Routine Blood Samples Research Articles

Quantitative PCR Measurement of miR-371a-3p and miR-372-p Is Influenced by Hemolysis.

Cell-free microRNAs have been reported as biomarkers for several diseases. For testicular germ cell tumors (GCT), circulating microRNAs 371a-3p and 372-3p in serum and plasma have been proposed as biomarkers for diagnostic and disease monitoring purposes. The most widely used method for quantification of specific microRNAs in serum and plasma is reverse transcriptase real-time quantitative PCR (RT-qPCR) by the comparative Ct-method. In this method one or several reference genes or reference microRNAs are needed in order to normalize and calculate the relative microRNA levels across samples. One of the pitfalls in analysis of microRNAs from serum and plasma is the release of microRNAs from blood cells during hemolysis. This is an important issue because varying degrees of hemolysis are not uncommon in routine blood sampling. Thus, hemolysis must be taken into consideration when working with circulating microRNAs from blood. miR-93-5p, miR-30b-5p, and miR-20a-5p have been reported as reference microRNA in analysis of the miR-371a-373 cluster. We here show how these three microRNAs are influenced by hemolysis. We also propose a new reference microRNA, miR-191-5p, which is relatively stable in serum samples with mild hemolysis. In addition, we show how hemolysis can have effect on the reported microRNA levels in patient samples when these reference microRNAs are used in samples with varying levels of hemolysis.

Comparison of preoperative serum VEGF in leiomyosarcoma and uterus myomatosus patients: A proof of concept study.

e17117 Background: The preoperative differentiation of a uterine fibroid from a sarcoma is still a pending clinical problem. Currently with the suspicion of a sarcoma (LMS) the more invasive open surgical approach is recommended to minimise the risk of LMS fragmentation and distribution. In a minority of cases, a malignancy is indicated by clinical suspicion, pre-surgical imaging or routine serum blood samples (SBS). Our hypothesis postulates higher VEGF levels in LMS compared to fibroid patients in the pre-surgical SBS. To assess this hypothesis, SBS were taken from patients with the clinical suspicion of LMS after informed consent and analysed after histology confirmed the diagnosis. Methods: Case series of patients with suspected LMS over a 4year time period. Analysis was performed via SBS collected from LMS- and fibroid patients before surgery. Serum VEGF protein was measured by ELISA. The final tumor histology was obtained from the report of the institutional pathologist. VEGF-serum levels were then compared between fibroid and LMS patients using the non-parametric Mann-Whitney U test. Results: 25 patient SBS were collected prior to surgery. In 9 cases the histopathology confirmed a LMS, with heterogeneous pTNM classifications. On average, VEGF serum levels were higher in the LMS patients as compared to the fibroid patients (628,96 pg/ml vs 351,91 pg/ml; further statistics see Table 1); however, the difference was not statistically significant (Mann-Whitney-U Test, p = 0.141). Conclusions: This proof-of-concept study with a small sample size of pre-surgical SBS indicates that VEGF serum levels may be increased in patients with histologically confirmed LMS; however, larger sample sizes are needed to validate our findings. If additional studies confirm a pronounced increase in pre-surgical serum VEGF levels in LMS patients, serum VEGF levels might routinely be used to assess the risk for a LMS in patients presenting with clinical uterine fibroids. [Table: see text]

Inflammatory, Serological and Vascular Determinants of Cardiovascular Disease in Systemic Lupus Erythematosus Patients.

Background and aim: Systemic lupus erythematosus (SLE) is associated with increased risk of cardiovascular disease (CVD). Among many mechanisms, accelerated atherosclerosis, endothelial dysfunction, and hypercoagulability play a main role. Here, we investigate whether inflammatory, serological and clinical markers of SLE determine and correlate with arterial stiffness in SLE patients. Materials and methods: Routine blood samples, inflammatory mediators, specific antibodies, and 24 h proteinuria were measured in 43 SLE patients and 43 age and sex-matched controls using routine laboratory assays. We also assessed arterial stiffness by measuring radial artery applanation tonometry-derived augmentation index (AI), normalized AI (AIx@75), aortic pulse pressure, central systolic, diastolic and peripheral blood pressure. Results: SLE patients showed a significantly greater arterial stiffness vs. controls, as demonstrated by the significantly higher AIx@75 and aortic pulse pressure. Interestingly, regression analysis showed that age, systolic pulse pressure, inflammatory markers (erythrocyte sedimentation rate and C-reactive protein), daily dose of glucocorticoids, and cumulative organ damage positively correlated with arterial stiffness. Conclusions: SLE patients show increased arterial stiffness which correlates with markers of inflammation, that is involved in early alterations in arterial walls. Applanation tonometry can be used to screen SLE patients for subclinical vascular damage to implement prevention strategies for CVD.

