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Blood Leukocytes (blood + leukocyte)
Kinds of Blood Leukocytes Selected AbstractsORIGINAL ARTICLE: Preterm Labor: CD55 in Maternal Blood LeukocytesAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 5 2009Stella Nowicki Problem, Intrauterine inflammation is a frequent and significant factor associated with the pathogenesis of preterm labor/birth (PTL/PTB). However, it remains unclear whether the intrauterine inflammatory responses activate the maternal peripheral circulation. We explored the association between PTL/PTB and the ,activation' of the peripheral circulatory system by determining whether CD55 mRNA expression within peripheral WBCs differed between PTL and control patients not in labor. Method of Study, RNA was purified from white blood cells collected from pregnant women with preterm labor (n = 45), and from pregnant (n = 30) control women. CD55 gene expression was evaluated by quantitative PCR. Results, The mean CD55 mRNA level within the PTL group (0.77 ± 0.03) was 1.48-fold higher than that observed (0.52 ± 0.02) within the control group (P < 0.0001); 71% of PTL patients and only 6.7% of control subjects expressed elevated CD55 mRNA. The receiver operating characteristics (with 95% CI) of CD55 as a marker for PTL were as follows: Sensitivity, 69% (53,82%); Specificity, 93% (78,99%); Positive Predictive Value, 94% (80,99%); and Negative Predictive Value, 67% (51,80%). In the patient population that delivered prematurely (before 37 weeks), 81% expressed elevated CD55 mRNA levels with a mean of 0.78 ± 0.03 and 95% CI of 0.71,0.84. The receiver operating characteristics were as follows: Sensitivity, 73% (54,88%); Specificity, 86% (71,95%); Positive Predictive Value, 81.5% (62,94%); and Negative Predictive Value, 80% (64,91%). Conclusion, Here we report for the first time that CD55 mRNA expression was elevated in the peripheral WBCs of subjects with preterm labor compared with control gestationally-matched pregnant woman and that elevated leukocyte CD55 may be a useful predictor of subsequent PTB. [source] Deletions removing the last exon of TACSTD1 constitute a distinct class of mutations predisposing to Lynch syndrome,HUMAN MUTATION, Issue 2 2009Marietta E. Kovacs Abstract Several different genetic alterations in the etiology of Lynch syndrome (hereditary nonpolyposis colorectal cancer [HNPCC]) are known, mostly point mutations and genomic rearrangements in 1 of at least 3 mismatch-repair (MMR) genes. However, no susceptibility factor has yet been identified in a significant part (30,50%) of clinicopathologically well-defined HNPCC families, suggesting the presence of other predisposing mechanisms. In a set of probands from 27 Lynch syndrome families who lacked evidence of a germline mutation in either the MSH2 or MLH1 gene, we performed genomic deletion screening with the use of multiplex ligation-dependent probe amplification (MLPA) and sequencing. We used immunohistochemistry (IHC) and microsatellite instability (MSI) analyses on samples of the probands of all families. Comparative analysis of mRNA transcripts was performed on blood leukocyte,derived samples from mutation carriers and noncarrier controls. We report that large germline deletions encompassing the last exons of the TACSTD1 gene, upstream of MSH2, cosegregate with the HNPCC phenotype in 19% (5/27) of families tested. The tumors of the carriers show high-level MSI and MSH2 protein loss. We show that these deletions, by removing the transcriptional termination sequences of the upstream gene, give rise to multiple TACSTD1/MSH2 fusion transcripts. Our results provide evidence that deletions removing the last exon of TACSTD1 constitute a distinct class of mutations predisposing to Lynch syndrome. Thus, analysis of the 3, region of the TACSTD1 gene should be included in the routine mutation screening protocols for HNPCC. Hum Mutat 30, 197,203, 2009. © 2009 Wiley-Liss, Inc. [source] Fabry disease in a heterozygote presenting as hand ischaemia and painful acroparaesthesiaAUSTRALASIAN JOURNAL OF DERMATOLOGY, Issue 1 2007Linda Martin SUMMARY A 48-year-old woman presented with acute unilateral ischaemia of the left hand. She had a background of chronic peripheral neuropathic pain, palpitations, anaemia and an episode of superficial thrombophlebitis. Physical examination revealed non-blanching purple discoloration of her left fingers and her left thumb, index finger and thenar eminance appeared ischaemic. Digital subtraction angiography of the left hand demonstrated reduced flow. Skin punch biopsy histology was unremarkable. The diagnosis of Fabry disease was made on urine lipid profile analysis and confirmed by reduced peripheral blood leukocyte ,-galactosidase A activity. [source] Near-infrared dyes for six-color immunophenotyping by laser scanning cytometryCYTOMETRY, Issue 3 2002Andreas O.H. Gerstner Abstract Background To adequately analyze the complexity of the immune system and reduce the required sample volume for immunophenotyping in general, more measurable colors for the discrimination of leukocyte subsets are necessary. Immunophenotyping by the laser scanning cytometer (LSC), a slide-based cytometric technology, combines cell detection based on multiple colors with their subsequent visualization without the need for physical cell sorting. In the present study, the filter setting of the LSC was adapted for the measurement of the far-red emitting dye cyanine 7 (Cy7), thereby increasing the number of measurable commercially available fluorochromes. Methods The optical filters of the LSC were replaced,photomultiplier (PMT) 3/allophycocyanin (APC): 740-nm dichroic long pass, and 670-/55-nm bandpass; PMT 4/Cy7: 810-/90-nm bandpass. Peripheral blood leukocytes were stained directly by fluorochrome-labeled antibodies or by indirect staining. The tandem dyes of Cy7 (phycoerythrin [PE]-Cy7, APC-Cy7) and the fluorochromes fluorescein isothiocyanate (FITC), PE, PE-Cy5, and APC were tested alone and in different combinations. Results With the new filter combination and tandem fluorochromes, Cy7 was measurable at 488-nm (argon laser) or 633-nm (helium-neon laser) excitation. Resolution was in the range of FITC for PE-Cy7 but approximately 30% lower for APC-Cy7; spillover into the respective donor fluorochrome channel for both tandem