 in women with POI, emphasizing its potential role as a contributing factor to diminished ovarian function. These effects seem to be independent of BMPR2 and FMR1 and regulated by other receptors and or ligands. Limitations, reasons for caution Our data are limited to mRNA-levels. So further experiments including protein-level and their phosphorylation-status are needed to receive an overall assessment of its impact on ovarian damage. It’s worth noting that the study is concentrated on leukocytes as potential diagnostic tool and results should be cross-checked in ovarian-tissue and cells. Wider implications of the findings The identified alteration in TGFß/BMP/SMAD pathway gene expression in women with POI suggest a role in ovarian damage. Lower expression levels in the aforementioned mRNAs could serve as a potential predictive biomarker for POI risks and putatively used in consultations of women prior to fertility treatment and/or preservation. Trial registration number not applicable

Diurnal fluctuations in biochemical parameters related to calcium homeostasis – the Bispebjerg study of diurnal variations

Purpose This aim of this study was to assess the possible association between diurnal oscillations and biochemical markers associated with calcium homeostasis. This included the markers parathyroid hormone (PTH), total calcium, total alkaline phosphatase, phosphate, and 25-hydroxyvitamin D (25-OH-D). By examining the influence of circadian rhythms on these parameters, the study aimed to deepen the understanding of calcium metabolism dynamics and its clinical implications. Patients and methods Blood samples from 24 Caucasian male volunteers aged 20 to 40 (mean age 26) with normal pulse, blood pressure, and BMI were analyzed for biochemical markers related to calcium homeostasis. Data was obtained from the Bispebjerg study of diurnal variations. Blood samples were collected every three hours over a 24-hour period. Patients were fasting from 22:00 to 09:00. The participants spent 24 h in the hospital ward, receiving regular meals and engaging in low-intensity activities. They experienced 15 h of daylight and 9 h of complete darkness during sleep. Diurnal oscillations were analyzed using cosinor analysis with statistical significance set at p < 0.05. Results Total calcium, phosphate, and PTH exhibited significant diurnal variations. Total calcium and PTH were inversely synchronized while PTH and phosphate oscillated in synchronization. The three parameters showed relatively large amplitude/reference range ratios from 25.4% to 41.5%. Conclusion This study found notable fluctuations in total calcium, phosphate, and PTH levels over a 24-hour cycle, while 25-OH-D and total alkaline phosphatase remained consistent. It highlights the importance of considering sampling times for total calcium, PTH, and phosphate in clinical settings.

Effects of dabigatran, rivaroxaban, and apixaban on fibrin network permeability, thrombin generation, and fibrinolysis

Introduction There are important pharmacological differences between direct oral anticoagulants (DOAC) and a deeper knowledge of how they influence different aspects of hemostasis in patients on treatment is desirable. Materials and methods Blood samples from patients on dabigatran (n = 23), rivaroxaban (n = 26), or apixaban (n = 20) were analyzed with a fibrin network permeability assay, a turbidimetric clotting and lysis assay, the calibrated automated thrombogram (CAT), plasma levels of thrombin-antithrombin complex (TAT) and D-dimer, as well as DOAC concentrations, PT-INR and aPTT. As a comparison, we also analyzed samples from 27 patients on treatment with warfarin. Results Patients on dabigatran had a more permeable fibrin network, longer lag time (CAT and turbidimetric assay), and lower levels of D-dimer in plasma, compared with patients on rivaroxaban- and apixaban treatment, and a more permeable fibrin network than patients on warfarin. Clot lysis time was slightly longer in patients on dabigatran than in patients on rivaroxaban. Warfarin patients formed a more permeable fibrin network than patients on apixaban, had longer lag time than patients on rivaroxaban (CAT assay), and lower peak thrombin and ETP compared to patients on treatment with both FXa-inhibitors. Conclusions Results from this study indicate dabigatran treatment is a more potent anticoagulant than apixaban and rivaroxaban. However, as these results are not supported by clinical data, they are probably more related to the assays used and highlight the difficulty of measuring and comparing the effect of anticoagulants.

