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Peripheral Blood Samples (peripheral + blood_sample)
Selected AbstractsAn examination of different fetal specific antibodies and magnetic activated cell sorting for the enrichment of fetal erythroblasts from maternal bloodCONGENITAL ANOMALIES, Issue 3 2002Xiao Xi Zhao ABSTRACT, The aim of the present study was to compare the rates of fetal cells obtained after separation from maternal blood by magnetic activated cell sorting (MACS) using different fetal specific antibodies, and to evaluate the potential role of this method in the prenatal diagnosis of fetal trisomies. Peripheral blood samples were obtained from 42 women carrying chromosomally normal fetuses and from 4 women with aneuploid fetuses (2 cases of 47,XX,+18 and 2 of 47,XY,+21) at 9,20 weeks of gestation. After fetal cells were enriched by MACS with three different monoclonal antibodies (GPA, CD71, CD14), fluorescence in situ hybridization (FISH) with chromosome X, and Y-specific probes was performed to detect the rates of fetal cells in the samples sorted. FISH with chromosome 13-, 18-, and 21-specific probes was carried out to compare proportions of cells with three-signal nuclei in chromosomally normal and abnormal groups. In male infants, X-and Y-positive cells were detected in 80%, 73.3%, and 66.6% of samples after the separation by antibodies CD14, GPA, and CD71, respectively. The percentage of nuclei with three signals was increased in pregnancies with trisomy, ranging between 2% and 5.18%. Pregnancies with normal fetuses showed 0 to 3.7% of nuclei with three signals. The data demonstrate that fetal cell detection varies depending on the antibodies used for cell sorting. This study provides further evidence on the feasibility of screening for fetal chromosomal abnormalities by enriching maternal blood for fetal cells and using FISH. [source] Total nucleated cell differential for blood and bone marrow using a single tube in a five-color flow cytometer,,CYTOMETRY, Issue 2 2008Sven Björnsson Abstract Background: Flow cytometry allows the use of several antibodies in addition to light scatter, and most flow cytometers will provide at least seven measurements on each cell passing through the laser beam. A skilled microscopist will classify at least 14 cell classes in bone marrow or blood. Our goal was to use the seven parameters available in our flow cytometer to provide a reliable differential count using only one tube. Methods: Peripheral blood samples were analyzed on the Beckman Coulter LH750 cell counter, and the flagging and messages from the cell counter were used to select normal or pathological samples. Samples without flags (N = 50), with >2% erythroblasts (N = 80), or with "Blast" or "Verify diff" flags (N = 54) were investigated. We used a lyse-no-wash method to ensure minimal loss of fragile cells with live gating on DRAQ5-positive cells to acquire only nucleated cells. The FL-1 to FL-4 channels were used for the antibodies CD36-FITC, CD203-PE, CD138-PE, CD45-ECD, CD16-Pcy5, and CD56-Pcy5. FL-5 was used for the DNA-stain DRAQ5. Results: Using live gate acquisition on DRAQ5, we were able to classify total nucleated cells into 10 classes. We were unable to identify megakaryocytes, but platelets could be studied by rerunning the sample after dilution and gating on DRAQ5-negative CD36-posive events. Validation against digitized microscopy and cell counter showed linear correlations within each cell class with correlation coefficients that seem reasonable for cellular classification. The lowest correlation was found for basophil granulocytes. Flow cytometry detected twice as many immature neutrophils compared to microscopy. Conclusions: We have designed a one-tube immunophenotyping panel for classification of total nucleated cells and platelets in blood or bone marrow. The seven parameters available in one single tube in our cytometer seem to be enough for reliable differential count even in difficult pathological samples. The analytical imprecision of the flow cytometer differential was much lower than that obtained with microscopy or cell counter differentials. © 2007 Clinical Cytometry Society. [source] Lack of genotoxicity induced by endogenous and synthetic female sex hormones in peripheral blood cells detected by alkaline comet assayENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 5 2007Mariana Gobbo Braz Abstract The etiology of hormone-induced cancers has been considered to be a combination of genotoxic and epigenetic events. Currently, the Comet assay is widely used for detecting genotoxicity because it is relatively simple, sensitive, and capable of detecting various kinds of DNA damage. The present study evaluates the genotoxic potential of endogenous and synthetic sex hormones, as detected by the Comet assay. Blood cells were obtained from 12 nonsmoking and 12 smoking women with regular menstrual cycles and from 12 nonsmoking women taking low-dose oral contraceptives (OC). Peripheral blood samples were collected at three phases of the menstrual cycle (early follicular, mean follicular, and luteal phases), or at three different moments of oral contraceptive intake. Three blood samples were also collected from 12 healthy nonsmoking men, at the same time as oral contraceptive users. Results showed no significant difference in the level of DNA damage among the three moments of the menstrual cycle either in nonsmoking and smoking women, or between them. No significant difference in DNA damage was also observed among oral contraceptive users, nonusers, and men. Together, these data indicate lack of genotoxicity induced by the physiological level of the female sex hormones and OC as assessed by the alkaline Comet assay. In conclusion, normal fluctuation in endogenous sex hormones and use of low-doses of oral contraceptive should not interfere with Comet assay data when this technique is used for human biomonitoring. Environ. Mol. Mutagen., 2007. © 2007 Wiley-Liss, Inc. [source] Splenic function and IgM-memory B cells in Crohn's disease patients treated with infliximabINFLAMMATORY BOWEL DISEASES, Issue 5 2008Antonio Di Sabatino MD Abstract Background: Under experimental chronic inflammation, tumor necrosis factor (TNF)-, plays a role in damaging spleen marginal zone. This latter has a crucial function in mounting B cell-dependent immune responses against infections by encapsulated bacteria. In Crohn's disease (CD), a chronic inflammatory disorder where TNF-, is centrally involved, impaired splenic function may increase the susceptibility to bacterial infections. On this basis, we aimed to investigate the influence of anti-TNF therapy on splenic function in CD patients. Methods: Peripheral blood samples were obtained from 15 CD patients before and after treatment with infliximab administered at weeks 0, 2, and 6 at a dose of 5 mg/kg. Counting of erythrocytes with membrane abnormalities (pitted red cells) was used as an indicator of splenic function. Multicolor flow cytometry was performed to analyze circulating B cells. Results: A substantial clinical improvement in 10 of the 15 CD patients was associated with a significant reduction of pitted red cells (from median 6.0% to 3.6%; P < 0.01) after 10 weeks of treatment. In responder patients the improvement of splenic function was accompanied by a parallel increase of circulating IgM-memory B cells (from median 6.9% to 13.3%; P < 