Collection Of Peripheral Blood Samples Research Articles

Nuclear Magnetic Resonance Analysis Implicates Sex-Specific Dysregulation of the Blood Lipids in Alzheimer's Disease: A Retrospective Health-Controlled Study.

The aging demographic landscape worldwide portends a heightened prevalence of neurodegenerative disorders. Foremost among these is Alzheimer's disease (AD), the foremost cause of dementia in older adults. The shortage of efficacious therapies and early diagnostic indicators underscores the imperative to identify non-invasive biomarkers for early detection and disease monitoring. Recently, blood metabolites have emerged as promising candidates for AD biomarkers. Leveraging nuclear magnetic resonance (NMR) spectroscopy on plasma specimens, we conducted a cross-sectional study encompassing 35 AD patients and 35 age-matched healthy controls. Cognitive function was evaluated using the mini-mental state examination in all participants, followed by peripheral blood sample collection. We utilized univariate and multivariate analyses to perform targeted lipidomic profiling via NMR spectroscopy. Our study revealed significant differences in the expression profiles of low-density lipoprotein-associated subfractions in females and high-density lipoprotein-associated subfractions in males between AD patients and healthy controls (all p<0.05). However, there was no significant metabolite overlap between males and females. Furthermore, receiver operating characteristic curve analysis demonstrated that the combination of lipid metabolites had good diagnostic values (all area under the curve>0.70; p<0.05). Our findings suggest that the blood plasma samples using NMR hold promise in distinguishing between AD patients and healthy controls, with significant clinical implications for advancing AD diagnostic methodologies.

The rs2018736 fibulin-5 polymorphism as a determinant for pelvic organ prolapse: a case–control study

Objective Fibulin-5 is a connective tissue component and may play a role in pelvic organ prolapse (POP) pathogenesis. This study aimed to verify the association of the rs2018736 polymorphism of the fibulin-5 gene with POP in postmenopausal Brazilian women, and to determine the risk factors for POP. Method This observational, cross-sectional, case–control study assessed postmenopausal women with advanced POP (stages III and IV) and control women (stages 0 and I) by examination and peripheral blood sample collection. DNA sequences were analyzed by real-time reverse-transcriptase polymerase chain reaction. A logistic regression model was used with p < 0.05 for significance. Results A total of 565 participants were evaluated (325 POP and 240 control). The homozygous C allele of rs2018736 (CC) was protective against POP (odds ratio [OR] 0.49, 95% confidence interval [CI] 0.26–0.91). Age (OR 1.09, 95% CI 1.05–1.13), number of pregnancies (OR 1.14, 95% CI 1.01–1.28), vaginal delivery (OR 5.32, 95% CI 2.58–11.01), forceps delivery (OR 3.34, 95% CI 1.72–6.47), weight of newborn (OR 1.0007, 95% CI 1.0002–1.0011), family history of POP (OR 2.35, 95% CI 1.24–4.44), hypertension (OR 1.74, 95% CI 1.01–3.00) and diabetes (OR 2.19, 95% CI 1.07–4.48)] were independent predictors for POP; cesarean (OR 0.02, 95% CI 0.005–0.09) was protective. Conclusion The rs2018736-CC genotype of the fibulin-5 gene has a protective role against POP.

Utility of clinical metagenomics in diagnosing malignancies in a cohort of patients with Epstein-Barr virus positivity.

Differentiation between benign and malignant diseases in EBV-positive patients poses a significant challenge due to the lack of efficient diagnostic tools. Metagenomic Next-Generation Sequencing (mNGS) is commonly used to identify pathogens of patients with fevers of unknown-origin (FUO). Recent studies have extended the application of Next-Generation Sequencing (NGS) in identifying tumors in body fluids and cerebrospinal fluids. In light of these, we conducted this study to develop and apply metagenomic methods to validate their role in identifying EBV-associated malignant disease. We enrolled 29 patients with positive EBV results in the cohort of FUO in the Department of Infectious Diseases of Huashan Hospital affiliated with Fudan University from 2018 to 2019. Upon enrollment, these patients were grouped for benign diseases, CAEBV, and malignant diseases according to their final diagnosis, and CNV analysis was retrospectively performed in 2022 using samples from 2018 to 2019. Among the 29 patients. 16 of them were diagnosed with benign diseases, 3 patients were diagnosed with CAEBV and 10 patients were with malignant diseases. 29 blood samples from 29 patients were tested for mNGS. Among all 10 patients with malignant diagnosis, CNV analysis suggested neoplasms in 9 patients. Of all 19 patients with benign or CAEBV diagnosis, 2 patients showed abnormal CNV results. The sensitivity and specificity of CNV analysis for the identification for tumors were 90% and 89.5%, separately. The application of mNGS could assist in the identification of microbial infection and malignancies in EBV-related diseases. Our results demonstrate that CNV detection through mNGS is faster compared to conventional oncology tests. Moreover, the convenient collection of peripheral blood samples adds to the advantages of this approach.

