Project description:We generated Ikkα-KA/KA knock-in mice (KA/KA), in which an ATP binding site of Ikkα Lys 44 was replaced by alanine. The knock-in mice develop severe skin lesions and begin to die after 6 to 10 months. We also found lung SCCs in some of the mice. To study lung SCC development, we stabilize the skin condition by crossing KA/KA with Lori.Ikkα transgenic mice to generate KA/KA-Lori.Ikkα mice, which 100% spontaneously developed lethal lung SCC at 4 to 6 months of age. Clodronate-liposomes obtained from Dr. N. van Rooijen (Vrije Universiteit, Amsterdam, Netherlands) were intravenously injected into mice twice every week, started in 4 weeks old mice, and continued for 3 months. The dose of clodronate-liposomes was 100µl during the first month and elevated to 150 100µl after that. The depletion efficacy was confirmed by detecting pulmonary macrophages with Flow Cytometry. Depletion of macrophages in these KA/KA-Lori.Ikkα mice could significantly reduce inflammation and prevent lung SCC developmrnt. This aim of this microarray assay is to identify differentially expressed genes among Wild type, KA/KA-Lori.Ikkα, and macophage-depleted KA/KA-Lori.Ikkα mice, which may highlight the important genes or parthways involved in inflammation-associated lung SCC carcinogenesis. Three samples were analyzed: 1. WT (lung tissue from wild type mouse); 2. KA2 (mixture tissue of lung SCC and tumor neighbor from KA/KA-Lori-Ikkα mouse ); 3. KAT (lung tissue from KA/KA-Lori-Ikkα mouse with macrophage depletion treatment)
Project description:NAP - neuroprotective peptide demonstrates increase in neuronal survival when injected into the hippocampus of rats in the model of epilepsy Microarray analysis was used to understand the expression of genes following KA treatment and the changes in gene expression following KA+NAP treatment Keywords: stress response KA was injected into the hippocampus of Sprague-Dawley rats. The other group of rats was injected with KA and NAP(10-13M). The third group was injected with NAP only and the last group was injected with PBS as a vehicle. CA3 area of hippocampus was removed 24h later and RNA extraction was done. The samples were subjected to microarray analysis.
Project description:Temporal lobe epilepsy (TLE) can develop from alterations in hippocampal structure and circuit characteristics, and can be modeled in mice by administration of kainic acid (KA). Adult neurogenesis in the dentate gyrus (DG) contributes to hippocampal functions and has been reported to contribute to the development of TLE. Some of the phenotypical changes include neural stem and precursor cells (NPSC) apoptosis, shortly after their birth, before they produce hippocampal neurons. Here we explored these early phenotypical changes in the DG 3 days after systemic KA administration to mice. Our specific aim was to understand the molecular mechanisms underlying altered apoptosis levels in NSPC following KA-induced status epilepticus (KA-SE). Accordingly, we chose a multi-omics experimental setup and analyzed DG tissue samples using proteomics, transcriptomics and microRNA profiling techniques. We here present a description of how these date were obtained and provide them to others for further analysis and validation. This may help to further identify and characterize molecular mechanisms involved in the alterations induced shortly after KA-SE in the mouse DG. Total RNA obtained from dentate gyrus 72h after mice were subjected to repeated low dose kainic acid induced status epilepticus or saline i.p. injections
Project description:Global gene expression analysis of induced pluripotent stem cell lines and their corresponding source cells Total RNA was harvested from H9 hESC (P51), non-integrated episomal CB-iPSC clones 6.2, 6.11, 6.13, (P14), 19.11, (P11), nonviral KER-iPSC clones KA.1, KA.3 (P13) nonviral FFB-iPSC: F.1, F.6 (P14) and viral fibroblast iPSC clones IMR1 (P66), IMR4 (P64). A single sample of each condition was used for this analysis.
Project description:We generated Ikkα-KA/KA knock-in mice (KA/KA), in which an ATP binding site of Ikkα Lys 44 was replaced by alanine. The knock-in mice develop severe skin lesions and begin to die after 6 to 10 months. We also found lung SCCs in some of the mice. To study lung SCC development, we stabilize the skin condition by crossing KA/KA with Lori.Ikkα transgenic mice to generate KA/KA-Lori.Ikkα mice, which 100% spontaneously developed lethal lung SCC at 4 to 6 months of age. Clodronate-liposomes obtained from Dr. N. van Rooijen (Vrije Universiteit, Amsterdam, Netherlands) were intravenously injected into mice twice every week, started in 4 weeks old mice, and continued for 3 months. The dose of clodronate-liposomes was 100µl during the first month and elevated to 150 100µl after that. The depletion efficacy was confirmed by detecting pulmonary macrophages with Flow Cytometry. Depletion of macrophages in these KA/KA-Lori.Ikkα mice could significantly reduce inflammation and prevent lung SCC developmrnt. This aim of this microarray assay is to identify differentially expressed genes among Wild type, KA/KA-Lori.Ikkα, and macophage-depleted KA/KA-Lori.Ikkα mice, which may highlight the important genes or parthways involved in inflammation-associated lung SCC carcinogenesis.