Assessment of prevalence of elevated blood lead levels and risk factors among children and pregnant women in Bihar, India

BackgroundWhile modeled estimates and studies in contaminated areas indicate high lead exposure among children in Bihar, India, local data on lead exposure in the child population is limited. ObjectivesTo characterize lead exposure, and assess potential sources of lead exposure among a state-representative sample of children and their pregnant mothers residing in Bihar. MethodsBlood samples were collected from 697 children under five and 55 pregnant women from eight districts in Bihar. Blood lead levels were determined using capillary blood and a portable lead analyzer. Household demographics, home environment, behavior, and nutrition information were collected through computer-assisted personal interviews with primary caregivers. Logistic regression was used to assess associations between potential risk factors and elevated blood lead levels. ResultsMore than 90% of children and 80% of pregnant women reported blood lead levels ≥5 μg/dL. Living near a lead-related industry and pica behavior of eating soil were significantly associated with increased odds of having elevated blood lead levels. Additional risk factors for having a blood level ≥5 μg/dL included the use of skin lightning cream (aOR = 5.11, 95%CI: 1.62, 16.16) and the use of eyeliners (aOR = 2.81, 95%CI: 1.14, 6.93). Having blood lead levels ≥10 μg/dL was also significantly associated with the household member who had an occupation or hobby involving the use of lead (aOR = 1.75, 95%CI: 1.13, 2.72). DiscussionElevated blood lead levels were prevalent among children and pregnant women in Bihar, indicating the urgent need for a comprehensive lead poisoning prevention strategy.

Consistency analysis of two fingertip capillary blood sampling methods for complete blood count

This study was performed to analyze fingertip capillary blood sampling in pediatric patients using microcapillary blood collection tubes and microhematocrit tubes and to compare the blood cell analysis results obtained via these two blood collection methods. Finger capillary blood was collected from 110 outpatients using microcapillary blood collection tubes and microhematocrit tubes and complete blood count analysis was performed with a Sysmex XS-900i hematology analyzer and manual microscopy for blood cell morphology. Paired data was evaluated for agreement and bias using the microhematocrit samples as the reference group and the samples from the microcapillary blood collection tubes as the observation group. The two blood collection methods demonstrated good agreement for measuring red blood cell (RBC) parameters (i.e., RBC, Hb, Hct, MCV, MCH and MCHC), wherein the relative bias was > allowable total error (TEa) in 0.91%, 1.82%, 11.82%, 1.82%, 0.91% and 8.18% of the parameter measures, respectively. According to industry requirements, the proportion of samples meeting the acceptable bias level should be > 80%. Additionally, the estimated biases at each medical decision level were within clinically acceptable levels for RBC, Hb, Hct, and MCV. However, the proportion of WBC and PLT counts with relative bias > TEa was 25.45% and 35.45%, respectively. Furthermore, the relative bias of the WBC count at the medical decision level of 0.5 × 109/L and that of the PLT counts at the medical decision levels of 10 × 109/L and 50 × 109/L were clinically significant. Bland–Altman analysis further showed a mean bias of 0.66 × 109/L (95% LoA, − 0.79 to 2.11) for the WBC count and 39 × 109/L (95% LoA, − 46 to 124) for the PLT count from the blood samples collected in the microcapillary blood collection tubes compared with the counts of those collected in the microhematocrit tubes. Neutrophil, monocyte, lymphocyte, eosinophil, and PLT counts increased significantly in the microcapillary blood collection tubes compared with those in the microhematocrit tubes, along with an elevated number of instrument false alarms (P < 0.05). The two capillary blood collection devices exhibit performance differences. Therefore, clinicians should pay attention to the variation in results caused by different blood collection methods.