Whole Exome Sequencing of 20 Spanish Families: Candidate Genes for Non-Syndromic Pediatric Cataracts.

Non-syndromic pediatric cataracts are defined as opacification of the crystalline lens that occurs during the first years of life without affecting other organs. Given that this disease is one of the most frequent causes of reversible blindness in childhood, the main objective of this study was to propose new responsible gene candidates that would allow a more targeted genetic approach and expand our genetic knowledge about the disease. We present a whole exome sequencing (WES) study of 20 Spanish families with non-syndromic pediatric cataracts and a previous negative result on an ophthalmology next-generation sequencing panel. After ophthalmological evaluation and collection of peripheral blood samples from these families, WES was performed. We were able to reach a genetic diagnosis in 10% of the families analyzed and found genes that could cause pediatric cataracts in 35% of the cohort. Of the variants found, 18.2% were classified as pathogenic, 9% as likely pathogenic, and 72.8% as variants of uncertain significance. However, we did not find conclusive results in 55% of the families studied, which suggests further studies are needed. The results of this WES study allow us to propose LONP1, ACACA, TRPM1, CLIC5, HSPE1, ODF1, PIKFYVE, and CHMP4A as potential candidates to further investigate for their role in pediatric cataracts, and AQP5 and locus 2q37 as causal genes.

Elevated complement component 8 gamma levels in astrocyte-derived exosomes are associated with cognitive impairment in obstructive sleep apnea patients without dementia

The complement system plays a crucial role in cognitive impairment in obstructive sleep apnea (OSA). The present study aimed to investigate the connections between complement component 8 gamma (C8G) levels in astrocyte-derived exosomes (ADEs) and cognitive impairment in OSA patients without dementia. This cross-sectional cohort study recruited 274 participants without dementia, including 124 OSA patients with mild cognitive impairment (MCI), 100 OSA patients without MCI, and 50 healthy control subjects. Enrolled participants underwent polysomnography (PSG) evaluation, neuropsychological scale assessment, magnetic resonance imaging scanning, and collection of peripheral blood samples for quantification of complement proteins in ADEs. The findings showed higher C8G concentrations in ADEs from OSA patients with MCI than in the controls and OSA without MCI group. Logistic regression analysis suggested that C8G levels in ADEs were independently associated with MCI in OSA patients. Multivariable linear regression analysis demonstrated that C8G levels in ADEs were significantly correlated with global cognitive scores and all cognitive subdomain scores after adjusting for demographic factors (age, sex, education), vascular risk factors (Body mass index, history of hypertension, diabetes, dyslipidemia), depressive symptoms measures, and apnea-hypopnea index (AHI) values. The levels of C8G were linearly positively related to the white matter hyperintensity (WMH) volumes in Pearson's correlation analysis. Our research confirmed that C8G levels are significantly associated with cognitive impairment in OSA patients, which paves the way for novel therapeutic targets for neurocognitive dysfunction progression in OSA patients in the future.

Autoimmune regulator (AIRE): Takes a hypoxia-inducing factor 1A (HIF1A) route to regulate FOXP3 expression in PCOS.

Autoimmune polyendocrinopathy-candidiasis- ectodermal dystrophy (APECED) pathology due to autoimmune regulator (AIRE) gene mutations leads to loss of central tolerance triggering immune attack, a factor causing infertility. One of the targets of autoimmune attack is ovary and its repercussion results in polycystic ovarian syndrome (PCOS). Although reduced Tregs have been reported in PCOS, a lacunae exists on the status of AIRE gene expression and its role in treg insufficiency via HIF1A-FOXP3 axis in PCOS. This is a case-control cohort study recruiting 40 normal and 40 PCOS volunteers for peripheral blood sample collection and PCOS diagnoses were based on Rotterdam Consensus criteria. AIRE and HIF1A expression status was analysed by qRT PCR and western blot. FACS analyses was conducted on AIRE silenced peripheral blood mononuclear cells (PBMCs) after Treg induction. Our results indicate a reduced AIRE (fold change log2 (RQ) = -2.6, P < .01) and increased HIF1A (fold change log2 (RQ) = 3.6, P < .02) in PBMCs of PCOS subjects compared to age-matched controls. Western blot of AIRE and HIF1A corroborates with qRT PCR data. Our CHIP data demonstrate AIRE mediated HIF1A promoter regulation. Silencing of AIRE in PBMCs contributes to the upregulation of HIF1A transcripts by two-fold (P < .0015) and downregulation in FOXP3 expression by three-fold (P < .0017). FACS analyses revealed that silencing of AIRE reduces Tcell to Treg conversion. Our consolidated results derive a new connection among AIRE-HIF1A-FOXP3 with AIRE reduction enabling increased HIF1A resulting in reduced FOXP3 in PBMCs of PCOS patients leading to Treg insufficiency.