Zinc and vitamin A deficiency in a cohort of children with autism spectrum disorder.

Studies suggest that trace element and vitamin deficiencies are common in children with autism spectrum disorder (ASD). Data describing the rates of vitamin and trace element deficiencies in the ASD population of the northwest of Ireland is lacking. We wished to determine the prevalence of zinc and vitamin A deficiency in the ASD population compared with controls within this geographical area. Parents of children aged 2-18years with ASD were invited to participate in the study. The control group consisted of well children attending the paediatric department for routine blood sampling. Children on vitamin supplements were excluded from both ASD and control groups. Informed written consent was obtained prior to recruitment. Samples were analysed for zinc and vitamin A levels according to standardized laboratory procedures. Seventy-four of the 150 children with ASD who were invited and 72 controls underwent blood sampling. Mean zinc and vitamin A levels were normal in both groups. There were significantly more males in the ASD group (88% versus 56%, p value<0.001). The mean (SD) zinc level was not different between the groups (ASD 11.7 [1.7] versus control 11.6 [2.1] μmol/L, p value=0.86). The mean (standard deviation) vitamin A level was higher in the ASD group (ASD 350.6 [82.6] versus 319.2 [82.8] μg/L, p value=0.03), but this was likely confounded by age. Children with ASD in the northwest of Ireland have mean zinc and vitamin A levels within the normal range. It is important that these findings are relayed to health professionals and to parents of children with ASD so that informed decisions on vitamin supplementation can be made.

European survey on preanalytical sample handling - Part 2: Practices of European laboratories on monitoring and processing haemolytic, icteric and lipemic samples. On behalf of the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) Working Group for the Preanalytical Phase (WG-PRE).

IntroductionNo guideline currently exists on how to detect or document haemolysis, icterus or lipemia (HIL) in blood samples, nor on subsequent use of this information. The EFLM WG-PRE has performed a survey for assessing current practices of European laboratories in HIL monitoring. This second part of two coherent articles is focused on HIL.Materials and methodsAn online survey, containing 39 questions on preanalytical issues, was disseminated among EFLM member countries. Seventeen questions exclusively focused on assessment, management and follow-up actions of HIL in routine blood samples.ResultsOverall, 1405 valid responses from 37 countries were received. A total of 1160 (86%) of all responders stating to analyse blood samples - monitored HIL. HIL was mostly checked in clinical chemistry samples and less frequently in those received for coagulation, therapeutic drug monitoring and serology/infectious disease testing. HIL detection by automatic HIL indices or visual inspection, along with haemolysis cut-offs definition, varied widely among responders. A quarter of responders performing automated HIL checks used internal quality controls. In haemolytic/icteric/lipemic samples, most responders (70%) only rejected HIL-sensitive parameters, whilst about 20% released all test results with general comments. Other responders did not analysed but rejected the entire sample, while some released all tests, without comments. Overall, 26% responders who monitored HIL were using this information for monitoring phlebotomy or sample transport quality.ConclusionStrategies for monitoring and treating haemolytic, icteric or lipemic samples are quite heterogeneous in Europe. The WG-PRE will use these insights for developing and providing recommendations aimed at harmonizing strategies across Europe.

Routine Surveillance Biopsies after Pediatric Heart Transplantation - Are They Still Necessary?