0.005). Splenic function was not ameliorated in nonresponder patients. Conclusions: Splenic function improved in CD patients who responded to infliximab and was accompanied by a concomitant restoration of the IgM-memory B cell pool responsible for the protection against encapsulated bacteria. Restoration of splenic function after infliximab treatment is intriguing and requires further investigation. (Inflamm Bowel Dis 2008) [source] In-vitro Fertilization and Embryo Transfer and Cellular Immunity: Study on Cytokines and T Lymphocyte subpopulations in IVF-ET CyclesJOURNAL OF OBSTETRICS AND GYNAECOLOGY RESEARCH (ELECTRONIC), Issue 3 2002Mitsutaka Murakami Objectives: To determine whether peripheral T lymphocyte subpopulations and cytokines change during in-vitro fertilization and embryo transfer (IVF-ET) cycles and to evaluate them with regard to pregnancy status and types of infertility. Methods: Peripheral T lymphocyte subpopulations and cytokines in 33 consecutive cycles of IVF-ET were examined. All the women were stimulated with purified FSH and hCG after pituitary suppression with GnRH agonist. Peripheral blood samples were collected before FSH administration, on the day of hCG administration, the day of ET (day 2), day 6 and day 15. We measured plasma estradiol and progesterone levels and plasma interferon-,, interleukin-4 (IL-4), IL-10 and IL-12 levels. Peripheral T lymphocyte subpopulations, T helper type 1 and 2 cells (Th1 and Th2) and T cytotoxic type 1 and 2 cells (Tc1 and Tc2), were analyzed with three-color flowcytometry. Results: There were no changes in the plasma levels of the cytokines or in the proportions of Th1 and Th2 and the proportions of Tc1 and Tc2 in peripheral blood lymphocytes during the IVF-ET cycles. In women with endometriosis, the ratios of Tc1 to CD8+ lymphocytes and the Tc1 to Tc2 ratios before FSH administration were much higher than in women without endometriosis. The ratios of Tc1 to CD8+ lymphocytes were significantly lower in the patients with endometriosis who became pregnant. Conclusions: Peripheral cellular immunity does not change during IVF-ET cycles. In women with endometriosis, the peripheral Tc1 subpopulation is more predominant before ovarian stimulation, suggest- ing that the ratio of Tc1 before ovarian stimulation could be an indicator of fecundity for women with endometriosis. [source] Cell-free mRNA concentrations of CRH, PLAC1, and selectin-P are increased in the plasma of pregnant women with preeclampsiaPRENATAL DIAGNOSIS, Issue 8 2007Yuditiya Purwosunu Abstract Objective To compare mRNA concentrations of corticotrophin-releasing hormone (CRH), placenta specific-1 (PLAC1), and selectin-P in preeclamptic and normal pregnancies. Methods Peripheral blood samples were obtained from 43 pregnant women with preeclampsia and 41 control subjects. Plasma was harvested from samples and RNA extracted. Plasma RNA was analyzed using reverse transcription polymerase chain reaction (PCR) assay. Median concentrations of CRH, PLAC1, and selectin-P mRNA in plasma were compared, to assess possible differences in distribution. Data were also stratified and compared according to clinical severity of preeclampsia. Finally, CRH, PLAC1, and selectin-P were plotted against quantitative distributions of blood pressure and proteinuria. Results All markers were differently distributed between cases and controls. Median values in subgroups correlated with severity of preeclampsia. All markers correlated with both. Selectin-P was identified as the marker with the highest degree of correlation. No correlation was found between any markers in the control group and proteinuria or blood pressure. Conclusion CRH, PLAC1, and selectin-P are distributed differently in preeclampsia cases compared to controls and correlate with signs of preeclampsia. Copyright © 2007 John Wiley & Sons, Ltd. [source] Melanoma antigen-1 mRNA combined with ,-fetoprotein mRNA levels in peripheral blood of patients with hepatocellular carcinoma: a predictor of postoperative recurrence or metastasis?ANZ JOURNAL OF SURGERY, Issue 1-2 2009Yuqing Zhang Abstract Background:, The aim of the study was to identify whether melanoma antigen (MAGE)-1 mRNA and ,-fetoprotein (AFP) mRNA expressed in peripheral blood could be used to predict the recurrence and metastasis of hepatocellular carcinoma (HCC) after hepatectomy. Methods:, One hundred and forty-two HCC patients underwent hepatectomy. The control group includes 27 patients with chronic virus hepatitis and cirrhosis and 10 healthy volunteers. Peripheral blood samples were collected on the seventh day before operation, seventh day after operation and 30th day after operation. MAGE-1 mRNA and AFP mRNA were tested by nested reverse transcription polymerase chain reaction. Median follow up was 25.5 months (range 4,40 months). Patient survival, disease-free survival and clinicopathological features were compared between patients with positive and negative MAGE-1 mRNA and/or AFP mRNA. Results:, The expression of MAGE-1 mRNA and/or AFP mRNA in peripheral blood was closely correlated to the pathological stage and the positive ratio of tumour cells in the peripheral blood (P < 0.01). There was recurrence and/or metastasis after operation in 55 of 142 HCC patients. Among the 55 patients who had recurrence or metastasis, MAGE-1 mRNA and/or AFP mRNA in peripheral blood were persistently detected after operation in 38 patients and MAGE-1 mRNA and AFP mRNA turned to positive after operation in 14 patients. In contrast, no recurrence was found in 62 patients whose MAGE-1 mRNA and/or AFP mRNA turned to negative after operation. 88.1% (52 of 59) of patients with MAGE-1 mRNA and/or AFP mRNA persistently positive after operation showed recurrence or metastasis, whereas only 3.6% patients (3 of 83) with the negative of MAGE-1 mRNA and/or AFP mRNA after operation showed recurrence or metastasis (P < 0.001). Conclusion:, Melanoma antigen-1 mRNA combined with AFP mRNA in peripheral blood after hepatectomy is more sensitive and specific than AFP mRNA singly for predicting the recurrence and metastasis of the HCC patients, whereas preoperative transient detection is not. [source] A prospective study of diagnosis of Toxoplasma gondii infection after bone marrow transplantation,APMIS, Issue 5 2008BENJAMIN EDVINSSON Active infection with Toxoplasma gondii in immunocompromised transplant recipients can lead to toxoplasmosis, which may have a rapid disease course and in some cases be fatal. It is of paramount importance to diagnose toxoplasmosis at an early stage, and to initiate specific treatment to improve the outcome. Polymerase chain reaction (PCR) is today the primary diagnostic tool to diagnose toxoplasmosis in immunocompromised patients. Timely diagnosis may, however, be difficult if toxoplasmosis is at first asymptomatic. To investigate the magnitude of toxoplasmosis after bone marrow transplantation (BMT), we conducted a screening study by PCR where 21 autologous and 12 allogeneic BMT recipients were included. Peripheral blood samples were taken one week prior to BMT; thereafter, blood samples were drawn weekly for the first 6 months, and monthly up to one year after BMT. The samples were analyzed by conventional PCR and real-time PCR. T. gondii DNA was detected in peripheral blood from one patient 5 days post allogeneic BMT. There were no clinical signs of toxoplasmosis. Medical records were reviewed and showed