Transcriptome of CDKN1C-siRNA-injected embryos were compared to sham-injected embryos using RNA-sequencing to determine the genes and pathways downstream of the silenced gene that may have been altered. Overall design: Transcriptome comparison between two pools of embryos (i.e. CDKN1C-siRNA-injected vs sham-injected embryos)
Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos.
Specimen part, Subject
View SamplesUsing fluorescence activated cell sorting, we isolated CD45+, CSF1R-GFP+, F4/80+, Ly6G- mouse lung monocytes and macrophages at 7 days after pneumonectomy procedure. We then used microfluidic single cell RNA-sequencing to transcriptional profile unique myeloid subsets. Using the pneumonectomy dataset, we identified 6 cell groups and 4 gene groups that marked several regenerative macrophage subsets including CCR2+, Ly6C+ monocytes and CD206+, Chil3+ M2-like macrophages. Overall design: individual macrophages 7 days post-pneumonectomy in a B6 CSF1R-GFP mouse
Recruited Monocytes and Type 2 Immunity Promote Lung Regeneration following Pneumonectomy.
Specimen part, Subject
View SamplesUsing fluorescence activated cell sorting, we isolated CD45+, CSF1R-GFP+, F4/80+, Ly6G- mouse lung monocytes and macrophages at 7 days after sham thoracotomy procedures. We then used microfluidic single cell RNA-sequencing to transcriptional profile unique myeloid subsets. Overall design: After sequencing 31 single cell transcriptomes were analyzed. Hierarcical and k-means clustering reveals several populations of macrophages are present in the lung.
Recruited Monocytes and Type 2 Immunity Promote Lung Regeneration following Pneumonectomy.
Specimen part, Subject
View SamplesFlow sorted mouse LNEP cells isolated from normal lungs and influenza-infected lungs (day 17 PR8) Overall design: Highly purified quiescent LNEPs (EpCAMposb4posCC10negFoxJ1neg) and activated LNEPs (Krt5-CreERT2 traced cells 17 days post infection) were flow sorted and extracted for RNA using ReliaPrepâ„¢ RNA Tissue Miniprep kit (Promega).
Local lung hypoxia determines epithelial fate decisions during alveolar regeneration.
Specimen part, Subject
View SamplesPurpose: The goals of this study were to identify the molecular alterations in the SCA1 inferior olive, and determine whether these changes are found in other affected tissues. Methods: mRNA profiling was conducted in two different SCA1 mouse models (Atxn1 154Q/2Q KI and ATXN1-82Q Tg), in two different affected tisues (inferior olive and cerebellum) during early disease initiation and progression (5 week and 12 week time-points). All analyses were conducted relative to appropriate wild-type controls. TopHat2 v2.1.0 was utilized to align reads to the mouse reference genome (mm10) before quantification and differential expression analysis with Cufflinks v2.2.1. Normalized expression values were generated using Cuffnorm. Results: Differentially regulated genes identified in the SCA1 inferior olive segregated into several enriched biological pathways, including the Defense Response at 12 weeks of age. Our study demonstrates that vulnerable tissues in SCA1 are not uniform in their gene expression changes, and express discrete and commonly enriched biological pathways. In addition, we found that brain region-specific differences occur early in disease initiation and progression at 5 weeks of age. Conclusions: The findings from this study suggest that different mechanisms of neurodegeneration are at work in the SCA1 inferior olive and cerebellum. Overall design: mRNA profiling was conducted on an Illumina HiSeq 2500. Three biological replicates were sequenced for each genotype (Atxn1 154Q/2Q KI mice and wild-type controls; ATXN1-82Q Tg mice and wild-type controls) in each brain region (inferior olive and cerebellum) at each time-point (5 weeks old and 12 weeks old), yielding a total of 48 biological samples.
Molecular pathway analysis towards understanding tissue vulnerability in spinocerebellar ataxia type 1.
Age, Specimen part, Cell line, Subject
View SamplesMotherhood involves a switch in natural rewards, whereby offspring become highly rewarding. Nucleus accumbens (NAC) is a key CNS region for natural rewards and addictions, but to date no study has evaluated on a large scale the events in NAC that underlie the maternal change in natural rewards. In this study we utilized microarray and bioinformatics approaches to evaluate postpartum NAC gene expression changes in mice. Modular Single-set Enrichment Test (MSET) indicated that postpartum (relative to virgin) NAC gene expression profile was significantly enriched for genes related to addiction and reward in 5 of 5 independently curated databases (e.g., Malacards, Phenopedia). Over 100 addiction/reward related genes were identified and these included: Per1, Per2, Arc, Homer2, Creb1, Grm3, Fosb, Gabrb3, Adra2a, Ntrk2, Cry1, Penk, Cartpt, Adcy1, Npy1r, Htr1a, Drd1a, Gria1, and Pdyn. ToppCluster analysis found maternal NAC expression profile to be significantly enriched for genes related to the drug action of nicotine, ketamine, and dronabinol. Pathway analysis indicated postpartum NAC as enriched for RNA processing, CNS development/differentiation, and transcriptional regulation. Weighted Gene Coexpression Network Analysis identified possible networks for transcription factors, including Nr1d1, Per2, Fosb, Egr1, and Nr4a1. The postpartum state involves increased risk for mental health disorders and MSET analysis indicated postpartum NAC to be enriched for genes related to depression, bipolar disorder, and schizophrenia. Mental health related genes included: Fabp7, Grm3, Penk, and Nr1d1. We confirmed via quantitative PCR Nr1d1, Per2, Grm3, Penk, Drd1a, and Pdyn. This study indicates for the first time that postpartum NAC involves large scale gene expression alterations linked to addiction and reward. Because the postpartum state also involves decreased response to drugs, the findings could provide insights into how to mitigate addictions.