Purpose Regular surveillance endo-myocardial biopsies (rsEMB) after heart transplantation (HTx) remain the standard of care to detect allograft rejection. However, in small children the procedure is highly invasive, increases hospitalization and may misdiagnose rejections. We studied efficacy and safety of a very restrictive rsEMB protocol. Methods Single center, retrospective observational study to evaluate the early follow-up of all pediatric HTx patients between 2009- 2018. Following our institutional protocol, rsEMB was performed only once, 12 months after HTx. Instead, echocardiography, ECG and routine blood samples including T-Cell count, HLA-antibodies and levels of immunosuppressants were monitored frequently. Calcineurininhibitors and Everolimus were the primary immunosuppressants, together with steroids for the first months. Results 47 patients (25 male) were transplanted at a median age of 10.1 (0.1- 17) years. Median time since HTx was 3.4 (0.1- 9.9) years. Six patients (12.8%) had a univentricular circulation prior to HTx, 15 patients (32%) were bridged with an LVAD. Overall mortality was 10.6% (n= 5). Seven patients have not yet completed the first year after HTx. 35 patients underwent their one year follow-up including an rsEMB without complications: Histology showed no acute cellular rejection (ACR) or antibody-mediated rejection (AMR) and revealed two (5.7%) mild chronic cellular rejections (CCR) without therapeutic consequences, both no longer detectable in the follow-up EMBs 3 months later. Mild cardiac allograft vasculopathy was seen in two samples (6.3%). At 1-year follow-up, there were no clinical signs of heart failure in any of the 35 patients. Echo revealed a mean LVEF of 70 (50- 78) % with normal strain and tissue Doppler values. Median serum creatinine level was 0.5 (0.2- 1.3) mg/dL, median NT-pro BNP 209 (50- 1575) pg/ml and normal ranges for Troponin T were found. No donor specific HLA- antibodies were detected. During the whole study period there were not more than seven clinically indicated, unscheduled biopsies, exposing one ACR 2R (14%) and two ACR 1R (29%). Tests for AMR, CCR and cardiac allograft vasculopathy were all negative. Conclusion Our findings suggest that rsEMB can safely be reduced in number and replaced with non-invasive methods without increasing the risk of overlooking acute allograft rejection in children. Regular surveillance endo-myocardial biopsies (rsEMB) after heart transplantation (HTx) remain the standard of care to detect allograft rejection. However, in small children the procedure is highly invasive, increases hospitalization and may misdiagnose rejections. We studied efficacy and safety of a very restrictive rsEMB protocol. Single center, retrospective observational study to evaluate the early follow-up of all pediatric HTx patients between 2009- 2018. Following our institutional protocol, rsEMB was performed only once, 12 months after HTx. Instead, echocardiography, ECG and routine blood samples including T-Cell count, HLA-antibodies and levels of immunosuppressants were monitored frequently. Calcineurininhibitors and Everolimus were the primary immunosuppressants, together with steroids for the first months. 47 patients (25 male) were transplanted at a median age of 10.1 (0.1- 17) years. Median time since HTx was 3.4 (0.1- 9.9) years. Six patients (12.8%) had a univentricular circulation prior to HTx, 15 patients (32%) were bridged with an LVAD. Overall mortality was 10.6% (n= 5). Seven patients have not yet completed the first year after HTx. 35 patients underwent their one year follow-up including an rsEMB without complications: Histology showed no acute cellular rejection (ACR) or antibody-mediated rejection (AMR) and revealed two (5.7%) mild chronic cellular rejections (CCR) without therapeutic consequences, both no longer detectable in the follow-up EMBs 3 months later. Mild cardiac allograft vasculopathy was seen in two samples (6.3%). At 1-year follow-up, there were no clinical signs of heart failure in any of the 35 patients. Echo revealed a mean LVEF of 70 (50- 78) % with normal strain and tissue Doppler values. Median serum creatinine level was 0.5 (0.2- 1.3) mg/dL, median NT-pro BNP 209 (50- 1575) pg/ml and normal ranges for Troponin T were found. No donor specific HLA- antibodies were detected. During the whole study period there were not more than seven clinically indicated, unscheduled biopsies, exposing one ACR 2R (14%) and two ACR 1R (29%). Tests for AMR, CCR and cardiac allograft vasculopathy were all negative. Our findings suggest that rsEMB can safely be reduced in number and replaced with non-invasive methods without increasing the risk of overlooking acute allograft rejection in children.

Data-independent acquisition-based quantitative proteomic analysis reveals differences in host immune response of peripheral blood mononuclear cells to sepsis.

This study was aimed to uncover proteins that are differentially expressed in sepsis. Data-independent acquisition (DIA) was used for analysis to identify differentially expressed proteins in peripheral blood mononuclear cells (PBMCs) of patients. A total of 24 non-septic intensive care unit (ICU) patients, 11 septic shock patients and 27 patients diagnosed with sepsis were recruited for the mass spectrometry (MS) discovery. PBMCs were isolated from routine blood samples and digested into peptides. A DIA workflow was developed using a quadrupole-Orbitrap liquid chromatography LC-MS system, and mass spectra peaks were extracted by Skyline software. Orthogonal partial least-squares discriminant analysis (OPLS-DA) and partial least-squares discriminant analysis (PLS-DA) were applied to distinguish the patient groups at the level of fragment ion and peptide. Differentially expressed proteins in the patient groups were verified by enzyme-linked immunosorbent assay (ELISA). Receiver-operating characteristic (ROC) curves were used to evaluate the protein expression. A total of 1062 fragment ions and 122 proteins were identified in the MS-DIA analysis conducted by Skyline software. Using gene ontology clustering analysis, we discovered that 51 of the 122 identified proteins were associated with biological processes, including carbon metabolism, biosynthesis of antibiotics, platelet activation, bacterial invasion of epithelial cells and complement, and coagulation cascades. Among them, five proteins (high-mobility group box1 [HMGB1], matrix metalloproteinase 8 [MMP8], neutrophil gelatinase-associated lipocalin [NGAL], lactotransferrin [LTF] and grancalcin [GCA]) were identified by ELISA as closely related to the development of sepsis. The ROC curves displayed good sensitivity and specificity.