Serum NfL and GFAP are weak predictors of long-term multiple sclerosis prognosis: A 6-year follow-up

BackgroundSerum neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP) are promising biomarkers that might be associated with clinical and radiological markers of multiple sclerosis (MS) severity. However, it is not known whether they can accurately identify patients at risk of disability progression in the medium and long term. ObjectivesWe wanted to determine the association between sNfL and sGFAP, Expanded Disability Status Scale score changes, and conversion to secondary progressive MS (SPMS) in a cohort of 133 patients with relapsing remitting MS. MethodsBlood samples were collected at inclusion to measure SNfL and sGFAP by single molecule array and their prognostic value was assessed using a linear mixed model. ResultsIn this cohort, 37 patients (27.8 % of 133) experienced disability progression and 12 patients (9.0 %) converted to SPMS during the follow-up (mean follow-up duration: 6.4 years). Only sNfL (p = 0.03) was associated with conversion to SPMS, and neither SNfL nor sGFAP was associated with disability progression. ConclusionSerum NfL and GFAP do not seem to accurately predict MS outcome in the long term. More studies are needed to determine how serum biomarkers, associated with other clinical and MRI biomarkers, might be used to improve MS prognostication.

BIOM-01. CARBONIC ANHYDRASE IX INTERPLAY WITH LOCAL AND SYSTEMIC IMMUNE RESPONSE IN PEDIATRIC BRAIN TUMORS

Abstract BACKGROUND Brain tumors (BT) remain the leading cause of death among childhood cancers. There is an urgent need to identify novel targets in these patients. BT progression depends on the adaptive responses to hypoxia. Carbonic anhydrase IX (CAIX) helps cancer cells survive in a hypoxic environment and can be potentially as a therapeutic target. The role of CAIX in the interplay between the tumor and immune cells requires in-depth investigation to design effective therapies. METHODS Fresh BT, blood, and intraoperative cerebrospinal fluid (iCSF) samples were collected prospectively from pediatric patients, who underwent brain tumor surgery at Vilnius University Hospital Santaros Klinikos from March 2022 to December 2023. All tumor types were included. Immunohistochemistry (IHC) and flow cytometry (FC) were performed to analyze the expression of CAIX and selected immunological markers in the fresh tumor, iCSF and peripheral blood mononuclear cells (PBMCs). Soluble CAIX content in serum and iCSF was assessed by ELISA. In addition, a retrospective analysis of previously diagnosed pediatric glioma specimens was performed by IHC. RESULTS Twelve patients with heterogeneous BT types have been enrolled prospectively and 17 patients with glioma were reviewed retrospectively. Glioma accounted for 72% (21/29) of all samples. IHC showed that 57% of gliomas were positive for CAIX as well as a moderate correlation between the CAIX score and the expression of CD44, CD206, and CCR2 markers. PD-1/PD-L1 expression was negligible. In the prospective cohort, 83% (10/12) of BT tissue was positive for CAIX by IHC and FC. Soluble CAIX was detectable in 40% of iCSF samples, but not in serum. We found a correlation trend of CAIX expression in the fresh tumor cells and iCSF with increased CCR2 expression on CD45+ PBMCs. CONCLUSIONS This interim analysis showed a correlation between CAIX tumor expression and immune biomarkers, prompting further investigation by enrolling more patients.

Evaluation of the neuroprotective effect of antipsychotics by serum quantification of protein S100B