dyes was prominent. A six-color panel for leukocyte subtyping was designed. Conclusions With this adaptation, it is possible to measure the tandem conjugates PE-Cy7 and APC-Cy7. This new setup opens the way for six-color immunophenotyping by LSC. Cytometry 48:115,123, 2002. © 2002 Wiley-Liss, Inc. [source] No increases in biomarkers of genetic damage or pathological changes in heart and brain tissues in male rats administered methylphenidate hydrochloride (Ritalin) for 28 days,,ENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 1 2010Kristine L. Witt Abstract Following a 2005 report of chromosomal damage in children with attention deficit/hyperactivity disorder (ADHD) who were treated with the commonly prescribed medication methylphenidate (MPH), numerous studies have been conducted to clarify the risk for MPH-induced genetic damage. Although most of these studies reported no changes in genetic damage endpoints associated with exposure to MPH, one recent study (Andreazza et al. [2007]: Prog Neuropsychopharmacol Biol Psychiatry 31:1282,1288) reported an increase in DNA damage detected by the Comet assay in blood and brain cells of Wistar rats treated by intraperitoneal injection with 1, 2, or 10 mg/kg MPH; no increases in micronucleated lymphocyte frequencies were observed in these rats. To clarify these findings, we treated adult male Wistar Han rats with 0, 2, 10, or 25 mg/kg MPH by gavage once daily for 28 consecutive days and measured micronucleated reticulocyte (MN-RET) frequencies in blood, and DNA damage in blood, brain, and liver cells 4 hr after final dosing. Flow cytometric evaluation of blood revealed no significant increases in MN-RET. Comet assay evaluations of blood leukocytes and cells of the liver, as well as of the striatum, hippocampus, and frontal cortex of the brain showed no increases in DNA damage in MPH-treated rats in any of the three treatment groups. Thus, the previously reported observations of DNA damage in blood and brain tissue of rats exposed to MPH for 28 days were not confirmed in this study. Additionally, no histopathological changes in brain or heart, or elevated serum biomarkers of cardiac injury were observed in these MPH-exposed rats. Environ. Mol. Mutagen. 2010. Published 2009 Wiley-Liss, Inc. [source] DNA damage in peripheral blood leukocytes of physically active individuals as measured by the alkaline single cell gel electrophoresis assayENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 4 2009Gursatej Gandhi Abstract DNA damage induced by physical activity and/or exercise has been reported under different conditions but not for individuals maintaining physical fitness by regular strenuous exercise. Therefore, we compared levels of DNA damage in blood leukocytes of 40 healthy individuals (35 males, 5 females) who regularly exercised in gymnasiums/health clubs and 15 healthy sedentary controls who had never exercised. The former group was selected (after informed consent) on the basis of how long they had been exercising on a regular basis as well as their exercise schedule and regimen. The length of time since starting a regular exercise regimen ranged from 2 months to 9 years, whereas the daily exercise duration ranged from 40 min to 3 hrs and warm-up sessions ranged from none to 90 min. The length of DNA migration (44.66 ± 2.68 ,m in males, 29.62 ± 1.69 ,m in females) and the percentage of cells with tails (79.86 ±1.27% in males, 67.20 ± 0.96% in females) in peripheral blood leukocytes of physically active individuals were increased significantly (P < 0.001) with respect to corresponding values in control males and females (18.85 ± 1.79 ,m, 23.37 ± 3.94 ,m; 24.50 ± 1.98%, 33.00 ± 4.44%, respectively). Highly significant differences for DNA damage were also observed between physically active males and females. These observations, in the absence of any other exposures, indicate a correlation between strenuous exercise to keep fit and increased levels of DNA damage. This finding may have relevance in terms of the ageing process, with diseases associated with aging, and with carcinogenesis. Environ. Mal. Mutagen. 2009. © 2009 Wiley-Liss, Inc. [source] DNA damage in Pakistani pesticide-manufacturing workers assayed using the Comet assayENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 8 2006Javed A. Bhalli Abstract The production and use of chemical pesticides has increased in recent years. Although the increased use of pesticides may benefit agriculture, they are also the potential source of environmental pollution, and exposure to pesticides can have negative consequences for human health. In the present study, we have assessed DNA damage in blood leukocytes from 29 Pakistani pesticide-factory workers and 35 controls of similar age and smoking history. The workers were exposed to various mixtures of organophosphates, carbamates, and pyrethroids. DNA damage was measured with the single cell gel electrophoresis (SCGE) assay or Comet assay, using the mean comet tail length (,m) as the DNA damage metric. Exposed workers had significantly longer comet tail lengths than the controls (mean ± SD 19.98 ± 2.87 vs. 7.38 ± 1.48, P < 0.001). Of the possible confounding factors, smokers had significantly longer mean comet tail lengths than nonsmokers and exsmokers for both the workers (21.48 ± 2.58 vs.18.37 ± 2.28, P < 0.001) and the controls (8.86 ± 0.56 vs. 6.79 ± 1.31, P < 0.001), while age had a minimal effect on DNA damage (P > 0.05 and P < 0.05 for workers and controls, respectively). The results of this study indicate that occupational exposure to pesticides causes DNA damage. Environ. Mol. Mutagen., 2006. © 2006 Wiley-Liss, Inc. [source] Ulcerative colitis: Correlation of the Rachmilewitz endoscopic activity index with fecal calprotectin, clinical activity, C-reactive protein, and blood leukocytesINFLAMMATORY BOWEL DISEASES, Issue 12 2009Alain M. Schoepfer MD Abstract Background: The accuracy of noninvasive markers for the detection of endoscopically active ulcerative colitis (UC) according the Rachmilewitz Score is so far unknown. The aim was to evaluate the correlation between endoscopic disease activity and fecal calprotectin, Clinical Activity Index, C-reactive protein (CRP), and blood leukocytes. Methods: UC patients undergoing colonoscopy were prospectively enrolled and scored independently