3357 TL1 Team Approach to Using a Combination of Ganglioside 2 and 3 as an Immunoaffinity Target for Circulating Osteosarcoma Cell Detection

OBJECTIVES/SPECIFIC AIMS: The objective of our collaboration is to develop a strong trans-disciplinary team consisting of microfluidics engineers, cancer biologists, and clinicians, to identify a universal marker to detect circulating osteosarcoma cells (COC) using microfluidic devices. Our goals are 3 fold: 1) Identify cell surface markers unique to osteosarcoma (OS) for COC isolation, 2) Develop a Geometrically Enhanced Mixing (GEM) device to isolate COCs, and 3) Evaluate the efficacy of GEM device to detect COCs in patients with OS. The long term goal of this collaboration is to utilize this cell detection approach to evaluate treatment efficacy and correlate the presence of circulating osteosarcoma cells with metastatic incidence. METHODS/STUDY POPULATION: In this phase of our study, we have identified an abundant and conserved cell surface marker across a panel of OS cell lines. Flow cytometry was used to evaluate the relative expression of Epithelial Cell Adhesion Molecule (EpCAM), and Ganglioside 2 or/and 3 (GD2/3) on a panel of OS cell lines. An antibody coated GEM microfluidic device is used to affirm the efficacy of GD2/3 to capture COCs. Further capture studies will be conducted using OS cell spiked blood samples. Analysis of variance (ANOVA) will be used to determine any significant difference in capture efficiency between EpCAM, GD2/3 cell surface markers. RESULTS/ANTICIPATED RESULTS: Our results demonstrate that EpCAM is not a suitable marker for COC detection. Results from our flow cytometry studies demonstrate that GD2/3 expression is significantly higher than EpCAM expression, across all OS cell lines within our panel. The cell capture efficiency strongly correlates with the cell surface expression data obtained from flow cytometry analysis. DISCUSSION/SIGNIFICANCE OF IMPACT: OS is the most common primary bone tumor and the third leading cause of pediatric cancer deaths. At diagnosis, 80% of patients will present with metastasis, however only 20% of these cases are clinically detectable. Innovative strategies to identify patients at risk of metastasis would allow for stratification of intervention therapies. Liquid biopsies are a novel alternative to current diagnostic imaging systems to monitor metastatic incidence and treatment efficacy. The detection of circulating tumor cells (CTCs) through routine blood sampling has the potential to be used clinically for earlier detection, monitoring the treatment of metastatic cancers and surveying the effect of therapeutic interventions on metastasis. To date, the majority of the studies on CTCs have evaluated their presence in carcinomas. Although sarcomas are rare, they generally have a poorer prognosis. This study will address one of the unmet medical needs in the field of CTC detection; the identification of cell surface OS makers to improve binding specificity, increase purity, and maintain a high capture efficiency.

Clinical, behavioral and biomarker predictors of PCSK9 levels in HIV-infected patients naïve of statin therapy: A cross-sectional analysis from the Swiss HIV cohort

Background and aimsBetter characterization of Proprotein Convertase Subtilisin/Kexin 9 (PCSK9) profile is currently needed to tailor appropriate lipid-lowering strategies in HIV patients. MethodsHIV-infected individuals aged ≥ 40 years and naive of statin therapy included in the Swiss HIV cohort study were screened for PCSK9 levels with a routine blood sample collection in 2014 at the Geneva University Hospitals. An exploratory linear regression model was built including clinical (age, sex, ethnicity, cardiovascular risk factors, body mass index, low CD4 defined as ≤200 cells/μl, leucocytes, lymphocytes, platelet, antiretroviral therapy), behavioral (tobacco and marijuana smoking, alcohol use and physical activity) and biomarker (CRP, TNF-α, IL-8, Il-10 and MCP-1) to investigate association with continuous PCSK9 levels. ResultsWe studied 239 HIV-infected individuals who met inclusion criteria and available PCSK9 levels with a mean age of 49 years. 35 subjects (14.6%) reported marijuana consumption, of whom 20 (57.1%) reported daily consumption and 15 (6.3%) occasional use. PCSK9 levels were correlated with low-density lipoprotein-cholesterol (LDL-C). Our exploratory model identified marijuana consumption (p=0.023) and low CD4 values (p=0.020) as significantly associated factors with higher PCSK9 levels. No association was found with Framingham risk score. Patients with marijuana consumption had significantly higher levels of PCSK9 with a dose-response effect (p < 0.001); the association persisted after adjustment for the calculated Framingham risk score (p=0.003) and additional adjustment for clinical variables (p=0.027). ConclusionsIn HIV-infected individuals naïve of statin treatment, marijuana consumption and low CD4 values are associated with higher PCSK9 levels independently of clinically relevant confounding factors.