ObjectiveThis research delves into the intricate interplay between antipsychotic medications and neuroprotection focusing on the S100B protein—a central player in the regulation of neuroapoptotic activity. MethodBlood samples were collected to assess serum S100B protein levels using an immunoassay of immunoelectrochemiluminescence. The first two samples were collected with a 3-month interval between each, and the third sample was obtained 6 months after the previous one. Changes in S100B protein levels throughout the study were assessed using Friedman's ANOVA test. This was followed by the Wilcoxon signed-rank test with Bonferroni correction to account for multiple comparisons. ResultsThis study involved 40 patients diagnosed with severe mental disorders (34 schizophrenia, 4 schizoaffective disorder, 1 bipolar disorder, and 1 borderline personality disorder). These patients had been receiving antipsychotic treatment for an average duration of 17 years. The results revealed that the S100B protein remained within physiological levels (median values 39.0 ng/L for the first sample, median values 41.0 ng/L for the second sample, and median values 40.5 ng/L for the third sample) with no significant changes (p = 0.287), with all anti-psychotic medicaments values consistently below 50 ng/L, a lower value compared to maximum range of 105 ng/L. Importantly, there were no significant differences in S100B protein levels between patients on monotherapy and those on combination antipsychotic therapy (p = 0.873), suggesting that combination therapy did not increase neuroapoptotic activity. ConclusionsThese findings provide compelling evidence for the potential neuroprotective effects of long-term antipsychotic treatment in individuals with severe mental disorders. By maintaining physiological levels of the S100B protein, antipsychotic medications may help protect against neuronal damage and dysfunction. This research contributes valuable insights into the neuroprotective mechanisms of antipsychotic drugs, enhancing our understanding of their potential benefits in the treatment of severe mental disorders.

Toxicology findings from drivers suspected of drug-impaired driving in Ontario (2008–2019)

Objective This study examines the results of toxicological tests performed on blood and urine samples collected from suspected drug-impaired drivers in Ontario from 2008 to 2019. The report examines the results of toxicological analysis of the samples submitted, the characteristics of those drivers from whom samples were collected, and the temporal and situational circumstances that led to police investigations and sample collection to better understand drug-impaired driving behavior and to assist in the development and implementation of countermeasure strategies and programs. Methods Blood and urine samples were sent to the Center of Forensic Sciences where they were analyzed using standardized comprehensive toxicological analysis to test for a wide variety of potentially impairing drugs. Demographic and temporal information for each case from which a sample was collected were also examined to describe the circumstances and characteristics of these driving incidents. Results During the 12-year period examined, 5,388 samples collected from suspected drug-impaired drivers were analyzed. The number of samples collected increased substantially following the implementation of the Drug Evaluation and Classification Program (DECP) in July 2008, the enactment of legislation facilitating the collection of blood samples from suspects, and the legalization of cannabis for nonmedical purposes in 2018. The number of samples submitted shows temporal correlation with the number of police officers certified as Drug Recognition Experts (DRE) in the province. Over the 12-year period of this study, cannabis was the most frequently detected substance in drivers (52.8% of cases), followed by cocaine (44.3%) and methamphetamine (24.8%). In 80% of cases, more than one substance was detected. Conclusions Examining the characteristics of suspected drug-impaired drivers, the temporal circumstances, and the drug findings throughout the large geographic area of Ontario and over the extended period of this study enhances our understanding of drug-impaired driving behavior. These characteristics can assist in the development and/or evaluation of enforcement strategies and enhanced countermeasure activities.

Extending culture time to improve Mitotic Index for cytogenetic dosimetry

Purpose To analyze the effects of extending lymphocyte cultivation time on the Mitotic Index, frequency of first-division cells, and dose estimation after irradiating blood samples with different doses of radiation. Materials and methods Blood samples from two healthy male volunteers were separately irradiated with three doses (3, 5, and 6 Gy) using a 60Co gamma source (average dose rate: 1.48 kGy.h−1) and cultivated in vitro for conventional (48 h) and extended (56, 68, and 72 h) amounts of time. Colcemid (0.01 µg.mL−1) was added at the beginning of the culture period. Cells were fixed, stained with fluorescence plus Giemsa (FPG), and analyzed under a light microscope. The effects of prolonged culture duration on the Mitotic Index (MI), frequency of first-division cells (M1 cells), and the First-Division Mitotic Index (FDMI) were investigated. The estimation of delivered doses was conducted using a conventional 48h-culture calibration curve. Results Overall, cells presented higher MI (up to 12-fold) with the extension of culture, while higher radiation doses led to lower MI values (up to 80% reduction at 48 h). Cells irradiated with higher doses (5 and 6 Gy) had the most significant increase (5- to 12-fold) of MI as the cultivation was prolonged. The frequency of M1 cells decreased with the prolongation of culture for all doses (up to 75% reduction), while irradiated cells presented higher frequencies of M1 cells than non-irradiated ones. FDMI increased for all irradiated cultures but most markedly in those irradiated with higher doses (up to 10-fold). The conventional 48h-culture calibration curve proved adequate for assessing the delivered dose based on dicentric frequency following a 72-hour culture. Conclusion Compared to the conventional 48-hour protocol, extending the culture length to 72 hours significantly increased the Mitotic Index and the number of first-division metaphases of irradiated lymphocytes, providing slides with a better scorable metaphase density. Extending the culture time to 72 hours, combined with FPG staining to score exclusively first-division metaphases, improved the counting of dicentric chromosomes. The methodology presented and discussed in this study can be a powerful tool for dicentric-based biodosimetry, especially when exposure to high radiation doses is involved.

