Transcription profiling of mouse GalT knockout recipient of GalT transducded allo-bone marrow cells after sublethal irradiation
ABSTRACT: Xenotransplantation holds the promise of providing an unlimited supply of donor organs for terminal patients with organ failure. The gal carbohydrate results in rejection of wild type pig grafts, however, chimerism established by expression of the GalT gene prior to transplantation in GalT knockout mice results in tolerance to Gal+ heart grafts. We used microarrays in order to further understand the early events that occur within grafts that demonstrate tolerance. Experiment Overall Design: The GalT BMT recipient is a GalT knockout mouse which recieved GalT gene transduced allo-bone marrow cells transplantation after sublethal irradiation. A heart of wild type C57BL/6 was heterotopically transplanted into the recipient after GalT BMT. Syngeneic Control recipient is a wild type C57BL/6 transplanted a heart of wild C57BL/6.
Project description:To better understand the pathophysiology of galactose-1-phosphate uridyltransferase (GALT) deficiency in humans, we studied the mechanisms by which a GALT-deficient yeast survived on galactose medium. Under normal conditions, GALT-deficient yeast cannot grow in medium that contains 0.2% galactose as the sole carbohydrate, a phenotype of Gal(-). We isolated revertants from a GALT-deficient yeast by direct selection for growth in galactose, a phenotype of Gal(+). Comparison of gene expression profiles among wild-type and revertant strains on galactose medium revealed that the revertant down-regulated genes encoding enzymes including galactokinase, galactose permease, and UDP-galactose-4-epimerase (the GAL regulon). By contrast, the revertant strain up-regulated the gene for UDP-glucose pyrophosphorylase, UGP1. There was reduced accumulation of galactose-1-phosphate in the galactose-grown revertant cells when compared to the GALT-deficient parent cells.
Project description:Ischemia reperfusion (I/R) injury is an unavoidable event occurring during heart transplantation, leading to graft failures and lower long-term survival rate of the recipient. microRNAs are major regulators of genes. IR causes apoptosis/death of cardiomyocytes, resulting from up-regulation of apoptotic genes and down-regulation of anti-apoptotic genes which are regulated by microRNA. We used microarrays to detail the global programme of gene expression underlying cellularisation and identified distinct classes of up-regulated genes during this process. We perserved donor hearts with university of Wisconsin solution for 18h at 4C degree before being implanted into recipients to create ischemia reperfusion injury. The preserved hearts were implanted into a syngeneic recipient mice. 24h after transplantation, heart grafts were harvested for microRNA extraction.
Project description:Comparison of gene expression profiles in GALT-deficient and GALT-reconstituted cells showed that cells lacking GALT activity responded to galactose challenge by activating a set of genes characteristic of endoplasmic reticulum (ER) stress. This response was specific to galactose insult, as cells grown in glucose or hexose-free media did not exhibit ER stress. Keywords: GALT infected cells Overall design: Cells were first grown and then infected in high glucose, then transferred to DMEM, 3 experiments with dye-swap
Project description:To understand the altered gene expression by D-galactose in galT cells, the cells were grown in the presence and absence of D-galactose and tiling array analysis was performed for transcriptome profiling. A galT defect causes the disease Galactosemia in human. This work helps us to study the human metabolic disease Galactosemia. Total RNAs were isolated from Escherichia coli galT strain grown in the absence and presence of D-galactose. The transcriptome profile of galT cells was figured out in the galactose-stressed condition.
Project description:BMCs from BMT recipient mice transplanted with BMCs-expressing EZH2-mutant were analyzed using Affymetrix GeneChip Mouse430_2 BMCs were extracted from BMT recipient mice transplanted with ABCG2-expressing BMCs and analyzed using Affymetrix GeneChip Mouse430_2 Overall design: EZH2-Mt expressing BMCs were sorted into GFP-positive and negative populations before RNA extraction. Whole BMCs were used for RNA extraction from ABCG2-expressing BMCs and compared to BMCs from control BMT mice.