Inflammatory Cytokine and Chemokine Patterns in Paediatric Patients with Suspected Serious Bacterial Infection.

Background and objectives: In children, acute infection is the most common cause of visits to the emergency department. Although most of them are self-limiting, mortality due to severe bacterial infections (SBI) in developed countries is still high. When the risk of serious bacterial infection is too high to ignore, yet too low to justify admission and hospital observation, clinicians try to improve diagnostic accuracy by performing various laboratory tests. The aim of the study was to investigate whether an early inflammatory cytokine and chemokine panel can add information in diagnostics of SBI and assessment of efficacy of early therapies in hospitalized children with fever. Methods: This study included 51 children with febrile infections that were admitted to the emergency department (ED). Clinical examination and microbiological and radiological tests were used as reference standards for the definition of SBI. Study population was categorized into two groups: (1) patients with SBI (n = 21); (2) patients without SBI (n = 30). Inflammatory cytokine and chemokine panels were analyzed from the first routine blood samples at hospital admission and after 24 h. Results: Out of 12 cytokines and chemokines, only Eotaxin and granulocyte colony-stimulating factor (G-CSF) had statistically significant differences between groups at the time of inclusion. Receiver operator characteristic analysis to predict SBI showed an area under the curve (AUC) of 0.679 for G-CSF. Conclusions: Analysis of inflammatory cytokine profiles may provide additional information in early diagnostics of SBI.

The Norwegian Cognitive impairment after stroke study (Nor-COAST): study protocol of a multicentre, prospective cohort study

BackgroundEarly and late onset post-stroke cognitive impairment (PCI) contributes substantially to disability following stroke, and is a high priority within stroke research. The aetiology for PCI is complex and related to the stroke itself, brain resilience, comorbid brain diseases, prestroke vulnerability and complications during the hospital stay. The aim of the Norwegian Cognitive Impairment After Stroke study (Nor-COAST) is to quantify and measure levels of cognitive impairments in a general Norwegian stroke population and to identify biological and clinical markers associated with prognosis for cognitive disorders following incident stroke. The study will be organised within five work packages: 1) Incidence and trajectories 2) Pathological mechanisms 3) Development of a risk score 4) Impact of physical activity and 5) Adherence to secondary prevention.MethodsNor-COAST is an ongoing multicentre (five participating hospitals), prospective, cohort study with consecutive inclusion during the acute phase and with follow-up at three and 18 months, and at three years. Inclusion criteria are stroke defined according to the WHO criteria. During the recruitment period from 18.05.2015 to 31.03.2017, 816 participants have been included. Cognitive impairment will be classified according to the DSM-5 criteria using a consensus group. Cognitive function is assessed by a standardised neuropsychological test battery, the Montreal Cognitive Assessment, Trail making A and B, ten-word immediate and delayed recall test, the Controlled Oral Word Association, Global Deterioration Scale and proxy based information by and the Ascertain Dementia 8 item informant questionnaire. Biomarkers include magnetic resonance imaging, routine blood samples and bio-banking. Clinical assessments include characteristics of the stroke, comorbidity, delirium, frailty and tests for cognitive and physical function, sensor based activity monitoring and adherence to secondary prophylaxis.DiscussionNor-COAST is the first Norwegian multicentre study to quantify burden of PCI that will provide reliable estimates in a general stroke population. A multidisciplinary approach aiming to identify biomarkers and clinical markers of overall prognosis will add new knowledge about risk profiles, including pre-stroke vulnerability and modifiable factors such as physical activity and secondary prophylaxis of relevance for clinical practice and later intervention studies.Trial registrationClinicalTrials.gov: NCT02650531. Retrospectively registered January 8, 2016. First participant included May 18, 2015.