according the endoscopic and clinical part of the Rachmilewitz Index. Patients and controls provided fecal and blood samples for measuring calprotectin, CRP, and leukocytes. Results: Values in UC patients (n = 134) compared to controls (n = 48): calprotectin: 396 ± 351 versus 18.1 ± 5 ,g/g, CRP 16 ± 13 versus 3 ± 2 mg/L, blood leukocytes 9.9 ± 3.5 versus 5.4 ± 1.9 g/L (all P < 0.001). Endoscopic disease activity correlated closest with calprotectin (Spearman's rank correlation coefficient r = 0.834), followed by Clinical Activity Index (r = 0.672), CRP (r = 0.503), and leukocytes (r = 0.461). Calprotectin levels were significantly lower in UC patients with inactive disease (endoscopic score 0,3, calprotectin 42 ± 38 ,g/g), compared to patients with mild (score 4,6, calprotectin 210 ± 121 ,g/g, P < 0.001), moderate (score 7,9, calprotectin 392 ± 246 ,g/g, P = 0.002), and severe disease (score 10,12, calprotectin 730 ± 291 ,g/g, P < 0.001). The overall accuracy for the detection of endoscopically active disease (score ,4) was 89% for calprotectin, 73% for Clinical Activity Index, 62% for elevated CRP, and 60% for leukocytosis. Conclusions: Fecal calprotectin correlated closest with endoscopic disease activity, followed by Clinical Activity Index, CRP, and blood leukocytes. Furthermore, fecal calprotectin was the only marker that reliably discriminated inactive from mild, moderate, and highly active disease, which emphasizes its usefulness for activity monitoring. Inflamm Bowel Dis 2009 [source] Activation of an IL-6:STAT3-dependent transcriptome in pediatric-onset inflammatory bowel diseaseINFLAMMATORY BOWEL DISEASES, Issue 4 2008Rebecca Carey MD Abstract Background: While activation of the IL-6-dependent transcription factor signal transducer and activator of transcription 3 (STAT3) has been implicated in the pathogenesis of inflammatory bowel disease (IBD), a direct effect on mucosal gene expression and inflammation has not been shown. We hypothesized that a proinflammatory IL-6:STAT3-dependent biological network would be up regulated in pediatric-onset IBD patients, and would be associated with the severity of mucosal inflammation. Methods: Patients with pediatric-onset IBD were enrolled at diagnosis and during therapy. Serum cytokine analysis was performed using Bioplex. STAT3 phosphorylation (pSTAT3) in peripheral blood leukocytes (PBLs) was assessed by flow cytometry. Immunohistochemistry of colonic mucosa was used to localize pSTAT3 and STAT3 target genes. Microarray analysis was used to determine RNA expression profiles from colon biopsies. Results: Circulating IL-6 was upregulated in active IBD patients at diagnosis and during therapy. STAT3 activation was increased in PB granulocytes, IL-6-stimulated CD3+/CD4+ lymphocytes, and affected colon biopsies of IBD patients. The frequency of pSTAT3+ PB granulocytes and colon epithelial and lamina propria cells was highly correlated with the degree of mucosal inflammation. Microarray and Ingenuity Systems bioinformatics analysis identified IL-6:STAT3-dependent biological networks upregulated in IBD patients which control leukocyte recruitment, HLA expression, angiogenesis, and tissue remodeling. Conclusions: A proinflammatory IL6:STAT3 biologic network is upregulated in active pediatric IBD patients at diagnosis and during therapy. Specific targeting of this network may be effective in reducing mucosal inflammation. (Inflamm Bowel Dis 2007) [source] Transmission of neozoic Anguillicoloides crassus and established Camallanus lacustris in ruffe Gymnocephalus cernuusJOURNAL OF FISH BIOLOGY, Issue 9 2008J. Unger In the present study, groups of ruffe Gymnocephalus cernuus, reared singly, were exposed to defined numbers of Anguillicoloides crassus or Camallanus lacustris under controlled laboratory conditions. Infection took place orally through feeding G. cernuus with axenically cultured and laboratory infected copepods, in which the parasites had developed to the infective third stage (L3). Mean prevalence (94·3%) and infection probability (38·5%) for the established C. lacustris were significantly higher than for the neozoic A. crassus (14·3 and 1·0%, respectively). Peripheral blood leukocytes were significantly increased in infected fish, apparently independent of exposure level, parasite species or intensity of infection compared to the controls. In infected fish, the gonado-somatic index (IG) was significantly reduced by c. 50%, and the spleen-somatic index (IS) was significantly increased compared to controls. Both parasites raised similar physiological and immunological responses in G. cernuus, which was able to effectively reject the neozoic A. crassus. [source] Cellular normalization of viral DNA loads on whole blood improves the clinical management of cytomegalovirus or Epstein Barr virus infections in the setting of pre-emptive therapyJOURNAL OF MEDICAL VIROLOGY, Issue 1 2009Céline Bressollette-Bodin Abstract Two quantitative duplex real-time PCR assays were developed for co-amplification of human albumin and cytomegalovirus (CMV) or Epstein Barr virus (EBV) genes after automated extraction on whole blood, and compared two units for expressing viral DNA loads (copies per ml of blood or per 106 peripheral blood leukocytes (PBLs)) on 1,138 positive samples. Both PCRs were characterized by high sensitivity, reproducibility, and linear range. Automated extraction by a MagNA Pure LC Instrument was shown to be more efficient when peripheral blood cell count was inferior to 5,×,109 PBLs/L. Albumin co-amplification allows the detection of PCR inhibitors and normalization of viral load according to the number of cells calculated in the sample. The two ways of expressing viral load results were highly correlated, but quantitative differences varied in relation to variations of blood cell count. As these two viruses are highly cell associated, viral loads can be underestimated in patients with leucopenia. In the setting of pre-emptive strategies during CMV infection, the units in which results are expressed can influence clinical management, as illustrated in this article. J. Med. Virol. 