Addiction and reward-related genes show altered expression in the postpartum nucleus accumbens.
Specimen part
View SamplesThe transition from the non-maternal to the maternal state is characterized by a variety of CNS alterations that support the care of offspring. The septum (including lateral and medial portions) is a brain region previously linked to various emotional and motivational processes, including maternal care. In this study, we used microarrays (PLIER algorithm) to examine gene expression changes in the septum of postpartum mice and employed gene set enrichment analysis (GSEA) to identify possible regulators of altered gene expression. Genes of interest identified as differentially regulated with microarray analysis were validated with quantitative real-time PCR. We found that fatty acid binding protein 7 (Fabp7) and galanin (Gal) were downregulated, whereas insulin-like growth factor binding protein 3 (Igfbp3) was upregulated in postpartum mice compared to virgin females. These genes were previously found to be differentially regulated in other brain regions during lactation. We also identified altered expression of novel genes not previously linked to maternal behavior, but that could play a role in postpartum processes, including glutamate-ammonia ligase (Glul) and somatostatin receptor 1 (Sstr1) (both upregulated in postpartum). Genes implicated in metabolism, cell differentiation, or proliferation also exhibited altered expression. Unexpectedly, enrichment analysis revealed a high number of microRNAs, transcription factors, or conserved binding sites (177 with corrected P-value <0.05) that were significantly linked to maternal upregulated genes, while none were linked to downregulated genes. MicroRNAs have been linked to placenta and mammary gland development, but this is the first indication they may also play a key role in sculpting the maternal brain. Together, this study provides new insights into genes (along with possible mechanisms for their regulation) that are involved in septum-mediated adaptations during the postpartum period.
Gene expression changes in the septum: possible implications for microRNAs in sculpting the maternal brain.
Specimen part
View SamplesIn order to determine the regulators of outer hair cell postnatal maturation, we utilized the RiboTag mouse model to perform a detailed transcriptomic analysis of outer hair cells at five postnatal developmental time points: P8, P14, P28, 6 weeks (6wk) and 10 weeks (10wk). This analysis resulted in consistent enrichment of outer hair cell expressed genes in the immunoprecipitated RNA compared to whole cochlear input RNA from each time point. Using transcription factor binding motif prediction on a set of defined outer hair cell enriched genes, we further use this dataset to identify the helios transcription factor as a regulator of the postnatal outer hair cell transcriptome. Overall design: Examination of the outer hair cell translatome by outer hair cell expressed HA-tagged ribosomal immunoprecipitation at 5 postnatal timepoints (P8, P14, P21, 6wk and 10wk). Immunoprecipitated samples were compared to input cochlear RNA controls in independent biological duplicates or triplicates.
Helios is a key transcriptional regulator of outer hair cell maturation.
Specimen part, Subject
View SamplesIn order to determine the transcriptional effect of Ikzf2 overexpression in mammalian auditory hair cells, mouse cochleae were transfected with either a control GFP or a Ikzf2 virus between postnatal day 1 and 3 (P1-3) and then harvested for single cell gene expression profiling at postnatal day 8 (P8) on the 10X Genomics Single Cell 3' v2 platform. Overall design: Dataset is composed of two separate single cell RNA-Seq samples captured on the 10X Genomics Chromium platform with the Single Cell 3' Solution v2 chemistry. Viral GFP- (control) or Ikzf2-transfected cochleae were harvested and single cell suspensions prepared from transgenic mice expressing tdTomato in hair cells. Hair cells expressing tdTomato were enriched by flow sorting and then captured on the Chromium system in parellel. Library preparation and sequencing was performed as defined by the 10X Genomics Single Cell 3' Solution v2 protocol.
Helios is a key transcriptional regulator of outer hair cell maturation.
Specimen part, Cell line, Treatment, Subject
View SamplesCoordinated gene expression changes across the CNS help to produce the mammalian maternal phenotype. Lateral septum (LS) is a brain region critically involved with aspects of maternal care, and we recently examined gene expression of whole septum (LS and medial septum) in selectively bred maternal mice. Here, we expand on the prior study by 1) conducting microarray analysis solely on LS in virgin and postpartum mice, 2) using outbred mice, and 3) evaluating the role of sensory input on gene expression changes. Large scale changes in genes related to neuronal signaling were identified, including nine GABAA receptor subunits (p<0.05). Subunits 4 and were downregulated in maternal LS, likely reflecting a reduction in the extrasynaptic, neurosteroid-sensitive 4/ containing receptor subtype. Conversely, subunits and were increased in maternal LS. Sixteen K+ channel related genes showed altered expression, as did dopamine receptors Drd1a and Drd2 (both downregulated), hypocretin receptor 1 (Hcrtr1), kappa opioid receptor 1 (Oprk1), and transient receptor potential channel 4 (Trpc4). Expression of a large number of genes linked to developmental processes or cell differentiation were also altered in postpartum LS, including chemokine (C-X-C) motif ligand 12 (Cxcl12), fatty acid binding protein 7 (Fabp7), plasma membrane proteolipid (Pllp), and suppressor of cytokine signaling 2 (Socs2). Additional genes that are linked to anxiety, such as glutathione reductase (Gsr), exhibited altered expression. Pathway analysis also identified changes in genes related to cyclic nucleotide metabolism, chromatin structure, and the Ras gene family. The sensory presence of pups was found to contribute to the altered expression of a subset of genes across all categories. This study suggests that both large changes in neuronal signaling and the possible terminal differentiation of neuronal and/or glial cells play important roles in producing the maternal state.
Large scale expression changes of genes related to neuronal signaling and developmental processes found in lateral septum of postpartum outbred mice.
Specimen part
View Samples