a previously undiagnosed eye infection in another allogeneic BMT recipient. These two patients were seropositive for T. gondii. We concluded that monitoring for T. gondii DNA in peripheral blood samples using PCR might be a valuable method for identifying toxoplasma-seropositive stem cell transplant recipients. [source] Interleukin-6 and type I interferon,regulated genes and chemokines mark disease activity in dermatomyositisARTHRITIS & RHEUMATISM, Issue 11 2009Hatice Bilgic Objective Up-regulation of whole blood type I interferon (IFN),driven transcripts and chemokines has been described in a number of autoimmune diseases. An IFN gene expression "signature" is a candidate biomarker in patients with dermatomyositis (DM). This study was performed to evaluate the capacity of IFN-dependent peripheral blood gene and chemokine signatures and levels of proinflammatory cytokines to serve as biomarkers for disease activity in adult and juvenile DM. Methods Peripheral blood samples and clinical data were obtained from 56 patients with adult or juvenile DM. The type I IFN gene signature in the whole blood of patients with DM was defined by determining the expression levels of 3 IFN-regulated genes (IFIT1, G1P2, and IRF7) using quantitative real-time reverse transcription,polymerase chain reaction. Multiplexed immunoassays were used to quantify the serum levels of 4 type I IFN,regulated chemokines (IFN-inducible T cell , chemoattractant, IFN,-inducible 10-kd protein, monocyte chemotactic protein 1 [MCP-1], and MCP-2) and the serum levels of other proinflammatory cytokines, including interleukin-6 (IL-6). Results DM disease activity correlated significantly with the type I IFN gene signature (r = 0.41, P = 0.007) and with the type I IFN chemokine signature (r = 0.61, P < 0.0001). Furthermore, the serum levels of IL-6 were significantly correlated with disease activity (r = 0.45, P = 0.001). In addition, correlations between the serum levels of IL-6 and both the type I IFN gene signature (r = 0.47, P < 0.01) and the type I IFN chemokine signature (r = 0.71, P < 0.0001) were detected in patients with DM. Conclusion These results suggest that serum IL-6 production and the type I IFN gene signature are candidate biomarkers for disease activity in adult and juvenile DM. Coregulation of the expression of IFN-driven chemokines and IL-6 suggests a novel pathogenic linkage in DM. [source] Gene expression signatures in polyarticular juvenile idiopathic arthritis demonstrate disease heterogeneity and offer a molecular classification of disease subsetsARTHRITIS & RHEUMATISM, Issue 7 2009Thomas A. Griffin Objective To determine whether peripheral blood mononuclear cells (PBMCs) from children with recent-onset polyarticular juvenile idiopathic arthritis (JIA) exhibit biologically or clinically informative gene expression signatures. Methods Peripheral blood samples were obtained from 59 healthy children and 61 children with polyarticular JIA prior to treatment with second-line medications, such as methotrexate or biologic agents. RNA was extracted from isolated mononuclear cells, fluorescence labeled, and hybridized to commercial gene expression microarrays (Affymetrix HG-U133 Plus 2.0). Data were analyzed using analysis of variance at a 5% false discovery rate threshold after robust multichip analysis preprocessing and distance-weighted discrimination normalization. Results Initial analysis revealed 873 probe sets for genes that were differentially expressed between polyarticular JIA patients and healthy controls. Hierarchical clustering of these probe sets distinguished 3 subgroups within the polyarticular JIA group. Prototypical patients within each subgroup were identified and used to define subgroup-specific gene expression signatures. One of these signatures was associated with monocyte markers, another with transforming growth factor ,,inducible genes, and a third with immediate early genes. Correlation of gene expression signatures with clinical and biologic features of JIA subgroups suggested relevance to aspects of disease activity and supported the division of polyarticular JIA into distinct subsets. Conclusion Gene expression signatures in PBMCs from patients with recent-onset polyarticular JIA reflect discrete disease processes and offer a molecular classification of disease. [source] The clinical utility of the prostate specific membrane antigen reverse-transcription/polymerase chain reaction to detect circulating prostate cells: an analysis in healthy men and womenBJU INTERNATIONAL, Issue 9 2002L. Llanes Objective,To evaluate the overall specificity of nested reverse transcriptase-polymerase chain reaction (RT-PCR) to detect prostate-specific membrane antigen (PSM) mRNA in peripheral blood samples of healthy donors. Subjects and methods,Peripheral blood samples were taken from 60 healthy blood-donors (30 men and 30 women aged < 50 years) and analysed for PSM-mRNA using nested RT-PCR (in ,hot-start' conditions and confirmed using nested EcoRI restriction enzyme). Intron-spanning primer pairs specific for human PSM were deduced from the GenBank sequence (M99487) using gene software. The outer primer pair for PSM was: fwd: 1368 5,-TCACCGGGACTCATGGGTGT-3,; reverse: 1860 5,-GCCTGAAGCAATTCCAAGTCGG-3,. Inner primer pair for PSM was: fwd: 1480 5,-AAGGAAGGGTGGAGACCTAG-3,; reverse: 5-ACTGAACTCTGGGGAAGGAC-3,. The integrity of cDNAs was checked using primer pairs specific for the housekeeping gene ,-actin. The specificity and false-positive rate were calculated assuming that the underlying prostate cancer incidence was nil. Results,The first PCR was negative for all samples (100% specificity; 0% false-positive rate). The nested PCR detected 23 positive samples (23/60, 38%) with an overall specificity of 62% (false positive rate, 38%). Conclusion,Nested RT-PCR of PSM-mRNA in peripheral blood is highly unspecific. Its clinical utility in the management of prostate cancer must be low. Further development is needed of quantitative RT-PCR, primers that identify prostatic PSM or another prostate-specific marker gene to differentiate PSM mRNA from circulating prostate cells and from non-prostatic tissues. [source] An assessment of the immunological status of patients with renal cell carcinoma based on the relative abundance of T-helper 1- and -2 cytokine-producing CD4+ cells in peripheral bloodBJU INTERNATIONAL, Issue 9 2001T. Onishi Objective To assess the immunological status of patients with renal cell carcinoma (RCC), by analysing the proportion of cluster-of-differentiation 4-positive (CD4+) cells showing intracellular cytokine production, i.e. interferon-, derived from T-helper (Th) 1 and interleukin-4 derived from Th2 cells, among peripheral blood lymphocytes from these patients Patients, subjects and methods Peripheral blood samples (5 mL) were collected from 36 patients (mean age 61 years, range 44,78) with RCC before and after they underwent nephrectomy. The proportion of cytokine-producing CD4+ cells was determined by flow cytometric analysis after stimulating the cells with phorbol 12-myristate 13-acetate, ionomycin and brefeldin A, and staining the cells with fluorescein isothiocyanate-labelled anti-interferon-,, anti-interleukin-4 and anti-immunoglubulin-2b antibodies. The results were expressed as the percentage of cytokine-producing cells in the CD4+ population. As a control, peripheral blood obtained from 35 healthy volunteers (mean age 34 years, range 22,49) was also analysed. Results The proportion of CD4+ cells producing interferon-, and interleukin-4 was significantly higher (P < 0.04 and P < 0.001, respectively) in patients with RCC than in controls. The Th1/Th2 ratio (i.e. the ratio of CD4+ cells producing each cytokine) was significantly lower in patients with RCC (P < 0.001). There was a significant correlation in the controls between interferon-, and interleukin-4 production (r = 0.489, P < 0.01) but not in patients with RCC. The proportion of CD4+ cells producing interleukin-4 was significantly higher and the Th1/Th2 ratio significantly lower in patients with high-stage than in those with low-stage RCC (P < 0.05). The percentage of CD4+ cells producing interleukin-4 was significantly less after nephrectomy in those with low-stage RCC (P < 0.01) and the Th1/Th2 ratio significantly greater (P < 0.05) than before nephrectomy; there was no such trend in patients with high-stage RCC. Conclusion An evaluation of the production of interferon-, and interleukin-4 in CD4+ peripheral blood lymphocytes is useful for assessing the immunological status of patients with RCC; there is a change in the predominant response from Th1 to Th2 with increasing stage of RCC. [source] Intraoperative decay profile of intact (1,84) parathyroid hormone in surgery for secondary hyperparathyroidism in a consecutive series of 50 patients on haemodialysisBRITISH JOURNAL OF SURGERY (NOW INCLUDES EUROPEAN JOURNAL OF SURGERY), Issue 9 2000J. Lokey Background The usefulness of rapid intraoperative monitoring of intact (1,84) parathyroid hormone (PTH) is not clearly defined in the surgical management of secondary HPT in the patients on haemodialysis. The aim of this study was to define the normal pattern of decay during surgery for secondary HPT using the rapid intact (1,84) PTH assay during operation. Methods Fifty patients on haemodialysis underwent neck exploration for secondary HPT. The therapeutic goal in all patients was the subtotal resection of four or more glands and bilateral transcervical thymectomy. PTH levels were monitored using a rapid immunochemiluminometric assay. Peripheral blood samples were assayed at induction of anaesthesia, after dissection but before resection, and 20 and 40 min after resection in all patients. All patients were followed up for at least 6 months. PTH levels were expressed as absolute values, as multiples of the upper limit of normal and as the percentage decline from pre-excision values. Results Forty-eight patients (96 per cent) were considered cured after surgery. Twenty patients (40 per cent) had a PTH level less than twice normal and 20 patients (40 per cent) had a PTH level between two and four times normal at 20 min. At late follow-up, all these patients were cured. Ten patients (20 per cent) had a PTH level greater than four times normal at 20 min. Eight of these patients were cured. Seven of these eight had a PTH level at 20 min, while not less than four times normal, less than 40 per cent of the original value. In contrast, the two failures had neither a decline to less than four times normal nor a decay to less than 40 per cent of the original value. One has been reoperated with resection of a fifth gland and one awaits reoperation. Conclusion The intraoperative decay of PTH during surgery for secondary HPT in patients on haemodialysis is slower than that in patients with normal renal function. However, 20 min after resection, a decline to less than four times the upper limit of normal is predictive of cure. Variability of decay slopes in individual patients may reflect molecular heterogeneity or biphasic metabolism of the hormone. © 2000 British Journal of Surgery Society Ltd [source] Assessment of in vitro immunity to Mycobacterium tuberculosis in a human peripheral blood infection model using a luciferase reporter construct of M. tuberculosis H37RvCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 3 2006R. Al-Attiyah Summary Protective immune responses to tuberculosis in man are primarily cell-mediated and require the interaction of specific T cells, cytokines and activated macrophages. In the present study, Mycobacterium tuberculosis H37Rv labelled with luciferase reporter enzyme was used to analyse the anti-mycobacterial immunity in man using an in vitro whole blood infection model. Peripheral blood samples obtained from M. bovis bacille Calmette,Guérin (BCG)-vaccinated tuberculin-positive healthy volunteers (n = 23) were cultured with M. tuberculosis H37Rv reporter strain. The growth of bacteria in the whole blood cultures was monitored after 48 and 96 h of infection. The results showed that the growth of M. tuberculosis was significantly inhibited after 96 h (P < 0·029) of culture. Among the cytokines studied, interleukin (IL)-10 and IL-12 were not detected at all, whereas low levels of interferon (IFN)-, after 96 h (0·4 IU/ml) and tumour necrosis factor (TNF)-, after 48 (135 pg/ml) and 96 h (47 pg/ml) of culture were detected in the supernatants of whole blood infected with M. tuberculosis. The magnitude of bacterial growth correlated directly with the concentration of TNF-, detected after 48 h (r = 0·722) and 96 h (r = 0·747) of culture (P , 0·0001 and P , 0·0001, respectively). However, the addition of monoclonal antibodies specific to TNF-, and IFN-, to the blood cultures did not alter mycobacterial growth indicating the role of other mechanisms/factors in restricting the growth of M. tuberculosis in whole blood cultures. [source] Inflammatory mediators in perinatal asphyxia and infectionACTA PAEDIATRICA, Issue 2002M Xanthou Aim: To determine serum levels of interleukin-6 (IL-6), IL-1,, tumor necrosis factor-, (TNF-,), soluble intercellular adhesion molecule-1 (sICAM-1) and C-reactive protein (CRP) in asphyxiated neonates and compare these inflammatory factors with those found in neonates with perinatal infection. Methods: 88 neonates were studied, of whom 36 were asphyxiated, 18 were infected and the remaining 34 were controls. Peripheral blood samples were obtained on the 1st, 3rd and 5th postnatal days. Results: Cytokines IL-6 and IL-1, as well as sICAM-1 serum levels did not differ between asphyxiated and infected neonates; however, at most time periods, their values were significantly higher than controls. TNF-, was similar in the three groups of neonates. CRP serum values were significantly higher in the infected neonates than in the asphyxiated or control subjects. Among the 54 asphyxiated and infected neonates, 15 were considered as severe cases and 39 as mild. The severe cases, at most time periods, had significantly higher IL-6, IL-1, and sICAM-1 levels compared with the mild ones. Through receiver operating characteristic curves the cut-off points, sensitivities, and specificities for distinguishing neonates at risk or at high risk for brain damage were established. Conclusion: Similar increases in serum levels of IL-6, IL-1, and sICAM-1 were found in perinatally asphyxiated and infected neonates. As these increases correlated with the severity of the perinatal insults, neonates at high risk for brain damage might be detected. [source] An optimized whole blood method for flow cytometric measurement of ZAP-70 protein expression in chronic lymphocytic leukemiaCYTOMETRY, Issue 4 2006T. Vincent Shankey Abstract Background: ZAP-70 protein expression has been proposed as a marker for immunoglobulin