81:90,98, 2009. © 2008 Wiley-Liss, Inc. [source] Identification of a unique BK virus variant in the CNS of a patient with AIDS,JOURNAL OF MEDICAL VIROLOGY, Issue 1 2003Gunn Eli Kimo Jørgensen Abstract Human polyomavirus BK (BKV; GenBank or EMBL or DDBJ accession no. NC001538) is often reactivated in immunosuppressed patients. Reactivation has been associated primarily with excretion of the virus in the urine, and there have been few reports of renal and/or neurological disease caused by BKV in patients with acquired immunodeficiency syndrome (AIDS). Polymerase chain reaction, Southern blotting, and sequencing were used to detect and identify the noncoding control region (NCCR) of BKV in different tissues in an AIDS patient with meningoencephalitis, retinitis, and nephritis. An undescribed reorganized NCCR variant of the virus, completely different from the variants detected in peripheral blood leukocytes (PBLs) and urine, was identified in the cerebrospinal fluid (CSF) and CNS tissues. These results suggest that rearrangements in the NCCR of the virus have resulted in a BKV variant, which is better adapted to the host cell machinery of the cells in CNS tissue. The rearranged variant (BKV CNS) might have been involved in the initiation and/or development of the pathological lesions observed in the CNS-related tissues of this patient. J. Med. Virol. 70: 14,19, 2003. © 2003 Wiley-Liss, Inc. [source] Quantitation of cytomegalovirus (CMV) DNA by real-time PCR for occurrence of CMV disease in HIV-infected patients receiving highly active antiretroviral therapyJOURNAL OF MEDICAL VIROLOGY, Issue 3 2003Karine Gourlain Abstract In HIV-infected patients treated with highly active antiretroviral therapy (HAART) included in the Predivir cohort, we have evaluated the usefulness of CMV DNA quantitation by a TaqMan® PCR assay from peripheral blood leukocytes (PBLs) to predict CMV disease occurrence. In parallel with the immune restoration after treatment by HAART, the percentage of positive samples decreased progressively from 7.3% at Day 0 to 3.5% at Month 12. Among the CMV markers, the smallest concordance with PBL CMV TaqMan® PCR, as evaluated by kappa, was observed with pp65 antigenemia, whereas concordance with all other CMV markers was high. Among the 16 patients with CMV DNA copies at least once >100/150,000 cells, CMV disease occurred in six during follow-up, whereas among the 159 patients with CMV DNA copies always <10/150,000 cells, CMV disease occurred in three and among the seven patients with CMV DNA copies >10 and <100 occurred in only one. In univariate Cox models, all the CMV markers including PBL CMV TaqMan® PCR >10/150,000 cells (RR: 27.6, IC95: 7.1,107.2), the CD4 cell count <75 cells/mm3 and the HIV viral load >100,000 copies/ml were predictive for CMV disease. In a stepwise multivariate analysis, which should be interpreted with caution due to the small number of events (n = 10), three covariates were associated independently with CMV disease: pp65 antigenemia >100 nuclei/200,000, PBL CMV TaqMan® PCR >10 copies/150,000 cells and HIV viral load remaining or increasing >100,000 copies/ml. J. Med. Virol. 69:401,407, 2003. © 2003 Wiley-Liss, Inc. [source] The mediterranean fever gene modifies the progression of disability in non-Ashkenazi Jewish multiple sclerosis patientsJOURNAL OF NEUROCHEMISTRY, Issue 2002Y. Shinar MS is an autoimmune, CNS demyelinating disease manifested in most patients with progressive disability. The progression rate varies between patients and may depend on modifier, immune related genes. The Mediterranean fever gene, expressed in peripheral blood leukocytes, is responsible for familial Mediterranean fever (FMF), a recessive, periodic autoinflammatory disease prevalent in Semitic populations, and less penetrant in Ashkenazi Jews. We related common, FMF associated MEFV mutations to the progression of disability in Jewish, relapsing remitting (RR) MS patients. The mutations 148Q, 694V, 695R and 726A were identified by enzymatic restriction of PCR-amplified MEFV DNA. The progression to statuses 3 and 6 of the expanded disability status scale (EDSS) was analyzed on survival plots. 35% of 48 non-Ashkenazi patients had one MEFV mutation. Compared to non-carriers (n = 31) the heterozygous cohort (n = 17) represented with an increased fraction reaching both EDSS statuses (p < 0.05), and with a shorter median time to reach both EDSS =,3 (2 years in carriers vs. 10 years in non-carriers, p < 0.01) and EDSS =,6 (6 vs. 23 years, respectively, p < 0.005). 17% of 71 Ashkenazi patients had one MEFV mutation. There was no significant difference in the fraction of disabled or in the progression of disability between Ashkenazi carrier patients and non-carriers. The susceptibility of the non-Ashkenazi group attributed, in part, to the detrimental non-Ashkenazi 694V mutation. The results suggest phenotypic expression of one mutated MEFV gene in non-ashkenazi patients, pertinent to the pathogenesis of disability. Acknowledgements:, Granted by the Israeli Ministry of Science (#6279). [source] ACE and MTHFR gene polymorphisms in unexplained recurrent pregnancy lossJOURNAL OF OBSTETRICS AND GYNAECOLOGY RESEARCH (ELECTRONIC), Issue 3 2008Venkatesan Vettriselvi Abstract Aim:, To assess the association between polymorphisms in angiotensin converting enzyme and methylene tetrahydrofolate reductase genes and recurrent pregnancy loss by a case-control study in South Indian women. Methods:, DNA was extracted from peripheral blood leukocytes of 104 women with Recurrent Pregnancy Loss (RPL) and 120 controls. Genotyping of ACE Insertion Deletion and MTHFR C677T polymorphism were carried out by PCR and PCR-RFLP, respectively. Results:, No statistically significant difference was observed in the distribution of genotypes between cases and controls for ACE and MTHFR polymorphisms. Further, the combination of MTHFR and ACE genotypes failed to reveal an association. Conclusion:, In conclusion, the present study reveals lack of association of MTHFR C677T and ACE I/D polymorphisms in RPL in South Indian women. However, we cannot exclude the possibility that other polymorphisms of ACE and MTHFR genes could be associated with the disease and might be clinically useful as a marker to assess risk for RPL. [source] Genetic variants in germline TP53 and MDM2 SNP309 are not associated with early onset colorectal cancer,JOURNAL OF SURGICAL ONCOLOGY, Issue 7 2008Sajid A. Khan MD Abstract Background and Objectives Colorectal cancer (CRC) arising in patients under age 30 is a rare