DIFFERENTIAL CELL COUNT AS DIAGNOSTIC AND PROGNOSTIC MARKER OF ACUTE MYOCARDIAL INFARCTION

Myocardial infarction is a necrosis of myocardial cells due to lack of blood and oxygen supply caused by obstruction of coronary arteries, mostly due to atherosclerosis processes. Increased inflammatory marker level is associated with poor cardiovascular prognosis. This study was aimed to know whether leukocytes count, differential cell count and the Ratio of Neutrophils-Lymphocytes (RNL) could distinguish between types of Acute Myocardial Infarction (AMI) and to evaluate its correlation with mortality. This was a cross-sectional retrospective study using medical records patients which were diagnosed as AMI by clinicians in Cardiac Centre of the Dr. Wahidin Sudirohusodo Hospital during the period of April 1st, 2015 - May 31st, 2016. Statistical analysis used the Mann-Whitney and Chi-Square test, p&lt;0.05 was considered as significant. The total subjects were 435 patients divided into 289 ST- Elevation Myocardial Infarction (STEMI) and 146 Non-ST-Elevation Myocardial Infarction (NSTEMI). There were significant differences in that mean of leukocytes, neutrophils, lymphocytes, monocytes, eosinophils counts and RNL between STEMI and NSTEMI (p &lt;0.05). Significant differences were also found in leukocyte, neutrophils, lymphocytes, eosinophils, basophils and RNL mean between those who died and survived (p &lt;0.05) and a significant correlation between increased leukocytes, neutrophils, basophils counts with mortality (p &lt;0.05). In conclusion, the number of leukocytes and leukocyte count can be used as diagnostic markers of AMI between STEMI and NSTEMI, as well as prognostic markers among patients who died and survived. Routine blood sampling cohort studies in patients with AMI can avoid the bias of the results obtained.

Biobank linked to SWEDEHEART quality registry—routine blood sample collection opens new opportunities for cardiovascular research

High-quality biobanking within routine health services, through the use of existing health-care practices and infrastructure, with respect to safety and integrity of patients in line with the Swedish Biobank Act, enables large-scale collection of biological material at reasonable costs. Complementing the extensive information on myocardial infarction patients from a national registry gives unique opportunities for research focusing on better understanding of cardiovascular disease occurrence and prognosis, developing of new diagnostic methods, and personalized treatments with greater efficacy and fewer side effects.

Quality, quantity and recovery of DNA content from routine blood samples and genotyping success rate: comparison between phenol chloroform method (PCM) with a new kit-based DNA extraction method (KBM)

Background: DNA extraction has become a baseline method for molecular biology studies. There are a variety of methods available for this purpose. Newer kit-based methods (KBM) are easy and less time consuming than traditional chemical methods of extraction like phenol chloroform method (PCM). Though estimates of quality from different methods are available in labels, this study compared the practical outcomes regarding quantity, quality, DNA recovery rate and assessed the outcomes at two different time points.Methods: This study was done as a secondary analysis from an ongoing project. The quantity and quality of DNA isolated from the same group of 100 deidentified blood samples by PCM and KBM were analysed using Multi analyzer and repeated after a period of 3 months. Genotyping of the samples were done by RT-PCR. The quantity, quality and amplification proportion were compared between two groups to reach the inference.Results: The median (range) concentration of DNA by PCM was 543.27 (960.59) µg/ml and that of KBM was 32.115 (36.73) µg/ml. The quality of DNA as measured by absorbance at 260/280 nm was 1.84 in PCM and 1.81 in KBM (p&gt;0.05). Genotyping success rate was 78% in PCM and 98% in KBM (p = 0.002). The DNA recovery rate was 96% in PCM and 80% in KBM (p=0.014).Conclusions: The median concentration of DNA obtained from PCM was more compared to KBM. The quality of DNA was comparable in both the groups. The genotyping success rate was more in KBM group. The DNA recovery rate at 3 months was more in PCM group.

The effect of hypoglycaemia on neurocognitive outcome in children and adolescents with transient or persistent congenital hyperinsulinism.

To examine the hypoglycaemic effect on neurodevelopmental outcome in patients with transient and persistent congenital hyperinsulinism (CHI) born in the 21st century. A cohort of 117 patients (66 males, 51 females) with CHI aged 5 to 16years (mean age 8y 11mo, SD 2y 7mo) were selected from a Finnish nationwide registry to examine all the patients with similar methods. Neurodevelopment was first evaluated retrospectively. The 83 patients with no risk factors for neurological impairment other than hypoglycaemia were recruited and 44 participated (24 males, 20 females; mean age 9y 7mo, SD 3y 1mo) in neuropsychological assessment with the Wechsler Intelligence Scale for Children, Fourth Edition and the Finnish version of the Developmental Neuropsychological Assessment, Second Edition domains of attention, language, memory, sensorimotor, and visual functioning. In retrospective analysis, transient and persistent CHI groups had similar prevalences of mild (22% and 18% respectively) or severe (5% and 7% respectively) neurodevelopmental difficulties. In clinical assessment, the neurocognitive profile was within the average range in both groups, but children with persistent CHI showed significant but restricted deficits in attention, memory, visual, and sensorimotor functions compared with the general population. The transient CHI group did not differ from the standardization samples. Besides the more apparent broader neurological deficits, children with persistent CHI have an increased risk for milder specific neurocognitive problems, which should be considered in the follow-up. Children with persistent congenital hyperinsulinism showed deficits in attention, memory, visual, and sensorimotor functions. The deficits were potentially of hypoglycaemic origin. Children with transient hyperinsulinism did not differ from the general population.

