CGvHD vs. non-cGvHD gene expression profiling in peripheral blood
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ABSTRACT: We performed peripheral blood gene expression profiling to identify an array of transcriptional biomarkers that discriminate presence of active cGvHD requiring immunosuppression following allogeneic HCT. Peripheral blood mononuclear cells were prepared by Ficoll gradient and cryopreserved from 63 patients (median age 50 yrs; range 19-64) following allogeneic HCT (median 475 days post-HCT) on an IRB approved protocol. Samples were randomly selected into a training set (23 cGvHD and 19 without cGvHD) and test set (12 cGvHD and 9 without cGvHD) and processed on the Aligent whole human genome 44k microarray platform. Bioinformatics programs including AILUN, GeneSpring, SAM, and PAM were used to select a minimum gene-set (FDR <5%) to predict cGvHD in the training set. Patient characteristics in the two groups were collected for confounder/interaction analysis across the following parameters: age, gender, disease histology, disease status at HCT, graft source, conditioning regimen, white blood count (WBC) at sample, and follow-up status including relapse and survival. Type, grade, and activity of acute cGvHD and cGvHD at time of sample and at most recent follow-up were determined by two independent investigators by standard clinical criteria. Three-condition experiment, 35 cGvHD vs. 28 non-cGvHD samples vs. 7 controls (cGvHDnoIS; no immunosuppression drug).
Project description:Background Immune tolerance and persistent mixed chimerism can be achieved reproducibly after combined organ and hematopoietic cell transplantation in mice conditioned with total lymphoid irradiation plus anti-thymocyte globulin. We studied the safety and reproducibility of this approach in a cohort of kidney transplant patients, and tried to identify immune monitoring procedures that can predict tolerance and guide complete immunosuppressive drug withdrawal. Methods Ten patients conditioned with 10 doses of total lymphoid irradiation and 5 doses of anti-thymocyte globulin were given kidney transplants and an injection of CD34+ hematopoietic progenitor cells and T cells from HLA matched donors. Blood cell monitoring included changes in chimerism, balance of T cell subsets, gene expression, and responses to donor alloantigens. Results Nine of 10 patients developed multi-lineage chimerism without graft versus host disease (GVHD), and all had excellent graft function at the last observation point. Five of these with chimerism persisting for at least 12 months were completely withdrawn from immunosuppressive drugs for 6 to 35 months. Blood cells from patients off drugs showed development of specific unresponsiveness to donor alloantigens, M-bM-^@M-^\toleranceM-bM-^@M-^] profiles on gene microarrays, early high ratios of regulatory versus conventional naM-CM-/ve T cells, and early high levels of chimerism among NK cells. Conclusions Total lymphoid irradiation, and anti-thymocyte globulin promoted the development of persistent chimerism and tolerance in a cohort of patients given kidney transplants and donor cell injections. Assays were identified that can assist in the safe withdrawal of immunosuppressive drugs. 45 Agilent Microarray samples were conducted, including 16 tolerance patients, 10 chronic rejection patients, 5 healthy normal controls, and 7 paired pre and post-transplant induced tolerance patients. The aim is to see whether induced tolerance patients show operational tolerance gene expression signature and can withdraw or minimize the immunosuppression regimens.
Project description:Genomic Analysis of more than 400 patients from multi-center transplant programs and clinical trials provides a non-invasive QPCR based gene expression test for operational renal allograft tolerance 3 group comparison of blood from TOL, CAN and Non-transplant healthy controls (HD) allografts. Biological replicates: 16 TOL, 10 CAN and 5 HC
Project description:To obtain genetic evidence about the interaction between MSCs and metastatic NB cancer cells in bone marrow (BM), we isolated bone marrow-derived MSCs from children with NB and compared their global expression patterns with MSCs obtained from normal pediatric donors using the Agilent 44K microarrays. A total of eight different donors (four NB patients and four healthy donors) were analyzed and three independent microarrays hybridizations were performed for each donor.
Project description:The ability to identify a robust means for both pediatric and adult liver transplant recipients for minimization and weaning of immunosuppression is greatly needed. We analyzed 298 samples from pediatric (n=83) and published adult (n=57) liver transplant recipients, and the public available cell or tissue specific samples (n=158). Biomarkers were first identified from the integration of the cross-platform analysis of Stanford pediatric and adult studies and validated in independent samples from UCLA on microarray. Furthermore, Q-PCR was performed for targets validation and delivers a robust diagnostic assay of predicting potential liver tolerance after liver transplantation. Thirteen unique genes were identified (FDR<5%) using nearest shrunken centroid classification methods as a minimum gene set to cross-validate and predict Stanford pediatric tolerance samples with 1misclassification, 5 misclassification for adult samples, and 100% sensitivity and 83% specificity for an independent UCLA pediatric samples in microarray platform. These subset of tolerant specific genes are highly expressed CD56+ natural killer cells (p=0.03). Furthermore, the subset can be narrowed down to 3 key genes with combined ROC=0.988 for the liver tolerance prediction from Q-PCR verification and 64% MIS and STA were predicted as tolerance with the >90% prediction probability. Specific peripheral transcriptional programs can be identified in operational tolerance in pediatric recipients of liver allografts, distinct from those previously identified in adult operationally tolerant liver recipients, and may provide a means to non-invasively monitor patients in a serial manner for immunosuppression minimization. These genes are highly expressed in specific peripheral blood lymphocyte subsets, and their coordinated may support the maintenance of operational tolerance in children, following liver transplantation. Two centers data was included, which are 41 samples from Stanford as the training set and 44 samples from UCLA as the test set. 26 samples from Stanford and 15 samples from UCLA were used in microarray Agilent whole human genome microarray
Project description:Gene expression profile of response to auxin at 3 h after treatment in rice root tips: IR64 (loss-of-function of DRO1 type) vs Near-isogenic line homozygous for the Kinandang Patong allele of DRO1 in an IR64 genetic background (Dro1-NIL; gain-of-function of DRO1 type) We used two rice varieties, IR64 and near-isogenic line homozygous for the Kinandang Patong allele of DRO1 in an IR64 genetic background (Dro1-NIL). We performed comprehensive microarray analysis of rice root tips with auxin treatment (3h) and pre-treatment (0h) in IR64 and Dro1-NIL. Seedling root tips of IR64 and Dro1-NIL were treated with 10 M-BM-5M 2,4-D. n = 3 biological repeats (15 seedlings per repeat).