disease, and few cases have been reported within Li-Fraumeni kindreds. To determine how often alterations in the p53 pathway genes contribute to disease susceptibility, we have evaluated patients with early onset CRC for the presence of germline variants in the p53 gene and MDM2 SNP309. Methods Thirty-five patients with CRC diagnosed before age 30 were included in this study-based on tissue availability. DNA samples from peripheral blood leukocytes were analyzed for constitutional mutations and polymorphisms in p53 as well as polymorphisms in MDM2 SNP309. Results No mutations were found in exons 4,10 of the p53 gene. The frequencies of polymorphisms in p53 and in MDM2 SNP309 did not differ from rates previously reported for normal control populations, and no polymorphism in either gene could be associated with early onset CRC. Conclusions Neither germline variants in p53 nor MDM2 SNP309 play an underlying role in the development of very early onset CRC. For the large majority of cases, the genetic basis of this disease remains unknown. J. Surg. Oncol. 2008;97:621,625. © 2008 Wiley-Liss, Inc. [source] Factor IX polypyrimidine tract mutation analysis using mRNA from peripheral blood leukocytesJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 11 2004N. S. Van De Water [source] Gene expression profiling of acute cellular rejection in rat liver transplantation using DNA microarraysLIVER TRANSPLANTATION, Issue 5 2009Naoki Hama Acute cellular rejection (ACR) is still a major problem in organ transplantation, and its genetic and molecular mechanisms remain poorly understood. We used DNA microarrays to investigate the gene expression profiles in ACR. We hypothesized that changes of gene expression in grafts could also be detected in peripheral blood leukocytes. We first compared the gene expression profiles in liver isografts (Lewis to Lewis) and allografts (Dark Agouti to Lewis) harvested from rats at days 1, 3, 5, and 7 after transplantation. Hierarchical clustering analysis indicated that gene expression started to change on day 3, and 89 differentially expressed genes were extracted from allografts in comparison with isografts at day 3. Most of the up-regulated genes were associated with graft-infiltrating leukocytes. We then confirmed the similarity of gene expression changes in peripheral leukocytes by quantitative real-time polymerase chain reaction. We also investigated the gene expression changes in other inflammatory and liver dysfunction models. Two interferon-gamma inducible genes, interferon regulatory factor 1 and guanylate nucleotide binding protein 2, were overexpressed in both the peripheral leukocytes and liver graft during ACR. Although further studies are necessary, these 2 genes in peripheral leukocytes could be potentially useful markers for rejection or immunosuppression. Liver Transpl 15:509,521, 2009. © 2009 AASLD. [source] In vivo behaviour of long-circulating liposomes in blood vessels in hamster inflammation and septic shock models,use of intravital fluorescence microscopyLUMINESCENCE: THE JOURNAL OF BIOLOGICAL AND CHEMICAL LUMINESCENCE, Issue 2 2001Jean-Marie Devoisselle Abstract This study aimed to observe liposome uptake by leukocytes in vivo. The study was performed on skin by using a dorsal skin-fold chamber implanted in golden hamsters using intravital microscopy. 5 and 6-CF-encapsulated polyethylene glycolated liposomes were injected intravenously. The skin microcirculation was observed with an intravital Eclipse E800 Nikon microscope (using ×40, ×80 magnification) fitted with a Xenon light source and an epifluorescence assembly (excitation, 470,nm, FWHM 40,nm; emission, 540,nm, FWHM 40,nm). An ultra-high sensitivity videocamera mounted on the microscope projected the image onto a monitor, and the images (720,×,576 pixels) were recorded for playback analysis with a digital video cassette recorder. An acute inflammatory response was obtained by removing one complete layer of skin and the underlying fascia and avascular tissue on the opposing side of the flap corresponding to an area equivalent to the window aperture. Using this model and set-up, leukocyte rolling and adhesion were easily observed and the entry of PEGylated liposomes into hamster blood leukocytes was studied for a period of 6,h. PEGylated liposomes were clearly identified alone inside the blood flow and inside the leukocytes as soon as the inflammatory reaction appeared. This study shows for the first time that blood leukocytes in their natural milieu of whole blood are capable of interacting with, and taking up, liposomes. This observation is in accordance with previous in vitro studies. Copyright © 2001 John Wiley & Sons, Ltd. [source] Cytokine production from sputum cells and blood leukocytes in asthmatics according to disease severityALLERGY, Issue 7 2010M. Manise To cite this article: Manise M, Schleich F, Gusbin N, Godinas L, Henket M, Antoine N, Corhay JL, Louis R. Cytokine production from sputum cells and blood leukocytes in asthmatics according to disease severity. Allergy 2010; 65: 889,896. Abstract Background:, Although mild to moderate asthma is known to be Th2 driven, cytokines produced in refractory asthma might not fit the classical Th2 pattern. Methods:, The aim of our study was to assess the cytokine production by sputum and blood cells from 15 refractory asthmatics (American Thoracic Society Criteria) compared to 15 mild untreated and 17 moderate treated asthmatics and 22 healthy subjects. Spontaneous production of interleukin (IL)-4, IL-6, IL-10, interferon-,, and tumor necrosis factor , was measured by immunotrapping after 24 h sputum or blood cell culture. Results:, Moderate and refractory asthmatics were both characterized by a lower production of IL-6 from their airway cells compared to healthy subjects. However, the difference was no longer significant when expressing the results per gram of sputum. No significant difference between the three groups was found regarding other cytokines. As for cytokine production from blood, the three groups of asthmatics exhibited raised production of IL-4 when compared to healthy subjects, and this was true when results were expressed per blood volume or after normalization for total leukocyte cell count. Moderate asthmatics exhibited greater production of IL-10 when compared to refractory asthmatics