Project description:Pulmonary exposure to high doses of nanoparticles (NP) leads to well characterized lung toxicity in addition to long-term NP retention. However, pulmonary NP accumulation and toxicity following low dose exposures are not well described. In the present study we sought to: (1) investigate particle retention in mouse lungs following intratracheal instillation of varying doses of nano-sized titanium dioxide (nano-TiO2) and (2) determine the effects of long-term particle accumulation on pulmonary systems. Female C57BL/6 mice were exposed to rutile nano-TiO2 (primary size of 20.6 nm and surface area of 107.7 m2/g) via single intratracheal instillations of 18, 54 and 162 M-BM-5g/mouse and sampled 1, 3 and 28 days post-exposure. The deposition of nano-TiO2 in the lungs was assessed using Nanoscale Hyperspectral Microscope. DNA microarrays, pathway-specific real-time RT-PCR (qPCR) and gene-specific qPCR arrays, and tissue protein analyses were employed to characterize pulmonary responses. Hyperspectral mapping showed dose-dependent retention of nano-TiO2 in the lungs up to 28 days post-exposure time. Retention did not correlate with the extent of inflammatory neutrophil influx into the lungs. DNA microarray analysis showed altered expression of approximately 3000 genes across all treatment groups (M-BM-11.3 fold; p<0.1). Several inflammatory mediators changed in a dose- and time-dependent manner at both the mRNA and protein levels. Although the low dose exposure failed to induce observable inflammation, significant changes in the expression of genes and proteins associated with inflammation were observed. Moreover, diminished (or absent) neutrophil influx in the low and medium dose groups was correlated with negative regulation of genes associated with ion homeostasis and muscle regulation. Gene expression changes for several inflammatory mediators have previously been noted in mice exposed to the same nano-TiO2 via inhalation. Our results suggest that retention of nano-TiO2 in the absence of inflammation and effective clearance can perturb calcium and ion homeostasis, and affect smooth muscle activities over time. This experiment consists of three different dosages of TiO2, e.g., low (18 ug), medium (54 ug) and high (162 ug), and one control. There are 3 time points for each treatment and control group, e.g., day 1, day 3 and day 28. Each dose or time point has 5-6 biological replicates. There are total 65 samples (arrays)

Project description:Pancreatic ductal adenocarcinoma is one of the most lethal cancers, with a 5 year-survival rate below 5%. Lack of curative treatment and failure of targeted therapies urge the need to identify novel efficient therapeutic strategy. Achievement of this goal will be obtained through the identification of diagnosis and prognosis biomarkers, identification of novel therapeutic targets and the knowledge of resistance mechanisms induced by these targeted therapies. PI3K/Akt/mTOR signalling, one of the most altered in cancers, is overactivated in pancreatic cancer and correlated with poor prognosis. In the Vertebrates, the family of class I phosphoinoitide-3-kinase (PI3K) includes four isoforms: p110α, p110β, p110δ and p110γ. Although they all perform the same biochemical reaction (phosphorylation of PIP2 in PIP3, a membrane lipid messenger), each isoform were demonstrated to have specific physiological roles. Global PI3K inhibitors are currently being tested in phase I/II clinical trials in advanced solid cancers, but show at maximal doses tolerated a limited therapeutic benefit. Isoform-selective PI3K inhibitors are currently the most promising agents because, at low doses, they are more efficient to inhibit one PI3K isoform, and thus, less toxic than pan-PI3K inhibitors. The objectives of this thesis are to determine isoform-specific PI3K roles and the therapeutic interest to target one or more isoforms in pancreatitis and PDAC, by the identification of isoform-specific pathways and the study of adaptive responses induced by targeting of one or all isoforms of PI3K. In a first part, my work has highlighted, validated and completed results obtained in the team, to demonstrate the significance of PI3K/Akt signalling in two physiological processes: chronic pancreatitis and initiation of pancreatic carcinogenesis. Precisely, the overactivation of PI3K/Akt pathway measured on human and murine chronic pancreatitis samples is correlated with a specific p110α activation gene expression signature. Moreover, genetic and pharmacologic inactivation of p110α during pancreatic chronic inflammation or cancerogenesis (by oncogenic Kras) prevents the formation of acino-ductal metaplasia, structures at the origin of pancreatic carcinogenesis initiation. Development of in vitro acino-ductal transdifferentiation protocol allowed me to demonstrate that only p110α is necessary at this initial step of pancreatic carcinogenesis by the regulation of Rho small GTPases, further regulating actin remodelling. In the second part, by a phosphoproteomic-based approach, I quantified PI3K downstream phosphorylation-regulated targets in a pancreatic cancer cell line treated or not by a pan- or selective PI3K inhibitor at different times. I demonstrated for the first time existence of targets, signalling pathways and adaptive responses regulated by each PI3K isoform. To conclude, all these results demonstrate the rational of combinatorial use of isoform-specific PI3K inhibitors in patients with pancreatic cancer for better clinical response