Project description:Nascent embryonic joints, interzones, contain a distinct cohort of progenitor cells responsible for the formation of the majority of articular tissues. However, to date the interzone has largely been studied using in situ analysis for candidate genes in the context of the embryo rather than using an unbiased genome wide expression analysis on isolated interzone cells, leaving significant controversy regarding the exact role of the intermediate and outer interzone layers in joint formation. Therefore, in this study, using laser capture microdissection (three biological replicates), we selectively harvested the intermediate and outer interzones of mouse embryos at gestational age 15.5 days, just prior to cavitation, when the differences between the layers should be most profound. Microarray analysis (Agilent Whole Mouse Genome Oligo Microarrays) was performed and the differential gene expression between the intermediate interzone cells and outer interzone cells was examined by performing a 2-sided paired student t-test and pathway analysis. 197 genes were differentially expressed (M-bM-^IM-% 2-fold) between the intermediate interzone and the outer interzone with a p-value M-bM-^IM-$ 0.01. Of these, 91 genes showed higher expression levels in the intermediate interzone and 106 were expressed higher in the outer interzone. Pathway analysis of differentially expressed genes suggests an important role for inflammatory processes in the interzone layers, especially in the intermediate interzone, and hence in joint and articular cartilage development. The high representation of genes relevant to chondrocyte hypertrophy and endochondral ossification in the outer interzone suggests that it undergoes endochondral ossification. 2 study groups (intermediate and outer interzone) with three biological replicates (1 biological replicate = 1 embryo)
Project description:To identify the direct molecular targets of 6S, gene microarrays were used to profile gene expression in HCT-116 cells treated with 6S (20 uM for 24 h) comparing with HCT-116 cells treated with DMSO (control). Two-class comparisons. HCT-116 cells treated with 6S (20 uM for 24 h) vs. HCT-116 cells treated with DMSO control;Biological replicates: 4 replicates for each group.
Project description:Nasonia vitripennis injects venom into its host organism Sarcophaga crassipalpis together with the eggs in order to make it suitable for the offspring to survive. The venom is known to suppress the hosts immune system, elevate the lipid levels, slow down development, et cetera.This microarray can uncover new transcriptomal effects on the host organism after natural envenomation that have not been discovered by bioassays. Since transcriptomal effects will vary during time, two different time points have been selected, 3 and 25 hours after parasitization. 16 individuals per sample, 4 replicates per group, loopdesign, 4 control individuals per time point, dye swap
Project description:Understanding the crosstalk between malignant B-cells and their milieu could provide clues on the molecular and clinical biology of Chronic Lymphocytic Leukemia (CLL). Aiming to generate novel therapeutic strategies, different groups have studied different CLL proliferative fractions. We previously, identified one of these proliferative subsets in the peripheral blood from progressive unmutated CLL patients. Since the presence of this small subset appears to be a hallmark of a proliferative disease in which B lymphocytes are being constitutively activated in the proliferative centers of these patients, we performed gene expression analysis comparing the global mRNA and microRNAs expression of this leukemic subpopulation. Our results suggest that the proliferative behaviour of this fraction appears to depend on microRNA-22 over-expression, which induces PTEN downregulation and PI3K/AKT pathway activation. Transfection experiments demonstrated that miR-22 overexpression in CLL B-cells switches on PI3K/AKT leading to FOXO1 inactivation, p27-Kip1 downregulation, and overexpression of Survivin and Ki-67 proteins. We also demonstrated that this pathway could be triggered by microenvironment signals like CD40L/IL4 and more importantly, that this regulatory loop is also present in lymph nodes from progressive UM patients. Altogether, these results underline the key role of PI3K/AKT pathway in the generation of the CLL proliferative pool and provide rationale for the usage of PI3K inhibitors. Two-subset of B cells, QF vs PF. 8 samples of 4 patients.
Project description:To study the difference of gene expression pattern of whole blood between klinefelterM-bM-^@M-^Ys syndrome and normal individuals and discovered the disease-related genes and biological pathway. 717 differentially expressed genes (480 up regulated, 237 down regulated) were identified between whole blood samples of the klinefelterM-bM-^@M-^Ys patients and normal volunteers. Function enrichment analysis identified a number of genes involved in immune response, metabolism etc. Three genes with significant down-regulation in KlinefelterM-bM-^@M-^Ys patients were also confirmed by qRT-PCR.Using the whole blood as material in patients with klinefelterM-bM-^@M-^Ys syndrome, the differentially expressed genes was identified. It is noticed that genes involved in metabolism pathways were significantly decreased. These observations may suggest aberrant of metabolism may be important for developing the klinefelterM-bM-^@M-^Ys syndrome. In addition, our results also show that no X-linked genes are overexpression in the peripheral blood cells of KS patients. In the present study, we examined gene expression in whole blood of 5 klinefelterM-bM-^@M-^Ys patients and 5 healthy control volunteers. The RNA of 10 samples was isolated and Gene expression was detected by gene microarray. The different expressed genes were analyzed by SAM software.