Matrix Metalloproteinase-9 Levels are Associated with Brain Lesion and Persistent Venous Occlusion in Patients with Cerebral Venous Thrombosis.

Elucidating mechanisms of brain damage in cerebral venous thrombosis (CVT) would be instrumental to develop targeted therapies and improve prognosis prediction. Matrix metalloproteinase-9 (MMP-9), a gelatinase that degrades major components of the basal lamina, has been associated to blood-brain barrier disruption. We aimed to assess, in patients with CVT, the temporal change in serum concentrations of MMP-9 and its association with key imaging and clinical outcomes. Pathophysiology of Venous Infarction-PRediction of InfarctiOn and RecanalIzaTion in CVT (PRIORITy-CVT) was a multicenter prospective cohort study of patients with newly diagnosed CVT. Serial collection of peripheral blood samples performed on day 1, 3, and 8, and standardized magnetic resonance imaging on day 1, 8, and 90. MMP-9 was quantified using enzyme-linked immunosorbent assay in 59 patients and 22 healthy controls. Primary outcomes were parenchymal brain lesion, early evolution of brain lesion, early recanalization, and functional outcome on day 90. CVT patients with parenchymal brain lesion had higher baseline concentrations of MMP-9 compared with controls (adjusted p = 0.001). The area under receiver operating characteristic curve value for MMP-9 for predicting brain lesion was 0.71 (95% confidence interval [CI]: 0.57-0.85, p = 0.009). Patients with venous recanalization showed early decline of circulating MMP-9 and significantly lower levels on day 8 (p = 0.021). Higher MMP-9 on day 8 was associated with persistent venous occlusion (odds ratio: 1.20 [per 20 ng/mL], 95% CI: 1.02-1.43, p = 0.030). We report a novel relationship among MMP-9, parenchymal brain damage, and early venous recanalization, suggesting that circulating MMP-9 is a dynamic marker of brain tissue damage in patients with CVT.

Collagen I and collagen III polymorphisms in women with pelvic organ prolapse.

Verify the presence of the single nucleotide polymorphisms rs1800012 of the collagen I (COL1A1) and rs1800255 of the collagen III (COL3A1) genes and their association with pelvic organ prolapse (POP) in Brazilian women and to determine risk factors for POP. We assessed 826 postmenopausal women divided into POP (stages III and IV) and control groups (stages 0 and I) by examination and peripheral blood sample collection. DNA sequences of interest were analyzed by real-time reverse-transcriptase polymerase chain reaction. We used logistic regression analyses, recessive and codominance models of inheritance, and P < .05 for significance. Six-hundred and thirty-four postmenopausal women were included: 348 (54.8%) POP cases and 286 (45.1%) controls. The frequencies of GG, GA, and AA genotypes for COL1A1 were 69.12%, 20.24%, and 10.59% in POP group and 71.79%, 20%, and 8.21% in controls; GG, GT, and TT for COL3A1 were 37.54%, 59.53%,and 2.93% in POP group and 36.24%, 60.14%, and 3.62% in controls. There were no genotypic or allelic association with POP phenotype that link both SNPs rs1800012 and rs1800255 to increased risk of POP. Vaginal delivery (odds ratio [OR] = 13; 95% confidence interval [CI] [4.00-47.08]), POP family history (OR = 3.1; 95% CI [1.49-6.50]), diabetes mellitus (OR = 2.3; 95% CI [1.08-5.21]), number of pregnancies (OR = 1.2; 95% CI [1.05-1.36]), and age (OR = 1.1; 95% CI [1.09-1.19]), were variables of increased risk for POP (P < .05 for all). Our study suggests lack of association between DNA polymorphisms rs1800012 of COL1A1 and rs1800255 of COL3A1 with advanced POP. Vaginal delivery, POP family history, diabetes mellitus, number of pregnancies, and age are risk factors for POP.