and healthy subjects when results were normalized for total leukocyte cell count. Conclusions:, Sputum cells from moderate and refractory asthmatics release less IL-6. While the systemic overproduction of IL-4 was observed through the all spectrum of asthma severity, moderate asthmatics exhibited greater systemic IL-10 production compared to refractory asthmatics. [source] Immunostimulating effects of the polyphenol-rich fraction of sugar cane (Saccharum officinarum L.) extract in chickensPHYTOTHERAPY RESEARCH, Issue 2 2007Kenji Hikosaka Abstract The phagocytic activity of peripheral blood leukocytes (PBL) in chickens orally administered sugar cane extracts (SCE) or polyphenol-rich fraction (PRF) of SCE (500 mg/kg/day) for 3 consecutive days increased significantly, when compared with that of saline-administered control chickens. Chickens orally administered SCE or PRF (500 mg/kg/day) for 3 consecutive days showed significantly higher antibody responses against sheep red blood cells and Brucella abortus than control chickens. In addition, oral administration of SCE or PRF also resulted in a significant increase in the number of IgM- and IgG-plaque forming cell responses of PBL, intestinal leukocytes and splenocytes, when compared with those of control chickens. Furthermore, delayed type hypersensitivity responses to human , globulin significantly increased in chickens orally administered SCE or PRF, compared with those of control chickens when evaluated on the basis of net increased wattle thickness at 24, 48 and 72 h after challenge. These results suggest that PRF of SCE has an immunostimulating effect in chickens. Copyright © 2006 John Wiley & Sons, Ltd. [source] Identification of repertoires of surface antigens on leukemias using an antibody microarrayPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 11 2003Larissa Belov Abstract We have previously described a microarray of cluster of differentiation (CD) antibodies that enables concurrent determination of more than 60 CD antigens on leukocytes. This procedure does not require protein purification or labeling, or a secondary detection system. Whole cells are captured by a microarray of 10 nL antibody dots immobilized on a nitrocellulose film on a microscope slide. Distinct patterns of cell binding are observed for different leukemias or lymphomas. These haematological malignancies arise from precursor cells of T- or B-lymphocytic, or myeloid lineages of hematopoiesis. The dot patterns obtained from patients are distinct from those of peripheral blood leukocytes from normal subjects. This microarray technology has recently undergone a number of refinements. The microarray now contains more CD antibodies, and a scanner for imaging dot patterns and software for data analysis provide an extensive immunophenotype sufficient for diagnosis of common leukemias. The technology is being evaluated for diagnosis of leukemias with parallel use of conventional diagnostic criteria. [source] REVIEW ARTICLE: Effects of Early Conceptus Signals on Circulating Immune Cells: Lessons from Domestic RuminantsAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 4 2010Troy L. Ott Citation Ott TL, Gifford CA. Effects of early conceptus signals on circulating immune cells: lessons from domestic ruminants. Am J Reprod Immunol 2010 While there are few similarities between mechanisms for extending corpus luteum (CL) function during early pregnancy in ruminants and primates, there is increasing evidence that conceptus-immune crosstalk in ruminants and primates affects the function of circulating immune cells at the very earliest stages of pregnancy. Most notable are changes in immune cell phenotypes with increased numbers of cells exhibiting the T regulatory phenotype and suppression of Th1 cytokines that promote tolerance to paternal alloantigens. Until recently, interferon , produced by the ruminant trophectoderm was thought to act exclusively on the uterine endometrium; however, it is now clear that this unique embryonic interferon escapes the uterus and alters gene expression in the CL and in peripheral blood leukocytes (PBL). In fact, a large number of interferon-stimulated genes are now known to be increased during early pregnancy in PBL. What is not known is how this conceptus-immune system cross-talk affects maternal immune status outside the reproductive tract. It is attractive to hypothesize that some of these effects are designed to counter-balance progesterone-induced immunosuppression so as not to place the dam at a greater risk of infection on top of the tremendous stresses already induced by pregnancy. Furthermore, recent evidence suggests that pregnancy induced changes in peripheral immune cells may aid in orchestrating establishment of pregnancy. Existing evidence points toward a greater convergence of systemic immune responses to early pregnancy signaling between ruminants and primates. [source] ORIGINAL ARTICLE: Cell-Surface CD200 May Predict Efficacy of Paternal Mononuclear Leukocyte Immunotherapy in Treatment of Human Recurrent Pregnancy LossAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 1 2009David A. Clark Problem, The allogeneic leukocytes in transfused blood can modulate the recipient's immune system so as to induce TGF-,-producing suppressor cells, and the cell-surface CD200 tolerance-signaling molecule on mononuclear dendritic cells is required for this effect. A subset of couples with unexplained recurrent pregnancy loss appears to benefit from transfusion of allogeneic paternal blood leukocytes (LIT), and considerable effort has been devoted to characterizing those who may benefit. Some data has been accumulated for LIT as sole therapy in patients with classical spontaneous abortions with respect to dose,response, duration of protection, need for boosting, excluding patients with autoimmunity, and inefficacy of paternal mononuclear cells stored at 4°C overnight before use which causes loss of cell-surface CD200. Recent data emphasize an important role of expression of the CD200 tolerance-signaling molecule on cells used to prevent abortions both in mice and humans. Method of study, An observational study of outcome as a function of the number of CD200+ paternal mononuclear cells was performed. Fourteen patients constituted the pilot group. Patients with autoimmunity who had failed