heavy chain mutational status, which some studies have correlated with disease course in B-cell chronic lymphocytic leukemia (CLL). Studies published to date measuring levels of expression of ZAP-70 intracellular protein using flow cytometry have demonstrated poor performance, as defined by the difference in signal in known positive and negative lymphocyte populations. Methods: A recently published method (Chow S, Hedley DW, Grom P, Magari R, Jacobberger JW, Shankey TV, Cytometry A 2005;67:4,17) to measure intracellular phospho-epitopes was optimized using a design of experiments (DOE) approach to provide the best separation of ZAP-70 expression in positive T- or NK-cells as compared to negative B-cells in peripheral blood samples. A number of commercially available anti-ZAP-70 antibody-conjugates were screened using this methodology, and the antibody-conjugate showing the best performance was chosen to develop a four-color, five antibody assays to measure ZAP-70 levels in whole blood specimens. Results: Using the optimized fixation and permeabilization method, improvement in assay performance (signal-to-noise, S/N) was seen in most of the antibodies tested. The custom SBZAP conjugate gave the best S/N when used in conjunction with this optimized fixation /permeabilization method. In conjunction with carefully standardized instrument set-up protocols, we obtained both intra- and interlaboratory reproducibility in the analysis of ZAP-70 expression in whole blood samples from normal and CLL patients. Conclusions: The development of a sensitive, specific and highly reproducible ZAP-70 assay represents only the first essential step for any clinical assay. The universal implementation of a validated data analysis method and the establishment of methodology-based cutoff points for clinical outcomes must next be established before ZAP-70 protein analysis can be routinely implemented in the clinical laboratory. © 2006 International Society for Analytical Cytology [source] Remission induction chemotherapy induces in vivo caspase-dependent apoptosis in bone marrow acute myeloid leukemia blast cells and spares lymphocytesCYTOMETRY, Issue 3 2006J.-P. Vial Abstract Background The goal of new therapeutic strategies is to adapt the treatment of acute myeloid leukemia (AML) patients to the prognostic and/or to the hematological response. Methods We analyzed in vivo apoptosis induction in blast cells and in lymphocytes of AML patients receiving remission induction treatment. Results We show, on 12 peripheral blood samples, that the increase of peripheral apoptotic blast cells cannot be considered as the earliest marker of the treatment efficiency, because the significant increase of apoptosis followed the white blood cell and the peripheral blast cell count reductions, probably due to an efficient clearance of circulating apoptotic cells. Furthermore, the study of 65 bone marrow samples at d15 showed that the treatment induced apoptosis of blast cells while sparing the lymphocytes. This apoptosis was evidenced both at the caspase and at the membrane levels using respectively fmk-VAD-FITC and Annexin V binding assays. We found that less than 50% of apoptosis, measured with the fmk-VAD-FITC, in the d15 residual bone marrow blast cells, correlated with lower disease-free survival probability. Conclusion More studies are needed in larger series and earlier during the remission induction treatment to confirm the possible prognostic significance of in vivo apoptosis induction. © 2006 International Society for Analytical Cytology [source] Feasibility of conducting the micronucleus test in circulating erythrocytes from different mammalian species: An anatomical perspectiveENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 9 2006Ion Udroiu Abstract The in vivo mammalian micronucleus test can be conducted easily on peripheral blood samples since the maturation of erythrocytes involves the loss of the major nucleus. In addition, mature erythrocytes are relatively long-lived, so that the test potentially can detect genotoxic damage caused by chronic exposures. However, some species have spleens that remove micronuclei from the peripheral circulation, making such measurements problematical. This report summarises haematological and mutagenesis studies dealing with this subject and provides an anatomical interpretation of the phenomenon. Anatomical features can be used to identify those species in which micronuclei are removed by the spleen. Environ. Mol. Mutagen., 2006. © 2006 Wiley-Liss, Inc. [source] DNA sequence analysis of interlocus recombination between the human T-cell receptor gamma variable (GV) and beta diversity-joining (BD/BJ) sequences on chromosome 7 (inversion 7)ENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 2 2002Scott W. Ballinger Abstract V(D)J recombinase-mediated recombination between the T-cell receptor (TCR) gamma variable (GV) genes at chromosome 7p15 and the TCR beta joining (BJ) genes at 7q35 leads to the formation of a hybrid TCR gene. These TCR gamma/beta interlocus rearrangements occur at classic V(D)J recombination signal sequences (RSS) and, because the loci are in an inverted orientation, result in inversion events that are detectable in the chromosome structure as inv(7)(p15;q35). Similar rearrangements involving oncogenes and either TCR or immunoglobulin genes mediated by the V(D)J recombinase are found in lymphoid malignancies. Oligonucleotide primers that allow polymerase chain reaction (PCR) amplification across the inv(7) genomic recombination junction sequence have been described. Southern blot analysis has been primarily used to confirm the GV/BJ hybrid nature of the product, with limited information on the DNA sequence of these recombinations. We have modified this PCR method using total genomic DNA from the mononuclear cells in peripheral blood samples to increase specificity and to allow direct sequencing of the translocation junction that results from the recombination between the GV1 and BJ1 families of TCR genes in 25 examples from 11 individuals (three adults, one child, six newborns, and one ataxia telangiectasia (AT) patient). We focused on samples from newborns based on previous studies indicating that the predominant hypoxanthine-guanine phosphoribosyl transferase (HPRT) mutations in newborns are V(D)J recombinase-mediated deletion events and that the frequency of these mutations decreases with increasing age. Although the dilution series-based PCR assay utilized does not yield sharply defined quantitative endpoints, results of this study strongly suggest that inv(7) recombinations in newborns occur at equal or lower frequencies than those seen in adults. Consistent with the PCR primer pairs, all sequenced products contain a GV1 and a BJ1 segment and most also contain a BD1 segment. GV1s2 and 1s4 were the most frequently found GV1 genes (8 and 9 examples, respectively) and BJ1s5 and 1s6 were the most frequently found BJ1 genes (9 and 10 examples, respectively). These results demonstrate the effectiveness of this methodology for assessing GV/BJ interlocus rearrangements mediated by V(D)J recombinase. Environ. Mol. Mutagen. 