Mechanisms of Adaptation to Ibrutinib in High Risk Chronic Lymphocytic Leukemia

Introduction. Ibrutinib inhibits the BTK molecule downstream the B-cell receptor (BCR). Though highly active in high risk chronic lymphocytic leukemia (CLL), the most typical response achievable in patients is a minimal residual disease (MRD) positive partial remission (PR) which is maintained until the development of genetically driven resistance caused by the acquisition of mutations in the BTK or PLCG2 genes. The study aims at characterizing the adaptation process allowing residual CLL cells to persist despite BTK inhibition. Methods. The IOSI-EMA-001 study (NCT02827617) is an observational study consisting in the prospective and longitudinal collection of peripheral blood samples and clinical data from high risk CLL patients treated with ibrutinib. Peripheral blood CLL cells longitudinally drawn from patients before treatment start and at fixed timepoints under ibrutinib were monitored by: i) next generation flow cytometry approaches for changes in proliferation rate, surfaceome, and pathway activation; and ii) CAPP-seq targeted deep next generation (sensitivity ~10-3) for clonal evolution. Results. The study cohort comprised 31 high risk CLL patients, including 15 treatment naïve, 16 relapsed, 80% IGHV unmutated, 42% 17p deleted and 55% TP53 mutated. Median duration of ibrutinib treatment was 45 weeks (24-72 weeks). All patients obtained a MRD positive PR that was maintained in all but one who progressed with a PLCG2 mutation (VAF 3%). Compared to baseline, under ibrutinib therapy CLL cells slowed down their proliferation, as suggested by the decreased expression of Ki-67, the reduction of the proliferating fraction (CXCR4dimCD5bright), and the increase of the resting fraction (CXCR4brightCD5dim). Compared to baseline, under ibrutinib therapy CLL cells also upregulated BCR and adhesion/homing proteins, and decreased the expression of BCR inhibitor proteins. Upon stimulation of the BCR with anti-IgM, the downstream path through pBTK and pPLCG2 was inhibited by ibrutinib, while conversely the downstream path through pAKT and pERK was still inducible throughout all the assessed timepoints. The proportion of CLL cells harboring nuclear localization of NF-kB progressively increased over time under ibrutinib. NF-kB nuclear localization was inducible throughout all the assessed timepoints by CD40L stimulation of the non-canonical NF-kB pathway, but not by anti-IgM stimulation of the BCR/canonical NF-kB pathway. Overall, 880 individual mutations were longitudinally discovered and monitored across a total of 121 sequential timepoints collected during ibrutinib treatment. Clonal evolution was observed in (67.7%) cases, a proportion rate previously documented in CLL treated with chemoimmunotherapy. Clonal evolution appeared to be heterogeneous involving different genes without a stereotypic targeting. Consistently, none of the main driver gene mutations was homogeneously selected or suppressed by ibrutinib suggesting that the biological adaptation of CLL cells under ibrutinib is not genetically driven. Clonal evolution propensity was not associated with any of the biomarkers of the disease, and it did not decrease over time under ibrutinib. Conclusions. Taken together these results suggest that residual CLL cells persisting under ibrutinib therapy adapt their phenotype by upregulating adhesion molecules, chemokine receptors and BCR molecules, and by maintaining a competence of BCR signaling through the PI3K/AKT/ERK pathway. The progressive selection of CLL cells having NF-kB in the nucleus, likely due to the BTK independent non-canonical NF-kB pathway, might explain their survival despite ibrutinib therapy. Finally, clonal evolution is not suppressed by ibrutinib chemotherapy, and despite does not seem to be directly involved in such adaptation process, may ultimately favor the acquisition of BTK and PLCG2 ibrutinib resistance mutations. DisclosuresZucca:Celltrion: Consultancy; AstraZeneca: Consultancy. Ghia:Sunesis: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; AbbVie, Inc: Honoraria, Research Funding; Acerta: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Gilead: Honoraria, Research Funding; BeiGene: Honoraria, Research Funding. Montillo:Janssen: Consultancy, Honoraria; Gilead: Consultancy, Honoraria, Speakers Bureau; AbbVie: Consultancy, Honoraria, Speakers Bureau; Roche: Consultancy, Honoraria, Research Funding. Tedeschi:Janssen: Consultancy, Speakers Bureau; Gilead: Consultancy; AbbVie: Consultancy. Gaidano:AbbVie: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Morphosys: Honoraria; Roche: Consultancy, Honoraria.

Frequent Tail-tip Blood Sampling in Mice for the Assessment of Pulsatile Luteinizing Hormone Secretion.