Project description:Recently, enamel peptides revealed their critical role in sex classification of old skeletal remains. In particular, the presence of protein AMELY (amelogenin isoform Y) within the enamel dental tissue is restricted to the male sex only, while AMELX (isoform X) can be found in both sexes. AMELY can be easily detected through peptide SM(ox)IRPPY by means of LC-MS/MS and retrieved from ion chromatograms (exact mass [M+2H]+2 440.2233 m/z). In this paper, we exploited the dimorphic features of the amelogenin protein to unravel the sex of the so-called ‘Lovers of Modena’, two Late Antique individuals whose skeletons were intentionally buried hand-in-hand.
Project description:Injury of the CA1 subregion induced by a single injection of kainic acid (1×KA) is attenuated when juvenile animals (P20) have a history of two sustained neonatal seizures on P6 and P9. To identify gene candidates involved in the spatially protective effects produced by early life conditioning seizures, we profiled and compared the transcriptomes of CA1 subregions from control, 1×KA, and 3×KA treated animals. More genes were regulated following 3×KA (9.6%) than after 1×KA (7.1%). Following 1×KA, genes supporting oxidative stress, growth, development, inflammation, and neurotransmission were upregulated (e.g., Cacng1, Nadsyn1, Kcng1, Aven, S100a4, GFAP, Vim, Hrsp12, Grik1). After 3×KA, protective genes were differentially over-expressed (e.g., Cat, Gpx7, GAD1, Hspa12A, Foxn1, adenosine A1 receptor, Ca2+ adaptor and homeostatic proteins, Cacnb4, Atp2b2, anti-apoptotic Bcl-2 gene members, intracellular trafficking protein, Grasp, suppressor of cytokine signaling (Socs3)). Distinct anti-inflammatory interleukins not observed in adult tissues (e.g., IL6 transducer, IL23 and IL33 or their receptors (ILF2)) were also over-expressed. Several transcripts were validated by real-time polymerase chain reaction (QPCR) and immunohistochemistry. QPCR showed that casp 6 was increased after 1×KA but reduced after 3×KA; pro-inflammatory gene cox1 was either upregulated or unchanged after 1×KA but reduced by ~70% after 3×KA. Enhanced GFAP immunostaining following 1×KA was selectively attenuated in the CA1 subregion after 3×KA. The observed differential transcriptional responses may contribute to early life seizure-induced pre-conditioning and neuroprotection by reducing glutamate receptor-mediated Ca2+ permeability of the hippocampus and redirecting inflammatory and apoptotic pathways which could lead to new genetic therapies for epilepsy.
Project description:Multiple transcription factors regulate B cell commitment, which coordinates with myeloid–erythroid lineage differentiation. One such factor, NF-kB, has long been speculated to regulate early B cell development; however, this issue remains controversial. IKKa is required for splenic B cell maturation, but not for bone marrow (BM) B cell development. Here, we unexpectedly found defective BM B cell development and an increased myeloid–erythroid lineages in kinase-dead IKKa (KA/KA) knock-in mice. Markedly increased cytosolic p100, an NF-kB2 inhibitory form, and reduced nuclear NF-kB p65, RelB, p50, and p52, as well as IKKa, was observed in KA/KA splenic and BM B cells. Several B- and myeloid–erythroid-cell regulators, including Pax5, were deregulated in KA/KA BM B cells. Using fetal liver and BM congenic transplants, and IKKa deletion from early hematopoietic cells in mice, this defect was identified as B cell intrinsic and as an early event during hematopoiesis. Re-expression of IKKa, Pax5, or combined NF-kB molecules promoted B cell development, but repressed myeloid–erythroid cell differentiation in KA/KA BM B cells. Together, these results demonstrate that IKKa regulates B-lineage commitment via combined canonical and noncanonical NF-kB transcriptional activity to target Pax5 expression during hematopoiesis.
Project description:We have reported that pulmonary infiltrating macrophages, which highly express inducible nitric-oxide synthase (iNOS/NOS2), play a critical role for driving spontaneous lung SCC development in a new mouse model (L-Ikka KA/KA; KA/KA) that mimics human lung SCC development. However, the role of NOS2 in lung SCC development is unknown. Our data suggests deletion of the gene iNOS prevents tumorogenesis in this mouse model. We use microarray data to compare levels of expression of genes associated with lung tumor development and prevention.
Project description:KA-PARP1 was expressed in HeLa-KO PARP1 cells. Protein targets of KA-PARP1 were determined in cell lysate by using a DTB-modified NAD+analog.