40:85,92, 2002. © 2002 Wiley-Liss, Inc. [source] Human hematopoietic stem/progenitor-enriched CD34+ cells are mobilized into peripheral blood during stress related to ischemic stroke or acute myocardial infarctionEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 6 2005E. Paczkowska Abstract:, The hematopoietic and non-hematopoietic stem/progenitor cells harvested directly from the bone marrow (BM) or G-CSF mobilized peripheral blood were demonstrated to play an important role in regeneration of damaged organs (1, 2). Here, we asked if the stroke- or acute heart infarct-related stress triggers mobilization of stem/progenitor-enriched CD34+cells from the BM into the peripheral blood, which subsequently could contribute to regeneration of damaged tissues. To address this question the peripheral blood samples were harvested from patients with ischemic stroke during the first 24 h of manifestation of symptoms and on the second and sixth day afterwards or during the first 24 h of acute cardiac pain as well as on the second and sixth day of infarct. We measured in these patients (i) percentage of circulating hematopoietic stem/progenitor-enriched CD34+ cells in peripheral blood by employing fluorescence activated cell sorter (FACS) and (ii) number of hematopoietic progenitor cells for the granulocyte-monocytic colony-forming unit (CFU-GM) and erythoid burst-forming unit (BFU-E) lineages circulating in peripheral blood. We concluded that stress related to ischemic stroke or acute myocardial infarction triggers the mobilization of hematopoietic stem/progenitor-enriched CD34+ cells from the BM into peripheral blood. These circulating stem/progenitor-enriched CD34+ cells may contribute to the regeneration of ischemic tissues, however, this possibility requires further studies. [source] A study of a single variant allele (rs1426654) of the pigmentation-related gene SLC24A5 in Greek subjectsEXPERIMENTAL DERMATOLOGY, Issue 2 2009Gerasimos Dimisianos Abstract:, The SLC24A5 gene, the human orthologue of the zebrafish golden gene, has been shown to play a key role in human pigmentation. In this study, we investigate the prevalence of the variant allele rs1426654 in a selected sample of Greek subjects. Allele-specific polymerase chain reaction was performed in peripheral blood samples from 158 attendants of a dermatology outpatient service. The results were correlated with pigmentary traits and MC1R genotype. The vast majority of subjects (99%) were homozygous for the Thr111 allele. Only two subjects from the control group (1.26%) were heterozygous for the alanine and threonine allele. Both of these Thr111/Ala111 heterozygotes carried a single polymorphism of MC1R (one with the V92M variant and another with the V60L variant). Following reports of the rs1426654 polymorphism reaching fixation in the European population, our study of Greek subjects showed a prevalence of the Thr111 allele, even among subjects with darker skin pigmentation or phototype. [source] Genetic polymorphisms of drug-metabolizing enzymes CYP2D6, CYP2C9, CYP2C19 and CYP3A5 in the Greek populationFUNDAMENTAL & CLINICAL PHARMACOLOGY, Issue 4 2007Kostas Arvanitidis Abstract The aim of the present study was to determine the prevalence of the most common allelic variants of the polymorphic cytochrome P450 (CYP) enzymes CYP2D6, CYP2C9, CYP2C19 and CYP3A5 and to predict the genotype frequency for each polymorphism in the Greek population. DNA isolated from peripheral blood samples derived from 283 non-related Greek ethnic subjects was used to determine the frequency of CYP2D6*3, CYP2D6*4, CYP2C9*2, CYP2C9*3 and CYP3A5*3 allelic variants by the polymerase chain reaction (PCR)-restriction fragment length polymorphism method, CYP2C19*2 and CYP2C19*3 with allelic specific amplification (PCR-ASA), and CYP2D6*2 (gene duplications) by long PCR analysis. The allelic frequencies (out of a total of 566 alleles) for CYP2D6*3 and CYP2D6*4, were 2.3% and 17.8%, respectively, while gene duplications (CYP2D6*2) were found in 7.4% of the subjects tested. For CYP2C9*2 and CYP2C9*3 polymorphisms the allelic frequencies were 12.9% and 8.13% respectively. For CYP2C19, the *2 polymorphism was present at an allelic frequency of 13.1%, while no subjects were found carrying the CYP2C19*3 allele. Finally, the CYP3A5*3 allele was abundantly present in the Greek population with an allelic frequency of 94.4%. Overall our results show that the frequencies of the common defective allelic variants of CYP2C9, CYP2C19 and CYP3A5 in Greek subjects are similar to those reported for several other Caucasian populations. Finally, a high prevalence of CYP2D6 gene duplication among Greeks was found, a finding that strengthens the idea that a South/North gradient exists in the occurrence of CYP2D6 ultrarapid metabolizers in European populations. [source] Therapy adapted to molecular response in patients with chronic myelogenous leukaemia in first chronic phase: results of the Duesseldorf study,HEMATOLOGICAL ONCOLOGY, Issue 4 2008Frank Neumann Abstract This study evaluates response-adapted treatment of chronic myelogenous leukaemia (CML) in chronic phase using molecular response criteria. bcr-abl/G6PDH ratios were assessed by Light-Cycler quantitative real-time polymerase chain reaction (PCR( in 277 peripheral blood samples from 33 patients, before and every 3 months during therapy. Sixty-six per cent (22/33) of the patients fulfiled our molecular response criterion of ,1 log decrease in bcr-abl transcript after 6 or ,2 log decrease after 9 and every following 3 months. Dose escalation was necessary for 33% (11/33) of the patients. Of these, 54% (6/11) achieved a reduction of bcr-abl mRNA by ,2 log (n,=,3) or ,3 log (n,=,3) with 800,mg Imatinib. Forty-five per cent (5/11) showed insufficient molecular response with 800,mg Imatinib and received Nilotinib. In conclusion, the assessment of molecular response permits an individual patient-tailored treatment of CML in first chronic phase, resulting in the majority of patients achieving a major molecular response after 2 years of therapy. Copyright © 2008 John Wiley & Sons, Ltd. [source] An optimized method to separate reticulocytes from peripheral blood for molecular analysisINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 3 2009R. PETRUZZELLI Summary A method based on immunomagnetic sorting of reticulocytes from peripheral blood was set up and combined to a commercial extraction kit for the isolation of total RNA from whole blood. This procedure resulted in high-quality RNA samples suitable for molecular analysis. We used this procedure to analyse erythroid-specific transcripts, starting from peripheral blood samples, to search for differently expressed mRNAs in patients with hereditary persistence of foetal haemoglobin. After erythrocyte lysis, CD15+and CD45+ peripheral cells were negatively sorted to remove leucocyte populations that could have affected the subsequent screening procedure. The cell sorting and RNA extraction procedure was completed within 1,2 h of erythrocyte lysis, which represents a consistent saving of time compared with other procedures. Moreover, it produced 1 ,g of total RNA per ml of blood samples, which is sufficient for molecular analysis. Therefore, our method is a reliable and efficient tool to isolate RNA from specific cell subpopulations poorly represented in peripheral blood, particularly when accurate detection and characterization of highly unstable and poorly expressed molecules is required. [source] Utility of reticulocyte maturation parameters in the differential diagnosis of macrocytic anemiasINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 5 2003A. Torres Gomez Summary