In many endocrine systems, circulating factors or hormones are not released continuously, but are secreted as a discrete pulse in response to a releasing factor. Single-point sampling measures are inadequate to fully understand the biological significance of the secretory pattern of pulsatile hormones either under normal physiologic conditions or during conditions of dysregulation. Luteinizing hormone (LH) is synthesized by the anterior pituitary gonadotropecells and secreted in a pulsatile pattern which requires frequent collection of blood samples for pulse assessment. This has not been possible in mice until recently, due to the development of a high-sensitivity LH assay and advancement in a technique for frequent low-volume sample collection, initially described by Steyn and colleagues.1 Here we describe a protocol for the frequent peripheral blood sample collection from mice with sufficient handling acclimatization to detect pulsatile secretion of LH. The current protocol details an expanded acclimatization period that allows assessment of robust and continuous pulses of LH over multiple hours. In this protocol, the tip of the tail is clippedand blood is collected from the tail using a hand-held pipette. For assessment of pulsatile LH in gonadectomized mice, serial samples are collected every 5-6 min for 90-180 min. Importantly, the collection of blood and measurement of robust pulses of LH can be accomplished in awake, freely behaving mice, given adequate handling acclimatization and effort to minimize environmental stressors. Sufficient acclimatization can be achieved within 4-5 weeks prior to blood collection. This protocol highlights advances in the methodology to ensure collection of whole blood samples for assessment of pulsatile LH secretion patterns over multiple hours in the mouse, a powerful animal model for neuroendocrine research.

Quantification of peripheral blood CD34+ cells prior to stem cell harvesting by leukapheresis: a single center experience

BackgroundDue to laboratory logistic issues, our center has traditionally scheduled peripheral blood stem cell harvests based on timing from the start of mobilization. This has proved to be useful in some cases, but also resulted in many fruitless harvests due to poor mobilization. In order to improve the efficiency of collections and compare the effectiveness of peripheral blood CD34+ cells as a predictor with data from other reports, this study analyzed the implementation of this routine. MethodsPeripheral blood and leukapheresis samples were quantified by flow cytometry and the association between these parameters was assessed. ResultsSixty-six consecutive leukapheresis samples were collected from 34 patients after the collection of peripheral blood samples for CD34+ quantification. A moderate positive correlation was observed between peripheral blood CD34+ cell count and total CD34+ cell count/kg (r=0.596; p-value<0.001). A multivariable regression model also confirmed this association and allowed the estimation that for every increase in five CD34+ cells/μL in the peripheral blood, a mean increase of 0.38×106CD34+cells/kg could be predicted. Demographic characteristics, baseline comorbidities and mobilization regimen did not influence final CD34+ cell count in this sample. ConclusionsAs observed in other centers, quantification of peripheral blood CD34+ progenitor cells is a strong predictor of effectiveness to guide stem cell harvesting. Due to the results of this study, a modification in the peripheral blood stem cell harvesting logistics was implemented at our center in order to incorporate this routine.

Collagen XVIII and LOXL-4 polymorphisms in women with and without advanced pelvic organ prolapse.

We verified the presence of single nucleotide polymorphisms (SNP) rs2236479 of the collagen 18 (COL18A1) and rs2862296 of the lysyl oxidase-like 4 (LOXL-4) genes and the association with pelvic organ prolapse (POP) in Brazilian women and determined risk factors for POP development. We assessed 532 postmenopausal women divided into POP (stages III and IV) and control (stages 0 and I) groups by examination and peripheral blood sample collection. DNA sequences of interest were analyzed by real-time reverse-transcriptase polymerase chain reaction (RT-PCR). We used logistic regression models for the analyses, with p < 0.005 for significance. The frequency of homozygous polymorphic alleles (AA) in COL18A1 and (GG) in LOXL-4 were similar in both groups (17.5% and 15.4% for COL18A1 and 18.9% and 20.6% for LOXL-4, respectively). There were no associations between those polymorphisms or other genotypes and POP. Multiple logistic regression analysis identified age [odds ratio (OR) = 1.10, confidence interval (CI) 95% = 1.07; 1.14), number of vaginal births (OR = 1.66, CI 95% = 1.36; 2.03), and family history (OR = 2.55 CI 95% = 1.43; 4.55) as independent risk factors for POP. Our study suggests lack of association between DNA polymorphisms rs2236479 of COL18A1 and rs2862296 of LOXL-4 with advanced POP in this population.

Convergent evidences from human and animal studies implicate angiotensin I-converting enzyme activity in cognitive performance in schizophrenia.

In schizophrenia (SCZ), higher angiotensin I-converting enzyme (ACE) levels have been reported in patient's blood and cerebrospinal fluid (CSF). Hereby, we propose to explore whether the ACE activity levels are associated to cognitive performance in SCZ. Seventy-two patients with SCZ or schizoaffective disorder diagnosis, and 69 healthy controls (HCs) underwent a cognitive battery with parallel collection of peripheral blood samples to measure ACE activity. Significant higher ACE activity levels were confirmed in the plasma of SCZ patients compared with HCs (Student's t=−5.216; P<0.001). ACE activity significantly correlated to Hopkins delayed recall measures (r=−0.247; P=0.004) and Hopkins total (r=−0.214; P=0.012). Subjects grouped as high ACE activity (above average) had worse performance compared with low ACE activity level group for Hopkins delayed recall measure, even after correction for clinical condition, age, gender and years of education (P=0.029). The adjusted R squared for this final model was 0.343. This result was evident only comparing extreme groups for ACE activity, when splitting the sample in three groups with similar number of subjects. To clarify this finding, we performed an evaluation of the cognitive performance of transgenic mice with three copies of ACE gene in novel object recognition (NOR) test, which showed that such animals presented impairment in NOR (P<0.05) compared with two copies of wild-type animals. The results observed in SCZ patients and animal model suggest both the association of ACE to cognitive deficits in SCZ. This finding may support the evaluation of novel treatment protocols and/or of innovative drugs for specific intervention of cognitive deficits in SCZ envisioning concomitant ACE activity and behavior evaluations.