Project description:Comparison of gene expression profiles in GALT-deficient and GALT-reconstituted cells showed that cells lacking GALT activity responded to galactose challenge by activating a set of genes characteristic of endoplasmic reticulum (ER) stress. This response was specific to galactose insult, as cells grown in glucose or hexose-free media did not exhibit ER stress. Experiment Overall Design: Cells were first grown and then infected in high glucose, then transferred to DMEM, 3 experiments with dye-swap
Project description:This study reports the evolution of the donor CD8 effector T cell transcriptomes at multiple sites and time points within the same host following allo-SCT. Donor derived CD8+ T cells were flow sorted from multiple organs in mice developing GVHD in two clinically relevant murine models of H-2b MHC-matched minor antigen-mismatched BMT: (1) B6>129 model – transfer of C57BL/6 polyclonal CD4+ and CD8+ T cells into 129/Sv BMT recipients; (2) F>M – transfer of monoclonal HY-specific TCR-transgenic MataHari CD8+ T cells into C57BL/6 male BMT recipients.
Project description:MicroRNAs are small non-coding RNA molecules that regulate the post-transcriptional expression of target genes. In addition to being involved in many biologic processes including development, cell differentiation, proliferation, and apoptosis, microRNAs are important regulators in innate and adaptive immune responses. Distinct sets of expressed miRNAs are found in different cell types and tissues and aberrant expression of microRNAs is associated with many disease states. MicroRNA expression was examined in a model of heterotopic heart transplantation by microarray analyses and a unique profile was detected in rejecting allogeneic transplants (BALB/C to C57BL/6) as compared to syngeneic transplants (C57BL/6 to C57BL/6). The microRNA miR-182 was significantly increased in rejecting cardiac allografts and in mononuclear cells that infiltrate the grafts. Forkhead Box (FOX) proteins are a family of important transcription factors and FOXO1 is a target of miR-182. As miR-182 increases after transplant, there is a concomitant post-transcriptional decrease in FOXO1 expression in heart allografts that is localized to both the cardiomyocytes and CD3+ T cells. The microRNA miR-182 is significantly increased in both PBMC and plasma during graft rejection suggesting potential as a biomarker of graft status. Our results identify microRNAs that may regulate alloimmune responses and graft outcomes. In total 14 microRNA microarrays data. For heart graft: each sample column corresponds to the expression profile of 3 pooled syngeneic or 3 pooled allogeneic heart grafts or 3 pooled normal heart. For graft infiltrating lymphocytes (GILs): each sample column corresponds to the expression profile of 3 pooled syngeneic or 3 pooled allogeneic GILs or 3 pooled normal PBMC.
Project description:To screen for candidate genes that may contribute to the pathogenesis of GalT-II deficiency Transcriptome-wide expression profiling using the Affymetrix Gene 1.0 ST platform comparing the gene expression patterns of skin fibroblasts of the two affected sisters with those of three healthy individuals. Comparison between two SEDMJL1 human fibroblasts and three healthy controls.
Project description:The frequency of delayed function of kidney transplants varies greatly and is associated with the quality of graft, donor age, and the duration of cold ischemia time. Body weight differences between donor and recipient can affect primary graft function. The underlying mechanism is poorly understood. Here, we have transplanted kidney grafts from commensurate body weight (L-WD) or reduced body weight (H-WD) donor rats into syngeneic or allogeneic recipients. 24 hours post-transplantation, serum creatinine level in H-WD recipients was significantly higher compared to that of L-WD recipients indicating impaired primary graft function. We detected a 10 fold higher transcription of IL-6 and dramatically increased tubular destruction in grafts from H-WD recipients. This was accompanied by decreased expression of genes associated with kidney function and an up-regulation of other genes such as cytochrome P450 isoforms, FosL and Trib3 as revealed by DNA microarray analysis. A single application of IL-6 into L-WD recipients is sufficient to impair primary graft function and to cause tubular damage. Whereas, immediate neutralization of IL-6 receptor signaling rescued primary graft function resulting in low serum creatinine levels, well-preserved kidney graft architecture and a normalized gene expression profile. These findings have strong clinical implication as anti-IL6R treatment of patients receiving grafts from lower-weight donors could be used to improve primary graft function. The dataset comprises eight samples divided into four sample groups. Each group represents rat kidneys collected after allogeneic transplantation under a certain condition and includes two biological replicates. The first group is characterized by a high body weight difference between donor and recipient, rats in the second group exhibit a low weight difference. Group three and four are similar to group one, but underwent an additional treatment with anti-IL6R mAb or prednisolone immediately after transplantation.