Differential GPR56 Expression in T Cell Subpopulations for Early-Stage Lung Adenocarcinoma Patient Identification

ABSTRACT Objective This study aimed to investigate the expression of GPR56 in the T cells of early-stage lung adenocarcinoma (LUAD) patients and clarify its diagnostic significance. Methods Blood samples were collected from 32 patients with stage IA LUAD and 31 healthy controls. GPR56 and perforin were analysed in circulating T-cell subsets by flow cytometry. In addition, a correlation between perforin and GPR56 expression was detected. Changes in GPR56+ cells in early LUAD patients were analysed, and the diagnostic significance of GPR56+ T cells for early LUAD was studied by receiver operating characteristic (ROC) curve analysis. Results The expression of GPR56 in CD8+ T cells from early-stage LUAD patients was significantly greater than that in CD4+ T cells. The percentage of perforin-positive GPR56+ cells in early-stage LUAD patients was high. GPR56 levels in the T cells of LUAD patients were significantly lower than those in healthy controls. ROC analysis revealed that the area under the curve for the percentage of GPR56-positive CD8+ TEMRA cells to distinguish early-stage LUAD patients from healthy individuals- reached 0.7978. Conclusion The decreased expression of GPR56 in the peripheral blood of early-stage LUAD patients correlated with perforin levels, reflecting compromised antitumor immunity and aiding early-stage LUAD screening.

#668 Combined biomarker testing for the assessment of acute kidney injury among myocardial infarction patients

Abstract Background and aims The utility of different biomarkers for the prediction of acute kidney injury following ST-elevation myocardial infarction (STEMI) has been evaluated in several studies however, very few data exist on the prognostic value of combining biomarkers. Among numerous biomarkers examined, C-reactive protein (CRP) and Neutrophil gelatinase associated Lipocalin (NGAL) stand high in their utility to assess AKI risk. The aim of this study was to assess the prognostic value of combined CRP and NGAL measurement for the assessment of the risk of AKI in reperfused STEMI patients Methods Blood samples for plasma NGAL and CRP were drawn 24 hours after admission. High NGAL and CRP were defined as values within the 4th quartile (&amp;gt;75percentile). Patients were stratified into 4 study groups: Low NGAL/Low CRP, LOW NGAL/high CRO, High NGAL /low CRP and high NGAL/high CRP. Patients were assessed for the occurrence of in-AKI based on the KDIGO criteria. Results A total of 311 patients were included (mean age 66 ± 14 years, 81% males). According to the 4 study groups, there was a stepwise increase in the proportion of AKI (9% vs. 21% vs. 40% vs. 68%; p 0.001; Fig. 1). In a multivariate regression model, compared to patients having low NGAL/low CRP we observed a graded, independent increase in the risk for AKI for low NGAL/high CRP, high NGAL/low CRP and high NGAL/high CRP(OR 2.89, p = 0.02 vs. OR 3.29, p &amp;lt; 0.001 vs. OR 17.6, p &amp;lt; 0.001 respectively, Table 1). Peak concentrations showed significant areas under the curves (AUCs) for the prediction of AKI for both CRP AUC (0.692, 95% CI 0.62-0.76) and NGAL (AUC 0.778, 95% CI 0.72-0.84). The combination of NGAL and CRP yielded a significant increase in AUC to 0.822 (95% CI 0.77 to 0.88) (combined biomarkers vs CRP: p &amp;lt; 0.001 and combined vs. NGAL: p = 0.02; Fig. 2). Conclusions In patients with reperfused STEMI, the combined assessment of NGAL and CRP provided incremental prognostic information for the prediction AKI when compared with single-biomarker measurement.