Cytopenias in the Early Post-Autologous Hematopoietic Cell Transplantation (aHCT) Period Predict for Subsequent Development of Therapy-Related Myelodysplasia/ Acute Myeloid Leukemia (t-MDS/AML) Among Patients with Lymphoma

t-MDS/AML is the most common cause of non-relapse mortality in patients undergoing autologous hematopoietic cell transplantation (aHCT) for Hodgkin lymphoma (HL) or non-Hodgkin lymphoma (NHL). Cytopenias are a common occurrence in the early post-aHCT period. To better understand the significance of cytopenias with respect to the risk of t-MDS/AML, we conducted a prospective, longitudinal study in patients undergoing aHCT for HL or NHL between 1999 and 2009 at City of Hope, with serial collection of peripheral blood samples from pre-aHCT to 10y. Patients with post-aHCT persistent disease (n=55) or solid second malignancies (n=9) or refusal to participate (n=18) were excluded from the analysis, leaving 292 study participants (HL: n=81; NHL: n=211). The median length of follow-up was 4.5y from aHCT. The cumulative incidence of t-MDS/AML approached 9% at 7y (Figure). Older age at aHCT (50+y: RR=3.6, 95%CI, 1.3-9.8, p=0.01) and exposure to total body irradiation (TBI: RR=2.5, 95%CI, 1.0-5.9, p=0.04) were associated with an increased risk of t-MDS/AML. Serial evaluation of peripheral blood parameters (hematocrit [Hct], mean corpuscular volume [MCV], red blood cell [RBC] count, hemoglobin [Hgb], red cell distribution width [RDW], white blood cell [WBC] count, and platelet [Plt] count) were abstracted from medical records at pre-aHCT, and after aHCT at d100, 6m, 1y, and annually thereafter for up to 10y (a total of 1,919 time points). This report focuses on alterations in peripheral blood parameters from pre-aHCT to several years post-aHCT among patients who developed t-MDS/AML (n=21; cases) and those who did not (n=271; controls). Values of peripheral blood parameters associated with post-aHCT relapse or development of t-MDS/AML were censored at 3m prior to the development of these events. As shown in the Figure, comparison of the peripheral blood parameters between cases and controls revealed that Hct, Hgb, Plt, and RBC values were significantly lower for cases compared to controls at d100, 6m, 1y, and 2y (p<0.001); WBC values for cases were lower than for controls at the pre-aHCT, d100, and 6m time points (p<0.01); RDW and MCV values did not differ between cases and controls. A Cox regression model was fitted to the data to examine the association between t-MDS/AML and specific low peripheral blood parameter values at d100, 6m, and 1y post-aHCT (adjusting for year of aHCT, age at aHCT, primary diagnosis, race/ethnicity, sex and TBI). The following parameters (with their cutpoints) emerged as significantly associated with increased risk of t-MDS/AML: Hgb<12, Plt<100k, and RBC<3. Compared with patients with Hgb, Plt and RBC values above these cutpoints at d100, 6m, or 1y, those with low values for all three blood parameters had a significantly increased risk of developing t-MDS/AML (d100: HR=10.5, 95%CI, 2.1-53.1, p=0.004; 6m: HR=9.9, 95%CI, 1.9-53.1, p=0.007; 1y: HR=10.7, 95%CI, 1.5-74.0, p=0.02). In summary, we consistently observed lower values for specific blood parameters during the post-aHCT period among patients who subsequently developed t-MDS/AML as compared to controls across multiple timepoints post-aHCT. These differences appeared soon after aHCT, persisted, and preceded the development of t-MDS/AML, providing evidence that bone marrow injury long predates the development of t-MDS/AML. Importantly, the combination of low Hgb, Plt, and RBC values at d100, 6m, or 1y predicted future development of t-MDS/AML, and could be used as early indicators of t-MDS/AML risk and the need for close monitoring.Abstract 2507. TablePeripheral blood parametersDay 100(HR, 95%CI)p-value6 month(HR, 95%CI)p-value1 year(HR, 95%CI)p-valueAll normalHgb≥12 and Plt ≥100 and RBC ≥31.000.0041.000.0071.000.02All lowHgb <12 and Plt<100 and RBC <310.5 (2.1-53.1)9.9 (1.9-53.1)10.7 (1.5-74.0) [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Aspirin resistance: Analysis of the incidence amongst stable patients on prophylactic therapy versus unstable patients experiencing acute ischaemic events utilising the multiplate analyser in a tertiary institution