inspite of treatment with IVIG + Heparin + Aspirin ± Prednisone were allowed to have paternal mononuclear cells added to their therapy. CD200 on purified paternal blood mononuclear cells was measured by flow cytometry. Results, The number of CD200+ cells administered was significantly greater in women achieving pregnancy (39.2 × 106 versus 20.8 × 106, P < 0.025) and in those who achieved a live birth (50.2 × 106 versus 20.8 × 106, P < 0.005) compared to those who did not achieve pregnancy, and % of paternal cells that were CD200+ was greater (11,12.5% versus 5.6%, P < 0.01). Amongst those achieving pregnancy which failed, the CD200+ cell dose was not significantly different from the non-pregnant group (30.5 × 106 versus 20.8 × 106). Conclusion, The number of CD200+ paternal mononuclear leukocytes may be an important determinant of subsequent reproductive outcome in a subset of patients. A lower % CD200+ cell number may also reflect hitherto unappreciated paternal factors bearing on reproductive success. It is feasible to recruit women to enter observational studies and to obtain useful data as a foundation for further studies. More complete patient characterization in a larger study is needed. [source] Deficiency of CXCR2, but not other chemokine receptors, attenuates autoantibody-mediated arthritis in a murine modelARTHRITIS & RHEUMATISM, Issue 7 2010Jonathan P. Jacobs Objective Chemokines coordinate leukocyte trafficking in homeostasis and during immune responses. Prior studies of their role in arthritis have used animal models with both an initial adaptive immune response and an inflammatory effector phase. We undertook analysis of chemokines and their receptors in the effector phase of arthritis using the K/BxN mouse serum,transfer model. Methods A time-course microarray analysis of serum-transferred arthritis was performed, examining ankle tissue, synovial fluid, and peripheral blood leukocytes. Up-regulation of chemokines was confirmed by quantitative reverse transcriptase,polymerase chain reaction. The functional relevance of chemokine induction was assessed by transferring serum into mice deficient in CCR1,7, CCR9, CXCR2, CXCR3, CXCR5, CX3CR1, CCL2, or CCL3. Further mechanistic analysis of CXCR2 involved treatment of arthritic mice with a CXCR2 antagonist, bone marrow (BM) cell transfers with CXCR2+/, and CXCR2,/, donors and recipients, flow cytometry of synovial cells, and competition experiments measuring enrichment of CXCR2-expressing neutrophils in arthritic joints of mice with mixed CXCR2+/+ and CXCR2,/, BM cells. Results Gene expression profiling revealed up-regulation of the CXCR2 ligands CXCL1, CXCL2, and CXCL5 in the joint in parallel with disease activity. CXCR2,/, mice had attenuated disease relative to CXCR2+/, littermates, as did mice receiving the CXCR2 inhibitor, while deficiency of other chemokine receptors did not affect arthritis severity. CXCR2 was required only on hematopoietic cells and was widely expressed on synovial neutrophils. CXCR2-expressing neutrophils were preferentially recruited to arthritic joints in the presence of CXCR2-deficient neutrophils. Conclusion CXCR2 (but not other chemokine receptors) is critical for the development of autoantibody-mediated arthritis, exhibiting a cell-autonomous role in neutrophil recruitment to inflamed joints. [source] MicroRNA-146a contributes to abnormal activation of the type I interferon pathway in human lupus by targeting the key signaling proteinsARTHRITIS & RHEUMATISM, Issue 4 2009Yuanjia Tang Objective MicroRNA have recently been identified as regulators that modulate target gene expression and are involved in shaping the immune response. This study was undertaken to investigate the contribution of microRNA-146a (miR-146a), which was identified in the pilot expression profiling step, to the pathogenesis of systemic lupus erythematosus (SLE). Methods TaqMan microRNA assays of peripheral blood leukocytes were used for comparison of expression levels of microRNA between SLE patients and controls. Transfection and stimulation of cultured cells were conducted to determine the biologic function of miR-146a. Bioinformatics prediction and validation by reporter gene assay and Western blotting were performed to identify miR-146a targets. Results Profiling of 156 miRNA in SLE patients revealed the differential expression of multiple microRNA, including miR-146a, a negative regulator of innate immunity. Further analysis showed that underexpression of miR-146a negatively correlated with clinical disease activity and with interferon (IFN) scores in patients with SLE. Of note, overexpression of miR-146a reduced, while inhibition of endogenous miR-146a increased, the induction of type I IFNs in peripheral blood mononuclear cells (PBMCs). Furthermore, miR-146a directly repressed the transactivation downstream of type I IFN. At the molecular level, miR-146a could target IFN regulatory factor 5 and STAT-1. More importantly, introduction of miR-146a into the patients' PBMCs alleviated the coordinate activation of the type I IFN pathway. Conclusion The microRNA miR-146a is a negative regulator of the IFN pathway. Underexpression of miR-146a contributes to alterations in the type I IFN pathway in lupus patients by targeting the key signaling proteins. The findings provide potential novel strategies for therapeutic intervention. [source] Evaluation of genotoxic effects in human peripheral blood leukocytes following an acute in vitro exposure to 900 MHz radiofrequency fieldsBIOELECTROMAGNETICS, Issue 4 2005O. Zeni Abstract Human peripheral blood leukocytes from healthy volunteers have been employed to investigate the induction of genotoxic effects following 2 h exposure to 900 MHz radiofrequency radiation. The GSM signal has been studied at specific absorption rates (SAR) of 0.3 and 1 W/kg. The exposures were carried out in a waveguide system under strictly controlled conditions of both dosimetry and temperature. The same temperature conditions (37.0,±,0.1 °C) were realized in a second waveguide, employed to perform sham exposures. The induction of DNA damage was evaluated in leukocytes by applying the alkaline single cell gel electrophoresis (SCGE)/comet assay, while structural chromosome aberrations and sister chromatid exchanges were evaluated in lymphocytes stimulated with