Project description:Technical replicates with 2 hybridizations (cat 350 and cat175) 1.04 agar Arabidopsis thaliana - columbia-0 plants, RNA-extracted by VIB, and amplified and hybridized at UNIL.
Project description:Combination of reverse- and chemical genetic screens reveals a network of novel angiogenesis inhibitors and targets Drug target identification and validation are bottlenecks in the drug discovery process. Accordingly there is a need to develop new methods to facilitate the development of much-needed innovative drugs. We have combined reverse- and chemical genetics to identify new targets modulating blood vessel development. Through mRNA expression profiling in mice we identified 155 drugable gene products that were enriched in the microvasculature. Orthologs of 50 of these candidates were knocked down in a reverse genetic screen in zebrafish. 16 of the 50 genes encoded products that affected angiogenesis. In parallel, screening of 300 known drugs and pharmacologically active compounds in a human cell-based angiogenesis assay identified 11 angiogenesis inhibitors. Strikingly the reverse- and chemical genetic screens identified an overlap of three gene products of the same superfamily of serine/threonine protein phosphatases and two compounds targeting that family. Furthermore, the gene products identified in the reverse genetic screen comprise an interacting network with the targets of the chemical genetic screen. Thus, combining reverse- and chemical genetic screens is a powerful approach to identify novel biological processes and drug targets in vertebrates. Keywords: Cell-type comparison 24 samples were analyzed, representing 8 sample groups. In every sample group, three biological replicates were hybridized separately. The microarrays were hybridized with a Cy3-labeled sample and Cy-5 labeled Common Reference (Universal Mouse Reference RNA, cat. No.: 740100, Stratagene) simultaneously.
Project description:In the present study, we demonstrate that hMSCs migrate toward human keratinocytes as well as toward conditioned medium from cultured human keratinocytes (KCM) indicating that the hMSCs can respond to signals from keratinocytes. Incubation of hMSCs with KCM induced dermal myofibroblast like differentiation characterized by expression of cytoskeletal markers vinculin and F-actin filaments with increased expression of alpha smooth muscle actin. We then examined the therapeutic efficacy of hMSCs in wound healing in two animal models representing normal and chronic wound healing. Accelerated wound healing, as determined by quantitative measurements of wound area, was observed when hMSCs and KCM exposed hMSCs (KCMSCs) were injected near the site of incisional/excisional wounds in nondiabetic athymic and NOD/SCID mice as compared with normal human fetal lung fibroblast WI38 cells or saline control induced wound healing. Experiment Overall Design: Keratinocyte conditioned media exposed MSCs(KCMSCs) were used in the experiment in triplicates as follows; Experiment Overall Design: KCM_1 Experiment Overall Design: KCM_2 Experiment Overall Design: KCM_3 Experiment Overall Design: Where as MSCs were exposed to Keratinocyte growth media for 30 days were used as a controls as follow; Experiment Overall Design: KGM_1 Experiment Overall Design: KGM_2 Experiment Overall Design: KGM_3 Experiment Overall Design: All the samples were analyzed.
Project description:We used gene microarray analysis to compare the global expression profile of genes involved in adaptation to exercise training in skeletal muscle from chronically strength/resistance trained (ST), endurance trained (ET) and untrained control subjects (Con). Resting skeletal muscle samples (~100mg) were obtained from the vastus lateralis of 20 subjects (Con n=7, ET n=7, ST n=6; trained groups >8 years specific training). Total RNA was extracted from tissue and microarrays completed, with test samples compared with standard human reference RNA. Subjects were characterised by performance measures of maximal oxygen uptake (VO2max) on a cycle ergometer and maximal concentric and eccentric leg strength on an isokinetic dynamometer. 263 genes were differentially expressed in trained (TR collectively ET + ST) subjects compared with Con (P<0.05) while 21 genes were different between ST and ET (P<0.05). Manual cluster analyses revealed significant regulation of genes involved in muscle structure and development in TR subjects compared with Con (P<0.05), and expression of these correlated significantly with measures of performance (P<0.05). ET had increased and ST decreased expression of gene clusters related to mitochondrial/oxidative capacity (P<0.05), and these mitochondrial gene clusters correlated significantly with VO2max (P<0.05). VO2max also correlated with expression of gene clusters that regulate fat and carbohydrate oxidation (P<0.05). We have demonstrated that chronic training has marked effects on basal gene expression by regulating levels of multiple mRNAs that transcribe genes for important functional groups in human skeletal muscle. Some specific gene clusters are expressed regardless of the training stimulus, whereas others exhibit divergent expression patterns as a result of specific training stimuli. Keywords: Comparative, cluster analysis, endurance training, strength training, muscle phenotype This was a crossectional study examining basal gene expression profiles of the human vastus lateralis. Twenty healthy males volunteered for this investigation. Seven were endurance trained cyclists (ET), who had been participating in endurance training for 8 yr. These subjects had no history of resistance training. Six subjects were strength trained power-lifters (ST) who had been participating exclusively in strength/resistance training for 9 yr. The final seven subjects were healthy controls (CON) that did not participate in any formal exercise. The study was approved by the Human Research Ethics Committee of RMIT University and Monash University Standing Committee on Ethics Research on Humans. After RNA extraction, amplification and indirect labelling, a dual colour micro-array analysis was conducted by hybridizing a test (muscle RNA; Cy5) sample and a reference (Universal human RNA; Cy3) sample on the AGRF glass slide human 8K micro-array. After data capture with the Genepix scanner and associated software data was normalised and the three populations compared using GeneSpring and simplified cluster analysis.