Aim To determine the incidence of aspirin resistance amongst stable patients treated with aspirin as primary prophylaxis for cardiovascular disease (control group) versus unstable patients experiencing acute arterial ischaemic events including acute coronary syndrome, transient ischaemic attack, cerebrovascular accident and critical limb ischaemia (event group). Methods Patients on aspirin therapy, presenting to cardiology outpatient departments, vascular surgical wards and coronary care units in atertiary institution were identified. Compliance with aspirin therapy was confirmed directly by verbal interview. Patient consent was obtained prior to the collection of peripheral blood samples into citrate and hirudin tubes. The samples were analysed to determine the platelet count and platelet aggregation response to arachidonic acid (ASPItest) and thrombin receptor activating peptide (TRAPtest) using impedence aggregometry on the Multiplate analyser Results Results of statistical analysis to follow Discussion Correlation between the incidence of aspirin resistance amongst stable versus unstable patients can be used to assess the role of the Multiplate analyser in the laboratory and the implications of these results in guiding patient therapy.

Abstract 4719: Exploratory biomarker program in Phase I clinical development of RG7155, a novel humanized anti-CSF-1R Mab, targeting Tumor Associated Macrophages (TAMs).

Abstract Early Phase I clinical development of RG7155, a humanized anti-CSF-1R monoclonal antibody binding to dimerization interface of CSF-1 receptor and inducing apoptosis in CSF-1R expressing macrophages is accompanied by an extensive biomarker program. Phase I biosampling aims to enable monitoring of mode of action, drug efficacy and potential tumor escape mechanisms by identifying response prediction and mechanistic efficacy markers and therefore to support optimal biological dose (OBD) definition and patient stratification for subsequent clinical testing. One biomarker hypothesis to support potential patient stratification assumes that response to treatment with RG7155 is enhanced in tumors with high CSF-1R positive macrophage infiltration and with high CSF-1 levels in the peripheral blood as the basis for tumor promoting macrophage recruitment and differentiation. The Phase I biomarker program comprises extensive biosampling including mandatory fresh baseline and on treatment tumor, skin biopsies as well as peripheral blood sampling to identify and monitor pharmacodynamic as well as mechanistic efficacy markers. Pre- and on-treatment tumor tissue analysis involves tumor infiltrating macrophage subtype characterization via CD68/CD163/CSF-1R/MHC II, CD4 and CD8 T cell assessment by multiplex IHC assays. In order to detect early tissue response prediction markers in surrogate tissue, wound associated macrophages (WAMs) are assessed in wounded skin tissue samples. Further, systematic collection of peripheral blood samples before and during RG7155 administration aims at identifying CSF-1R expressing target populations, other potential RG7155 susceptible immune cell populations by flow cytometry using CD8/CD4/CD3/CD45/MHC II/CD14/CD16/CD56/CD19 markers and an extended cytokine profiling by Luminex 17-plex panel. These markers will be correlated to direct anti-tumor effects assessed using KI67 IHC, FDG-PET, DC-Ultrasound and clinical response. Here we describe the development of the clinical biomarker strategy based on preclinical data and the specific assay developments, to enable the selection of the OBD, the correlation of TAM elimination with anti-tumor effects and the optimization of the biomarker sampling to minimize the burden of diagnostic procedures for patients in future clinical trials. Citation Format: Michael A. Cannarile, Kai Habben, Florian Heil, Ann-Marie Broeske, Dominik Ruettinger, Lisa Culton, Friedrich Feuerhake, Carola H. Ries. Exploratory biomarker program in Phase I clinical development of RG7155, a novel humanized anti-CSF-1R Mab, targeting Tumor Associated Macrophages (TAMs). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4719. doi:10.1158/1538-7445.AM2013-4719

Lower Volume Peripheral Blood Sample Collection: Key Considerations for Nurses - Roundtable Discussion

A group of seven nurses from a range of laboratory and hospital settings across the United States were invited to present opinions and clinical case study examples that reflect the key considerations in peripheral blood sample collection involving lower volume samples and patients affected by difficult venous access (DVA). Panelists were asked to review and compare prevailing standard operating procedures (SOPs) in sample collection and challenges in efforts to expand the use of lower volume sample collection and processing. The discussion identified achievable goals to improve standard of care in lower volume blood sample collection and the treatment of DVA patients.