The significant value of predicting prognosis in patients with colorectal cancer using 18F-FDG PET metabolic parameters of primary tumors and hematological parameters.

The purpose was to evaluate the correlation of the pre-treatment hematological parameters with metabolic parameters of primary tumor in baseline 18F-FDG PET/CT in patients with colorectal cancer (CRC) and estimate the prognostic value of both. We retrospectively investigated 231 patients with CRC who underwent baseline 18F-FDG PET/CT. Routine blood sampling was tested in the same term. PET parameters in term of hematological parameters and pathological characteristics of primary tumor were compared. Kaplan-Meier survival analysis was performed in the patients without distant metastasis. The differences of disease-free survival between groups were compared by log-rank tests. Neutrophil-to-lymphocyte ratio (NLR) and lymphocyte-to-monocyte ratio (LMR) were significantly correlated with all the metabolic parameters including maximum standardized uptake value (SUVmax), metabolic tumor volume (MTV) and tumor lesion glycolysis (TLG). The patients with NLR > 3 had higher MTV (24.82 ± 18.16 vs 19.06 ± 13.30, P = 0.039) and TLG (219.04 ± 186.94 vs 166.45 ± 146.39, P = 0.047) than those whose NLR ≤ 3. NLR in those patients with distant metastasis was significantly higher than those without distant metastasis (P = 0.018) while LMR in those patients with distant metastasis was significantly lower than those without distant metastasis (P = 0.032). Survival analysis showed that those patients with low MTV (P = 0.015), low NLR (P = 0.008) and high LMR (P = 0.027) revealed significant survival benefit. There was a significant association between the pre-treatment hematological parameters and metabolic parameters of baseline 18F-FDG PET/CT in the patients with CRC. It might be helpful in those patients with high NLR and low LMR to undergo 18F-FDG PET/CT to detect distant metastasis and predict prognosis.

A General Lack of IgG Against Pertussis Toxin in Chinese Pregnant Women and Newborns.

To investigate the level of pertussis-related antibodies in pregnant women and newborns. A total of 286 serum samples from healthy pregnant women and 221 cord samples from newborns were collected in 2016 in Beijing. A routine blood sample from pregnant women was obtained at 35 weeks of gestational age, and cord samples were collected in 20 minutes after delivery. The values from cord samples were used as the infant values. Anti-pertussis toxin (PT) IgG concentration was measured by ELISA (Euroimmun, Lübeck, Germany) using purified PT as a coating antigen. Newborns with anti-PT IgG ≤40 IU/mL in cord samples were considered to be unprotected against pertussis. Anti-PT IgG ≥100 IU/mL was considered to be indicative of a recent pertussis infection in pregnant women. The anti-PT IgG concentration below the lower limit of detection (<5 IU/mL) occurred in 74.1% (212/286) of pregnant women and 66.5% (147/221) of newborns. Even with detectable anti-PT antibodies, the majority of pregnant women (79.7%, 59/74) and newborns (73.0%, 54/74) had antibody level of 5 to <20 IU/mL, and 13.5% (10/74) of pregnant and 14.9% (11/74) of newborns had antibody level of 20 to <40 IU/mL. The 75% percentiles for anti-PT IgG of pregnant women and newborns were 5.08 and 6.98 IU/mL, respectively. The prevalence of unprotected newborns as defined by anti-PT IgG ≤40 IU/mL was 95.9% (202/211). The prevalence of recent pertussis infection in pregnant women as defined by anti-PT IgG ≥100 was 0.7% (2/286). The pregnant women and newborns were generally lack of protective antibody and are vulnerable to pertussis in Beijing, China. Although acellular pertussis vaccine is administrated in infancy in China, a booster vaccination to pregnant women should be considered for protecting young infants who are too young to start pertussis vaccination.

Assessing prevalence of alcohol consumption in early pregnancy: Self-report compared to blood biomarker analysis