The aim of this study was to test the clinical utility of reticulocyte maturation parameters in the differential diagnosis of macrocytic anemias. Using an automated reticulocyte counter, we analyzed immature reticulocyte fraction (IRF), mean reticulocyte volume (MRV) and mean fluorescence index (MFI) in peripheral blood samples from healthy donors (n = 30), patients diagnosed with myelodysplastic syndromes (MDS, n = 35), with megaloblastic anemia (MA, n = 10) and with non-megaloblastic macrocytic anemias (NMMA, n = 30). Macrocytic anemias due to ineffective erythropoiesis (MA and MDS) showed reticulocytes skewed to a more immature fraction. Therefore, they have a larger volume and a greater RNA content than healthy controls. Interestingly, reticulocytes in both low and high risk MDS are significantly larger (127.3 vs. 118.3 fl, P < 0.01) and have a greater RNA content (MFI 20.5 vs. 12.9, P < 0.01 and IRF 22.5 vs. 9.1%, P < 0.01) than NMMA patients. We conclude that measurement of reticulocyte maturation parameters may be a very useful tool in the differential diagnosis of macrocytic anemia. The presence of extremely high values of IRF (>16%), MFI (>18) and MRV (>129 fl), makes the diagnosis of NMMA very unlikely. An underlying MDS should, therefore, be sought. [source] The presence of local and circulating autoreactive B cells in patients with advanced periodontitisJOURNAL OF CLINICAL PERIODONTOLOGY, Issue 4 2002Tord Berglundh Abstract Aim: The aim of the present investigation was to study the local (gingival) and systemic occurrence of autoreactive B cells (CD5+CD19 positive) in subjects with a high or low susceptibility to periodontitis. Material and Methods: 2 groups of subjects (Group A and B) susceptible to periodontitis were included. Group A consisted of 22 adult patients (7 females and 15 males, aged 24,66 years) with advanced and generalized chronic periodontitis and group B comprised 7 children (4 girls and 3 boys aged 9,13 years) with localized aggressive periodontitis. 26 periodontally healthy subjects, Group C (aged 23,80 years, mean 49.6±16.3), were also recruited. Assessment of clinical and radiographical characteristics of periodontal disease was performed. Gingival biopsies and peripheral blood samples were obtained and prepared for immunohistochemical analysis. Blood samples only were obtained from the periodontally healthy subjects (group C). Results: The proportion of autoreactive B cells (CD5+CD19 positive) of peripheral blood lymphocytes was about 6 times higher in group A and 4 times higher in group B than in the samples from the control subjects (group C). About 40,50% of the B cells in the peripheral blood of the periodontitis susceptible individuals expressed markers for autoreactive features while less than 15% of the circulating B cells in the subjects of group C exhibited such markers. The periodontitis lesion in the adult periodontitis patients contained a substantial number of B cells out of which about 30% demonstrated autoreactive features. Conclusion: It is suggested that both circulating and local B cells in periodontitis susceptible individuals have a higher propensity to autoreactive properties than B cells of patients with a low susceptibility to periodontitis. Zusammenfassung Zielsetzungen: Untersuchung des lokalen (in der Gingiva) und systemischen Vorkommens autoreaktiver B-Zellen (CD5 und CD19 positiv) bei Individuen mit hoher und niedriger Anfälligkeit für Parodontitis. Material und Methoden: 2 Gruppen von Personen, die anfällig für Parodontitis waren, nahmen an der Studie teil: Gruppe A: 22 erwachsenen Patienten (im Alter von 24,66 Jahren; 7 weiblich) mit fortgeschrittener generalisierter chronischer Parodontitis; Gruppe B: 7 Kinder (9,13 Jahre; 4 Mädchen) mit lokalisierter aggressiver Parodontitis. Zusätzlich wurden 26 parodontal gesunde Personen (23,80 Jahre) untersucht. Klinische und röntgenologische parodontale Parameter wurden erhoben. In den Gruppen A und B, wurden Gingivabiopsien und periphere Blutproben, in Gruppe C nur Blutproben entnommen. Ergebnisse: Der Anteil autoreaktiver B-Zellen an den Lymphozyten im peripheren Blut war etwa 6 mal höher in gruppe A und 4 mal höher in Gruppe B als in Proben der Kontrollgruppe (Gruppe C). Etwa 40,50% der B-Zellen im peripheren Blut der für Parodontitis anfälligen Patienten exprimierten Marker für autoreaktive Eigenschaften während weniger als 15% der zirkulierenden B-Zellen der Individuen aus Gruppe C solche Marker aufwiesen. Die parodontalen Läsionen der erwachsenen Parodontitispatienten enthielten eine hohe Zahl von B-Zellen, von denen etwa 30% autoreaktive Eigenschaften aufwiesen. Schlussfolgerungen: Sowhol lokale als auch zirkulierende B-Zellen von für Parodontitis anfälligen Patienten zeigen mit größerer Häufigkeit autoreaktive Eigenschaften als die B-Zellen von Patienten mit geringer Parodontitisanfälligkeit. Résumé But: Le but de cette recherche était d'étudier la présence locale (gingivale) et systémique de cellules B auto réactives (CD5+CD19 positives) chez des sujets présentant une forte ou une faible susceptibilitéà la parodontite. Matériaux et méthodes: 2 groupes de sujet (A et B) susceptible à la parodontite furent inclus. Le groupe A était constitué de 22 patients adultes (7 femmes et 15 hommes âgés de 24 a 66 ans) présentant une parodontite chronique avancée et généralisée et le groupe B était constitué de 7 enfants (4 filles et 3 garçons ages de 9 à 13 ans) présentant une pardontite agressive localisée. 26 sujets sains d'un point de vue parodontal (groupe C, âgés de 23 à 80 ans, age moyen 49.6±16.3) furent également recrutés. L'observation des caractéristiques cliniques et radiographiques de la maladie parodontale fut réalisée. Des biopsies gingivales et des échantillons sanguins furent prélevées et préparées pour des analyses immunohistochemiques. Seuls des prélèvements sanguins furent pris sur le groupe des patients sains. Résultats: La proportion de cellules B auto réactives (CD5+CD19 positives) des lymphocytes du sang périphérique était 6× plus élevée dans le groupe A et 4× plus élevée dans le groupe B que chez les sujets contrôles du groupe C. Environ 40 a 50% des cellules B du sang périphérique des individus susceptibles à la parodontite exprimaient des marqueurs pour des caractéristiques auto réactives alors que moins de 15% des cellules B circulantes des sujets du groupe C présentaient de tels marqueurs. La lésion parodontale de patients atteints de parodontite de l'adulte contenait un nombre substantiel de cellule B parmi lesquels environ 30% présentaient des caractéristiques auto réactives. Conclusions: Cela suggère que les cellules B locales et circulantes des individus susceptibles à la maladie parodontale aient une puls grande propension aux propriétés auto réactives que les cellules B des patients ayant une susceptibilité faible à la parodontite. [source] Human papillomavirus DNA detected in peripheral blood samples from healthy Australian male blood donorsJOURNAL OF MEDICAL VIROLOGY, Issue 10 2009Alice Che-Ha Chen Abstract Recent studies have shown that human papillomavirus (HPV) DNA can be found in circulating blood, including peripheral blood mononuclear cells (PBMCs), sera, plasma, and arterial cord blood. In light of these findings, DNA extracted from PBMCs from healthy blood donors were examined in order to determine how common HPV DNA is in blood of healthy individuals. Blood samples were