Beta Thalassemia In Different Tribal Population In West Bengal, India

AbstractAbstract 5170 Background:The Thalassemias are a group of anemia that results from a genetic defect whch reduces the rate of synthesis of normal globin chains. Beta Thalassemia results in severe microcytic, hypochromic, haemolytic anaemia among affected patients. It has emerged as one of the most common health problems among the tribal populations in West Bengal. A project entitled, “Status of Prevalence and Awarness Among Thalassaemia Population: Special reference to various tribes belonging in different Districts of West Bengal” has been sanctioned by the Department of Biotechnology, Govt. of India(BT/PR10336/SPD/24/293/2007). This study is aimed to observe the spectrum of various beta mutations among the tribes. In the screening schedule thalassemia awarness programme was conducted in various rural areas of West Bengal. We mainly confronted with the Sardar tribes in extreme south of West Bengal. In North Bengal there were Toto, Rabha, Oraon, Munda, Toppo, Baraik, Nagbanshi, Khalko, Kheria, Kerketta and Soren. Method:In the year 2009–2010, 1289 tribes were screened. Mass awarness programme were followed by collection of peripheral blood samples in EDTA vials and transported to the laboratory in ice packs. NESTROFT was done on spot using 0.36% Saline Buffer solution (Sodium chloride, Sodium dihydrogen phosphate, Disodium hydrogen Phosphate). Complete Blood Count was performed within 24 hours of collection. HPLC (High Performance Liquid Chromatography) was done to identify the beta samples. DNA was extracted from the blood of beta carrier and beta major and subjected to ARMS (Amplification Refractory Mutation System) PCR (Polymerase Chain Reaction) to detect the point mutations. Result:Name of the TribeLocationIndividuals screenedNormalHbE CarrierHbE HomozygousBeta CarrierBeta PatientOthersTotoToto para, North Bengal548276195752NilNilRabhaRabha basti, North Bengal100303733NilNilNilOraonNorth Bengal147922241757MundaNorth Bengal119791751125ToppoNorth Bengal72471319Nil2NagbanshiNorth Bengal10364172182NilSardarSouthern part of Bengal2001231724810Nil Conclusion:Molecular Characterization of Beta globin gene mutations among these tribes have confirmed the presence of the following mutations: IVS-1 nt5 (G>C), IVS-1 nt1(G>T), codon 15 (G>A), codon 26 (G>A), the mutation which leads to HbE, codon 19 (A>G), codon 17 (A>T), -28 (A>G). The most common mutation observed among Totos and Rabhas were codon 26 (G>A) of North Bengal. The prevalence of Beta carrier among them is negligible. The mutation IVS-1 nt5 (G>C) is prevalent among the Oraon, Nagbanshi and Sardar tribes of Bengal. The other mutations which are present among them are codon 15 (G>A) and codon 30 (G>C). There are two codon 17 (A>T) and one -28 (A>G) mutations which are present among Mundas. The rest of them are mutation IVS-1 nt5 (G>C). In few cases presence of sickle cell anemia was observed. Our results have added to the existing data on the common beta globin gene defects which are prevalent among the general population of West Bengal, India. Disclosures:No relevant conflicts of interest to declare.

P164 Enabling next day processing of blood samples with T-SPOT.TB

IntroductionThe T-SPOT®.TB assay involves the collection of peripheral blood samples for analysis of the T cell response to TB-specific antigens. Traditionally the blood sample has been required to be processed...

Genetic characteristics of familial type 2 diabetes pedigrees: a preliminary analysis of 4468 persons from 715 pedigrees

To analyze the inheritance character of type 2 diabetes mellitus (T2DM) pedigrees. 4468 persons from 715 T2DM pedigrees (including the spouses) undergo peripheral blood sample collection to examine blood sugar and physical examination. Questionnaire survey was conducted to explore the family history. Type 1 DM and maturity-onset DM of young people were to be ruled out. Pedigree chart were made. The prevalence rates of T2DM and impaired glucose regulation (IGR) was 47.62%, including 218 T2DM and 422 IGR newly discovered. The prevalence rates of T2DM and IGR were 38.33% and 14.25% in the siblings, and 56.81% and 12.58% in the parents, all significantly higher than those in the second-degree relatives (9.55% and 6.10%) and spouses (10.57% and 9.55% respectively, all P < 0.01). The prevalence and newly discovered rates of IGR in the offspring were 12.46% and 11.73% respectively, both significantly higher than those in the spouses (9.55% and 9.55% respectively, all P < 0.01). There is significant familial aggregation in T2DM. The first-degree relatives of T2DM patients are high risk populations, so long term monitoring and early screening should be performed.