Providing appropriate antenatal and postnatal care for women who drink alcohol in pregnancy is only possible if those at risk can be identified. We aimed to compare the prevalence of alcohol consumption in the first trimester of pregnancy using self-report and blood biomarker analysis. Six-hundred routine blood samples from 2014, taken at the antenatal booking appointment, in the first trimester of pregnancy, were anonymously analysed for the presence of Carbohydrate Deficient Transferrin (CDT), a validated marker of chronic alcohol exposure (normalising 2–3 weeks from abstinence) and Gamma-glutamyltransferase (GGT), a liver enzyme elevated for up to 8 weeks after alcohol exposure. In a separate sample of women, from 2015, data taken during the antenatal visit, documenting women's self-reported alcohol consumption, were collected. The percentage of women who reported alcohol intake in the first trimester was 0.8%. This compared to 74.1% of women who reported consuming alcohol before pregnancy. CDT analysis revealed a prevalence rate of 1.4% and GGT a prevalence rate of 3.5% in the first trimester of pregnancy. Although those with elevated CDT generally had high levels of GGT, only one person was positive for CDT and GGT. Results from CDT analysis and self-report may underestimate prevalence for different reasons. GGT appeared to lack specificity, but it may have value in supporting findings from CDT analysis. Further studies using additional blood biomarkers, or a combination of blood biomarkers and self-report, may be beneficial in accurately detecting alcohol drinking history in pregnancy.

Assessment of Immature Granulocytes Percentage to Predict Severe Bacterial Infection in Latvian Children: An Analysis of Secondary Data.

Background: Detection of small proportion of serious bacterial infections (SBI) with a potentially life threating course in a large group of children with fever admitted to emergency department (ED) is still complicated. Measurement of immature granulocytes (IG) percentage may be used as a marker of bacterial infections. The aim of the study was to evaluate whether the IG percentage is a useful additional predictive marker of SBI. Methods: This study included 258 children with febrile infections that were admitted to the ED. Clinical follow-up, microbiological and radiological tests were used as reference standards for the definition of SBI. Study population was categorized into two groups: (1) infected patients with no suspicion of SBI (n = 75); (2) patients with suspicion of SBI (n = 183). IG percentage, white blood cell count (WBC), and C-reactive protein (CRP) levels were analyzed from the first routine blood samples at hospital admission. Results: A statistically significant difference in IG percentage levels was observed in children with SBI and those without—the mean IG percentage was 1.2% for the SBI group, 0.3% for those without SBI. The cutoff level of IG percentage to predict SBI was 0.45 (84% specificity, 66% sensitivity, 90% positive predictive value). We combine variables and evaluate their additive values. The sensitivity of WBC to detected SBI improved from 74% to 85% when IG percentage was added to the prediction models. When CRP, WBC, and IG percentage were combined, the sensitivity to predict SBI increased to 93%, the specificity to 86% (95% CI 77%–93%). Receiver operator characteristic analysis to predict SBI showed an area under the curve (AUC) of 0.80 for IG percentage. Conclusion: Addition of IG percentage to traditionally used markers of SBI as WBC and CRP may help to identify children with serious bacterial infections. Furthermore, IG percentage can be rapidly obtained from the traditional full blood count without any extra sampling and costs.

Associations between trabecular bone score and biochemistry in surgically vs conservatively treated outpatients with primary hyperparathyroidism: A retrospective cohort study

PurposeTrabecular Bone Score (TBS) is a software-based method for indirect assessment of trabecular bone structure of the spine, based on analysis of pixels in dual energy x-ray absorptiometry (DXA) images. Few studies describe the use of TBS in patients with primary hyperparathyroidism (PHPT). This study aimed at further describing this relationship, investigating possible correlations between biochemistry, body mass index (BMI), fracture incidence and TBS. MethodsCross-sectional study of 195 patients with verified PHPT, surgically (27) or conservatively (168) treated at the Department of Endocrinology, Aalborg University Hospital. TBS was acquired by reanalyzing DXA-images of the included subjects from the outpatient clinic. Biochemical variables were obtained from clinical routine blood samples taken in relation to the DXA-scans. History of fractures and medical history was obtained from radiology reports and medical charts. ResultsPatients with active PHPT had a TBS-score signifying a partly degraded bone structure, whereas surgically treated patients had a normal bone structure as judged by TBS, though the difference in TBS-score was not statistically significant. Use of antiresorptive treatment was negatively associated with BMD but not TBS. No correlations between the biochemical variables and TBS were found. A negative correlation between TBS and BMI in patients with PHPT was present. Patients experiencing a fragility fracture had a significantly lowered TBS, BMD and T-Score. ConclusionBiochemistry does not seem to predict bone status in terms of TBS in patients with PHPT. TBS is negatively correlated to BMI, which is also seen in patients not suffering from PHPT. The lack of a predictive value for antiresorptive treatment for TBS may raise concern. TBS appears to have a predictive value when assessing risk of fracture in patients with PHPT. Mini abstractThis cross-sectional study investigates possible correlations between biochemical variables, body mass index (BMI) and trabecular bone score (TBS) in 195 patients with primary hyperparathyroidism. It finds no correlation between biochemical variables and TBS, but finds a negative correlation between TBS and BMI and a clear association between fracture incidence and low TBS-score.