collected from 180 healthy male blood donors (18,76 years old) through the Australian Red Cross Blood Services. Genomic DNA was extracted and specimens were tested for HPV DNA by PCR using a broad range primer pair. Positive samples were HPV-type determined by cloning and sequencing. HPV DNA was found in 8.3% (15/180) of the blood donors. A wide variety of different HPV types were isolated from the PBMCs; belonging to the cutaneous beta and gamma papillomavirus genera and mucosal alpha papillomaviruses. High-risk HPV types that are linked to cancer development were detected in 1.7% (3/180) of the PBMCs. Blood was also collected from a healthy HPV-positive 44-year-old male on four different occasions in order to determine which blood cell fractions harbor HPV. PBMCs treated with trypsin were negative for HPV, while non-trypsinized PBMCs were HPV-positive. This suggests that the HPV in blood is attached to the outside of blood cells via a protein-containing moiety. HPV was also isolated in the B cells, dendritic cells, NK cells, and neutrophils. To conclude, HPV present in PBMCs could represent a reservoir of virus and a potential new route of transmission. J. Med. Virol. 81:1792,1796, 2009. © 2009 Wiley-Liss, Inc. [source] Biochemical Analysis of Pericardial Fluid and Whole Blood in Dogs with Pericardial EffusionJOURNAL OF VETERINARY INTERNAL MEDICINE, Issue 6 2005Armelle M. de Laforcade Studies evaluating pericardial fluid analysis in dogs to determine the etiology of pericardial effusions have yielded conflicting results. The purpose of this prospective study was to compare acid-base status, electrolyte concentrations, glucose, and lactate of pericardial fluid to peripheral blood from dogs with pericardial effusion and to compare these variables between dogs with neoplastic and nonneoplastic pericardial effusion. Acid-base status, electrolyte concentrations, glucose, hematocrit, urea nitrogen, and lactate concentrations were evaluated in peripheral blood samples and in pericardial effusion samples of 41 client-owned dogs with pericardial effusion. Common abnormal findings in the peripheral blood of dogs with pericardial effusion included hyperlactatemia (n = 38 [of 41]; 93%), hyponatremia (n = 25/41; 61%), hyperglycemia (n = 13/41; 32%), and hypermagnesemia (n = 13/41; 32%). Bicarbonate, sodium, ionized calcium, glucose, and hematocrit were all significantly lower in the pericardial fluid compared with peripheral blood, whereas lactate, chloride, and PCO2 were significantly higher in the pericardial fluid. When comparing the concentrations of variables in the pericardial fluid of dogs with neoplasia (n = 28) to those without neoplasia (n = 13), pH, bicarbonate, and chloride were significantly lower in dogs with neoplasia, whereas lactate, hematocrit, and urea nitrogen were significantly higher in the pericardial fluid of dogs with neoplasia. The difference between peripheral and pericardial glucose concentrations was significantly larger in dogs with neoplasia than in dogs without neoplasia. Although differences between variables in dogs with neoplastic and nonneoplastic pericardial effusion were documented, clinical relevance is likely limited by the degree of overlap between the 2 groups. [source] Specific IgE to allergens in cord blood is associated with maternal immunity to Toxoplasma gondii and rubella virusALLERGY, Issue 11 2008M. J. Ege Background:, Various studies have found reduced prevalences of atopic sensitization and atopic diseases in children previously exposed to infections or living conditions with a high microbial burden, such as the farming environment. Objective:, We sought to determine the relationships of cord blood immunoglobulin E (IgE) with maternal health conditions before and during pregnancy. Methods:, Pregnant women living in rural areas in five European countries were recruited in the third trimester of pregnancy. Information on maternal health during pregnancy was collected from maternity records and by questionnaires (n = 497). Specific IgE for inhalant and food allergens was assessed in cord blood and peripheral blood samples of the mothers. Results:, Inverse associations of cord blood IgE to seasonal allergens with positive maternal records for Toxoplasma gondii (adjusted odds ratio = 0.37 [0.17,0.81]) and rubella virus (adjusted odds ratio = 0.35 [0.13,0.96]) were found. The previously described effect of prenatal farm exposure on IgE to seasonal allergens was partly confounded by a positive maternal record for T. gondii. The number of maternal siblings, maternal contact to cats during pregnancy or during her first year of life, predicted a positive maternal record for T. gondii. Conclusions:, Maternal immunity to T. gondii and rubella may impact on atopic sensitization in the fetus. A positive T. gondii record explained the previously identified effect of prenatal farm exposure on IgE to seasonal allergens only to a minor extent. [source] Prenatal RHD gene determination and dosage analysis by PCR: clinical evaluationPRENATAL DIAGNOSIS, Issue 4 2001F.-Y. Chan Abstract Background , Use of the polymerase chain reaction (PCR) for detection of the RHD gene can measure the RHD gene status for unborn babies at risk for hemolytic disease of the newborn (HDN). The occurrence of D gene variants has led to errors in prenatal typing. Previous reports have highlighted the danger of assigning a positive fetus as negative, resulting in intrauterine fetal deaths. Objective , To evaluate the effectiveness of a testing strategy whereby PCR was not only performed to determine the presence/absence of the RHD gene, but also used to assess the D gene copy number (zero, one or two RHD genes) in family studies for at risk pregnancies. Methods , Samples comprising maternal (57) and paternal (42) peripheral blood samples, amniotic fluid (64), and matching cord blood (64) were collected. Rhesus (Rh) serotyping was performed on all blood samples. For RHD genotyping, DNA was extracted from all samples except for 28 cord samples, where only serotyping was performed (total 199 DNA genotyping). RHD gene PCR amplified exon 4 and exon 7 regions of the RHD gene. The dosage of RHD gene was determined by comparing the intensity of the RHD gene to that of the RHCE gene. Results , A total of 197/199 samples showed concordance between exon 4 and exon 7 PCR results. Two discrepant results occurred in one family: the father carried one normal D gene and one D gene variant where PCR was tested to be positive using exon 4 but negative using exon 7. One of a pair of dizygotic twins inherited this abnormal D gene and was mildly affected by HDN. This was correctly identified antenatally and the pregnancy successfully managed. The concordance rate between serotypes and genotypes for 135 blood samples was 100%. Amongst the family groups, 8/14 heterozygous fathers transmitted the D gene and 26/26 homozygous fathers transmitted the D gene to the babies. The concordance rate between RHD genotypes from amniotic fluid and Rh D serotypes from cord blood was also 100%. Conclusion , The present study demonstrates the effectiveness of using PCR in a clinical setting. It verifies the importance of testing more than one region of the gene, and also the need for a testing strategy where both maternal and paternal testing for RHD gene dosages are performed. Copyright © 2001 John Wiley & Sons, Ltd. [source] | |||||||||||||||||||||||||||||||