Distribution of ABO, Rh, and MNS blood groups from students in Walailak University, Thailand: A descriptive cross-sectional study

BackgroundThe presence or absence of blood group antigens can lead to alloimmunization, causing hemolytic transfusion reactions and hemolytic disease of the fetus and newborn. The distribution of these antigens varies among different ethnic populations, making it essential to study their prevalence in specific regions. ObjectivesThis study aimed to investigate ABO, Rh, and MNS blood group antigens, phenotypes, and allele frequencies among university students in Southern Thailand. Materials and methodsBlood samples were collected and typed for blood group antigens using serological tests. Antigen, phenotype, and allele frequencies were calculated based on the observed phenotypes. ResultsThe most common ABO blood group was O (37.29%), followed by B (35.98%), A (19.47%), and AB (7.26%). Allele frequencies for ABO system were found to be 0.6107 for ABO*O, 0.2453 for ABO*B, and 0.1427 for ABO*A. Rh blood group distribution revealed a high prevalence of the D antigen (99.01%), followed by e (96.37%), C (95.05%), c (39.27%), and E (29.37%). The allele frequencies for D and d were 0.9005 and 0.0995, respectively. The MNS system showed M and N antigen frequencies of 85.81% and 67.66%, respectively. The most common phenotype was M+N+ (53.47%), followed by M+N− (32.34%) and M−N+ (14.19%). Mia antigen was present in 10.56% of our population. ConclusionsThis study provides an understanding of blood group distribution and gene frequency in our young population, which is crucial for blood management in community transfusion. Additionally, this knowledge holds significant value for population genetics studies.

ICU NURSES AND VAD ADHERENCE TO INS 2021

Abstract Background Continuous updates and evidence–based nursing practices are vital for quality patient care. In 2021, the Infusion Nursing Standard (INS) have updated vascular access device (VAD) guidelines. Evaluating nurses’ VAD knowledge according to these guidelines will improve caring quality, particularly for ICU patients that need a functioning VAD. Aim To evaluate the knowledge of ICU nurses regarding VADs according to the latest INS 2021 guidelines. Material and methods A cross–sectional study was conducted on a sample of nurses providing direct care to ICU patients. A web survey was administered using the “Nurses’ Knowledge and Self–Efficacy on Vascular Access” instrument, which assesses nurses’ knowledge and self–efficacy in relation to short (SPC) and long peripheral canula (LPC), PICC, and PORT, as for INS 2021 guidelines. The survey consisted of 32 items with four closed–answer options, of which only one was correct. Competence and skill levels were calculated by summing the correct responses and standardizing the result to 100. Results Some 35 nurses have enrolled, with an average age of 37 years (SD= 10.80), including 16 men and 19 women, working in UTIC, ICU and Stroke Unit. The highest adherence score (97%) has been registered for the PICC flushing technique before and after each use. The lowest result (14%) has been obtained regarding the performing method of blood sampling from a central VAD. Other positive results have been achieved in the selection of the correct placement sites for all VAD types, with score exceeding 90%. The second part of the questionnaire, which focused of nurses’ self–confidence, showed that nurses felt most confident in SPC section, followed by the PORT section, LPC, and lastly, PICC. Conclusions ICU nurses have a good level of competence and self–efficacy about the VAD management, adhering to the 2021 INS guidelines, although gaps have been identified in the management of SPC and some techniques and despite the high self–confidence expressed directly by nurses. Healthcare organizations should continuously monitor the ICU nurses’ competencies and skills level to implement targeted intervention and ensure the application of evidence in clinical practice, thereby improving the patients’ quality of care.