Project description:An estimated 1% or less of the nanoparticles (NPs) deposited in lungs translocate to systemic circulation and enter other organs; however this estimation may not be accurate considering the low sensitivity of the existing in vivo NP detection methods. Moreover, the biological outcomes of such low levels of translocation are not elucidated. The objectives of the present study were to employ a Nano-scale Hyperspectral Microscope to spatially observe and spectrally profile NPs in tissues, and characterize the effects of NPs in blood, liver and heart following pulmonary deposition and subsequent translocation from lungs. Adult female C57BL/6 mice were exposed via intratracheal instillation to 18 and 162 µg per mouse of industrially relevant non-doped titanium dioxide nanoparticles (nano-TiO2). Using the Nano-scale Hyperspectral Microscope translocation to heart and liver was confirmed at both doses and to blood at the highest dose at 24 hours post-exposure time-point. The analysis of biological effects using DNA microarrays, RT-qPCR and ELISA revealed activation of complement cascade and inflammatory process in heart and specific activation of complement factor 3 in blood, potentially suggestive of activation of early innate immune response essential for particle opsonisation and clearance. The liver showed subtle response with changes in the expression of few genes associated with acute phase genes. This study establishes a direct link between particle translocation and systemic effects. An estimated 1% or less of the nanoparticles (NPs) deposited in lungs translocate to systemic circulation and enter other organs; however this estimation may not be accurate considering the low sensitivity of the existing in vivo NP detection methods. Moreover, the biological outcomes of such low levels of translocation are not elucidated. The objectives of the present study were to employ a Nano-scale Hyperspectral Microscope to spatially observe and spectrally profile NPs in tissues, and characterize the effects of NPs in blood, liver and heart following pulmonary deposition and subsequent translocation from lungs. Adult female C57BL/6 mice were exposed via intratracheal instillation to 18 and 162 µg per mouse of industrially relevant non-doped titanium dioxide nanoparticles (nano-TiO2). Using the Nano-scale Hyperspectral Microscope translocation to heart and liver was confirmed at both doses and to blood at the highest dose at 24 hours post-exposure time-point. The analysis of biological effects using DNA microarrays, RT-qPCR and ELISA revealed activation of complement cascade and inflammatory process in heart and specific activation of complement factor 3 in blood, potentially suggestive of activation of early innate immune response essential for particle opsonisation and clearance. The liver showed subtle response with changes in the expression of few genes associated with acute phase genes. This study establishes a direct link between particle translocation and systemic effects. This experiment consists of one dose of nano-TiO2 (162 ug) and one control. There are 2 time points for each treatment and control group, e.g., day 1 and day 28. Each dose or time point has 5-6 biological replicates. There are total 22 samples (arrays)

			Action	DRS
	Experimentaldesign.xlsx	Xlsx
	MQ_TiO2_1.zip	Other
	MQ_TiO2_2.zip	Other
	MQ_TiO2_3.zip	Other
	MQ_TiO2_4.zip	Other

Dataset Information

Time-resolved phosphoproteomic analysis of the TCR signaling pathway in primary T cells

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LymphoAtlas: a dynamic and integrated phosphoproteomic resource of TCR signaling in primary T cells reveals ITSN2 as a regulator of effector functions.

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