phytohemagglutinin. Alterations in kinetics of cell proliferation were determined by calculating the mitotic index. Positive controls were also provided by using methyl methanesulfonate (MMS) for comet assay and mitomycin-C (MMC), for chromosome aberration, or sister chromatid exchange tests. No statistically significant differences were detected in exposed samples in comparison with sham exposed ones for all the parameters investigated. On the contrary, the positive controls gave a statistically significant increase in DNA damage in all cases, as expected. Thus the results obtained in our experimental conditions do not support the hypothesis that 900 MHz radiofrequency field exposure induces DNA damage in human peripheral blood leukocytes in this range of SAR. Bioelectromagnetics 26:258,265, 2005. © 2005 Wiley-Liss, Inc. [source] Genotoxicity of radiofrequency signals.BIOELECTROMAGNETICS, Issue 2 2002Abstract As part of a comprehensive investigation of the potential genotoxicity of radiofrequency (RF) signals emitted by cellular telephones, in vitro studies evaluated the induction of DNA and chromosomal damage in human blood leukocytes and lymphocytes, respectively. The signals were voice modulated 837 MHz produced by an analog signal generator or by a time division multiple access (TDMA) cellular telephone, 837 MHz generated by a code division multiple access (CDMA) cellular telephone (not voice modulated), and voice modulated 1909.8 MHz generated by a global system of mobile communication (GSM)-type personal communication systems (PCS) cellular telephone. DNA damage (strand breaks/alkali labile sites) was assessed in leukocytes using the alkaline (pH>13) single cell gel electrophoresis (SCG) assay. Chromosomal damage was evaluated in lymphocytes mitogenically stimulated to divide postexposure using the cytochalasin B-binucleate cell micronucleus assay. Cells were exposed at 37±1°C, for 3 or 24 h at average specific absorption rates (SARs) of 1.0,10.0 W/kg. Exposure for either 3 or 24 h did not induce a significant increase in DNA damage in leukocytes, nor did exposure for 3 h induce a significant increase in micronucleated cells among lymphocytes. However, exposure to each of the four RF signal technologies for 24 h at an average SAR of 5.0 or 10.0 W/kg resulted in a significant and reproducible increase in the frequency of micronucleated lymphocytes. The magnitude of the response (approximately four fold) was independent of the technology, the presence or absence of voice modulation, and the frequency (837 vs. 1909.8 MHz). This research demonstrates that, under extended exposure conditions, RF signals at an average SAR of at least 5.0 W/kg are capable of inducing chromosomal damage in human lymphocytes. Bioelectromagnetics 23:113,126, 2002. © 2002 Wiley-Liss, Inc. [source] Oxidoreductase macrophage migration inhibitory factor is simultaneously increased in leukocyte subsets of patients with severe sepsisBIOFACTORS, Issue 4 2008Lutz E. Lehmann M.D. Abstract The oxidoreductase Macrophage Migration Inhibitory Factor (MIF) is discussed as a promising target for immunomodulatory therapy in patients with severe sepsis. Moreover, MIF expresses tautomerase as well as thiol-protein oxidore-ductase activities and has a potential role in cellular redox homeostasis, apoptosis inhibition, endotoxin responsiveness as well as regulation of nuclear transcription factors. To further elucidate a potential role of intracellular MIF in severe sepsis, we assessed alterations of intracellular MIF content in peripheral blood leukocytes of patients with severe sepsis in comparison to healthy controls and non-septic patients after major surgery. Intracellular MIF was significantly elevated simultaneously in lymphocytes, B-cells, macrophages and granulocytes of patients with severe sepsis when compared to healthy control individuals (p < 0.05) and increased when compared to non-septic patients after major surgery. In parallel, plasma MIF levels were elevated in severe sepsis (p < 0.05). There was no difference of intracellular MIF in lymphocytes, B-cells, macrophages or granulocytes between surviving and non-surviving patients with severe sepsis (p > 0.05). However, in survivors LPS ex vivo stimulation increased MIF secretion but not in non-survivors of sepsis (p < 0.05). This finding underlines the role of intracellular MIF in inflammatory diseases. It suggests monitoring of intracellular MIF in further clinical and non-clinical research valuable. [source] Cytochrome P450 gene expression levels in peripheral blood mononuclear cells in comparison with the liverCANCER SCIENCE, Issue 6 2004Motonobu Furukawa Cytochromes P450 (CYPs) compose a superfamily of similar proteins involved in detoxification and elimination, as well as activation of a wide variety of compounds. Most CYP family members are localized in the liver. In order to assess whether peripheral blood leukocytes (PBL) are available as a surrogate for the determination of CYP gene expression levels in the liver, we compared CYP gene expression levels in PBL with those in liver tissues from patients with hepatocellular carcinoma (HCC). We measured CYP1A1, 1A2, 1B1, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 2F1, 2J2, 3A4, 3A5, 3A7, 4A11, 4B1 and CYP27 gene expressions in PBL and in the liver by real-time reverse-transcription (RT)-PCR. We could detect expression of CYP1A1, 1A2, P1B1, 2A6, 2B6 and 2E1 genes in PBL and all the genes except for CYP2F1 in the liver. Although gene expression levels within each subfamily were closely correlated within PBL and within the liver, a clear correlation of gene expression levels between PBL and liver tissues was found only for CYP4B1. Although inter-individual variation of the expression level of each CYP gene was wide, the induced level was proportional to the basal expression level. Therefore, monitoring of CYP gene expression levels in PBL, especially those of CYP4B1, could be available as a biomarker for monitoring of exposure to environmental pollutants and assessing the associated risk. Compared with non-tumor tissue, HCC tissues tended to show overexpression of multiple CYP genes, indicating that individualized selection and more effective administration of chemotherapeutic agents could perhaps be based on the pattern of CYP overexpression. [source] | ||||||||||||||||